Abstract 3134: Crizotinib inhibits tumor growth and metabolic inactivation of gemcitabine in new patient derived orthotopic pancreatic tumors with c-Met overexpression: A step toward personalized treatment in pancreatic cancer
Authors: Avan Amir , Caretti Viola , Funel Niccola , Galvani Elena , Maftouh Mina , Honeywell Richard J. , Lagerweij Tonny , Campani Daniela , Verheul Henk M. , Schuurhuis Gerrit Jan , Boggi Ugo , Peters Godefridus J. , Würdinger Thomas , Giovannetti Elisa 
Published: May 2015

Abstract
Pancreatic ductal adenocarcinoma (PDAC) remains a major unsolved health problem.Most drugs that pass preclinical tests fail in patient, emphasizing the need of improved preclinical models. This study aimed to (1) develop orthotopic models employing primary PDAC cells genetically engineered to express Firefly- and Gaussia- luciferase (Fluc/Gluc), enabling monitoring of tumor growth, and (2) evaluate the therapeutic potential of c-Met inhibitors.Four primary PDAC-cultures were successfully transduced with lentiviral vectors containing Fluc and Gluc, and then injected orthotopically into pancreas. The bioluminescence intensities increased overtime in all the models. Histopathological and immunohistochemical analyses were carried out on paraffin-embedded slices of pancreas, as well as on metastatic lesions in liver, lungs and lymph nodes, showing that the xenografts recapitulated the phenotype of human PDAC. Genetic characteristics of the xenografts were similar to their originator tumor/cells, as assessed by array-CGH. Genomic and immunohistochemical analysis revealed c-Met increased copy-number and overexpression in one of our models (PDAC3). Therefore we explored several c-Met inhibitors in vitro and in vivo.
Crizotinib emerged as the most active inhibitor of cell growth, with an IC50 of 0.5 μM in the PDAC3. Median drug-effect analysis revealed strong synergistic interaction of crizotinib with gemcitabine. The mice receiving this combination had significantly lower BLI signals compared to mice receiving either gemcitabine or crizotinib alone (e.g., at day 30, Fluc was 7x108 vs. 1x108 photon/second/cm2 in gemcitabine and combination group, respectively), which was reflected in a significant longer survival of mice treated with combination. Importantly, we observed a significant increase of gemcitabine in the combination group with respect to the mice treated with gemcitabine alone (6.3 vs. 2.5nm/mg, as measured by LC/MS-MS). Correspondingly, crizotinib reduced the activity of cytidine deaminase both in tissue specimens, and blood samples. Since intact c-Met signaling is a critical factor in the protection against excessive generation of endogenous ROS, we demonstrated that ROS induction caused c-Met inhibition was responsible for CDA degradation after exposure to crizotinib, resulting in the increased stabilization of gemcitabine in the PDAC-3 cells. Our orthotopic imaging models showed genetic, histopathological and metastatic features similar to their originator tumors and enabled the identification of c-Met as a potential therapeutic target. Crizotinib and gemcitabine are a promising drug combination against this target, acting synergistically also through the increase of drug delivery by CDA modulation, and warranting further investigation for the treatment of PDAC patients.
Citation Format: Amir Avan, Viola Caretti, Niccola Funel, Elena Galvani, Mina Maftouh, Richard J. Honeywell, Tonny Lagerweij, Daniela Campani, Henk M. Verheul, Gerrit Jan Schuurhuis, Ugo Boggi, Godefridus J. Peters, Thomas Würdinger, Elisa Giovannetti. Crizotinib inhibits tumor growth and metabolic inactivation of gemcitabine in new patient derived orthotopic pancreatic tumors with c-Met overexpression: A step toward personalized treatment in pancreatic cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3134. doi:10.1158/1538-7445.AM2014-3134