Strategies to overcome resistance in NSCLC with driver mutations Federico Cappuzzo Istituto Toscano Tumori Ospedale Civile Livorno-Italy

First-line therapy for metastatic NSCLC in 2012 Stratification for EGFR, ALK and histology EGFR mutated ALK+ EGFR WT non-squamous EGFR WT squamous EGFR-TKI Crizotinib Platinum + pemetrexed +/- bevacizumab Platinum-based doublet

Mut+ NSCLC: EGFR-TKI Acquired Resistance Malgrado gli TKIs siano la terapia di prima scelta in presenza di mutazioni di EGFR, nessun paziente viene guarito e tutti inevitablmente recidivano e muoiono. Questo è quello che tipicamente avviene in un paziente con mutazione di EGFR. Inizialmente si verifica una rapida regressione, sino al 90% dei casi, e dopo mediamente 9 mesi la malattia progredisce. In questi pazienti si pone il problema delle terapie successive e della sospensione del trattamento con TKIs. Si pone quindi il problema deela “disease flare”, ossia ……….. Baseline Tumor regression (RR up to 90%) Progression (median 9 months) Disease Flare: Hospitalization and/or death attributable to disease progression after discontinuation of gefitinib or erlotinib and before initiation of study drug

Risk of disease flare in EGFR mut+ NSCLC with acquired resistance: Chaft J et al. (O 19.05) Characteristic: Total patients=61 N (% or range) Male sex – N (%) 13 (21) Age at diagnosis (years) Median (range) 58 (26-78) EGFR mutation – N (%) Exon 19 deletion Exon 18 G719A Exon 21 L858R 41 (67) 1 (2) 19 (31) Time on gefitinib or erlotinib (months) 19 (7-78) Age in years - Median (Range) 61 (27-80) Karnofsky Performance Status (%) 90% 80% 70% 37 (61) 11 (18) Il rischio di disease flare è significativamente + alto in coloro che hanno avuto un più basso PFS (9 verso 15 mesi, p=0.002) Questi dati chiaramente indicano due cose: Se si dispone di una terapia ulteriore il tempo di sospensione deve essere + breve possibile Se no si hanno terapie ulteriori è preferibile continuare con un inibitore anche in presenza di PD. Infatti ……dati Reily 14 of 61 patients (23%, 95% CI 14-35%) had a disease flare (hospitalization or death) Flare & no flare group – same 30 day pretrial hospitalization rate Median time from last TKI to flare was 8 days (range 3-21 days) 3 patients went on to trial treatment

Changes in Tumor Diameter (RECIST) After Discontinuation and Re-introduction of EGFR TKI 50% 20% Change from baseline 0% -30% EGFR TKI stop re-start 3 weeks 3 weeks Riely et al, CCR 2007

Mechanisms responsible for EGFR-TKI resistance Sequist et al, Science Transl Med 2011

EGFR-TKI resistance A B

T790M Mutation causes drug resistance by increasing affinity for ATP Yun PNAS 2008

T790M mutations in EGFR-TKI naive NSCLC Present in up to 50% of NSCLC with EGFR-TKI acquired resistance Rare event in EGFR-TKI naive NSCLC (<3%) using low sensitive methods Detected in up to 40% of EGFR-TKI naive patients using high sensitive methods

Presence of T790M mutation predicts poor outcome to EGFR-TKI Detected using high sensitive methods Su et al. JCO 2012; Rosell et al. Clin Cancer Res 2011;

T790M mutation and acquired resistance to gefitinib therapy Irreversible EGFR-TKI are still effective Kobayashi et al. NEJM, 352, 786-792, 2005

Afatinib: Dual irreversible EGFR-HER2 inhibitor Orally bioavailable, small molecule tyrosine kinase inhibitor (TKI) Designed to irreversibly bind to the ATP binding pocket of EGFR and HER2 Highly specific for EGFR and HER2 EGFR IC50: 0.50 nM HER2 IC50: 14 nM Afatinib EGFR or HER2 ATP binding pocket +

Afatinib: active against resistance mutation BIBW2992 but not erlotinib is active against cells expressing T790M EGFR mutation: NCI-H1975 Li et al. Oncogene. 2008;27:4702–4711

Afatinib + cetuximab as the best option in presence of EGFR T790M mutation Regales et al. JCI 2009

Afatinib + cetuximab for metastatic NSCLC: Study Design Phase Ib, open-label, multicenter trial in the US and The Netherlands Stop erlotinib/ gefitinib for ≥72 hours3 Disease progression2 NSCLC with EGFR mutation1 OR SD 6 months with erlotinib/gefitinib Partial or complete response to erlotinib/gefitinib MTD cohort expanded up to 80 EGFR mutation-positive patients4: 40 T790M+ and 40 T790M– Dose escalation schema 3–6 patients per cohort Afatinib p.o. daily + escalating doses of i.v. cetuximab q 2 weeks Dose levels starting at: afatinib 40 mg + cetuximab 250 mg/m2 Predefined maximum dose: afatinib 40 mg + cetuximab 500 mg/m2 This was a Phase Ib, open-label, multicenter trial in the US and the Netherlands Protocol-defined dose reductions were as follows: First reduction • Decrease afatinib 40 → 30 mg and cetuximab 500 → 450 mg/m2 Second reduction • Continue afatinib 30 mg, decrease cetuximab 450 → 400 mg/m2 1EGFR G719X, exon 19 deletion, L858R, L861Q; 2Progression of disease (Response Evaluation Criteria in Solid Tumors v1.1) on continuous treatment with erlotinib or gefitinib within the last 30 days; 3Amended from original 14-day interval; 4Acquisition of tumor tissue after the emergence of acquired resistance was mandated. i.v.=intravenous; MTD=maximum tolerated dose; NSCLC=non-small cell lung cancer; SD=stable disease.

Tumor Regression by T790M Mutation Status at Recommended Dose 39 patients with proven EGFR T790M mutation: confirmed RR=31%

Acquired resistance to EGFR-TKIs Acquired drug resistance is almost inevitable (~10 months) About 30% of resistant mechanisms are unknown. Mitsudomi, et al. Cancer Sci., 2007

Resistant mechanisms in 33 tumors from 6 patients with EGFR-TKI acquired resistance Number of Tumors MET gene copy numbers (folds) Tumors with T790M Tumors without T790M <2.0 2.0 - 4.0 < 4.0 93% 8% 80% Suda et al. Clin Cancer Res 2010

Inhibition of both EGFR and MET is necessary for growth inhibition of HCC827 GR cells 125 Gefitinib 100 PHA665752 Gefitinib/PHA665752 75 % of control 50 25 0.01 0.1 1 10 Drug Concentration ( μ M) Irreversible EGFR inhibitors have no effect on HCC827 GR MET shRNA restores sensitivity to gefitinib Engelman et al. Science 2007

EML4-ALK fusion oncogene in NSCLC 3–7% of patients with NSCLC have an EML4-ALK gene fusion1 detection test available mainly seen in adenocarcinomas (mutually exclusive with EGFR mutations)2 phase I/II trial of crizotinib, oral c-MET and ALK inhibitor in selected patients: DCR = 70%3 further potential for personalising therapy in NSCLC 1. Koivunen, et al. Clin Cancer Res 2008 2. Shaw, et al. ASCO 2009; 3. Bang, et al. ASCO 2010

ALK secondary mutations and crizotinib resistance Sasaki et al. Cancer Res 2011

New ALK inhibitors TAE684 and AP26113 overcome crizotinib resistance in H3122 CR cell line Ci sono nuovi inibitori attivi sulle mutazioni secondarie di ALK ma altre strategie sembrano offrire risultati molto promettenti come la combinazione con anti-EGFR o gli inibitori di HSP90 Katayama et al. PNAS 2011

Cell lines with ALK secondary mutation and ALK and EGFR co-dependency In alcuni modelli sperimentali, linee cellulari con mutazione secondaria di ALK sono risultati dipendenti da ALK e anche da EGFR. Questo ha risvolti clinici interessanti in quanto offre la possibilità di ripristinare la sensibilità tumorale utilizzando una combinazione di anti-ALK e anti-EGFR. Sasaki et al. Cancer Res 2011

ALK amplification or ALK FISH loss as mechanisms of crizotinib resistance Katayama et al. PNAS 2011, Katayama et al Science Transl Med 2012, Doebele et al. Clin Cancer Res 2012

Several mechanisms responsible for crizotinib resistance: clinical implications L’esistenza di multipli meccanismi responsabili di resistenza a crizotinib ha delle importanti conseguenze cliniche in quanto rende necessario identificare nel singlo paziente l’evento che ha causato la resistenza e sulla base di questo pianificare il programma di trattamento. Inoltre la presenza nello stesso paziente di di eventi driver differenti general la domanda se tali eventi sono presenti nella stessa cellula tumorale o in cellule diverse. Nel modello uno viene mostratta la prima ipotesi, ovvero nella stessa cellula sono presenti due drivers differenti: questo è quanto sappiamo succede nel NSCLC con mutazione secondaria MET amplification. Nel modello due si ammette che più drivers siano presenti nello stesso paziente ma in popolazioni cellulari differenti. Questo è il caso almeno di alcuni pazienti ALK positivi dove abbiamo sia la traslocazione ALK che la mutazione di EGFR o di KRAS. Doebele et al. Clin Cancer Res 2012

Conclusions Different mechanisms are responsible for acquired resistance to novel targeted therapies So far no proven efficacy of irreversible EGFR-TKIs in NSCLC with acquired resistance to reversible agents No clinically available strategies for crizotinib resistant patients New drugs and new strategies are under investigation