Recombinant Endoglin-Single-Chain Variable Fragment/ Induced Protein 10 Fusion Protein Potently Boosts the Anti-Tumor Efficacy of Adoptively Transferred TRP2-Specific CD8+ CD28+ Cytotoxic T Lymphocytes in Mice

In this research, we studied the therapeutic efficacy of a newly designed fusion protein containing Endoglin single-chain variable fragment and IP10 (Endoglin-scFv/IP10), together with our recently generated TRP2-specific CD8+ CD28+ CTLs (CD8+ CD28+ CTLs) in controlling melanoma growth in mice. The recombinant Endoglin-scFv/IP10 was expressed in E. coli, purified by affinity chromatography, and characterized in vitro for its chemotactic movement and immunoreactivity with endoglin-expressing cells. In vivo, melanoma xenografts were established in mice (C57BL/6) using B16F10 cells. After that, mice were treated with intravenous injections of vehicle (PBS), Endoglin-scFv/IP10 alone, CD8+ CD28+ CTLs alone, or Endoglin-scFv/IP10+ CD8+ CD28+ CTLs. The therapeutic efficacy was assessed by monitoring tumor growth, mouse survival and cellular biomarkers. Endoglin-scFv/IP10 fusion protein combined with CD8+ CD28+ CTLs observed a reduction in tumor growth, resulting in improved survival. On the cellular level, the combination treatment dramatically reduced the number of systemic and tumor associated myeloid-derived suppressor cells or regulatory T cells, increased tumor-responsive interferon-γ-producing lymphocytes and tumor-associated CD8+ CXCR3+ T cells, and inhibited proliferation and angiogenesis but stimulated apoptosis within melanoma tissue. This study demonstrates the therapeutic potential of Endoglin-scFv/IP10 fusion protein in combination with CD8+ CD28+ CTLs in melanoma treatment.