Regulation of ERBB2 receptor by t-DARPP mediates trastuzumab resistance in human esophageal adenocarcinoma.

Hong J, Katsha A, Lu P, Shyr Y, Belkhiri A, El-Rifai W
Cancer Res. 2012 72 (17): 4504-14

PMID: 22745369 · PMCID: PMC3432752 · DOI:10.1158/0008-5472.CAN-12-1119

Esophageal adenocarcinoma (EAC) is an aggressive malignancy with a poor outcome. Although targeting ERBB2 with trastuzumab has been evaluated in clinical trials, the molecular mechanisms of trastuzumab resistance remain uncharacterized in EAC. The dopamine and cyclic AMP-regulated phosphoprotein of MR 32000 (DARPP-32), also known as PPP1R1B, is located together with ERBB2 at the 17q12-q21 amplicon. We evaluated the expression of a transcript variant of DARPP-32 (t-DARPP) and ERBB2 in 141 primary tumors and investigated the role of t-DARPP in trastuzumab resistance using OE19 and OE33 EAC cell models. Overexpression of t-DARPP mRNA was detected in two-thirds of tumors with a correlation between ERBB2 and t-DARPP overexpression levels (r = 0.58, P = 0.003). Cell viability and clonogenic survival assays showed that t-DARPP increased survival by 40% in response to trastuzumab (P < 0.01). The Annexin-V staining and Western blot analysis indicated that t-DARPP effectively abrogated trastuzumab-induced apoptosis, inhibited cleavage of caspase-3, and blocked trastuzumab-induced dephosphorylation of ERBB2 and AKT proteins. The knockdown of endogenous t-DARPP reversed these effects and sensitized cells to trastuzumab (P < 0.01). The cycloheximide-based protein degradation analysis indicated that t-DARPP extended the half-life of ERBB2, explaining the increase in the basal levels of ERBB2, p-ERBB2(Y1248), and p-AKT(S473). Coimmunoprecipitation and Western blot analysis showed that t-DARPP associated with ERBB2 in a protein complex, and interfered with trastuzumab binding to the ERBB2 receptor. Using EAC-xenografted mouse model, t-DARPP enhanced tumor growth and rendered tumors unresponsive to trastuzumab. This study establishes t-DARPP as a mediator of trastuzumab resistance and underscores its potential importance in clinical trials of EAC.

©2012 AACR.

MeSH Terms (25)

Adenocarcinoma Animals Antibodies, Monoclonal, Humanized Antineoplastic Agents Apoptosis Caspase 3 Cell Line, Tumor Cell Survival Dopamine and cAMP-Regulated Phosphoprotein 32 Drug Resistance, Neoplasm Enzyme Activation Esophageal Neoplasms Female Gene Expression Regulation, Neoplastic Gene Silencing Humans Mice Mice, Nude Protein Binding Protein Stability Receptor, ErbB-2 Signal Transduction Trastuzumab Tumor Burden Xenograft Model Antitumor Assays

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