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ESSENTIALS OF GENETIC DISORDERS

Genetic factors play a significant role in all cardiovascular disorders (see also Chap. 10). Genetic defects areresponsible for malformations of the heart and blood vessels, which account for the largest number of human birth

defects. The estimated incidence of congenital heart disease is approximately 1% of all live births.1 The prevalence isestimated to be 10-fold higher among stillbirths.2 Genetic defects are responsible for familial cardiovasculardisorders, such as cardiomyopathies and the long QT (LQT) syndrome, as well as nonfamilial sporadic forms.Evidence is now becoming available indicating genetic predisposition for common complex phenotypes, such asatherosclerosis and hypertension. Molecular genetics in conjunction with cytogenetics provide the opportunity todecipher the genetic basis and pathogenesis of cardiovascular diseases. Given the rapid pace of genetic discoveries, itis expected that genetic diagnosis and screening will become incorporated into standard practice in the near future. Itis thus imperative that cardiologists understand the basis for genetic disorders and the medical and ethicalimplications of genetics.

Basis for Genetic TransmissionAll hereditary information is transmitted through DNA, a linear polymer composed of purine (adenine, guanine) andpyrimidine (cytosine, thymine) bases. The gene is the basic hereditary unit. It consists of a distinct fragment of DNA,

which encodes a specific polypeptide (protein). There are approximately 30,000 genes in the human genome.3 Eachindividual has two copies of each gene, called alleles. The genes are localized in a linear sequence along 23 pairs ofchromosomes, including 22 pairs of autosomes (chromosomes 1 to 22) and 1 pair of sex chromosomes, X and Y.Females have two X chromosomes, whereas males carry one X and one Y chromosome. Each parent contributes oneof each chromosome pair (the members of the pair are referred to as homologous chromosomes) and thus one copyof each gene. The site at which a gene is located on a particular chromosome is referred to as the genetic locus. Agiven gene always resides at the same genetic locus on a particular chromosome, so the loci on homologouschromosomes are identical. However, alleles residing at these loci may be identical or different, leading tohomozygous (identical alleles) and heterozygous (two different alleles present at the locus) states.

The genetic information is encoded by the linear sequence of the four bases of the DNA. Translation of thisinformation into protein is through a translational code passed on through messenger ribonucleic acid (mRNA). Eachunit of three bases, referred to as a codon, encodes a specific amino acid. The transcribed mRNA serves as thetemplate that determines the sequence of the amino acids in the resulting polypeptide. Both autosomal alleles areusually transcribed into mRNA and translated into protein. However, expression of a gene can be restricted to specificcells and organs or regulated during a developmental stage because of regulation by cell- and tissue-specifictranscription factors. In cells that carry two X chromosomes, only one X is active and the other X is silent after earlyembryogenesis.

Classification of Genetic DisordersIn general, DNA nucleotide sequences remain stable during transmission to offspring. Nonetheless, occasional basesequence changes do occur, which are referred to as mutations. Mutations represent stable, heritable alterations inDNA. Somatic mutations, however, are not heritable. A number of mutagenic factors'such as environmental agents,radiation, chemicals, and errors by the DNA synthetic and editing enzymes—can induce mutations. Mutations caninvolve a visible alteration at the level of the chromosome (chromosomal abnormalities), which can result in thedeletion or translocation of a portion of the chromosome, whereby several genes are often eliminated or altered. Incontrast, mutations can be restricted to minor alterations in the DNA sequence, which vary from the substitution of asingle nucleotide to that of the deletion or addition of multiple nucleotides. Thus hereditary and congenital diseasesare conventionally classified into three broad categories: (1) chromosomal abnormalities, (2) single-gene ormonogenic disorders, and (3) polygenic disorders or complex traits.

CHROMOSOMAL ABNORMALITIESEach human cell has two copies of each chromosome (diploids) and each chromosome has two arms, referred to asthe long, or "q," and the short, or "p," arms. The arms of the chromosomes meet at a primary constriction referred toas the centromere. Mutations typically occur during meiosis when chromosomes separate. Mutations can involve largedeletions, duplications, translocations, rearrangements, and aneuploidy (too few or too many chromosomes).Chromosomal abnormalities are relatively common during embryonic life and lead to spontaneous abortion, oftenduring the first trimester of pregnancy. However, a significant number of fetuses with chromosomal abnormalities

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survive. Chromosomal aberrations, numerical or structural, occur in approximately 1 in 150 liveborn infants.4 Mostdiseases caused by chromosomal abnormalities are detected in neonates or infants because involvement of manygenes causes phenotypes that are easily diagnosed on physical examination. Chromosomal abnormalities often lead

to structural heart defects and are found in 5% to 13% of liveborn children with congenital heart disease.1

The usual cause for gain of a chromosome is nondisjunction because of failure of a homologous pair of chromosomesto separate during meiosis. When an additional copy of the chromosome is added during fertilization, three copies ofthe same chromosome (or only one copy) are found in the new zygote instead of the chromosome pair. Two of themost common chromosomal disorders causing heart disease in the adult, namely Down syndrome (trisomy 21) andTurner syndrome (XO), are both commonly caused by nondisjunction. Chromosomal rearrangements occur when achromosome breaks and rejoins within itself incorrectly, which can result in an inversion of the genetic material.Inversion occurs when a chromosome breaks at two points and the intermediate segment reunites in invertedorientation. Typically, there is no apparent phenotype in persons carrying an inversion, but their offspring may havesevere abnormalities due to the disruption in chromosome pairing that can take place during meiosis. Isochromes areformed when two short or long arms join with loss of the other arm. Chromosomal translations occur when breaksarise in two chromosomes that are reunited after exchange of segments. Chromosome duplications or gains ofchromosomal material may also be associated with phenotypic abnormality, but most commonly, they cause noobvious aberration.

Chromosome deletions are large deletions (equal to or greater than 106 base pairs) that commonly lead to loss of alarge amount of DNA and loss or disruption of multiple genes. Consequently, a series of phenotypes in a singleindividual may be present as a result of interruptions in a series of genes within the loci of a single chromosome.

Each human genome also contains a large number of structural variations that include insertions, deletions,

duplications, and rearrangement.5 The size of the structural variations could vary from 2 base pairs to several millionbase pairs. Insertion or deletion structural variations that change the copy number of the genes are commonlyreferred to as copy number variants (CNVs). Structural variants or CNVs are relatively common in the population butdo not necessarily cause a gross phenotype, as typically observed in chromosomal abnormalities. However, structuralvariations including CNVs play significant roles in susceptibility to complex phenotypes, such as developmentalabnormalities associated with congenital heart defects and neurologic disorders.

SINGLE-GENE DISORDERSA single-gene disorder is an inherited disease that can be caused by a mutation in a single gene. Single-genedisorders show a mendelian pattern of inheritance. They are classified as autosomal dominant, autosomal recessive,or X-linked (dominant or recessive). The majority of monogenic diseases exhibit an autosomal-dominant mode ofinheritance. Therefore, in a given family, approximately half of the members are affected. Monogenic disorders withan autosomal-recessive inheritance are caused by mutations in both copies of the gene. Therefore, in a given familyonly 25% of the offspring exhibit the phenotype, 50% carry the mutation, and 25% are normal. In X-linkedinheritance, males exhibit the disease and females are usually free of the phenotype but carry the mutation.However, if the mutation involves a major protein, the effect of the mutation may be dominant and females canexhibit the clinical phenotype. In diseases caused by mitochondrial DNA mutations, inheritance is from the mother(no male-to-male transmission), because mitochondrial DNA is predominantly inherited from the ovum.

Only a fraction of cardiovascular disorders are monogenic. The DNA mutation gives rise to a change in thecorresponding amino acids of the encoded protein and exerts its deleterious effects via functional alterations. Achange in even one amino acid located in a critical domain of the protein can enhance the function (gain-of-functionmutation) or impair the function (loss-of-function mutation), with a concomitant change in the phenotype. On

average, a mutation occurs every 106 cell divisions or once every 200,000 years. Only mutations occurring in thegametes are transmitted.

In single-gene disorders, although the presence of the causal mutations is necessary for the development of thedisease, other factors also affect the phenotypic expression of the disease. Modifier genes (the genetic background ofthe affected subjects) and the environmental factors are major determinants of phenotypic expression of a single-gene disorder.

COMMON POLYGENIC DISORDERSPolygenic disorders such as coronary artery disease and related traits are caused by the interaction of geneticvariants and nongenetic factors. Therefore, in this setting, the presence of a single variant is not sufficient to cause adisease, nor will its absence prevent development of the disease. Essentially, all of the common disorders have agenetic predisposition due to genetic variants that occur more frequently than that of rare single-gene disorders.Polygenic disorders account for the majority of the cardiovascular diseases, including atherosclerosis, hypertension,obesity, and diabetes mellitus. In polygenic diseases, multiple genes predispose to the condition. The mutations




responsible for genetic predisposition in polygenic disorders usually involve single nucleotides distributed throughout

the human genome with a frequency of approximately 1 per 1000 base pairs (bp).6,7 These single nucleotidesimparting changes to the DNA sequence are referred to as single-nucleotide polymorphisms (SNPs). The effects ofSNPs could confer protection against or susceptibility toward a complex phenotype and may be located in exons,

introns, or intergenic regions.8 Technology made available in 2005 provided the opportunity to perform genome-wideassociation studies, which has led to the mapping of hundreds of loci predisposing to polygenic diseases, includingcoronary artery disease (discussed later).

Classification of MutationsMost human diseases exhibit genetic heterogeneity, defined as being caused by different genes and mutationscausing the same phenotype. The heterogeneity may arise from multiple mutations in one gene (allele heterogeneity)or in two or more genes (locus heterogeneity). Within any one family, however, typically there is one causal gene andmutation in all affected members; uncommonly there are two different causal mutations or genes transmitted for thesame disease. A good example is familial hypertrophic cardiomyopathy (HCM), which involves more than a dozendifferent genes (locus heterogeneity) with multiple mutations in each (allelic heterogeneity). In less than 10% of thecases, HCM is caused by double mutations.

Mutations can involve a microscopically visible alteration, such as deletion or translocation of a portion of thechromosome (chromosomal abnormalities), or a minute change in the DNA sequence, such as alteration of one purineor pyrimidine base. Mutations involving only a single nucleotide are known as point mutations and are responsible for70% of all adult single-gene disorders. A point mutation may be a substitution of one nucleotide for another,

changing the amino acid sequence (missense mutation), or it may change from encoding an amino acid to become astop codon, which will truncate the protein (truncated or nonsense mutation), or it may eliminate a stop codon so theprotein is elongated (elongated mutant). Finally, it may change the codon without changing the amino acid sequence(synonymous mutation). All genes during transcription and translation are read from 5' to 3' orientation, with eachtriplet of bases (codon) coding for a specific amino acid. If a nucleotide is deleted (deletion) or an additionalnucleotide is inserted (insertion), it will shift the reading frame. The resulting protein would be entirely different(frameshift mutation) and usually nonfunctional. If a purine nucleotide is substituted for a pyrimidine or vice versa,the mutation is referred to as a transversion. If purine or pyrimidine substitutes for another purine or pyrimidine,respectively, it is called a transition. Other mutations may result from the deletion or addition of several nucleotides.In one form of myotonic dystrophy, for example, a triplet repeat of several thousand nucleotides in length is insertedinto the 3' end of the gene. Another type of mutation is known as a gene conversion, where two genes interact andpart of the nucleotide sequence of one gene becomes incorporated into that of the other. Mutations in genes exerttheir deleterious effects via a structural alteration of the protein that has functional consequences, as noted.

Genetic Penetrance and ExpressivityThe percentage of individuals within a family who have inherited the causal mutation and have one or more featuresof the disease is referred to as the penetrance. Penetrance is an all-or-none phenomenon. Any manifestation,however minute, indicates that the gene has penetrance in that individual. Nonpenetrance refers to lack of anyobservable phenotype. This feature is to be distinguished from expressivity, which refers to the variable nature of theclinical phenotype, such as the severity. Thus, by definition, to have expressivity, the trait must be penetrant.Numerous genetic and environmental factors can affect expression of a gene, making it nearly impossible todetermine which factor is most important in a specific individual or disease. Table 82–1 shows these factors.

Patterns of Inheritance

Table 82–1. Factors Affecting the Phenotype in Genetic Disorders

1. Causal genes and mutations

2. Modifier genes (genetic background)

3. Age

4. Sex

5. Exogenous or environmental factors such as exercise or diet

6. Maternal factors

7. Epigenetic alterations (such as DNA methylation)

8. Posttranscriptional and posttranslation modifications

9. Gene–gene (epistasis) and gene–environmental interactions

10. MicroRNAs




Inherited disorders caused by a single abnormal gene are transmitted to offspring in a predictable fashion, termedmendelian transmission. As previously noted, each individual has two copies of each gene, referred to as alleles, onetransmitted from each parent. Per Mendel's first law, each of the two alleles, located on separate chromosomes,segregates independently and is transmitted unchanged to offspring. Thus the chance of inheriting the mother's alleleversus the father's is 50%. Mendel's second law states that genes on the same chromosome also assert themselvesindependently through the process of crossover between segments of chromosomes (discussed below). The greaterthe distance between two loci, the more likely they are to be separated during genetic transmission. Mutant geneslocated on any of the 22 autosomal pairs or the 2 sex chromosomes may produce phenotypes inherited by simplepatterns classified as autosomal (dominant or recessive) or X-linked, respectively. The terms dominant inheritanceand recessive inheritance refer to characteristics of the phenotype. Dominant inheritance implies that a person withone copy of a mutant allele and one copy of the normal allele develops the phenotype associated with the mutantallele. Recessive traits, on the other hand, require both alleles to be mutant in order to produce a phenotype.

AUTOSOMAL-DOMINANT INHERITANCEDominant disorders are those exhibiting a phenotype in heterozygous individuals, as noted. Males and females areequally affected, and offspring of an affected heterozygote have a 50% chance of inheriting the mutant allele. In asporadic case, the mutation occurs de novo and in one of the germ lines of parents (typically sperm). By definition, itis absent in the somatic cells of parents. Autosomal-dominant inheritance can be misdiagnosed as sporadic if there islow expressivity in the phenotypically normal parent carrying the mutant allele or if extramarital paternity hasoccurred. Table 82–2 lists the features characteristic of autosomal-dominant inheritance.

Table 82–2. Characteristic Features of Patterns of Inheritance

A. Autosomal-dominant transmission

1. Each affected individual has an affected parent unless the disease occurred because of a new mutation or theheterozygous parent has low expressivity.

2. Equal proportions (ie, 50-50) of normal and affected offspring are likely to be born to an affected individual.

3. Normal children of an affected individual bear only normal offspring.

4. Equal proportions of males and females are affected.

5. Both sexes are equally likely to transmit the abnormal allele to male and female offspring, and male-to-maletransmission occurs.

6. Vertical transmission through successive generations occurs.

7. Delayed age of onset.

8. Variable clinical expression.

B. Autosomal-recessive transmission

1. Parents are clinically normal (in alternate generations) but genetically are heterozygotes.

2. Alternate generations are affected, with no vertical transmission.

3. Both sexes are affected with equal frequency.

4. Each offspring of heterozygous carriers has a 25% chance of being affected, a 50% chance of being anunaffected carrier, and a 25% chance of inheriting only normal alleles.

C. X-linked transmission

1. No male-to-male transmission.

2. All daughters of affected males are carriers.

3. Sons of carrier females have a 50% risk of being affected, and daughters have a 50% chance of being carriers.

4. Affected homozygous females occur only when an affected male and carrier female have children.

5. The pedigree pattern in X-linked recessive traits tends to be oblique because of the occurrence of the trait in thesons of normal carriers but not in the sisters of affected males (ie, uncles and nephews affected).

D. Mitochondrial transmission

1. Equal frequency and severity of disease for each sex.

2. Transmission through females only, with offspring of affected males being unaffected.

3. All offspring of affected females may be affected.

4. Extreme variability of expression of disease within a family (may include apparent nonpenetrance).

5. Phenotype may be age-dependent.




AUTOSOMAL-RECESSIVE INHERITANCEAutosomal-recessive phenotypes are clinically apparent when the patient carries two mutant alleles (ie, ishomozygous) at the locus responsible for the disease (Fig. 82–1). The disease-causing gene is found on 1 of the 22autosomes. Thus both males and females are equally affected. Clinical uniformity is typical, and disease onsetgenerally occurs early in life. Recessive disorders are more commonly diagnosed in childhood than are dominantdiseases. On average, only one in four children (25%) will be affected (see Table 82–2).

X-LINKED INHERITANCEX-linked inherited disorders are caused by defects in genes located on the X chromosome. Because females have 2 Xchromosomes, they may carry either one mutant allele (heterozygote) or two mutant alleles (homozygote). The traitmay therefore display dominant or recessive expression. Males have a single X chromosome (and one Ychromosome). Consequently, a male is expected to display the full syndrome whenever he inherits the abnormalgene from his mother. Hence the terms X-linked dominant and X-linked recessive apply only to the expression of thegene in females. Because a male must pass on his Y chromosome to all male offspring, he cannot pass on a mutant Xallele to his sons. Therefore, no male-to-male transmission in X-linked disorders can occur. On the other hand, amale must contribute his one X chromosome to all daughters (see Fig. 82–1). All females receiving a mutant Xchromosome are known as carriers, and those who become affected clinically with the disease are known asmanifesting female carriers. Table 82–2 lists the characteristic features of X-linked inheritance. Examples of X-linkeddisorders of the heart include X-linked cardiomyopathy, Barth syndrome, and Duchenne, Becker, and Emery-Dreifussmuscular dystrophies.

MITOCHONDRIAL INHERITANCESpermatocytes contribute few or no mitochondria to the zygote. The entire mitochondrial DNA in an embryo isderived from the mitochondria already present in the cytoplasm of the oocyte. Thus phenotypes caused bymitochondrial DNA mutations demonstrate only maternal inheritance (see Fig. 82–1). Table 82–2 lists thecharacteristic features of mitochondrial inheritance.

6. Organ mosaicism is common.

Figure 82–1.

This typical set of pedigrees outlines the usual inheritance patterns for autosomal-dominant and autosomal-recessive traits,X-linked inheritance, and mitochondrial inheritance. Squares signify males; circles signify females. Filled-in circles andsquares are affected females and males, respectively.




OVERVIEW OF GENE MAPPING AND MUTATION DETECTION

Chromosomal Mapping in Single-Gene DisordersUntil the 1980s, identification of a disease-causing gene without knowing the causal protein was nearly impossible.For the majority of diseases, neither the defect nor the protein was known. Technical advances that madechromosomal mapping feasible include (1) computerized linkage analysis, (2) development of highly informative DNAmarkers spanning the entire genome, and (3) detection of markers by polymerase chain reaction (PCR). The 46chromosomes of the human genome contain 3.2 billion bp of DNA. To locate a particular gene, one must first map thechromosomal locus, which requires knowledge of certain chromosomal landmarks, referred to as DNA markers. ADNA marker is a polymorphic sequence of DNA with a known chromosomal position, which can be detected byanalyzing an individual's DNA (discussed in detail below). Initial genetic linkage studies were performed using short-tandem repeat (STR) DNA markers. STR markers are available that span each chromosome at intervals of not morethan 4 million base pairs (Mbp) on all chromosomes (a set of approximately 800 markers). In recent years, SNPshave been used for genetic linkage as well. The typical SNP linkage panel contains several thousands SNPs that arepositioned throughout the genome, with an average distance of approximately 0.5 Mbp. Genetic distance is measuredin terms of centimorgans (cM), named after the geneticist T.H. Morgan. One cM approximates 1 Mbp. DNA markerslike genes have two alleles in a given individual and are transmitted to offspring according to Mendel's law, with theindividual being heterozygous or homozygous for that marker. If a marker is homozygous, it is not informative forgenetic linkage. When all of the markers are placed together on each chromosome and the genetic distance betweenthem is estimated, a genetic map is produced. Several maps of more than 5000 highly informative DNA markers that

span the entire genome have been developed.9

Identification of a particular locus is made possible by showing that the causal gene of interest is in close proximity toa DNA marker on the same chromosome, a method referred to as genetic linkage analysis. A fundamentalrequirement for linkage analysis is a family in which the disease of interest is transmitted to offspring over at leasttwo and preferably three generations. At least six affected individuals are required for analyzing cosegregation ofDNA markers with inheritance of the disease, although a larger number of affected individuals is preferable.

The homologous pairs of chromosomes are assorted, and one from each parent is transmitted to the offspring bychance. Each gene, allele, or marker is transmitted independently. Thus the odds of any two genes (or a marker anda gene) being coinherited is 50% (chance alone). Even genes on the same chromosome are transmittedindependently by the mechanism of crossover between homologous chromosomes (Fig. 82–2), unless they are inclose physical proximity to each other. In the latter case, they cosegregate together. Homologous recombinationprovides for continual mixing of the genes during every meiosis. It is the predominant reason why no two individualshave the same genotype for all DNA markers unless they are identical twins. Before meiosis, the two homologouschromosomes come together and form bridges (chiasmata) such that segments of equal proportion are exchangedbetween them, giving rise to crossover between homologous regions of various genes. In genetic parlance, crossingover is referred to as recombination. The loci occupy the same chromosomal position on the homologouschromosome on which they are combined as they had on their original homologous chromosome. There is no net lossof chromosomal material or genes, but crossover leads to a constant intermixing of the chromosomes such that notwo offspring will ever be identical. Crossovers occur only between homologous chromosomes. On average, there are33 crossovers between homologous chromosome pairs per meiosis.

Figure 82–2.




FAMILY HISTORY AND EVALUATIONThe most important part of an evaluation for genetic disease is the family history. First, this may give clues to thediagnosis of a particular phenotype and inheritance patterns within an individual family. For instance, an individual'sethnic background may suggest the need for specific types of genetic screening, such as for hemoglobinopathies inindividuals of African or Mediterranean ancestry or for Tay-Sachs disease in individuals of eastern European(Ashkenazi) Jewish ancestry. The individual with the medical problem who brought the family to the attention of thephysician is referred to as the proband or propositus (proposita for females) or index case. Information shouldgenerally be collected on all individuals who are first-, second-, or third-degree relatives of the proband. First-degreerelatives of the proband are the parents and children. Second-degree relatives are aunts and uncles, grandparents,and grandchildren of the proband. Third-degree relatives are first cousins, great aunts and uncles, great-grandparents, and great-grandchildren. A pedigree chart (see Fig. 82–1) is then generated. This information shouldinclude medical problems and pregnancies. If relatives are deceased, the age at death and the cause of death shouldbe recorded. With a pedigree chart and specific family information, general questions are asked, including whetherother family members have the same or similar problems. Information about various types of birth defects, mentalretardation, early infant deaths, miscarriages, stillbirths, or other diseases or handicaps in the family is sought. Withsome disorders, there may be a variability of a particular condition (ie, clinical heterogeneity), even within a family.For example, with a possible diagnosis of HCM, one should ask about premature death or syncope. A pregnancyhistory may provide information to support a possible teratogenic exposure. The date of the last menstrual period,whether the pregnancy was planned, whether contraception was used immediately before pregnancy, the time whenthe pregnancy was recognized, and when the mother sought prenatal care should be noted. Problems during thepregnancy'such as bleeding, spotting, cramping, fevers, rashes, or illnesses; drug exposures (both prescribed andnonprescribed), alcohol intake, or "recreational" drug use; and exposures to potent chemicals in the workplace orwhile involved in various hobbies'should be explored. Pregnancy and family histories can then be used in conjunctionwith the findings on physical examination to derive a potential etiologic diagnosis and to plan for further diagnosticstudies. The term etiologic diagnosis should suggest whether a specific cardiac defect is familial (by family history),genetic but not familial (sporadic), teratogenic (by pregnancy history), or multifactorial. Prognosis and recurrence riskare linked strongly to an accurate diagnosis and its probable etiology. In sum, accurate phenotypic characterization isessential for all genetic studies.

CONCEPT OF GENETIC LINKAGE ANALYSIS

Linkage analysis. Loci A (disease locus) and B (DNA marker locus) are located in close proximity, with minimal chance ofcrossover between them. Thus even when crossover occurs between homologous segments of chromosomes during meiosis,A and B loci cosegregate together and thus are considered genetically linked.




Despite the independent assortment of chromosomes and genes during meiosis, genes (alleles) on two or more lociare often coinherited because they are so close together that a chiasmatic bridge does not form between them. Twoloci coinherited more than 50% of the time are considered genetically linked. To map the chromosomal locusresponsible for a causal gene, DNA markers that are evenly distributed across the chromosomes are selected. DNA iscollected from all members of a family (normal and affected) and genotyped for the selected markers. If a DNAmarker is coinherited with the phenotype in the affected individuals, the chromosomal locus where the DNA markerresides is in close physical proximity to the locus of the causal gene. This is referred to as genetic linkage betweenthe disease (causal gene) and the DNA marker. Figure 82–2 illustrates the concept of linkage analysis. Shown in theleft panel is an illustration of genetic linkage between a locus for a DNA marker and that of a disease that is inheritedin a mendelian dominant fashion. The locus, designated with an "A," carries the allele responsible for the disease. Thecorresponding locus, "a," on the homologous chromosome has the allele that codes for the same protein but has notundergone a mutation and is thus the normal allele. The loci designated "B" and "b" represents alleles of a DNAmarker of known location that has nothing to do with the disease. In the right panel, the disease and the marker lociare so close that they tend to be coinherited within the family. In contrast, in the left panel, the "A" and "b" loci areso far apart that recombination and crossover occur between the two markers, and thus they segregateindependently. The calculation necessary to prove definitively that genetic linkage does or does not exist between aDNA marker and a disease-related locus is sophisticated and requires advanced computer programs. The odds for andagainst linkage are calculated. Linkage exists if the odds in favor of linkage are at least 1000:1. Commonly, thelogarithm of the odds, referred to as the LOD score (log of the odds), is used, and a LOD score of 3 indicateslinkage. A LOD score of –2 (ie, 102 or 100:1 odds against linkage) excludes the linkage. The likelihood of two genesbeing separated by recombination increases in proportion to the distance between them. The distance between amarker and a disease-causing gene when genetically linked is quite variable and may be anywhere from 1 to 50 Mbpbut is usually within 1 to 10 Mbp. Thus the inherent resolution of genetic linkage analysis is not better than 1 Mbp.

It is possible on the basis of linkage analysis alone to construct a chromosomal map of all of the DNA markers, withthe distance between the various markers estimated in centimorgans. This is a complex calculation derived from thenumber of recombinations between the DNA markers during meioses. The recombination frequency between twomarkers, two genes, or a gene and a marker is the ratio of the number of crossover events to the total number ofmeioses. The lower the recombination frequency between the locus of a DNA marker and that of a disease-causinggene, then the closer those two are in physical distance on the chromosome. However, despite the close physicalproximity of the loci of the DNA marker and the disease-causing gene, recombination still may occur. The extent towhich recombination does occur reflects roughly the physical distance between the two loci. The recombinationfraction (or theta) is used to develop a means of estimating the genetic distance (in centimorgans) betweengenetically linked loci. A recombination frequency or crossover of 1% between two loci, whether occupied by twogenes or one gene and a DNA marker, reflects a physical distance of approximately 1 cM between them. For a markerand a gene separated by 1 cM, this means the chance of a crossover between them during meiosis is only 1%; thusthe chance of their being coinherited is 99%. This is a statistically derived genetic map, however, and the distancesare only approximate.

IDENTIFICATION OF THE GENE AND CAUSATIVE MUTATIONOnce the chromosomal location of a gene has been mapped, the first technique in attempting to identify the gene isreferred to as the positional candidate gene approach. There are approximately 230,000 genes in the genome,however, more than 100,000 expressed sequenced tags (ESTs) have been mapped. These ESTs are unique DNAsequences of 100 to 200 bp, each of which is claimed to represent a portion of the expressed sequences of a gene.These genes and ESTs are available through a worldwide network of databases.

Known genes or ESTs in the mapped chromosomal region are amplified as candidate genes that contain the causativemutation that segregates with the disease. If none of the candidate genes in the region are shown to have a mutationthat cosegregates with the disease, it may be necessary to clone the region. This approach is referred to as positionalcloning, so named because a region is cloned knowing only its position relative to the genetically linked DNA markers.Positional cloning is usually unnecessary, as most genes in the human genome have been mapped and identified.However, if attempted, it is necessary to reduce the region (containing the gene) to 1 cM. It is often necessary toexpand the family with the hope of finding crossovers such that DNA markers common to all affected would span onlya short distance (<1 cM). The cloned genes or PCR-amplified DNA is then analyzed, commonly by direct sequencing,for the presence of the mutation. To strengthen the causality, the mutation must be shown to cosegregate with thedisease and not with the unaffected members in the family. In addition, it is crucial to show that the variant is absentin large number of normal individuals with the same ethnic background, and hence it is not a polymorphism. Finally,to establish the causality, in vivo and in vitro functional studies are necessary. Table 82–3 summarizes the approachto chromosomal mapping of hereditary diseases by linkage analysis and subsequent isolation of the gene. Mapping of

genes predisposing to polygenic disorders10-12 is discussed under Genetics of Coronary Artery Disease.




Table 82–3. Chromosomal Mapping and Gene Identification in Single-Gene Disorders

1. Identification of a family with a familial disease.

2. Collection of clinical data from the family.

3. Clinical assessment to provide an accurate diagnosis of the disease using a consistent and objective criterion toseparate normal individuals from those affected and from those who are indeterminate or unknown.

4. Collection of blood samples for immediate DNA analysis and development of lymphoblastoid cell lines for arenewable source of DNA.

5. Development of a family pedigree.

6. DNA analysis for markers of known chromosomal loci that span the human genome in an attempt to find amarker locus linked to the disease.

7. Identification of the gene.

8. Identification of mutation(s) causing the disease.

9. Demonstration of a causal relationship between the mutant gene and the disease.

10. Development of a convenient test to screen for the mutation.

GENETIC COUNSELING PRINCIPLES

Genetic counseling should provide information about the diagnosis, possible etiology, and prognosis of a disease. Inaddition, psychosocial issues, reproductive options, and the availability of prenatal diagnosis should be discussed.Genetic counseling should be nondirective, providing information in a nonjudgmental, unbiased manner. The familyshould then be able to make decisions based on medical information in the context of their religious, moral, cultural,and social backgrounds and their financial situation. Although a genetic counselor may occasionally feel frustratedwith a specific couple's decision, an effective counselor does not let personal biases interfere with the counseling role.Conflicts leading to major ethical issues and disputes may arise, however, and may be particularly apparent regardingissues of nonpaternity, sex selection, pregnancy termination, and selective nontreatment of malformed infants.Couples have many potential reproductive options, but not all may be acceptable religiously or culturally.Nevertheless, potential options should be mentioned in a sensitive manner. A common misunderstanding amongfamilies in genetic counseling is the issue of prenatal diagnosis and its relationship to abortion. Prenatal diagnosisdoes not imply that a parent should or would terminate the pregnancy. In many circumstances the information fromprenatal diagnosis may help to reassure a couple that their risk of having another handicapped child is, in fact, muchlower than expected. Conversely, if defects are found, the subspecialist may use more diagnostic approaches to makerational decisions about medical management of the infant before or immediately after delivery.

The accelerated pace of progress in gene discovery, molecular medicine, and molecular diagnostics has begun toallow for improved genetic counseling and portends the possibility of future genetic therapy. As knowledge about thegenetic basis of disease grows, however, so does the potential for discriminatory health insurance policies to excludeindividuals who are at risk for an illness or to charge prohibitively high rates on the basis of predetermined illness.For this reason planners of the Human Genome Project recognized the need to protect individuals who volunteer forgenetic study as well as those diagnosed by molecular methods in the future. Also for this reason, the NationalInstitutes of Health–Department of Energy Working Group on Ethical, Legal, and Social Implications of the HumanGenome Project was developed. After 13 years of debate in Congress, on May 21, 2008, President Bush signed intolaw the Genetic Information Nondiscrimination Act. The bill protects Americans against discrimination based ongenetic information for hiring or insurance.

CARDIOVASCULAR ABNORMALITIES CAUSED BY CHROMOSOMAL DEFECTS

Table 82–4 lists chromosomal defects that cause cardiovascular abnormalities. The most common chromosomaldefects are described briefly below.

Table 82–4. Partial List of Chromosomal Abnormalities Associated with Heart Disease

Chromosome Defects Syndromes Cardiac Phenotype

45X Turner syndrome Coarctation of the aorta, ASD, aorticstenosis

Trisomy 5 Interrupted aortic arch

Trisomy 13 Patau syndrome CHD, VSD

Trisomy 18 Edwards syndrome CHD, VSD




Turner SyndromeTurner syndrome is characterized by a constellation of findings that result from partial or complete monosomy of the

X chromosome.13 It is the most common chromosomal abnormality in females, with an incidence of 1 per 2500 to3000 liveborn girls, which corresponds to approximately 2 million cases worldwide.14 It is characterized bycardiovascular anomalies, short stature, low-set ears, excess nuchal skin, broad chest with widely spaced nipples,peripheral lymphedema, and ovarian dysgenesis. Cardiac abnormalities are common, with a prevalence estimated to

be between 23% and 40%.15 The most common cardiovascular abnormalities are bicuspid aortic valve, which ispresent in 10% to 20%, and coarctation of aorta, present in 10% of adult cases. The prevalence of theseabnormalities is higher in children. Less common cardiovascular anomalies include aortic stenosis, systemichypertension, mitral valve prolapse, conduction defects, partial anomalous venous drainage, and ventricular septaldefect (VSD). Aortic dilatation and dissection, partly because of concomitant hypertension, also occur (see also Chap.

106). Women with Turner syndrome are more susceptible to aortic aneurysms and ischemic heart disease.14,15

Patients with Turner syndrome should undergo periodic cardiovascular evaluation, including 12-lead ECG andechocardiography.

Turner syndrome is caused by complete or partial absence of an X chromosome. The most common karyotype is

monosomy X (45,X).13 Approximately 5% to 10% of cases have duplication of the long arm of 1 X (46,X,i[Xq]) andthe rest have mosaicism.13 The pathogenesis of Turner syndrome is not fully understood. It likely entailshaploinsufficiency of genes (located on the X chromosome) that, under normal conditions, escape inactivation.Inactivation of 1 copy of the X chromosome during early embryogenesis is partial, and several genes escapeinactivation. Specific genes that account for cardiovascular phenotype in Turner syndrome are unknown. SHOX (shortstature homeobox-containing gene) or PHOG (pseudoautosomal homeobox-containing osteogenic gene), whichencode two isoforms of a homeodomain protein, are considered responsible for the short stature in Turner

syndrome.14 Zinc finger protein X and zinc finger protein Y genes (ZFX/ZFY), which are involved in sexdetermination, are also candidate genes for Turner syndrome.16

Down SyndromeDown syndrome, or trisomy 21, is a major cause of mental retardation and congenital heart disease, with acharacteristic set of facial and physical features. The incidence of Down syndrome is approximately 1 in 700

livebirths, affecting more than 350,000 individuals in the United States alone.17 The risk of having a live-born infantwith Down syndrome increases with maternal age. It is estimated at 1 in 1000 at age 30 years and 10-fold higher at

age 45 years.18 The recurrence rate in the offspring is approximately 1%. Clinical manifestations include congenitalanomalies of the heart and gastrointestinal tract, epicanthal folds, flattened facial profile, small and rounded ears, up-slanted palpebral fissures, excess nuchal skin, and brachycephaly. An increased risk of leukemia, immune systemdefects, and an Alzheimer-like dementia are associated with Down syndrome. Cardiac abnormalities are present in

approximately half of the cases.19,20 The most common cardiac abnormalities are atrioventricular canal defect andisolated VSD, which occur in 45% and 35% of cases, respectively.19,20 Isolated secundum atrial septal defect (ASD)is present in 8% and tetralogy of Fallot in 5% of cases (see also Chap. 9).19

Partial trisomy 20q Dextrocardia

Trisomy 21 Down syndrome CHD, ASD, VSD, PDA

Trisomy 22 VSD

Partial tetrasomy 22 Schmid-Fraccaro syndrome CHD

Anomalous pulmonary venous return

Deletion 4p Wolf-Hirschhorn syndrome CHD

Deletion 7q11.23 Williams syndrome CHD, supravalvular aortic stenosis,hypertension, MVP

Deletion paternal 15q11 Prader-Willi syndrome CHD

Deletion 17p Miller Dieker syndrome CHD, ASD

Deletion 22q11 CATCH-22, DiGeorge, and velocardiofacialsyndromes

CHD

Rearrangement 5p15.1-3 Cri du chat CHD

Recombinationchromosome 8

San Luis Valley syndrome Tetralogy of Fallot

ASD, atrial septal defect; CHD, congenital heart disease; MVP, mitral valve prolapse; PDA, patent ductus arteriosus;VSD, ventricular septal defect.

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Down syndrome is caused by trisomy 21. It is full trisomy in 95%, chromosomal translocation in 2%, and mosaic in

3%.21,22 The vast majority of errors in meiosis leading to trisomy 21 are of maternal origin and occur during the firstmeiosis in two-thirds and during second meiosis in one-fifth of the cases. The exact causal genes responsible for the

cardiovascular defects are unknown. However, four Down syndrome critical regions (DSCRs) have been mapped.22-25

DSCR1 encompasses an area of approximately 5 million bp and approximately 20 genes.22 DNA markers in thisregion are associated with mental retardation, and most of the facial features of the syndrome.22 Among thecandidate genes is DSCR1, which is abundantly expressed in the heart and brain.26 It is a candidate for cardiacanomalies and mental retardation.26 High-resolution map of the phenotype of congenital heart defects in patientswith the Down syndrome locus restricts the region to a 1.7-Mbp interval, which contains 10 genes. Among thecandidates in this region is DSCAM, which encodes Down syndrome cell adhesion molecule. It is the only gene in this

region that is expressed in the heart.27 The pathogenesis of Down syndrome is unknown. Down syndrome isconsidered a contiguous gene syndrome.17 It is expected to involve increased expression of multiple contiguousgenes. Overexpression of DSCR1, a product of DSCR, is shown in the brains of patients with Down

syndrome.28DSCR1 encodes calcipressin 1, which functions through direct binding and inhibition of calcineurin A, thecatalytic subunit of the Ca2+/calmodulin-dependent serine threonine protein phosphatase (PP2B).29 Calcineurindephosphorylates nuclear factor of activated T cells (NFAT), which leads to its nuclear localization and induction of

gene expression.29 Inhibition of calcineurin by DSCR1 is expected to increase levels of phosphorylated NFAT andreduce nuclear localization of NFAT and NFAT-mediated gene expression. Inhibition of calcineurin by DSCR1 may be

one of the multiple mechanisms involved in the pathogenesis of Down syndrome.30

Edwards SyndromeEdwards syndrome, or trisomy 18, is the second most common trisomy, with a prevalence of approximately 1 in 4000

to 8000 livebirths.31 The majority of the infants die within a couple of weeks and approximately 10% survive morethan a year.32 The syndrome is characterized by anomalies of the heart and microcephaly with a prominent occiput, anarrow forehead, low-set and malformed ears, micrognathia, clefting of the lip and palate, clenched hand with

overlapping digits, rocker-bottom feet, and various hernias.21 Cardiovascular anomalies are present in approximately90% of cases. They include VSD, ASD, patent ductus arteriosus (PDA), pulmonary stenosis, tetralogy of Fallot,

transposition of the great arteries, bicuspid aortic valve, dysplastic valves, and coarctation of the aorta.19,21 Fulltrisomy occurs in more than 85%, chromosomal translocation in 3%, and mosaicism in 5% of cases.21 The causalgene(s) for the cardiovascular anomalies remains unknown.

Patau SyndromePatau syndrome, or trisomy 13, is a rare disorder with an incidence of 1 per 5000 to 1 in 20,000 livebirths and a high

early mortality.21 Approximately 50% of affected infants die within the first month and 85% within first year of life.21

Patau syndrome is characterized by cardiac, urogenital, craniofacial, and central nervous system anomalies. Specificanomalies include microcephaly with sloping forehead, microphthalmia, cleft lip and palate, overlapping fingers withpostaxial polydactyly, and renal abnormalities, including polycystic kidney disease. Cardiac abnormalities are presentin approximately 80% of the cases. They include VSD, ASD, PDA, pulmonary stenosis, coarctation of the aorta,

dextrocardia, and truncus arteriosus.19,21

Patau syndrome is caused by nondisjunction of chromosome 13 during meiosis in the vast majority of cases andrarely by translocation. Five percent of the cases are mosaic. The causal genes for cardiovascular anomalies intrisomy 13 are unknown.

DiGeorge (Catch-22) and Velocardiofacial SyndromesDiGeorge and velocardiofacial syndromes are autosomal-dominant congenital anomalies caused by hemizygousmicrodeletion of a large segment of the long arm of chromosome 22 (22q11). The deletion leads to anomalies ofmultiple organs, including the heart and facial bones. The prevalence is approximately 1 in 4000, accounting for

approximately 15% of all congenital heart defects.32,33 The term CATCH-22 denotes cardiac, abnormal facies, thymichypoplasia, cleft palate, hypocalcemia (as a result of parathyroid hypoplasia), and the 22nd chromosome. Cardiacanomalies are present in approximately two-thirds of cases. A diverse array of congenital heart defects, includingtetralogy of Fallot, interrupted aortic arch, truncus arteriosus, and PDA, have been described. Tetralogy of Fallot is

the most common abnormality.32 Patients with velocardiofacial syndromes exhibit craniofacial anomalies, cleft palate,and a variety of cardiac abnormalities, such as aortic arch anomalies, tetralogy of Fallot, and VSD. Cardiac valves andthe myocardium are usually spared.

DiGeorge syndrome is caused by microdeletion of approximately 3 Mbp of DNA encompassing approximately 30

genes.32 Genetic analysis in mouse and mutation analysis of the candidate genes in the region have led toidentification of mutations in TBX1, which encodes a T-box transcription factor.34 TBX1 is critical for embryogenesisof aortic and pulmonary outflow tracts. Loss-of-function mutations in TBX1 result in haploinsufficiency. The




downstream target genes of TBX1 and the pathways involved in the pathogenesis of cardiac phenotype are mostlyunknown.

Another candidate gene is UFD1L, which encodes a protein involved in degradation of ubiquitinated proteins. It isexpressed during the embryogenesis of cell lines typically associated with DiGeorge syndrome. Similarly, UFD1L isexpressed in association with the conotruncus and the fourth embryologic aortic arch. A large deletion in humantranscription factor UFD1L in a single patient with a phenotype similar to that of DiGeorge syndrome has been

identified.35 Deletion of Ufd1L in mice produced some of the typical cardiac phenotypes that result from defectivedevelopment of the fourth branchial arch.36 However, several other studies have excluded mutations in UFD1L inpatients with DiGeorge syndrome.37 Other genes, such as ZNF74, which encodes a zinc-finger transcription factor,also have been implicated in the pathogenesis of DiGeorge syndrome.38 However, the causal role remains to beestablished.

GENETIC BASIS OF SPECIFIC CONGENITAL HEART DISEASES

A significant number of congenital heart diseases occur in isolation and are not part of complex phenotypes asobserved in chromosomal abnormalities. Recently, the causal genes for several congenital heart diseases have beenidentified. Preliminary studies depict a common theme in the pathogenesis of isolated congenital heart defects, whichimplicate deficiency of several transcriptional factors that regulate cardiac gene expression during embryogenesis.However, there is considerable phenotypic, locus, and allelic heterogeneity.

Supravalvular Aortic Stenosis

Supravalvular aortic stenosis39 is an autosomal-dominant disease characterized by discrete narrowing of theascending aorta above the level of the sinus of Valsalva. It commonly occurs as a phenotype of Williams syndrome(or Williams-Beuren syndrome) in conjunction with mental retardation in some, and exceptional talents in others,hypercalcemia, characteristic facial appearance, and stenosis of other major arteries. The prevalence of supravalvularaortic stenosis is estimated to be 1 in 25,000 livebirths.

The gene responsible for supravalvular aortic stenosis was initially mapped to chromosome 7q11.23 and

subsequently identified as ELN, encoding elastin.41 Almost all cases of isolated supravalvular aortic stenosis arecaused by ELN mutations, which comprise a variety of point and deletion mutations.40 Mutations result in elastindeficiency, which in the vascular system leads to inelasticity of the vessel wall and subsequent fibrosis as a result ofan altered stress–strain relation (elastin arteriopathy). Thus haploinsufficiency underlies the pathogenesis ofsupravalvular aortic stenosis.

Patients with Williams syndrome may exhibit additional cardiovascular phenotypes, including pulmonary arterial

stenosis, aortic and mitral valve abnormalities, and tetralogy of Fallot.41 In 98% of cases of Williams syndrome, thedeletion mutation includes 1.5 Mbp of DNA comprising ELN and another 20 contiguous genes. Contribution of thesegenes to pathogenesis of specific phenotypes in Williams syndrome remains unknown. The genetic basis of William-

Beuren syndrome has been reviewed in detail in Schubert.42

Familial Atrial Septal Defect

ASD is among the most common congenital heart diseases, with an estimated incidence of 1 in 1000 livebirths.1 ASDis usually sporadic. However, familial ASD with an autosomal-dominant mode of inheritance has also been

described.43,44 Individuals with ASD are commonly asymptomatic until the third or fourth decades. Commonsymptoms are palpitations, commonly caused by supraventricular arrhythmias, and symptoms associated withpulmonary hypertension and right-sided volume overload resulting in left-to-right shunt. Uncorrected ASD can lead toheart failure and premature death in the fourth or fifth decade of life.

The first gene identified for familial ASD is NKX2–5 (CSX1), which is the human homologue of Nkx2.5 in mouse and

tinman in Drosophila melanogaster.45 The gene is located on 5q35 and encodes NKX2.5, a predominantly cardiac-specific transcription factor that regulates expression of several cardiac genes.46 A multiplicity of mutations has beendescribed in patients with secundum ASD and conduction defects.45,47 Mutations often result in haploinsufficiency.Point mutations in the DNA-binding domain reduce the affinity of NKX2.5 for the promoter regions and hence

decrease expression of cardiac-specific genes.48 The spectrum of clinical phenotypes caused by mutations in NKX2.5extends beyond secundum ASD and comprises VSDs, tetralogy of Fallot, subvalvular aortic stenosis, pulmonary

atresia, and others.49

The second causal gene for familial ASD with an autosomal-dominant mode of inheritance is GATA4 on chromosome

8p22-23.50 The mutations diminish DNA-binding affinity and transcriptional activity of GATA4 transcription factor andblock its physical interaction with TBX5, another transcription factor involved in the pathogenesis of congenital heart

disease.50




The third causal gene for familial ASD is MYH6, which is located in chromosome 14q12 and encodes myosin heavy

chain 6.51 A missense mutation in MYH6 in a family with atrial fibrillation mapped to 14q12 locus has beenidentified.51 The MYH6 protein is expressed at high levels in atrial tissues and plays an important role in formation ofinteratrial septum.51

Holt-Oram SyndromeHolt-Oram syndrome is a rare autosomal-dominant inherited disorder characterized by anomalies of the heart and

upper extremities, hence the name hand–heart syndrome (see also Chap. 84).52 The most common congenital heartdefects are ASD and VSD, followed by conduction system abnormalities and atrial fibrillation.53 Less common cardiacabnormalities include truncus arteriosus, mitral valve defect, PDA, and tetralogy of Fallot.53 Anomalies of the upperlimb vary from mild malformation of the carpal bones to phocomelia, but upper limb preaxial radial abnormalities are

commonly present.53

Mutations in TBX5 on chromosome 12q24, which codes for transcription factor TBX5, are responsible for the cardiac

and skeletal abnormalities in Holt-Oram syndrome.53,54 A number of mutations have been described, and most arenonsense, frameshift, or splice-junction abnormalities. The proposed molecular mechanism is haploinsufficiency,resulting in reduced expression level of TBX5. Haploinsufficiency because of truncation or frameshift mutations resultsin severe birth defects in the heart and hands, whereas point mutations predominantly affect either hand or heart

development.55 Mutations in the 5' end of the gene exhibit a preponderance of cardiac abnormalities with mildskeletal abnormalities, and those in the 3' end lead to severe skeletal and mild cardiac abnormalities.

Ellis–Van Creveld SyndromeEllis–van Creveld syndrome is an autosomal-recessive skeletal dysplasia, which is associated with congenital heartdisease in the majority of cases. Skeletal anomalies include short limbs, short ribs, postaxial polydactyly, anddysplastic nails and teeth. ASD and common atrium are the typical cardiac anomalies present in two-thirds of thecases (see also Chap. 84).

The gene responsible for Ellis–van Creveld syndrome was mapped to chromosome 4p16.156 near an area proximal tothe FGFR3 gene, which is known to cause hypochondroplasia and achondroplasia. Subsequently, splice donor,

truncation, and missense mutations in a novel gene, EVC, were identified.57 Mutations in EVC account forapproximately 20% of the cases of Ellis–van Creveld syndrome.58 Recently, mutations in a second gene, namedEVC2, for Ellis–van Creveld syndrome, were identified.58 The pathogenesis of Ellis–van Creveld syndrome remainsunknown.

Familial Patent Ductus Arteriosus or Char SyndromePDA can occur as a sole cardiac anomaly or in conjunction with other congenital heart disease. Familial PDA with anautosomal-dominant inheritance has been described in patients with Char syndrome. Char syndrome is a congenitaldisease that was first described by Florence Char in 1978 and is characterized by a constellation of facialdysmorphism, fifth-finger middle phalangeal hypoplasia, and PDA. Variation of this syndrome is associated withbicuspid aortic valve, distinctive facial appearance, polydactyly, and fifth-finger clinodactyly. The predominant clinicalfeatures are those of PDA, which include symptoms and signs of left-heart failure and pulmonary hypertension.

The gene responsible for Char syndrome in two families was recently mapped to chromosome 6p12-21.59

Subsequently, mutations in TFAP2B, which encodes a neural crest-related helix-span-helix transcription factor, were

identified.60 These findings suggest that Char syndrome results from derangement of neural crest-cell derivatives.60

A second locus for familial PDA has been mapped to a 3-cM interval on chromosome 12q24.61 The causal generemains unknown.

Noonan and Leopard SyndromesNoonan syndrome is an uncommon autosomal-dominant disorder characterized by dysmorphic facial features, HCM,

pulmonic stenosis, mental retardation, and bleeding disorders.62 LEOPARD syndrome (lentigines, electrocardiographicconduction abnormalities, ocular hypertelorism, pulmonic stenosis, abnormal genitalia, retardation of growth, and

deafness) is an allelic variant of the Noonan syndrome.62 Pulmonic stenosis and HCM are the primary cardiacphenotypes. Others include atrioventricular septal defects, aortic coarctation, ASD, mitral valve defects, PDA, andfibroelastosis. Noonan syndrome is also seen in conjunction with cardiofaciocutaneous syndrome and other congenitalabnormalities, such as neurofibromatosis (see also Chap. 9).

Noonan syndrome is sporadic in half of the cases and autosomal-dominant in the other half. The gene responsible forautosomal-dominant Noonan and LEOPARD syndromes was mapped to chromosome 12q22 and subsequently

identified as encoding protein-tyrosine-phosphatase, nonreceptor type 11 (PTPN11).63,64 With the exception ofdeletion of amino acid glycine 60, all mutations in PTPN11 are missense mutations.62 Most mutations are recurrent




and localized to exons 3, 7, 8, and 13.62 The N308D mutation is the most common and accounts for approximately25% of cases.62 Overall, mutations in PTPN11 are found in approximately two-thirds of cases of Noonan syndrome.62

Mutations are located in interacting portions of the amino-terminal src-homology 2 (N-SH2) and protein-tyrosine-

phosphatase (PTP) domains.63,64 Both gain-of-function and dominant-negative mechanisms have been implicated inthe pathogenesis of Noonan syndrome caused by PTPN11 mutations.

Mutations in KRAS, SOS1, and RAF1 also have been identified in patients with Noonan syndrome. Collectively,mutations in the above three genes account for approximately 25% of all Noonan syndrome cases.

Familial Myxoma Syndrome (Carney Complex)Myxomas are the most common cardiac tumors and are generally sporadic. Myxomas are familial, with an autosomal-

dominant mode of inheritance in approximately 10% of cases (see also Chap. 84).65 Familial myxoma commonlyoccurs as a part of Carney complex, with the constellation of cardiac myxoma, endocrine disorders, and skin

pigmentation.66 LAMB (lentigines, atrial myxoma, mucocutaneous myxoma, and blue nevi) and NAME (nevi, atrialmyxoma, myxoid neurofibromata, and ephelides) syndromes are considered variants of Carney complex. Atrial,ventricular, and skin myxomas; endocrine tumors and disorders such as Cushing syndrome; and skin lesions such aslentiginosis are part of the phenotypic expression of Carney complex. Clinical features of atrial myxoma may includefever, arthralgia, dyspnea, diastolic rumble, tumor plop, and systemic embolisms.

Carney complex exhibits locus heterogeneity, and at least two loci on chromosome 17q24 and 2p16 have been

mapped.67,68 The majority of familial cardiac myxomas (Carney complex) are caused by mutations in the PRKRA1Agene on chromosome 17q24.69 It encodes the -regulatory subunit of cyclic adenosine monophosphate (cAMP)–dependent protein kinase. Frameshift mutations in PRKRA1A result in haploinsufficiency, which suggests that thePRKRA1A functions as a tumor-suppressor gene. Recently, a missense mutation in the perinatal myosin heavy-chaingene (MYH8) was identified in members of a family with Carney complex and trismus-pseudocamptodactyly

syndrome.70

Situs InversusSitus inversus is a reversal of the asymmetric anatomic position of visceral organs. In situs inversus totalis, allvisceral organs are reversed in a mirror-image manner. It is part of the immotile cilia syndrome (primary ciliarydyskinesia). Kartagener syndrome is situs inversus, bronchiectasis, and male sterility. Most cases of situs inversusare sporadic. Autosomal-recessive, autosomal-dominant, and X-linked forms have been reported.

Situs inversus, as a component of immotile cilia syndrome, such as that in Kartagener syndrome, is caused by

mutations in dyneins.71 Dyneins are large proteins with adenosine triphosphatase (ATPase) activity that interact withintermediary filaments to produce energy and motion. Mutations in dynein axonemal intermediate chain 1 (DNAI1) onchromosome 9p13-p21, dynein axonemal heavy chain 5 (DNAH5) on chromosome 5p, and dynein axonemal heavychain type 11 (DNAH11) on chromosome 7p21 have been found in patients with primary ciliary dyskinesia (and situs

inversus).71-73

Situs inversus has also been mapped to chromosome Xq26.2. Mutations in ZIC3, encoding a zinc-finger protein of the

cerebellum, are associated with situs ambiguus in male and situs solitus or inversus in females.74 Other causal genesfor right-left axis abnormality include CFC1 on chromosome 2, LEFTB (also known as LEFTY2) and ACVR2B, encoding

activin receptor IIB.75,76

Alagille Syndrome (Arteriohepatic Dysplasia)Alagille syndrome is an autosomal-dominant disorder characterized by anomalies of the right side of the heart anddevelopmental abnormalities of eyes, skeleton, and kidney. Cardiac abnormalities are present in approximately 70%of cases; the most common is diffuse pulmonary artery stenosis. Others include hypoplastic pulmonary circulation,

pulmonary atresia, tetralogy of Fallot, coarctation of aorta, secundum ASD, PDA, and VSD.77 The most commoncausal gene is Jagged-1 gene (JAG1), located on chromosome 20p12.78,79 Deletion or point mutations in JAG1 arefound in approximately 90% of the patients with Alagille syndrome. JAG1 is a cell surface protein that is a ligand forthe Notch receptor. The Notch intercellular signaling pathway mediates cell fate decisions during development. Theproposed molecular mechanism is haploinsufficiency leading to defective cell adhesions. Recently, mutations in

NOTCH2 were found in those who did not have JAG1 mutations.80 Collectively, the findings indicate that Alagillesyndrome is disease of Notch signaling pathway.

GENETIC DISEASES OF CARDIAC MUSCLE

The term cardiomyopathy denotes an exclusive group of disorders in which the primary defect is in the myocardium,affecting cardiac myocyte structure and/or function. The primary defect, however, does not need to be exclusive tothe heart. It can also involve other tissues and organs, as in cardiomyopathies arising from metabolic disorders and




mitochondrial myopathies. Myocardial dysfunction can also occur because of systemic, infiltrative, toxic, andendocrine disorders; coronary atherosclerosis; and valvular pathologies. In such conditions, the primary defect is notin the myocardium. Thus myocardial involvement is considered secondary. In a sense, cardiac involvement in suchdisorders does not meet the pure definition of cardiomyopathy. In the broader definition, the Council on ClinicalCardiology, Heart Failure and Transplantation Committee of the American Heart Association provided the followingdefinition for cardiomyopathies: "Cardiomyopathies are a heterogeneous group of diseases of the myocardiumassociated with mechanical and/or electrical dysfunction that usually (but not invariably) exhibit inappropriate

ventricular hypertrophy or dilatation and are due to a variety of causes that frequently are genetic"81 (see Chap. 84).

Cardiomyopathies are classified according to their phenotypic characteristics. The common form groups arehypertrophic, dilated, restrictive, and arrhythmogenic right ventricular cardiomyopathy. Phenotypic classification,although clinically convenient and useful, does not sufficiently reflect the molecular and genetic basis ofcardiomyopathies. Future classification of cardiomyopathies is expected to be based on our understanding of theirmolecular pathogenesis.

Genetic Basis of Hypertrophic CardiomyopathyHCM is a relatively common autosomal-dominant disease diagnosed clinically by the presence of unexplained cardiac

hypertrophy.82 Commonly, a left ventricular wall thickness of 13 mm, in the absence of hypertension or valvularheart disease, is used to define HCM. The prevalence of HCM is approximately 1 in 500 in young adults.83 It is likelyhigher in the elderly population because of age-dependent penetrance.

Cardiac hypertrophy, the clinical hallmark of HCM, is asymmetric in approximately two-thirds of cases, withpredominant involvement of the interventricular septum (Fig. 82–3). Hence the term asymmetric septal hypertrophyis used to describe this condition. Rarely, hypertrophy is restricted to apex of the heart (apical HCM). Morphologically,the left ventricular cavity is small, and left ventricular ejection fraction, a measure of global systolic function, is

increased. However, more sensitive indices of myocardial function show impaired contraction and relaxation.84

Diastolic function is commonly impaired, leading to an increased left ventricular end-diastolic pressure and thus,frequently, to symptoms of heart failure (see Chap. 84).

Figure 82–3.




Patients with HCM exhibit protean clinical manifestations ranging from minimal or no symptoms to severe heartfailure. The clinical manifestations often do not develop until the third or fourth decade of life, but the onset isvariable. The majority of patients are asymptomatic or mildly symptomatic. Predominant symptoms include dyspnea,chest pain, palpitations, and/or syncope. Severe systolic heart failure is uncommon. It occurs in a small fraction of

patients in whom the disease evolves into a dilated cardiomyopathy (DCM) phenotype.85 In contrast, cardiac diastolicfunction is usually impaired and left ventricular end-diastolic pressure is elevated. A dynamic left ventricular outflowis present in approximately 25% of patients. It could contribute to mitral regurgitations and symptoms of heartfailure. Cardiac arrhythmias, in particular atrial fibrillation and nonsustained ventricular tachycardia, are relatively

common and are associated with adverse clinical outcome.86,87 Wolff-Parkinson-White (WPW) syndrome is present ina small percentage of patients with HCM. Its presence suggests the possibility of a phenocopy, typically a glycogen

storage disease.88-91

Syncope is a serious symptom. It is often a result of serious cardiac arrhythmias and associated with an increased

risk of sudden cardiac death (SCD).92-94 HCM, although uncommon, is the most common cause of SCD in young,competitive athletes.95,96 It accounts for almost half of all cases of SCD in athletes younger than 35 years of age inthe United States.95,96 SCD is often the first and tragic manifestation of HCM in young, apparently healthyindividuals.95,96 Table 82–5 lists the factors associated with an increased risk of SCD.97 Overall, in the assessment ofthe risk of SCD, combination of all known risk factors should be considered.94 In the absence of major risk factors for

Main pathologic features of hypertrophic cardiomyopathy. A. Gross cardiac hypertrophy with predominant involvement of theinterventricular septum and a small left ventricular cavity. B. Myocyte disarray and hypertrophy. C. Interstitial fibrosis.




SCD, HCM has a relatively benign course, with an estimated annual mortality of approximately 1% in the adult

population.98-100 Apical HCM, characterized by giant T-wave inversion in the precordial leads on theelectrocardiogram, is also a relatively benign disease.101,102

The pathologic hallmark of HCM is cardiac myocyte disarray.103 It is defined as malaligned, distorted, and often shortand hypertrophic myocytes oriented in different directions (see Fig. 82–3). Myocyte disarray often comprises more

than 20% of the ventricle, as opposed to <5% of the myocardium in the normal hearts.103-105 It is more prominentin the interventricular septum, but commonly is found throughout the myocardium.103,105 Other pathologic featuresof HCM include myocyte hypertrophy, interstitial fibrosis, thickening of media of intramural coronary arteries, andsometimes malpositioned mitral valve with elongated leaflets. Cardiac hypertrophy, interstitial fibrosis, and myocyte

disarray are associated with the risk of SCD, mortality, and morbidity in patients with HCM.106-109 Other pathologicfeatures of HCM include thickening of the media of intramural coronary arteries, abnormal positioning of the mitralvalve apparatus, and elongated mitral valve leaflets.

MOLECULAR GENETICS OF HYPERTROPHIC CARDIOMYOPATHYHCM is a genetically heterogeneous disease with an autosomal-dominant mode of inheritance. Approximately two-

thirds of patients have a family history of HCM.110-112 In the remainder, the disease is sporadic. Familial andsporadic cases both are caused by mutations in contractile sarcomeric proteins.109 In sporadic cases, mutations arede novo and could be transmitted to the offspring of the index cases.113,114 Because hypertrophy is a commonresponse of the heart to all forms of injury or stimuli, a phenotype of hypertrophy in the absence of an increasedexternal load could also occur because of mutations in nonsarcomeric proteins. As such, unexplained cardiac

hypertrophy, which clinically denotes HCM, could also occur in storage disorders,115 metabolic disorders,116

mitochondrial diseases,117 and triplet repeat syndromes,118 as well as congenital heart diseases.63 Although thegross phenotype is similar, the pathogenesis of HCM caused by different classes of mutant proteins, at least in part,could differ. Therefore, such conditions are considered phenocopy (diseases mimicking HCM).

Causal Genes and MutationsThe pioneering works of Christine and Jonathan Seidman have led to elucidation of the molecular genetic basis ofHCM. In 1990, an arginine-to-glutamine substitution at codon 403 (R403Q) in the -myosin heavy chain (MHC) wasidentified as the first causal mutation.119 Since then, several hundred different mutations in more than a dozen genesencoding sarcomeric proteins have been identified (Table 82–6). Consequently, HCM (excluding phenocopy) is

considered a disease of contractile sarcomeric proteins (Fig. 82–4).120 Systematic screening of sarcomeric genessuggests that mutations in MYHC and MYBPC3, which encode -MHC and myosin-binding protein C (MBP-C),respectively, are the most common causes of human HCM, together accounting for approximately half of all

cases.121-123 Mutations in TNNT2 and TNNI3, encoding cardiac troponin T and I, respectively, are relativelyuncommon, each accounting for approximately 3% to 5% of the HCM cases.123-125 Thus mutations in MYH7,MYBPC3, TNNT2, and TNNI3 collectively account for approximately 60% of all HCM cases. A small fraction of HCMcases are caused by mutations in genes encoding -tropomyosin (TPM1), titin (TTN), cardiac -actin (ACTC),telethonin (TCAP), and essential and regulatory light chains (MYL3 and MYL2, respectively).120,126-131 Rare

Table 82–5. Potential Risk Factors for SCD in Patients with HCM

Established risk factors

Prior episode of aborted SCD (sudden cardiac arrest)

Family history of SCD (more than one victim of SCD)

Causal mutations, including double mutations

Modifier genes (genetic background)

History of syncope

Severe cardiac hypertrophy

Sustained and repetitive nonsustained ventricular tachycardia

Less-established risk factors

Left ventricular outflow tract gradient

Histologic phenotypes (interstitial fibrosis and myocyte disarray)

Early onset of clinical manifestations (young age)

Abnormal blood pressure response to exercise

Presence of myocardial ischemia

HCM, hypertrophic cardiomyopathy; SCD, sudden cardiac death.




mutations in cardiac troponin C (TNNC1), -MHC (MYH6), myosin light chain kinase (MYLK2), phospholamban (PLN),and caveolin 3 (CAV3) have been reported in patients with HCM.132-136 Finally, mutations in MYOZ2 and TCAPencoding Z disc proteins myozenin 2 (or calsarcin 1) and telethonin, respectively, are also uncommon causes of HCM137,138 Overall, the causal genes and mutations for approximately two-thirds of HCM cases have been identified. Theremainder are yet to be identified or are a result of genes inducing a phenocopy.

Table 82–6. Causal Genes for Hypertrophic Cardiomyopathy (Sarcomeric Genes)

Gene Symbol Locus Frequency Predominant Mutations

-Myosin heavy chain MYH7 14q12 25% Missenses

Myosin binding protein-C MYBPC3 11p11.2 25% Splice-junction and insertion/deletion

Cardiac troponin T TNNT2 1q32 3%-5% Missenses

Cardiac troponin I TNNI3 19p13.2 3%-5% Missense and deletion

-Tropomyosin TPM1 15q22.1 <3% Missenses

Essential myosin light chain MYL3 3p21.3 <3% Missenses

Regulatory myosin light chain MYL2 12q23-24.3 <3% Missense and one truncation

Cardiac -actin ACTC 15q11 <3% Missense mutations

Titin TTN 2q24.1 <3% Missense mutations

Telethonin (Tcap) TCAP 17q2 Rare Missense mutations

-Myosin heavy chain MYH6 14q1 Rare Missense and rearrangement mutations(association)

Cardiac troponin C TNNC1 3p21.3-3p14.3

Rare Missense mutations (association)

Cardiac myosin light peptidekinase

MYLK2 20q13.3 Rare Point mutations (association)

Caveolin 3 CAV3 3p25 Rare Point mutations (association)

Phospholamban PLN 6p22.1 Rare Point mutations (association)

Myozenin 2 MYOZ2 4q26-q27 1:250 Point mutations

Figure 82–4.




More than 100 different mutations in the -MHC, a major component of thick filaments in sarcomeres, have beenidentified. The majority are missense mutations, localized in the globular head of the myosin molecule. Codons 403

and 719 are potential hotspots for mutations.139,140 Missense, deletion, and insertion/deletion mutations in the rodand tail regions have also been described but are uncommon.141-145 Overall, the frequency of each particular MYHCmutation is relatively low, and most mutations are private. Accordingly, a founder effect (sharing of a commonancestor) is uncommon.

Mutations in MYBPC3 account for approximately 25% of all HCM cases. Mutations are scattered throughout the gene

without a particular predilection.122,146-149 Unlike mutations in MYHC, a significant proportion of MYBPC3 mutationsare deletion/insertion or splice-junction mutations.122 Insertion/deletion mutations could result in frameshift andtruncation of the MBP-C protein. The mutant protein harbors severe structural and functional defects or becomesdegraded. The frequency of each particular mutation is relatively low, and a founder effect is uncommon.

Mutations in TNNT2 are relatively common causes of human HCM, accounting for approximately 3% to 5% of all

cases.120,125,126,150,151 More than 20 mutations in TNNT2 have been described, and codon 92 is considered ahotspot for mutations.120,150,151 The majority of the mutations are missense, but deletion mutations that involvesplice donor sites and could lead to truncated proteins also have been described.120

Mutations in TNNI3 are also relatively uncommon and estimated to account for approximately 3% to 5% of all HCM

cases.123,124,152,153 Mutations in other components of thin and thick filaments are uncommon causes of HCM andcollectively account for approximately 5% of all HCM cases. Mutations in other sarcomeric genes, namelyTPM1,TNNC1, TTN,ACTC,MYL3, and MYL2, are very uncommon, and those in MYH6, myosin light chain kinase,SERCA2A, and phospholamban are rare.

Modifier Genes and PolymorphismsA remarkable feature of HCM is the presence of considerable variability in its phenotypic expression, whether it is thedegree of cardiac hypertrophy or the risk of SCD. The molecular basis of such variability is not fully known. It isprobably partly because of the diversity of the causal genes and mutations, which impart a spectrum of functional

and structural defects.154 In addition, the presence of multiple mutations, detected in a small fraction of patients, isassociated with a severe phenotype.155,156 Environmental factors, such as competitive sports and exercise, couldpotentially contribute to the phenotypic expression of HCM.157,158 However, there are insufficient data to supporttheir contributions to the phenotype.

The presence of considerable phenotypic variability among affected members of different families with identical causal

Schematic representation of sarcomeric proteins involved in cardiomyopathies.




mutations emphasizes the significance of the genetic background to phenotypic expression of HCM. Genes other thanthe causal genes that affect the phenotype are referred to as the modifier genes. Unlike the causal genes, modifier

genes are neither necessary nor sufficient to cause HCM.159 However, they influence the severity of cardiachypertrophy, risk of SCD, and expression of other cardiac phenotypes in HCM. DNA polymorphisms, includingputatively functional SNPs (ie, SNPs located in the coding or regulatory regions or splice junctions) in genes involvedin cardiac hypertrophy, are the prime candidates. The identity of the modifier genes for HCM and the magnitude oftheir effects remain largely unknown. Five modifier loci have been mapped, and several genes have been

implicated.160,161 The effect sizes of the modifier loci are considerable, as the loci in homozygous form drasticallyinfluence expression of cardiac hypertrophy in patients with the causal mutation. A modifier locus also has been

mapped in a genetically engineered mouse model.162 Functional variants of genes coding for the components of therenin-angiotensin-aldosterone system are the most extensively studied candidates. ACE, encoding angiotensin-I

converting enzyme 1 (ACE-1), was the first gene implicated as a modifier of cardiac phenotype in human HCM.163

ACE-1 catalyzes conversion of angiotensin I to angiotensin II and inactivates bradykinin, both of which are

biologically active agents with opposing effects on cardiac growth and cellular proliferation.164ACE contains more than30 different polymorphisms, which contribute to interindividual variation in plasma, tissue, and cellular levels of ACE-

1.165 The most commonly studied ACE polymorphism is an insertion (I) or deletion (D) of a 287-bp Alu repeat inintron 16. The I/D polymorphism, probably because of being in LD with other functional SNPs, is associated with

variation in plasma, cellular, and tissue levels of ACE-1 in a codominant manner (DD > ID > II).165

ACE I/D polymorphism has been associated with the severity of cardiac hypertrophy and risk of SCD in HCM in most,

but not all, studies.162-172 The DD genotype is more common in HCM families with a high incidence of SCD, ascompared with those with a low incidence, and is associated with the severity of cardiac hypertrophy.163,166 Theobserved association is gene-dose dependent, consistent with the biologic effect of the I/D variants on plasma and

tissue levels of ACE.166 An interaction between the modifying effect of the I/D genotypes and the underlying causalmutations also has been reported.167,172

Variants of endothelin-1 (EDN1), tumor necrosis factor- (TNF- ), angiotensinogen (AGT), angiotensin II receptor 1(AGTR1), and platelet-activating factor acetylhydrolase (PLA2G7) have been associated with the severity of the

cardiac hypertrophy.173-175 The results, however, have been inconsistent, partly because of the small sample size ofthe studies, population characteristics, and presence of confounders that are common in SNP-association studies.176

MOLECULAR GENETICS OF HCM PHENOCOPY

Mutations in nonsarcomeric proteins can cause a phenotype grossly similar to HCM, referred to as a phenocopy.115

The distinction between true HCM and HCM phenocopy is important because the pathogenesis, as well as histologicphenotypes, of the two conditions differ. Table 82–7 provides a partial list of HCM phenocopy. The prevalence of HCMphenocopy is not precisely known. Given the prevalence of each particular HCM phenocopy, that phenocopy is

expected to comprise approximately 5% to 10% of the cases with the clinical diagnosis of HCM.115 A prototypicexample of HCM phenocopy is Fabry disease, an X-linked lysosomal storage disease.177,178 Fabry disease is presentin approximately 3% of cases with the clinical diagnosis of HCM in adult population.179 The phenotype results fromdeficiency of -galactosidase A ( -Gal A), also known as ceramide trihexosidase.180 The deficiency of the enzymeresults in deposits of glycosphingolipids in multiple organs, including the heart. The causal gene is GLA on

chromosome Xq22, which codes for lysosomal hydrolase -Gal A protein.180 The phenotype is characterized byangiokeratoma, renal insufficiency, proteinuria, neuropathy, transient ischemic attack, stroke, anemia, corneal

deposits, and cardiac hypertrophy.180 Cardiac hypertrophy, which is often indistinguishable from the true HCM, isassociated with high QRS voltage, conduction defects, and cardiac arrhythmias. Other cardiac phenotypes include

valvular regurgitation, coronary artery disease, myocardial infarction, and aortic annular dilatation.179,181 Thedisease predominantly affects males. Female carriers could exhibit a milder form.181 The diagnosis is established bymeasuring -Gal A levels and activity in leukocytes. Fabry disease can be treated with enzyme replacement therapyusing human -Gal A (agalsidase ) or recombinant human -Gal A (agalsidase ).177,178,182,183

Table 82–7. Genes Known to Cause Hypertrophic Cardiomyopathy Phenocopy

Gene Gene Symbol Chromosome Frequency

Protein kinase A, subunit PRKAG2 7q22-q31.1 1%-2%

-Galactosidase A {2468} GLA Xq22 3%

Unconventional myosin 6 MOY6 6q12 Rare

Lysosome-associated membrane protein 2 LAMP2 Xq24 1%-2%

Mitochondrial genes MTTG, MTTI MtDNA Rare

Frataxin (Friedreich ataxia) FRDA 9q13 Rare




Glycogen storage disease, caused by mutations in PRKAG2 gene, is another HCM phenocopy.88-91 Cardiachypertrophy results predominantly from storage of glycogen in myocytes. The gene encodes the 2 regulatory subunit

of adenosine monophosphate (AMP)–activated protein kinase (AMPK), which is considered the energy biosensor of

the cell. Mutations in PRKAG2 lead to cardiac hypertrophy, conduction defects, and WPW.88-90

HCM phenocopy also occurs in trinucleotide repeat syndromes, a group of genetic disorders caused by expansion of

naturally occurring trinucleotide repeats.184 HCM phenocopy occurs in Friedreich ataxia, an autosomal-recessiveneurodegenerative disease caused by expansion of GAA repeat sequences in the intron of FRDA.185

HCM phenocopy also occurs in patients with Noonan syndrome. The phenotype is characterized by dysmorphic facialfeatures, pulmonic stenosis, mental retardation, bleeding disorders, and cardiac hypertrophy. LEOPARD syndrome isan allelic variant of Noonan syndrome. Noonan syndrome is an autosomal-dominant disorder caused by mutations inPTPN11, KRAS, SOS1, and RAF1 in approximately three-quarters of the cases, as discussed earlier (see Noonan and

LEOPARD Syndromes above).63,64

Metabolic diseases also cause HCM phenocopy. Refsum disease, Pompe disease (glycogen storage disease type II),Danon disease, Niemann-Pick disease, Gaucher disease, hereditary hemochromatosis, and CD36 deficiency are

examples of metabolic disorders that cause HCM phenocopy.186-189 Defective mitochondrial oxidativephosphorylation pathways also cause HCM phenocopy. Kearns-Sayre syndrome is a mitochondrial diseasecharacterized by a triad of progressive external ophthalmoplegia, pigmentary retinopathy, and cardiac conduction

defects and less frequently HCM phenocopy.117

GENE EXPRESSION IN HYPERTROPHIC CARDIOMYOPATHYIn keeping with the diversity of cardiac phenotypes in HCM, expression levels of a variety of genes, in response to themutant protein, are altered. Expression of genes encoding contractile sarcomeric proteins, cytoskeletal proteins, ionchannels, intracellular signaling transducers, proteins maintaining the reduction–oxidation state, and those involved

in transcriptional and translation machinery are changed.190,191 Expression levels of the markers of "secondary"cardiac hypertrophy, such as skeletal -actin, isoforms of myosin light chain, and atrial natriuretic factor, areupregulated.190 Upregulation of markers of secondary cardiac hypertrophy suggest that hypertrophy in HCM is also a"secondary" phenotype. Accordingly, common pathways are involved in induction of cardiac hypertrophy in geneticand acquired forms. The diversity of molecular phenotype is in accord with the diversity of pathologic and clinicalphenotypes in HCM that encompass not only hypertrophy and disarray, but also interstitial fibrosis and others.

DETERMINANTS OF CARDIAC PHENOTYPE IN HYPERTROPHICCARDIOMYOPATHYCollective data from genotype–phenotype correlation studies indicate that mutations exhibit highly variable clinical,

ECG, and echocardiographic manifestations and that no particular phenotype is mutation specific.192 Observationaldata show that cardiac hypertrophy accelerates during puberty and adolescence in patients with HCM.193 The findingsuggests that growth factors contribute to expression of cardiac hypertrophy. Similarly, experimental and clinicalstudies suggest that cardiac hypertrophy, the clinical hallmark of HCM, is a compensatory phenotype and likely to be

modulated by a large number of genetic and nongenetic factors.194 Overall, the final phenotype in HCM is determinednot only by the causal mutations but also by the effects of modifier genes, environmental factors, epigenetic andepistatic factors, and posttranscriptional and posttranslational modifications of the proteins.

Impact of Causal Genes and MutationsCausal genes and mutations are the primary determinant of expressivity of cardiac phenotype, including the severity

of hypertrophy and the risk of SCD.121,122,149,150,195-199 Collectively, the data suggest gene- and mutation-specificeffects. In general, mutations in MYH7 are associated with an early onset, extensive hypertrophy, and a high

incidence of SCD, which are variable among different MYH7 mutations.195,196,198,200 MYH7 mutations are consideredmajor prognosticators in HCM (Fig. 82–5). Topography of the mutations and their impact on -MHC protein functionare likely to be important determinants of the severity of cardiac hypertrophy as well as the risk of SCD. However, itis important to note that there is a significant degree of variability, which is partly independent of the causalmutations and partly reflects the effects of modifier genes. Given the relatively low frequency of each causalmutation, results of genotype–phenotype correlation studies have had limited utility. Consistent correlations havebeen observed for only a few mutations, such as R403Q and R719W, which are associated with a high incidence of

SCD and severe hypertrophy (see Fig. 82–5).195,201 In contrast, G256Q and L908V are associated with a benign andQ930L with an intermediate prognosis.195,196

Myotonin protein kinase (Myotonic dystrophy) DMPK, DMWD 19q13 Uncommon

Protein tyrosine phosphatase, nonreceptor type 11 PTPN11 12q24 Uncommon, higher in children




The phenotype in the majority of patients with MYBPC3 mutations is relatively mild. Commonly, the age of onset ofclinical symptoms is late, the degree of cardiac hypertrophy is relatively mild, and the incidence of SCD is

low.122,145,198 Accordingly, the penetrance, although age-dependent, is relatively low. Thus a normal physicalexamination, ECG, and echocardiogram have low negative predictive value in early life. The clinical phenotype oftendevelops in the fifth or sixth decades of life. It may be unmasked by the presence of concomitant hypertension.Indeed, hypertensive HCM of the elderly could be a form of HCM caused by mutations in MYBPC3 and unmasked by

hypertension.145 Nonetheless, there is a significant degree of variability in the phenotypic expression of HCM causedby MYBPC3, as "malignant" mutations, associated with severe hypertrophy and a high incidence of SCD, also have

been described.122

The risk of SCD in HCM caused by mutations in MYH7 and MYBPC3 is partially reflective of the severity of

hypertrophy.153 Mutations associated with mild hypertrophy generally carry a relatively benign prognosis, and thosewith severe hypertrophy indicate a high incidence of SCD. This is in contrast to HCM caused by mutations in TNNT2,

which is characterized by mild cardiac hypertrophy, a high incidence of SCD, and extensive myocyte disarray.109,150

Inadequate genotype–phenotype correlation data are available regarding mutations in TNNI3, TPM1, TNNC1, TTN,ACTC, MYL3, and MYL2.

It is important to note that the results of genotype–phenotype correlation studies are subject to a large number ofconfounding factors, including the small size of the families, small number of families with identical mutations, lowfrequency of each mutation, phenotypic variability, homozygosity for the causal mutations or compound mutations,

and the influence of modifier genes and environmental factors.158,202,203 Collective data indicate that mutationsexhibit highly variable clinical, ECG, and echocardiographic manifestations, and no particular phenotype is mutation-

specific.193

Impact of Modifier Genes and PolymorphismsExpression of cardiac hypertrophy in HCM is modulated by interactions between the causal mutations, modifier genes,environmental factors, and epigenetic elements, as well as posttranscriptional and posttranslational modifications of

proteins.194 Given the complexity of cardiac hypertrophy, a large number of genes are likely to modify phenotypicexpression of HCM, each contributing a small fraction of the phenotype.

ACE I/D polymorphism, which is the most commonly studied polymorphism in HCM,162,165-171 accounts forapproximately 3% to 5% of the total variability of left ventricular mass in genetically unrelated populations and for

approximately 10% to 15% in members of the same family.174 Similarly, contributions of other potential modifiergenes appears to be small.172-174

Figure 82–5.

Kaplan-Meier survival curves in patients with hypertrophic cardiomyopathy. Survival curves in two families with two differentmutations, namely arginine-to-glutamine substitution at amino acid 403 (R403Q) and glutamine-to-lysine change at aminoacid 930 (Q930L), in the MYH7 are shown.




Impact of Environmental FactorsEvidence for the effect of environmental factors on cardiac phenotypes in HCM in humans, although expected, is

circumstantial. Experimental data in animal models of HCM have not shown consistent effects.156,157,204 Becausehypertrophy is considered a secondary phenotype, one could speculate that heavy physical exercise, particularlyisometric exercise, could stimulate the development of severe hypertrophy in HCM. Although there is no directevidence in humans, the common finding of HCM in young competitive athletes who succumb to SCD suggests thatheavy physical exercise may worsen the cardiac phenotypes.

PATHOGENESIS OF HYPERTROPHIC CARDIOMYOPATHYAs the diversity of the causal mutations suggests, there is no single initial defect common to all mutations (Table 82–8). The diversity of the clinical phenotypes, such as hypertrophic, dilated, or restrictive cardiomyopathy arising frommutations in the same gene, further adds to the complexity of the pathogenesis. Topography of the causal mutationis likely to be important, as the initial defect is likely to be domain, but not protein, specific. Mutations located in aspecific functional domain of a given protein are expected to confer similar initial defects, which may be organ-specific because of expression of the partner proteins. Given that each sarcomeric protein could have multiplefunctions, usually as a result of having multiple domains, mutations in the same protein could impart various initialdefects.

The causal mutation initiates a series of molecular events, which begins with alteration of the molecular structure andfunction of the protein (see Table 82–8). Because the majority of mutant sarcomeric proteins differ from the wild typeonly by a single amino acid (missense mutations), the mutant proteins incorporate into the sarcomere, albeit

sometimes inefficiently.205 After incorporation, mutant sarcomeric proteins exert diverse functional defects, such asalterations in myofibrillar Ca2+ sensitivity and ATPase activity.206-210 Functional phenotypes lead to activation ofsecondary molecules, which are largely unknown but expected to include activating of many intracellular signalingpathways. The secondary molecular phenotypes mediate induction of the morphologic and histologic phenotypes.Accordingly, hypertrophy and fibrosis are considered secondary phenotypes because of activation of intermediarymolecular phenotypes (Fig. 82–6).

Table 82–8. Initial Defects Caused by Mutations in Sarcomeric Proteins

1. Mechanical defect

Impaired actomyosin interaction

Impaired cardiac myocyte and myofibril contractile performance

2. Biochemical defects

Impaired protein-protein binding

Impaired Ca2+ affinity of myofibrillar force generation

3. Bioenergetics

Impaired myofibrillar adenosine triphosphatase activity

4. Structural defects

Impaired sarcomere assembly

Impaired subcellular localization of sarcomeric proteins

Altered stoichiometry

Figure 82–6.




Many HCM mutations involve deletions or truncations that are considered null alleles because of the possible

expression of unstable mRNA and proteins.143,211 Whether haploinsufficiency causes HCM by altering thestoichiometry of the sarcomeric proteins remains unknown. Myocyte dropout caused by apoptosis also has been

implicated in animal models.212 However, the significance of myocyte apoptosis in the pathogenesis of human HCMremains to be established. Regardless of the initial primary defect, cardiac hypertrophy, the clinical hallmark of HCM,is considered a compensatory phenotype. Evidence suggesting the compensatory nature of cardiac hypertrophyincludes upregulation of expression of molecular markers of secondary cardiac hypertrophy, such as atrial and brain

natriuretic peptides,189,213,214 endothelin-1,215 transforming growth factor 1 (TGF 1), and insulin-like growth factor

1 (IGF-1).216 The predominant involvement of the left ventricle and its frequent absence in the low-pressure rightventricle, despite equal expression of mutant sarcomeric protein in both, suggest contribution of the environment tothe development of hypertrophy. Furthermore, variation in hypertrophic response because of the genetic background,its absence early on in life, and its attenuation through pharmacologic interventions, at least in animal models,

supports the secondary nature of hypertrophy.217,218 The primary impetus for hypertrophy is not well defined. It islikely to involve activation of myofibrillar signaling pathways in response to increased cell mechanical stress, altered

moiety of the mutant proteins for the signaling molecules, and/or altered Ca2+ homeostasis.

POTENTIAL NEW THERAPEUTIC INTERVENTIONS

Current pharmacologic interventions in HCM are empiric (class IIa).82 None of the current pharmacologic agents havebeen shown to induce regression of hypertrophy, fibrosis, and disarray, which are associated with increased mortality

and morbidity (class IIb).106,107 Currently, there is no suitable method to correct the underlying genetic defect.Therefore, the emphasis has been on prevention, reversal, and attenuation of the phenotype through pharmacologicinterventions aimed at blockade of intermediary molecular phenotypes. Recent studies have shown potential clinicalusefulness of angiotensin II receptor blockers, beta-hydroxy-beta-methylglutaryl-coenzyme A (HMG-CoA) reductaseinhibitors, and antioxidants in prevention, attenuation, and reversal of cardiac phenotypes in animal models of HCM

(class IIb).217-220 For example, blockade of angiotensin II receptor 1 in cardiac troponin T-Q92 transgenic micereduced interstitial collagen volume, expression levels of collagen 1 (I) mRNA, and TGF 1 protein to normal

levels.219 Treatment with HMG-CoA reductase inhibitors also has been shown to prevent and attenuate cardiacphenotype in a transgenic rabbit model of human HCM.217,218 Treatment with simvastatin, a pleiotropic HMG-CoAreductase inhibitor, reduced left ventricular mass, wall thickness, and collagen volume fraction in the -MHC-Q403transgenic rabbit model of human HCM.217 In addition, indices of left ventricular filling pressure were improvedsignificantly. Similarly, administration of atorvastatin prevented the development of cardiac hypertrophy in the same

transgenic rabbit model.218 Treatment with antioxidant N-acetylcysteine (NAC) reversed interstitial fibrosis in amouse model of HCM.220 Likewise, treatment with NAC resolved cardiac hypertrophy and fibrosis in a transgenicrabbit model of HCM and reduced vulnerability to cardiac arrhythmias.221 Calcineurin inhibitors, although effective inattenuation of experimental cardiac hypertrophy,222 were found to worsen cardiac phenotype in an -MHC mousemodel of HCM.206 Pretreatment with diltiazem, an L-type Ca2+ channel blocker, prevented the exaggerated cardiachypertrophic response to calcineurin inhibitors in the -MHC mouse model.206 Collectively, the results in geneticallyengineered animal models raise the possibility of pharmacologic interventions to prevent, attenuate, and reverseevolving cardiac phenotype in HCM. Clinical trials in humans are needed to determine potential salutary effects ofHMG-CoA reductase inhibitors, angiotensin II blockers, and N-acetylcysteine in human patients with HCM.

Genetic Basis of Dilated Cardiomyopathy

Sequence of phenotype characterization in the pathogenesis of cardiomyopathies.




DCM is a primary disease of the myocardium that manifests by dilatation of the left ventricular along with a gradualdecline in contractility. The diagnosis is based on a left ventricular ejection fraction of less than 0.45 and a left

ventricular end-diastolic diameter of more than 2.7 cm/m2. It has a prevalence of 40 cases per 100,000 individualsand an incidence of 5 to 8 cases per 100,000 persons.223 Patients with DCM are often asymptomatic in the earlystages but gradually develop symptoms and signs of heart failure, syncope, cardiac arrhythmias, and SCD. Thus anormal history and physical examination in a subject at risk, particularly in the early decades of life, does not excludeDCM. A significant number of affected relatives of patients with DCM are asymptomatic and are diagnosed for the first

time on additional testing (such as an echocardiogram).224 A family history of DCM is present in approximately half ofall index cases with idiopathic DCM.224,225 In the remainder, DCM is considered sporadic. Familial DCM is commonlyinherited as an autosomal-dominant disease,224 which clinically manifests during the third and fourth decades of life.

An X-linked DCM is suspected when only male members of a family exhibit symptoms and signs of DCM and there isno male-to-male transmission. Three common forms of X-linked DCM have been identified, including Duchenne andBecker muscular dystrophies, Emery-Dreifuss syndrome, and Barth syndrome. DCM also occurs in multiorgandisorders, such as mitochondrial DNA mutations, triplet repeat syndromes, and metabolic disorders.

MOLECULAR GENETICS OF DILATED CARDIOMYOPATHYDCM is an extremely heterogeneous disease, as indicated by the heterogeneity of the mapped loci and genes forfamilial DCM (Table 82–9). The predominant mode of inheritance is autosomal dominant; however, autosomal-

recessive and X-linked DCM also occur. Several causal genes for autosomal-dominant DCM have been identified.220

Several causal genes encode sarcomeric proteins, which are also known to cause HCM.221 Thus, despite thecontrasting phenotypes of HCM and DCM, mutations in sarcomeric genes can cause either of the phenotypes.Because many of the known causal genes for DCM involve the myocyte cytoskeleton, DCM has been considered to bea disease of cytoskeletal proteins. However, mutations in proteins other than the cytoskeletal proteins also can causeDCM.

Table 82–9. Causal Genes for Dilated Cardiomyopathy (DCM)

Gene Symbol Locus Inheritance Mutations/Frequency/Context

Sarcomeric/cytoskeletal

Cardiac -actin ACTC 15q11-14 Autosomaldominant

Missense/uncommon; also causes HCM

-Myosin heavy chain MYH7 14q11-13 Autosomaldominant

Missense/ 5%; also causes HCM

Cardiac troponin T TNNT2 1q32 Autosomaldominant


-Tropomyosin TPM1 15q22.1 Autosomaldominant

Missense/rare; also causes HCM

Cypher/ZASP (LIMdomain binding 3)

LDB3 10q22.3-q23.2

Sporadic andfamilial

Titin TTN 2q24.1 Autosomaldominant


Telethonin (T-cap) TCAP 17q12

Cytoskeletal

-Sarcoglycan SGCA 17q21 Autosomaldominant

Limb–girdle muscular dystrophy

-Sarcoglycan SGCB 4q12 Autosomaldominant

-Sarcoglycan SGCD 5q33-34 Autosomaldominant

Autosomalrecessive

Dystrophin DMD Xp21 X-linked Muscular dystrophy

Muscle LIM protein MLP(CSRP3)

11q15.1 Autosomaldominant

Rare, founder effect in families described

Ankyrin repeatdomain 1

ANKRD1 10q23.31 Sporadic 1%




Causal Genes and Mutations

The gene encoding cardiac -actin (ACTC) was the first causal gene identified for autosomal-dominant DCM.228

Subsequently, mutations in genes encoding additional components of the sarcomere, namely MYH7, TNNT2, TTN, and

TCAP, were found in patients with DCM.225,226,229 Because mutations in ACTC,MYH7, and TNNT2 are also known tocause HCM, these findings point to the commonality of the genetic basis of DCM and HCM. The diversity of thephenotype may reflect the topography of the causal mutations on the protein, as well as the genetic background ofthe individuals.

Mutations in cytoskeletal proteins—namely, delta sarcoglycan,230 beta sarcoglycan,231 metavinculin,232 anddystrophin233—are also important causes of DCM. Mutations in alpha sarcoglycan (adhalin) cause an autosomal-recessive form of DCM that occurs in conjunction with limb-girdle muscular dystrophy.231 Recently, a mutation inmuscle LIM protein (MLP) was identified in several related families with DCM.229 MLP interacts with telethonin, a titin-interacting protein, and colocalizes with it to the Z disk. Mutations in another Z disk protein named ZASP or Cypher or

LIM domain binding protein 3 (LDB3) also have been identified in patients with DCM.234,235 These findings, togetherwith the results of studies in the LIM-deficient mouse model, implicate the Z disk is a mechanosensor for cardiacmyocytes. Furthermore, rare mutations in ABCC9, which encodes the regulatory SUR2A subunit of the cardiac K(ATP)

channel, have been identified in patients with DCM.236 Mutant SUR2A proteins show aberrant redistribution ofconformations in the intrinsic adenosine triphosphate (ATP) hydrolytic cycle and induce abnormal K(ATP) channel

phenotypes.236

An intriguing causal gene for familial DCM is the lamin A/C gene,237 which encodes a nuclear envelope protein. Theobserved phenotype resulting from mutations in the rod domain of lamin A/C is progressive conduction disease, atrialarrhythmias, heart failure, and SCD. Moreover, mutations in the intermediary filament desmin and its associated

protein alphaB-crystallin have been identified in patients with DCM.238,239 Often such mutations lead to a phenotypeof cardiac and skeletal myopathy that is referred to as desmin-related myopathy.239 Collectively, these findingssuggest that mutations affecting the integrity of the sarcomeric, cytoskeleton, and Z disk proteins are the maincauses of DCM.

Intermediary filaments

Desmin DES 2q35 Autosomaldominant

Also causes RCM and desminopathies

-B-Crystallin CRYAB 11q35 Desminopathy

Nuclear proteins

Lamin A/C LMNA 1q21.2 Autosomaldominant

DCM, conduction defect, muscular dystrophy,lipodystrophy, insulin resistance

Emerin EMD Xq28 X-linked

Vinculin VCL 10q22.1-q23

Sporadic Metavinculin isoform

Cell junction molecules

Desmoplakin DSP 6p23-25 Autosomalrecessive

Also causes ARVC

Unknown

Taffazin (G4.5) TAZ Xq28 X-linked Ventricular noncompaction

1q32

2q14-22

2q31

3p22-25

6q23-24

9q13-22

10q21-23 Autosomaldominant

7p12.1-7q21

ARVC, arrhythmogenic right ventricular cardiomyopathy; DCM, dilated cardiomyopathy; HCM, hypertrophiccardiomyopathy; RCM, restrictive cardiomyopathy.




Duchenne and Becker Muscular DystrophiesThe phenotype is characterized by progressive degeneration of muscle function. It commonly manifests itself as mildbut progressive skeletal myopathy, early contractures, and cardiomyopathy. The incidence of Duchenne/Becker

muscular dystrophy is 1 in 3500 newborn males.240 Duchenne muscular dystrophy is a severe form and Beckermuscular dystrophy a milder form of the disease. The disease commonly manifests itself during the first or seconddecades of life in male patients. Female family members are commonly spared. However, they may exhibit a mildphenotype, typically late in life. Cardiac involvement includes progressive atrioventricular block, arrhythmia, loss of P-wave amplitude on the ECG, atrial standstill, DCM, akinesis/dyskinesis of the posterobasal wall of the left ventricle,and SCD. Often, DCM is the primary feature of Duchenne and Becker muscular dystrophies. Approximately 90% of

patients will eventually develop DCM.241 Death often occurs by the third decade of life.

The gene responsible for Duchenne and Becker muscular dystrophies is dystrophin, located on Xp21, which encodes a

large cytoskeletal protein.242 A variety of point, deletion, and insertion mutations or gene rearrangements indystrophin have been described.233,241 Approximately two-thirds of the mutations are either deletions orduplications. DCM can also occur in the absence of skeletal myopathy. Mutations leading to a frameshift induce asevere form, whereas missense mutations often lead to a mild form of the disease. Mutations in the 5' region of the

dystrophin gene can cause DCM without skeletal involvement.243

Emery-Dreifuss Muscular DystrophyEmery-Dreifuss muscular dystrophy is an X-linked degenerative disorder characterized by mild but progressive

skeletal and cardiac myopathy.244,245 Clinical features include muscle weakness and atrophy, flexion deformities ofthe elbows, and mild pectus excavatum. Cardiac phenotypes include cardiomyopathy, arrhythmia, SCD, conduction

defects, loss of P-wave amplitude on the ECG, and atrial standstill.246

The causal gene is EMD, which encodes emerin. Emerin is located along the nuclear rim of many cell types. It is a

member of the nuclear lamina-associated protein family.247

Barth SyndromeDCM is also a major phenotypic component of the Barth syndrome, another X-linked disorder. The characteristicphenotype of Barth syndrome includes skeletal and cardiac myopathy, neutropenia, and abnormal mitochondria.Barth syndrome is caused by point, deletion, and splice-junction mutations in the tafazzin (TAZ) or G4.5 gene,

located on Xq28.248

Modifier Genes and MutationsThe phenotype of DCM is determined not only by the causal mutations, but also by the modifier genes and theenvironmental factors. Genetic studies to identify the modifier genes for DCM are largely restricted to SNP-associationstudies and have limitations similar to those described for HCM. Several potential candidates, including ACE, havebeen identified. None have been established to modify cardiac phenotypes in DCM. A modifier locus in a calsequestrin

mouse model of DCM has been mapped. 249

GENOTYPE–PHENOTYPE CORRELATION IN DILATED CARDIOMYOPATHYThere is no large-scale systematic study to delineate the impact of causal and modifier genes and mutations on theDCM phenotype. Therefore, the available genotype–phenotype data may be specific to a family or small number offamilies. Overall, a diverse array of phenotypes caused by mutations in different genes is observed in DCM families.Typically, mutations in cardiac -actin, -MHC, and cardiac troponin T (cTnT) cause DCM without other phenotypes,such as conduction defects or deafness.226 In contrast, mutations in the rod domain of lamin A/C cause DCM inconjunction with progressive conduction defects, atrial arrhythmias, and SCD.237 Mutations in the lamin A/C gene canalso cause an autosomal-dominant form of Emery-Dreifuss syndrome.250 Mutations in desmin and alphaB-crystallingenes are commonly associated with skeletal myopathy as well as DCM with unique pathologic features, a phenotype

referred to as the desmin-related myopathy.251 Mutations in the dystrophin gene commonly lead to skeletal andcardiac myopathy. The severity of the myopathic phenotype is partly determined by the type of mutation. Those thatare frame-shift mutations—for example, insertion or deletion of a single base—cause a severe form, whereasmissense mutations often lead to a mild form of DCM and muscular dystrophy. Mutations in the 5' region of the

dystrophin gene can cause DCM without skeletal involvement.243

Cardiac involvement is quite common in triplet repeat syndromes and includes DCM, conduction disorders, andarrhythmia. Prevalence of cardiac involvement increases with advancing age, and approximately three-quarters ofadult patients exhibit conduction defects, such as first-degree atrioventricular block and intraventricular conduction

defects.252 There is also a direct relationship between the severity of the disease and the severity of cardiacinvolvement and the number of CTG repeats.252




PATHOGENESIS OF DILATED CARDIOMYOPATHYMutations in cardiac -actin, -myosin heavy chain, cardiac troponin T, and cytoskeletal proteins are expected toimpart a dominant-negative effect on transmission of the contractile force to the extracellular matrix proteins.227,228

Identification of mutations in MLP, LDB3, and TCAP emphasize the significance of the Z disk in maintaining normal

cardiac function.229,234,235 Similarly, identification of mutations in the dystrophin-associated protein complex ascauses of DCM signifies the role of sarcolemma in the pathogenesis of DCM.230 Mutations in the dystrophin gene leadto decreased expression levels of dystrophin, a major cytoskeletal protein in skeletal and cardiac muscles. Decreased

levels of dystrophin are expected to impair mechanical coupling and myocyte shortening.243 The molecularpathogenesis of other X-linked DCMs because of mutations in EMD and TAZ is unknown.

Pathogenesis of DCM resulting from mutations in desmin and alphaB-crystallin involves deposition of desmin and

alphaB-crystallin aggregates in the myocardium.253 Molecular pathogenesis of DCM caused by mutations in lamin A/Cor emerin remain largely unknown. It is likely to involve disruption of integrity of the cytoskeleton. The pathogenesisof cardiomyopathies in patients with the triplet repeat syndromes is also unclear. Expansion of the CTG (CUG inmRNA) repeats in the genes responsible for triplet repeat syndromes could indirectly affect transcription, transport,

splicing, and translation of mRNAs of cardiac genes.183

Genetic Basis of Arrhythmogenic Right VentricularCardiomyopathy/DysplasiaArrhythmogenic right ventricular cardiomyopathy (ARVC) is an uncommon cardiomyopathy with characteristic clinical

and pathologic features.254-257 The clinical phenotype comprises ventricular arrhythmias, primarily originating fromthe right ventricle, SCD, and heart failure.254,256,257 The pathologic phenotype is characterized by the gradualreplacement of the cardiac myocytes by adipocytes and fibrosis (Fig. 82–7).254,257 A comprehensive approach for thediagnosis of ARVC has been developed by the European Society of Cardiology and the Scientific Council on

Cardiomyopathies of the International Society and Federation of Cardiology.258

The disease often has a "concealed" stage, which is characterized by minor ventricular arrhythmias and subtlepathologic findings. It is followed by symptomatic ventricular arrhythmias and gradual progression to right-heart

failure and, finally, global cardiac failure.257

Electrocardiographic features include the characteristic and yet uncommon epsilon wave, depolarization andrepolarization abnormalities in the right precordial leads, and ventricular arrhythmias originating from the right

ventricle.256,257

Figure 82–7.

Histologic features of arrhythmogenic right ventricular dysplasia. Fibrofatty infiltrate in the right ventricle is shown.




ARVC is an important cause of SCD in apparently healthy individuals.254,259-261 In the US population, it accounts for3% to 4% of SCD associated with physical activity in young athletes.95 In some reports, ARVC was found in up to25% of cases of nontraumatic SCD.259-262 Collectively, the data suggest that ARVC is an important cause of SCD inyoung competitive athletes.

ARVC PHENOCOPYThe presence of myocardial fat alone is distinct from ARVC. Significant fatty infiltration of the myocardium could be

present in normal individuals, particularly in the elderly.263,264 Cor adiposum (fatty infiltration of the myocardium) isdistinguished from true ARVC by the absence of right ventricular myocardial thinning, myocyte atrophy andapoptosis, patchy fibroadipocytic replacement of myocytes, predominantly in the right ventricle, and lymphocytic

myocarditis.263 Right ventricular dilatation, fibrosis, myocyte atrophy, and excess adipocytes have been observed inpatients with Becker muscular dystrophy, Emery-Dreifuss muscular dystrophy, and myotonic dystrophy.257 Thedistinction between muscular dystrophies and ARVC is usually not problematic because of the skeletal involvement inmuscular dystrophies.

Idiopathic right ventricular outflow tract tachycardia and stress-induced (catecholaminergic) polymorphic ventriculartachycardia, caused by mutations in cardiac ryanodine receptor (RYR2), often present with arrhythmias resembling

those in ARVC.265,266 The absence of structural or histologic cardiac abnormalities suggests phenocopy and not trueARVC.257

MOLECULAR GENETICS OF ARRHYTHMOGENIC RIGHT VENTRICULARCARDIOMYOPATHY/DYSPLASIA

ARVC is a genetic disease that is estimated to be familial in approximately 30% to 50% of cases.255 The mostcommon mode of inheritance is autosomal dominant. Recessive forms in conjunction with keratoderma and woollyhair (Naxos disease) or with predominant involvement of the left ventricle (Carvajal syndrome) also have been

described and referred to as cardiocutaneous syndrome.267,268

The genetic basis of ARVC is partially known, with many chromosomal loci having been mapped (Table 82–10).269-275 Mutations in DSP, JUP, PKP2,DSC2 and DSG2, encoding desmosomal proteins desmoplakin (DP), plakoglobin(PG), plakophilin 2 (PKP2), desmocollin 2, and desmoglein 2 (DSG2), have been identified (see Table 82–10).267,276-

279,280 Mutations in PKP2 appear to be the most common causes of ARVC, accounting for approximately 20% of thecases.276,279 Mutations in DSG2 and DSP each account for approximately 10% to 15% of cases of ARVC.279 Themajority of causal mutations cause frameshift and hence are expected to lead to premature termination of theproteins.

The causal gene for an autosomal-recessive form of ARVC, palmoplantar keratoderma, and peculiar woolly hairs

(Naxos syndrome) is JUP, which encodes plakoglobin.266 Plakoglobin is also a desmosome protein and, along with

Table 82–10. Chromosomal Loci and Causal Genes for Arrhythmogenic Right VentricularDysplasia

Chromosome Symbol Protein Function

ARVC1a

14q24.3 TGF 3 Transforming growth factor- 3 Mitotic and trophic factor

ARVC2a

14q42.2-q43 RYR2 Ryanodine receptor 2 Calcium channel

ARVC3 1q12-q22

ARVC4 2q32.1

ARVD5 3p23 TMEM43 Trans-membrane protein 43 Unknown

ARVD6 10p12-p14

ARVD7 10q22

ARVC8 6p24 DSP Desmoplakin Desmosomes

ARVC9 12p11 PKP2 Plakophilin 2 Desmosomes

18q12.1 DSG2 Desmoglein 2 Desmosomes

18q21 DSC2 Desmocollin 2 Desmosomes

Naxos disease 17q21 JUP Plakoglobin Desmosomes

aPhenocopy. RYR2 mutations cause catecholaminergic polymorphic ventricular tachycardia and not truearrhythmogenic right ventricular cardiomyopathy.




desmoplakin, anchors intermediate filaments to desmosomes. The phenotype was described first in a family from theisland of Naxos in Greece and was mapped to 17q21. Mutational analysis detected a 2-bp deletion in JUP, whichencodes PG, a major component of desmosomes and adherens junctions. Another recessive form of ARVC is Carvajal

syndrome, which is a cardiocutaneous syndrome with predominant involvement of the left ventricle.268 It is causedby mutations in DSP.

Recently, a point mutation in TMEM43 encoding transmembrane protein 43 was identified in the affected members of

a large family with ARVC mapped to chromosome 3p region.279 The mutation was fully penetrant and was associatedwith high mortality. The function of this protein is unknown.

A point mutation in the 5' untranslated region (UTR) of TGFB3 gene also has been identified in a family mapped to

14q24.3.281,282 Whether this point mutation in the 5UTR is a true causal mutation or simply a DNA marker remainsto be established.283

Mutations in RYR2 cause catecholaminergic polymorphic ventricular tachycardia, which was initially considered avariant of ARVC. However, it is now considered a distinct phenotype because of the absence of fibrofatty infiltration in

the myocardium.257,265,266

PATHOGENESIS OF ARRHYTHMOGENIC RIGHT VENTRICULARCARDIOMYOPATHY

The molecular pathogenesis of ARVC is unknown. Infection and inflammation,264,284,285 apoptosis,286 myocytetransdifferentiation,287 and myocyte detachment257 have been proposed. Apoptosis has been detected in autopsyspecimens collected from patients with ARVC.288,289 However, apoptosis alone does not explain the mechanism forfibrofatty infiltration of the myocardium. Experimental data suggest that suppression of the canonical Wnt signaling

by PG is a mechanism for the pathogenesis of ARVC.290 Accordingly, mutations in desmosomal protein impairdesmosome assembly and lead to excess free (unincorporated) PG and PKP2, which can translocate into the nucleus.Nuclear PG and PKP2, because of structural similarities to -catenin, the effector of the canonical Wnt signaling,compete with -catenin for binding to Wnt core protein complex, resulting in suppression of the canonical Wntsignaling. The latter leads to enhanced fibroadipogenesis and myocyte apoptosis, which are the characteristic findings

in human ARVC.290 Genetic fate-mapping experiments in animal models of ARVC have identified the second heartfield cardiac progenitor cells as a main source of excess adipocytes in ARVC. These cells in the presence of

suppressed canonical Wnt signaling and increased expression of adipogenic markers differentiate to an adipocytes.291

Genetic Basis of Restrictive CardiomyopathyRestrictive cardiomyopathy (RCM) is a heart-muscle disease characterized by severely enlarged atria as a result ofelevated right and left ventricular filling pressures, normal or reduced ventricular volumes, and, usually, preserved

global systolic function.292 The clinical manifestations are those of heart failure, often with predominance of right-sided signs and symptoms. The age of onset of the disease is variable, and the prognosis is relatively poor. RCM canoccur because of systemic infiltrative disorders, such as amyloidosis and sarcoidosis, and storage diseases, such as

Fabry disease.292 Although such disorders are also genetic in etiology, RCM in such disorders is an indirectconsequence and not a primary myocardial abnormality.

MOLECULAR GENETICS OF RESTRICTIVE CARDIOMYOPATHYFamilial RCM, with an autosomal-dominant form of inheritance in conjunction with skeletal myopathy and

atrioventricular conduction defects, has been described.293-295 Two causal genes for RCM—DES,296 encoding desmin,and TNNI, 3,297 encoding cardiac troponin I'have been identified. Desmin is an intermediary filament that is alsoinvolved in desminopathies involving skeletal muscles as well as the heart. Mutations in TNNI3, which are known to

cause HCM and DCM, also cause RCM.297 RCM also occurs in patients with Noonan syndrome,298 which is caused bymutations in the protein tyrosine phosphatase, nonreceptor type II.63 The pathogenesis of RCM remains largelyunknown.

Genetic Basis of Cardiomyopathies in Trinucleotide Repeat SyndromesTrinucleotide repeat syndromes are a group of genetic disorders caused by expansion of naturally occurring GC-rich

triplet repeats in genes.287,299 The group comprises more than 10 different diseases, including myotonic musculardystrophy and Huntington disease.183,299 Cardiac involvement is common in several forms of triplet repeatsyndromes and is a major determinant of morbidity and mortality.184 The phenotype commonly includes DCM, HCM,conduction disorders, and arrhythmias. Average life expectancy of the affected individuals is approximately 30 to 40years.

GENETIC BASIS OF CARDIOMYOPATHIES IN MYOTONIC DYSTROPHY

Myotonic dystrophy (DM) is an autosomal-dominant disorder with highly variable penetrance.118 The estimated




prevalence of DM is approximately 1 in 8000 in the North American population.118,252 It is the most common form ofmuscular dystrophy in adults. DM commonly manifests itself as progressive degeneration of muscles and myotonia,cardiomyopathy, conduction defects, male-pattern baldness, infertility, premature cataracts, mental retardation, and

endocrine abnormalities.118,252 Cardiomyopathy is a common phenotypic manifestation of myotonic DM.252 Cardiacconduction defects, such as first-degree atrioventricular block and intraventricular conduction defects, are present in

approximately three-quarters of adult patients.252

Mutations in two genes have been identified for DM, including expansion of CTG (CUG in mRNA) trinucleotide repeats

in the 3' untranslated region of DM protein kinase (DMPK), located on chromosome 19q3.300-302 The number of CTGrepeats in normal individuals varies between 5 and 37. It expands from 50 to more than several thousand in patients

with DM.121 Expansion of the repeats can interfere with DMPK transcription, RNA processing, and/or translation,resulting in decreased levels of expression of DMPK protein. Several proteins that bind to the trinucleotide repeats

have been identified, including CUGBP1, ETR3-like factors (CELFs), and muscle blind-like (MBNL) proteins.303

Increased activities of these proteins affect splicing and processing of several mRNAs, including splicing of cTnT.304

The length of the CTG repeats often correlates with the severity of clinical phenotypes, including conduction defects

and cardiomyopathy.305

The second gene responsible for DM is zinc-finger protein 9 (ZNF9), located on 3q21.306 Expansion of a CCTGtetranucleotide repeats in intron 1 of ZNF9 leads to expression of abnormal RNA, which binds and alters activities ofRNA-binding proteins. Increased activities of RNA-binding proteins affect splicing and expression of multiple targetgenes, resulting in the subsequent multiorgan phenotype.

GENETIC BASIS OF CARDIOMYOPATHIES IN FRIEDREICH ATAXIAFriedreich ataxia (FRDA) is an autosomal-recessive neurodegenerative disease. It primarily involves the central and

peripheral nervous system and less frequently manifests as cardiomyopathy and occasionally as diabetes mellitus.307

FRDA is caused by the expansion of the GAA trinucleotide repeats in intron 1 of FRDA.307 The encoded protein isfrataxin, which is a soluble mitochondrial protein with 210 amino acids. Cardiac involvement can manifest as either

DCM or HCM. The severity of clinical manifestations of FRDA also correlates with the size of the repeats.308 Thepathogenesis of cardiomyopathies in FRDA is likely to involve impaired iron homeostasis and increased oxidative

stress.309

Genetic Basis of Cardiomyopathies in Metabolic DisordersMetabolic cardiomyopathies encompass a group of disorders in which there is the primary metabolic abnormality inthe heart (Table 82–11). This metabolic abnormality may also involve other organs; however, cardiac involvement isdirect and not a consequence of secondary changes in other organs. Secondary involvement of the myocardium insystemic metabolic disorders is not considered a metabolic cardiomyopathy.

A prototype of metabolic cardiomyopathies is glycogen storage disease type II (glycogenosis type II or Pompedisease). Pompe disease is an autosomal-recessive disorder caused by deficiency of -1,4-glucosidase (acid maltase),which degrades -1,4 and -1,6 linkages in glycogen, maltose, and isomaltose.310 Deficiency of the enzyme leads tostorage of glycogen in lysosomal membranes. Phenotypic expression of Pompe disease includes HCM, DCM,conduction defects, and muscular hypotonia. The cause is mutations in the acid maltase gene. Mutations lead todeficiency of acid -glucosidase. A high-protein diet and recombinant acid -glucosidase have been used effectivelyto treat this disorder.311

Mutations in the gene encoding the AMP-activated 2 noncatalytic subunit of protein kinase A (PRKAG2) have beenidentified in families with HCM and WPW syndrome.88-91 AMP-activated kinase is a biosensor of the cellular energystate. Cardiac involvement varies from a predominant phenotype of preexcitation and conduction abnormalities to a

predominant phenotype of cardiac hypertrophy.88,91 Nonetheless, the primary phenotype appears to be a deposit ofglycogen in the myocardium, which is responsible for cardiac enlargement as well as facilitated atrioventricular (AV)

Table 82–11. Examples of Causal Genes for Metabolic Cardiomyopathies

Protein Symbol Locus Frequency Mutations/Phenotype

AMP-activated protein kinase,2 regulatory subunit

PRKAG2 7q35-q36 Rare Point and insertion mutations, HCM, WPW,and conduction defect

Acid maltase gene GAA 7q25.2-q25.3

Rare Pompe disease, DCM, HCM, conductiondefects

Phytanoyl-CoA hydroxylase PAHX orPHYH

10p13 Rare DCM, HCM, and conduction defects

AMP, adenosine monophosphate; CoA, coenzyme A; DCM, dilated cardiomyopathy; HCM, hypertrophiccardiomyopathy; WPW, Wolff-Parkinson-White syndrome.




conduction.90,91

Refsum disease is an autosomal-recessive disorder characterized clinically by a tetrad of retinitis pigmentosa,

peripheral neuropathy, cerebellar ataxia, and elevated protein levels in the cerebrospinal fluid.312 Cardiacinvolvement includes ECG abnormalities, which are common. Cardiac hypertrophy and heart failure are uncommon.Mutations in the gene encoding phytanoyl-coenzyme A (CoA) hydroxylase (PAHX or PHYH) are responsible for Refsum

disease.313 Mutations reduce the enzymatic activity and lead to accumulation of phytanic acid, an unusual branched-chain fatty acid in tissues and body fluids.314

Cardiac involvement in patients with mucopolysaccharidosis, Niemann-Pick disease, Gaucher disease, hereditary

hemochromatosis, and CD36 deficiency have also been described (reviewed in Watkins et al195).

Genetic Basis of Cardiomyopathies in Mitochondrial DisordersCardiomyopathies are common in patients with mitochondrial disorders. Mitochondrial cardiomyopathy exhibits amatrilineal transmission. Because nuclear genes encode for proteins that primarily regulate mitochondrial function,mutations in nuclear DNA can also cause mitochondrial myopathies. Mitochondrial DNA is a circular double-strandedgenome of approximately 16.5 kb, encoding 13 polypeptides of the respiratory chain complexes I, III, IV, and Vsubunits; 28 ribosomal RNAs; and 22 tRNAs (transfer ribonucleic acids). Mutations in mitochondrial oxidative

phosphorylation pathways often result in a complex phenotype involving multiple organs, including the heart.315

Cardiac involvement can lead to hypertrophy as well as dilatation. Each mitochondrion has multiple copies of its ownDNA, and each cell contains thousands of mitochondria. Therefore, mutations result in a significant degree ofheteroplasmy, which increases over time as the mitochondria multiply. In general, approximately 80% to 90% of

mitochondrial DNA must mutate in order to affect mitochondrial function and lead to a clinical phenotype.315

Kearns-Sayre syndrome is a mitochondrial disease caused by sporadically occurring mutations in mitochondrial

DNA.316 Kearns-Sayre syndrome is characterized by a triad of progressive external ophthalmoplegia, pigmentaryretinopathy, and cardiac conduction defects.316 The classic cardiac abnormality in Kearns-Sayre syndrome isconduction defects; however, DCM and HCM are also often observed, but at a lower frequency.

l-Carnitine deficiency is a cause of mitochondrial myopathy resulting from mutations in nuclear DNA. The phenotypeis characterized by skeletal myopathy, congestive heart failure, abnormalities of the central nervous system and liver,

and, rarely, HCM.317,318 Carnitine is an important component of fatty acid metabolism and is necessary for the entryof long-chain fatty acids into mitochondria. Mutations in the chromosomal gene encoding solute carrier family 22,member 5 (SLC22A5), or OCTN2 transporter impair transport of carnitine to mitochondria and cause systemic

carnitine deficiency.318 Similarly, mutations in genes encoding enzymes involved in the transfer and metabolism ofcarnitine can cause carnitine deficiency. The list includes carnitine mitochondrial carnitine palmitoyltransferase I(CATI or CPT-1), located in the outer mitochondrial membrane; carnitine-acylcarnitine translocase (SLC25A20),located in the inner membrane; and carnitine palmitoyltransferase 2 (CPT-2).

Mutations in acyl-CoA dehydrogenase also impair mitochondrial fatty acid oxidation and can lead to

cardiomyopathy.317,319 The clinical manifestations are remarkable for cardiac hypertrophy, with diminished systolicfunction, fasting hypoglycemia, inadequate ketotic response to hypoglycemia, hepatic dysfunction, skeletal

myopathy, and SCD.317,319 The majority of cases of medium-chain acyl-CoA dehydrogenase deficiency are caused bysubstitution of glutamic acid for lysine in the mutant protein, whereas the molecular genetic basis of short-chain acyl-CoA dehydrogenase deficiency is more heterogeneous.

GENETIC DISEASES OF CARDIAC RHYTHM AND CONDUCTION

Cardiac rhythm and conduction abnormalities could occur as the primary phenotypes of genetic disorders orsecondary phenotypes resulting from genetic diseases that primarily affect structure of the heart. In general, cardiacarrhythmias and conduction defects result from abnormalities in three main families of proteins: contractilesarcomeric proteins, such as that in HCM; the cytoskeletal proteins, which are responsible for DCM; and the ion

channels and their regulators, which are responsible for familial arrhythmias and conduction defects.320 As discussedearlier, there is significant phenotypic overlap as mutations in the same gene could cause a variety of cardiac rhythmand conduction disorders. This is best exemplified by mutations in sodium channel SCN5A, which could cause long QT

syndrome (LQT), Brugada syndrome, and familial conduction disease.321 Therefore, a simplistic classification ofgenetic disorders is considered preliminary, and some of the key genetic findings used to risk-stratify for SCD orarrhythmias are based on studies in only few families. Table 82–12 summarizes the list of genetic disorders in whichthe primary phenotype is cardiac arrhythmias and conduction defects.

Table 82–12. Genetic Disorders Causing Cardiac Arrhythmias in the Absence of StructuralHeart Disease (Primary Rhythm Disorders)




Rhythm Inheritance Locus Gene

Supraventricular

Atrial fibrillation AF AD 10q22 —

AD 6q14-16 —

AD 10p11-q21 —

AD 12p12 —

AD 5p15 —

AD 11p15 KCNQ1

AD 21q22 KCNE2

AD 11q13 KCNE3

AD 17q23 KCNJ2

AD 12p13 KCNA5

AD 1q21 GJA5

AR 5p13 NUP155

AD 3p21 SCN5A

AD 1p36-35 NPPA

Atrial standstill SND, AF AD 3p21 SCN5A

Sick sinus syndrome SND AD 15q24 HCN4

AR 3p21 SCN5A

Absent sinus rhythm SND, AF AD — —

WPW AVRT AD — —

Familial PJRT AVRT AD — —

Conduction disorders

PCCD AVB AD 19q13 —

3p21 SCN5A

Ventricular

LQT syndrome (RW) TdP AD

LQT1 11p15 KCNQ1

LQT2 7q35 HERG

LQT3 3p21 SCN5A

LQT4 4q25 ANKB

LQT5 21q22 minK

LQT6 21q22 MiRP1

LQT7 17q23 KCNJ2

LQT8 12p13 CACNA1C

LQT9 3p25 Cav3

LQT10 11q23 SCN4B

LQT11 7q21-q22 AKAP9

LQT12 20q11.2 SNTA1

LQT syndrome (JLN) TdP AR 11p15 21q22 KCNQ1 minK

SQT syndrome VF AD

SQT1 7q35 HERG

SQT2 11p15 KCNQ1

SQT3 17q23 KCNJ2

Catechol-aminergic PVT VT AD AR 1q42 1p13-p11 RYR2 CASQ2

Brugada syndrome VT/VF AD 3p21 SCN5A




Ion Channelopathies as the Basis for Cardiac Arrhythmias and ConductionDefectsIon channels, crucial units in cardiac excitability, are glycoproteins embedded in the membrane of the cardiacmyocytes, which allow flux of ions in and out of the cell to modulate the electrical gradient. Many different ionchannels are orderly activated to give rise to the electrical current that will ultimately be responsible for thedevelopment of the myocyte excitability. This is a complex process and requires a very well-controlled ionic balanceto prevent arrhythmogenesis.

The subunit of the cardiac sodium channel gene, SCN5A, which is responsible for phase 0 of the cardiac actionpotential, has been studied extensively during the past 5 years. SCN5A was first cloned and characterized in 1995

and localized to 3p21.322 The gene is comprised of 28 exons that code for a 2016–amino acid protein. It contains fourhomologous domains (DI-DIV), each of which contains six membrane-spanning segments (S1-S6).320 In 1995, Wanget al323 linked mutations in SCN5A to LQT syndrome, a disease characterized by prolongation of the QT interval andsudden death at a young age. Subsequently, mutations in SCN5A were linked to idiopathic ventricular fibrillation,324

the Brugada syndrome,321 progressive conduction defect,325 sudden infant death syndrome (SIDS),326 and suddenunexpected death syndrome (SUDS).327 The latter is a phenotype identified 2 decades ago in Southeast Asia thatwas causing sudden death in males, usually at night.327 Figure 82–8 shows the phenotypes arising from mutations inSCN5A.

AD 3p22 GPD-1L

AD 12p13 CACNA1c

AD 10p12 CACNB2b

AD 11q13 KCNE3

AD, autosomal dominant; AF, atrial fibrillation; AR, autosomal recessive; AVB, atrioventricular block; AVRT,atrioventricular reentrant tachycardia; JLN, Jervell and Lange-Nielsen; LQT, long QT; PCCD, progressive cardiacconduction defect; PJRT, paroxysmal junctional reentrant tachycardia; RW, Romano-Ward; SND, sinus nodedysfunction; TdP, torsade de pointes; VF, ventricular fibrillation; VT, ventricular tachycardia; WPW, Wolff-Parkinson-White syndrome.

Figure 82–8.




Brugada Syndrome and Its VariantsBrugada syndrome is identified by a characteristic ECG pattern consisting of right bundle-branch block and ST-

segment elevation in V1 to V3 (Fig. 82–9) and sudden death at a young age.328 It was described originally in 1992based on electrocardiographic pattern and occurrence of syncope or sudden death episodes in patients with a

structurally normal heart.328 The episodes of syncope and sudden death are caused by fast polymorphic ventriculartachycardia.328,329

Schematic structure of INa sodium channel (SCN5A) and phenotypes arising from mutations in SCN5A.

Figure 82–9.




Brugada syndrome often manifests in subjects in the third or fourth decades of life, and occasionally in infants as

SIDS.330 Recent studies suggest SUDS, which is prevalent in Southeast Asia, is a form of Brugada syndrome. SUDSis estimated to affect up to 1% of the population, and it is the most common cause of death in young males in

Thailand.331 Death often occurs at night and more commonly in male subjects (male-to-female ratio is 10:1).Electrocardiographically, the disease is identical to Brugada syndrome. As in Brugada syndrome, mutations in SCN5A

are responsible for SUDS, and biophysical data indicate a nonworking SCN5A or accelerated inactivation.327

GENETICS OF THE BRUGADA SYNDROME

SCN5A was identified in 1998 as the first, and thus far the only, causal gene for Brugada syndrome.324 More than 60different mutations in SCN5A have been identified that collectively account for approximately 25% of all cases ofBrugada syndrome.

Mutations of SCN5A can lead to a large spectrum of phenotypes, including Brugada syndrome, LQT3, isolated

progressive cardiac conduction defect, idiopathic ventricular fibrillation, atrial standstill, and SUDS.321 Thephenotypes are all considered allelic variants caused by mutations in SCN5A. Electrocardiographic, clinical, genetic,and biophysical data have clarified the relationship between these phenotypes. The distinction between the LQT3 andBrugada syndromes is difficult to ascertain in some cases, and one family has been described manifesting the

phenotype of both Brugada and LQT3 syndrome.329 Likewise, progressive conduction disease and Brugada syndromehave been described in members of a single family.330 Collectively, these data suggest that mutations in SCN5Acause variable phenotypic manifestations that span Brugada syndrome, LQT3, and progressive conduction defects(see Fig. 82–6).

As in many other genetic disorders, Brugada syndrome also exhibits locus heterogeneity, and a second locus on

chromosome 3 has been mapped.332 The gene involved has been recently described, the glycerol-3-phosphatedehydrogenase 1-like (GPD-1L), which seems to affect the trafficking of the cardiac sodium channel to the cellsurface. In fact, the responsible mutation (A280V) reduces inward sodium currents by approximately 50% and

SCN5A cell surface by approximately 31%.333

Recent reports demonstrate that not only mutations leading to a loss of function in the sodium channel (eitherthrough SCN5A or GPD-1L) can cause the Brugada syndrome, but also loss-of-function mutations in the cardiaccalcium channel CACNA1c (Cav1.2) and its alpha-subunit CACNB2b can be responsible for a syndrome overlapping

short-QT and the Brugada ECG pattern.334 These findings open up new lines of research, in which the concept ofBrugada syndrome as a pure sodium channelopathy gives way to the concept of the syndrome as an ionic imbalancebetween the inward and outward currents during phase 1 of the action potential. Supporting this hypothesis, the

Typical electrocardiogram of Brugada syndrome. Note the pattern resembling a right bundle-branch block and the ST-segment elevation in leads V1 to V2.




same group has very recently identified the first family with Brugada syndrome carrying a mutation (R99H) in theKCNE3 gene, which encodes a -subunit that is thought to modulate Kv4.3 channels, responsible for transientpotassium Ito currents.335 Functional studies demonstrate that cotransfection of the mutation R99H-KCNE3 withKCND3 results in a significant increase in the Ito intensity and underlies the development of Brugada syndrome inthis family.

PATHOGENESIS OF BRUGADA SYNDROMEThe identification of mutations in SCN5A in patients with Brugada syndrome suggests decreased availability of sodiumions could shift the ionic balance in favor of Ito during phase 1 of the action potential. Biophysical characterization of

mutations in SCN5A suggests that mutations decrease the Na1+ current availability by two main mechanisms:decreased expression of the mutant channel or acceleration of inactivation of the channel. In addition, the alterationin the ionic currents that worsen at higher temperatures has been implicated for certain mutations, such as the

T1620M.336 The clinical relevance of this mechanism is corroborated by the observation of several cases ofventricular fibrillation during febrile illnesses in patients with Brugada syndrome. Compared with LQT3, thepathogenesis of the Brugada syndrome could be considered a mirror image. Biophysical data indicates that LQT3

mutations cause a delayed inactivation of the channel,320 which is exactly the opposite as in Brugada syndrome,where there is an accelerated inactivation.336

GENOTYPE–PHENOTYPE CORRELATION IN BRUGADA SYNDROMELimited data is available regarding the correlation between genotypes and phenotype in patients with Brugadasyndrome. This may partly reflect very recent identification of the first causal gene, phenotypic variability of SCN5A,and allelic and locus heterogeneity of Brugada syndrome. It has been suggested that electrocardiographicparameters, such as longer conduction intervals (PQ and HV) on baseline ECG could distinguish the carriers of sodium

channel mutations from the noncarriers.337

In the last years, polymorphisms are acquiring greater importance to explain certain phenotypes of genetic diseases.In the SCN5A locus, the common H558R polymorphism has been shown to restore (at least partially) the sodium

current impaired by other simultaneous mutations causing either cardiac conduction disturbances (T512I)338 orBrugada syndrome (R282H).339 Thus this polymorphism seems to give rise to less severe phenotypes by mitigatingthe effect of nearby mutations. Recently the common variant H558R was confirmed as a genetic modulator of the

Brugada phenotype in patients with an SCN5A mutation.340

Long QT Syndrome

LQT syndrome is a disease of ventricular repolarization identified by the prolongation of the QT interval on ECG.320 Itis characterized by syncopal episodes, malignant ventricular arrhythmias, and ventricular fibrillation. The majority ofpatients with the LQT syndrome are asymptomatic. However, approximately one-third present with syncope oraborted malignant ventricular arrhythmias including torsade de pointes, which is the most typical ventriculararrhythmia in LQT syndrome. SCD is relatively common. Prognosis of the symptomatic cases, if untreated, is poor.Approximately one-fifth of patients who present with syncope and remain untreated die within 1 year, and 50% diewithin 10 years.

The LQT syndrome is either acquired, which is iatrogenic and commonly induced by drugs, or congenital. A commoncause of the acquired disease is the use of medications such as antiarrhythmics, antidepressants, and phenothiazines(Table 82–13). In addition, electrolyte imbalance, such as hypokalemia, hypomagnesemia, and hypocalcemia,especially in the presence of predisposing medications, could cause LQT syndrome.

Table 82–13. Selected Medications Associated with Prolonged QT Interval

Antiarrhythmic drugs

Quinidine

Procainamide hydrochloride

Disopyramide phosphate

Sotalol hydrochloride

Amiodarone

Ibutilide fumarate

Dofetilide

Propafenone

Anesthetics/antiasthmatics




Two patterns of inheritance have been described in the congenital LQT syndrome: (1) autosomal-recessive disease,described by Jervell and Lange Nielsen in 1957, which is associated with deafness, and (2) autosomal-dominantdisease, described by Romano and Ward, which is not associated with deafness and is more common than therecessive form.

The pathogenesis of LQT syndrome could be summarized as mutations in K1+ channel resulting in inadequateopening and decreased potassium outward current, whereas mutations in Na1+ channels lead to inadequate closureof the channels and excessive sodium inward currents. The ensuing result is inadequate maintenance of electricalgradient (loss of function) during an action potential and prolongation of the QT interval.

AUTOSOMAL-DOMINANT LQT SYNDROME (ROMANO-WARD SYNDROME)

The first locus for the autosomal-dominant disease was mapped to chromosome 11 in 1991.341 Since then, seven locihave been mapped and six genes identified (see Table 82–12). All encode proteins that are responsible forautomaticity of the electrical activity in the cardiac cells. Mutations cause a disruption in the formation of thechannels, altering the cardiac action potential and creating a voltage gradient especially at the ventricular level, whichis responsible for reentrant arrhythmias.

LQT Syndrome 1The causal gene for LQT1 is the KVLQT1 (or KCNQ1), which encodes a voltage-gated potassium channel subunitand is strongly expressed in the heart.342 It consists of 16 exons spanning 400 kb, which form 6 transmembranesegments. It coassembles with the subunit min K (KCNE1) to form the slow-activating potassium current IKs.

Mutations in this gene disrupt the normal function of the protein, causing a decrease in the potassium current.Several mutations have been described in KCNQ1 to date.

LQT Syndrome 2The LQT2 gene is HERG ("human ether-a-go-go related" gene), which was isolated in 1994 from hippocampus andnamed human ether-a-go-go related gene because of its homology to Drosophila "ether-a-go-go" gene. The gene islocalized on chromosome 7q35-q36. It contains 16 exons spanning approximately 55 Kb of genomic sequence. It

encodes a protein that forms six transmembrane segments.343 The protein is responsible for the rapidly activatingdelayed rectifier potassium current IKr after coassembly with MIRP1 (KCNE2). As in the case of LQT1, mutations in

HERG cause an abnormal protein with a resulting loss of potassium current.

LQT Syndrome 3

Droperidol

Adrenaline

Antibiotics

Clarithromycin

Erythromycin

Pentamidine

Trimethoprim-sulfamethoxazole

Ketoconazole

Fluconazole

Antihistamines

Terfenadine

Diphenhydramine

Antihyperlipidemic

Probucol

Central nervous system active drugs

Droperidol

Haloperidol

Pimozide

Risperidone

Gastrointestinal stimulants

Cisapride




The causal gene is SCN5A, located on chromosome 3, which encodes for the cardiac sodium channel.323 Mutations inSCN5A also cause Brugada syndrome and progressive conduction system disease, as discussed earlier.Electrophysiologic studies following expression of the mutant proteins in xenopus oocytes indicate a gain of functionmutation evidenced by delayed inactivation and persistent leaking of sodium ions after phase 0 of the actionpotential.

LQT Syndrome 4A locus for a French family with 65 affected members with LQT and sinus node dysfunction was mapped to 4q25-q27

in 1995.344 Very recently, the causal gene was identified as ANKB (also known as ANK2), which encodes ankyrin-B.345 Mutations in ankyrin B disrupt cellular localization of the sodium pump, the sodium/ calcium exchanger andinositol-1,4,5-triphosphate receptors, reduce their expression levels and affect Ca2+ signaling in adult cardiacmyocytes. This finding suggest that not only do mutations in ion channels cause cardiac arrhythmias, but alsomutations in proteins associated with ion channels, such as ankyrin B, could induce a similar phenotype.

LQT Syndrome 5MinK (minimal potassium ion channel), located on chromosome 21q22.1-q22.2, is the causal gene. It contains three

exons. MinK coassembles with KVLQT1 to form the cardiac IKschannel.342 Mutations in this gene have been identifiedas causing both the autosomal- dominant and autosomal-recessive disease.

LQT Syndrome 6LQT6 is caused by mutations in KCNE2 or MirP1 (minK-related peptide). It is mapped to 21q22.1, next to minK,

arrayed in opposite direction. KCNE2 assembles with HERG to form the IKr current.346 Mutations in KCNE2 decreasepotassium current availability, with slower activation.

LQT Syndrome 7Andersen syndrome is a rare autosomal-dominant inherited disorder characterized by constellation of periodicparalysis, cardiac arrhythmias, LQT, and dysmorphic features such as short stature, scoliosis, clinodactylism,

hypertelorism, low-set or slanted ears, micrognathia, and broad forehead.347. The causal gene is KCNJ2, located onchromosome 17q23. It encodes the inward rectifier potassium channel Kir2.1, expressed in skeletal and cardiacmuscles. Kir2.1 is a strong inward rectified channel that prevents passage of any current at potential greater than 40mV. Electrophysiologic studies indicate that the mutant protein exerts a dominant negative effect on Kir2.1 function,with an ultimate decrease in potassium current.

LQT Syndrome 8LQT8, also known as Timothy syndrome, is characterized by the presence of facial dysmorphic features, syndactyly,small teeth, mental retardation, and severe QT prolongation. Mutations in CACNA1C, encoding for the subunit of L-type calcium channel, have been identified as responsible for the syndrome. The mutations cause a gain-of-function

defect, increasing the inward current and prolonging the action potential.348

Long QT Syndrome 9Caused by mutations in Caveolin-3, this occurs in less than 2% of individuals with LQT syndrome. The effect isassociated with the role of this protein in modification of the Na channel. As in LQT3, mutations in Cav-3 prolong

repolarization by increasing the late INa.349

Long QT Syndrome 10This form of the LQT syndrome is caused by mutations in SCN4B, the -subunit of the sodium channel (NaV 4). Themutation induces a positive shift in inactivation of the Na channel, which increases INa.

350

Long QT Syndrome 11

In 2007, Chen et al351 identified a rare Yotiao (AKAP9) missense mutation in an LQTS family that disrupts bindingbetween KCNQ1 and Yotiao, reduces PKA phosphorylation of KCNQ1, eliminates the response of KCNQ1 to cAMP, andprolongs the action potential.

Long QT Syndrome 12

Ueda et al352 proposed recently a new susceptibility gene (SNTA1) for inherited LQTS. The mutation caused a markedincrease in late INa, comparable to the increases seen in patients with LQT3 mutations.353 The data, showing aplausible molecular mechanism for the effects of the mutation on the regulation and function of the cardiac sodiumchannel through the nNOS complex, further support the view that the mutation in this gene is pathogenic. If thegenotype-phenotype relationship is supported by further studies in additional patients, then SNTA1 would join the

LQT9 gene CAV3354 and the LQ10 gene SCN4B355 as a rare LQTS-susceptibility gene (LQT12) that, together withCAV3 and SCN4B, produce LQTS through actions on SCN5A to cause a net gain of function with increased late INa.




AUTOSOMAL-RECESSIVE LQT SYNDROME (JERVELL AND LANGE-NIELSENSYNDROME)

The autosomal-recessive forms of the LQT syndrome have been linked to mutations in the genes encoding IKscurrent, namely KVLQT1 and minK.356 For the LQT phenotype, which is also associated with sensorineuronaldeafness, to express, patients must inherit a mutation from both parents. Consequently, it is less common than theRomano-Ward syndrome but is associated with a more malignant course and a longer QT interval. The phenotypecould also arise in recessive forms when different mutations in the same gene are inherited from the parents(compound heterozygote).

GENOTYPE–PHENOTYPE CORRELATION IN LQT SYNDROMEGiven the availability of a large number of families with the LQT syndrome, several genotype–phenotype correlationstudies have been performed to identify the genetic determinants of triggering events, electrocardiographicphenotype, and response to therapy. The studies predominantly encompass the three most common forms of LQTsyndrome—LQT1, LQT2, and LQT3—and have significant limitations inherent to genotype–phenotype correlationstudies as described earlier. Despite their limitations, characteristic features have emerged that could guide the

analysis of patients toward a specific genetic defect (Table 82–14).357

Triggering EventsCharacteristic features have emerged that help in part guide the analysis of patients toward a specific genetic defect.In general, individuals with LQT1 exhibit symptoms during physical activity, especially in swimming activities.

Individuals with LQT3 are symptomatic during sleep.358

Electrocardiographic PhenotypesLQT1 patients have a T wave that usually begins just after the QRS, becoming long and broad based. LQT2 patients

have a small or notched T wave, and LQT3 patients show a very late T wave with a prolonged ST segment.359

Clinical Phenotypes According to GenotypeMutations also carry prognostic significance and in all three groups (LQT1, LQT2, and LQT3), there is a correlationbetween cardiac events and the QT interval. In general, patients with LQT1 and LQT2 have a higher risk of cardiacevents than patients with LQT3. The latter, despite having fewer events, have a relatively higher mortality, which

indicates higher lethality of the events.360

Biophysical PhenotypesThere have been few studies that have assessed the biophysical effect and location of the mutation and correlated itwith the severity of the phenotype. In 2002, a study made a correlation between the presence of a pore mutation in

KCNH2 and increased phenotype severity.361 In 2007, a new study associated the phenotype with the type andlocation of KCNQ1 mutations, showing that dominant-negative mutations and transmembrane mutations had a higher

incidence of events.362

Response to TherapyIn addition, response to drug therapy seems to be correlated with the genotype (class IIa). Although -blockers areconsidered the first line of therapy in patients with LQT1, they have not been shown to be beneficial in patients with

LQT3. Preliminary data suggest that LQT3 patients might benefit from Na1+ channel blockers, such as mexiletine, butlong-term data are not yet available (class IIb).363

INDUCED OR ACQUIRED LONG QT SYNDROMEInduced or acquired LQT syndrome is iatrogenic, caused by a long list of medications (see Table 82–13) andelectrolyte abnormalities. A large number of factors determine the risk of developing LQT syndrome in an individual inresponse to drug therapy. They include bioavailability of the drug, the interaction with other medications that affectthe same repolarizing current, and the presence of SNPs. SNPs play a major role in determining pharmacodynamicsand pharmacokinetics of drugs and thus the risk of LQT syndrome. The final effect on the repolarization will dependon the so-called repolarizing reserve, or the degree of alteration that the ionic currents can sustain before

Table 82–14. Genotype–Phenotype Correlation in Long-QT Syndromes

Phenotype Gene T Wave Trigger

LQT1 KCNQ1 Early onset, broad-base T Emotion, swimming

LQT2 HERG Low amplitude Auditory

LQT3 SCN5A Late T, normal amplitude Sleep




repolarization is compromised. Any combination of genetic and environmental factors (drug, electrolyteabnormalities) that decrease this repolarization reserve below a safe threshold will place the individual at risk of

arrhythmia.364

Epidemiologic studies have led to identification of mutations and SNPs in genes known to cause LQT syndrome. TheseSNPs are typically silent until unmasked by the use of IKr blockers. Thus one may consider these subjects as those

with unexpressed congenital LQT syndrome.365 Recent identification of a common SNP that predisposes to inducedarrhythmias in the African American population serves as an example.365 A recent study has provided the first link ofthe usefulness of genome-wide association studies in the identification of genes modulating cardiac repolarization.Genetic variants in the NOS1AP, encoding nitric oxide synthase 1, have been associated with a variation of 1.5% of

the QT interval in the population.366

Short QT SyndromeThe short QT syndrome is a newly described disease characterized by the presence of shortening of the QT intervalon ECG and clinically by episodes of syncope, paroxysmal atrial fibrillation, and/or life-threatening cardiacarrhythmias. Short QT syndrome usually affects young and healthy individuals with no structural heart disease. It

may be present in sporadic cases, as well as in families. It was originally described in 2000.367 In 2003, a link wasprovided between the short QT syndrome and familial sudden death with the first clinical report of two families with

short QT syndrome and a high incidence of SCD.368

CLINICAL MANIFESTATIONSMost patients with short QT syndrome have a history of familial sudden death and/or atrial fibrillation, short

refractory periods, and inducible ventricular fibrillation at electrophysiologic study.367,368 The age at onset of clinicalmanifestations could be extremely young. Malignant forms of short QT syndrome responsible for neonatal SCD have

been attributed to SIDS.368

The characteristic sign of the disease is the presence of a very short QT interval on ECG. The T wave remains upright,and the interval between the peak and end of the T wave is not prolonged. The appearance of a well-separated Uwave has also been reported in several cases. It is difficult to define the normal QT interval because the correctingequations have several limitations. Nevertheless, at a heart rate of 60 beats/min, the uncorrected QT interval is

usually higher than 360 milliseconds.369 From the data shown in the familial forms of the short QT syndrome, it isprobably reasonable to postulate that the presence of a QT of <330 milliseconds should raise high suspicion aboutthe disease. The severity of the clinical manifestations of short QTs is highly variable, ranging from asymptomatic toatrial fibrillation, recurrent syncope to sudden death.

MOLECULAR GENETICS AND ELECTROPHYSIOLOGYGenetics and biophysical analysis have provided important information regarding the pathophysiologic mechanismsthat cause the short QT syndrome. Short QT syndrome, as are the majority of primary familial electrical diseases, iscaused by mutations in genes encoding cardiac ion channels. Three genes have so far been discovered, thereby

proving that the disease is genetically heterogeneous.370-372

Short QT Syndrome 1

KCNH2 expresses a protein that makes up the channel responsible for the rapidly activating outward K+ currentinvolved in phase 3 repolarization. The protein is often referred to as HERG. The first genetic basis for the diseasewas obtained with the identification of two different missense mutations in the same residue in KCNH2 in three

unrelated families.370,373 Both mutations resulted in the substitution of asparagine for lysine at codon 588 (N588K),an area at the outer mouth of the channel pore.

Analysis of the current generated after transferring the mutated channels into human mammalian cells showed thatthe mutation abolished the inactivation of the channels, thus resulting in an increased developing current. Thebiophysical analysis therefore showed that the mutation induced a gain of function in IKr current, thus causing a

shortening of the action potential.370

The presence of paroxysmal atrial fibrillation in some affected individuals and especially in one of the families as theonly clinical manifestation suggested that the increased heterogeneity would also be present at the atrial level andcould be responsible for the arrhythmia.

Short QT Syndrome 2

The KCNQ1 gene encodes a subunit of the proteins responsible for the slowly activating delayed outward K+ current.A missense mutation in this gene causing short QT syndrome was first identified by Bellocq et al371 in a 70-year-oldindividual who suffered ventricular fibrillation and had a QT interval of 290 milliseconds after resuscitation. He wasnot inducible at electrophysiologic study and had no cardiac structural abnormalities. A second mutation in KCNQ1




was identified in a baby girl born at 38 weeks after induction of delivery that was prompted by bradycardia and

irregular rhythm.374 The ECG revealed atrial fibrillation with slow ventricular response and short QT interval.

Biophysical analysis showed that both mutations were leading to a gain of function in the outward current thatexplains the short QT syndrome phenotype.

Short QT Syndrome 3The third form of short QT syndrome has been linked to mutations in the KCNJ2 gene, which codes for the channel

protein responsible for the inward rectifier current.372 The proband and her father with the mutation both displayedshort QT intervals of 315 and 320 milliseconds, respectively. ECG recordings showed asymmetrical T waves with anabnormally rapid terminal phase. When expressed in Chinese hamster ovary cells, the mutated channels generatedelectrical currents that did not rectify (decreased) as much as the normal channels in their functional positive rangeof potentials (80- 30 mV). Such a range of voltages corresponds to the very end of phase 3 repolarization and tophase 4. Simulation of the effects of the mutated channels on the morphology of the ventricular action potentialshowed a selective speeding of late repolarization, thus shortening significantly the action potential duration at 90%repolarization.

PHENOTYPE–GENOTYPE CORRELATIONRobust genotype–phenotype correlation data are not yet available for short QT syndrome. The disease is clinically

highly heterogeneous, as indicated by the fact that in the three families with the same mutation,370,373 there istremendous variation in symptoms and presentation.

TREATMENT STRATEGY

Preliminary data show that there may be effective pharmacologic therapy for this disease (class IIb).375,376 However,the high incidence of SCD warrants the implantation of an intracardiac cardioverter defibrillator, especially inindividuals with aborted sudden death.

Because QT shortening is likely caused by an increase in the outward current, it was suggested that blocking thecurrent with class III antiarrhythmic drugs (known to increase the QT interval) could be a potential therapeutic

approach for the treatment of short QT syndrome (class IIb). More recently, in a clinical study, Gaita et al375 showedthat treatment with quinidine prolonged the QT interval, decreased inducibility, and therefore had the potential to bean effective therapy for these patients (class IIb). Clinical follow-up in one family with paroxysmal atrial fibrillation

indicates that the episodes respond well to treatment with class Ic agent propafenone (class IIb).373

Progressive Familial Heart BlockFamilial heart block is autosomal-dominant progressive disease of cardiac conduction system characterized by initialdevelopment of bundle branch block and gradual progression to complete heart block. Two forms have beenrecognized. In type I, the onset is early and the disease is rapidly progressive. In type II, the onset is later in life,and commonly the QRS complex is narrow and AV nodal block predominates. Clinical features of the disease includesyncope, SCD, and Stokes-Adams attacks. A locus was identified in a large family of Portuguese descent on

chromosome 19q13.377 The gene has not been identified yet. As discussed earlier, mutations in SCN5A have beenshown in some families with familial heart block.378 In addition, AV block in conjunction with congenital heartdisease, such as ASD (NKX2.5 mutations) and DCM (lamin A/C mutations), have been described; these werediscussed earlier.

Catecholaminergic Polymorphic Ventricular TachycardiaRyanodine receptors are responsible for release of calcium from the sarcoplasmic reticulum and are activated by the

incoming calcium; therefore, they are Ca2+-activated Ca2+ channel. Mutations in ryanodine receptors (RYR2) havebeen shown to cause a phenotype electrically resembling ARVC (phenocopy),379 as discussed earlier, and familialpolymorphic ventricular tachycardia.379

Familial polymorphic ventricular tachycardia is an autosomal-dominant inherited disease with a mortality rate ofapproximately 30% by the age of 30 years. Phenotypically, it is characterized by runs of bidirectional andpolymorphic ventricular tachycardia in response to vigorous exercise in the absence of evidence of structuralmyocardial disease.

A recessive form of familial polymorphic ventricular tachycardia also has been described and mapped to 1p13.3-

p11.380 Mutation screening identified a missense mutation in calsequestrin 2 (CASQ2) as responsible for the disease.CASQ2 is involved in the same pathway as RYR2 to control calcium release from the sarcoplasmic reticulum.

Sick Sinus SyndromeSick sinus syndrome is characterized by the occurrence of sinus bradycardia, sinus arrest, and chronotropic




incompetence. Sinus node dysfunction has been linked to loss-of-function mutations in HCN4381,382 and in recessiveform to SCN5A.383 HCN4 contributes to native f-channels in the sinoatrial node, the natural cardiac pacemakerregion. In 2006, a loss-of-function defect in HCN4 was also linked to familial sinus bradycardia.384

Familial Atrial FibrillationAtrial fibrillation (AF) is the most common sustained rhythm disorder or arrhythmia encountered in clinical practice,with a prevalence of 1% in the general population, which increases with age to approximately 6% in people over the

age of 65 years.385 It is responsible for more than one-third of all cardioembolic episodes.386

AF was not generally appreciated to be a familial inherited disorder. AF was first reported as a familial form in

1943.387 In last 5 years, published studies have shown several lines of evidence supporting the geneticallydetermined heritable basis to AF.388,389 A study by Darbar et al390 indicated that the familial form of the diseasemay have a higher prevalence than previously suspected and emphasizes the importance of expanding geneticsstudies. Familial AF may be a monogenic disorder and is often identified as AF being present in many members of the

same family. The first locus for familial atrial fibrillation was identified in 1997, in 10q22.391 Several other loci andgenes have been identified since then.392-394

ATRIAL FIBRILLATION ASSOCIATED WITH GENETIC ALTERATIONS INPOTASSIUM CURRENTS

Most of the single-gene mutations that have been discovered in families with AF cause cardiac K+-channel defects.The first gene responsible for familial AF, KCNQ1, was identified in 2003, linking the disease to an ion

channelopathy.395KCNQ1 encodes the pore-forming -subunit of the cardiac slow delayed-rectifier (IKs) channel, and

its loss of function had been previously associated with long QT syndrome. The analysis of KCNQ1 identified amissense mutation resulting in the amino acid change S140G. Electrophysiologic studies revealed a gain of function inIKs current when the mutated channel was expressed with the subunits minK and MirP1. This gain of function

explained well the shortening of the action potential duration and effective refractory period, which are thought to bethe culprits of the disease. In an interesting new finding, a gain-of-function mutation in codon 141, next to the onedescribed in the above-mentioned family, was found to be responsible for a severe form of AF in utero and short QT

syndrome.374 In a family with AF, Otway et al396 identified a mutation in KCNQ1 (R14C) that induces a mutantprotein that only showed increase IKs upon cell stretch, with a hypotonic solution opening a new and stimulating

debate on the role of genetic-environmental interaction in the development of the disease.

Identification of mutations in KCNE2 in two families with AF397confirmed that defects in genes encoding for potassiumcurrents are responsible of AF. The mutation R27C caused a gain of function when coexpressed with KCNQ1 but hadno effect when expressed with HERG.

A third genetic defect was described in KCNE3398; however, the functional analysis did not demonstrate a differentbiophysical effect caused by the mutant genetic defect, indicating that it could be a rare polymorphism.

Finally, a gain-of-function mutation in Kir2.1 caused by a mutation in KCNJ2399 was found in 2005 in a new kindred.The biophysical findings therefore indicated a role of gain-of-function mutations in potassium channels in AF,

highlighting the pathophysiologic role of shortened atrial action potentials. When Olson et al400 described a loss-of-function mutation in KCNA5, the gene that encodes KV1.5, the debate became more stimulating, as it initiated thehypothesis that a prolongation of the action potential can also be a basic mechanism for AF progress.

ATRIAL FIBRILLATION ASSOCIATED WITH GENETIC ALTERATIONS IN SODIUMCURRENTThe -subunit of the cardiac sodium channel gene, SCN5A, responsible for phase 0 of the cardiac action potential,has been studied extensively. Because of its key role in phase of the cardiac action potential, SCN5A is involved inseveral primary arrhythmia syndromes. Gain-of-function mutations in SCN5A, mainly due to its inability to inactivate,have been associated with long QT syndrome type 3, and loss-of-function mutations have been associated with

Brugada syndrome, SIDS, Lev-Lenègre syndrome, SUDS in Southeast Asia, and DCM.401 Variants in SCN5A havebeen linked with lone and familial AF402 and also associated with LQT syndrome.403

ATRIAL FIBRILLATION ASSOCIATED WITH GENETIC ALTERATION IN NON-IONCHANNELSAtrial natriuretic peptide precursor (NPPA) encodes atrial natriuretic peptide (ANP). ANP modulates ionic currents incardiac myocytes and can play a role in shortening of the atrial conduction time, which could be a potential substrate

for atrial reentrant arrhythmias. In 2008, Hodgson-Zingman et al404 identified a frameshift mutation in NPPA in alarge family with AF.

The latest gene to be linked to the disease is the one identified by Zhang et al.405 The clinical phenotype was




characterized by a neonatal onset, with autosomal-recessive inheritance. They identified a mutation in NUP155, whichencodes a member of the nucleoporins. Although still unknown, the mechanism by which NUP155 may be associatedwith AF could be related to modulation of calcium handling proteins and ion channel and expression of its possibletarget genes, such as HSP70. The mutation was associated with inhibition of both export of HSP70 mRNA and nuclearimport of Hsp70 protein, indicating that loss of function of NUP155 caused the disease by altering mRNA and proteintransport. This gene is located in 5p13 and had been also associated with sudden death in the family.

SOMATIC MUTATIONS IN ATRIAL FIBRILLATIONIn 2006, three missense mutations in GJA5 were identified in atrial tissue specimens from patients with idiopathic

AF.406GJA5 encodes the gap junction protein connexin 40, which is involved in electrical coupling, and its disruptionmay cause atrial arrhythmias.

Monomorphic Ventricular TachycardiaA somatic point mutation (F200L) in the inhibitory subunit 2 of G protein was identified in a patient with sustainedmonomorphic ventricular tachycardia that was unresponsive to vagal maneuvers and adenosine.407 The mutation waspresent in cardiac tissue at the arrhythmogenic locus and was shown to increase intracellular cAMP concentration andinhibit suppression of cAMP by adenosine.

Familial Wolff-Parkinson-White SyndromeFamilial WPW syndrome is a rare syndrome with an autosomal-dominant mode of inheritance. It occurs in isolation orin conjunction with other disorders, such as HCM and Pompe disease. It is characterized by evidence of preexcitationon electrocardiogram, palpitation, and syncope as a result of supraventricular arrhythmias. The phenotype of WPW inconjunction with HCM and conduction defect was found in patients with mutations in PRKAG2, as discussed

earlier.408-410 It has also been reported in patients with Pompe disease caused by mutations in -1,4-glucosidase,411

in patients with HCM caused by mutations in TNNI3 and MYBPC3, and in Leber hereditary optic neuropathy, which is

caused by mutations in mitochondrial DNA.412

GENETIC BASIS OF CARDIAC DISEASE IN CONNECTIVE TISSUE

DISORDERS

Marfan SyndromeMarfan syndrome is a primary disorder of connective tissue characterized by cardiovascular, ocular, and skeletal

abnormalities.413 There is significant variability in the clinical manifestations of Marfan syndrome, but thepredominant features are progressive dilatation of the aortic root, aortic aneurysm, dissection, and aortic and mitral

valve regurgitation. The estimated incidence of Marfan syndrome is 1 per 5000 population.414 The age of onset ofclinical manifestations of Marfan syndrome is variable, but cardiac phenotypes commonly occur in the third or fourthdecades of life. Aortic dissection is the leading cause of premature death in patients with Marfan syndrome. Inaddition to cardiovascular abnormalities, marfanoid habitus (increased height, disproportionately long limbs anddigits), lens dislocation or subluxation, arachnodactyly, thoracic abnormalities, and increased joint laxity are commonclinical features (see also Chap. 106).

GENETIC BASIS OF MARFAN SYNDROMEMarfan syndrome is an autosomal-dominant disease that exhibits locus and allelic heterozygosity. The first causal

gene to be identified is the FBN1, which is located on 15q15.23 and encodes fibrillin.414,415 Fibrillin is a cysteine-richprotein with a molecular mass of 350 kDa; it is the major component of extracellular microfibrils in both elastic andnonelastic connective tissues. More than 600 nonrecurring unique mutations in FBN1 have been described that

encompass missense, nonsense, and deletion mutations, as well as abnormal splicing or exon skipping.416 Mutationsare spread throughout most of the gene, and the frequency of each particular mutation is relatively low, which makesscreening for mutations tedious.

There is a significant variability in the phenotypic expression of Marfan syndrome. The phenotypic variability may bepartly caused by locus and allelic heterogeneity and partly by the effect of modifier genes and perhaps environmentalfactors. The clinical spectrum varies from ectopia lentis in the absence of any other phenotype to neonatal Marfansyndrome and premature death, often within the first 2 years of life. Mutations inducing premature termination of theprotein result in approximately a 50% reduction in the level of fibrillin and more frequent ocular manifestations.

A phenocopy or a variant of Marfan syndrome is congenital contractural arachnodactyly. It is characterized by severekyphoscoliosis, generalized osteopenia, flexion contractures of the fingers, abnormally shaped ears, and, lessfrequently, mitral regurgitation and congenital heart disease. Recently, point mutations in the FBN2 gene have been

described as causes of contractural arachnodactyly.417FBN2 mutations clustered in limited regions alter amino acidsin the calcium-binding consensus sequence in the epidermal growth factor–like domains. Mutations affect either the




conserved cysteine residues or residues of the calcium-binding consensus sequence of the cbEGF motifs and oftenresult in premature termination of the protein. In addition, mutations in transforming growth factor (TGF) receptor1 (TGFBR1) and TGF receptor 2 (TGFBR2) have been identified in individuals with Marfan-like syndrome.418

The pathogenesis of Marfan syndrome entails decreased expression levels of the fibrillin protein and reduceddeposition of fibrillin in vascular adventitia, which results in weakening of the adventitia and aneurysm formation.

Recently, increased TGF signaling was detected in a mouse model of aortic aneurysm.419 Treatment with aangiotensin II receptor blocker, known to inhibit TGF , prevented aortic aneurysm formation in this mouse model.419

Ehlers-Danlos SyndromeEhlers-Danlos syndrome (EDS), a relatively uncommon disorder, encompasses a group of conditions characterized by

increased elasticity of the skin and connective tissue diseases (see also Chap. 84).413,420 The classic form of EDS ischaracterized by joint hypermotility and fragile, bruisable skin that heals with peculiar "cigarette-paper" scars. Otherclinical features include translucent elasticity of the skin, mitral valve prolapse, spontaneous rupture and aneurysm oflarge arteries, kyphoscoliosis, atrophic scars, and hematomas in the joint areas, especially in the knees and elbows.In the severe form, spontaneous rupture of the intestines and arteries is common. In the benign form, the onlymanifestations may be hyperextensibility of joints and easy bruisability. The age of onset of clinical manifestations isvariable and ranges from childhood to late adulthood.

GENETIC BASIS OF EHLERS-DANLOS SYNDROMEEDS has several different forms that are inherited in three different patterns of transmission—autosomal-dominant,

autosomal-recessive, and X-linked recessive.413,420 Cardiovascular abnormalities are more common in forms I and IVand include congenital malformations, such as tetralogy of Fallot, atrial septal defects, and valvular abnormalities

such as mitral and tricuspid valve prolapse.413

Mutations in genes encoding collagen components are responsible for EDS.420 For example, Ehlers-Danlos type IV,which is considered the most malignant form because of proneness to spontaneous rupture of the bowel and large

arteries and a high incidence of pregnancy-related complications, is caused by mutations in COL3A1.421 The gene islocated on 2q31 and encodes type III procollagen. Cardiac manifestations include aortic and coronary arteryaneurysms with a high incidence of rupture. Some case of EDS type I are caused by mutations in COL5A2, coding for

collagen -2(V), and COL1A1, encoding collagen -1(I).413 EDS is considered a primary disorder of collagendeficiency. Point and deletion mutations lead to either deficiency of collagen-processing enzymes, haploinsufficiency,or expression of dominant-negative collagen chains. Consequently, there is decreased collagen synthesis and lossof connective tissue resiliency.

Ellis–Van Creveld SyndromeThis syndrome is discussed above, in the text concerning congenital heart diseases.

Cutis LaxaCutis laxa comprises a heterogeneous group of acquired and genetic disorders characterized by redundant, wrinkled,loose, sagging skin that slowly returns to normal after stretching. Cardiac manifestations include pulmonic stenosis,aortic aneurysms, and right-sided heart failure. Vessels are very tortuous, resembling corkscrews on the angiogram.

Autosomal-dominant, autosomal-recessive, and X-linked forms have been described, and mutations in the elastin

gene (ELN) have been identified in the autosomal-dominant form.422 A homozygous missense mutation in the fibulin-5 (FBLN5) gene has been identified as responsible for the recessive form.423

Pseudoxanthoma ElasticumPseudoxanthoma elasticum is a genetic disorder characterized by dermatologic, ocular, and cardiovascularabnormalities resulting from degeneration of the elastic fibers. Manifestations include pseudoxanthoma, especially inareas of the neck and axillae, angioid streaks in the optic fundus, and gastrointestinal hemorrhagic and occlusivedisease. Cardiovascular abnormalities include calcification of the peripheral arteries, with resulting intermittentclaudication, coronary artery disease, mitral valve prolapse, and hypertension.

Recently, mutations in the ATP-binding cassette (ABC) transporter gene (ABCC6) on chromosome 16p13 were

identified as the cause of pseudoxanthoma elasticum.424 The exact biologic function of ABCC6 protein and themechanism(s) by which mutations in ABCC6 cause pseudoxanthoma elasticum are unknown.

Osteogenesis ImperfectaOsteogenesis imperfecta comprises a heterogeneous class of connective tissue disorders characterized by bonefragility. Bone fragility results from defective collagen synthesis, which leads to decreased bone mass, disturbedorganization of bone tissue, and altered bone geometry (size and shape). Cardiovascular abnormalities include




valvular lesions, such as mitral and aortic regurgitation, and an increased fragility of the blood vessels.

Mutations in one of the two genes encoding type I collagen, namely COL1A1 and COL1A2, have been identified as the

cause.425,426

GENETIC DISORDERS OF THE PULMONARY CIRCULATION

Familial Primary Pulmonary HypertensionPrimary pulmonary hypertension (PPH) is diagnosed when mean resting pulmonary artery blood pressure is greaterthan 25 mm Hg in the absence of known secondary causes such as lung disease or pulmonary venous congestionsecondary to heart failure (see also Chap. 71). Dyspnea is the most common symptom; it is the first symptom in

60% of the cases.427 Other clinical manifestations include exercise intolerance, fatigue, cyanosis, syncope, andSCD.428 Clinical manifestations often start in the third and fourth decades of life and are about twice as common infemales.427 Median survival in those with established PAH is approximately 3 years.429 Prevalence is 1 to 2 per1,000,000 individuals.428

PPH is a familial disease with an autosomal-dominant mode of inheritance in 5% to 10% of cases.427,430 PPH is agenetically heterogenous disease. Mutations in bone morphogenic protein receptor type II (BMPR2), mapped tochromosome 2q31-33, is responsible for approximately 50% of familial PPH and 10% to 15% of sporadic

cases.428,430-433 The spectrum of mutations includes nonsense and frameshift mutations, expected to producedysfunctional protein. Bone morphogenic protein (BMP) receptor is a cell-surface receptor that belongs to the TGFfamily.433 Binding of ligands to BMPR2 activates signaling through Smad molecules. Mutations in another member ofthe TGF receptor family, namely, activin-receptor–like kinase 1 (ALK1), are responsible for a small fraction offamilial PPH.434 Finally, an association between serotonin transporter (5-HTT) SNPs and pulmonary hypertension hasbeen documented.435

The pathogenesis of pulmonary hypertension caused by mutations in BMPR2 includes haploinsufficiency, whereindefective TGF signaling via Smad molecules results in proliferation of smooth muscle cells and reducedapoptosis.433,436,437 Molecular pathogenesis of PPH as a consequence of mutations in ALK1 and serotonin transporteralso entails smooth muscle cell proliferation.

MONOGENIC LIPID DISORDERS

The majority of common dyslipidemias are complex traits caused by interaction of multiple genes and environmentalfactors. SNPs in a variety of genes encoding protein components of cholesterol and fatty acid biosynthesis have beenimplicated in susceptibility to dyslipidemia; these are discussed elsewhere (see Chap. 52). Monogenic forms ofdyslipidemias are described briefly below.

Familial HypercholesterolemiaFamilial hypercholesterolemia (FH) is an autosomal-dominant disorder with a prevalence of 1 in 500 in the mild formand in 1 in 100,000 people in its severe form. It is characterized by severely elevated plasma levels of low-density

lipoprotein cholesterol (LDL-C; type IIa hyperlipidemia) and premature atherosclerosis.438 Plasma levels of totalcholesterol are in the range of 300 to 400 mg/dL in affected heterozygous individuals and greater than 500 mg/dL inhomozygous subjects. The affected individuals develop severe atherosclerosis involving multiple vascular territories,tendon xanthomata, and corneal arcus. Subjects homozygous for the causal mutations exhibit clinical atherosclerosisin the first or second decades of life and heterozygous subjects in the fourth or fifth decades. These patients oftensuffer from ischemic symptoms and/or cardiac events requiring revascularization procedures very early in life (seealso Chap. 64).

The causal gene is LDLR, which is located on chromosome 19p13.439 It encodes LDL-C receptors. More than 1000point, deletion, and splice mutations in LDLR have been identified in patients with FH. Approximately 60% of themutations are missense mutations, 20% are minor rearrangements, 13% are major rearrangements, and 7% are

splice-junction mutations.440 Mutations cause FH by perturbing the function of LDL-C receptors. Mutations couldaffect synthesis and targeting to the cell membrane, binding of the receptor to LDL-C, internalization of the receptorafter binding to LDL-C, and recycling of the receptors. The ensuing biologic effect is impaired removal of

apolipoprotein B (apoB) and apoE from the circulation.439 Mutations affect LDL-C receptor function to variabledegrees, leading to variable clinical manifestations.441 In general, there is an inverse correlation between plasmalevels of LDL-C and the level of residual LDLR activity. Mutations that completely inactivate the receptors lead tosevere premature atherosclerosis in childhood. Frameshift mutations, by markedly altering the structure of the LDL-Creceptors, cause severe phenotype. In contrast, mutations that partially inactivate the receptors cause mild tomoderate hypercholesterolemia. Thus the development and severity of coronary atherosclerosis vary according tocausal mutations, that is, residual LDL-C receptor activity. Genetic background, diet, environmental factors, andepigenetic factors are also likely to contribute to the phenotype.




Other forms of FH are type B hypercholesterolemia or familial defective apolipoprotein B and autosomal-dominanthypercholesterolemia type 3, which are discussed separately.

Familial Defective Apolipoprotein B100Familial defective apolipoprotein B100 (FDB) is an autosomal- dominant disease, expressed as increased plasma

levels of LDL-C and very low-density lipoprotein C (VLDL-C).442,443 Phenotypically it is similar to heterozygous FHand includes premature coronary artery disease and tendon xanthoma, in addition to elevated plasma levels of LDL-C. However, the LDL receptor activity is normal in these individuals. FDB is a relatively common disorder, with anestimated frequency of 1 in 1000 people, but the prevalence varies worldwide. The frequency of the mutation variesin different regions of the world.

The causal gene is APOB, located on chromosome 2q24.444 The causal mutation involves amino acid 3500 in morethan 99% of cases, with the predominant mutation being R3500Q and, rarely, R3500W.429-431 Mutations decreasethe affinity of LDL receptors for apolipoprotein B, which result in accumulation of VLDL-C and LDL-C in the plasmaand blood vessels.

Autosomal-Dominant Hypercholesterolemia Type 3The phenotype is characterized by severely elevated plasma LDL-C levels and is similar to that in FH and FDB. Thecausal gene is PCSK9, which encodes proprotein convertase subtilisin/kexin type 9, also known as neural apoptosis-

regulated convertase (NARC1).445 Pathogenesis of autosomal-dominant hypercholesterolemia type 3 involvesenhanced degradation of the LDL receptors by PCSK9.446 It is also noteworthy that SNPs in PCSK9 are alsoassociated with plasma LDL-C levels and risk of coronary artery disease in the general population.447,448

Autosomal-Recessive HypercholesterolemiaAutosomal-recessive hypercholesterolemia is a rare disease with a phenotype similar to FH. The causal gene is ARH,which encodes an adaptor protein that binds to LDL receptors and hence is called LDL receptor adaptor protein 1

(LDLRAP1).449 The protein contains a phosphotyrosine-binding domain that interacts with the cytoplasmic tail of LDLreceptors. The pathogenesis of the phenotype involves impaired clearance of plasma LDL-C, despite normal activity ofLDL receptors.

HypobetalipoproteinemiaHypobetalipoproteinemia, or abetalipoproteinemia, is a rare disease characterized by extremely low plasma levels of

apolipoprotein B, total cholesterol, and LDL-C.450-452 High-density lipoprotein cholesterol (HDL-C) levels are high,and atherosclerosis is very uncommon. The phenotype often presents in childhood with failure to thrive, fatmalabsorption, celiac disease, vitamin A and E deficiency, ataxia, demyelination of the central nervous system, and

low plasma LDL-C levels.451 Sporadic and familial cases with an autosomal-dominant mode of inheritance have beenreported. A causal gene is MTTP on chromosome 4q22-24, which encodes the microsomal triglyceride transfer

protein.442,453 MTTP is a heterodimer of a unique large subunit and the protein disulfide isomerase, which catalyzesthe transport of triglyceride, cholesteryl ester, and phospholipid from phospholipid surfaces. Mutations encode

truncated nonfunctional protein, thus leading to very low levels of apoB, LDL-C, and total cholesterol.453

Familial hypobetalipoproteinemia also can arise because of truncation mutations in APOB.454 It is also a rare disordercharacterized by very low plasma levels of LDL-C and total cholesterol.451

Another form of hypobetalipoproteinemia is chylomicron retention disease, which is an autosomal-dominant disease

characterized by the selective absence of apoB-48.451 Chylomicrons are absent in plasma of the affected individualsafter a fat-containing meal. The phenotype is characterized by steatorrhea, growth retardation, malnutrition, and

acanthocytosis. The causal gene is SAR1B (SARA2) located on chromosome 5q31.455 The encoded protein is involvedin intracellular trafficking of proteins in COP (coat protein complex) coated vesicles.442

Fish-Eye DiseaseFish-eye disease is a rare autosomal-dominant condition caused by deficiency of lecithin:cholesterol acyltransferase

(LCAT).456,457 The LCAT gene is located on chromosome 16q22.1 and codes for a protein involved in the synthesisfrom pre-alpha lipoprotein A1 and conversion of HDL3 to HDL2 cholesterol. Deficiency of LCAT leads to premature

coronary atherosclerosis, proteinuria, anemia, renal failure, and corneal opacification.

Tangier DiseaseTangier disease is an autosomal-codominant disease characterized by the virtual absence of HDL-C and very lowplasma levels of apoAI. Deposition of cholesteryl esters results in characteristic hypertrophic orange-colored tonsils,

hepatosplenomegaly, and premature coronary artery disease.458 Mutations in the ATP-binding cassette transporter(ABCA1) gene cause Tangier disease and familial hypoalphalipoproteinemia, its allelic variant.459-462ABCA1 gene is




located on chromosome 9q31 and codes for an mRNA of 6783 bp and a protein of 2261 amino acids.459,461,462

ABCA1 is a transmembrane protein with 12 transmembrane domains. It acts as a flippase at the plasma membrane,

stimulating cholesterol and phospholipid efflux to apoAI and HDL-C.458 Normally, ABCA1 transports free cholesterolto the extracellular space where it binds to apoAI synthesized by the liver and forms nascent HDL particles fromVLDL. In the absence of ABCA1, free cholesterol is not transported extracellularly and lipid-poor apoAI rapidlydegrades. Common polymorphisms in the ABCA1 gene have been associated with coronary atherosclerosis in the

general population.463-465

MONOGENIC FORMS OF HYPERTENSION

The predominant form of hypertension is essential hypertension, which accounts for 95% of all cases. Hypertension isa complex phenotype caused by the interactions of multiple genes and environmental factors. Several genes, inparticular those coding for the components of the renin–angiotensin–aldosterone system, have been implicated in

essential hypertension.466-468 They are discussed in Chap. 70; only monogenic forms of hypertension are describedbriefly here.469

Glucocorticoid-Remediable AldosteronismGlucocorticoid-remediable aldosteronism is a rare autosomal-dominant disorder and the first described familial form

of hyperaldosteronism.468 It is caused by a chimeric mutation that joins the promoter region of the 11 -hydroxylase(CYP11B1) gene to the coding region of the aldosterone synthase (CYP11B2) gene.470 The new chimeric gene,located on chromosome 8q24, lacks the negative feedback regulation imparted by angiotensin II. The promoter of thefusion gene, which is made up of the 5' fragment of CYP11B1 gene, is responsive to adrenocorticotropic hormone.Thus expression of aldosterone remains unchecked, and excess aldosterone synthesis leads to the retention ofsodium and salt and consequent hypertension. Glucocorticoid-remediable aldosteronism responds to treatment withglucocorticoids, which suppress the production of adrenocorticotropic hormone. Alternatively, treatment withmineralocorticoid receptor blockers also controls the hypertension.

Apparent Mineralocorticoid ExcessApparent mineralocorticoid excess is a rare autosomal-recessive disease of peripheral metabolism of cortisol. Clinicalmanifestations of apparent mineralocorticoid excess, in addition to hypertension, include hypokalemia, low plasmarenin activity, and responsiveness to spironolactone. There are two types of apparent mineralocorticoid excess,defined on the basis of severity of the biochemical phenotype. Both clinical variants are caused by mutations in the

HSD11B2 gene on chromosome 16q22, which encodes 11 -hydroxysteroid dehydrogenase II.471,472 The enzyme isresponsible for the peripheral conversion of biologically active cortisol to inactive cortisone. Point mutations inHSD11B2 reduce or abolish the activity of 11 -hydroxysteroid dehydrogenase in the conversion of cortisol tocortisone. Thus cortisol accumulates, leading to retention of salt and fluid through activation of mineralocorticoid

receptors and hypertension.459 Accordingly, patients with apparent mineralocorticoid excess respond to blockade ofmineralocorticoid receptors.

Liddle SyndromeLiddle syndrome is a rare autosomal-dominant disease characterized by hypertension, hypokalemic metabolic

alkalosis, low plasma renin activity, and suppressed aldosterone secretion.473 The phenotype usually develops earlyin life, and hypertension is frequently severe. The first gene identified was the SCNN1B, located on locus 16p12,

which encodes the subunit of the amiloride-sensitive Na1+ channel.473 The renal epithelial Na1+ channel has threesubunits: , , and . Subsequently, mutations in the subunit of epithelial sodium channels were also identified.474

The mutations activate the channel (gain-of-function mutations) and lead to sodium retention and hypertension.

Pseudohypoaldosteronism Type IIPseudohypoaldosteronism type II, also known as the Gordon hyper-kalemia–hypertension syndrome, is a rareautosomal dominant disorder characterized by hypertension and hyperkalemia early in life, mild hyperchloremia,metabolic acidosis, and suppressed plasma renin activity. Two causal genes for pseudohypoaldosteronism type II—

WNK4 on chromosome 17q21 and WNK1 on chromosome 12p—have been identified.475 WNK4 and WNK1 encodeserinethreonine kinases expressed in the distal nephron.475 Missense and deletion mutations exert a gain-of-functioneffect, increasing the expression levels of the proteins in the kidney and leading to increased renal salt reabsorption

and reduced renal K+ excretion.475

GENETICS OF CORONARY ARTERY DISEASE: COMMON GENETIC RISK

VARIANTS

Genetics Necessary for Prevention and Management




The results of modifying lifestyle and drug intervention have documented the importance and the effect of bothprimary and secondary prevention in common polygenic disorders such as coronary artery disease (CAD) and relatedtraits. Drug therapy alone has been shown repeatedly in multiple clinical trials to decrease the incidence of clinical

events from CAD by at least 30% to 40%.476,477 It has been stated476 that CAD could be eliminated in the 21stcentury just by treating the risk factors. However, it is well recognized, as indicated below, that at least 50% of

susceptibility for CAD is due to a genetic component.478 Furthermore, most risk factors such as hypertension,hypercholesterolemia, and obesity have an inherent significant genetic component. Identification of the genes that

predispose to CAD and related traits may be necessary for comprehensive prevention of these traits and CAD.479

Second, elucidation of the molecular pathways through which these genes mediate their effects should provide

targets for development of new therapies. The discovery of 9p21480 (discussed later) illustrates both of these points,namely, a risk factor, independent of all known risk factors, exerting its predisposition through a mechanism as of yetunknown. Increasing knowledge of pharmacogenetics indicates that the variable response to therapy for CAD is in

large part genetically determined.481 In the deCODE study of the Icelandic population, 20% of individuals withhypercholesterolemia did not have a significant response to statin.482 It is estimated that 20% of individuals areresistant to aspirin.483 Secondary prevention after a myocardial infarction, although effective, is far from adequate,as assessed from a public health viewpoint. Finally, identification of genetic risk variants is a prerequisite for

personalized medicine.479

Evidence Supporting a Genetic Basis for Coronary Artery DiseaseFAMILIAL AGGREGATION STUDIESThe power of a family history of CAD is strikingly illustrated in the state of Utah. Fourteen percent of the population ofUtah has a family history of heart disease, and it is in this cohort that 72% of all premature heart disease occurs and

48% of all cases of CAD.484 Similarly 11% of the population has a history of stroke, and it is in this cohort that 86%of premature strokes occur in Utah.484 Case-control family studies show a two- to three-fold increase in risk for CADin first-degree relatives.485-489 Family history of CAD in a first-degree relative before the age of 60 years is anindependent risk factor for early myocardial infarction (MI), even after controlling for traditional risk factors.490,491

Several prospective studies have shown up to a two-fold increase in CAD risk associated with a family history of CAD

after adjusting for traditional risk factors.492-497 There is also a clustering of susceptibility to CAD in families thathave risk factors associated with abnormalities such as lipid metabolism, hypertension, diabetes, and obesity,

indicating a genetic basis for these conditions and risk factors.498-503 The extent of coronary occlusion in patientswith CAD also relates to a parental history of MI.504 In families with CAD onset before age 46, heritability is 92% to100%, whereas within families of older cases, heritability is 15% to 30%.505 Premature CAD in the young istransmitted as an even greater genetic load to their offspring.506 The Danish twin registry of 8000 twin pairs shows ahigher incidence of CAD and deaths in monozygotic twins than in dizygotic twins: 44% versus 14%.507

GENETIC FACTORS PREDOMINATE IN PREMATURE CORONARY ARTERYDISEASE

CAD incidence increases with age; however, 10% of patients are diagnosed before the age of 50 years.508,509 In astudy of 207 cases of MI occurring before the age of 55 years and 621 matched controls, a family history of MI

occurring in a first-degree relative before the age of 55 years increased the risk of MI by 7.1-fold.510 This studyestimated heritability for early onset of CAD at 0.63; after exclusion of apparent lipid abnormalities, the heritabilityestimate was 0.56, suggesting that more than half of CAD diagnosed before the age of 55 years is genetic. Inanother study of 916 MI cases and 1106 controls, if an MI occurred early (<55 years) and in at least two relatives,

the relative risk was elevated 20-fold.511 Early-onset CAD occurs eight times more frequently in men than in women,suggesting that women are protected from early onset.508 Conversely, early-onset CAD in women may be associatedwith additional genetic factors compared with those that predispose men.

SINGLE GENE VERSUS POLYGENIC DISORDERSThis chapter has so far detailed the progress in rare single-gene disorders occurring in less than 0.1% of thepopulation, all of which exhibit mendelian patterns of inheritance. The technique to map the chromosomal locationresponsible for single-gene disorders, as indicated previously, is genetic linkage analysis of families of two to threegenerations. Using 300 to 800 DNA markers, one determines whether one or more markers segregates with affectedmembers more often than by chance. Genetic linkage analysis has the resolution for single-gene disorders in whichthe phenotype is dominated by a single gene. In single-gene disorders, the gene is both necessary and sufficient toinduce this disorder, as proven repeatedly by expressing human genes as transgene genes. This is well illustrated by

examples of expressing human genes responsible for familial hypertrophic cardiomyopathy in the mouse512,513 orrabbit514 and inducing the phenotype observed in humans with the disease. In contrast, none of the genespredisposing to a polygenic disorder are necessary or sufficient to induce the disease. Common polygenic disorders




such as CAD are due to common genes, and the predisposition is the result of multiple genes, each contributing as

little as 1% to 10% of the phenotype.479 The technique to map genes predisposing to such common disorders suchas CAD is referred to as a Genome-Wide Association Study (GWAS). In contrast to the hundreds of markers requiredfor genetic linkage analysis, this method requires hundreds of thousands of DNA markers spanning the genome togenotype thousands of unrelated cases and controls. Because of the high incidence of false-positive associations, it isessential that those markers showing an association be replicated in an independent population of the same ethnicity

and of similar or greater sample size.515,516

Background to Genome-Wide Association StudiesMost genes exist in many forms, referred to as alleles. All of us have two alleles for each gene, one from each parent.If they are identical, then one is homozygous; if different, heterozygous. These alleles usually differ by only a singlenucleotide. Collectively, these nucleotides responsible for the polymorphism of these alleles are referred to as SNPs.

The nucleotide sequence of DNA is 99.5% identical in all human genomes.517 The difference is due to 3 million SNPs(0.1% of the genome) and the remainder (0.4%) due to structural variation including long DNA repeats that vary in

length and are referred to as copy number variants (CNVs).518 It is currently believed that in human variation,regardless of whether it is in attributes such as height, hair color, or susceptibility to disease, 80% is due to SNPs,

with the remainder due to CNVs.519 The Human Genome Project520 and, more recently, the International HapMapProject521 has annotated more than 3 million SNPs. From these SNPs, the first microarray was formed, initially with500,000 SNPs and currently with 1 million SNPs and 1 million probes for CNV.522 These SNPs or CNV probes wereselected as markers to span the genome to perform GWAS. Each individual's DNA sample is genotyped with thearray. The fundamental basis for the GWAS is simple and well founded. Any SNP that associates with the casessignificantly more frequently than with the controls is said to be in or near a risk variant for the disease, and SNPsoccurring more frequently in controls are in or near a protective variant. There has been great concern regardingwhat P value is required to establish a significant association. This is because, using 1 million SNPs, one will havemany false positives simply because of the large number of SNPs. If one accepts the conventional P value of 0.05,one would expect 50,000 false positives on the basis of chance alone. Therefore, it has become conventional to

perform a Bonferroni correction, namely, dividing 0.05 into 1 million, which is 5 x 10–8. Based on this correction, it isnow accepted that a SNP, to show significant association in the initial population (referred to as the discovery

population), must have a P value of 5 x 10–8. Thus SNPs that exhibit a difference between cases and controls thathave a P value 5 x 10–8 are said to show a genome-wide significant association.523 The second component to case-control association studies involves genotyping the SNPs that show a genome-wide association in an independent

population of cases and controls of similar ethnicity and with similar sample size.524,525 If the association is notconfirmed, then it is considered a false positive. In the second population, the association must undergo a Bonferronicorrection, namely, 0.05 is divided by the number of SNPs genotyped. Obviously, if there is a single SNP, there is noneed for correction.

The Discovery of 9p21: The First Common Risk Variant for Coronary ArteryDiseaseIn 2007, the results of the first GWAS for CAD was the discovery of the 9p21 locus, the first common risk variant for

CAD.510 Since then, more than 400 loci have been mapped to be associated with various traits and diseases.526

Multiple loci have been identified to be associated with many of the common chronic diseases, including more than 40

loci for diabetes.500 The 9p21 locus is by far the most robust and validated of the identified loci for CAD. The 9p21locus illustrates both the limitations as well as the future hope for these genetic predisposing variants. The 9p21 locus

has been confirmed in populations throughout the world involving more than 100,000 individuals.479,527-530 Theoriginal association was with CAD in which the cases were confirmed by coronary angiography and the controls wereelderly asymptomatic individuals. This was confirmed simultaneously by the deCode group, in whom the phenotype of

the cases was MI rather than documented CAD.525 These two studies were followed by multiple further studies inwhich the phenotype was either CAD, confirmed by coronary angiography, or a coronary event, primarily MI. All ofthese studies were in white patients of European ancestry. Subsequently, studies showed that 9p21 is also a risk

factor for Chinese,531 Korean,532 and East Asian individuals.533 Currently the only populations exempt from the riskof 9p21 are Africans and African Americans.533 The 9p21 locus is an extremely common variant that occurs in 75% ofthe population, with 50% being heterozygous and 25% homozygous. Individuals homozygous for 9p21 haveapproximately a 50% increased risk for CAD, and those heterozygous have approximately a 25% increased risk. It isparticularly noteworthy, as shown originally and confirmed now in multiple studies involving more than 100,000individuals, that the risk exerted by 9p21 is independent of all known risk factors, including blood pressure,

cholesterol, diet, diabetes, and obesity.480 This emphasizes another reason to pursue genetic risk variants, namely,novel mechanisms as yet unknown that contribute to the etiology of CAD. These mechanisms provide targets,previously unknown, for development of new therapies to prevent and treat CAD. The 9p21 locus reflects thecomplexity of determining biologic function of variants contributing to polygenic disorders. Preliminary studies




indicate that the risk region for 9p21 consists of a DNA region of 58,000 base pairs, with no annotated protein-coding

genes.534 This region overlaps with a large noncoding RNA (ANRIL), whose function is also unknown. Genomicanalysis indicates that the 9p21 region did not evolve until the evolution of the chimpanzee. The ANRIL noncodingtranscript has no homology in the opossum or mouse. Functional analysis suggests that there is an enhancer

sequence that appears to normally suppress the expression of downstream genes that induce cellular proliferation.534

The enhancer associated with risk appears to be associated with increased expression of genes that induceproliferation, which may be the mechanism whereby 9p21 mediates its risk. It is of some significance that the 9p21

risk variant is also associated with increased risk of aortic aneurysm,535 intracranial aneurysm, and stroke.536 Takinginto consideration all of these findings, a reasonable hypothesis would be that 9p21 risk variant is associated with adefect in the vessel wall rather than associated with increased plaque rupture or other associated sequelae, such asMI. This is also in keeping with results of several recent genome-wide association studies showing that 9p21

associates more significantly with CAD and atheroma than with MI. Recent findings537 confirm that 9p21 is associatedwith increased severity of coronary atherosclerosis, with homozygous individuals more likely to have triple-vesseldisease and worse Duke and Gensini scores.

Other Genetic Risk Variants Associated with Coronary Artery DiseaseResults of several GWAS for CAD and related traits have been published and are summarized in Table 82–15. Twelveloci have been mapped and replicated to be associated with the phenotype of CAD, as documented by coronary

angiography, or a proven clinical event, primarily MI.526 All of the loci shown in Table 82–15 have been replicated inindependent populations of European ancestry. The frequency of these risks variants in the population and the degreeto which they increase risk for CAD are minimal to moderate. They all fall into the category of common variants,namely, occurring in at least 5% of the population. In fact, all 12 occur in greater than 10% of the population, withan average frequency, except for 9p21, of approximately 20%. The relative increased risk of each variant asexpected is very small, with relative odds ratios varying from 1.2 to 1.3. A surprising finding of the 12 loci is theobservation that only 4 of the 12 act through known mechanisms. These four act primarily through increased oraltered lipids. A second surprise of the DNA loci mapped to be associated with CAD is that most are in nonprotein-coding regions. Several are in either promoter regions or RNA noncoding regions. The frequency of common riskvariants so far discovered occurring in at least 10% of the population is not accidental. This may reflect a sample sizelimitation of the current GWAS. Genetic risk variants occurring in 5% to 10% of the population with increased riskratios of 1.1 require much larger sample sizes than previously anticipated. To overcome this limitation in sample size,a new consortium, referred to as Coronary Artery Disease Genome-wide Replication and Meta-analysis(CARDIoGRAM), comprises 13 GWAS pursuing genes for CAD. CARDIoGRAM includes almost all of the GWAS in theworld. The discovery population of more than 80,000 cases and controls has already been recruited and phenotyped,

with a replication population of similar sample size.538 This sample size, the largest in the world, should be sufficientto detect all common risk variants of CAD occurring with a frequency 5% with increased risk as low as 10%. Despitethe rapid accumulation of genetic variants, the 12 loci for CAD account for less than 5% of the genetic predisposition.It is expected that the remainder are in common variants with frequencies between 5% and 10% with minimal effect,and others are in rare variants that occur by definition in less than 5% of the population. Variants with a frequency of

less than 5% are not expected to be detected by GWAS.523 These rare variants have to be identified through directDNA sequencing, which is now possible with rapid high-throughput parallel sequencing.523

Table 82–15. Summary of 12 CAD Susceptibility Loci Identified to Date through GWAS

Lead SNP Locus Risk Allele Freq Genes in or Near Locus OR (95% CI) P value

rs4977574 9p21 0.56 CDKN2A-CDKN28 1.29 (1.25-1.34) 2.7 x 10-44

rs646776 1p13 0.81 CELSR2-PSRC1-S0RT1 1.19 (1.13-1.26) 7.9 x 10-12

rs17465637 1q41 0.72 MIA 3 1.14 (1.10-1.19) 1.4 x 10-9

rs1746048 10q11 0.84 CXCL12 1.17(1.11-1.24) 7.4 x 10-9

rs9982601 21q22 0.13 SLC5A3-MRPS6- KCNE2 1.2 (1.14-1.27) 6.4 x 10-11

rs12526453 6p24 0.65 PHACTR1 1.12 (1.08-1.17) 1.3 x 10-9

rs6725887 2q33 0.14 WDR12 1.17 (1.11-1.23) 1.3 x 10-8




Genetic Risk Variants: Their Function and Role in Clinical ApplicationThe genetic risk variants are expected to have clinical application in genetic screening to stratify individuals with highand low risk. This could be performed using the SNPs as markers to detect risk loci already replicated. However,there has not yet been a proper assessment of their clinical utility. Until such an assessment is performed, clinicalapplication must be regarded as investigational. The most exciting use of these genetic risk loci is likely to be astargets to develop new therapies for improved prevention and treatment. For these variants to serve as targets fornew therapies, it will be necessary to know the precise causative sequence in the variant, as well as its biologicfunction. It is of note that currently, although all of these loci have been mapped, the precise causative sequenceremains unknown. Assuming that most of these variants are due to an SNP, it would thus be one among thousands.Nevertheless, although this is a current limitation, there is considerable hope for the future. Of the 12 loci identified,only 4 appear to act through known mechanisms for atherosclerosis or CAD. Thus the implication for the remainingloci is that new mechanisms not yet discovered are actively involved in the underlying process of atheroma and itssequelae. Identifying these new mechanisms and the molecular pathways through which their risk is manifested willundoubtedly lead to new therapies both for prevention and treatment of this disease.

rs1122608 19p13 0.75 LDLR 1.15 (1.1-1.2) 1.9 x 10-9

rs11206510 1p32 0.81 PCSK9 1.15 (1.1-1.21) 9.6 x 10-9

rs9818870 3q22 0.154 MRAS 1.15 (1.1-1-1.19) 7.44 x 10-13

CTTG hap 6q26-q27 0.156 SLC22A3-LPAL2-LPA 1.2 (1.3-1.27) 1.19 x 10-9

CCTC hap 6q26-q27 0.018 SLC22A3-LPAL2-LPA 1.82 (1.57-2.11) 4.20 x 10-15

rs3184504 12q24 0.42 SH2B3 1.13 (1.08-1.18) 8.6 x 10-8

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