Tumor-secreted AGR2 induces dendritic cell dysfunction and impairs immunotherapy efficacy in LKB1-deficient cancer.

Abstract

Background: LKB1 (STK11)-deficient tumors exhibit an immunosuppressive microenvironment that limits the efficacy of immunotherapies such as anti-programmed cell death protein 1 (PD-1) antibodies. However, the underlying mechanisms driving immune evasion remain unclear. Dendritic cells (DCs), especially conventional DC 1 (cDC1), play a crucial role in antigen presentation and CD8⁺ T-cell activation, yet their dysfunction in LKB1-deficient tumors has not been well characterized.

Methods: Tumor-intrinsic LKB1 deficiency was modeled by subcutaneous inoculation of CRISPR/Cas9-engineered Stk11-knockout tumor cell lines into syngeneic mice. DC infiltration and function were assessed through a series of flow cytometry-based in vivo and in vitro assays, including analyses of infiltration, migration, antigen uptake, maturation, and CD8⁺ T-cell priming. Naïve CD8⁺ T-cell activation by cDC1s was evaluated via adoptive transfer of CD45.1⁺ OT-I T cells. Secretome proteomics and functional rescue experiments identified anterior gradient 2 (AGR2) as a key immunosuppressive mediator, and arginase 1 (ARG1) was validated as its functional interactor through liquid chromatography-tandem mass spectrometry (LC-MS/MS), co-immunoprecipitation, and imaging flow cytometry. Upstream regulation of AGR2 was investigated by AMPKα/FOXA1 silencing and chromatin immunoprecipitation (ChIP) quantitative PCR, revealing a tumor-intrinsic AMPKα-FOXA1-AGR2 axis driving DC dysfunction.

Results: LKB1-deficient tumors exhibited significantly reduced cDC1 infiltration, and cDC1s were functionally impaired in antigen uptake, migration, and naïve CD8⁺ T-cell priming. AGR2, a secreted protein transcriptionally upregulated via the tumor-intrinsic AMPKα-FOXA1 pathway, was identified as a key mediator of DC dysfunction. Mechanistically, AGR2 interacted with ARG1, stabilizing its expression and promoting ARG1 accumulation in DCs. Restoring DC infiltration and activity through FLT3L-driven expansion and tumor-antigen-specific DC supplementation significantly enhanced the efficacy of anti-PD-1 treatment in LKB1-deficient lung adenocarcinoma models.

Conclusion: Our study identifies profound DC dysfunction-characterized by impaired antigen uptake, migration, and naïve CD8⁺ T-cell priming-as a key mechanism of immune evasion in LKB1-deficient tumors. This dysfunction is driven by tumor-secreted AGR2, which stabilizes ARG1 in DCs and suppresses CD8⁺ T-cell activation. Targeting the AMPKα-FOXA1-AGR2-ARG1 axis or restoring DC competence offers a promising strategy to enhance immunotherapy efficacy in LKB1-deficient subtype.