A CGA/EGFR/GATA2 positive feedback circuit confers chemoresistance in gastric cancer
Tianyu Cao, Yuanyuan Lu, Qi Wang, Hongqiang Qin, Hongwei Li, Hao Guo, Minghui Ge, Sarah E. Glass, Bhuminder Singh, Wenyao Zhang, Jiaqiang Dong, Feng Du, Airong Qian, Ye Tian, Xin Wang, Cunxi Li, Kaichun Wu, Daiming Fan, Yongzhan Nie, Robert J. Coffey, Xiaodi Zhao
Tianyu Cao, Yuanyuan Lu, Qi Wang, Hongqiang Qin, Hongwei Li, Hao Guo, Minghui Ge, Sarah E. Glass, Bhuminder Singh, Wenyao Zhang, Jiaqiang Dong, Feng Du, Airong Qian, Ye Tian, Xin Wang, Cunxi Li, Kaichun Wu, Daiming Fan, Yongzhan Nie, Robert J. Coffey, Xiaodi Zhao
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Research Article Gastroenterology Oncology

A CGA/EGFR/GATA2 positive feedback circuit confers chemoresistance in gastric cancer

Abstract

De novo and acquired resistance are major impediments to the efficacy of conventional and targeted cancer therapy. In unselected gastric cancer (GC) patients with advanced disease, trials combining chemotherapy and an anti-EGFR monoclonal antibody have been largely unsuccessful. In an effort to identify biomarkers of resistance so as to better select patients for such trials, we screened the secretome of chemotherapy-treated human GC cell lines. We found that levels of CGA, the α-subunit of glycoprotein hormones, were markedly increased in the conditioned media of chemoresistant GC cells, and CGA immunoreactivity was enhanced in GC tissues that progressed on chemotherapy. CGA levels in plasma increased in GC patients who received chemotherapy, and this increase was correlated with reduced responsiveness to chemotherapy and poor survival. Mechanistically, secreted CGA was found to bind to EGFR and activate EGFR signaling, thereby conferring a survival advantage to GC cells. N-glycosylation of CGA at Asn52 and Asn78 is required for its stability, secretion, and interaction with EGFR. GATA2 was found to activate CGA transcription, whose increase, in turn, induced the expression and phosphorylation of GATA2 in an EGFR-dependent manner, forming a positive feedback circuit that was initiated by GATA2 autoregulation upon sublethal exposure to chemotherapy. Based on this circuit, combination strategies involving anti-EGFR therapies or targeting CGA with microRNAs (miR-708-3p and miR-761) restored chemotherapy sensitivity. These findings identify a clinically actionable CGA/EGFR/GATA2 circuit and highlight CGA as a predictive biomarker and therapeutic target in chemoresistant GC.

Authors

Tianyu Cao, Yuanyuan Lu, Qi Wang, Hongqiang Qin, Hongwei Li, Hao Guo, Minghui Ge, Sarah E. Glass, Bhuminder Singh, Wenyao Zhang, Jiaqiang Dong, Feng Du, Airong Qian, Ye Tian, Xin Wang, Cunxi Li, Kaichun Wu, Daiming Fan, Yongzhan Nie, Robert J. Coffey, Xiaodi Zhao

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Figure 5

Reciprocal positive regulation between GATA2 and CGA/EGFR signaling.

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Reciprocal positive regulation between GATA2 and CGA/EGFR signaling. (A ...
(A and B) Correlations between CGA and TFs in KM plotter (A) and CCLE (B) databases. Data evaluated using Pearson’s correlation coefficient. (C) Kaplan-Meier analysis of correlation between GATA2 expression and overall survival of GC patients using the KM plotter database. (D) RT-qPCR and immunoblotting of GATA2 in MDR and SGC7901 cells. (E and F) Immunoblotting of GATA2 and CGA in MDR cells transfected with 2 independent siRNAs against GATA2 (siGATA2) or a control siRNA (siCtrl) and in SGC7901 cells transfected with a GATA2 expression vector or empty vector (E). Viability was measured in the indicated cells treated with chemotherapy (F). (G) Left: Diagram of consecutive deletion and mutation constructs spanning the CGA promoter. GBE mutations shown in red boxes. Right: Luciferase reporter driven by the WT, deletion, or mutant (MUT) promoter was transfected into SGC7901ADR cells. Luciferase activity was measured with or without GATA2 cotransfection. (H) ChIP with anti-GATA2 antibody in SGC7901 cells with or without GATA2 transfection. (I–K) Immunoblotting of CGA, GATA2, EGFR, and p-EGFR in indicated cells. (L) GATA2 and CGA expression in SGC7901 and NCI-N87 cells treated with low concentrations of fluorouracil (1 μg/mL) for the indicated times. (M) ChIP with anti-GATA2 antibody in SGC7901 cells treated with low-concentration chemotherapy. Data are presented as mean ± SEM. **P < 0.01 by 1-way ANOVA with Bonferroni’s post hoc test (F) or by Student’s t test (G and L).