Targeting asparagine potentiates anti-PD-L1 immunotherapy in gastric cancer by enhancing CD8⁺ T cell anti-tumor response

Mingpai Ge^1,2Longfei Wang³Bowen Zheng⁴Lu Zhan¹Lu Cui¹Han Wang¹Enyuan Huang¹Yuan Xu¹Xusheng Chang¹Zhaorui Liu¹Jun Xu^1*Kai Yin^1,2*

• ¹Changhai Hospital, Second Military Medical University, Shanghai, China
• ²Shidong Hospital, Shanghai 200082, China, Shanghai, China
• ³Shanghai Engineering Research Center of Immunotherapeutics, Fudan University, Shanghai, China, Shanghai, China
• ⁴Tianjin Medical University General Hospital, Tianjin 300052, China, Tianjin, China

Introduction: While immunotherapy holds considerable promise for cancer treatment, its clinical efficacy in gastric cancer (GC) is frequently constrained by the intricate tumor microenvironment (TME), a milieu profoundly influenced by aberrant tumor metabolism. Asparagine, an amino acid indispensable for neoplastic cell proliferation, is also implicated in modulating the metabolic programming of CD8+ T cells. This study aimed to delineate the impact of targeting asparagine on the GC immune microenvironment and the mechanisms underpinning the antitumor activity of its combination with immunotherapy. Methods: GC tumor models were established to evaluate the therapeutic efficacy of asparagine targeting. Flow cytometry was employed to meticulously analyze CD8+ T cells within the TME, and the concentrations of IFN-γ, GZMB, CXCL9, and CXCL10 were precisely quantified by ELISA. The effects of asparagine targeting on CD8+ T cell activation and antitumor function were rigorously assessed in vitro and in vivo. GC models were utilized to evaluate the therapeutic benefit afforded by combining asparagine targeting with anti-PD-L1 therapy. The indispensable role of CD8+ T cells in the combined treatment was definitively established through T cell depletion experiments utilizing anti-CD4/CD8 antibodies. Results: Targeting asparagine demonstrated profound inhibitory effects on GC growth both in vitro and in vivo, strongly implicating immune system participation in its antitumor activity. Mechanistic investigations unveiled that asparagine targeting significantly augmented the proportion of CD8+ T cells within the TME and robustly upregulated the expression of IFN-γ, GZMB, CXCL9, and CXCL10. These findings suggest a potential association with enhanced CD8+ T cell proliferation and intrinsic antitumor effects elicited by asparagine targeting. Targeting asparagine in conjunction with anti-PD-L1 therapy exhibited remarkable synergistic antitumor activity. Conversely, selective depletion of CD8+ T cells substantially attenuated the therapeutic efficacy of the combined regimen. Conclusion: This study suggests that targeting asparagine promotes CD8+ T cell activation and infiltration, thereby effectively remodeling the GC immune microenvironment and significantly augmenting host antitumor immune responses. The combination of asparagine targeting and anti-PD-L1 therapy elicits potent anti-tumor effects in vivo, and its therapeutic efficacy is demonstrably contingent upon the presence of CD8+ T cells.