Abstract

<div>Abstract<p><b>Purpose:</b> Expression of CBL, an ubiquitin ligase, is decreased in 60% of human pancreatic ductal adenocarcinomas (PDAC) and is associated with shorter overall survival. We sought to determine how low CBL directly contributes to clinically more aggressive PDAC.</p><p><b>Experimental Design:</b> Human PDACs were stained for CBL, pEGFR, and EGFR. CBL-low was modeled in PDAC cells (Panc-1, L3.6pl, and AsPC-1) via transient transfection (siRNA) or stable knockdown (shRNA). Cell viability and apoptosis were measured by MTT assays and FACS. Immunoblot and a phospho-receptor tyrosine kinase (pRTK) array were used to probe signal transduction. NOD-<i>scid</i>-<i>IL2Rγ<sup>null</sup></i> mice were subcutaneously implanted with PDAC or PDAC<sup>CBL-low</sup> cells on opposite flanks and treated with gemcitabine ± erlotinib for ≥4 weeks.</p><p><b>Results:</b> There was an inverse correlation between CBL and pEGFR protein expression in 12 of 15 tumors. <i>CBL</i> knockdown increased PDAC resistance to gemcitabine and 5-fluorouracil (5-FU) by upregulating pEGFR (Y1068), pERK, and pAKT. A pRTK array of PDAC<sup>CBL-low</sup> cells revealed additional activated tyrosine kinases but all to a much lower magnitude than EGFR. Increased chemoresistance from low CBL was abrogated by the EGFR inhibitor erlotinib both <i>in vitro</i> and <i>in vivo</i>. Erlotinib+gemcitabine–treated PDAC<sup>CBL-low</sup> cells exhibited greater apoptosis by cleaved PARP, caspase-3, and Annexin V/PI.</p><p><b>Conclusions:</b> Low CBL causes chemoresistance in PDAC via stress-induced EGFR activation that can be effectively abrogated by EGFR inhibition. These results suggest that dysregulation of ubiquitination is a key mechanism of EGFR hyperactivation in PDAC and that low CBL may define PDAC tumors likely to respond to erlotinib treatment. <i>Clin Cancer Res; 21(1); 157–65. ©2014 AACR</i>.</p></div>

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