Objective The objective of this study is to improve the efficacy of CLDN18.2/CD3 bispecific T-cell engagers (BiTEs) as a promising immunotherapy against pancreatic ductal adenocarcinoma (PDAC). Design Humanised hCD34+/hCD3e+, Trp53R172HKrasG12DPdx1-Cre (KPC), pancreas-specific Cldn18.2 knockout (KO), fibroblast-specific Fcgr1 KO and patient-derived xenograft/organoid mouse models were constructed. Flow cytometry, Masson staining, Cell Titer Glo assay, virtual drug screening, molecular docking and chromatin immunoprecipitation were conducted. Results CLDN18.2 BiTEs effectively inhibited early tumour growth, but late-stage efficacy was significantly diminished. Mechanically, the Fc fragment of BiTEs interacted with CD64+ cancer-associated fibroblasts (CAFs) via activation of the SYK-VAV2-RhoA-ROCK-MLC2-MRTF-A-α-SMA/collagen-I pathway, which enhanced desmoplasia and limited late-stage infiltration of T cells. Molecular docking analysis found that vilanterol suppressed BiTEs-induced phosphorylation of VAV2 (Y172) in CD64+ CAFs and weakened desmoplasia. Additionally, decreased cyclic guanosine-adenosine monophosphate synthase/stimulator of interferon genes (STING) activity reduced proliferation of TCF-1+PD-1+ stem-like CD8+ T cells, which limited late-stage effects of BiTEs. Finally, vilanterol and the STING agonist synergistically boosted the efficacy of BiTEs by inhibiting the activation of CD64+ CAFs and enriching proliferation of stem-like CD8+ T cells, resulting in sustained anti-tumour activity. Conclusion Vilanterol plus the STING agonist sensitised PDAC to CLDN18.2 BiTEs and augmented efficacy as a potential novel strategy. Data are available upon reasonable request.

中文翻译：

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CD64 + 成纤维细胞靶向维兰特罗和 STING 激动剂通过减少结缔组织和富集干细胞样 CD8 + T 细胞来增强 CLDN18.2 BiTES 对胰腺癌的疗效


目的 本研究的目的是提高 CLDN18.2/CD3 双特异性 T 细胞接合器 （BiTE） 作为胰腺导管腺癌 （PDAC） 有前途的免疫疗法的疗效。设计 构建人源化 hCD34+/hCD3e+、Trp53R172HKrasG12DPdx1-Cre （KPC）、胰腺特异性 Cldn18.2 敲除 （KO）、成纤维细胞特异性 Fcgr1 KO 和患者来源的异种移植物/类器官小鼠模型。进行流式细胞术、 Masson 染色、 细胞滴度 Glo 测定、虚拟药物筛选、分子对接和染色质免疫沉淀。结果 CLDN18.2 BiTEs 可有效抑制早期肿瘤生长，但晚期疗效显著降低。在机械上，BiTES 的 Fc 片段通过激活 SYK-VAV2-RhoA-ROCK-MLC2-MRTF-A-α-SMA/Collagen-I 通路与 CD64 + 癌症相关成纤维细胞 （CAF） 相互作用，从而增强结缔组织增生并限制 T 细胞的晚期浸润。分子对接分析发现，维兰特罗抑制 BiTEs 诱导的 CD64+ CAFs 中 VAV2 （Y172） 磷酸化并减弱结纤维增生。此外，干扰素基因的环鸟苷-腺苷单磷酸合酶/刺激物 （STING） 活性降低降低了 TCF-1+PD-1+ 干细胞样 CD8+T 细胞的增殖，从而限制了 BiTEs 的后期效应。最后，维兰特罗和 STING 激动剂通过抑制 CD64+ CAFs 的激活和丰富干细胞样 CD8+ T 细胞的增殖，协同提高 BiTEs 的疗效，从而产生持续的抗肿瘤活性。结论 Vilanterol 联合 STING 激动剂使 PDAC 对 CLDN18.2 BiTES 敏感，并作为一种潜在的新策略增强疗效。数据可根据合理要求提供。