Sunday, June 3, :30 PM – 8:30 PM Hyatt Regency Chicago Chicago, Illinois Translational Research 2012: Using Leading-Edge Science to Make the Correct Treatment Choice Today and Tomorrow Jointly sponsored by Postgraduate Institute for Medicine and Clinical Care Options, LLC This program is supported by educational grants from Image: suravid/Copyright©2012 Shutterstock Images LLC. All Rights Reserved Image: Gunilla Elam/Copyright©2012 Photo Researchers, Inc. All Rights Reserved Image: Carol and Mike Werner/Copyright©2012 Photo Researchers, Inc. All Rights Reserved Image: Science Photo Library/Copyright©2012 Custom Medical Stock Photo, Inc. All Rights Reserved In partnership with Translational Research in Oncology

Jordan D. Berlin, MD Associate Professor of Medicine Vanderbilt University Clinical Director, GI Oncology The Vanderbilt-Ingram Cancer Center Nashville, Tennessee Current and Future Directions in the Treatment of GI Cancers: Optimizing Treatment by Genetic Features of Disease Image: suravid/Copyright©2012 Shutterstock Images LLC. All Rights Reserved Image: Gunilla Elam/Copyright©2012 Photo Researchers, Inc. All Rights Reserved Image: Carol and Mike Werner/Copyright©2012 Photo Researchers, Inc. All Rights Reserved Image: Science Photo Library/Copyright©2012 Custom Medical Stock Photo, Inc. All Rights Reserved

clinicaloptions.com/oncology Translational Research 2012 Outline  EGFR-targeted antibodies in CRC –Rationale for and data supporting KRAS mutation testing  Biomarkers for future treatment algorithms –Impact of BRAF mutations on EGFR-targeted therapy –Other potential biomarkers –CTC levels  Role of HER2 in gastric cancer –Rationale for and data supporting use of HER2-targeted therapies  Biomarkers for future treatment algorithms –Other potential biomarkers

EGFR-Targeted Antibodies in Colorectal Cancer

clinicaloptions.com/oncology Translational Research 2012 EGFR-Targeted Antibodies in CRC  EGFR-targeted antibodies approved in CRC –Cetuximab –Panitumumab  EGFR expression status insufficient for discerning which patients derive most benefit  KRAS: signaling molecule downstream of EGFR –Mutated KRAS becomes constitutively active, negating effect of EGFR inhibition [1] –~40% of CRCs have mutated KRAS genotype [1] –Mutated KRAS associated with poor response to EGFR-targeted agents [2] 1. Lièvre A, et al. Oncogene. 2010;29: Dahabreh IJ, et al. Ann Intern Med. 2011;154:37-49.

clinicaloptions.com/oncology Translational Research 2012 KRAS Testing: What Are the Recommendations?  NCCN guidelines [1] –Strongly recommend KRAS testing in all patients with mCRC at the time of diagnosis of metastatic disease –Testing should be performed in a CLIA-certified lab –Testing can be performed on either primary or metastatic tissue  ASCO Provisional Clinical Opinion [2] –All patients with mCRC who are candidates for anti-EGFR antibody therapy should have their tumor tested for KRAS mutations in a CLIA-accredited laboratory  In both cases, anti-EGFR agents (cetuximab and panitumumab) are recommended for wild-type KRAS patients only [1,2] 1. NCCN. Clinical practice guidelines in oncology: colon cancer Allegra CJ, et al. J Clin Oncol. 2009;27:

clinicaloptions.com/oncology Translational Research 2012 Studies Evaluating Addition of EGFR- Targeted Antibody to Chemotherapy  Benefit in KRAS wild-type CRC patients only –CRYSTAL: cetuximab plus FOLFIRI [1] –OPUS: cetuximab plus FOLFOX-4 [2] –PRIME: panitumumab plus FOLFOX-4 [3]  No benefit in either KRAS wild-type or mutated CRC patients –COIN: cetuximab plus oxaliplatin/fluoropyrimidine [4] 1. Van Cutsem E, et al. J Clin Oncol. 2011;29: Bokemeyer C, et al. Ann Oncol. 2011;22: Douillard JY, et al. J Clin Oncol. 2010;28: Maughan TS, et al. Lancet. 2011;377:

clinicaloptions.com/oncology Expert Insight Into the First-line Treatment of Metastatic Colorectal Cancer CRYSTAL: KRAS Status in Response to Cetuximab  Randomized, multicenter phase III trial [1] –Significant improvement in PFS with addition of cetuximab to FOLFIRI vs FOLFIRI alone for first-line mCRC treatment  Updated survival analysis of CRYSTAL [2] –Included only subset of KRAS-evaluable patients (n = 1063) 1. Van Cutsem E, et al. N Engl J Med. 2009;360: Van Cutsem E, et al. J Clin Oncol. 2011;29: FOLFIRI* 2-wk cycle (n = 609) FOLFIRI* 2-wk cycle + Cetuximab † (n = 609) Stratified by geographic region, ECOG PS EGFR-positive patients with previously untreated mCRC (N = 1218) *FOLFIRI: irinotecan 180 mg/m 2, folinic acid 400 mg/m 2, 5-FU 400 mg/m 2 bolus, 5-FU infusion 2.4 g/m 2 over 46 hrs. † Cetuximab: 400 mg/m 2 on Day 1, then 250 mg/m 2 /wk.

clinicaloptions.com/oncology Expert Insight Into the First-line Treatment of Metastatic Colorectal Cancer CRYSTAL: Updated Survival Analysis Efficacy Results  Significant interactions between KRAS status and treatment outcome observed with PFS, OS, and ORR efficacy outcomes –Survival and response benefits with addition of cetuximab only achieved in KRAS wild-type patients Van Cutsem E, et al. J Clin Oncol. 2011;29: Outcome Cetuximab + FOLFIRI FOLFIRIHazard RatioP value Wild-type KRAS  Median PFS, mos  Median OS, mos  ORR, % * <.001 Mutated KRAS  Median PFS  Median OS  ORR, % * *Odds ratio

clinicaloptions.com/oncology Expert Insight Into the First-line Treatment of Metastatic Colorectal Cancer OPUS: KRAS Status in Response to Cetuximab  Randomized, open-label, multicenter phase II trial [1] –Significant improvement in ORR with addition of cetuximab to FOLFOX-4 vs FOLFOX-4 alone for first-line mCRC treatment  Biomarker analysis of OPUS [2] –Included only subset of KRAS-evaluable patients (n = 315) 1. Bokemeyer C, et al. J Clin Oncol. 2009;27: Bokemeyer C, et al. Ann Oncol. 2011;22: FOLFOX-4* (n = 168) FOLFOX-4* + Cetuximab † (n = 169) EGFR-positive patients with previously untreated mCRC (N = 344) *FOLFOX-4: oxaliplatin 85 mg/m 2, leucovorin 200 mg/m 2, 5-FU 400 mg/m 2 bolus, 5-FU infusion 600mg/m 2 over 22 hrs. † Cetuximab: 400 mg/m 2 on Day 1, then 250 mg/m 2 /wk.

clinicaloptions.com/oncology Expert Insight Into the First-line Treatment of Metastatic Colorectal Cancer OPUS: Biomarker Analysis OS Results Bokemeyer C, et al. Ann Oncol. 2011;22:

clinicaloptions.com/oncology Expert Insight Into the First-line Treatment of Metastatic Colorectal Cancer Meta-Analysis: KRAS Status in Response to Cetuximab  Pooled efficacy analysis of CRYSTAL and OPUS studies –CRYSTAL: FOLFIRI + cetuximab vs FOLFIRI alone [2] –OPUS: FOLFOX + cetuximab vs FOLFOX alone [3]  90% of samples were subjected to KRAS genotype testing –HRs for benefit of addition of cetuximab highly statistically significant in patients with wild-type KRAS –PFS –HR: 0.66 (P <.0001) –OS –HR: 0.81 (P =.0062) Bokemeyer C, et al. ASCO Abstract 3506.

clinicaloptions.com/oncology Expert Insight Into the First-line Treatment of Metastatic Colorectal Cancer PRIME: KRAS Status in Response to Panitumumab  Randomized, global, open-label, phase III trial Douillard JY, et al. J Clin Oncol. 2010;28: Panitumumab 6.0 mg/kg q2w + FOLFOX4 q2w (n = 593) FOLFOX4 q2w (n = 590) Stratified by ECOG PS (0-1 vs 2) and geographic region (Western Europe, Canada, and Australia vs all other locations) Patients with previously untreated mCRC (N = 1183)

clinicaloptions.com/oncology Expert Insight Into the First-line Treatment of Metastatic Colorectal Cancer PRIME: Efficacy Results  Wild-type KRAS patients –PFS significantly improved with FOLFOX4 + panitumumab –HR: 0.80,; 95% CI: ; P =.01 Douillard JY, et al. J Clin Oncol. 2010;28: Outcome, mos Panitumumab + FOLFOX4 FOLFOX4P value Wild-type KRAS  Median PFS  Median OS Mutated KRAS  Median PFS  Median OS  Mutated KRAS patients –Worse PFS outcome with panitumumab addition –HR: 1.27; 95% CI: ; P =.02

clinicaloptions.com/oncology Expert Insight Into the First-line Treatment of Metastatic Colorectal Cancer MRC COIN: KRAS Status in Response to Cetuximab  Randomized, open-label trial –Largest to assess addition of cetuximab to chemotherapy in first-line treatment of mCRC –Prospective analysis of efficacy in relation to KRAS mutation status OxFP* (n = 815) OxFP* + Cetuximab † (n = 815) Patients with previously untreated mCRC irrespective of EGFR status (N = 344) *OxFP: oxaliplatin 130 mg/m 2 plus capecitabine 850 mg/m 2 twice daily for 2 weeks, or oxaliplatin 85 mg/m 2 plus leucovorin 175 or 350 mg and 5-FU 400 mg/m 2 bolus then 5-FU infusion 2400 mg/m 2 over 46 hrs. † Cetuximab: 400 mg/m 2 on Day 1, then 250 mg/m 2 /wk. 3 rd treatment arm randomized patients to interittent OxFP (n = 815) Maughan TS, et al. Lancet. 2011;377:

clinicaloptions.com/oncology Expert Insight Into the First-line Treatment of Metastatic Colorectal Cancer MRC COIN: Survival Efficacy Results  No improvement in either OS or PFS with addition of cetuximab to chemotherapy in KRAS wild-type patients Maughan TS, et al. Lancet. 2011;377: KRAS wild-type KRAS mutated OxFP OxFP + Cetuximab

clinicaloptions.com/oncology Expert Insight Into the First-line Treatment of Metastatic Colorectal Cancer KRAS: Not all Mutations are Equal  Mutation pattern [1] –79% of CRC tumors have mutations in codon 12 –17.6% of CRC tumors have mutations in codon 13  Anecdotal reports suggest < 10% of KRAS-mutated CRC patients can respond to anti-EGFR therapy [2,3] –Codon 13 mutations overrepresented in these tumors  Codon 13 mutations show weaker transforming activity in vitro [4]  KRAS G13D mutation associated with longer OS and PFS vs other KRAS mutations [1] 1. De Roock W, et al. JAMA. 2010;304: Benvenuti S, et al. Cancer Res. 2007;67: Moroni M, et al. Lancet Oncol. 2005;6: Guerrero S, et al. Cancer Res. 2000;60:

A Vision of Tomorrow’s Treatment Algorithm: Colorectal Cancer

clinicaloptions.com/oncology Translational Research 2012 Impact of BRAF Mutations on Anti-EGFR Therapy  5% to 9% of CRC tumors have specific BRAF V600E mutation [1,2] –Results in constitutive activation of BRAF protein –Typically do not occur in tumors with KRAS exon 2 mutations [2]  Outcomes with BRAF V600E mutation –Non-first-line setting: BRAF V600E mutation associated with resistance to EGFR-targeted agents [3-5] –First-line setting: BRAF V600E mutation associated with poorer prognosis, but some patients may still benefit from addition of cetuximab to chemotherapy [6-7] 1. Van Cutsem E, et al. ASCO Abstract Tol J, et al. NEJM. 2009;361: Di Nicolantonio F, et al. J Clin Oncol. 2008;26: Laurent-Puig P, et al. J Clin Oncol. 2009;27: Loupakis F, et al. Br J Cancer. 2009;101: Van Cutsem E, et al. J Clin Oncol. 2011;29: Bokemeyer C, et al. ASCO Abstract 3506.

clinicaloptions.com/oncology Translational Research 2012 BRAF Mutations in First-Line Setting  CRYSTAL and OPUS meta-analysis –Patients with wild-type KRAS/mutated BRAF –Worse outcomes vs wild-type KRAS/wild-type BRAF –Still experienced non-significant improvements (small N) Outcome Wild-type KRAS/ Mutated BRAF Wild-type KRAS/ Wild-type BRAF Cetuximab + CT CT OnlyP Value Cetuximab + CT CT OnlyP Value Median PFS, mos <.001 Median OS, mos Median ORR, % <.0001 Bokemeyer C, et al. ASCO Abstract 3506.

clinicaloptions.com/oncology Translational Research 2012 Other Potential Biomarkers in CRC  PI3K: 39% [1] –Associated with significantly decreased PFS in patients treated with FOLFIRI plus cetuximab [1]  NRAS: 2% [2] –Prognostic/predictive value unclear [2]  EGFR ligands –Surrogate markers for EGFR inhibition with cetuximab [3]  DNA hypermethylation: 30% [4] –Associated with poor prognosis [5]  Defective mismatch repair: 15% [6] –Improved prognosis and stage- dependent survival [7] –Predictive for lack of efficacy of 5- FU-based adjuvant therapy [8]  VEGF receptors and ligand –VEGF expression may be associated with poorer prognosis [9] 1. Linot B, et al. ASCO Abstract Irahara N, et al. Diagn Mol Pathol. 2010;19: Cremolini C, et al. ASCO Abstract Kulendran M, et al. Cancers. 2011;3: Barault L, et al. Cancer Res. 2008;68: Kerr DJ. J Clin Oncol. 2010;28: Popat S, et al. J Clin Oncol. 2005;23: Sargent D, et al. J Clin Oncol. 2010;28: Bendardaf R, et al. Anticancer Res. 2008;28:

clinicaloptions.com/oncology Translational Research 2012 CTC Levels  mCRC patients treated with chemotherapy plus targeted agents in CAIRO2 phase III trial (N = 467) [1] –Higher CTC count at baseline and during treatment associated with lower survival vs lower CTC count –PFS HR: 1.5 –OS HR: 2.2  mCRC patients (N = 430) [2] –Higher CTC count at baseline and during treatment associated with lower survival vs lower CTC count –PFS: 4.4 vs 7.8 months, P =.004 –OS: 9.4 vs 20.6 months, P < Tol J, et al. Ann Oncol. 2010;21: Cohen SJ, et al. Ann Oncol. 2009;20:

Role of HER2 in Gastric Cancer

clinicaloptions.com/oncology Translational Research 2012 Rationale for Targeting HER2 in Gastric Cancer  10% to 25% tumors overexpress HER2 1  HER2 overexpression associated with poor prognosis 2 –Poor survival –Serosal invasion –Lymph node metastases –Disease stage –Distant metastases 1. Yano T, et al. Oncol Rep. 2006;15: Jorgensen JT, et al. J Cancer. 2012;3:

clinicaloptions.com/oncology Translational Research 2012 Chemotherapy Treatment of HER2- Positive Gastric Cancer Patients  Patients with initially metastatic or recurrent gastric cancer, treated with first-line mFOLFOX-6  TTP particularly worse in HER2-positive patients without diffuse-type histology Outcome, mosHER2-positiveHER2-negativeP value Median TTP Median OS Kim JW, et al. Anticancer Res. 2012;32:

clinicaloptions.com/oncology Translational Research 2012 Challenges Determining HER2 Status in Gastric Cancer  HER2 testing in gastric cancer differs from breast cancer –More frequent HER2 heterogeneity (focal staining) –Incomplete membrane staining  Testing protocol –Initial testing with IHC –Retest IHC 2+ samples with FISH Rüschoff J, et al. Mod Pathol. 2012;25:

clinicaloptions.com/oncology Translational Research 2012 HER2 Positivity: IHC Score Surgical Specimen Staining Pattern Biopsy Specimen Staining Pattern HER2 Overexpr. Assessment 0 No reactivity or membranous reactivity in < 10% of tumor cells No reactivity or no membranous reactivity in any tumor cell Negative 1+ Faint or barely perceptible membranous reactivity in ≥ 10% of tumor cells; cells are reactive only in part of their membrane Tumor cell cluster with faint or barely perceptible membranous reactivity irrespective of % of tumor cells stained Negative 2+ Weak to moderate complete, basolateral or lateral membranous reactivity in ≥ 10% of tumor cells Tumor cell cluster with weak to moderate complete, basolateral or lateral membranous reactivity irrespective of % of tumor cells stained Equivocal 3+ Strong complete, basolateral or lateral membranous reactivity in ≥ 10% of tumor cells Tumor cell cluster with strong complete, basolateral or lateral membranous reactivity irrespective of % of tumor cells stained Positive Bang YJ, et al. Lancet. 2010;376: Bang YJ, et al. ASCO Abstract 4556.

clinicaloptions.com/oncology Translational Research 2012 Trastuzumab + Chemotherapy in Advanced HER2+ Gastric Cancer: ToGA  Rationale: a subpopulation of gastric cancers overexpress HER2  Primary endpoint: OS *Selected at investigator’s discretion: 5-FU 800 mg/m 2 /day infusional on Days 1-5 q3w x 6; capecitabine 1000 mg/m 2 BID on Days 1-14 q3w x 6. (n = 584) R Patients with advanced gastric cancer screened for HER2 status (N = 3803) Stratified by ECOG PS, advanced vs metastatic, gastric vs GEJ, measurable disease, capecitabine vs 5-FU Patients with HER2+ advanced gastric cancer (n = 810; 22% of successful screenings) 5-FU or Capecitabine* + Cisplatin 80 mg/m 2 q3w x 6 + Trastuzumab 6 mg/kg q3w until PD (8 mg/kg loading dose) (n = 294) 5-FU or Capecitabine* + Cisplatin 80 mg/m 2 q3w x 6 (n = 290) Bang YJ, et al. Lancet. 2010;376:

clinicaloptions.com/oncology Translational Research 2012 ToGA: HER2 Positivity Worldwide Bang YJ, et al. ASCO Abstract HER2 positivity in ToGA: 22.1% Asian/Pacific Europe South/Middle America Other Patients, % Australia (n = 61)Japan (n = 410) Korea (n = 687) China (n = 590)India (n = 118)Taiwan (n = 34) South Africa (n = 27) Russia (n = 459)Mexico (n = 67) Guatemala (n = 53) Panama (n = 40)Brazil (n = 97)Peru (n = 168)Costa Rica (n = 59)Denmark (n = 33)Italy ( n = 99)France (n = 78)UK (n = 132)Germany (n = 173)Portugal (n = 67)Spain (n = 122)Belgium (n = 14)Turkey (n = 58)Finland (n = 21)

clinicaloptions.com/oncology Translational Research 2012 ToGA: HER2 Positivity Histological Type Bang YJ, et al. ASCO Abstract SubtypeHER2 Positivity (%)P Value Histological typeIntestinal Diffuse Mixed <.001 LocalizationGEJ Gastric <.002 Histological subtype and tumor sublocalization are important factors for HER2 expression/amplification in gastric cancer

clinicaloptions.com/oncology Translational Research 2012 ToGA: HER2 Positivity GEJ vs Gastric Cancer Gastric cancer GEJ cancer Samples (%) BelgiumDenmarkFinlandFranceGermanyUKAustraliaPortugalMexicoItalyIndiaSpainPanamaCosta RicaChinaRussiaTurkeyPeruBrazilGuatemalaJapanKoreaTaiwanSouth Africa N = 2759; *P <.001Gastric CancerGEJ CancerTotal HER2 positive, % *21.3 Bang YJ, et al. ASCO Abstract 4556.

clinicaloptions.com/oncology Translational Research 2012 Events ToGA: Primary Endpoint OS Mos Pts at Risk, n Survival Probability FC + T FC HR % CI P Value.0046 Median OS Bang YJ, et al. Lancet. 2010;376:

clinicaloptions.com/oncology Translational Research 2012 ToGA: Efficacy Outcomes  Preplanned subgroup analysis indicated improved OS benefit with increasing HER2 expression by IHC  Exploratory analysis of IHC 2+/FISH+ and IHC 3+ cohort demonstrated a 4-mo increase in OS with trastuzumab –HR: 0.65 (95% CI: ) OutcomeChemotherapy + Trastuzumab (n = 294) Chemotherapy Alone (n = 290) HR (95% CI)P Value Median OS, mos ( ).0046 Median PFS, mos ( ).0002 ORR, %  CR  PR Bang YJ, et al. Lancet. 2010;376:

clinicaloptions.com/oncology Translational Research 2012 ToGA: OS by HER2 Status HER2 Status SubgroupMedian OS, Mos (CT + T vs CT Alone) HR* (95% CI) All patients (N = 584)13.8 vs ( ) Preplanned analysis  IHC 0/FISH+ (n = 61)10.6 vs ( )  IHC 1+/FISH+ (n = 70)8.7 vs ( )  IHC 2+/FISH+ (n = 159)12.3 vs ( )  IHC 3+/FISH+ (n = 256)17.9 vs ( )  IHC3+/FISH- (n = 15)17.5 vs ( ) Exploratory analysis  IHC 0 or 1+/FISH+ (n = 131)10.0 vs ( )  IHC 2+/FISH+ or IHC 3+ (n = 446)16.0 vs ( ) Bang YJ, et al. Lancet. 2010;376: *HR 1 favors chemotherapy alone.

clinicaloptions.com/oncology Translational Research 2012 Events Mos Pts at Risk, n Survival Probability FC + T FC HR % CI Median OS ToGA: OS in IHC 2+/FISH+ or IHC 3+ (Exploratory Analysis) Bang YJ, et al. Lancet. 2010;376:

clinicaloptions.com/oncology Translational Research 2012 ToGA: Select Toxicities Adverse Event, %Chemotherapy + Trastuzumab (n = 294) Chemotherapy Alone (n = 290) Grade 3/4 hematologic events  Neutropenia2730  Anemia1210 Grade 3/4 nonhematologic events  Diarrhea94  Nausea77 Cardiac events66  Grade 3/413 LVEF reduction of ≥ 10% to absolute value < 50%* 51 Bang YJ, et al. Lancet. 2010;376: *Chemotherapy plus trastuzumab: n = 237; chemotherapy alone: n = 187.

A Vision of Tomorrow’s Treatment Algorithm: Gastric Cancer

clinicaloptions.com/oncology Translational Research 2012 Other Potential Biomarkers in Gastric Cancer  MicroRNAs –Unique microRNAs associated with distinct prognostic and predictive values [1,2] –7-microRNA signature independent predictor of OS and RFS [3]  DNA methylation –Prognostic/predictive value [4]  CEA –Elevated levels correspond with worse prognosis and survival [5]  Cyclin D1 –Elevated levels correspond with worse prognosis [6] 1. Ueda T, et al. Lancet Oncol. 2010;11: Li X, et al. Gut. 2010;59: Brenner B, et al. World J Gastroenterol. 2011;17: Sapari NS, et al. PLoS One. 2012;7:e Mihmanli M, et al. Hepatogastroenterology. 2004;51: Gao P, et al. World J Gastroenterol. 2004;10:

clinicaloptions.com/oncology Translational Research 2012 Summary  KRAS mutation status critical determinant of benefit with EGFR-targeted antibody therapy –BRAF mutations potentially predictive of reduced benefit in non-first-line setting  Future biomarkers in CRC –PI3K, NRAS, EGFR ligand, DNA hypermethylation, mismatch repair, VEGF, CTC levels  HER2 expression confers worse prognosis in gastric cancer –HER2 expression challenging to assess and should be performed only by experienced pathologists  Future biomarkers in gastric cancer –MicroRNAs, DNA methylation, CEA, cyclin D1

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