Department of Clinical Pathomorphology, Medical University of Lublin Agnieszka Fronczek, Justyna Śliwińska, Justyna Szumiło Adenocarcinoma of the stomach – epidemiology, prevention and new therapeutic strategies Gruczolakorak żołądka – epidemiologia, zapobieganie i nowe strategie leczenia Key words: adenocarcinoma, stomach, gastric carcinoma, histopathology, epidemiology, prevention, treatment

Abstract Carcinoma of the stomach is the fourth most common malignant neoplasm in the word but the second most common cause of death from cancer. Histologically, majority of them are adenocarcinomas. Many histological classification systems of gastric carcinomas have been developed. The most commonly used are the World Health Organization and Lauren classifications. Significant risk factors for intestinal type of gastric carcinoma are: Helicobacter pylori infection, excessive salted and smoked food consumption with increased concentration of nitrates, diet low in fruits and vegetables, smoking, alcohol and excess body weight. Unfortunately, the risk factors for diffuse type remain unclear. Gastric carcinomas are associated with poor prognosis, despite advances in surgical techniques and application of adjuvant or neoadjuvant therapy. At present, the only chance for successful treatment is complete resection with cancer-free margins, however most patients are diagnosed in advanced stage with unresectable tumors. The increasing knowledge on the role of genetic and epigenetic abnormalities in pathogenesis and behavior of the tumor can be applied in diagnosis and treatment of gastric carcinomas. One of the promising is HER2 targeted therapy using monoclonal antibody. Therefore, evaluation of HER2 receptor status should be performed in every case of unresectable and recurrent carcinoma of the stomach or gastroesophageal junction. The most important strategy to lower mortality should be prevention and screening programs.

Streszczenie Rak żołądka jest czwartym, co do częstości występowania nowotworem złośliwym na świecie, ale jednocześnie drugą przyczyną zgonu z powodu choroby nowotworowej. Zdecydowaną większość raków żołądka stanowią gruczolakoraki. Opracowano wiele

klasyfikacji histologicznych tych nowotworów, jednak najbardziej rozpowszechniona jest klasyfikacja Światowej Organizacji Zdrowia oraz Laurena. Najważniejszymi czynnikami ryzyka w typie jelitowym raka żołądka są: zakażenie Helicobacter pylori, nadmierne spożycie solonych i wędzonych potraw ze zwiększoną zawartością azotanów, dieta uboga w owoce i warzywa, palenie tytoniu, alkohol i nadmierna masa ciała. Natomiast czynniki ryzyka w typie rozlanym nie zostały w pełni poznane. Rokowanie w raku żołądka jest złe, mimo postępów w chirurgii

i

intensywnej

terapii

oraz

zastosowania

leczenia

adjuwantowego

lub

neoadjuwantowego. Obecnie jedyną metodą dającą szanse na wyleczenie jest całkowita resekcja z pozostawieniem wolnych marginesów, jednak większość pacjentów jest diagnozowana w zaawansowanym stadium choroby, kiedy interwencja chirurgiczna nie jest możliwa. Pogłębienie wiedzy o zaburzeniach genetycznych i epigenetycznych w raku żołądka może wpływać na skuteczniejszą diagnostykę i terapię. Jedną z bardziej obiecujących metod leczenia jest spersonalizowana terapia celowana z użyciem przeciwciał monoklonalnych skierowanych przeciwko receptorowi HER2. Oceny statusu receptora HER2 powinno się dokonywać u wszystkich pacjentów ze wznową lub nieresekcyjnym rakiem żołądka i połączenia żołądkowo-przełykowego. Najważniejszymi strategiami pozwalającymi na zmniejszenie śmiertelności z powodu raka żołądka są zapobieganie i badania przesiewowe.

Introduction Carcinoma of the stomach is the fourth most common malignant neoplasm in the word but the second most common cause of death from cancer (Khushalani, 2008). Histologically, majority of them are adenocarcinomas. They can be classified according to the location into: gastroesophageal junction, proximal and distal stomach tumors. The incidence of carcinomas in the body and the distal stomach decreased, but the occurrence of the lesions in gastroesophageal junction increased. Surgical resection can be regarded as curative treatment but exclusively in patients with early stage cancer. However, most patients are diagnosed in advanced stage with unresectable tumors (Khushalani, 2008). The survival of patients with advanced but resectable lesions remains poor, as well. In developed countries more than a half of patients die during five years after presentation, despite additional treatment strategies like preoperative chemotherapy or adjuvant chemoradiation (MacDonald et all, 2001). Therefore, new therapeutic strategies are needed. A better understanding of the molecular basis of the cancer will allow the development of molecular targeted, personalized treatment. Decrease of gastric cancer mortality can be achieved also by effective prevention strategies, based on

evaluation of specific risk profiles including: Helicobacter pylori genotype, host gene polymorphism and environmental factors (Hartgrink et all, 2009).

Clinical and morphological features of gastric adenocarcinoma Clinically, gastric carcinoma is usually asymptomatic until advanced stage. In the early stages indigestion and stomach discomfort, mild nausea and loss of appetite may occur. The late symptoms include: weight loss, abdominal pain, anorexia, vomiting, anemia and hemorrhage, abdominal distention secondary to ascites and bowel obstruction. Sporadically metastatic lymph nodes can be palpated in the left supraclavicular area (Virchow node) and left axillary region (Irish node). Bilateral enlargement of the ovaries can be the consequence of gastric cancer metastatses (Krukenberg tumors). In the course of the tumor some paraneoplastic syndromes are also noticed including acanthosis nigricans or seborrheic keratosis (LeserTrelat sign) (Khushalani, 2008). The most frequent location of gastric adenocarcinomas is antro-pyloric region. Lesions of the body are usually located along the greater or lesser curvature. Many histological classification systems of gastric carcinomas have been developed. The most commonly used are the World Health Organization (WHO) and Lauren classifications (Lauwers et all, 2010). According to WHO gastric adenocarcinoma is subdivided based on the predominant histological pattern into: papillary, tubular, mucinous, poorly cohesive (including signet-ring cell carcinoma) and mixed types (Lauwers et all, 2010). The old but widely accepted and clinically useful, Lauren classification subdivides gastric carcinomas into three main categories. The most frequent is the intestinal type characterized by formation of neoplastic glands, resembling those of colonic adenocarcinoma and lined by pleomorphic tall columnar or cuboidal cells (Fig. 1A). Diffuse type presents with dispersed, poorly differentiated cells, with little if any formation of glandular structures (Fig. 1B). Tumor cells contain clear vacuoles filled with mucus, pushing the nucleus to the cell periphery (signet ring cell carcinoma). This type displays frequently typical gross pattern, so-called linitis plastica (“lather bottle”) which is caused by dispersed interstitial infiltration with enhanced desmoplastic reaction and formation of extremely thick and rigid wall of the organ (Hartgrink et all, 2009). In the mixed type usually both patterns are observed (Stelzner et all, 1997). The basis of Goseki classification is evaluation and combination of two features: formation of neoplastic glands and amount of intracellular mucin in one tumor. It was proved, that survival of patients with carcinomas infiltrating subserosa (T3) but rich in mucus (Goseki II and IV) is

worse than that poor in mucus (Goseki I and II) (Lauwers et all, 2010). The Japanese Classification of Gastric Carcinoma is significantly similar to WHO one (Nishi et all, 1995). The old macroscopic classification of Borrmann distinguished four types of advanced gastric cancers: polypoid, fungating, ulcerated and infiltrative (Lauwers et all, 2010). Early gastric carcinomas are macroscopically classified as protruded (type I), superficial (type II) and excavated (type III). The superficial one is further divided into elevated (type IIa), flat (type IIb) and depressed (type II c) (Nishi et all, 1995).

Epidemiology and risk factors of gastric adenocarcinoma About two-third cases of gastric carcinomas is diagnosed in developing countries. High-risk areas include: East Asia (China, Japan), Eastern Europe as well as Central and South America. It affects about one million people per year (Hatrgrink et all, 2009). In most countries the neoplasm is more frequent in males (M:F is 2:1). Sex discrepancy is probably the consequence of protective effect of endogenous ovarian hormones (Duell et all, 2010). The 5-year survival rate for advanced gastric cancer is about 20%. The frequency of diffuse type of gastric carcinoma is almost invariable but, the frequency of intestinal type in well developed countries decreases (Khushalani, 2008). The risk factors for intestinal type of gastric carcinoma are relatively well-known. The most significant are: Helicobacter pylori infection, excessive salted and smoked food consumption with increased concentration of nitrates, diet low in fruits and vegetables, smoking and alcohol (Lauwers et all, 2010). Excess body weight (body mass index >25) is also associated with an increasing risk of gastric cancer, especially that located in the proximal part of the organ (Yang et all, 2009). Other risk factors include: pernicious anemia, hypochlorchydria favoring colonization of gastric mucosa by H. pylori, partial gastrectomy and blood group A. It was proved that carcinoma developed through a sequence of precursor lesions induced by Helicobacter pylori. Infection caused chronic inflammation, followed by atrophy of the mucosa, intestinal metaplasia, dysplasia and carcinoma (Lauwers et all, 2010). These alterations are the consequence of both bacterial proteins and host immune response. It is also likely that growth of cancer is strongly associated with more virulent strains of H. pylori (i.e. the Cag A strain). Risk of gastric adenocarcinoma in infected individual is 6x greater than in person free of H. pylori (Hartgrink et all, 2009). The risk factors for diffuse gastric carcinoma are unclear. It is more likely present in younger patients and has a genetic basis. Hereditary conditions that raise gastric carcinoma risk are Lynch syndrome, Li-Fraumeni syndrome with germline mutation of TP53 and Peutz -

Jeghers syndrome (Lauwers et all, 2010). Hereditary diffuse gastric cancer is a familial carcinoma syndrome caused by germline mutations of E-cadherin/CDH1 gene in 30-40% of cases (Lauwers et all, 2010). It accounts for about 1-3% of all gastric carcinomas. Estimated life time risk of gastric cancer in carriers of a CDH1 mutation is 67% in men and 83% in woman (Hartgrink et all, 2009).

Molecular markers The staging of gastric carcinoma is the most important prognostic factor. However, prognosis varies among patients in the same stage. The increasing knowledge on the role of genetic and epigenetic abnormalities in pathogenesis and behavior of the tumor can be applied in diagnosis and treatment of gastric carcinomas. Finding of molecular markers that are able to predict the potential of tumor recurrence and prognosis is important for appropriate individualized therapy. Some of these markers like p53 protein and nm23, are related to invasiveness of the tumor and poorer post-operative prognosis (Yasui et all, 2001). Genetic instability is one of the pathways in which the genomic changes arise in sporadic gastric cancer. Silencing of MLH1 (MutL homolog 1) proteins through promoter hypermetylation is the most frequent cause of the microsatellite instability, associated with less frequent lymph node metastases and extended survival (Yasui et all, 2001). CDKN2A (p16) (cyclindependent kinase inhibitor 2A) gene hypermetylation is noted in 12-30% of gastric carcinomas. Reduced expression of CDKN2A correlates with depth of invasion and metastasis (Lauwers et all, 2010). One of the new molecular markers is PRL-3 (phosphatase regenerating liver 3). Its expression in gastric carcinoma from surgical specimen seems to be an independent factor in prediction of recurrence and survival (Dai, 2009). Detection of mRNA for keratin 19 or CEA (carcinoembrionic atigen) is useful for evaluation of micrometastases in the lymph nodes and free cancer cells in peritoneal washes negative for abnormal cytology (Yasui i wsp. 2001). Other markers providing additional prognostic data in gastric carcinoma are number of DNA copy in neoplastic cells and expression of EGFR (epidermal growth factor receptor), thymidylate synthese, CDH1 (cadherin 1), β-catenin, VEGFR (vascular endothelial growth factor receptor), matrix metalloproteinases, TGFα (transforming growth factor α), and cyclin E (Scatozzi et all, 2004). Some of these markers can be a molecular target to chemotherapeutics or to biologic targeted therapies (Scatozzi et all, 2004). The HER2 protein (p185, HER2/neu, Erb-2) is a transmembrane tyrosine kinase receptor. In carcinomas HER2 acts as oncogene since high level of gene amplification induces

protein overexpression. This phenomenon is also observed in gastric carcinomas (Fig. 2A-B). The results of immunohistochemical reaction with this marker differed significantly by histological subtype (34, 6 and 20% positive cases in intestinal, diffuse and mixed type, respectively) and site of the tumor (32% and 18% positive cases in gastro-esophageal junction and gastric localization, respectively). It was correlated with more aggressive behavior of the tumor and poor outcome. HER2 expression is crucial for identifying of patients who could respond to HER2 targeted therapy using the fully humanized monoclonal antibody (Gravalos et all, 2008). The evaluation of HER2 receptor should be performed in every case of unresectable and recurrent carcinoma of the stomach or gastro-esophageal junction. The HER2 status is evaluated immunohistochemically using modified criteria established especially for gastric carcinoma (Ruschoff et all, 2010). In borderline status (+2) evaluation of gene amplification by FISH (fluorescence in situ hybridization) is proposed.

Treatment Surgery is the standard approach for localized gastric cancer. Extend of the operation depends on the stage of the tumor. Patients with early gastric cancer, i.e. confined to the mucosa or submucosa, irrespective of regional lymph node status, can undergo local excision by endoscopic mucosal resection. Endoscopic submucosal dissection is a new technique suitable for obtaining even large tumors in one piece and indicated for well differentiated intramucosal carcinomas. Complete tumor removal (R0 resection) is the only potentially curative treatment in localized, non-metastatic gastric carcinomas, however loco-regional recurrence and metastatic spread is frequent (Hartgrink et all, 2005). Additional treatment like neoadjuvant or adjuvant radio-chemotherapy and perioperative chemotherapy has been advocated to reduce postoperative relapse rates (MacDonald et all, 2006). Chemotherapy is applied in advanced gastric carcinoma, but in phase III studies overall survival does not exceed 1 year (Hartgrink et all, 2009). Some of tumor markers, e.g. microsatellite and chromosomal instability or overexpression of thymidylate synthetase, are thought to be predicting for therapy response and might improve quality of life for non-responders and reduce time until surgery (Hartgrink et all, 2009). The rapid development of targeted therapies in many malignancies has been observed recently. Specific antibodies against molecular targets such as HER2, EGFR and VEGFR are investigated in clinical trials. Trastuzumab (Herceptin®, Roche) is a monoclonal antibody which specifically binds the extracellular domain of HER2 receptor (Gravalos et all, 2008). In randomized trial addition of trastuzumab to conventional chemotherapy with capecitabine and

cisplatin significantly prolonged survival in patients with high expression of HER2 advanced gastric or gastro-esophageal junction carcinoma (overall survival 11 vs. 16 months for control and study groups, respectively) (Fujita et all, 2010). Other biologic agent used in targeted therapy of gastric and gastro-esophageal junction adenocarcinoma is EGFR inhibitor – Erlotinib® (Roche). The response rate in the Southwest Oncology Group 1237 trial was 9% and all responses occurred in patients with gastroesophageal junction carcinoma (Dragovich et all, 2007). Inhibitors of angiogenic factors have also been explored in clinical trials. Bevacizumab® (Genetech/Roche) is an anti-VEGF humanized monoclonal antibody. It was applied in combination with cisplatin and irinotecan in patients with advanced gastro-esophageal junction adenocarcinoma with 65% of overall response rate and median survival – 12,3 months (Shah et all, 2006).

Prevention The main strategy for decreasing gastric cancer mortality should be primary prevention. Targets for prevention are modifiable risk factors, i.e. high salt and nitrite consumption, low fruit and vegetable intake, cigarette smoking and H. pylori infection. The term chemoprevention is referred to the prevention of cancer using specific agents to suppress or reverse the carcinogenic process. Recently, special attention had been focused on the anticancer properties of non-steroidal anti-inflammatory drugs, H. pylori eradication and antioxidant supplementation (Nardone et all, 2004). In families with at least two members suffering from diffuse gastric carcinoma including one diagnosed >50 years of life, mutation analysis is recommended. In patients with mutation in CDH1 gene and fulfilling criteria for hereditary diffuse gastric carcinoma prophylactic gastrectomy should be also considered (Hartgrink et all, 2009). Initial screening tool in some countries are still barium swallow and serum pepsinogen level evaluation. In case of abnormal results patient is screened by endoscopy. Screening by endoscopic examination seems to be the most accurate method for detection of gastric cancer (Lauwers et all, 2010).

Conclusions The most important strategies to decrease mortality from gastric carcinoma should be prevention and screening programs. In Western countries most of patients are diagnosed in advanced stage with unresectable tumors. The only chance for successful treatment of gastric

carcinoma is complete resection with cancer-free margins. High mortality rate, despite advances in surgical techniques and application of adjuvant or neoadjuvant therapy, indicated development of modern, personalized and targeted treatment. The increasing knowledge on the role of genetic and epigenetic abnormalities in pathogenesis and behavior of the tumor can be applied in diagnosis and treatment of gastric carcinomas.

(A)

(B)

Fig.1 Intestinal (A), and diffuse (B) type of gastric adenocarcinoma according to Lauren classification (hematoxylin&eosin; objective magn. (A)x10, (B) x20).

(A)

(B)

Fig. 2 Positive (A), and negative (B) immunostaining for HER2 in intestinal and diffuse type of gastric adenocarcinoma, respectively (Herceptest and EnVisionTM+/HRP, DakoCytomation; objective magn. (A)x20, (B)x10).

References 1. Carneiro F, Huntsman DG, Smyrk TC, Owen DA, Seruca R, Pharoah P, Caldas C, Sobrinho-Simoes M. (2004) Model of the early developement of diffuse gastric cancer in Ecadherin mutation carriers and its implications for patient screening. Journal of Pathology. Vol. 203, s. 681-687. 2. Dai N. (2009) Expression of phosphatase regenerating liver 3 is an independent prognostic indicator for gastric cancer. World Journal of Gastroenterology. Vol. 15, s. 1499-1505 3. Davesa S, Fraumeni JF Jr. (1999) The rising incidence of gastric cardia cancer. Journal of National Cancer Institute. Vol. 91, s. 747-749. 4. Dragovich T, McCoy S, Fenoglio-Preiser CM. (2007) Phase II trial of erlptinib in gastroesophageal junction and gastric adenocarcinomas. Journal of Clinical Oncology. Vol. 25, s. 2334. 5. Duell E, Travier N, Lujan-Barroso L. (2010) Menstrual and reproductive factors, exogenous hormon use, and gastric cancer risk in cohort of women from European Prospective Investigation Into Cancer and Nutrition. American Journal of Epidemiology. doi:10.1093/aje/kwq321. 6. Fuccio L, Zagri RM, Eusebi LH, Laterza L, Ceroni L. (2009) Meta-analysis : can Helicobacter pylori eradication treatement reduce the risk for gastric cancer? Annales of Internal Medicine. Vol. 151, s. 121-128. 7. Fujita, T. (2010) Trastuzumab for gastric cancer treatment. Lancet. Vol. 367, s. 1735-1736. 8. Gonzales C, Riboloi E. (2010) Diet ad cancer prevention, contributions from the European Prospective Investigation into Cancer and Nutrition (EPIC) study. European Journal of Cancer. Vol 46, s. 2555-2562. 9. Gravalos C, Jimeno A. (2008) HER2 in gastric cancer: a new prognostic actor and noveltherapeutic target. Annals of Oncology. Vol.19, s. 1523-1529. 10. Hartgrink HH, Jansen E P, van Grieken N, van de Velde C. (2009) Gastric cancer. Lancet. Vol. 374, s. 477-90. 11. Hartgrink HH, van de Velde DJ. (2005) Status of extended lymph node dissection: locoregional control is the only way to survive gastric cancer. Journal of Surgical Oncology. Vol. 90, s, 153-65. 12. Khushalani N. (2008) Cancer ofthe esophagus and stomach. Mayo Clinlinical Procedures. Vol. 83, s. 712-22.

13. Lauwers GY, Carneiro F, Graham Dy, Curado M. (2010) Gastric carcinoma. In: WHO classification of tumours of the digestive system. Edited by: Carneiro F, Hruban R, Theise N, Bosman F. Lyon. International Agency for Research on Cancer (IARC), s. 46-94. 14. Leung WK, Wu MS, Kakugawa Y, Kim JJ, Yeoh KG, Goh KL, Wu DC, Sollano J, Kachintorn U, Gotoda T, Lin JT, You WC. (2008) Screening for gastric cancer in Asia: current evidence and practice. Lancet oncology. Vol. 9, s. 279. 15. MacDonald JS, Smalley SR, Benedetti J. (2001) Chemoradiotherapy after surgery compared with surgery alone for adenocarcinoma of stomach or gastroesophageal junction. New England Journal of Medicine. Vol 345, s. 725-30. 16. Nardone G, Rocco A. (2004) Chemoprevention of gastric cancer: role of COX-2 inhibitors and other agents. Digestive Diseases. Vol. 22, s.320-326. 17. Nishi M, Omori Y. Miwa K. Histological findings. (1995) Japanese Classification of Gastric Carcinoma. Tokyo. Japanese Reserch Society for Gastric cancer. Kanehara and Co. First English edition. 18. Ruschoff J, Dietel M, Baretton G, Arbogast S, Walch A, Monges G, Chenard MP, Penault-LorcaF, Nagelmeier I, Shlake W. (2010) Her2 diagnostics in gastric cancer – guideline validation and development of standardized immunohitochemical testing. Virchows Archives. Vol. 457, s. 299-307. 29. Scatozzi M, Galizia E, Freddari F, Berardi R, Cellerino R, Cascinu S. (2004) Molecular biology of sporadic gastric cancer: prognostic indicators and novel therapeutic approaches. Cancer Treatemet Review. Vol. 30, s. 451-459. 20. Shah MA, Ramanathan RK, Ilson DH. (2006). Multicenter phase II study of irinotecan, cisplatin and bevacizumab in patients with metastatic gastric or gastroesophageal junctionadenocarcinoma (FOLCETUX study). Journal of Clinical Oncology. Vol. 24, s. 52015206. 21. Stelzner S, Emmrich P. (1997) The mixed type in Lauren’s Classification of gastric carcinoma. Histologic description and biologic behavior. General and Diagnostic Pathology. Vol.143, s. 39-48. 22. Yang P, Zhou Y, Chen B, Wan HW, Jia GQ, Bai HL, Wu XT. (2009) Overweight, obesity and gastric cancer risc: results from a meta-analysis of cohort studies. European Journal of Cancer.Vol. 45, s. 2867-73.