癌相关成纤维细胞（CAFs）是肿瘤微环境中最丰富的间质细胞之一，介导结缔组织反应并是免疫抑制微环境的主要驱动因素，导致三阴性乳腺癌（TNBC）免疫治疗失败。因此，减少CAF数量可能增强免疫治疗（如PD-L1抗体）的效果。松弛素（RLN）已被证明能显著改善转化生长因子-β（TGF-β）诱导的CAF活化和肿瘤免疫抑制微环境，但其短半衰期和全身血管扩张限制了其体内功效。因此，将编码松弛素（pRLN）的质粒通过新增的阳离子聚合物聚甲糖（PolyMet）投入表达局部的松弛素，可显著增强基因转移效率，并且低毒性已由我们的实验室证实。为了提高pRLN在体内的稳定性，此复合物进一步形成了脂多聚-γ-谷氨酸（PGA）/PolyMet-pRLN纳米粒子（LPPR）。LPPR粒径为205.5 ± 2.9 nm，Zeta电位为+55.4 ± 1.6 mV。LPPR在体外展示了优异的肿瘤穿透效能，并在4T1luc/CAF肿瘤球中削弱了CAF增殖。在体内，它通过减少纤维化细胞因子的表达逆转异常激活的CAF，消除物理屏障来重新塑造肿瘤间质微环境，使得肿瘤内细胞毒性T细胞浸润增加了2.2倍，而免疫抑制细胞浸润则下降了。因此，LPPR单独对4T1肿瘤承载小鼠的肿瘤生长被抑制，并且当它与PD-L1抗体（aPD-L1）联合时，重新塑造的免疫微环境进一步促进了抗肿瘤效应。总之，本研究提出了一种新的治疗方法，使用LPPR针对肿瘤间质，实现与免疫检查点阻断治疗的组合方案，以应对结缔组织性TNBC模型。©2023年沈阳药科大学。

Cancer-associated fibroblasts (CAFs) are one of the most abundant stromal cells in the tumor microenvironment which mediate desmoplastic response and are the primary driver for an immunosuppressive microenvironment, leading to the failure of triple-negative breast cancer (TNBC) immunotherapy. Therefore, depleting CAFs may enhance the effect of immunotherapy (such as PD-L1 antibody). Relaxin (RLN) has been demonstrated to significantly improve transforming growth factor-β (TGF-β) induced CAFs activation and tumor immunosuppressive microenvironment. However, the short half-life and systemic vasodilation of RLN limit its in vivo efficacy. Here, plasmid encoding relaxin (pRLN) to locally express RLN was delivered with a new positively charged polymer named polymeric metformin (PolyMet), which could increase gene transfer efficiency significantly and have low toxicity that have been certified by our lab before. In order to improve the stability of pRLN in vivo, this complex was further formed lipid poly-γ-glutamic acid (PGA)/PolyMet-pRLN nanoparticle (LPPR). The particle size of LPPR was 205.5 ± 2.9 nm, and the zeta potential was +55.4 ± 1.6 mV. LPPR displayed excellent tumor penetrating efficacy and weaken proliferation of CAFs in 4T1luc/CAFs tumor spheres in vitro. In vivo, it could reverse aberrantly activated CAFs by decreasing the expression of profibrogenic cytokine and remove the physical barrier to reshape the tumor stromal microenvironment, which enabled a 2.2-fold increase in cytotoxic T cell infiltration within the tumor and a decrease in immunosuppressive cells infiltration. Thus, LPPR was observed retarded tumor growth by itself in the 4T1 tumor bearing-mouse, and the reshaped immune microenvironment further led to facilitate antitumor effect when it combined with PD-L1 antibody (aPD-L1). Altogether, this study presented a novel therapeutic approach against tumor stroma using LPPR to achieve a combination regimen with immune checkpoint blockade therapy against the desmoplastic TNBC model.© 2023 Shenyang Pharmaceutical University.