Regulation of ERBB2 Receptor by t-DARPP Mediates Trastuzumab Resistance in Human Esophageal Adenocarcinoma
Menée sur 141 échantillons tumoraux prélevés sur des patients atteints d'un adénocarcinome de l'œsophage et à l'aide de xénogrefffes, cette étude évalue le rôle de t-DARPP dans la régulation de l'expression du récepteur ERBB2 et l'apparition d'une résistance au trastuzumab
Esophageal adenocarcinoma (EAC) is an aggressive malignancy with a poor outcome. Although targeting ERBB2 with trastuzumab is evaluated in clinical trials, the molecular mechanisms of trastuzumab resistance remain uncharacterized in EAC. The dopamine and cyclic AMP-regulated phosphoprotein of Mr 32,000 (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). Co-immunoprecipitation and Western blot analysis demonstrated 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.