Ghosh Arnab, Bhoumick Avinandan, Paul Subhojit, Chatterjee Akash, Mandal Subhasis, Basu Abhimanyu, Mukhopadhyay Soma, Das Kaushik, Sen Prosenjit
School of Biological Sciences, Indian Association for the Cultivation of Science, Jadavpur, Kolkata, India.
IPGMER, SSKM Hospital, Bhowanipore, Kolkata, India.
J Thromb Haemost. 2025 Mar;23(3):903-920. doi: 10.1016/j.jtha.2024.11.027. Epub 2024 Dec 10.
Treatment of breast cancers with immunotherapy has so far achieved limited success. Traditional immunotherapies focusing on cytotoxic T cells have attained modest success, while the approval of phagocytic checkpoint blockers is still pending. Coagulation proteases are crucial to cancer growth and proliferation, but their relevance in altering the immunologic topography in tumors remains largely unknown.
In this study, we aimed to examine whether factor VIIa (FVIIa)-driven protease-activated receptor 2 (PAR2) activation and its subsequent signaling pathways assist cancer cells in evading phagocytic macrophages.
Peripheral blood mononuclear cell- or THP-1-derived macrophages were cocultured with MDA-MB-468 cells that were pretreated with or without FVIIa. The phagocytic activity of macrophages was assessed through flow cytometry and immunofluorescence. Additionally, an allograft model using wild-type and PAR2-deleted 4T1 cells was employed to investigate the impact of PAR2 activation on immune escape from macrophages in vivo.
We found evidence that FVIIa-induced PAR2 cleavage activates downstream signaling cascades and augments cellular levels of microRNA221, which transcriptionally activates both CD47 and stanniocalcein 1 expression, thereby assisting the escape from phagocytosis by macrophages. Stanniocalcein 1 decreases the surface expression of calreticulin, a dominant prophagocytic signal, thereby tilting it in favor of phagocytic evasion. Mouse models using PAR2-depleted cells displayed smaller tumor volumes and corresponding greater phagocytic events when combined with anti-CD47/anti-PD-L1 antibodies.
PAR2 signaling initiates an intrinsic mechanism of immune escape by diminishing phagocytosis of cancer cells.
迄今为止，用免疫疗法治疗乳腺癌取得的成功有限。专注于细胞毒性T细胞的传统免疫疗法取得了一定成效，而吞噬细胞检查点阻断剂的批准仍在等待中。凝血蛋白酶对癌症的生长和增殖至关重要，但其在改变肿瘤免疫格局方面的相关性仍 largely unknown。
在本研究中，我们旨在研究因子VIIa（FVIIa）驱动的蛋白酶激活受体2（PAR2）激活及其随后的信号通路是否有助于癌细胞逃避吞噬性巨噬细胞。
将外周血单核细胞或THP-1来源的巨噬细胞与用或不用FVIIa预处理的MDA-MB-468细胞共培养。通过流式细胞术和免疫荧光评估巨噬细胞的吞噬活性。此外，使用野生型和PAR2缺失的4T1细胞的同种异体移植模型来研究PAR2激活对体内巨噬细胞免疫逃逸的影响。
我们发现证据表明，FVIIa诱导的PAR2裂解激活下游信号级联反应并增加microRNA221的细胞水平，后者转录激活CD47和钙网蛋白1的表达，从而有助于逃避巨噬细胞的吞噬作用。钙网蛋白1降低了钙网蛋白的表面表达，钙网蛋白是一种主要的促吞噬信号，从而使其倾向于吞噬逃避。使用PAR2缺失细胞的小鼠模型与抗CD47/抗PD-L1抗体联合使用时，肿瘤体积较小，相应的吞噬事件更多。
PAR2信号传导通过减少癌细胞的吞噬作用启动免疫逃逸的内在机制。
