Pathophysiology of Gastric Cancer

Cancer is currently the second leading cause of death in the world behind

cardiovascular diseases. It is estimated that more than 1.6 million new cases

of cancer are diagnosed in every year (American Cancer Society, 2012). Cancer

is characterized by the proliferation of abnormal cells that fail to respond

correctly to normal regulatory mechanisms. Carcinogenesis, a term used to

describe cancer development, is a multiple-step process consisting of

initiation, promotion, and progression of uncontrolled cells. At the initiation

step, damage to DNA occurs. Finally, cells begin to proliferate and expand

into abnormal cells during the promotion step and during the progression

step, further changes occur to these abnormal cells leading to formation of

malignant cells (Klaunig J, et al., 2004).

The incidence of gastric cancer is different throughout the world and 60% of

deaths from gastric cancer occur in developing countries(Forman D, et al.,

2006; Liu T S, et al., 2008). Cancers of the gastrointestinal tract, including

esophageal, stomach, liver, colon, and pancreas are responsible for

approximately 3 million new cases and over 2 million deaths each

year(Hamilton S R, et al., 2000). Malignancies of the G.I. tract are relatively

resistant to radiation therapy while chemotherapy has modest benefit so an

effective therapy still remains elusive in the treatment of gastroduodenal

ulceration(Luck G D, 1998). Early diagnosis of human gastric cancer or tumor

recurrence is primarily based on endoscopy, biopsy and pathological

examination. Endoscopy is a widely used method for detecting early stages of

gastric cancer (Tashiro A, et al., 2006; Lu X et al., 2008).

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 Pathophysiology of Gastric Cancer

Pathology

The primary epithelial tumor of the stomach is the adenocarcinomas

develop from the stomach mucosa, usually maintaining glandular

differentiation. Most common tumors 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.

The diagnosis of gastric cancer is often delayed by the lack of early

symptoms, with early gastric cancer causing non-specific gastrointestinal

complaints, such as dyspepsia, in only 50% of patients. Up to 90% of Western

gastric cancer patients first present with advanced carcinomas, which have

more serious symptoms such as abdominal pain, bleeding, vomiting, or

severe weight loss. Endoscopic screening is considered to be the most

sensitive and specific diagnostic test for gastric cancer. Dysplasia may

present as a flat lesion or exhibit polypoid growth, with depressed, reddish or

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 Pathophysiology of Gastric Cancer

discoloured mucosa. Endoscopic detection of changes in colour, relief, and

architecture of the mucosal surface enables the classification of gastric

cancers according to their macroscopic growth pattern. Early gastric cancers

may feature protruded (Type I), elevated (Type IIa), flat (Type IIb),

depressed (Type IIc) or excavated (Type III) growth (Hamilton S R, et al.,

2000), whereas advanced gastric carcinomas are classified into polypoid

(Type I), fungating (Type II), ulcerated (Type III) or infiltrative (Type IV)

growth patterns (Borrmann R, 1926; Hamilton S R, et al., 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.

Epidemiology

Gastric cancer is one of the most common cancers worldwide, ranking

fourth in overall frequency, and accounting for over 870,000 new cases and

over 650,000 deaths annually in world. (Stewart B W, et al., 2003). Mortality

from gastric cancer is second only to lung cancer. Gastric cancer occurs more

frequently in men than in women, with the estimated number of new cases

worldwide being 558,000 for males and 317,000 for females, respectively

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 Pathophysiology of Gastric Cancer

accounting for 5.5% and 3.1% of all malignancies, excluding skin cancer

(Hamilton S R, et al., 2000). The geographic distribution of gastric cancer

varies from an annual incidence of more than 300,000 new cases in the more

developed regions, to nearly 550,000 new cases per year in the developing or

less developed regions of Africa, Latin America and the Caribbean, Asia

(excluding Japan), Micronesia, Polynesia and Melanesia. In high risk areas,

the intestinal type adenocarcinoma is more frequent, whereas the poorly

differentiated diffuse type carcinoma predominates in low risk areas.

The incidence and mortality rates of gastric carcinoma are steadily

declining. However, due to the aging population, the absolute number of new

cases per year is increasing (Munoz N, et al., 1971; Hamilton S R, et al., 2000).

Below the age of 30, the incidence of gastric carcinoma is extremely rare, but

thereafter rises quickly and continuously, with the oldest age groups having

the highest rates. In males, the intestinal type is more common than the

diffuse type and the incidence rises faster with age, whereas the diffuse-type

mainly impacts younger individuals, frequently females. A decline in

incidence of the intestinal-type carcinomas is largely responsible for the

decline in overall incidence rates (Kaneko s, et al., 2001; Henson D E, et al.,

2004), and has been correlated with the corresponding decrease in

prevalence of H. pylori infection (The Eurogast Study Group, 1993; Konturek

P C, et al., 2003).

Both gastric cancer and H. pylori infection affect patients from low

socioeconomic backgrounds, associated with low social class, poor education,

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 Pathophysiology of Gastric Cancer

low hygiene standards, a diet lacking fresh fruit and vegetables, but rich in

starch and preserved meats, and atrophic gastritis. Indeed, the distinctive

epidemiological characteristics of gastric cancer, in particular, the regional

differences and chronological changes in incidence may be, in part, related to

H. pylori infection (Nagel G, et al., 2007). However, the incidence of the

diffuse-type carcinoma may be increasing (Craanen M E, et al., 1992; Henson

D E, et al., 2004), which is worrying given that these types of tumors have a

worse prognosis (Blok P, et al., 1997). An increase has also been observed for

cancers localized to the gastro-oesophageal junction, some probably

originating from the distal esophagus caused by gastro-oesophageal reflux

(Yamada Y, et al., 1989). Cancers of the cardia and gastroesophageal junction

are conspicuously increasing in incidence and frequently exhibit a different

pathogenesis to non-cardia carcinomas.

Pathogenesis

The pathogenesis of gastric cancer involves multiple risk factors

including dietary, infectious, occupational, genetic and preneoplastic risk

factors, most of which act on the gastric mucosal microenvironment over a

prolonged time period. The resultant sequential changes in the gastric

mucosa that precede the development of invasive cancer are known as the

“precancerous cascade”, first described in 1975 (Correa P, et al., 1992),

where normal gastric mucosa is transformed by chronic atrophic gastritis

and develops multifocal atrophy and intestinal metaplasia, followed by the

appearance of dysplasia and finally invasive carcinoma.

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 Pathophysiology of Gastric Cancer

Past research has concentrated on the identification of the complex

etiology of environmental and genetic risk factors, which may influence the

initiation, promotion, and progression of gastric cancer (Stadtländer C T, et

al., 1999; Chan A O, et al., 2001; Correa P, 2002; Kelley J R, et al., 2003).

Histological Classification

Various systems have been applied to the classification of gastric

carcinomas, including the WHO (Hamilton S R, et al., 2000), Ming (Ming S C,

1977), Lauren (Lauren P, 1965), and Goseki (Goseki N, et al., 1992)

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 (Wittekind C et al., 2002), is the most important prognostic

factor in clinical practice (Alekseenko S A, 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 Lauren classification (Lauren P, 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. Typically arising on a

background of intestinal metaplasia, these tumours exhibit an intestinal,

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 Pathophysiology of Gastric Cancer

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

Lauren classification.

World Health Organization (WHO) Classification of Gastric Cancer

The World Health Organization (WHO) classification issued in 2010

appears to be the most detailed among all pathological classification systems.

It is remarkable that the WHO classification includes not only

adenocarcinoma of the stomach but also all other types of gastric tumors of

lower frequency (Flejou JF, et al.,2011). The gastric adenocarcinoma type is

divided into several subgroups including papillary, tubular, mucinous and

mixed carcinoma, which can be compared to the indeterminate type in the

Lauren classification. The poorly cohesive carcinoma type includes the signet

ring cell carcinoma. All other classified gastric adenocarcinomas can be

designated as uncommon because of their minor clinical relevance. In the

WHO classification, the most common type of gastric cancer is the tubular

adenocarcinoma, followed by the papillary and mucinous types. The signet

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 Pathophysiology of Gastric Cancer

ring cell carcinoma accounts for approximately 10% of gastric cancers and is

defined by the presence of signet ring cells in over 50% of the tumor (Flejou J

F, et al., 2011; Werner M, et al., 2001). The prognosis of the signet ring cell

carcinoma is controversial. Most authors have described a worse prognosis

for the signet ring cell carcinoma compared to other subtypes of gastric

cancer (Ribeiro M M, et al., 1981, Hass H G, et al 2011). Recent studies

indicate that, on the contrary, signet ring cell carcinoma of the stomach does

not differ in prognosis from the other types of gastric cancer (Hass H G, et al.,

2012). Furthermore, signet ring cell carcinoma was shown to have an

irregular uptake of 18F-fluorodeoxyglucose during positron emission

tomography radionuclide imaging; consequently, this tumor as well as any

metastases cannot be detected reliably (Alakus H, et al., 2010). Patients with

a papillary adenocarcinoma experience a poor prognosis, a tendency for

metastatic disease, a higher age at diagnosis and location in the upper third

of the stomach (Yasuda K, et al., 2000). Another study that employed the

previous WHO classification found that poorly differentiated and mucinous

adenocarcinomas have a worse prognosis than the papillary and tubular

subtypes. In the same study, the WHO classification appeared to be an

independent prognostic factor (Zheng H C,et al 2010). Kawamura H, et al.,

(2001) also found a poor prognosis associated with mucinous

adenocarcinoma, which suggests a link with advanced stage and metastatic

disease. However, unlike most common types of gastric malignancies, the

WHO classification is more widely used for studies of infrequent types of

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 Pathophysiology of Gastric Cancer

gastric cancer. For adenosquamous carcinomas of the stomach, a poor

prognosis and a case of simultaneous gastric adenocarcinoma are described

(Toyota N, et al., 996; Faria G R, et al., 2010; Su J S, et al., 2013). Most of the

infrequent types of gastric malignancies are described in case reports, so a

systematic investigation of their prognoses is not readily available. As the

previous WHO classification was renewed in 2010, it is expected that more

gastric cancer studies that refer to the most recent WHO classification will be

conducted in the near future. An indication for the significance of the WHO

classification can be seen in a similar Japanese classification system. Although

the Japanese classification divides the common types of gastric

adenocarcinoma into additional subtypes, (e.g., tubular adenocarcinoma is

divided into well differentiated and moderately differentiated

adenocarcinoma), a dependence on the WHO classification system is evident

(Japanese Gastric Cancer Association, 2011). This particular subdivision of

tubular adenocarcinoma was based on differences in the submucosal

invasion rate, lymph node metastasis and size of the lesions (Fujii T, et al.,

1994).

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 Pathophysiology of Gastric Cancer

Lauren and World Health Organization classification

Intestinal type  Papillary adenocarcinoma

 Tubular adenocarcinoma
 Mucinous adenocarcinoma
Diffuse type  Signet-ring cell carcinoma and other poorly
cohesive carcinoma
Intermediate type  Mixed carcinoma
 Adenosquamous carcinoma
 Squamous cell carcinoma
 Hapaloid adenocarcinoma
 Carcinoma with lymphoid stroma
 Choriocarcinoma
 Carcinosacoma
 Parietal cell carcinoma
 Malignant rhabdoid tumor
 Mucoepidermoid carcinoma
 Paneth cell carcinoma
 Undifferentiated carcinoma
 Mixed adeno-neuroendocrine carcinoma
 Endodermal sinus tumor
 Embryonal carcinoma
 Pure gastric yolk sac tumor
 Oncocytic adenocarcinoma

Prognosis

The prognosis of gastric cancer depends on various pathological

factors, such as the macroscopic type, the depth of invasion, cancer-stromal

relationship, histological growth pattern, lymph node involvement, lymphatic

invasion, vascular invasion and tumour site with the main prognostic factors

being the TNM staging, along with the presence and extent of lymph node

metastases (Yokota T, et al., 2004). Diagnostic improvements and advances in

treatment options have improved the long-term survival of early gastric

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 Pathophysiology of Gastric Cancer

cancer patients. Prognosis is correlated with the tumour staging, worsening

as the degree of infiltration increases (Lello E, et al., 2007). The 5-year

survival rates in stages Ia, Ib, II, IIIa, and IIIb/IV are 91%, 64%, 27%, 18%,

and 0%, respectively with the survival rate of patients with early gastric

cancers invading but limited to the mucosa or submucosa being 90-100%,

compared to 60-80% for tumours reaching the muscularis propria, and 41-

50% for tumours limited to the subserosa or serosa (Antonioli D A, 1994;

Yoshikawa K, et al., 1985; Tanaka k, et al., 2004; Lello E, et al., 2007).

However, the prognosis of advanced gastric cancer remains poor, with

survival rates lower than 23% (Tanaka k, et al., 2004) and rarely exceeding

15% (Stewart B W, et al., 2003). The depth of infiltration correlates with the

presence of lymph node metastases, and the presence of regional lymph node

metastases reduces the 5-year survival rate of early gastric cancer patients

from 90% to 70% in tumours invading the submucosa (Antonioli D A, 1994;

Inoue K, et al., 1991).

The lymph node status and the ratio of metastasispositive/ metastasis-

negative lymph nodes are the strongest markers of gastric cancer prognosis

(Ichikura T, et al., 1999; Yokota T, et al., 2004), and the N-ratio

(metastatic/examined lymph nodes) has been validated as an independent

prognostic factor in a large multi-centre series, even where less than the

recommended 15 lymph nodes have been examined (Marchet A, et al., 2007).

The 5-year survival rate for patients with metastases in 1-6 lymph

nodes is 44%, and drops to 30% for 7-15 lymph node metastases, ending

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 Pathophysiology of Gastric Cancer

with 11% for more than 15 lymph nodes metastases. The N-ratio

classifications 0, 1, 2, and 3 exhibit 5-year survival rates of 83.4%, 66.3%,

468%, and 19.0%, respectively. Unfortunately, most patients presenting with

advanced gastric cancer already have lymph node metastases.

Other prognostic factors include lymphatic and vascular invasion, both

being associated with lower survival rates (Hamilton S R, et al., 2000; Yokota

T, et al., 2004), and the histological classification, whereby diffuse type

(Lauren classification) and mucous-rich (Goseki classification) tumours may

predict a worse prognosis (Martin I G, et al., 1994; Songun I, et al., 1999;

Hamilton S R, et al., 2000.)

Causes of Gastrointestinal carcinoma

Helicobacter pylori

Helicobacter pylori, a Gram-negative microaerophilic, spiral bacterium found

in the gastric mucosa in patients with severe gastritis and chronic atrophic

gastritis, has been recognized as an important risk factor for gastric cancer

(Catalano V, et al.,2009; Houghton J, et al., 2005). The results of several meta-

analyses concluded that H. pylori infection is associated with an

approximately two-fold increased risk of developing gastric cancer (Eslick G

D, 2006). In a prospective study involving 1526 Japanese patients who had

duodenal ulcers, gastric ulcers, gastric polyps or non-ulcer dyspepsia, 2.9% of

H. pylori infected patients subsequently developed gastric cancer while none

of the uninfected patients developed tumors (Uemura N, et al., 2001). In

1994, the International Agency for Research on Cancer categorized H. pylori

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 Pathophysiology of Gastric Cancer

as a “Group 1 human carcinogen” based on a plethora of studies

(Schistosomes. 1994).

Currently, approximately 50% of the world’s population is infected by

H. pylori. The prevalence of H. pylori infection varies markedly in different

countries in Asia with seroprevalence rates higher in developing countries

than in industrialized, developed nations (Fock K M, et al., 2010) The

identification of H. pylori as a risk factor for gastric carcinogenesis has

stimulated extensive research on the mechanisms by which H. pylori induces

carcinogenesis. A combination of a virulent organism, a permissive

environment, and a genetically susceptible host is considered essential for H.

pylori-induced gastric cancer. H. pylori has been suggested to trigger a

cascade of events that promote the sequential progression of normal gastric

epithelium through atrophic gastritis, intestinal metaplasia, and dysplasia to

carcinoma (Kim S S, et al., 2011; Amieva M R, et al., 2008; Hofman P, et al.,

2004) The bacterium secretes several products that cause gastric mucosal

damage such as urease, protease, phospholipase, ammonia, and

acetaldehyde. H. pylori disrupt gastric barrier function via urease-mediated

myosin-II activation (Wroblewski L E, et al., 2009).

Generation of oxidative stress is recognized as a virulence factor in H.

pylori-infected hosts. H. pylori infection induces the production of reactive

oxygen and nitrogen species and suppresses the host antioxidant defense

mechanisms, leading to oxidative DNA damage. However, H. pylori, which is

endowed with a variety of antioxidant enzymes is spared from oxidative

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 Pathophysiology of Gastric Cancer

stress and the damage is solely restricted to the gastric mucosa of the

susceptible host (Suzuki H, et al., 2012). H. pylori although not directly

mutagenic, has been suggested to favor the formation of mutagenic

substances through inflammatory mediators or by impairing the mismatch

repair pathway (Yao Y, et al., 2006). Kim s s, et al.,(2011) demonstrated that

H. pylori infection promotes gastric carcinogenesis by increasing endogenous

DNA damage whilst decreasing repair activities and by inducing mutations in

the mitochondrial and nuclear DNA. Aberrant DNA methylation induced by H.

pylori infection has been found to be a significant risk factor for gastric

cancer (Perrin D, et al., 2010). Epidemiological evidence suggests that H.

pylori strains containing the cag pathogenicity island (cagPAI) are more

virulent. The cagPAI is a 40-kb genome segment that encodes approximately

30 genes including the cytotoxin-associated gene A (cagA). The virulent cagA

positive strains increase the risk of non-cardia gastric cancer of both

intestinal and diffuse types, but not the risk of cardia cancer. The CagA

protein is delivered into gastric epithelial cells where it undergoes tyrosine

phosphorylation by SRC family kinases. Phosphorylated CagA specifically

binds to and activates SHP2, a phosphatase that transmits positive signals for

cell growth and motility. Thus H. pylori acting via cagA activates growth

factor receptors, increases proliferation, inhibits apoptosis, and promotes

invasion and angiogenesis (Hatakeyama M, et al., 2004)

Gene expression profiling of gastric antral mucosa samples from H.

pylori infected patients by microarray analysis followed by quantitative real-

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 Pathophysiology of Gastric Cancer

time PCR assays have revealed differential expression of 38 genes, indicating

that H. pylori infection leads to evasion of host defense, enhanced

inflammatory and immune responses, activation of NF-κB and Wnt/β-catenin

signaling pathways, perturbation of metal ion homeostasis, and induction of

carcinogenesis (Yang Z M, et al., 2012)

Epstein-Barr virus infection

The human herpes virus 4, or Epstein-Barr virus (EBV), is an

icosahedral herpes virus containing double stranded DNA that has been

connected with gastric cancer. The EBV has been classified as a Group I

carcinogen by the WHO and IARC, and is ubiquitous in all human populations.

EBV is the cause of Burkitt’s lymphoma, sino-nasal angiocentric T-cell

lympoma, Hodgkin’s disease and nasopharyngeal carcinoma (IARC, 1997).

EBV-associated carcinomas are found in all geographic regions (Stadtländer

et al., 1999), and are approximately three-fold more frequently found in

Japanese than in American populations (Watanabe et al, 1997). EBV is

associated with both intestinal- and diffusetype gastric cancers (Shibata D, et

al., 1992), but may be more prevalent in the male than in the female

(Tokunaga et al, 1993). The mechanism of EBV-mediated gastric

carcinogenesis is as yet unclear. Virus replication occurs in pharynx and

salivary gland epithelial cells. The subsequent infection of lymphoid B-cells is

mediated by the interaction of the gp350 viral envelope glycoprotein and

CD21, the C3d complement component CR2 (IARC, 1997). The viral

glycoproteins gp85, gp25 and gp42 are involved in host cell binding and viral

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 Pathophysiology of Gastric Cancer

envelope fusion,with the virus persisting in a latent state until triggering of

the host cell results in shedding of infectious virus particles (IARC, 1997). Up-

regulation of p53 is rarely observed in EBV-positive carcinomas, but found in

over 30% of EBV-negative carcinomas (Ojima H, et al., 1997; Chang M S, et al.,

2005) and p27 loss, p16 loss, cyclin D1 expression and NF-xB nuclear

positivity are found more frequently in EBV-positive gastric carcinomas

(Chang M S, et al, 2005). Despite the association of EBV infection with the

development of gastric carcinoma, there is no correlation with bcl-2

expression and p53 accumulation (Gulley et al, 1996), leading to the

conjecture that EBV induces gastric carcinomas via different mechanisms

than EBV-negative carcinomas (Ojima H, et al., 1997).

Dietary factors

A survey of literature on the role of diet in the pathogenesis of gastric

cancer using Pub Med as a search platform has revealed over 2000

epidemiological and experimental studies. Populations at high risk for

stomach cancer have been shown to consume diets rich in starch and poor in

protein quality, and are not inclined to eat fresh fruits and vegetables. Both

high starch and low protein diet may favor acid-catalyzed nitrosation in the

stomach and cause mechanical damage to the gastric mucosa (Krejs G J, et al.,

2010; Berretta M, et al., 2012; Tsugane S et al., 2007). Using an ecological

approach, Park B, et al., (2011) found a negative association between

refrigerator use, fruit intake, and gastric cancer mortality and positive

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 Pathophysiology of Gastric Cancer

associations between salt/ sodium intake and gastric cancer mortality and

incidence in Korea.

Both epidemiological and experimental studies strongly support the

role of excessive salt intake in gastric carcinogenesis. D’Elia L, et al.,(2012)

reported a direct correlation between dietary salt intake and risk of gastric

cancer with progressively increasing risk across consumption levels based on

a meta analysis of prospective studies. Consumption of large amount of salted

fish, soy sauce, pickled vegetables, cured meat and other salt-preserved foods

enhances H. pylori colonization, and increases the risk of gastric cancer

through direct damage to the gastric mucosa resulting in gastritis. Salt is also

known to induce hypergastrinemia and endogenous mutations, promoting

epithelial cell proliferation which eventually leads to parietal cell loss and

gastric cancer progression (Tsugane S, et al., 2004 Wang X Q, et al., 2009).

Reports from this laboratory as well as by other workers have demonstrated

that saturated sodium chloride (S-NaCl) promotes the development of N-

methyl-N’-nitro-N-nitrosoguanidine (MNNG)-induced rat gastric carcinomas

(Velmurugan B, et al.,2005; Kim D J, et al.,1997) Dietary nitrates are found

either naturally in foods such as cabbage, cauliflower, carrot, celery, radish,

beets, and spinach or added during preservation. In addition, the nitrate

content of fertilizers, soil, and water also contribute to dietary nitrate. Nitrite,

nitrate, and nitro sating agents can be synthesized endogenously by reactions

mediated by bacteria and activated macrophages. Nitrosation of a number of

naturally occurring guanidines and L-arginine-containing polypeptides

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 Pathophysiology of Gastric Cancer

produces mutagenic compounds. Dietary nitrate is converted to carcinogenic

N-nitroso compounds (NNC) by gastric acid thereby increasing gastric cancer

risk. Small quantities of preformed NNC and nitrosamines may also be

present in some foods including cured meats, dried milk, instant soups, and

coffee dried on direct flame (Suzuki H, et al., 2005; Mitacek E J, et al., 2008;

Liu C, et al., 2008)

In addition to specific components of the diet, certain cooking

practices are also associated with increased risk of gastric cancer. These

include boiling of meats, roasting, grilling, baking, and deep frying in open

furnaces, sun drying, salting, curing, and pickling, all of which increase the

formation of NNC. Polycyclic aromatic hydrocarbons such as benzo(a)pyrene

formed in smoked food have been incriminated in many areas of the world

with high stomach cancer rates (Jedrychowski W, et al., 2003; Wogan G N, et

al., 2004)

Lifestyle

Alcohol, a gastric irritant is an important risk factor for gastric cancer.

Zaridze D, et al., (2000) have reported an increased risk of stomach cancer in

men and women who regularly consume strong alcoholic beverages. A direct

correlation was observed between consumption of alcohol and tobacco and

the risk of gastric cancer in a population-based prospective cohort study

(Sjodahl K, et al., 2007). A study from this laboratory demonstrated a positive

correlation between alcohol consumption and cigarette smoking with the

blood lipid profile in gastric cancer patients (Manoharan S, et al., 1997). The

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 Pathophysiology of Gastric Cancer

European Prospective Investigation into Cancer and Nutrition (EPIC) project

found a significant association between the intensity and duration of

cigarette smoking and gastric cancer risk (Gonzalez C A, et at., 2003).

Smoking history was found to be a significant independent risk factor for

death from gastric cancer in patients who had undergone curative surgical

resection (Smyth E C, et al., 2012). Smoking is known to decrease

prostaglandins that maintain gastric mucosal integrity (Mc Cready D R, et al.,

1985). Tobacco smoke has been reported to induce the development of

precursor gastric lesions such as gastritis, ulceration, and intestinal

metaplasia. Smokers tend to have a higher incidence of H. pylori infection and

gastroduodenal inflammation than non-smokers (Gonzalez C A, et al., 2010).

Family history

Gastric cancer is a known manifestation of inherited cancer

predisposition syndromes similar to hereditary nonpolyposis colon cancer

and Li-Fraumeni syndrome. According to the OMIM database, 90% of gastric

cancers are sporadic, whereas 10% are hereditary. The first documented

report of familial predisposition to gastric cancer was described for Napoleon

Bonaparte’s family with Napoleon, his father, grandfather, brother, and three

sisters, all dying of stomach cancer at a relatively early age (Bevan S, et al.,

1999). The Scandinavian twin study in the Swedish, Danish, and Finnish twin

registries found an increased risk of stomach cancer in the twin of an affected

person (Lichtenstein P, et al., 2000). Family members usually share the same

environment and have similar socioeconomic status. These risk factors act

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 Pathophysiology of Gastric Cancer

independently or in conjunction with genetic factors thereby increasing the

risk of stomach cancer.

Occupations

A positive correlation has been recognized between increased stomach

cancer risk and a number of occupations including mining, farming, refining,

and fishing as well as in workers processing rubber, timber, and

asbestos(Krstev S, et al., 2005; Straif K, et al.,2000). Occupational exposure to

dusty and high temperature environments such as in cooks, wood processing

plant operators, food and related products machine operators was associated

with a significant increased risk of gastric cancer of the diffuse subtype

(Santibanez M, et al., 2012). A German uranium miner cohort study however

found a positive statistically non-significant relationship between stomach

cancer mortality and occupational exposure to arsenic dust, fine dust, and

absorbed dose from α and low-linear energy transfer radiation (Kreuzer M,et

al.,2012).

Blood Group-A

An association between gastric carcinomas and the blood group A has

been reported (Aird I, et al., 1953; Haenszel W, et al., 1976), which may be

related to the interaction between the Lewis blood group antigen and H.

pylori (Carneiro F, et al., 1996). The association of the blood group A with

males, with diffuse-type gastric cancer is stronger than with females, or

intestinal-type gastric cancer (Kramer B S, et al., 1995).

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 Pathophysiology of Gastric Cancer

Gene polymorphism

In recent years, genetic polymorphisms have come to be recognized as

crucial factors determining disease susceptibility. Host gene polymorphisms

frequently influence the magnitude of the host response, and this

interindividual variation contributes to the clinical outcome. The

development of gastric cancer in a milieau of chronic inflammation induced

by H. pylori may be significantly influenced by host gene polymorphisms.

The proinflammatory cytokine interleukin-1 (IL-1) gene cluster

containing IL-1β and IL-1RN encodes IL-1 β and the IL-1β receptor

antagonist, respectively, and the risk of gastric cancer and its precursor

lesions is increased in the presence of H. pylori by polymorphisms in these

genes. An increased risk of developing H. pylori-mediated hypochlorhydria

and gastric atrophy is associated with the IL-1B-31*C or -511*T, and the IL-

1RN*2/*2 genotypes (El-Omar E M, et al., 2000). These genotypes are also

associated with a 2-3 fold increase in the risk of developing gastric cancer,

compared to individuals with less proinflammatory genotypes (El-Omar E M,

et al., 2000, 2003; Figueiredo C, et al., 2002). Although a range of studies have

reported lower risks, recent meta-analyses have supported the findings of

the higher risk of the IL-1B-511*T, and the IL-1RN*2 genotypes, particularly

in association with ethnic group and tumour type (Camargo M C, et al.,

2006;Kamangar F, et al., 2006; Wang P, et al., 2007).

Although IL-1 is one of the most important proinflammatory cytokines

mediating the effects of H. pylori infection (El-Omar E M, et al., 2000),

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 Pathophysiology of Gastric Cancer

polymorphisms in other proinflammatory cytokines, such as TNFα (308*A)

and IL-10 (ATA/ATA), have also been associated with increased risk for

gastric cancer (El-Omar E M, et al., 2003). The more proinflammatory

genotypes an individual has, the higher the risk of developing gastric cancer

(Figueiredo C et al., 2002). Gastric cancer and H. pylori infection has been

linked with the human leukocyte antigen (Magnusson et al, 2001), with the

*1601 allele significantly increasing the risk of gastric cancer (odds ratio 8.7;

95% CI 2.7-28). This association is seemingly independent of H. pylori

infection, being stronger in H. pylori-negative patients and in diffuse-type

carcinomas (Correa P, 2002).

The MUC-1 mucin is a glycoprotein involved in the protection and

lubrication of epithelial surfaces, detecting potential external insults and

interacting with signal transduction and cell adhesion proteins (Gendler S J,

2001).MUC-1 contains a variable number of tandem repeats, with higher

numbers of repeats encoding larger proteins better able to respond to

external stimuli. Gastric cancer patients exhibit higher proportions of smaller

MUC-1 proteins, with smaller alleles linked to gastric atrophy and intestinal

metaplasia (Carvalho F, et al., 1999; Correa P, 2002).

Diagnosis of Gastrointestinal Cancer

The diagnosis of cancer involves the analysis of tissue and cytology

specimens obtained through several procedures, including surgical biopsy,

core or aspirational needle biopsy, venipuncture, pleural or ascitic tap,

scraping of tissue surfaces and collection of exfoliative cells from urine and

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 Pathophysiology of Gastric Cancer

sputum. Conventional histopathology based on assessing morphology has

remained the standard diagnostic method for many years but development of

advanced sophisticated technologies like mass spectrometry microarray and

automated DNA sequencing have opened new avenues in cancer diagnosis

and therapeutics.

1. Clinical Symptoms

Clinical symptoms of cancer vary according to the type and nature of

the cancer and its location in different organs. These include gastrointestinal

obstruction which may be accompanied by bleeding which is presented as

diarrhea and vomiting (commonly associated with tumors invading the

stomach, small intestine, large intestine, or colon), hematuria (in tumors of

the kidney or bladder), Cushing's disease, hypoglycemia, etc. (in hormone-

producing tumors such as some pancreatic, thymic and hepatic tumors),

hematological disturbances (Staszewski H, et al., 997) as anemia,

polycythemia, granulocytosis etc. and neurologic symptoms (Darnell R B, et

al., 2006; Rees J H, et al.,2004) such as loss of coordination or seizures (in

tumors of the brain or spinal cord). Cancers producing non-specific

symptoms are extremely difficult to be diagnosed for their location, referred

to as paraneoplastic disorders. These include weight loss, low-grade fever,

seizures, lethargy, loss of appetite, diarrhea, skin rash, hair loss, and general

arthritic-like symptoms. These types of cancers require specialized

diagnostic techniques such as laboratory screening tests, X-rays, CT scan, MRI

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 Pathophysiology of Gastric Cancer

etc. which can provide a means for earlier diagnosis and perhaps better long-

term prognosis.

2. Imaging

A diagnosis of malignancy is frequently suspected based on imaging

information, later confirmed on histology. Until now, exploratory surgery or

limited radiologic evaluations are most commonly used techniques for cancer

diagnosis and staging. With the advent of computed tomography (CT) and

magnetic resonance imaging (MRI), it became possible to obtain important

structural and anatomic information. Molecular imaging with magnetic

resonance spectroscopy (MRS) and positron emission tomography (PET) is

currently possible in clinical practice. These modalities permit functional,

biochemical and physiologic assessment of important aspects of malignancy.

Sites in which imaging plays a key role for the diagnosis include brain, breast,

lung and mediastinum, the tumors arising from the abdominal organs, retro

peritoneum and bones. Conventional radiography provides the easiest way to

diagnose the tumors of gastro intestinal tract, lungs, brain, liver, urinary

bladder, breast, bone, joints etc in the pet and domestic animals (Cole R, et al.,

2007; Drost, W T, et al., 1996; Russo M, et al.,1999; Steinberg H, et al., 2006;

Wuersch K, et al., 2009)

3. Ultrasound

Ultrasonography uses high frequency broadband sound waves in the

megahertz range that are reflected by tissue to varying degrees to produce

images (up to 3D). It used in the diagnosis of cancers of abdominal organs,

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 Pathophysiology of Gastric Cancer

heart, breast, muscles, tendons, arteries and veins. It is useful in aiding the

characterization of lesions (shape, size, and density) found on screening

mammograms in women with dense breasts (Kolb T M, 2000.) and in bitches

having mammary tumors (Gonzalez de Bulnes A, et al., 1998). While it may

provide less anatomical detail than techniques such as CT or MRI, it has

several advantages which make it ideal in numerous situations, in particular

that it studies the function of moving structures in real-time, emits no

ionizing radiation, and contains speckle that can be used in elastography.

Ultrasound has also been used for the diagnosis of the tumourous conditions

of the abdominal organs of domestic and wild animals (Ferreira V L, et al.,

2010). Tumours of the hollow organs such as urinary bladder can be

diagnosed easily by the ultrasonographic technique (Hoque M, et al., 2002).

Singh C, et al., (2009) used ultrasound guided biopsies (USGB) and

ultrasound guided fine needle aspiration biopsies (USGFNAB) to diagnose the

hepatic affections in dogs and benefits of these techniques can be utilized for

the diagnosis of tumours of other internal organs. High resolution ultrasound

can be used to evaluate the tumour volume accurately in the murine

orthotopic tumour models without sacrifice (Kraaij R, et al., 2002; Pezold J C,

et al., 2006).

4. Computed Tomography

Recent innovations include spiral (helical) CT, multiphase imaging and

multi detector scanning. Potential patient benefits include rapid data

acquisition and improved detection and characterization of lesions. Spiral CT

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 Pathophysiology of Gastric Cancer

currently is the preferred technique for detecting cancerous lesions in

pulmonary organs and liver prior to metastasectomy and for surgical

planning of pancreatic and renal cancer treatment. Cases of nasal tumours in

dogs (Kondo Y, et al., 2008) and horses (Veraa S, et al., 2008) can be

diagnosed and evaluated on the basis of the CT scan New roles for spiral CT

include the detection of pulmonary emboli, CT angiography and endoscopic

viewing of hollow organs. Dogs and cats are prone to brain and spine

tumours which are life threatening and need to be diagnosed as early as

possible before they attain incurable stage. Brain tumours includes the

tumours of pituitary, cerebrum, cerebellum, hypothalamus etc. and tumours

of spine divide as extradural, intradural - extramedullary, or intramedullary,

among them 50% are extradural, whereas 35% and 15% are

intraduralextramedullary and intramedullary, respectively (Narama I, et al.,

1992). So it is certain that CT scan can play an important role in the diagnosis

of tumours of brain and spine in dogs and cats (Bast R C, et al., 2003;

Iwamoto K S et al., 1993; Mauldin G N, et al., 1990; Pollard R E, et al., 2010).

5. Magnetic Resonance Imaging

In this technique powerful magnets are used to polarize and excite

hydrogen nuclei in water molecules in tissue, producing a detectable signal

which is spatially encoded, resulting in images of the body. MRI has a number

of imaging benefits including superb soft tissue contrast, multiplanar and 3D

image acquisition, freedom from ionizing radiation and bony artifacts, and

ability to acquire biological and physiological information. Recent advances

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 Pathophysiology of Gastric Cancer

with the use of supercoils have resulted in an increase in sensitivity and

specificity (Kaiser W A, et al., 1992). Contrast agents used in it are chelates of

gadolinium, a lanthamide with three unpaired electrons, which has a very

strong magnetic field. MRI is the imaging technique of choice for evaluating

tumors in brain, head and neck, spine, breast (when mammography is

technically difficult owing to dense breast, silicone implants and scarring due

to surgery/ trauma), liver and adrenal glands. Recent advances include

increased speed of data acquisition and the ability to visualize function

superimposed on anatomical changes. Breast MRI has been shown to be

capable of detecting early breast cancer, with sensitivities in the range of 95–

100%, i.e. a low falsenegative rate (Harms S, et al., 1999; Orel S G, et al.,

1994). In veterinary field MRI has been used frequently in detecting

macrotumours of brain and spine in dogs (Duesberg C A, et al., 1995;

Kippenes H, et al., 1999). Dynamic contrast-enhanced magnetic resonance

imaging (DCE-MRI) is a newer technique by which one can evaluate the

kineticparameters such as blood flow, perfusion, vascular permeability, and

the fraction of interstitial space within a tumor. These parameters derived

from DCE-MRI present information which are appropriate to noninvasively

differentiate canine brain tumors (Zhao Q, et al., 2010).

6. Metabolic and Functional Imaging

Functional imaging is a recent tool used in oncology and its uses

includes characterization of indeterminate lesions on conventional imaging,

cancer staging and monitoring response to treatment.

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 Pathophysiology of Gastric Cancer

7. Positron emission tomography (PET)

PET can detect biologic changes in vivo using radiolabeled tracers. It

represents the metabolic activity of underlying tissue processes such as

glucose, oxygen and amino acid metabolism or measures receptor density

status. Fluorine-18 fluorodeoxyglucose (18FFDG) is used most commonly,

and closely mimics endogenous molecules (Grahek D, et al., 2004; Kaiser W A

et al., 1992). FDG enters cells and is phosphorylated to FDG-6-phosphate,

which becomes trapped within malignant tumor cells with high glucose

metabolism. PET is the most accurate non-invasive technique for detecting

and staging lung cancer. It is superior to CT arterial portography in detecting

intrahepatic metastases in colorectal cancers and can identify metastatic

deposits in lymph nodes that are still <1 cm in size and considered benign by

CT. In contrast, PET may recognize large masses, such as post therapy fibrotic

tissue, as benign if minimal FDG uptake is demonstrated. Limitation with PET

to tumor detection is that increased FDG uptake can also be demonstrated in

inflammatory tissue.

8. Magnetic resonance spectroscopy

MRS is a non-invasive method for studying tumor biochemistry and

physiology. It measures signals from chemical compounds within tissues;

P31 MRS provides information on tissue energetic and pH while H1- MRS

conveys information on cell membrane synthesis and degradation, reflecting

cellular proliferation and necrosis. MRS resonances can provide diagnostic

information on tumor grade and are used to monitor tumor response to

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 Pathophysiology of Gastric Cancer

therapy. In a review by Katz-Brull R, et al., (2002) in a studies of five breast

H-MRS performed in four independent centers around the world to date, the

combined analysis of the data from a total of 153 tumors demonstrated

sensitivity and specificity as high as 92% in distinguishing malignant from

benign tumors using the choline signal. Of special interest, in a subgroup of

20 younger women the sensitivity and specificity of the method approached

100% (Katz-Brull R, et al., 2002).

9. Cytologic and Histopathological Technique

Histopathology is still a gold standard for diagnosis of tumours but it

alone does not provide sufficient details of the cellular changes which could

predict the clinical behaviour of the tumour. Even then histopathological

examination of the tumour cells by any expert oncologist can give an accurate

diagnosis about the type of tumour and possible malignancy status. Serous

effusions from pleural, peritoneal or pelvic cavity can act as biopsy material

for diagnosis of tumours of those regions as these may consist the cancerous

cells. Oyafuso M S, et al., (1996) compare the results of cytological diagnosis

of tumours in Brazil, and found that sensitivity, specificity, efficiency, as well

as positive and negative predictive values for cytologic diagnosis were

44.5%, 95.7%, 50.1%, 98.7% and 20%, respectively. Presence of

hyperchromatic nuclei, more nucleus to cytoplasm ratio, disorientation of the

cells, etc. are the cellular changes observed in cancer cell. Presences of

mitotic figures in the cells are also related with neoplastic change in the

tissues. During studies on the canine mammary and skin tumours it was

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 Pathophysiology of Gastric Cancer

found that number of mitotic figures in the malignant tumours was

significantly higher than the benign tumours (Pawan K, et al., 2010; Reddy G,

et al., 2009). Special staining procedures can differentiate the different types

of tumours and thus help in diagnosis such as toluidine blue stain

differentiate mast cell tumour from other tumours as it stains the

metachromatic granules present in mast cells.

10. Serological Methods

Serological methods used in estimation of serum tumor markers are

ELISA and RIA (Schrohl A S, et al., 2003, Wu J T, et al., 2007). The ELISA is

typically used to detect and quantify antigen within biological fluids, in which

the Dual-Antibody Sandwich ELISA is being used for measuring the

concentration of 80% of tumor markers in blood or serum. RIA is one of the

most sensitive technique for detecting antigen or antibody. The principle

involves competitive binding of radiolabelled antigen and unlabelled antigen

to a high-affinity antibody. Gamma emitting isotope such as Iodine and beta

emitting isotope such as tritium are also routinely used as labels. The

important step in the RIA is the determination of the amount of antibody

needed to bind 60% - 70% of a fixed quantity of radioactive antigen.

Determination of amount of bound labeled antigen can be done by

precipitating the Ag- Ab complex to separate it from free antigen and the

radio activity in the precipitate can be measured (Chan D W, et al., 1997). The

presence of CEA, AFP, PSA and other markers in the serum of the cancer

patients can be detected with the help of ELFA (Enzyme linked florescent

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 Pathophysiology of Gastric Cancer

assay).The test measures the amount of CEA that may appear in the blood of

some people who have certain kinds of cancers, especially large intestine

(colon and rectal) and breast cancer. It may also be present in people with

cancer of the pancreas, ovary, or lung (Blast R C, 2003; Thriveni K, et al.,

2007).

11. Fluorescence in Situ Hybridization (FISH) Technique

This technique involved the specific hybridization of a labeled nucleic

acid probe to complementary gene sequence and subsequent visualization by

autoradiographic or immunocytochemical method in tissue section, smears

or cytocentrifuged cell suspensions. Chromosome abnormalities are

frequently found in malignant cells. Chromosome rearrangements can be

duplications (addition of chromosome), deletions (loss of whole or parts of

chromosomes), segmental amplifications (random reiteration of segments or

extra fragments), translocations (exchange between chromosomes) and

inversions (reversal of orientation). It is applicable to interphase cells and is

more sensitive compared to conventional cytogenetic. Comparative genomic

hybridization (CGH) is a newly described method developed in 1992 and

used globally for studies of chromosomal gains and losses in genomic

complement. In CGH, test and reference genomic DNA are first differentially

labeled with different fluorescent dyes and co hybridized to normal

metaphase chromosomes. Then, fluorescent signals along each chromosome

are examined and analyzed to provide a cytogenetic pattern of gains and

losses. Several investigators have found this method to be useful in cancer

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 Pathophysiology of Gastric Cancer

studies, suggesting that different tumor types or different stages of tumor

progression have distinct CGH patterns (Forozan F, et al., 1997). These

quantitative changes are related to modification in expression level of genes

located in the target region. They found a substantial degree of correlation

between the two levels of information. To increase resolution, several groups

have adapted array technology to CGH, leading to socalled array-CGH

(Pollack J R, et al., 1999). Array-CGH has now been established as a new

method for molecular characterization of cancers. Moreover, it can serve as a

starting point for further screening investigations, such as genome-wide gene

expression profiling.

12. Polymerase Chain Reaction (PCR)

Molecular oncology studies the alterations in genetic and biochemical

processes at the molecular level. It helps in establishing a definitive diagnosis

and classification of tumors based on the recognition of complex profiles

('finger-prints') or unique molecular alteration that occur in specific tumor

types. The changes can be studied on chromosomes, DNA or RNA.

Microsatellite markers, also known as simple sequence repeats or SSRs (Litt

M, et al., 1989) are scattered widely within the biological genomes and

closely linked with many important genes. In carcinogenesis, microsatellites

often display loss of heterozygosity (LOH) as tumour suppressor genes.

These are highly polymorphic repetitive DNA sequences that are randomly

distributed throughout eukaryotic genomes displaying high levels of

variation and are having high mutation rates (1-4 per generation) (Weber J et

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 Pathophysiology of Gastric Cancer

al., 1993). The PCR process was originally developed to amplify short

segments of a longer DNA molecule (Robertson L E, et al., 1992). PCR allows

early diagnosis of malignant diseases such as leukemia and lymphomas,

which is currently the highest developed in cancer research and is already

being used routinely. PCR assays can be performed directly on genomic DNA

samples to detect translocation-specific malignant cells at a sensitivity which

is at least 10,000 fold higher than other methods. Quantitative PCR methods

allow the estimation of the amount of a given sequence present in a sample a

technique often applied to quantitatively determine levels of gene

expression. Real-time PCR is an established tool for DNA quantification that

measures the accumulation of DNA product after each round of PCR

amplification. Mackay et al. (Martins A, et al., 2010) opined that Real-time

quantitative PCR is a very powerful and accurate technique to examine

expression patterns of different oncogene, suppressor genes in different

cancerous conditions. Real-time PCR has engendered wider acceptance of the

PCR due to its improved rapidity, sensitivity, reproducibility and the reduced

risk of carry-over contamination. There are currently five main chemistries

used for the detection of PCR product during real-time PCR. PCR act as an

important tool for the diagnosis of the virus induced tumours of the animals

such as cutaneous papillomatosis in cattle, urinary bladder tumours of cattle

(De Villiers E M, et al., 2004; Pawaiya R, et al., 2005) and buffaloes (Sylvestre

O, et al., 2009), equine sarcoids (Nasir L, et al., 2008), papillomatosis in dogs

(Goldschmidt M N, et al., 1998; Narama I, et al., 1992) etc.

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 Pathophysiology of Gastric Cancer

13. Microarray

Microarray has emerged as a powerful tool to increase the potential of

standard methods through genome wide biology studies. DNA microarray

technology is a promising approach that allows both qualitative and

quantitative screening for sequence variations in the genomic DNA of cancer

cells. DNA microarray-based sequence analysis uses comparative

hybridization to obtain information ranging from mutational detection to

polymorphism genotyping. Sequencing by hybridization is conceptually

based on the construction of unknown sequences from hybridization data

(Wallraff G, et al., 1997). Labeled DNA for analysis binds strongly only to

those targets that are fully complementary to one of its subsequences.

Specific binding profile is further checked by a computational algorithm to

deduce the whole original sequence.

14. Mutational Analysis

Detection of mutations in cancer is of major importance for both basic

understanding of the disease process and clinical practice. High-density

oligonucleotide arrays are commonly used to achieve this purpose. Many

early applications of this method concerned breast cancer–associated genes

BRCA1 and BRCA2 (Favis R et al., 2000; Hacia J G, 1996, 2000). From this

initial success, one can easily predict the impact of specific “mutation arrays,”

which test for a variety of known mutations in numerous oncogenes, tumor

suppressors, and other genes shown to be of interest in cancer.

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 Pathophysiology of Gastric Cancer

15. Polymorphism Genotyping

Microarray is an appropriate tool to understand how sequence

polymorphism may impact biologic functions and be associated with

heritable phenotypes. Single nucleotide polymorphisms (SNPs) are the most

abundant form of DNA polymorphisms. SNP microarray is an oligoarray in

which SNPs are screened by a set of oligonucleotide probes. In a first

approach, different oligonucleotides can be used to identify several thousand

SNPs and then specific oligonucleotides can be used to genotype these SNPs

in various samples (Sapolsky R J, et al., 1999). SNP microarrays have

potential applications in loss of hetero zygosity (LOH) analysis, in disease

susceptibility and pharmaco- and toxicogenetic studies.

Characteristics of Free radicals and Oxidants

Free radicals are defined as an atoms or molecules containing one or

more unpaired electrons which are highly unstable and reactive substances.

The most important ROS are the superoxide anion radical O 2, hydrogen

peroxide (H2O2), alkoxyl (RO.), peroxyl (ROO.), hydroxyl radicals (.OH), and

hypochlorous acid (HOCl).Other non-oxygen species exists as reactive

nitrogen species(RNS), such as nitric oxide (NO.) and peroxynitrite having

important bioactivity. ROS is continuously generated in physiological

conditions but are effectively eliminated by intracellular and extracellular

antioxidant systems (Halliwell B, et al., 1999).

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 Pathophysiology of Gastric Cancer

Sources of Reactive Oxygen Species

ROS are produced from molecular oxygen as a result of normal cellular

metabolism. ROS can be divided into 2 groups: free radicals and nonradicals.

Molecules containing one or more unpaired electrons and thus giving

reactivity to the molecule are called free radicals. When 2 free radicals share

their unpaired electrons, non-radical forms are created. The 3 major ROS that

are of physiological significance are superoxide anion (O2-), hydroxyl radical

(OH.), and hydrogen peroxide (H2O2).

Endogenous and exogenous sources of reactive oxygen species (ROS)

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 Pathophysiology of Gastric Cancer

Major Reactive Oxygen Species (ROS)

Radical Non radical

Superoxide O2* Hydrogen peroxide H2O2

Hydroxyl radical OH* Ozone O3
Organic radical R* Organic ROOH
hydrogen peroxide
Peroxyl radical Roo* Hypochlorous acid HOCl
Alkoxyl radical Ro* Singlet Oxygen 1O2

Thiyl radical RS*

Sulphonyl radical ROS*
Thiyl peroxyl radical RSOO*

Superoxide anion radical and hydroxyl radical

Superoxide anion is formed by the addition of one electron to the

molecular oxygen. (Miller D M, et al., 1990) This process is mediated by

nicotine adenine dinucleotide phosphate [NAD(P)H] oxidase or xanthine

oxidase or by mitochondrial electron transport system. The major site for

producing superoxide anion is the mitochondria, the machinery of the cell to

produce adenosine triphosphate. Normally, electrons are transferred through

mitochondrial electron transport chain for reduction of oxygen to water, but

approximately 1 to 3% of all electrons leak from the system and produce

superoxide. NAD(P)H oxidase is found in polymorphonuclear leukocytes,

monocytes, and macrophages. Upon phagocytosis, these cells produce a burst

of superoxide that lead to bactericidal activity. Superoxide is converted into

hydrogen peroxide by the action of superoxide dismutases (SODs, EC

1.15.1.1). Hydrogen peroxide easily diffuses across the plasma membrane.

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 Pathophysiology of Gastric Cancer

Hydrogen peroxide is also produced by xanthine oxidase, amino acid oxidase,

and NAD(P)H oxidase (Dupuy C, et al., 1991; Granger D N, et al.,1991 ) and in

peroxisomes by consumption of molecular oxygen in metabolic reactions.

In a succession of reactions called Haber–Weiss and Fenton reactions,

H2O2 can breakdown to OH- in the presence of transmission metals like Fe2+

or Cu2+. (Fenton H, et al., 1984)

Fe3++ .O2 → Fe2+ + O2 Haber-Weiss

Fe3++ H2O2 → Fe3++OH ¯ + .OH Fenton reaction

O2- itself can also react with H2O2 and generate OH ¯ (Haber F, et al.,

1934; Liochev S I, et al., 2002). Hydroxyl radical is the most reactive of ROS

and can damage proteins, lipids, and carbohydrates and DNA. It can also start

lipid peroxidation by taking an electron from polyunsaturated fatty acids.

Hydrogen peroxide

Hydrogen peroxide can be generated through a dismutation reaction

from superoxide anion by superoxide dismutase. Enzymes such as amino

acid oxidase and xanthine oxidase also produce hydrogen peroxide from

superoxide anion. Hydrogen peroxide is highly diffusible and crosses the

plasma membrane easily.Hydrogen peroxide is the least reactive molecule

among reactive oxygen species and is stable under physiological pH and

temperature in the absence of metal ions. Hydrogen peroxide is a weak

oxidizing and reducing agent and is thus regarded as being poorly reactive.

Hydrogen peroxide can generate the hydroxyl radical in the presence of

metal ions and superoxide anion (·O2– + H2O2 → ·OH + OH– + O2) (Halliwell B,

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 Pathophysiology of Gastric Cancer

1997). Hydrogen peroxide can produce singlet oxygen through reaction with

superoxide anion or with HOCl or chloroamines in living systems. Hydrogen

peroxide can degrade certain heme proteins, such as hemoglobin, to release

iron ions.

Figure: Sources, generation of reactive oxygen species and the defense mechanisms

against damage by active oxygen species in human disease. (Jose et al, 1999).

Hypochlorous acid

Granulocytic enzymes further expand the reactivity of H2O2 via eosinophil

peroxidase and myeloperoxidase (MPO). In the presence of chloride ion, H2O2

is converted to hypochlorous acid (HOCl). HOCl is highly oxidative and plays

an important role in killing of the pathogens in the airways. (Klebanoff S J, et

al., 2005) However, HOCl can also react with DNA and induce DNA–protein

interactions and produce pyrimidine oxidation products and add chloride to

DNA bases. (Whiteman M, et al., 1997; Kulcharyk P A, et al., 2001) Eosinophil

peroxidase and MPO also contribute to the oxidative stress by modification of

proteins by halogenations, nitration, and protein cross-links via tyrosyl

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 Pathophysiology of Gastric Cancer

radicals. ( Brennan M L, et al., 2001; Denzler K L, et al., 2001; van Dalen C J, et

al., 2000)

Peroxyl radicals

Other oxygen-derived free radicals are the peroxyl radicals (ROO.).

Simplest form of these radicals is hydro-peroxyl radical (HOO.) and has a role

in fatty acid peroxidation. Free radicals can trigger lipid peroxidation chain

reactions by abstracting a hydrogen atom from a side chain methylene

carbon. The lipid radical then reacts with oxygen to produce peroxyl radical.

Peroxyl radical initiates a chain reaction and transforms polyunsaturated

fatty acids into lipid hydroperoxides. Lipid hydroperoxides are very unstable

and easily decompose to secondary products, such as aldehydes (such as 4-

hydroxy-2,3-nonenal) and malondialdehydes (MDAs). Isoprostanes are

another group of lipid peroxidation products that are generated via the

peroxidation of arachidonic acid and have also been found to be elevated in

plasma and breath condensates of asthmatics. (Wood L G, et al., 2000;

Montuschi P, et al., 1999) Peroxidation of lipids disturbs the integrity of cell

membranes and leads to rearrangement of membrane structure.

Antioxidants

An Antioxidant is a molecule capable of slowing or preventing the

oxidation of other molecules. Oxidation reactions can produce free radicals,

which start chain reactions that damage cells. Antioxidants (enzymatic &

non- enzymatic) terminate these chain reactions by removing radical

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 Pathophysiology of Gastric Cancer

intermediates and inhibiting other oxidation reactions by being oxidized

themselves. So, antioxidants are often reducing agents such as thiols or

polyphenols (Duarte T L, et al.,2005 ) Although oxidation reactions are

crucial for life, they can also be damaging plants and animals contain various

antioxidants, such as glutathione, vitamin C, and vitamin E as well as enzymes

such as catalase, superoxide dismutase and peroxidases. Low levels of

antioxidants or inhibition of the antioxidant enzymes causes oxidative stress

and may damage or kill cells (Valko M, et al., 2007)

The antioxidant defense systems function through blocking the initial

production of free radicals, scavengering the oxidants, converting the

oxidants to less toxic compounds, blocking the secondary production of toxic

metabolites or inflammatory mediators, blocking the chain propagation of

the secondary oxidants, repairing the molecular injury induced by free

radicals or enhancing the endogenous antioxidant defense system of the

target. These defense mechanisms act cooperatively to protect the body from

oxidative stress. The antioxidant defense system consists of powerful

enzymatic and non-enzymatic antioxidants (Halliwell B. 2007)

Enzymatic and Non-enzymatic antioxidants

Enzymatic Acronym Non-enzymatic Acronym
Superoxide SOD Glutathione GSH
dismutase
Catalase CAT Glutaredoxin GRX
Glutathione GPx Thioreoxin TRX
peroxidase
Glutathione-S- GST Peroxiredoxin PRX
transferase

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 Pathophysiology of Gastric Cancer

Glutathione GR Sulfiredoxin SRX

reductase
Peroxiredoxin APX Phytochemicals
Thioreoxin TRXR Vitamin A,C,E
reductase
Ceruloplsmin

Enzymatic Antioxidants

All cells in the body contain powerful antioxidant enzymes. The three major

classes of antioxidant enzymes are the superoxide dismutases, catalases and

glutathione (GSH) peroxidases. In addition, there are numerous specialized

antioxidant enzymes reacting with and detoxifying oxidants (Valko M, et al.,

2007)

Superoxide dismutases (SOD)

They are a class of closely related enzymes that catalyse the

breakdown of the superoxide anion into oxygen and hydrogen peroxide

(Zelko I, et al., 2002). They are present in almost all aerobic cells and in the

extracellular fluids. They contain metal ions that can be copper, zinc,

manganese or iron. In humans, the copper/zinc superoxide dismutase is

present in the cytosol, while manganese superoxide dismutase is present in

the mitochondria. There also exists a third form of superoxide dismutase in

extracellular fluids, which contains copper and zinc in its active sites

(Johnson F, 2005). Superoxide dismutase removes O2. – by catalyzing a

dismutation reaction. In the absence of superoxide dismutase, this reaction

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 Pathophysiology of Gastric Cancer

occurs non-enzymatically but at a very slow rate (Nozik-Grayck E, et al.,

2005)

Catalase

Catalase (H2O2 oxidoreductase) is a tetramer of four polypeptide

chains, each over 500 amino acids long, contains four porphyrin heme (iron)

groups that allow the enzyme to react with the hydrogen peroxide. Catalase

can decompose hydrogen peroxide (H2O2) in reactions catalyzed by two

different modes of enzymatic activity: the catalatic mode of activity (2 H 2O2 →

O2 + 2 H2O) and the peroxidatic mode of activity (H2O2 + A H2 → A + 2 H2O).

Catalase has one of the highest turnover rates of all enzymes; one molecule of

catalase can convert millions of molecules of hydrogen peroxide to water and

oxygen per second. Decomposition of H2O2 by the catalytic activity of catalase

follows the fashion of a first-order reaction and its rate is dependent on the

concentration of H2O2 (Valko M, et al., 2007; Berg J M, et al., 2002).

Catalase is an unusual enzyme since, although hydrogen peroxide is its

only substrate, it follows a ping-pong mechanism. Here, its cofactor is

oxidised by one molecule of hydrogen peroxide and then regenerated by

transferring the bound oxygen to a second molecule of substrate (Kabel A M,

et al., 2013)

Catalase is present in all prokaryotes and eukaryotes. With the

exception of erythrocytes, it is predominantly located in peroxisomes of all

types of mammalian cells where H2O2 is generated by various oxidases. Since

H2O2 serves as a substrate for certain reaction that generate the highly

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 Pathophysiology of Gastric Cancer

reactive hydroxyl radical, catalase is believed to play a role in cellular

antioxidant defense mechanisms by limiting the accumulation of H2O2 (Ho Y

S, et al., 2004)

The role of catalase in defending cells and tissues against oxidative

stress has been studied extensively. Over expression of catalase renders cells

more resistant to toxicity of H2O2 and oxidant-mediated injury. In addition,

transgenic mice overexpressing catalase are protected against myocardial

injury following administration of adriamycin and development of

hypertension from treatment with norepinephrine or angiotensin. Catalase-

deficient patients are phenotypically normal with the exception of an

increased tendency to development of progressive oral gangrene as a result

of tissue damage from H2O2 produced by peroxide-generating bacteria such

as streptococci and pneumococci as well as by the phagocytic cells at the sites

of bacterial infection (Yang H, et al., 2003)

Glutathione

The glutathione system includes glutathione, glutathione reductase

(GR), glutathione peroxidases (GPx) and glutathione S-transferases.

Glutathione peroxidase is an enzyme that catalyzes the breakdown of

hydrogen peroxide and organic hydroperoxides. Glutathione S-transferases

are another class of glutathione-dependent antioxidant enzymes that show

high activity with lipid peroxides (Sharma R, et al., 2004) These enzymes are

at high levels in the liver and also help in detoxification metabolism (Hayes J,

et al., 2005).

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 Pathophysiology of Gastric Cancer

Glutathione reductase (GR) is a crucial enzyme that reduces

glutathione disulfide (GSSG) to the sulfhydryl form (GSH) by the NADPH-

dependent mechanism, an important cellular antioxidant system. Due to its

significance, the enzyme has been purified from a number of animals, plants

and microbial sources and studied in an effort to identify and explain its

structure, kinetic mechanism and molecular properties (Linster C L, et al.,

2007). Its kinetic mechanism is known to be a ping-pong/sequential ordered

model. GR is a flavoprotein that contains two FAD molecules as a prosthetic

group, which is reducible by NADPH. GR is one of the thermostable enzymes.

GR belongs to the defense system protecting the organism against chemical

and oxidative stress. Deficiency of GR is characterized by hemolysis due to

increased sensitivity of erythrocyte membranes to H2O2 and contributes to

oxidative stress which plays a key role in the pathogenesis of many diseases

(Ulusu N N, et al., 2007)

Non- Enzymatic antioxidants-

Ascorbic acid

Ascorbic acid or vitamin C is a monosaccharide antioxidant found in

both animals and plants but cannot be synthesised in humans and must be

obtained from the diet. In cells, it is maintained in its reduced form by

reaction with glutathione. Ascorbic acid is a reducing agent that can reduce

and thereby neutralize reactive oxygen species such as hydrogen peroxide

(Linster C L, et al., 2007).

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 Pathophysiology of Gastric Cancer

Vitamin E

Vitamin E (α-tocopherol) is the most important lipid-soluble

antioxidant and protects cell membranes against oxidation by reacting with

the lipid radicals produced in the lipid peroxidation chain reaction and

removing the free radical intermediates. Tocotrienols may have a specialised

role in neuroprotection (Sen C, et al., 2006)

Carotenoids

Carotenoids are compounds with lipophilic properties that have

antioxidant functions in lipid phases. Beta-carotene besides being a

precursor to vitamin A has potent antioxidant properties as it removes

singlet oxygen thus protects against free radical attack. They are present in

liver, egg yolk, milk, butter, spinach, carrots, tomato and grains (Linster C L,

et al., 2007).

Fig. - Mutual association between oxidants and antioxidants.

(NOS – NO-synthase, ROS – reactive oxygen species, CAT – catalase, SOD – superoxide dismutase, GPx –

glutathione peroxidase, GST – glutathione S-transferase, UA – uric acid, GSH – glutathione reduced)

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 Pathophysiology of Gastric Cancer

Oxidative stress

Oxidative stress can be defined as any disturbance in the balance of

antioxidants and pro-oxidants in favor of the later due to different factors

such as aging, drug actions and toxicity, inflammation and/or addiction (Sies

H, 1985). It is in general, excess formation or/and insufficient removal of

highly reactive molecules such as reactive nitrogen species (RNS) and

reactive oxygen species (ROS) (Johansen et al., 2005). Oxygen is highly

reactive specie that has the ability to become part of potentially harmful and

damaging molecules (Free Radicals). Oxidative stress causes healthy cells of

the body to lose their function and structure by attacking them. Up until now,

pathogenesis of about more than 50 diseases has been implicated by free

radicals. It is when the antioxidant level is limited that this damage can

become debilitating and cumulative (Mark P, 1996). Damage to DNA,

proteins, and other macromolecules due to oxidation has been implicated in

the pathogenesis of a wide variety of diseases, most notably cancer and heart

disease (Halliwell B, 1994).

Oxidative stress induced organ damage

Lungs Asthma, chronic bronchitis

Kidneys Glomerulonephritis, chronic renal failure
Joints Arthritis, rheumatism
Brain Alzheimer’s disease, Parkinson’s disease, memory loss,
depression, stroke
Eyes Cataract, retinal diseases

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 Pathophysiology of Gastric Cancer

Fetus Preeclampsia, IU growth restriction

Heart Arteriosclerosis, hypertension, ischemia,
vessels cardiomyopathy, heart failure

Multiorgans Cancer, diabetes, inflammation infection, aging

Oxidative stress and gastrointestinal diseases

Oxidative stress plays important roles in pathogenesis of gastro-intestinal

diseases, which include mucosal damage, gastro-intestinal ulcers, and cancer.

Although gastric ulcer can be generated by different factors, e.g., non-

steroidal anti-inflammatory drugs (NSAIDs), thermal stress, ethanol, and H.

pylori infection, leading to oxidative damage through free radical generation

(specially OH·) and subsequent apoptotic responses of gastric mucosa (Ding

S Z, et al., 2007). In addition, gastro-intestinal diseases are associated to

increased oxidative stress and oxidants levels, such as glutathione, lipid

peroxidation, myeloperoxidases, protein carbonyl, etc (Ganguly K, et al.,

2012). Furthermore, pathogens are directly involved in aggravating oxidative

stress; for example, complete eradication of H. pylori is reported to attenuate

oxidative stress in gastric mucosa (Banerjee A, et al., 2013)

Although certain types of gastro-intestinal inflammations, like

ulcerative colitis, hepatitis, H. pylori infection, are more prone to develop

cancer, the reasons are still not well elucidated. Inflammation and

subsequent elevated oxidative stress might be the factors for aggravating

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 Pathophysiology of Gastric Cancer

chronic inflammation and inducing malignant transformation. Transgenic

mice expressing hepatitis B protein in liver develop chronic hepatitis with

elevated levels of 8-oxo-dG, leading to hepatocellular carcinoma (Hagen TM

et al., 1994). It is well accepted that inflammation is always accompanied

with elevated oxidative stress in cancer. Gastric cancer patients (with normal

renal and hepatic functions) are found to have significantly increased lipid

peroxidation levels (Reddy E P, et al., 2010). Gastric carcinoma patients have

significantly higher myeloperoxidase activity than controls, both before and

after operation, although total antioxidant status was decreased post-

operation (Czygier M, et al., 2010). Gastric cancers are also associated with

augmented protein oxidation, although no differences are found in oxidative

stress parameters and antioxidant enzyme activities between anti-H. Pylori

IgG positive and negative gastric cancer patients (Noyan T, et al., 2009).

Gastric enzyme - Pepsinogen

Pepsinogen is an inactive form of pepsin, which is the most important

proteolytic enzyme of gastric juice. (Tanaka Y, et al., 1991). Pepsinogen is

synthesised in the gastric mucosa by the chief and neck cells of the glandular

ducts and is secreted into the lumen of the gastric gland (Simpson L, et al.,

1980). Human pepsinogen can be separated on polyacrylamide gel

electrophoresis into seven isozymogens, consisting of two immunochemically

distinct groups: pepsinogen A (pepsinogen-I) and pepsinogen C (pepsinogen-

II). In contrast to pepsinogen C, pepsinogen A is secreted in the urine and

shows considerable interindividual heterogeneity. Several genetic models

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 Pathophysiology of Gastric Cancer

have been proposed to explain the inheritance of urinary pepsinogen A

(Korsnes L, et al., 1980).Increased activation of pepsinogen into pepsin by

enhanced acidity of gastric contents can cause ulcers in humans and animals

(Tanaka Y, et al., 1991; Saez-Alquezar A, et al., 1978; Vianello F, et al., 1988).

Pepsinogen-C/ Pepsinogen-II

Progastriscin/ Pepsinogen-C/ pepsinogen-II/ PGC, acts as a proteolytic

enzyme of gastric secretion (EC 3.4.23.3). It is a 40-kDa zymogen, an inactive

form of aspartic protease (Foltmann B, et al., 1982). PGC imparts aspartyl

proteinaselike features and is mainly synthesized in the gastric mucosa. After

synthesis, it is secreted into the gastric lumen where the proenzyme is

converted into its active proteolytic form under certain acidic conditions

(Moore S A, et al., 1995). It is widely distributed in the gastrointestinal tract

of various species and constitutes a major proteolytic enzyme present in the

gastric fluid (Samloff I M, 1989). The primary structure deduced from its c-

DNA and genomic clones reveals that PGC is a single polypeptide chain of 388

amino-acid residues and shares a high degree of sequence identity to that of

other aspartyl proteinases, e.g. pepsinogen A, procathepsin D, procathepsin E

and prorenin (Hayano T, et al., 1988; Taggart R T, et al., 1989). Another

biochemical study shows that the conversion of human progastricsin to

gastricsin is accompanied by two intermediates (Foltmann B, et al., 1982).

The first intermediate species has formed rapidly after a sudden drop in Ph

,4.0. Recently, the second intermediate has been characterized structurally

(Khan A R, et al., 1997).

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 Pathophysiology of Gastric Cancer

Role of PGC in the gastric tumor progression

Gastric adenocarcinoma is most common disease in Asian countries

and common cause of mortality in Europe (Pinto-Correia A L, et al., 2006). It

is well documented that any change in PGC could reflect the degree of gastric

disease and differentiation (Tsukamoto T, et al., 2007). It is evident from

epidemiological studies that Helicobacter pylori carriers have a significantly

greater risk of developing gastric cancer (Zhang L, et al., 1996). It was found

that PGC is a mature marker of stomach cells, and change in its expression

could reflect the degree of gastric lesions. Recently, the expression of PGC in

different gastric diseases was studied (Ning P F., et al., 2005), and the results

showed that PGC expression was 100% in normal gastric mucosa and only

2.4% in the gastric cancer. Moreover, such expression profile of PGC

decreased gradually from benign lesions to precancerous lesions to gastric

cancer.

The presence of PGC in metastasis is significantly correlated with a

primary tumor of the stomach, prostate, and pancreas (Duffy M J, 1996). The

overexpression of PGC has also been determined in the cancerous cells, basal

cell, and squamous cell carcinomas of the eyelids (Alvarez M L, et al., 2004).

Similarly, extra-digestive expression of PGC has been reported in several

tumors such as cervical, endometrial and ovarian (Serra D, et al., 1999),

prostate (Konishi N, et al., 1999), gastric (Fernandez R, et al., 2000), stomach

(Tatematsu M, et al., 1990), gall bladder (Tatematsu M et al., 1988), breast

carcinomas (Serra D, et al., 1999), and cutaneous malignant melanomas

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 Pathophysiology of Gastric Cancer

(Quintela I, et al.,2001). Interestingly, these expressions are significantly

associated with a longer overall survival of patients with breast carcinomas

(Merino A M, et al., 2000), Zhang X H, et al., (2006) observed that the serum

PGC-level abnormality was accompanied with a higher risk of precancerous

lesions. The presence of PGC in cancer cells implicated some mature

secreting function in them. However, the decrease of PGC expression

indicated dedifferentiation or malignancy of cancer cells, and it was also

closely related to prognosis and metastasis (Nakamura T, et al., 1997).

PGC Gene structure and regulation

The gene sequence of PGC from human was determined by many

groups independently (Evers M P, et al., 1989; Ishihara T, et al., 1989;

Takahashi K, 1992). Chromosomal localization was assigned to the PGC gene,

and subsequently its evolutionary mechanism has been established

(Kageyama, T, 2002). Hormonal regulation of PGC in breast cancer cells

suggests that androgens, glucocorticoids, and progesterone are capable of

inducing the expression of PGC gene in different breast cancer cell lines

(Balbin M, 1996). The PGC gene was cloned and expressed from various

species, e.g. human (Boudi F H, et al., 1990), rabbit (Kageyama T, et al., 1990),

rat (Ishihara T, et al., 1989), chicken (Hayashi K, et al., 1988), and

Epinephelus coioides (Feng S, et al.,2008). PGC gene was extensively

characterized, and it was found that it consists of 9 exons and 8 introns

(Takahashi k, 1992). The polypeptide sequence has been divided into two

parts, one 59-residue-long prosegment and another 329-residue-long pepsin

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 Pathophysiology of Gastric Cancer

moiety. Interestingly, the proposed substrate-binding sites were well

conserved among fish. The gene of human PGC is located on chromosome 6

and there is only a single copy in the human genome project sequence data.

Furthermore, it is suggested that PGC genes are derived from a common

pepsin-like ancestral gene (Kageyama T, 2002).

Although gene expression of PGC depends on various factors,

hormonal regulation is quite important among them (Ichinose M, et al.,

1988). Sakamoto N, et al. (2000) found that there are four GATA and one Sox-

binding motifs in 1.1 kb of the 5′ flanking region of the PGC gene, which are

essential to the organ-specific expression of the PGC. Hormonal regulation of

human PGC gene in the breast cancer cells is well known for a long time

(Balbin M, 1996). It was observed that a special factor regulates the

transcription of PGC. This factor is a 25-kDa protein that potentially binds to

a specific site in the 5′ flanking region of the gene only in the presence of

Mg2+ ion (Ishihara T, et al., 1989).

Direct actions of hormones on PGC genes are unknown. As suggested

by the fact that mesenchyme is necessary for enzyme induction

glucocorticoids may act indirectly on PGC genes through other factors

(Tsukada S, et al., 1998). In the other study, it was suggested that

glucocorticoids presumably inhibit the proliferation of mucosal cells and in

turn enhance the expression of pepsinogen gene (Tsukada S, et al., 1994).

Recently, the amino-acid sequences of three PGCs (PGC1, PGC2 and PGC3)

were deduced by cloning and nucleotide sequencing of Pacific bluefin tuna

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 Pathophysiology of Gastric Cancer

(Thunnus orientalis) (Tanji M, et al., 2009). It was found that all these forms

contained 16 signal sequences, an activation pro-peptide and mature peptide

of 321, 323, and 332 residues long in PGC1, PGC2, and PGC3, respectively.

(A) Three-dimensional structure of human PGC. The catalytic Asp residues are (light green) and

cysteine residues (yellow) are shown in ball and stick model. (B). Superimposition of human PGC

(green) with procine pepsin C (sky blue). The catalytic Asp residues of human PGC and procine pepsin C

are shown in green and yellow, respectively. Structure was drawn from the coordinates of PDB id: 1HTR

and 2PSG (Moore et al., 1995)

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