Inhibiting The xCT Antiporter Sensitizes High Risk DLBCL to Doxorubicin-Containing Therapy

Abstract Background. Although outcomes in diffuse large B-cell lymphoma (DLBCL) have dramatically improved following the introduction of R-CHOP two decades ago, one-third of the patients fail to attain durable remission and require salvage and additional therapies. Many attempts to improve upon R-CHOP efficacy by incorporating targeted agents have failed, suggesting a need to better understand the mechanisms underlying R-CHOP resistance/responsiveness. We have previously reported that Doxorubicin (Dox), the main cytotoxic ingredient in CHOP, has subtype-specific mechanisms of cytotoxicity in DLBCLs due to differences in its subcellular distribution pattern. Specifically, Dox-induced cytotoxicity in the therapy refractory ABC-DLBCLs is largely dependent on oxidative stress rather than the DNA damage response. These findings suggest that R-CHOP outcome could be modulated by manipulating oxidative stress response in DLBCL cells. This study focuses on xCT, the light chain of the cystine/glutamate antiporter xc` that controls the rate-limiting step for glutathione (GSH) biosynthesis. We demonstrate novel roles of the xCT-GSH-ROS axis in maintenance and therapeutic response of high risk DLBCLs. Methods. Prognostic significance of xCT protein expression in a cohort of 87 DLBCL patients treated with R-CHOP were evaluated. Dynamic changes of the cellular GSH pool to Dox treatment were examined. xCT inhibition was achieved using a small molecular inhibitor sulfasalazine (SASP) and through shRNA-mediated knock-down. The ability of SASP to sensitize ABC-DLBCL/double-hit lymphomas to Dox-containing therapy was investigated in cell line and NSG-based xenograft models. Results. Dox elicits a rapid antioxidant response characterized by a GSH spike that can be eliminated by suppressing the xCT antiporter function. Mechanistically, xCT inhibition depleted the total GSH pool and markedly enhanced the p38-dependent apoptosis response following Dox treatment. Furthermore, supporting a role of the xCT-GSH axis in clinical response to Dox-containing therapies, xCT expression predicts poor prognosis among ABC-DLBCL but not GCB-DLBCL patients treated with R-CHOP. Finally, we demonstrate that combinatorial treatment with Dox and SASP produced marked synergy in xenograft models for ABC-DLBCL/double-hit lymphomas. Conclusions. In ABC-DLBCL, the xCT-GSH-p38 axis plays a critical role in survival response to Dox-containing therapies. xCT inhibition may hold significant promise to improve the first-line treatment outcome of patients with high risk DLBCL.