cancer: a disease example that generates questions to ......cancer • overall, twin of a person...
TRANSCRIPT
The Environment’s Influence on Colon Cancer: A Disease Example that
Generates Questions to Answer for the Near Future
October 31, 2019
John M. Carethers, M.D., M.A.C.PC. Richard Boland Distinguished University Professor
John G. Searle Professor of Internal MedicineUniversity of Michigan
Disclosures• NONE (except NIH and University of Michigan research
funding)
Adenoma (polyp)-to-Carcinoma (cancer)
Adenoma Adenoma Adenoma
Adenoma Cancer Cancer
Familial Colorectal Cancer Risk
Sporadic (~65%) Familial (~35%)
Lynch (3-5%)
FAP (<1%)
Rare syndromes (~4%)
Colorectal Cancer145,600 cases annually in the US
Genetics of Adenomatous Polyposis Syndromes(dysplastic tissue)
AdenomatousSyndrome
Chromosomal Location(s)
Mutated Gene(s) Inheritance Pattern
Familial Adenomatous PolyposisGardner’s variantTurcot’s variant
5q21 APC Autosomal Dominant
MYH-Associated Polyposis 1p32-34 MYH (bi-allelic) Autosomal Recessive
POLE/POLD1 Polyposis 12q24.319q13.3
POLE (L424V)POLD1 (S478N)
AutosomalDominant
Lynch SyndromeMuir-Torre variantTurcot’s variant
2p16, 3p21, 2q32, 7p22, 2p16, 3p22
MSH2, MLH1, PMS2, MSH6, EPCAM (TACSTD1)
Autosomal Dominant
MSH3-Associated Polyposis(CMMRD)
5q14.1 MSH3 (bi-allelic) Autosomal Recessive
Familial CRC Type X 8q12 + 1q22 RPS20 + SEMA4A + HNRNPA0 + WIF1 + others
Autosomal Dominant
1991
2003
1993-1997
2005, 2014-
2013
2016
Genetics of Hamartomatous Polyposis Syndromes(disorganized but mature tissue or cells indigenous to the site of origin)
Hamartomatous Syndrome Chromosomal Location
Mutated Gene Frequency in Germline
PTEN Hamartoma SyndromeCowden’s Disease
Lhermitte-Duclos variantBannayan-Riley-Ruvalcaba Syndrome
10q22-23
10q22-23
PTEN/MMAC1/TEP1
PTEN/MMAC1/TEP1
>80%
~60%
Juvenile Polyposis Syndrome(with HHT overlap)
18q21.110q22-239q34
SMAD4BMPR1A/ALK3ENG
~20%~25%?
Peutz-Jeghers Syndrome 19p13.3 STK11/LKB1 70-90%
Hereditary Mixed Polyposis Syndrome
15q13-q14 duplication
GREM1 overexpression ?
Hyperplastic (Serrated) Polyposis Syndrome
17q22?1p
RNF43 (extremely rare)May not be familial
?
1997
1998, 2001, 2005
1997
2003/*2012
1996-*Nature Genetics 2012;44:699-703
Environmental and Heritable Factors for Cancer
Lichtenstein P et al. N Eng J Med 2000;343:78-85
• 44,788 pairs of twins• 10,803 persons among
9512 pairs of twins with cancer
• Overall, twin of a person with cancer had increased risk for that same cancer
• Only 3 tumors showed statistical heritablecomponent (prostate, colorectum, breast) among 28 sites
• Environment has overall principal role in causing sporadic cancer
Examples of Gene – Environmental Interactions for Cancer Development
VirusesHPVHBV
UV Exposure
Tobacco
GeneticallySusceptibleIndividual
Exposure Types of Cancer
High Fat Diet
Inflammation
Cervical, Anal, Head & NeckLiver
Skin, Melanoma
Lung, others
Colon, Breast
Colon, Pancreas, Stomach
DNA Damage Occurs Constantly• Block DNA
replication, transcription
• Genomic instability• Impair gene
expression• Mutagenic: human
disease• Stem cell
cytosol
nucleus
environment
UVOxidativeDamage
Carcinogens Cytosine DeaminationTo Uracil
DNAReplication
Errors
MUTATIONor
CELL DEMISE(without any repair)
Patient Risk Factors for CRC Development
Carethers JM. Dig Dis Sci 2015;60:711-721.
Comprehensive Mutation
Frequencies in 224 Primary
CRCs
Cancer Genome Atlas NetworkNature 2012;487:330-337
Hypermutated: MLH1 and POLε deficiencyNon-hypermutated: APC, TP53, KRAS, TTN, PIK3CA
~15% ~85%
Diet and CRC Risk in African Americans
O’Keefe SJD at al. Nat Comm 2015;6:6342
• 2 week food exchanges; mucosal biopsy• AA: fed high fiber, low-fat African-style diet• Rural Africans: fed high fat, low-fiber Western-style diet• Observed reciprocal changes in mucosal biomarkers
Butyrate(short chain fatty acid)
Deoxycholic acid (DCA)(secondary bile acid)
• During Dietary Intervention (DI)• AA: increased butyrate and decreased DCA• Rural Africans: decreased butyrate and increased DCA
ED: endoscopyHE: home environmentDI: dietary intervention
Aspirin Use Reduces CRC Death with Mutant PIK3CA
NEJM 2012;367:1596-1606
• Aspirin inhibits COX-2, down-regulates WT PI3K
• Mutant PI3K is predictor for aspirin effectiveness
Lynch Normal Accelerated (1-3 years)
80% Risk
FAP Normal 100% Risk
Normal Colonocyte
Tumor Initiation
Tumor Progression Carcinoma
Sporadic Normal(10-20 years)
5% Risk
Mean Age ~ 40 yrs
Mean Age ~ 40 yrs
Mean Age ~ 68 yrsNormal
(30-50 years)
Accelerated(5-20 years)
Normal colon Tubular adenoma High grade dysplasia CancerGrady W & Carethers JM. Gastroenterol 2008;135:1079-1099.
• Microbiome?• Inflammation Inflammation?• Microbiome?
• InflammationMetastasis
Type, Density and Location of Immune Cells Better Predictor Than Stage for Colorectal Cancer
• Immune cells– Can release mediators with pro-
angiogenic and pro-metastatic effects
• Tumor-infiltrating lymphocytes (CD3+)– Higher densities in center (CT)
and at invasive margin (IM) of tumor associated with improved patient survival
• CD8+ and granzyme B+ cytotoxic T lymphocytes (TH1 via IFNγ)
• CD45RO memory T cells
Galon et. al. Science 2006;313:1960-1964
Genetics Changing the Environment
Some Common Features of Cancer with Defective DNA MMR
• 15% of all sporadic CRCs• Tissues display Microsatellite Instability (frameshifts)
• Lost of DNA MMR protein expression
• CRCs more common in proximal (Right) colon• CRCs often show poor differentiation and mucin• Tumors are hypermutated, containing 102-103 somatic mutations• Tumors are mostly diploid; lack p53 mutations• CRCs attract T cells and form sub-epithelial lymphoid nodules
(neo-antigens from novel frameshifted peptides)• More susceptible to PD-1 immune checkpoint blockade
• CRCs are resistant to 5-fluorouracil treatment• Patients with MSI CRCs show longer survival, earlier stage
• Caveat: EMASTCarethers and Jung. Gastroenterology 2015;149:1177-1190.
Normal AdenomaHypermutable
PhenotypeCancer
Wnt:APC/β-catenin
Detection of Microsatellite
instability-High
Immunogenic neo-peptides fromACVR2, TGFBR2,BAX, MMR genes,
IGF2R, E2F4, MBD4, etc.frameshifts
Genomic/Epigenomicinstability
-Lynch (germline MMR mutation)-Lynch-like (bi-allelic somatic mutation)-Sporadic (hMLH1 hypermethylation)
B-RAFMutation
(non-Lynch)
Loss of DNA Mismatch Repair
somatic frameshift mutations
MSI Pathway and CRC Progression
Carethers JM. Curr Colorectal Cancer Rep 2017;13:73-80
PD-1
Induction of “protective” T cells
Schema for MSI Neo-peptide Inflammation
Induction of T Cell “Protective” Inflammation
Alteration of DNA MMR(somatic/germline MLH1, MSH2, MSH6, PMS2, EPCAM)
Target Gene Mutations (mononucleotide; flanking sequences)TGFBR2, ACVR2, etc.
Biological consequences(reduced metastasis)
Estimate of patients anti-PD-1 benefits
Le DT et al. Science 2017;357:409-413
Environment Changing the Genetics
Some Common Features of EMAST CRCs
• Observed in 50% of all sporadic CRCs (plus other cancers)• Tissues display EMAST (elevated microsatellite alterations at
selected tetranucleotide repeats)• Heterogeneous (reduced) MSH3 nuclear expression
• CRCs appear distributed throughout the colon• Tumors are non-hypermutated (excluding complete MMR defects)• CRCs associated intimately with inflammatory cells
• Could be a driver for EMAST
• CRCs are sensitive to 5-fluorouracil treatment• Patients with EMAST CRCs show shorter survival, more
metastases, poor prognosis, advanced staged disease• Higher frequency among African American cancers
Carethers and Jung. Gastroenterology 2015;149:1177-1190.
IL-6 Changes MSH3 Subcellular Location and Induces EMAST
Tseng-Rogenski SS et al. Gastroenterology 148:579-589, 2015.
MYCL1
Schema for Inflammatory-Associated Microsatellite Alterations AKA EMAST
Inflammation (IL-6 driven)
Initiation of Neoplasia and tumor advancement
Alteration of DNA MMR(MSH3 “loss of function”)
Potential Target GeneMutations (di, tri, tetranucleotide),
Double strand breaks
Biological consequences(increased metastasis)
Carethers JM et al. Genes 6:185-205, 2015
Microbiome?
50% of CRCs
Microbes and CRC Progression
Brennan CA, Garrett WS. Annu Rev Microbiol 2016;70:395-411.
CRC-Associated Microbes
Brennan CA, Garrett WS. Annu Rev Microbiol 2016;70:395-411.
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Earlyneoplasia
Lateneoplasia
pks+E. Coli and ETBF in FAP Patients and Mutant APC mice
Dejea CM et al. Science 2018;359:592-597.
FAP Patients
Mutant APC mice
Combination of pks+E. coli and ETBF• increased inflammation• faster tumor onset • poor survival• IL17-dependent
Colonized with both pks+E. coli and ETBF and assessed at 15 weeks
Colon inflammation score
Biofilm • Found in majority of Rt. Side
CRCs and associated normal mucosa
• Biofilm-negative CRCs are associated with normal mucosa that does NOT possess a biofilm
• Risk of proximal CRC development is 5-fold higher with biofilm
• Rt. Side: MSI, hypermutated, hypermethylation of MLH1, BRAFV600E
Right-sided Location of Biofilm in Sporadic CRCs
**
Dejea CM et al. PNAS 2014;111:18321-18326.
Biofilms Trigger Immune Response and Proliferation
Dejea CM et al. PNAS 2014;111:18321-18326.
Fusobacterium Persistence in Primary CRC and Liver Metastasis
Bullman S et al. Science 2017;258:1443-1448.
Primary CRC
Paired Liver Mets
Primary CRC
Paired Liver Mets
RNA ISH
Serial PDX Passage
Antibiotics Affect Fusobacterium Infected Tumor Growth
Bullman S et al. Science 2017;258:1443-1448.
Fusobacterium ROS
8-oxoGCpG islands
Translocation of MSH3 from nucleus to
cytoplasm
Recruitment of MSH2/MSH6
DNMTsPRC4
Replication errors at microsatellite
with di- or tetranucleotide
repeats
Hypermethylation of promoter CpG
islands
Hypomethylation of CpG at non-
promoter region
CIMP
Hypermethylation of MLH1 MSI-H
MSI-LEMAST
IL6PGE2
DNA damage
Proposed Pathways of Genetic/Epigenetic Alterationsin CRC Induced by F. nucleatum Infection
Koi et al. JARC 2018;2:37-46
Diet and Fusobacterium nucleatum status in CRC
Liu L et al. Clin Gastroenterol and Hepatol 2018;16:1622-1631.
EDIP: empiric dietary inflammatory pattern score (weighted intake of 18 foods constructed to predict plasma levels of IL6, CRP, TNFRSF1B, or TNF alpha-receptor 2)
CRC Trends by Birth Year or Year Diagnosis
Wolf AMD et al. CA Cancer J Clin 2018;68:250-281 and Murphy CC et al. Gastroenterol 2018;155:1716-1719
45-49 yrs
40-44 yrs
35-39 yrs30-34 yrs
Triclosan Increases Tumorigenesis (AOM/DSS)
Yang H et al. Science Translational Medicine 2018;10:1-10
Summary• The environment plays a major role in cancer
formation, progression, and metastasis– Colorectal cancer is a great example
• Understanding role and pathways involved affords intervention strategies– Diet intervention– Aspirin optimal usage– Microbiome intervention– Inflammation prevention or modification
Future Addressable Questions• How does the environment influence different
genetic susceptibilities?– e.g. aspirin and PIK3CA mutations in colorectal
cancer• How does the environment contribute to disease
formation?– e.g. microbiome and adenomas in colorectal cancer
• How does the environment generate changes in cellular pathways (that could be targeted)?– e.g. IL-6 inflammation and loss of DNA repair in CRC
Future Addressable Questions• How do we identify the genetically susceptible?
– Prior to disease formation (e.g. germline)– With disease (e.g. somatic mutation analysis)
• What in the environment leads to changes in the genome for disease formation?– e.g. what is increasing young-onset CRC
Thank you!