VISTA×MSLN双特异性抗体导向疗法作为上皮样恶性胸膜间皮瘤生物标志物指导的策略

Background Malignant pleural mesothelioma (MPM) is a rare, lethal cancer largely linked to asbestos exposure, with a 5-year overall survival <20%. Until the approval of nivolumab plus ipilimumab (CheckMate 743), platinum-based chemotherapy was the only systemic standard of care. The epithelioid subtype (~70% of cases) of MPM derives limited benefit from current ICIs, underscoring the need for biomarker-guided therapies. We identified co-expression of the immune checkpoint V-domain Ig suppressor of T-cell activation (VISTA) and the tumor-associated antigen mesothelin (MSLN) in epithelioid MPM and validated this axis by scRNA-seq, which demonstrated concordant VISTA and MSLN transcription within malignant epithelioid clusters. Based on this biomarker pair, we engineered an anti-VISTA×MSLN bispecific antibody (BsAb) and explored multiple modalities, including an IFN-beta mutein immunocytokine and bispecific ADC concepts. Method Anti-VISTA×MSLN BsAb formats were designed and screened for optimal binding/cell-surface coverage on VISTA/MSLN co-expressing cells. A stability/production/PK-enhanced IFN-beta mutein relative to recombinant IFN-beta was fused to the BsAb to generate a trispecific immunocytokine. In vitro binding, internalization, and effector assays were performed in human MPM cell lines. In vivo antitumor activity and TME modulation were tested in xenografts and a syngeneic mouse model; a subset used human IFNAR1/2 knock-in (hIFNAR1/2-KI) mice. To assess suitability for payload delivery, we analyzed antigen-dependent internalization and early bispecific ADC feasibility in vitro. Results scRNA-seq confirmed VISTA/MSLN co-expression in malignant epithelioid populations, supporting VISTA×MSLN as a therapeutic axis. The anti-VISTAxMSLN BsAb showed enhanced binding to dual-positive cells relative to the parental monospecific antibodies and efficient, antigen-driven internalization. The trispecific immunocytokine retained BsAb targeting and induced type I IFN-dependent tumor-cell killing and immune activation in vitro. In vivo, it produced significant antitumor activity in MPM models, including in hIFNAR1/2-KI mice, and remodeled the TME consistent with enhanced antitumor immunity. Internalization/trafficking supported feasibility of developing anti-VISTA×MSLN BsAb ADCs as an additional therapeutic modality. Conclusion VISTA/MSLN co-expression defines a biomarker-guided therapeutic axis in epithelioid MPM. The engineered anti-VISTAxMSLN BsAb is a versatile scaffold enabling multiple modalities- including an IFN-beta mutein immunocytokine with in vivo efficacy and TME reprogramming-and exhibits internalization compatible with bispecific ADC development. Collectively, these findings support anti-VISTA×MSLN-based therapeutics as promising preclinical candidates for epithelioid MPM.