MCW Regional Cancer Therapy Program IHC-Microarray Analysis of Patients With Peritoneal Metastases (PM) of Appendiceal or Colorectal Origin Green D.E., Hwang M., Jayakrishnan T.T., Pappas S.G., Gamblin T.C., Turaga K.K. MCW Regional Cancer Therapy Program

We have no relevant financial disclosures

Background Appendiceal vs. Colorectal Cancer Genomically different via microarray, phenotypic clusters predict prognosis -High Risk Colorectal, no survival @ 5 years -Low Risk Appendiceal, 70% survival @ 116 mo -High Risk Appendiceal, 25% survival @ 116 mo Levine 2012

Background Colorectal Cancer KRAS mutated in 34.5% Median survival 27.7 months BRAF mutated in 6.5% Median survival 11 months Yokota, 2011

Background Mutational analysis of patients with appendiceal adenocarcinoma (n=80) BRAF 8% KRAS 45% EGFR 24% PIK3CA 19% Lilleberg 2009

Background: IHC and Clinical Benefit Predictive Biomarker Associated Treatments * BRAF vemurafenib (cetuximab, panitumumab) KRAS (wt) cetuximab, panitumumab, sorafenib (erlotinib, gefitinib) ERCC 1 cisplatin, carboplatin, oxaliplatin PTEN erlotinib, gefitinib, cetuximab, panitumumab, trastuzumumab SPARC nab-paclitaxel TOP2A doxorubicin, liposomal-doxorubicin, epirubicin TOPO1 irinotecan, topotecan TS capecitabine, pemetrexed, fluorouracil * Expression of biomarker determines if chemotherapy is associated with response

Hypothesis We hypothesized that it is feasible to assess the immunohistochemistry and mutational analysis of patients with peritoneal carcinomatosis of appendiceal and colorectal primaries using a commercially available molecular profiling service.

Methods Retrospective chart review from clinical data April 2008 -June 2012 Inclusion: Appendiceal or colorectal primary, malignant PM, sufficient tissue biopsy 16 patients Exclusion: Insufficient tissue Molecular Profiling Caris Target Now: Phoenix, Arizona/ Irving, Texas Stratified data PCI score, histology, treatment

Methods 23 Biomarkers examined with immunohistochemistry ERCC1, RRM1, TS, COX-2, and TOP2A 88 Biomarkers examined with microarray analysis [Illumina Method]

Demographics n=16 Age (years) 51 (42-60) Female Gender 11 (69%) T4 Neoadjuvant Chemotherapy PCI Score 17 (13-26) CRS + HIPEC 8 (50%)

Comparison of General Characteristics of Colon and Appendix Cancer Patients Colon (n=11) Appendix (n=5) P-value Age (years) 51 (38-72) 51 (40-56) 0.42 Female Gender 8 (73%) 3 (60%) 1.00 T4 Neoadjuvant Chemotherapy 9 (82%) 2 (40%) 0.24 PCI Score 17 (8-27) 17 (12-20) 0.82 CRS + HIPEC 4 (36%) 4 (80%) 0.28 Median Survival (months) 30 60 0.50 9 (82%) 2 (40%)

Results Immunohistochemistry performed on 16 patients Sufficient tissue for microarray analysis on 7 patients (44%)

Immunohistochemistry Biomarker (IHC) * Colon (N=11) Appendix (N=5) P-value COX-2 80% 100% 0.52 ERCC1 1.00 PTEN SPARC 36% 20% TOP2A 0.31 TOPO1 TS 82% *Significance as reported by the commercial test based on percent staining and staining intensity threshold

Mutational Analysis 10% 20% 20% 10% 40% 20% 40% 40%

Survival Analysis Biomarker Hazards Ratio (95% CI) P-value COX-2 0.06 (0.005-0.71) 0.02 * RRM1 0.26 (0.04-1.57) 0.14 SPARC (Poly) 0.15 (0.02-1.3) 0.08 TS 1.00 * Survival not significant when adjusted for multiple comparisons

Targeted Therapy & Survival Biomarker Chemotherapy Number Median Survival (months) TS + 5FU/LV* 9 30 KRAS WT Cetuximab/panitumimab 4 ERCC1 + Oxaliplatin 11 60 TOP2A + Irinotecan 10 * 5 FU/LV = Fluorouracil/Leucovorin

Conclusions IHC and mutational analysis using commercially available molecular profiling testing is feasible for patients undergoing surgery in setting of PM 40% of patients with colorectal primaries and PM have BRAF mutations (KRAS wt) Possible survival benefit with COX-2 expression

Limitations Small sample size (n=16) Limited microarray analyses: done on 44% patients (n=7)

Future Directions Prospective evaluation of impact of microarray analysis on response to systemic chemotherapy using multi-institutional data.

References Levine E.A., et al. Gene expression profiling of peritoneal metastases from appendiceal and colon cancer demonstrates unique biologic signatures and predicts patient outcomes. J Am Coll Surg. 2012;214(4):599-606; discussion 606-7. Lilleberg SL, et al (2009, January). Use of mutation analysis of Pseudomyxoma peritonei and appendiceal adenocarcinomas to identify somatic variants within MAPK, AKT and WNT signaling pathways: Potential markers for focused treatment strategies. Poster session presented at Gastrointestinal Cancers Symposium in San Francisco, CA. Yokota T, et al. BRAF mutation is a powerful prognostic factor in advanced and recurrent colorectal cancer. Br J Cancer. 2011;104(5):856-862.

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