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Epidemiology and Genetics of Gastric Cancer New York Stomach Cancer Education Symposium Epidemiology and Genetics of Gastric Cancer Lecturer – Manish A. Shah, MD The Bartlett Family Associate Professor of Gastrointestinal Malignancies Director, Gastrointestinal Oncology Program Co-Director, Center for Advanced Digestive Care Weill Cornell Medical College/ New York-Presbyterian Hospital Debbie’s Dream Foundation Medical Advisory Board Member

Esophageal and Gastric Carcinoma US Incidence in 2016 U.S. : 43,280 new cases (2.6% all U.S. cancers) Gastric: 26,370 (61%) Esophagus: 16,910 (39%) Decline in Gastric Cancer Incidence Increase in Esophageal , GE JX, cardia adeno OS improvement, 1975-77, 1984-86, 1999-2006 Gastric: 16%  18%  27% Esophageal: 5%  10%  19% Siegel RL et al., CA Cancer J Clin. 2016; 66:7-30

Risk Factors for GI Cancer Diet Alcohol Hot drinks Tobacco smoking Acid reflux Obesity H. Pylori Atrophic gastritis Esophagus GE, gastroesophageal. —proximal nondiffuse, diffuse, and distal nondiffuse gastric cancer. + adeno oesophageal – Manish Shah Gastroesophageal junction Cardiac sphincter Liver

Factors Associated With Increased Risk of Developing Stomach Cancer Nutritional/environmental Salted or smoked foods High dietary nitrates Low intake of fruits, vegetables, and vitamin A and C consumption Low serum selenium Social Low socioeconomic status Tobacco use Medical Prior gastric surgery Helicobacter pylori infection Cag-1 Gastric atrophy and gastritis Pernicious anemia

Hereditary Diffuse Gastric Cancer Caused by germline mutation in CDH1, E-cadherin Cell-adhesion protein critical to development, cell differentiation, and cell architecture 60-80% penetrance Prophylactic gastrectomy – numerous in situ foci of signet ring cell adenocarcinoma

E. Cadherin/ Beta-catenin/ Wnt The E. Cadherin/ Beta-catenin/ Wnt signaling pathway Truncating mutation is responsible for DHGC ~ 2- 20% of sporadic diffuse GC have CDH1 mutations.

Criteria for testing for E-cadherin gene mutation Updated recommendations from the International Gastric Cancer Linkage Consortium (IGCLC)* Two or more documented cases of gastric cancer in first degree relatives, with at least one documented case of diffuse gastric cancer diagnosed before the age of 50 years 2. Three or more cases of documented diffuse gastric cancer in first- or second-degree relatives, independent of age of onset 3. Diffuse gastric cancer before the age of 40 years without a family history 4. Families with diagnoses of both diffuse gastric cancer and lobular breast cancer, with one case before the age of 50 years *In addition, in cases where expert pathologists detect carcinoma in situ adjacent to diffuse-type gastric cancer, genetic testing should be considered since this is rarely, if ever, seen in sporadic cases. Fitzgerald R et al:J Med Genet 47: 436-444, 2010

Disease Heterogeneity Gastric and Esophageal Cancers are not one disease. Esophagus Histology (squamous v adeno) Etiology (tobacco/EtOH v HPV) Gastric Histology (Intestinal vs Diffuse) Location (Cardia/GEJ vs Antrum) Biology (MET, CDH1, others?) Etiology (H. pylori related, others?)

Helicobacter pylori Robin Warren Barry Marshall

Helicobacter pylori Endemic infection present ~half of all humans. Particularly prevalent in developing countries Gram-negative bacteria that infects the antrum mucosa lining of the stomach leading to increase in pH levels Causes chronic gastritis, gastric ulceration, and gastric atrophy World Health Organization class-I carcinogen

Proximal vs Distal Gastric Cancer Two Distinct Patterns Cardia/ GEJ Cancers Male: Female (5:1) white: black (2:1) wide age range industrialized nations Non-Cardia Cancers Male: Female (2:1) white: black (1:4) incidence increases with age (peak age 50-70)

Epidemiology Summary Wide variation in global incidence Diet, lifestyle, H. pylori Falling incidence of gastric and squamous cancer Barrett’s/esophageal carcinoma Rising incidence Screening strategies being studied Needs better markers to predict risk of progression Knowledge of the molecular drivers will lead to new therapies.

GC Genetic Predisposition Syndromes Genetic Considerations GC Genetic Predisposition Syndromes Account for 10% -15% of all Gastrointestinal Cancers Hereditary Diffuse GC (~3-5%) Lynch Syndrome (~1-2%) germline mutations in one of the mismatch repair genes MLH1, MSH2, MSH6, and PMS2 Stomach cancers occur in ~11% of Lynch syndrome families FAP (Familial Adenomatous Polyposis) (~1%) Germline mutation in APC – adenomatous polyposis coli Fundic gland polyps Li Fraumeni’s syndrome (< 1%) p53 mutation Peutz-Jeghers syndrome (<1%) Autosomal dominant, hamartomatous polyps of GI tract and mucocutaneous melanin deposits Germline mutations in STK11 – serine threonine kinase 11.

Genetic Considerations Disease Heterogeny Global Heterogeneity More prevalent in Asia H. pylori variance More advanced disease in US, West. Different underlying biology? Upper GI Cancers cannot be treated as one disease. Histology (Intestinal vs Diffuse) Location (Cardia/GEJ vs Antrum) Biology (MET, CDH1, others?) Etiology (H. pylori related, others?)

Carcinogenesis Conceptual Model Distal non diffuse Gastric CA Genetic Immune SNPs IL1, IL4, etc. Distal non diffuse Gastric CA Family History CDH1 MMR APC TP53 Genetic Risk Environment H. Pylori cag A strain Behavior Proximal non diffuse Gastric Cancer Diffuse Gastric CA Tobacco use/ diet (salt) (fruits/vegetables )

Prevalent Risk Factors (estimated OR) Disease Types Gastric Cancer Subtype Prevalent Risk Factors (estimated OR) Non-Cardia Environmental High dietary salt Eating Fruits / Vegetables (OR ~0.7) Tobacco (OR ~1.5) Age (peak at age 50-70) Clinical H. pylori infection (OR ~3.0) Use of NSAIDs/ Aspirin Genetic Immune regulatory SNPs Diffuse none specifically identified ? CDH1 mutation Family history (non- CDH1 mutant) Proximal Tobacco Use Alcohol Obesity/ High BMI GERD Shah and Kelsen, JNCCN 2010

Disease Types Gastric cancer subtypes each differ significantly from normal and from each other. Type 1=proximal Type 2=diffuse Type 3=distal False Discovery Rate=5% Fold Change > 2.0 Probes that are significantly different between NORMAL stomach and GC subtype

Molecular evidence for 3 types of gastric cancer Data Molecular evidence for 3 types of gastric cancer 36 microarrays (15 of type 1, 13 of type 2 and 13 of type 3). All arrays were processed using Bioconductor R-package. To perform supervised classification, pdmclass R-library was used that implements ridge regression with the optimal scoring algorithm CONCLUSION: Samples from different gastric cancer subtypes are quite well separated and very tightly grouped. Distal/ intestinal Proximal/ GEJ Diffuse > 85% prediction Shah et al Clin Cancer Res 2011;17:2693-701

Data Genomic Data Tan et al Gastroenterology 2011 Expression analysis of 37 gastric cancer cell lines 4 independent patient cohorts (n = 521 patients) Identified a 171-gene set which robustly classifies tumors into predominantly intestinal and diffuse GC subtypes.

Unsupervised Cluster Analysis Data Unsupervised Cluster Analysis Two major types of Gastric cancer: G- INT - Blue G- DIF - Red A. Hierarchical dendogram B. Silhouette widths of individual cell lines C. Heat map

Two major GC signatures: association with Lauren’s subtype Data Gastric Cancer Subtypes Two major GC signatures: association with Lauren’s subtype

Molecular Subtypes of GC and Key Features The Cancer Genome Atlas Research Network, Nature. 2014;513:202-209

Four Genomic Subsets Emerge: Genome Atlas Genomically unstable (50%) Intestinal, present in most GEJ tumors High rate of p53 mutation, amplification of RTK’s High rate of microsatellite instability (MSI), gene mutation, and promoter hypermethylation (22%) Genomically stable (20%) Associated with diffuse histology, CHD-1 and RHOA mutation High Epstein-Barr virus burden (9%) High rate of PIK3CA mutation, PD-L1 and 2 amplification, strong IL-12 signaling indicating an immune presence The Cancer Genome Atlas Research Network., Nature. 2014;513:202-209

Current Disease Paradigm Proximal non diffuse Gastric Ca - Chromosome Instable EBV Associated Gastric Ca Immune SNPs IL1, IL4, etc. Family History: CDH1 MMR APC TP53 Genetic Risk Environment H. Pylori (cag A strain) EBV Behavior Distal non diffuse Gastric CA Chromosome Instable MSI Diffuse Gastric CA Genomically Stable Tobacco use/ diet (salt) (fruits/vegetables) The Cancer Genome Atlas Research Network, Nature. 2014;513:202-209

PD-L1 and PD-L2 Expression Are Elevated in Gastric Cancer PD-L1 (CD274) 7 6 5 4 3 2 1 mRNA Expression (RNA Seq RPKM)(log2) CIN EBV GS MSI Molecular Subtype PD-L2 (PDCD1LG2) 3.5 3.0 2.5 2.0 1.5 1.0 0.5 mRNA Expression (RNA Seq RPKM)(log2) CIN EBV GS MSI Molecular Subtype CIN = chromosome instability; EBV = Epstein-Barr virus; GS = gnomically stable; MSI = microsatellite instability. Bass A. Comprehensive molecular characterization of gastric cancer. Presented at: TCGA Symposium, May 13, 2014.

PD-L1 Status and Outcome: Gastric Cancer PD-L1 is a negative prognostic factor for overall survival in gastric cancer PD-L1 expression 1.0 Negative Positive Negative-censored 0.8 Positive-censored PD-L1 Negative (n=65) 0.6 Cumulative Survival 0.4 PD-L1 Positive (n=67) 0.2 PD-1, programmed death-1; PD-L1, programmed death ligand-1. 0.0 24 48 72 96 120 144 168 Time after surgery (months) Zhang L et al. Int J Clin Exp Pathol. 2015;8(9):11084-11091.

Implications Genomic Analysis Gastric cancer is not one disease. Subtypes of gastric cancer exist. Observed differences in response to therapy and prognosis may be explained. Different molecular drivers Different therapeutic targets