Prince Stuart, Viitala Miro, Sjöroos Riikka, Bendes Ábris Á, Rannikko Jenna H, Patten Daniel A, di Benedetto Ilaria, Turpin Rita, Ylitalo Arno, Tyni Laura, Figueiredo Carlos R, Boström Pia, Koskivuo Ilkka, Salminen Tiina A, Shetty Shishir, Hollmén Maija
MediCity Research Laboratory and InFLAMES Flagship, University of Turku, Turku, Finland.
Structural Bioinformatics Laboratory and InFLAMES flagship, Faculty of Science and Engineering, Åbo Akademi University, Turku, Finland.
Theranostics. 2025 Jun 23;15(15):7501-7527. doi: 10.7150/thno.110544. eCollection 2025.
Clever-1 is a multifunctional scavenger receptor that promotes immunosuppressive activity in macrophages, contributing to tumor immune evasion. Its high expression correlates with resistance to immune checkpoint inhibitors, and co-targeting Clever-1 with anti-PD-1 enhances therapeutic efficacy in refractory tumor models. The humanized anti-Clever-1 IgG4 antibody, bexmarilimab, is under clinical investigation for treating solid tumors (NCT03733990) and hematological malignancies (NCT05428969). To assess the impact of Clever-1 in cancer, we analyzed plasma samples from breast cancer patients (n=139) and bexmarilimab-treated clinical trial participants (n=193) using TRFIA-based ELISA to quantify secreted Clever-1 (sClever-1). A recombinant sClever-1 protein was produced and characterized biophysically. Functional assays, including flow cytometry, Western blotting, T cell activation, and Jurkat reporter systems, were used to assess interactions with T cells. Mechanistic studies involved extracellular vesicle isolation, pulldown assays, and mass spectrometry. Inhibitor studies and patient-derived tumor explants were used to evaluate the immunomodulatory impact of sClever-1 and its effect on anti-PD-1 responses. sClever-1 was significantly enriched in the plasma of cancer patients and reduced following bexmarilimab treatment. Its release was induced by IFNγ/LPS via serine protease-dependent cleavage. The recombinant sClever-1 bound selectively to activated T cells via mannose-6-phosphate-mediated interaction with IGF2R, impairing TCR signaling and Th1 expansion. sClever-1 was also associated with macrophage-derived extracellular vesicles and contributed to T cell tolerance and reduced anti-PD-1 efficacy. In tumor explants, sClever-1 bound to activated CD4 and CD8 T cells and increased TGFβ secretion. These findings identify sClever-1 as a previously unrecognized, immunosuppressive mediator in cancer that operates independently of cellular Clever-1 expression. sClever-1 may serve as both a therapeutic target and biomarker to guide immunotherapy strategies.
Clever-1是一种多功能清道夫受体，可促进巨噬细胞的免疫抑制活性，导致肿瘤免疫逃逸。其高表达与对免疫检查点抑制剂的耐药性相关，在难治性肿瘤模型中，将Clever-1与抗PD-1共同靶向可提高治疗效果。人源化抗Clever-1 IgG4抗体bexmarilimab正在进行治疗实体瘤（NCT03733990）和血液系统恶性肿瘤（NCT05428969）的临床试验。为了评估Clever-1在癌症中的作用，我们使用基于时间分辨荧光免疫分析（TRFIA）的酶联免疫吸附测定（ELISA）分析了乳腺癌患者（n = 139）和接受bexmarilimab治疗的临床试验参与者（n = 193）的血浆样本，以定量分泌型Clever-1（sClever-1）。制备了重组sClever-1蛋白并对其进行了生物物理特性鉴定。使用包括流式细胞术、蛋白质免疫印迹、T细胞活化和Jurkat报告系统在内的功能分析来评估与T细胞的相互作用。机制研究涉及细胞外囊泡分离、下拉分析和质谱分析。使用抑制剂研究和患者来源的肿瘤外植体来评估sClever-1的免疫调节作用及其对抗PD-1反应的影响。sClever-1在癌症患者血浆中显著富集，bexmarilimab治疗后降低。IFNγ/LPS通过丝氨酸蛋白酶依赖性切割诱导其释放。重组sClever-1通过与IGF2R的甘露糖-6-磷酸介导的相互作用选择性地结合活化的T细胞，损害TCR信号传导和Th1细胞扩增。sClever-1还与巨噬细胞来源细胞外囊泡相关，并导致T细胞耐受和抗PD-1疗效降低。在肿瘤外植体中，sClever-1与活化的CD4和CD8 T细胞结合并增加TGFβ分泌。这些发现表明，sClever-1是癌症中一种先前未被认识的免疫抑制介质，其作用独立于细胞Clever-1的表达。sClever-1可能既是治疗靶点，也是指导免疫治疗策略的生物标志物。
