1. Randomized phase III study of capecitabine, oxaliplatin and bevacizumab with or without cetuximab in advanced colorectal cancer CAIRO2 study of the Dutch Colorectal Cancer Group (DCCG) CJA Punt, J Tol, CJ Rodenburg, A Cats, GJ Creemers, JG Schrama, FLG Erdkamp, A Vos, L Mol, NF Antonini

2. Treatment of advanced colorectal cancer (CRC) Fluoropyrimidine-based chemotherapy plus the VEGF antibody bevacizumab is currently considered as the standard 1 st line treatment Cetuximab is a chimaeric monoclonal antibody against the epidermal growth factor receptor (EGFR) which has shown efficacy as a single agent and in combination with chemotherapy

3. Background VEGF and EGFR share common downstream signaling pathways, and preclinical models have shown additive effects for VEGF and EGFR inhibition In irinotecan-resistant CRC the combination of irinotecan, cetuximab, and bevacizumab (BOND2 study) 1 was feasible and suggested greater efficacy compared to irinotecan + cetuximab (BOND study) 2 Therefore, targeting both VEGF and EGFR in CRC appears a promising strategy and warrants evaluation in a prospective study 1 Saltz et al. J Clin Oncol 2007 2 Cunningham NEngl J Med 2004

4. Study design CAIRO2 Arm A Arm B Randomization Capecitabine Oxaliplatin Bevacizumab Capecitabine Oxaliplatin Bevacizumab Cetuximab

5. Endpoints Primary endpoint progression-free survival Secondary endpoints overall survival, response rate, toxicity, translational research

6. Statistical design Study was designed to detect a difference in median progression-free survival of 3 months (11m  14m) (HR 0.79), power 80%,  =0.05, 2- tailed test Stratification parameters - prior adjuvant chemotherapy - serum LDH - number of affected organs - institution

7. Histologically proven colorectal cancer Advanced disease not amenable to curative surgery Measurable disease parameters No previous systemic treatment for advanced disease Previous adjuvant chemotherapy should be completed  6 months prior to randomization Age  18 years WHO performance score 0-1 Adequate hepatic, bone marrow, and renal function No therapeutic dose of anticoagulant drugs No significant cardiovascular or other disease Main inclusion criteria

8. Dose and schedule all cycles given 3-weekly Arm A Cycle 1-6: oxaliplatin 130 mg/m² day 1 capecitabine 1000 mg/m² b.i.d. day 1-14 bevacizumab 7.5 mg/kg day 1 Cycle ≥ 7: capecitabine 1250 mg/m² b.i.d. day 1-14 bevacizumab 7.5 mg/kg day 1 Arm B oxaliplatin, capecitabine, bevacizumab: as in arm A cetuximab weekly 250 mg/m² (400 mg/m² 1 st dose)

9. Evaluation of response Evaluation of tumor response every 3 cycles (RECIST) Evaluation of toxicity prior to each cycle (NCI-CTC criteria, version 3.0) Central review was performed of all patient files when death occurred ≤ 30 days of the last administration of study drugs and which was accompanied by any other event than disease progression, irrespective of the causality reported for this event All serious adverse events and results of central review were submitted to the IDMC

10. Accrual Participation of 79 Dutch hospitals 755 patients were randomized between June 2005 and December 2006 736 patients were eligible 731 patients received ≥ 1 treatment cycle Median duration of follow-up 18.7 months

11. Arm A Arm B n = 368 n = 368 Age, median (range)62 (27-83)62 (33-80) Gender * male female 56% 44% 64% 36% Prior adjuvant treatment15% Serum LDHnormal abnormal 57% 43% 56% 44% Number of organs affected 1 > 1 45% 55% 44% 56% WHO performance status 059%66% * p = 0.035 Baseline characteristics

12. Arm A Arm B p value n = 368 n = 368 Median PFS (months) (HR; 95% CI) 10.7 (9.7-12.5) 9.6 (8.5-10.7) 0.018 (1.21;1.03-1.45) Median OS (months) (HR; 95% CI) 20.4 (18.1-26.1) 20.3 (17.9-21.6) 0.21 (1.15;0.93-1.43) Response rate (CR + PR) 44% 0.88 Disease control rate (CR + PR + SD) 83%81%0.39 Efficacy results Results were confirmed in the subgroup of patients with EGFR+ tumors

13. Arm A (without cetuximab) 10.7 months (9.7-12.5) Arm B (with cetuximab) 9.6 months (8.5-10.7) Progression-free survival Hazard ratio for progression 1.21 p value 0.018

14. Overall survival Arm A (without cetuximab) 20.4 months (18.1-26.1) Arm B (with cetuximab) 20.3 months (17.9-21.6) Hazard ratio for survival 1.15 p value 0.21

15. Arm A n = 366 Arm B n = 365 p value All grade 3-472%82%0.0013 Skin toxicity excluded (except HFS) 72%75%0.37 Hematological toxicity2% 0.99 Diarrhea19%26%0.026 Vomiting9%6%0.20 Hand-foot syndrome19% 0.98 Neurotoxicity10%8%0.37 GI perforation0.3%1.4%0.10 Venous thromboembolic events7%8%0.66 Toxicity (grade 3-4)

16. Arm A n = 366 Arm B n = 365 All grade acneiform skin reactions4%84%* Grade 3 acneiform skin reactions0.5%25%* All grade nail changes13%32%* Grade 3 nail changes0.3%4%* * p<0.001 Skin toxicity associated with cetuximab

17. Arm A n = 368 Arm B n = 368 Deaths ≤ 30 days of last treatment (any cause) 18 (5%) Treatment related4 (1%)2 (0.5%) - GI perforation31 - respiratory insufficiency1 - neutropenic fever1 60-day mortality7 (1.9%)9 (2.4%) Mortality

18. Arm A n = 366 Arm B n = 365 p value Number of cycles (range) median10 (1-39)9 (1-40)0.02 mean12.010.6 Reasons for treatment discontinuation - disease progression including death53%50%0.32 - toxicity27%32%0.22 - secondary metastasectomy5% - patient refusal7%6% - other8%7% Treatment characteristics

19. Arm B (with cetuximab): skin grade 3 (n=104) Arm B (with cetuximab): skin grade 0-1 (n=109) Arm B (with cetuximab): skin grade 2 (n=152) PFS according to skin toxicity Arm B grade 0-1 vs 2 vs 3: p ≤ 0.01

20. Arm B (with cetuximab): skin grade 0-1 (n=109) Arm B (with cetuximab): skin grade 2 (n=152) Arm B (with cetuximab): skin grade 3 (n=104) Arm A (without cetuximab): overall (n=366) PFS according to skin toxicity Arm A vs arm B grade 0-1: p < 0.0001

21. Wildtype n = 305 (61%) Mutation n = 196 (39%) Arm A152 (50%)103 (53%) Arm B153 (50%)93 (47%) Genotyping by Q-PCR - based assay No difference in baseline characteristics between patients with KRAS wildtype and mutation (except higher serum LDH in wildtype) No correlation between KRAS status and cetuximab-related skin toxicity KRAS genotyping (n=501)

22. Wildtype n = 305 (61%) Mutation n = 196 (39%) p value Arm A152 (50%)103 (53%) Arm B153 (50%)93 (47%) Median PFS (months) Arm A10.712.50.92 Arm B10.58.60.47 p value0.100.043 KRAS genotyping (n=501)

23. KRAS and PFS Arm A (without cetuximab); KRAS mutant Arm B (with cetuximab); KRAS mutant Arm A (without cetuximab); KRAS wildtype Arm B (with cetuximab); KRAS wildtype

24. Wildtype n = 305 (61%) Mutation n = 196 (39%) p value Arm A152 (50%)103 (53%) Arm B153 (50%)93 (47%) Median OS (months) Arm A23.024.90.90 Arm B22.219.10.52 p value0.490.35 KRAS genotyping (n=501)

25. Conclusions - I The addition of cetuximab to capecitabine, oxaliplatin and bevacizumab results in a significantly decreased progression-free survival, without affecting overall survival The addition of cetuximab to chemotherapy and bevacizumab results in a significant increase of skin toxicity and diarrhea, however the toxicity is acceptable in both treatment arms The grade of cetuximab-related skin toxicity significantly correlates with PFS

26. The results of cetuximab are not significantly influenced by KRAS status In patients with KRAS mutation the addition of cetuximab to chemotherapy and bevacizumab results in a significant decrease in PFS Toxicity as a reason for discontinuation of treatment does not significantly differ between treatment arms, therefore a negative interaction between anti-VEGF and anti-EGFR antibodies cannot be excluded Conclusions - II

27. DCCG CAIRO2 study - acknowledgements Investigators: C. Smorenburg, Alkmaar; R.Hoekstra, Almelo; C.Rodenburg, Amersfoort; J.van der Hoeven, Amstelveen; A.Cats, M.Geenen, C. van Groeningen, D.Richel, B.de Valk, N.Weijl, Amsterdam; J.Douma, Arnhem; P.Nieboer,Assen; F.Valster, Bergen op Zoom; R.Rietbroek, Beverwijk; A.Ten Tije, Blaricum; O.Loosveld, Breda; D.Kehrer, Capelle a/d IJssel; M.Bos, Delft; Z.Erjavec,Delfzijl; H.Sinnige, C.Knibbeler, Den Bosch; W.Van Deijk, F.Jeurissen, H.Sleeboom Den Haag; H.de Jong, Den Helder; A.Imholz, Deventer; E.Muller, Doetinchem; J. vanden Bosch,Dordrecht; S.Hovenga, Drachten; E.Balk, Ede; G.Creemers, M.Dercksen, Eindhoven; M.Legdeur, Enschede; A.Smals, Geldrop; H.van Halteren, Goes; M. van Hennik, Gorinchem; A.van der Torren, Gouda; G.Hospers, R.de Jong, Groningen; G.de Klerk, Haarlem; P.Zoon, Harderwijk; J.Wals, Heerlen; V.Derleyn, Helmond; H.Dankbaar, Hengelo; S.Luyckx, Hilversum; C. de Swart, Hoofddorp; J.Haasjes, Hoogeveen; W.Meijer, Hoorn; M.Polee, Leeuwarden;M.Tesselaar, Leiden; H.Oosterkamp,Leidschendam; J.Bollen, Lelystad; R.Jansen, Maastricht; P. van der Velden, Middelharnis; P.Slee, Nieuwegein; C.Punt, C.Mandigers, Nijmegen; A.Vos, Oss; M.den Boer, Roermond; D. de Gooyer, Roosendaal; A.Planting, A.vander Gaast, J.Pegels, T.Kok, F.de Jongh, Rotterdam; J.Braun, Schiedam; F.Erdkamp, Sittard; G.Veldhuis, Sneek; A.van Reisen, Terneuzen; C.Kruijtzer, Tiel; H.Roerdink, J. van Riel, Tilburg; D.ten Bokkel Huinink, E.Voest, Utrecht; M. van Diemen, Veghel; G.Vreugdenhil, Veldhoven; M.Werter, Venlo; L.Kerkhofs, Vlissingen; P.Schiphorst, Winterswijk; A.van Bochove, Zaandam; A.Ogilvie, Zoetermeer; A.Honkoop, Zwolle Pathology: J.van Krieken, J.Dijkstra, E.Börger, Nijmegen Statisticians: N.Antonini, O.Dalesio, Amsterdam Central Datamanagement: L.Mol, F.van Leeuwen, IKO Nijmegen, Independent Data Monitoring Committee: P. De Mulder †, D.Sleijfer, G.Stoter, Supported by Dutch Cancer Foundation, and unrestricted scientific grants from Merck Serono, Roche, Sanofi-Aventis