Chapter 2

REVIEW OF LITERATURE

Cancer

Cancer is a genetic disease, arising from an accumulation of mutations that promote

clonal selection of cells with increasingly aggressive behavior. The vast majorities of mutations

in cancer are somatic and are found only in an individual’s cancer cells. However, about 1% of

all cancers arise in individuals with an unmistakable hereditary cancer syndrome (Fearon, 1997).

Cancer is a progressive disease, occurring in a series of well-defined steps, typically arising as a

consequence of activating mutations (oncogenes) or deactivating mutations (tumor suppressor

genes) in proliferating cells (Lisa et al., 2001). Cancer is an English term, dissimilated from

Greek word 'Karakinos' (Sanskrit Karkita) for crab, the symbol for fourth zodiacal constellation

(the CANCER) or Cancer is a generic term that refers to a group of chronic diseases

characterized by the uncontrolled growth of abnormal cells within the body.. The word was

believed to be first used by Hippocrates, who attributed this disease to an excess of black bile.

Cancer was known in old ages, being described in early writings of Greeks and Romans.

Pathological evidences support the bone tumors in dinosaurs and other prehistoric animals but

the tumors in Egyptian mummies dating back to 5000 years represent the first known human

malignancies. Later, the Roman physician Celsus (28-50 BC) used the Latin term for crab,

cancer. Another Roman physician, Galen, (130-200 AD) used the Greek word for swelling,

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oncos, to describe tumours -the root of the modern English word, oncology. Cancer has been

defined in terms of an autonomous growth that is unresponsive to normal growth factors and

antigrowth signals; in provisions of irreversibility with which the cancer cells progressively lose

the differentiated characteristics and functions of normal tissue of origin; on the basis of

morphologic and cytogenetic features; and on the basis of reversion to growth and antigenic

properties characteristics of fetal cells. Normally, cells divide and replicate to replace worn-out

cells or to repair some form of injury to tissues of the body. After a predictable period, normal

cells wear out and die. Cancer cells do not grow, divide and die in the same predictable fashion

as normal cells. Rather, they grow, divide and create more abnormal cells, which outlive normal

cells. The abnormal cells often spread to other body parts, invading other organs or systems.

Gastric cancer

Gastric cancer refers to cancer arising from any part of the stomach. Gastric cancer is

often either asymptomatic (producing no noticeable symptoms) or it may cause only nonspecific

symptoms (symptoms which are not specific to just gastric cancer, but also to other related or

unrelated disorders) in its early stages. By the time symptoms occur, the cancer has often reached

an advanced stage and may have also metastasized (spread to other, perhaps distant, parts of the

body), which is one of the main reasons for its relatively poor prognosis.

Gastric cancer can cause the following signs and symptoms:

Stage 1 (Early)

Indigestion or a burning sensation (heartburn)

Loss of appetite, especially for meat

Abdominal discomfort or irritation

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Stage 2 (Middle)

Weakness and fatigue

Bloating of the stomach, usually after meals

Stage 3 (Late)

Abdominal pain in the upper abdomen

Nausea and occasional vomiting

Diarrhea or constipation

Weight loss

Bleeding (vomiting blood or having blood in the stool)

Dysphagia; this feature suggests a tumor in the cardia.

Pathology

The primary epithelial tumour of the stomach is the adenocarcinoma, and develops from

the stomach mucosa, usually maintaining glandular differentiation. Other less common tumours

of the stomach are the squamous cell carcinomas, and the adenosquamous carcinomas,

combining characteristics of both the adenocarcinoma and the squamous cell carcinoma to

approximately equal extent. Undifferentiated carcinoma lacks any differentiated features and

does not fit into any of the above categories. Gastric carcinomas can be classified according to

their localization in the stomach. The antral-pyloric region of the stomach is the most common

site of stomach cancer, and carcinomas of the body or corpus are located along the greater or

lesser curvature. Cancers of the cardia are often unable to be distinguished from cancers of the

gastroesophageal junction, and are believed to be a separate entity, probably originating from the

distal oesophagus. Early gastric cancers may feature protruded (Type I), elevated (Type IIa), flat

10

(Type IIb), depressed (Type IIc) or excavated (Type III) growth (Hamilton and Aaltonen,

2000), whereas advanced gastric carcinomas are classified into polypoid (Type I), fungating

(Type II), ulcerated (Type III) or infiltrative (Type IV) growth patterns (Borrmann, 1926;

Hamilton and Aalton, 2000). Type II or III advanced gastric cancers are commonly ulcerating,

and the risk of penetration of the submucosa is highest in early gastric cancers with a depressed

growth pattern (Type IIc), and in infiltrative advanced gastric carcinomas (Type IV). The

superficial spread of Type IV infiltrative (diffuse) tumours through the mucosa and submucosa

result in flat, plaque-like lesions, which may exhibit shallow ulcerations. Serosal, lymphatic, and

vascular invasion and lymph node metastases are most frequent in the diffusely growing tumours

(Mori et al., 1995; Carneiro, 1996).

Histological classification

Various systems have been applied to the classification of gastric carcinomas, including

the WHO (Laurén, 1965; Ming, 1977; Goseki and Koike, 1992; Hamilton and Aalton, 2000)

classifications. The clinical significance of these classifications is limited, with only the Lauren

and perhaps the Goseki classifications providing prognostic assessments (Alekseenko et al.,

2004). The TNM staging of the gastric carcinoma, according to the guidelines set out by the

International Union Against Cancer (UICC) (Wittekind and Sobin, 2002), is the most

important prognostic factor in clinical practice (Alekseenko et al., 2004). However, the Lauren

classification has been the most successful system, as it defines two distinct histological entities,

which clearly exhibit different clinical and epidemiological characteristics, even in advanced

gastric cancers (Satoh et al., 2007). In the Laurén classification(Laurèn, 1965), intestinal-type

carcinomas maintain the glandular phenotype, with well- to moderately-differentiated tumours

forming identifiable glands, often with poorly differentiated tumour cells at the invasive front.

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Typically arising on a background of intestinal metaplasia, these tumours exhibit an intestinal,

gastric and gastrointestinal mucinous phenotype. Diffuse-type carcinomas form no or very few

glandular structures, instead usually infiltrating the gastric wall, appearing diffusely distributed

as small, round single cells or poorly cohesive cell clusters. They may resemble signet-ring cells,

and may contain small amounts of intestinal mucin. Additionally, mixed tumours exhibit both

intestinal and diffuse characteristics, and undifferentiated tumors are classified as indeterminate.

The natural history of gastric carcinoma, in particular the association with environmental factors,

incidence trends, and precursor lesions, is often evaluated with respect to the Laurén

classification.

Descriptive epidemiology

One of the notable features of the descriptive epidemiology concerning gastric cancer is

that it establishes some clear distinction between cancer localized to the gastric cardia and cancer

of the rest of the stomach, as discussed below.

International variations

Despite a major decline in the incidence and mortality over several decades, gastric

cancer is still the fourth most common cancer and the second to third most frequent cause of

cancer death in the world (Brenner et al., 2009; Herszenyi and Tulassay, 2010). There is

marked geographic variation in the incidence of gastric cancer. International Agency for

Research on Cancer data for 1996, demonstrate age-standardized incidence rates in males

ranging from 95.5/10.5 in Yamagata, Japan,to 7.5/10.5 in Whites in the United States. High-risk

areas include China and large parts of central and South America (Parkin and Ferlay, 1997).

Most of the geographic variation is accounted for by differences in the incidence of non cardia

cancer. Cancer localized to the cardia has a more uniform distribution. Gastric cardia cancer

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accounts for only 4% of total gastric cancer cases in males in Osaka, Japan, compared to 39% in

white males in the United States (Parkin and Ferlay, 1997). On a histologic level, the incidence

of diffuse adenocarcinomas is reported to be similar in most populations, while the intestinal

type predominates in the high-risk geographic regions and is the type that has declined

significantly in incidence in many countries (Munoz and Cuello, 1968; Sipponen and Kekki,

1987). Ethnic groups who have migrated from high- to low-incidence countries have an overall

risk intermediate between that of their homeland and that of their new country. First generation

migrants tend to maintain their high-risk while subsequent generations have risk levels

approximating that of the host country (Haenszel, 1968). The prognosis is rather poor, with a

five-year survival below 30%. More than 90% of gastric cancers are adenocarcinomas, which are

malignant epithelial tumors, originating from glandular epithelium of the gastric mucosa. In the

Lauren classification, two major histological types of gastric adenocarcinoma can be

distinguished histopathologically: the diffuse and the intestinal type (Vauhkonen et al., 2006).

Intestinal metaplasia with goblet cells is considered to be a precursor lesion of the intestinal type

of gastric adenocarcinoma, which shows tubular differentiation (Vauhkonen et al., 2006). The

diffuse type gastric adenocarcinoma is characterized by non-cohesive single mucocellular cancer

cells (signet-ring cells) diffusely infiltrating the stroma (Vauhkonen et al., 2006). The diffuse

type gastric adenocarcinoma shows some predominance in the fundus and corpus of the stomach,

whereas the intestinal type gastric adenocarcinoma prevails in the antrum (Vauhkonen et al.,

2006). Furthermore, the remaining 10% of gastric malignancies are lymphomas or originate from

gastrointestinal stromal tissue (soft tissue tumors) (Verbeke et al., 2012).

Indian variations

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Cancer is one of the leading causes of adult deaths worldwide. In India, the International

Agency for Research on Cancer estimated indirectly that about 635 000 people died from cancer

in 2008, representing about 8% of all estimated global cancer deaths and about 6% of all deaths

in India (Ferlay et al., 2010). 7137 of 122 429 study deaths were due to cancer, corresponding to

556 400 national cancer deaths in India in 2010. 395 400 (71%) cancer deaths occurred in people

aged 30–69 years (200 100 men and 195 300 women). At 30–69 years, the three most common

fatal cancers were oral (including lip and pharynx, 45 800 [22·9%]), stomach (25 200 [12·6%]),

and lung (including trachea and larynx, 22 900 [11·4%]) in men, and cervical (33 400 [17·1%]),

stomach (27 500 [14·1%]), and breast (19 900 [10·2%]) in women. Tobacco-related cancers

represented 42·0% (84 000) of male and 18·3% (35 700) of female cancer deaths and there were

twice as many deaths from oral cancers as lung cancers. Age-standardised cancer mortality rates

per 100 000 were similar in rural (men 95·6 [99% CI 89·6–101·7] and women 96·6 [90·7–

102·6]) and urban areas (men 102·4 [92·7–112·1] and women 91·2 [81·9–100·5]), but varied

greatly between the states, and were two times higher in the least educated than in the most

educated adults (men, illiterate 106·6 [97·4–115·7] vs most educated 45·7 [37·8–53·6]; women,

illiterate 106·7 [99·9–113·6] vs most educated 43·4 [30·7–56·1]). Cervical cancer was far less

common in Muslim than in Hindu women (study deaths 24, age-standardised mortality ratio 0·68

[0·64–0·71] vs 340, 1·06 [1·05–1·08] (Rajesh et al., 2012).

Kashmir variations

The valley of Kashmir is one of the divisions of Jammu and Kashmir State, situated in

the Himalayas. In Kashmir valley where incidences of almost all types of organ cancers have

shown a drastic increase in last couple of decades particularly gastric cancer. Gastric cancer is

the leading one with an average frequency of 19.2% followed by esophagus and lung as 16.5%

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and 14.6%, respectively. In cancer types common to both sexes, the proportion in the men

exceeds that in the women particularly in lung and bladder cancer where around 80% were

males. Stomach (23%) and lung (21%) are the leading cancers in men while as esophageal

cancer tops (18.3%) in women followed by breast cancer (16.6%). Those tumors affecting the

stomach and extending to the esophageal lumen were considered gastric cancers. An interesting

finding was the presence of around 30% adenocarcinoma of GE junction among the total

frequency of stomach cancers. Around 21% of GE junction cancers were of sqamous cell

carcinoma of esophageal origin while as around 8% were recorded as adenocarcinoma of

esophageal origin. The age-standardized rates (ASR) of incidence of stomach cancer tops the list

with 10.2 cases/100,000/year followed by esophageal cancer (9.4/100,000) and lung cancer

accounts for third incident cancer (7.8 cases/100,000/year) (Arshad et al., 2012).

Age, sex and Race

The incidence of gastric cancer rises progressively with age, with most patients being

between the ages of 50 and 70 years at presentation. Cases in patients younger than 30 years are

very rare. Noncardia cancer is more common in males than females by a ratio of approximately

2:1. Gastric cardia cancer has a higher male-to-female ratio, of up to nearly 6:1 in U.S. Whites

(Parkin and Ferlay, 1997). There are significant variations in the overall incidence of gastric

cancer between different ethnic groups living in the same region (Parkin and Ferlay, 1997).

The ethnic distribution for cardia cancer is different, with a preponderance in Whites over Blacks

in the United States and non-Maoris over Maoris in New Zealand (Parkin and Ferlay, 1997).

Socioeconomic status

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Low socioeconomic status has been consistently shown to be associated with an

increased risk of gastric cancer overall (Howson and Wynder, 1986). Remarkably, the increase

in incidence of cardia cancer has been predominantly in professional classes (Powell, 1992).

Aetiology and risk factors

Aetiological factors

Migrants from high- to low-incidence countries tend to maintain the high risk of the

population of origin. However, subsequent generations of migrants have risk levels

approximating those in the host country (Stewart BW 2003). It has been suggested that

approximately 66–75% of stomach cancer risk could be reduced with high intake of fruit and

vegetables and low consumption of salted foods (AICR 1997). The World Cancer Research Fund

(WCRF) and Association for International Cancer Research (AICR) panel of experts reached the

following conclusions:

The evidence that diets high in vegetables and fruits, collectively and separately, decrease

the risk of stomach cancer is convincing. There is consistent evidence that raw

vegetables, allium vegetables and citrus fruits have a protective effect. A negative role

has been ascribed to salted nd pickled vegetables.

There is also convincing evidence that the availability of refrigeration—which facilitates

year-round consumption of vegetables and fruits and may also reduce the need for salt as

a preservative—protects against stomach cancer.

Diets high in salted foods and a high intake of salt, added in manufacture, in cooking, or

at the table, probably increase the risk of stomach cancer.

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Diets high in wholegrain cereals and green tea possibly decrease the risk of stomach

cancer.

Although the data regarding wholegrain cereals and stomach cancer are somewhat

limited in number, they are almost entirely consistent.

In terms of the most important vitamins and compound contained in vegetables and

fruits, the WCRF and AICR report considered that Vitamin C probably, and carotenoids possibly

decrease the gastric cancer risk, while it is doubtful that Vitamin E is involved. Alcohol

consumption possibly increases the risk of cancer of the gastric cardia (AICR 1997). Some

evidence has been provided by chemoprevention trials. An intervention study carried out in

volunteers(Correa P 2000) to assess the effect of Vitamin C and beta-carotene supplements

found a protective effect, that is an increase in the rate of regression of confirmed histological

diagnoses of intestinal metaplasia and/or atrophy (premalignant lesions). In another

chemoprevention trial, conducted in male smokers in Finland (Varis and Sipponen, 1998),

supplementation with alpha tocopherol and beta-carotene did not affect the risk of stomach

cancer and prevalence of premalignant lesions. The relationship between smoking and stomach

cancer has recently been recognised. The European Prospective Investigation Into Cancer and

Nutrition (EPIC) project (Gonzalez and Agudo, 2003), found a significant association between

cigarette smoking and gastric cancer risk: the hazard ratio (HR) was 1.45, 1.7 and 1.8 for ever

smokers, current male and current female smokers, respectively, and HR increased with intensity

and duration of cigarette smoking. An international population study performed by the

EUROGAST study group found that countries with a high incidence of stomach cancer have a

high prevalence of Helicobacter pylori infection (Group, 1993). However, the geographical

variation in gastric cancer risk in Europe cannot be explained by differences in the prevalence of

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infection, since only a small fraction of those infected by Helicobacter develop stomach cancer.

In Japan, the country with the highest incidence of stomach cancer in the world, it has been

estimated that of the 60 million people infected by Helicobacter only 0.4% had stomach

cancer(Asaka M 1998). H. pylori was isolated in 1982, and was recognised as a human

carcinogen by IARC in 1994, but the specific mechanisms of action in the complex process of

stomach cancer are not known (Gonzalez, 2002). Moreover, H. pylori infection does not

increase the risk of cancer in the gastric cardia. Gastric ulcers, atrophic gastritis and autoimmune

gastritis with pernicious anaemia (Hsing and McLaughlin, 1993; Stewart, 2003), are

conditions which cause an excessive rate of cell proliferation in the gastric epithelium. Gastritis

is associated with increased production of oxidants and reactive nitrogen intermediates, including

nitritic oxide. Gastritis and atrophy alter gastric acid secretion, elevates gastric pH, changes the

gastric flora and allows anaerobic bacteria to colonize the stomach. The evidence for a role of

ionizing radiation in the aetiology of stomach cancer comes from the study of survivors of the

atomic bombings of Hiroshima and Nagasaki (Thompson, 1994). A linear dose–response effect

was observed, although the excess risk was small and the attributable risk was low (6.5%).

Studies of patients undergoing therapeutic radiation to the stomach for peptic ulcer disease (an

old treatment modality) and for testicular cancer, found a two- to four-fold increased risk in

patients exposed to radiation doses of 15–30 Gy (Griem and Boice, 1994). The risk of stomach

cancer is increased in first-degree relatives of patients with the disease by approximately two-to

three-fold(Lissowska J 1999). From the Scandinavian Twin Study, an increased risk of stomach

cancer in the twin of an affected person was found (Lichtenstein and Verkasald, 2000). Model

fitting to assess the contribution of hereditary and environmental factors found that inherited

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genes contributed 28%, shared environmental factors 10%, and environmental factors 62%

(Vincenzo et al., 2005).

Dietary factors

Distinct variations in the incidence and mortality of gastric cancer, over time, between

and within countries, in differing socioeconomic groups, and in migrants and their offspring

suggest that diet may be etiologically important and its role has been extensively investigated

with inconclusive results.

Fruit and vegetables

The dominant dietary hypothesis is that fresh fruits and vegetables, or contained

micronutrients, are protective against gastric cancer. Numerous studies have shown, almost

uniformly, a protective association with fresh fruits and vegetables, independent of other dietary

factors. The association has been less pronounced in limited cohort studies (Kono, 1996).

Possible protective micronutrients include vitamins C (ascorbate) and E (alpha-tocopherol),

carotenoids (particularly beta carotene), and selenium (Kono, 1996). The evidence is strongest

for vitamin C, with an approximate halving of risk associated with high intake vs. low intake

demonstrated in case– control studies (Neugut et al., 1996). However, a 5-year intervention trial,

involving 30,000 40 to 69 year olds in China, did not show any change in risk of gastric cancer

in subjects receiving supplemental vitamin C (Blot et al., 1993).

Salt

The hypothesis that excess salt intake could be involved in the etiology of stomach cancer

was first presented in 1965. It was postulated that the continuous use of high doses of salt would

result in early atrophic gastritis, thereby increasing the later risk of stomach cancer (Joossens et

al., 1996). Since that time high salt consumption has been reasonably consistently associated

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with an increased risk of gastric cancer in ecologic and analytical studies, although good

quantitative data are lacking (Joossens et al., 1996; Kono, 1996).

Nitrite and Nitrate

Many N -nitroso compounds have been shown to be carcinogenic in animal experiments.

Such compounds may be formed in the human stomach from dietary nitrite or nitrate. Hence, the

hypothesis that a diet high in nitrite or nitrate may predispose to gastric cancer. The major

sources of nitrate and nitrite are vegetables and preserved meats, respectively. Drinking water is

an additional source of nitrate, but usually contains negligible nitrite. In general, daily nitrate

intake is approximately 100 times that of nitrite. Small quantities of pre-formed N -nitroso

compounds may also be contained in some foods including cured meats (Gonzalez and Badosa,

1994; Kono, 1996). Case–control studies examining dietary intake of nitrate and the risk of

gastric cancer have consistently found a negative association. In such studies, vegetable intake

has been consistently related to a decreased risk of gastric cancer. Nitrate intake was probably an

index of vegetable intake and the negative association is not surprising in that context (Kono,

1996). Recent case– control studies have all reported a weak, statistically nonsignificant

increased risk of gastric cancer (relative risks from 1.12 to 1.28) for high vs. low nitrite intake

(Gonzalez and Badosa, 1994). Nitrite intake probably reflects consumption of preserved meats,

typically high-salt foods so isolating an effect of nitrite consumption is difficult. Further

evidence suggests that the interaction of the above dietary components is important. For

example, a diet high in nitrite does not appear to confer an increased risk if that diet is also high

in antioxidants from fruit and vegetables (Buiatti et al., 1990).

Other dietary factors

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The collective literature on diet and gastric cancer provides data for a comprehensive

array of food groups, nutrients, micronutrients, and food-storage methods. Difficulties in

gathering and interpreting this evidence limit the conclusions that can be drawn. The advent of

widely available refrigeration, the consequent availability of fresh food and the decreased

consumption of preserved foods may have contributed to the decline in gastric cancer incidence

in the second half of this century. Dixon has pointed out how virulent strains of H. pylori release

reactive oxygen metabolites that could destroy neighboring glandular tissue leading to gastric

glandular atrophy, hastened by factors such as bile reflux or a high-salt intake but retarded by

antioxidants such as ascorbic acid, alpha tocopherol, beta carotene, and cysteine (Dixon, 1997).

Ionizing radiation

The best evidence concerning the role of ionizing radiation in the etiology of gastric

cancer comes from the study of survivors of the atomic bombings of Hiroshima and Nagasaki. In

a prospective incidence study of this cohort of approximately 80,000, Thompson et al. identified

more than 2,600 cases of gastric cancer (Thompson, 1994). A linear dose–response effect (P

>0.001) was observed between radiation dose and risk of gastric cancer, although the excess risk

was small (0.32 at one Sievert, 95% CI, 0.16 to 0.50) and the attributable risk was low (6.5%), in

the setting of a high background rate of gastric cancer in this Japanese population. Studies of

patients undergoing therapeutic radiation to the region of the stomach for peptic ulcer disease

(Griem and Boice, 1994) (a treatment modality used from the late 1930s to the mid-1960s) and

for testicular cancer (Moller et al., 1993; Van et al., 1993), also provide significant support for

this association. These studies suggest a two- to fourfold increased risk in patients exposed to

radiation doses of 15 to 30 Gy. Studies of occupational radiation exposure in radiographers

(Wang et al., 1990) and radiologists (Matanoski et al., 1975; Smith, 1981) have not

21

demonstrated increased risks, presumably due to the much lower radiation doses involved

compared to the atomic bomb survivors and the therapeutically irradiated. Differentiation of risk

based on the type of gastric cancer is not possible from the available evidence.

Pernicious anemia

An association between pernicious anemia and gastric cancer has long been recognized.

In the most recent and largest study of this topic, Hsing et al. observed a threefold increase in the

risk of gastric cancer in a cohort of 4,517 pernicious anemia patients, followed for up to 20 years

(Hsing and McLaughlin, 1993).

Smoking

The relationship between smoking and gastric cancer has been extensively examined yet

remains unclear; while most studies have reported a weak to moderate association, a few have

found none (Hansson et al., 1994; McLaughlin et al., 1995; Nomura et al., 1995; Ji et al.,

1996). In the positive studies the increased relative risks reported have generally been less than

twofold, and only a few studies have found a dose–response relationship (Hansson et al., 1994;

McLaughlin et al., 1995; Ji et al., 1996). A particular limitation of the available studies has

been a lack of control for confounding, particularly by H. pylori infection, which is positively

correlated with smoking, and by fruit and vegetable intake, which is inversely associated. Most

of the evidence does not allow any differentiation of risk by anatomic subsite or by histologic

type. One case– control study found a stronger association for gastric cardia cancer than for other

gastric cancer across multiple categories of smoking, whereas another did not (Ji et al., 1996).

Alcohol

A 1994 review of the experimental, descriptive, and analytical evidence relating to

alcohol and gastric cancer found little to support an association (Franceschi, 1994). After

22

examining over 50 mostly negative cohort and case–control studies, the authors concluded that

alcohol consumption was unlikely to be materially involved in the etiology of gastric cancer.

Subsequently, studies have not challenged that conclusion (Hansson et al., 1994; Ji et al.,

1996). Five case–control studies showed no association between alcohol consumption and cancer

of the gastric cardia (Unakami et al., 1989; Gray et al., 1992; Palli et al., 1992; Guo et al.,

1994; Ji et al., 1996) and one a doubling of risk in drinkers vs. nondrinkers (Kabat et al., 1993).

Epstein-Barr virus infection

Epstein-Barr virus has been isolated from gastric adenocarcinomas and poorly

differentiated carcinomas with lymphoid infiltrate by a number of investigators. Epstein-Barr

virus infection may contribute to the development of gastric carcinoma, but the data are limited.

Helicobacter pylori

The final sentence of Marshall’s, now classic, letter to the Lancet in June 1983

(Marshall, 1983) suggests that the, then unidentified, curved bacilli found in human gastric

epithelia ‘may have a part to play in poorly understood, gastritis associated diseases (i.e. peptic

ulcer and gastric cancer)‘. Exactly eleven years later, in June 1994, the International Agency for

Research on Cancer convened a Working Group which reviewed over 350 papers on the

relationship between Helicobacter pylori and gastric cancer and came to the conclusion that this

relationship was causal (IARC, 1994). This conclusion was reached after considering a wealth of

evidence concerning the epidemiology of H. pylori, specific studies of the association between

infection and the risk of cancer and mechanistic evidence about the pathogenicity of the

bacterium. It is extremely rare for such a body of evidence to accumulate in a short time period

and permit a definitive judgment about the carcinogenicity of an environmental exposure.

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Marshall’s conjecture in 1983 has proven to be remarkably prophetic, so much so that H. pylori

eradication is now envisaged as a method of cancer prevention.

Asbestos

Several studies of workers with occupational exposure to asbestos have reported limited

evidence of an association (Frumkin, 1988; Andersen et al., 1993; Cocco et al., 1994).

However, methodologic problems cast doubt on the association. A case–control study of heavily

exposed asbestos miners and millers in Western Australia found no association between gastric

cancer mortality and intensity of exposure, duration of employment, or time since employment

began (De et al., 1989).

Other risk factors

The risk of gastric cancer is increased in first-degree relatives of patients with the disease

by approximately two- to threefold (La et al., 1992; Palli et al., 1994; Lissowska et al., 1999).

other suggested risk factors include blood group A and gastric polyps. However, the final word

on heredity in gastric cancer may well be the Scandinavian Twin Study of 44,788 pairs of twins

in the Swedish, Danish, and Finnish twin registries (Lichtenstein and Verkasald, 2000). This

found an increased risk of gastric cancer in the twin of an affected person. Model fitting to assess

the contribution of hereditary and environmental factors found that inherited genes contributed

28% (95% C.I. 0–51%), shared environmental factors 10% (95% CI, 0–34%), and environmental

factors 62% (95% CI, 0–76%). The statistical model used provided a perfect fit (P>1.0).

Chronic inflammation

Chronic inflammation plays a multifaceted role in carcinogenesis. Mounting evidence

from preclinical and clinical studies suggests that persistent inflammation functions as a driving

force in the journey to cancer. The possible mechanisms by which inflammation can contribute

24

to carcinogenesis include induction of genomic instability, alterations in epigenetic events and

subsequent inappropriate gene expression, enhanced proliferation of initiated cells, resistance to

apoptosis, aggressive tumor neovascularization, invasion through tumor-associated basement

membrane and metastasis, etc. Inflammation-induced reactive oxygen and nitrogen species cause

damage to important cellular components (e.g., DNA, proteins and lipids), which can directly or

indirectly contribute to malignant cell transformation. Overexpression, elevated secretion, or

abnormal activation of proinflammatory mediators, such as cytokines, chemokines,

cyclooxygenase-2, prostaglandins, inducible nitric oxide synthase, and nitric oxide, and a distinct

network of intracellular signaling molecules including upstream kinases and transcription factors

facilitate tumor promotion and progression. While inflammation promotes development of

cancer, components of the tumor microenvironment, such as tumor cells, stromal cells in

surrounding tissue and infiltrated inflammatory/immune cells generate an intratumoral

inflammatory state by aberrant expression or activation of some proinflammatory molecules.

Many of proinflammatory mediators, especially cytokines, chemokines and prostaglandins, turn

on the angiogenic switches mainly controlled by vascular endothelial growth factor, thereby

inducing inflammatory angiogenesis and tumor cell-stroma communication. This will end up

with tumor angiogenesis, metastasis and invasion. Moreover, cellular microRNAs are emerging

as a potential link between inflammation and cancer (Joydeb et al., 2012). Inflammation is

regarded as seventh hallmark of cancer (Francesco et al., 2009) .

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Fig 1.

Fig. 1. Inflammation as the seventh hallmark of cancer. An integration to the six hallmarks of cancer [modified from Hanahan and Weinberg (Hanahan, 2000) and Mantovani (Mantovani, 2009) and Francesco (Francesco et al., 2009).

Chronic inflammation as a predisposing factor for malignant transformation

of cells

Chronic inflammation represents a major pathologic basis for the majority of human

malignancies. The role of inflammation in carcinogenesis has first been proposed by Rudolf

Virchow in 1863, when he noticed the presence of leukocytes in neoplastic tissues (Balkwill and

Mantovani, 2001). Since the Virchow’s early observation that linked inflammation and cancer,

accumulating data have supported that tumors can originate at the sites of infection or chronic

inflammation (Mueller, 2004). Approximately, 25% of all cancers are somehow associated with

chronic infection and inflammation (Jackson, 2006; Perwez, 2007). Although inflammation acts

as an adaptive host defense against infection or injury and is primarily a self-limiting process,

inadequate resolution of inflammatory responses often leads to various chronic ailments

26

including cancer (Jackson, 2006; Schottenfeld,2006). Multiple lines of evidence from

laboratory and population based studies suggest that organ-specific carcinogenesis is partly

associated with a persistent local inflammatory state (O’Byrne et al., 2001; Itzkowitz et al.,

2004; Nelson et al., 2004; Whitcomb, 2004). For instance, the development of carcinomas of

stomach, liver, gallbladder, prostate and pancreas has been attributed to Helicobacter pylori-

induced gastric inflammation, chronic hepatitis, cholecystitis, inflammatory atrophy of the

prostate and chronic pancreatitis, respectively (Philpott, 2004; Jackson, 2006; Matsuzaki et al.,

2007). Patients suffering from inflammatory bowel disorders, such as ulcerative colitis and

Crohn’s disease, have an increased risk of developing colorectal cancer (Seril et al., 2003;

Itzkowitz et al., 2004; Herszenyi and Tulassay, 2007), while the management of colitis with

anti-inflammatory drugs reduces this risk (Eaden et al., 2000).

Inflammation-associated carcinogenesis: roles of reactive oxygen and nitrogen

species

Sustained cellular injuries can cause inflammation, which may lead to carcinogenesis.

Various inflammatory and innate immune cells (e.g., mast cells, neutrophils, leukocytes,

macrophages, monocytes, eosinophils, dendritic cells, phagocytes, and natural killer cells) are

often recruited at the site of infection or inflammation. In response to proinflammatory stimuli,

activated inflammatory/immune cells generate reactive oxygen species (ROS) and reactive

nitrogen species (RNS), which can function as chemical effectors in inflammation-driven

carcinogenesis. Thus, one of the plausible mechanisms by which chronic inflammation can

27

initiate tumorigenesis is the generation of ROS and/or RNS in the inflamed tissue and subsequent

DNA damage leading to activation of oncogenes and/or inactivation of tumor suppressor genes.

Chronic exposure to ultraviolet (UV) B radiation is known to precipitate inflammatory tissue

damage and skin cancer (GM, 2005). Mutational changes in ras and p53 have been observed in

many types of human cancer (Rajalingam et al., 2007; Strano et al., 2007). The activation of

ras oncogene and loss-of-function of p53 tumor suppressor gene have been implicated in UVB-

induced mouse skin carcinogenesis (Hattori et al., 1996). ROS-induced DNA damages

including DNA strand breaks, DNA base modifications, and DNA cross-links result in the

replication errors and the genomic instability and hence contribute to tumor initiation. Nitric

oxide (NO), another reactive species, plays a role in inflammation-associated carcinogenesis by

direct modification of DNA and inactivation of DNA repair enzymes. 8-Oxo-7,8-dihydro-20-

deoxyguanosine (8-oxo-dG), a major biochemical hallmark of oxidative and mutagenic DNA

damage (Hoki et al., 2007), has been found to be produced in association with H. pylori induced

gastric (Xu et al., 2004) and tumor necrosis factor-a (TNF-a)-induced pulmonary carcinogenesis

(Babbar et al., 2006). Peroxynitrite, a product formed by a reaction between NO radical and

superoxide anion, causes DNA damage by forming 8-nitroguanine (8-NG), which is another

potential biomarker of inflammation-associated cancers (Kawanishi, 2006). Thus, oxidative and

nitrosative DNA damage products, such as 8-oxo-dG and 8-NG, have been implicated in the

initiation of inflammation-driven carcinogenesis. ROS and RNS can induce lipid peroxidation to

generate other reactive species, such as manoldialdehyde and 4-hydroxynonenal (4-HNE), which

are capable of forming DNA-adducts. 4-HNE forms an adduct preferentially at the codon 249 of

the p53 gene. Elevated intracellular ROS (e.g., superoxide anion, H2O2, and hydroxyl radical) and

RNS (e.g., peroxynitrite, NO, and S-nitrosothiols) also cause alterations in cellular protein

28

functions, such as perturbation of DNA-protein cross-links and post-translational modification of

proteins involved in maintaining cellular homeostasis. For example, NO has been shown to

hyperphosphorylate and inactivate retinoblastoma protein resulting in increased proliferation of

human colon cancer cells. In colon tissues from patients with ulcerative colitis, a positive

correlation between the expression of iNOS and the phosphorylation of p53 at serine 15 residue,

as well as the activation of p53 transcriptional activity has been noted (Hofseth et al., 2003).

Nitrosative stress also plays a critical role in inflammation-associated carcinogenesis by

activating activator protein-1 (AP-1), a representative redox sensitive transcription factor, which

is involved in cell transformation and proliferation. Paradoxically, ROS and RNS can cause

apoptotic or necrotic cell death (Halliwell, 2007).

Major mediators linking inflammation and cancer

Chronic inflammation is implicated in all stages of carcinogenesis, i.e., initiation,

promotion and progression. In a persistently inflamed tissue, excessive generation of ROS can

cause genomic instability which leads to initiation of cancer (Philip et al., 2004; Hussain,

2007). A single initiated cell undergoes proliferation to produce a clone of mutated cells which

form premalignant mass, the event generally termed tumor promotion. Some of the preneoplastic

cells encounter additional mutations and become malignant. This process is referred to as tumor

progression. Proliferating tumor cells, their surrounding host stromal cells and tumor-infiltrating

inflammatory/ immune cells create a tumor microenvironment that reflects a persistent

inflammatory state (Balkwill and Mantovani, 2001; Ariztia et al., 2006). Within the tumor

microenvironment, various proinflammatory mediators participate in a complex inflammatory

29

signaling that facilitates extravasation of tumor cells through the stroma, thereby fostering tumor

progression (Balkwill and Mantovani, 2001; Ariztia et al., 2006)(Fig. 2).

Fig. 2. A journey to cancer: inflammation as the driving force. Inflammation is implicated in multi-stage carcinogenesis. ROS/RNS or other reactive species derived from inflammatory stress can attack DNA and cause mutations in oncogenes/tumor suppressor genes or other genetic alterations. This will lead to initiation of carcinogenesis. Inflammation also contributes to promotion and progression stages by stimulating the proliferation of initiated or premalignant cells, enhancing angiogenesis and metastasis, rendering precancerous or neoplastic cells resistant to apoptosis, etc., through epigenetic mechanisms (Joydeb et al., 2012).

Inflammation acts as a key regulator of tumor promotion and progression by several

mechanisms including acceleration of cell cycle progression and cell proliferation, evasion from

apoptotic cell death, and stimulation of tumor neovascularization (Philip et al., 2004). Among

the major molecular players involved in the inflammation-to-cancer axis, the notable members

are cytokines, chemokines, COX-2, prostaglandins, prostanoid receptors (EP 1–4), iNOS, NO,

and NF-kB. Table 1 represents the mechanisms by which the key inflammatory mediators

contribute to carcinogenesis.

30

Table 1 Key mediators linking inflammation and cancer (Joydeb et al.,2012).

CytokinesCytokines including interlukins, TNF-α, growth factors and differentiation factors are

secreted or membrane bound small protein molecules that regulate diverse physiological

processes, such as growth, development, differentiation, wound healing and immune response

(Lu and Huang, 2006; Miki et al., 2007). Cytokine signaling is initiated upon binding of

specific cytokines to cell-specific cognate receptors followed by activation of intracellular

kinases, such as Janus activated kinase (JAK), phosphatidylionositol-3-kinase (PI3/K)/ Akt, IKK,

and MAP kinases, with subsequent activation of transcription factors, predominantly STAT, NF-

kβ, and AP-1 (Jung et al., 2002; Yoshimura, 2006). The pleiotropic nature of cytokine

functions is evident from cross-regulation of one cytokine by other cytokines, differential

31

response of the same cytokine depending on the cell type, and synergistic or antagonistic effects

elicited by combined cytokine stimulation of cells (Szlosarek et al., 2006). Despite a complex

nature of their function, cytokines can broadly be classified as inflammatory (e.g., IL-1, IL-6, IL-

8, IL-17) and anti-inflammatory (e.g., IL-10) ones. Some cytokines have been reported to play a

role in inflammation associated carcinogenesis (Lin, 2007; Rigby et al., 2007). For example,

mice genetically modified to disrupt SOCS3 exhibit enhanced colonic crypt formation, crypt

proliferation, and the increased number and the size of colon tumors after challenge with dextran

sulfate sodium (DSS) or azoxymethane (AOM) plus DSS (Rigby et al., 2007). While persistent

local inflammation leads to cell transformation, a tumor cell further augments inflammatory

responses in its vicinity by secreting cytokines and chemokines, thereby creating a positive loop

between inflammation and cancer. Both cytokines and chemokines facilitate the communication

between tumor cells and tumor-associated host stromal tissue, thereby accelerating tumor

progression (Ben, 2003; Ariztia et al., 2006; Porta et al., 2007).

Chemokines

Chemokines are soluble chemotactic cytokines, which are classified as four major

groups, i.e., CXC, CC, XC and CX3C primarily based on the positions of conserved cysteine

residues (Balkwill and Mantovani, 2001; Lu and Huang, 2006; Allen and Handel, 2007). In

chronic inflammation, chemokines are usually produced by proinflammatory cytokines.

The central role of chemokines is to recruit leukocytes at the site of inflammation (Lu

and Huang, 2006). Most tumor cells can produce CXC and CC chemokines, which again differ

in selectivity for particular leukocytes. While lymphocytes represent a common target of both

CXC and CC, neutrophils are targeted only by CXC chemokines. CC chemokines can also act on

32

other leukocyte subtypes, such as monocytes and eosinophils as well as dendritic cells and

natural killer cells (Balkwill and Mantovani, 2001). Like cytokines, chemokines also act by

interacting with specific receptors expressed by both infiltrated leukocytes and tumor cells in an

autocrine or a paracrine fashion (Balkwill and Mantovani, 2001; Lu and Huang, 2006).

Several studies have reported the involvement of chemokines and chemokine receptors in cell

proliferation, migration, invasion and metastasis of different types of tumors. Overexpression of

CXCL-1/GROa, CXCL-2/GROb or CXCL-3/GROg promotes soft agar colony formation and

transformation of melanocytes in culture as well as tumorigenicity of transplanted melanoma

cells in nude mice.Treatment of cultured melanoma cells with anti-IL-8Rb antibody inhibited the

cell growth. Chemokine regulation of tumor angiogenesis results from a balance between

proangiogenic and angiostatic activities. Besides their role in chemoattraction of leukocytes,

chemokines direct the migration of tumor cells to the distal organs via circulation. The metastatic

potential of chemokines is attributed to their ability to induce the expression of matrix

metalloproteinases (MMPs), which facilitate tumor invasion. A stromal cell derived factor (SDF-

1)/CXCL-12 promoted the migration of colon adenocarcinoma (CT26) cells in culture and the

growth of implanted CT26 cells in BALB/c mice in vivo through angiogenesis-dependent

induction of tumor cell proliferation and inhibition of apoptotic cell death. Moreover, silencing

of endogenous CXCR4 gene expression by CXCR4-shRNA resulted in the inhibition of the

proliferation, adhesion, chemotaxis and invasion of mucoepidermoid carcinoma cells (Wen et

al., 2007).

COX-2 and prostaglandins

COX-2, an inducible form of cyclooxygenase, serves as an interface between

inflammation and cancer (Aggarwal et al., 2006; Surh, 2007). In response to various external

33

stimuli, such as proinflammatory cytokines, bacterial LPS, UV, ROS and phorbol ester, COX-2

is transiently elevated in certain tissues (Surh, 2007). Abnormally elevated COX-2 causes

promotion of cellular proliferation, suppression of apoptosis, enhancement of angiogenesis and

invasiveness, etc., which account for its oncogenic function (Fig. 3).

Fig. 3. Role of COX-2 and PGs in inflammation-induced carcinogenesis. Inflammatory signaling triggers induction of COX-2 expression and subsequently production of an array of prostaglandins. While some prostaglandins, especially PGE2, are implicated in carcinogenesis, others (e.g., PGI2) have cytoprotective effects. Still another group of prostaglandins, including PGD2 and 15d-PGJ2, have dual effects on carcinogenesis. PGDH by inactivating PGE2 can protect against carcinogenesis and is recognized as a tumor suppressor. EP and FP denote PGE2 and PGF2a receptors, respectively (Joydeb et al., 2012).

Cytokine polymorphisms and gastrointestinal malignancy

Genetic polymorphisms have emerged in recent years as important determinants of

disease susceptibility and severity. This is particularly true for cytokine gene polymorphisms and

gastrointestinal malignancy. Perhaps the most compelling evidence for the role of inflammation

in GI malignancy comes from studies showing that proinflammatory cytokine gene

polymorphisms increase the risk of cancer and its precursors. An excellent example of this is the

role of these polymorphisms in the pathogenesis of H. pylori-induced gastric cancer. H. pylori

causes its damage by initiating chronic inflammation in the gastric mucosa. This inflammation is

34

mediated by an array of pro- and anti-inflammatory cytokines. Genetic polymorphisms directly

influence interindividual variation in the magnitude of cytokine response, and this clearly

contributes to an individual’s ultimate clinical outcome. In the case of H. pylori infection, we

speculated that the most relevant candidate genes would be ones whose products were involved

in handling the H. pylori attack (innate and adaptive immune responses) and ones that mediated

the resulting inflammation. H. pylori-induced gastritis is associated with three phenotypes that

correlate closely with clinical outcome. The first is an antrum-predominant/corpus-sparing

pattern associated with high acid secretion and increased risk of duodenal ulcer disease. Second

is mild mixed antrum/corpus gastritis with no major effect on acid secretion and, generally, no

serious clinical outcome. The last is a corpus-predominant or severe pangastritis pattern that is

associated with gastric atrophy, hypochlorhydria, and an increased risk of gastric cancer.

Inhibition of gastric acid pharmacologically can lead to a shift from an antrum-predominant

pattern to a corpus-predominant one with onset of gastric atrophy. Thus it was clear that an

endogenous agent that was upregulated in the presence of H. pylori, has a profound

proinflammatory effect, and was also an acid inhibitor would be the most relevant host genetic

factor to be studied. IL-1β fitted this profile perfectly, for not only is it one of the earliest and

most important proinflammatory cytokines in the context of H. pylori infection, it is also the

most powerful acid inhibitor known (El Omar, 2001). We have shown that proinflammatory IL-

1 gene cluster polymorphisms (IL-1β encoding IL-1β and IL-1RN encoding its naturally

occurring receptor antagonist) increase the risk of gastric cancer and its precursors in the

presence of H. pylori (El Omar et al., 2000). Individuals with the IL-1β-31C or -511T and IL-

1RN2/2 genotypes are at increased risk of developing hypochlorhydria and gastric atrophy in

response to H. pylori infection. This risk is extended to gastric cancer itself with a two- to

35

threefold increased risk of malignancy compared with subjects who have the less

proinflammatory genotypes (El-Omar et al., 2000; El-Omar et al., 2003). The association of

IL-1 gene cluster polymorphisms and gastric cancer has been confirmed in other reports

(Figueiredo et al., 2002). In addition to IL-1 gene cluster polymorphisms, proinflammatory

genotypes of TNF-α and IL-10 have also been identified as risk factors for gastric cancer, and, as

is the case with IL-1 gene cluster polymorphisms, this is restricted to noncardia adenocarcinomas

(El-Omar et al., 2003). We have shown that having an increasing number of proinflammatory

genotypes (IL-1β -511T, IL- 1RN2/2, TNF-α-308A, and IL-10 ATA/ATA) progressively

increases the risk of gastric cancer. Indeed, by the time three to four of these polymorphisms are

present, the risk of gastric cancer is increased 27-fold (El-Omar EM 2003). The fact that H.

pylori is a prerequisite for the association of these polymorphisms with malignancy demonstrates

that in this situation, inflammation is indeed driving carcinogenesis. It is likely that other

proinflammatory cytokine gene polymorphisms will be relevant to gastric cancer initiation and

progression. This exciting field has expanded greatly over the past few years, and the search is

now fully on for the full complement of risk genotypes that dictate an individual’s likelihood of

developing cancer. This approach has now been adopted for many other cancers as described

below. In Japanese patients with chronic HCV infection, the IL-1β - 511 T/T genotype has been

associated with an increased risk of progression to hepatocellular carcinoma (Tanaka et al.,

2003). Because the T/T proinflammatory genotype is related to greater IL-1β production, it is

feasible that risk of malignant transformation is higher. IL-1β leads to the production of PGE2

and hepatocyte growth factor and has angiogenic influence via inducible NO and COX-2

expression. Furthermore, the degree of HCV induced liver inflammation and fibrosis has been

correlated with hepatic expression of Th1 cytokines. At present, there is relatively little

36

information on the relationship between other gastrointestinal malignancies and cytokine

polymorphisms. Some studies have addressed the influence of polymorphisms on cancer

outcome. Barber et al. (Barber et al., 2000) found that possession of a genotype resulting in

increased IL-1β production was associated with shortened survival in pancreatic cancer. Park et

al. (Park et al., 1998) investigated TNF-A and –B polymorphisms in 136 colorectal cancer

patients and 325 healthy controls in an Asian population. Their results indicated that

TNF-B1/TNF-B1 genotypes showed an increased risk for colorectal cancer. De Jong and

colleagues (De Jong et al., 2002) recently performed pooled analyses on 30 polymorphisms in

20 lowpenetrance genes and identified an additional three studies investigating TNF-α

polymorphisms and colorectal cancer.Associations were detected for the a2, a5, and a13 TNF-α

alleles and colorectal cancer.

Mechanisms of inflammation associated tumor development in the gastro

intestinal (GI) tract

The mechanisms employed by ROS, COX-2, and cytokines to promote neoplasia are

discussed, these mechanisms include direct DNA damage, inhibition of apoptosis, subversion of

immunity, and stimulation of angiogenesis. In addition, chronic inflammation in the GI tract is

also known to affect proliferation, adhesion, and cellular transformation. Deregulation of cellular

proliferation is one of the hallmarks of cancer cells and is the outcome of interaction between a

variety of endogenous and exogenous factors that are active during the inflammatory process.

These include luminal contents, bacteria, inflammatory cytokines, and mediators such as the

matrix metalloproteinases. Direct mechanical irritation can also lead to epithelial proliferation,

and when this is combined with the effects of an additional inflammatory stimulus, such as a

37

bacterium for example, the resulting hyperproliferation can push the tissue further along the

pathway toward cancer. H. pylori infection, although initially enhancing apoptosis, ultimately

leads to a compensatory proliferation (Yanai et al., 2003). Pathways through which H. pylori

may influence apoptosis include those involving COX-2 and peroxisome proliferator-activated

receptor-γ(Gupta RA 2001). Proinflammatory cytokines, particularly TNF-α, are also able to

modulate apoptosis through altering the levels of the pro and anti apoptotic proteins Bcl-2 and

Bax. As well as affecting proliferation and apoptosis, the same mediators impact on cellular

adhesion and angiogenesis. Cancer cells responding to proinflammatory cytokines released from

macrophages may exploit the same mechanism used by leukocytes to migrate through the

vasculature. Upregulation of cell adhesion molecule expression is seen on exposure of colon

cancer cells to LPS, and COX-2 has also been shown to promote cell adhesion (Tsujii, 1995;

Sutton et al., 2000). Macrophages are important sources of VEGF, and studies have shown that

this can be augmented in tumors by the humoral antitumor immune response (Barbera et al.,

2002). Thus a Th2 environment promotes angiogenesis, and conversely, CMI/Th1 immune

responses tend to be inhibitory. Although infection and inflammation initially generate Th1

cytokines, a cycle involving COX-2-mediated upregulation of Th2 cytokines and subsequent

chronic downregulation of the Th1/CMI immune response can develop in neoplasia. This is

elegantly illustrated by Dalgleish and O’Byrne (Dalgleish, 2002) in their review of chronic

immune activation and inflammation as the cause of malignancy, from which Fig. 2.6a has been

modified.

38

Fig. 4. Mechanisms of inflammation-associated tumor development in the gastrointestinal tract: angiogenesis. Chronic inflammatory stimuli such as Helicobacter pylori gastritis and inflammatory bowel disease (IBD) initially generate a Th1 proinflammatory cytokine response. However, via cyclooxygenase (COX)-2 generation, Th2 cytokine production is upregulated and there is negative feedback inhibition of the Th1 response. The presence of COX-2 and the change to a predominantly Th2 environment promote angiogenesis. COX-2 stimulates angiogenic growth factors, in particular VEGF, which is also stimulated by a number of other inflammatory mediators including nitric oxide (NO) and certain cytokines. All may be acting through the transcription factor hypoxia-inducible factor-1α (HIF-1α), which is regulated by NO, certain cytokines, and growth factors and which, in turn, binds to the VEGF promoter region leading to its activation (Mairi et al., 2004).

The transition to a predominantly Th2 immune environment favors angiogenesis, and

COX-2 itself has proangiogenic activity. Hypoxia is a potent inducer of VEGF, and this is

mediated by the transcription factor hypoxia-inducible factor- 1α(HIF-1α). The VEGF gene

contains a number of HIF-1α-binding sites in its regulatory region, and HIF-1αis able to activate

the VEGF promoter. Liu and colleagues (Liu et al., 2002) demonstrated that PGE2 production

via COX-2-catalyzed pathways plays a critical role in HIF-1α regulation by hypoxia. They

showed that tumors treated with a COX-2 inhibitor were smaller, with increased apoptosis,

decreased microvessel density, and decreased tumor VEGF levels. ROS, NO, certain cytokines,

and growth factors are also regulators of HIF-1α expression, and this may explain their

proangiogenic activity. The significance of HIF-1αin inflammation has been highlighted by

Cramer et al.(Cramer et al., 2003), who revealed that it controls the redness and swelling of

39

injured tissues and the ability of leukocytes to enter inflamed areas. In the low oxygen

concentration of injured, inflamed, or neoplastic tissue, HIF-1α is required to generate ATP in

leukocytes and thus enable them to function. It also increases the production of NO, which acts

back to further increase HIF-1α activity. Thus acting through HIF-1α in a hypoxic environment,

various inflammatory mediators including growth factors, NO, cytokines, chemokines, COX-2,

and its products may “switch” on angiogenesis. They may do this via the generation of VEGF,

either directly or indirectly, and may also aid the process by activating other factors such as

proteases, which degrade the extracellular matrix. Therefore, it is not difficult to see that in

chronic GI inflammation, the development of hypoxic areas may increase the generation of

proangiogenic stimuli that tip the balance in favor of angiogenesis and further drive tissues

toward carcinogenesis. Finally, in addition to impacting on cellular proliferation, apoptosis,

adhesion, and angiogenesis, the stimuli and mediators of chronic inflammation can cause cellular

transformation. A number of viruses such as HBV, Epstein-Barr, and HPV are known to directly

bind to certain genes and affect protein activity, including transcriptional factors and oncogenes.

Animal models have also demonstrated that bacteria can lead to ultrastructural changes within

the colonic epithelia and subsequent hyperplasia (Luperchio, 2001). This may precede the

development of colonic adenomas. Fig.2.6b, provide an overview of how chronic inflammation

in the GI tract may ultimately lead to malignancy. This takes into account the various cellular

processes and pathways through which inflammation and its by products are thought to exert

their effects. There is ample epidemiological evidence to support links between chronic

inflammation and carcinogenesis of the GI tract, but increasingly, the basic molecular pathways

of this association are being uncovered. Inflammatory cells produce a wide range of mediators

40

including proinflammatory cytokines, chemokines, ROS, growth factors, and eicosanoids. COX-

2 may be a linchpin in orchestrating many of the mutagenic effects of these products, and this is

supported by studies showing the chemopreventative benefits of COX inhibitors. Cytokine gene

polymorphisms undoubtedly contribute to individual risks of malignancy, but their importance

lies in their contribution to the understanding of inflammation-mediated carcinogenesis. The fact

that chronic inflammation impacts crucial cellular processes such as proliferation, adhesion,

apoptosis, angiogenesis, and transformation highlights its pivotal role in the pathogenesis of GI

malignancy (Mairi et al., 2004).

41

Fig.5. Overview of mechanisms of inflammation-associated gastrointestinal carcinogenesis. This diagram illustrates the complex interactions between inflammatory mediators and the common cellular processes that could lead to cancer. When the factors are considered in layers, the environment within the gastrointestinal tract forms the outermost layer. The presence of a rapidly proliferating mucosa exposed to exogenous substances and a vast quantity of LPS-rich bacteria provides an ideal environment for chronic inflammation to get established. Moving inward, the next layer involves the release of mediators such as proinflammatory cytokines, chemokines, growth factors, and NO, many of which share transcriptional factors such as HIF-1α and NF-kB. These may act independently or in combination, leading to the activation of COX-2. At the center are 5 cellular mechanisms that could form a pathway to malignant transformation, either independently or more likely in concert. COX-2 is known to impact on each of them and is placed in an inner layer because of the growing evidence supporting its central role in gastrointestinal carcinogenesis. However, the relationships among NO, cytokines, and activation of COX-2 and their impact on cellular mechanisms are not purely linear. As described in the review, all of these factors are capable of exerting effects independent of COX-2, and thus this layer may not always be a necessary component in revealing the final core of pathways to cancer (Mairi et al., 2004).

42

Overall cancer statistics indicate that gastric cancer (GC) is the fourth most common

cancer worldwide, with 934,000 cases per year in 2002 (8.6% of total cases), and has the second

highest mortality after lung carcinoma (about 700,000 deaths annually) (Parkin et al., 2005).

Over the past 10 years GC has become significantly more common in Great Britain, the United

States, and other countries (Wayman et al., 2001). In Russia, GC is the second most widespread

malignancy, with approximately 48,300 cases per year (Aksel et al., 2001). Unfortunately, GC

prognosis remains poor. According to modern statistical data, the survival rate is around 25–30%

worldwide (mean lifetime, 17.2 months) after chemotherapy and radiotherapy (Parkin et al.,

2005). For unknown reasons, males are more predisposed to GC than females (Parkin et al.,

2005). Gastric carcinogenesis is a multifactorial process that includes environmental, host,

genetic, and bacterial factors. The main and foremost environmental etiological factors for GC

risk are nutrition pattern and individual dietary choices. High intakes of red or processed meat

(Larsson et al., 2006) and salty foods (Panel et al., 1997), excessive alcohol consumption (Liu

et al., 2008), and tobacco smoking (Tredaniel et al., 1997) may contribute to GC development.

Obesity has also been determined to be an etiological factor, but only for the cardiac subtype of

GC (Lagergren et al., 1999). A connection between Helicobacter pylori (HP) status and high

risk of GC has also been established (Group, 1993), and after numerous investigations this

bacterium was officially adopted as an etiological factor by the International Agency for

Research on Cancer (Hxcker et al., 2003). Modern cancer research is concerned with the

investigation of individual susceptibility of the human genome to various risk factors. During the

past decade many correlations between single nucleotide polymorphisms (SNPs) in DNA

structure and the risk of various diseases, including cancer, were reported (Tsigris et al., 2007).

43

Therefore, these polymorphisms can be applied as specific markers of predisposition for tumor

prevention.

Interleukins in gastric cancer development

Interleukins (ILs) belong to a diverse family of cytokines and represent small, specific

cell signaling protein molecules, which regulate the immune system of an organism. ILs are

synthesized predominantly by T cells, monocytes, macrophages, and endothelial cells. Their

functions are plural: facilitating communication between immune cells, controlling genes,

regulating transcription factors, and governing the inflammation, differentiation, proliferation,

and secretion of antibodies (Salazar et al., 2007). Thus, humoral regulation of the immune

response is greatly dependent on the functioning of interleukins. ILs implement cell coordination

by creating signals (Salazar et al., 2007), which provide autocrine or paracrine regulation. Each

IL has a specific ligand-dependent receptor (IL-R), which is expressed on the surface of the

target cell and directly participates in signaling. These receptors are membrane glycoproteins,

which consist of an external immunoglobulin-like domain, a transmembrane region, and a

cytoplasmic domain (McMahan et al., 1991). The impact of IL on a cell is a multistep process.

First, IL binds to the receptor on the cell surface and forms a single complex, which leads to

conformational changes in IL-R, bringing the JAKs (specific tyrosine kinases) close enough to

autophosphorylate themselves. Autophosphorylation of JAKs induces a conformational alteration

in its own structure, enabling it to further phosphorylate and activate transcription factors called

signal transducers and activators of transcription (STATs). Activated STATs dissociate from the

receptor and form dimers before translocating to the cell nucleus, where they regulate the

transcription of selected genes (Fig. 2.7). There are 35 ILs identified in the human body (Doan,).

Their coordinated work ensures the correct and effective functioning of immune system. Any

44

dysregulation between IL interactions or disruptions in the JAK/STAT pathway may lead to DNA

damage, excessive production of tumor-inducing factors, immune disorders, angiogenesis, and

dysplasia (Lederle et al., 2011). In addition, they often result in malignant transformation and

further formation of metastasis (El-Omar, 2006). Disruptions and dysregulations may be caused

by several factors, including activities of intracellular agents (Brauer et al., 2010), viruses

(Tacke et al., 2011), infections (Group, 1993), or smoking (Macha et al., 2011). Genetic

factors play an essential role in cytokine balance, including genetic polymorphisms. Thus, it

becomes clear that ILs may contribute to cancer development. In fact, many studies have proven

that the presence of a tumor is often connected with a high serum concentration of certain ILs.

An increased serum level of proinflammatory ILs was identified in patients with hepatocellular

carcinoma, oral squamous cell carcinoma, and prostate cancer (Al-Wabel et al., 1995; Tsai et

al., 1999; Michalaki et al., 2004). Anti-inflammatory ILs were also reported to contribute to the

development of cancer: high serum levels of IL-10 and IL-4 were indicated among melanoma,

gastric, pancreatic, and prostate cancer patients (Fortis et al., 1996; Takeshi et al., 2005).

Hence, a dramatic alteration of the IL level may reflect dysregulations in the production of ILs.

Individual genetic differences caused by SNPs may be closely related to these disruptions and

eventually play a role in carcinogenesis (Arseniy, 2011).

45

Fig. 6 Schematic plan illustrates how interleukins (ILs) mediate cell functioning. 1. IL binds with the corresponding IL receptor (IL-R), inducing conformational change. 2. Activated JAK phosphorylates STAT with mediation of the IL/IL-R complex. 3. STAT forms the dimer, which is able to enter the nucleus and bind to specific DNA sequences in the promoters of genes that begin transcription of needful genes.

Several inflammatory interleukins have been linked with tumorigenesis, which suggests

that inflammation is associated with cancer development (Table 2). These interleukins include

IL-1, IL-6, IL-8,IL-10 and IL-18. Interleukins mediate different steps in the pathway leading to

tumorigenesis. Secretion of IL-1α promotes growth of cervical carcinoma (Woodworth et al.,

1995) and can also induce anchorage independence in embryo fibroblasts and tumor cell

revertants (Vanhamme et al., 1993). Autocrine production of interleukin IL-1β promotes

growth and confers chemoresistance in pancreatic carcinoma cell lines (Arlt et al., 2002). High

levels of IL-1β have been identified as a key mediator of this activation in two of the

chemoresistant pancreatic cell lines. IL-1β secretion into the tumor milieu also induces several

46

angiogenic factors from tumor and stromal cells that promotes tumor growth through

hyperneovascularization in lung carcinoma growth in vivo (Saijo et al., 2002). IL-6 acts as a

paracrine growth factor for multiple myeloma, non-Hodgkin’s lymphoma, bladder cancer,

colorectal cancer, and renal cell carcinoma (RCC) (Klein et al., 1989; Okamoto et al., 1995;

Voorzanger et al., 1996; Angelo et al., 2002). Autocrine IL-6 production in RCC has been

linked with the involvement of p53. RCC cell lines containing mutant p53 produced higher

levels of IL-6 than those containing wild-type p53 (Angelo et al., 2002). Another important pro-

inflammatory cytokine IL-8 has been reported to promote growth and metastasis of wide variety

of tumors. Expression of IL-8 by human melanoma cells and human ovarian cancer cells

correlates with their metastatic potential (Luca et al., 1997; Xu, 2000; Huang et al., 2002). IL-8

has been detected in astrocytomas, anaplastic astrocytomas, glioblastomas, and central nervous

system cervical carcinoma metastasis. Thus, IL-8 secretion could be a key factor involved in the

determination of the lymphoid infiltrates observed in brain tumors and the development of

cerebrospinal fluid pleocytosis in persons with meningoencephalitides (Van et al., 1992).

Polymorphisms in the IL-8 gene contributes to a high risk of gastric cardia adenocarcinoma

(GCC) and esophageal squamous cell carcinoma (ESCC) among the population of Linxian in

north-central China (Savage et al., 2004). IL-8 has been found to be transcriptional target of Ras

signaling. Ras-dependent IL-8 secretion was required for the initiation of tumor-associated

inflammation and neovascularization(Sparmann A 2004). Constitutive production of IL-18,

RANTES, and MIP-1b, has been linked to disease progression in large granular lymphocyte

(LGL) leukemia (Kothapalli et al., 2005).

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Table 2 – Role of inflammatory interleukins and chemokines in tumorigenesis

IL-1β polymorphisms and gastric cancer

IL-1 plays a significant role in immune response signaling pathways (Dinarello, 1994).

The IL1 cluster consists of 3 related genes: IL1A, IL-1β, and IL1RA, which encode the signal

proteins IL-1α, IL-1β, and their receptor, IL-1RA, respectively. The relation of IL-1β and IL1RA

gene polymorphisms to GC development was observed in numerous studies (El-Omar et al.,

2000). Overexpression of the IL-1β gene is suggested to be promoted by H.pylori infection, as

well as chronic atrophic gastritis or duodenal ulcer (Blanchard et al.,1998). Overexpression of

IL-1β inhibits the secretion of gastric acid, which consequently results in the overexpression of

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gastrin and may stimulate neoplastic growth (Rozengurt et al.,2001). However, overexpression

of the IL-1β gene may also be caused without H pylori exposure because of SNPs(Kuipers et al.,

1995). There are 3 diallelic polymorphisms at positions –511 (C-T, rs16944), –31 (T-C,

rs1143627), and-3954 (C-T, rs1143634) base pairs from the transcription start site, which are

involved in the regulation of IL-1β expression. Other SNPs of the IL-1β gene have been

investigated less intensively. Four comprehensive meta-analyses on the IL-1β gene were chosen

to assess the role of these SNPs in GC. Researchers have been studying the T allele of the -511

SNP for a long time. First, Camargo et al. (Camargo et al., 2006) demonstrated that –511 TT

carriers had increased GC risk in comparison with CC wild-type genotype carriers in 14 studies

(odds ratio [OR] -1.21). The effect was more evident for T allele carriers with noncardiac and

intestinal GC subtypes (OR -1.66 and 1.80, respectively) among Caucasians. Interestingly, there

were no correlations among Asian populations. An analysis of 39 case–control studies conducted

by Wang et al. (Wang et al., 2006) revealed similar results: the T allele of –511 was more

frequent among patients with GC (OR -1.26). Histopathologic stratification revealed that the

association was stronger for patients with intestinal subtype (OR -1.76) but not with diffuse type

(OR -1.16), which confirmed the results of Camargo et al. (Camargo et al., 2006). The authors

also indicated that there was no impact of ethnicity and H. Pylori status on cancer risk in this

study. The results of Xue et al. (Xue et al., 2010) also confirmed the findings of previous

authors: the T allele correlated with higher intestinal and noncardiac GC risk in 18 studies versus

CC genotype (OR -1.55 and 1.33, respectively). Interestingly, these significant results were also

obtained among Caucasians but not in Asian or Hispanic populations (OR -1.33). The fourth

meta-analysis, which included 28 studies, was performed by Kamangar et al. (Kamangar et al.,

2006). They reported an absence of correlations between CT/TT genotypes and the GC genotype

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in comparison with the CC wild-type genotype (OR-1.07 and 1.16, respectively). Ethnic

stratification also indicated a lack of significant differences, as well as analysis of histopathology

or tumor site. Such controversial results may be explained by possible differences in study

design, inclusion criteria of investigations, errors during the statistical analysis, differences in

stratification, sample size, and chance. However, an overwhelming majority of studies

characterize the T allele of the IL-1β –511 polymorphism as frequent among Caucasian

individuals with noncardiac GC, preferably for the intestinal subtype of this cancer. Therefore, it

is feasible to suggest this SNP as a potential predictive marker for GC. Investigations of the IL-

1β –31 TATA-box polymorphism continue to be controversial even after several meta-analyses

have been carried out. Camargo et al. (Camargo et al., 2006) revealed a slight nonsignificant

connection between the C variant allele and GC risk compared with TT homozygotes (OR -1.04)

in 14 studies. Again, there was no association among Asian populations (OR -0.91) in

comparison with Hispanic or Caucasian populations (OR -0.91, 1.52, and 1.11, respectively).

Histologic stratification indicated a moderate increase of intestinal GC subtype among C allele

carriers in Caucasian populations (OR -1.61), but this statement was not true for the diffuse

subtype of GC. No associations were observed in the studies of Wang et al. (Wang et al., 2006),

Xue et al. (Xue et al., 2010), and Kamangar et al. (Kamangar et al., 2006) (OR -1.00, 0.97, and

0.99, respectively). Subgroup analysis did not indicate any correlations. Investigations on the IL-

1β –3954 SNP have indicated discordant results. Camargo et al. (Camargo et al.,2006) indicated

a slight non significant increase of GC risk among T mutant allele carriers compared with CC

individuals (OR-1.26), and the effect was more evident among Asian populations (OR -1.73).

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However, the small number of studies and the deficiency of T and CT allele carriers in some

studies could distort the results. Wang et al.(Wang et al., 2006) also determined that the T allele

of the -3954 gene polymorphism contributes to cancer risk (OR -1.37) in comparison with the

CC genotype, although no analyses of H.pylori status, tumor location, or subtype were

performed. Xue et al. (Xue et al., 2010) reported a lack of association between the -3954

polymorphism and GC. A small number of studies on the IL-1β -3954C/T polymorphism have

been conducted. A correlation of the T allele with GC was identified, which indicates a potential

role in gastric carcinogenesis. Recently, Lee et al. (Lee et al., 2004) discovered a new promoter

IL-1β –1473 G to C (rs1143623) polymorphism and reported an association between its G allele

and increased risk of the intestinal type of GC (OR -1.8 for the CG genotype and 2.1 for the GG

genotype) among a Korean population. The significance of this finding should be proven by

further functional and genetic association studies. It is important not to overlook the fact that the

above mentioned meta-analyses (Kamangar et al., 2006; Wang et al., 2006; Xue et al., 2010;

Camargo et al., 2006) are mostly composed of the same original studies, and therefore some

degree of overlap exists. First, meta-analyses devoted to the IL-1β –511C/T polymorphism are

composed of materials from 10 same case–control studies. Materials from 11 studies were

included in the above-mentioned meta-analyses on the IL-1β –31T/C SNP and data from 7

papers were involved in the meta-analyses on the IL-1β -3954C gene polymorphism. Such

overlapping of data creates sufficient difficulties in comparison of these meta-analyses among

each other, although the main trends and relations could be defined. The role of the T allele of

the IL-1β –511 polymorphism may be defined as cancer predictive. The impact of the –31 C

allele remains unclear based on contradictory results. Several epidemiologic studies have

reported that the T allele of the IL-1β –31 SNP is associated with vulnerable to persistent

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H.pylori infection, which can be modified by smoking (Hamajima et al., 2001), but according

to case–control studies, this allele apparently does not play a role in the development of GC,

even taking into account the fact that some studies confirm this link.

The IL-1β gene encoding IL-1β has two diallelic polymorphisms in the promoter region

at positions -511 and -31, representing C/T and T/C transitions, respectively, in near total linkage

disequilibrium (Hamajima et al., 2001; Machado et al., 2001; Machado et al., 2003). The less

common alleles of these loci (IL-1β -511T and IL-1β -31C) have been found to be associated

with Gastric carcinoma (Bidwell et al., 1999; El Omar et al., 2000; Machado et al., 2003). The

capacity to produce different cytokines varies among different individuals and may be

genetically determined. Such interindividual differences can be attributed to several molecular

mechanisms, including single nucleotide polymorphisms (SNPs) in the functional regions of

cytokine or cytokine receptor genes. These SNPs may affect the overall expression and secretion

of cytokines and may account for some of the heterogeneity of infectious diseases. These two

SNPs were associated with an increased risk of gastric cancer in Scottish and Polish subjects

( El-Omar and Bream 2000) and were subsequently confirmed by studies in other ethnic groups

from the USA (El-Omar et al., 2003) and Portugal (Machado et al., 2003). However, several

other studies failed to demonstrate the correlation (Kato et al., 2001; Zeng et al., 2003). The

diverse ethnic background with different sample size, different environmental exposures

infection in the population and genetic heterogeneity in the pathogenesis of gastric cancer may

account for the variability in different studies. Therefore, whether these polymorphisms of IL-1β

gene were related to the risk of gastric cancer remains inconclusive and needs to be replicated in

ethnically diverse populations.

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IL-8 polymorphisms and gastric cancer

Interleukin-8 (IL-8), a member of the CXC chemokine family, is a chemoattractant of

neutrophils and lymphocytes. A wide variety of normal and tumour cells could express IL-8, and

the principal role of IL-8 is to initiate and amplify acute inflammatory reactions. Additionally,

growing evidence has shown that the important roles IL-8 may play in the pathogenesis of

cancer, including angiogenesis, tumour growth, and metastasis. The interleukin-8 (IL-8) gene,

located on chromosome 4q13– q21 in humans, is composed of four exons, three introns, and a

proximal promoter region. IL-8enhances cell proliferation and migration and acts like a

chemoattractant for neutrophils and leukocytes, as well as a proangiogenic factor and mediator of

chronic inflammatory processes (Brat et al., 2005). The angiogenic and tumorigenic properties

of IL-8 were proven in experiments with mice long ago (Kitadai et al., 1999), and recent studies

revealed its involvement in adhesion, migration, and invasion in human GC cells in vitro (Ju et

al., 2010). The mucosal levels of IL-8 were reported to be elevated among GC patients in

comparison with healthy individuals, and the prognosis for patients with a high expression of IL-

8 was significantly poorer compared with that of patients who had a moderate level of this

protein (Kido et al., 2001). IL-8 may stimulate the expression of Reg protein in stomach cells,

which intensifies the proliferation of gastric mucosal cells and may indirectly promote GC

initiation (Yoshino et al., 2005). Moreover, IL-8 was reported to be involved in lung cancer

etiopathogenesis as well as in the initiation and progression of multiple myeloma(Xie JY 2010;

Vetvicka V 2011). Undoubtedly, these observations indicate a carcinogenic role of IL-8 and

underscore the significance of studying its tumorigenic properties. Fifteen functional

polymorphisms were identified in the IL-8 gene locus, and several may alter gene expression

(Hull et al., 2004). These “altering” SNPs were investigated in several case–control studies: IL-

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8–251A/T (rs4073), IL-8-396T/G (rs2227307), and IL-8-781C/T (rs2227306). First, the A allele

of the IL-8–251A/T polymorphism was associated with elevated IL-8 synthesis and also

contributed to the development of breast and bladder carcinomas (Hull et al., 2004; Lu et al.,

2007; Ahirwar et al., 2010; Snoussi et al., 2010). This allele correlated with increased cancer

risk compared with the TT genotype, according to the study of Lu et al. (Lu et al., 2007) (OR -

1.21). Kang et al. (Kang et al., 2009) also observed that the AA genotype of H.pylori. positive

individuals is associated with a higher GC risk (OR - 2.0) in comparison with H pylori.positive

controls. Unfortunately, no data regarding ethnic impact, histopathology, or tumor site were

available. These findings forward a hypothesis that the presence of a high producing allele

dramatically increases the production of the IL-8protein, which may lead to malignant

transformation resulting from the angiogenic and iatrogenic properties of this cytokine.

However, the analysis of Sugimoto et al. (Sugimoto et al., 2010) indicated an absence of

association between A allele carriers and noncardiac GC risk (OR -0.99), whereas peptic ulcer

risk was significantly higher among Western compared with Asian non-GC patients (OR-1.49).

Most studies in the meta-analysis have reported positive associations for Asians and negative

associations for Caucasians. The sample size was approximately equal to that in the study of Lu

et al. (Lu et al., 2007), and discrepancies between these investigations are difficult to explain.

Possible reasons may be differences in ascertainment, study design, and populations, as well as

features of the analysis. The relationship between other polymorphisms of the IL-8 gene (-396

and -781) and GC risk has not been established by the combined analysis of Sugimoto et al.

(Sugimoto et al., 2010), although Savage et al. (Savage et al., 2004) demonstrated a correlation

of the IL-8-251/-396/-781 AGT/ AGC haplotype with a 4-fold increased risk of cardiac GC (OR

- 4.14). Taking into account the fact that the AA genotype of –251 was associated with a 2-fold

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greater risk of GC (OR - 1.96), the contribution of -396 and -781 polymorphisms is obvious.

Sugimoto et al., (Sugimoto et al., 2010) demonstrates the efficiency of studying the cumulative

effect of haplotype, which may indicate a more sensitive result or reflect a lesser or greater share

of each polymorphism in its total impact. According to the above, the role of IL-8 gene

polymorphisms in GC remains obscure because of contradictory findings. However, the -251

gene polymorphism seems to play a major role in GC development and requires further

investigation.

Alterations in various genetic factors are important in increasing gastric cancer risk (Jung

et al., 2011).IL-8 a chemoattractant of neutrophils and lymphocytes, a wide variety of normal

and tumour cells could express IL-8, and role of IL-8 is to initiate and amplify acute

inflammatory reactions. Growing evidence has shown that the important roles IL-8 may play in

the pathogenesis of cancer, including angiogenesis, tumour growth, and metastasis (Lin et al.,

2010). Number of molecular epidemiological studies have been done to evaluate the association

between IL-8 –251 A/T polymorphism and tumour risk in diverse populations. The tumour types

included gastric cancer, (Taguchi et al., 2005) breast cancer(Kamali et al., 2007) colorectal

cancer, (Cacev et al., 2008) and lung cancer, (Campaet al., 2005). Taguchi and colleagues

reported that the IL-8–251AA genotype of IL-8 was associated with a significantly increased risk

of gastric cancer in a Japanese population (Taguchi et al., 2005); nevertheless, Savage and

colleagues did not find any significant association between –251A/T polymorphism of IL-8 and

gastric cancer in a case–control study based on a Polish population.(Savage et al., 2006). Many

studies have demonstrated the relationship between IL-8 and the risk of GC. IL-8 expression is

also strongly correlated with neovascularization in the tissues from GC patients. The -251 A/T

polymorphism in the IL-8 promoter region has been associated with increased expression of IL-8.

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Many researchers reported that the -251 A genotype is associated with the risk of GC as well as

antral atrophy and metaplasia compared with the T genotype. Furthermore, the association

between the -251 A genotype and the risk of GC varied according to histological type and tumor

location. Meta analysis done by Liu et al showed IL-8 -251 A/T polymorphism was not

associated with the risk of GC and the association may be varied when stratified for histological

type, tumor location and ethnicity/country (Liu et al., 2010).

IL-10 polymorphisms and gastric cancer

IL-10 generally functions as an immunosuppressor and anti-inflammatory mediator and is

also known as cytokine synthesis inhibitory factor for its ability to decrease the synthesis of

proinflammatory cytokines by activated T cells and NK cells. IL-10 also blocks the antigen-

presenting abilities of macrophages and stimulates the proliferation of B cells, T cells, and mast

cells (Rousset et al., 1992). Many recent in vitro and in vivo investigations observed an impact

of IL-10 on autoimmune diseases and progression of malignancies (Asadullah et al., 2003).

Throughout the past 2 decades intensive genetic association studies have demonstrated

controversial results on polymorphisms in the IL-10 gene, but maintain that they promote cancer

appearance and development one way or another. There are 3 functional promoter SNPs in the

IL-10 locus at –1082 (A to G, rs1800896), –819 (C to T, rs1800871), and –592 (A to C,

rs1800872) pairs from the transcriptional start site. Most studies on these polymorphisms have

investigated haplotypes, but not distinct polymorphisms, which is important. The GCC haplotype

for IL-10 –1082/-819/-592 polymorphisms is associated with higher cytokine production

compared with the ATA haplotype. Alteration of the IL-10 protein level may be directly linked

to the fact that polymorphisms of the IL-10 gene are associated with susceptibility to prostate,

cervical, bladder, and breast carcinoma (Ahirwar et al., 2009; Gerger et al., 2010; Liu et al.,

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2010; Matsumoto et al., 2010). Furthermore, a high serum level of IL-10 was reported to

contribute to chronic hepatitis C and several autoimmune diseases (Edwards et al., 1999). Wu et

al.(Wu et al., 2003) revealed that the high-producing GCC haplotype of IL-10 -1082/-819/-592

genetic polymorphisms was more frequent among GC patients in comparison with healthy

controls in a Thai population (OR - 2.67). The effect was more evident for cardiac GC (OR -

3.21) compared with a noncardiac tumor location (OR- 1.96). A high GC risk was also reported

for smokers and H pylori.-infected carriers of the GCC haplotype, whereas for both smokers and

H pylori.-positive patients the risk was extremely high (OR -3.05, 2.32, and 6.00, respectively).

Sugimoto et al. (Sugimoto et al., 2007) also identified the GCC combination as a risk haplotype

for GC in comparison with the ATA haplotype among H pylori.-positive Japanese individuals

(OR- 2.805). In contrast with previous papers, El-Omar et al. (El-Omar et al., 2003)

demonstrated the hypoactive ATA haplotype to be associated with noncardiac GC (OR -2.5).

Carriers of the low-producing haplotype apparently have a stronger inflammatory response to H

pylori. infection, which may result in the reduction of acid production, atrophy of the stomach,

and progressive growth of neoplastic cells. However, it remains unclear how the high-producing

GCC haplotype correlates with increased cancer risk. Perhaps this correlation may be explained

by the immunosuppressive properties of IL-10 or by the inhibitory effect on other cytokines

involved in antitumor defense. Finally, separate gene polymorphism investigations, among

which are the IL-10 –1082A/G, IL-10 –592A/C, and IL-10 –819C/T polymorphisms,

demonstrated the absence of significant differences in genotype distribution between patients

with GC and healthy controls (Zhou et al., 2008; Chen et al., 2010; Zhuang et al., 2010). In

addition, Kang et al. (Kang et al., 2009) reported that CC carriers of the IL10–592A/C

polymorphism have a decreased risk of intestinal GC (OR -0.4). These findings revealed the

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advantage of haplotype analysis, which allows to define plausible and consistent results. The

investigations on IL-10 gene polymorphisms allow to suggest that they impact GC (Arseniy,

2011).

Though several genetic changes may occur in gastric cancer, but polymorphisms in the

IL-1, IL-8 and IL-10 genes are the most common and most specific genetic abnormalities in

gastric cancer; this motivated us to study the aberrations of these genetic events in the

etiopathogenesis of gastric cancer.

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