Anti-VEGF Immunotherapy with HEBERSaVax Suppresses Melanoma Growth and Metastasis via Angiogenesis Blockade and Enhanced T-cell Infiltration

Yanelys Morera-Díaz^1*Josune Garcia²Camila Canaán-Haden¹Mónica Bequet-Romero¹Isabel Gonzalez-Moya¹Marta Ayala-Avila¹Srdan Tadic²Pablo Garrido²Judit Narro-Íñiguez²Dasha Fuentes-Morales³Johana Bernáldez-Sarabia⁴Blanca J Valdovinos-Navarro⁴Alexei F Licea Navarro⁴Alfredo Martínez^2*

• ¹Center for Genetic Engineering and Biotechnology (CIGB), Playa, Cuba
• ²Centro de Investigacion Biomedica de La Rioja, Logroño, Spain
• ³National Center for the Production of Laboratory Animals (CENPALAB), Havana, Cuba
• ⁴Centro de Investigacion Cientifica y de Educacion Superior de Ensenada, Ensenada, Mexico

Targeting tumor angiogenesis through vascular endothelial growth factor (VEGF) blockade represents a promising strategy for melanoma treatment. Here, we evaluate the therapeutic potential of HEBERSaVax, an anti-VEGF active immunotherapy, in aggressive B16-F10 syngeneic melanoma models. The antitumor activity of HEBERSaVax, formulated with aluminum phosphate adjuvant, was evaluated in C57BL/6 mice using two distinct B16-F10 melanoma models: (1) subcutaneous inoculation to assess primary tumor growth inhibition, and (2) intravenous inoculation to quantify lung metastasis suppression. Tumor vasculature and microenvironment changes were analyzed via immunohistochemistry (CD31, α-SMA, CD4+, CD8+). HEBERSaVax significantly reduced primary tumor volume and weight in subcutaneous implants compared to adjuvant controls. Histopathological analysis revealed potent angiogenesis inhibition, decreased CD31+ vessel density, and vascular remodeling. Concomitant with tumor control, we observed changes in the tumor microenvironment, including a reduction in α-SMA+ pericytes and an increase in the infiltration of CD4+ and CD8+ T cells. In the metastatic model, HEBERSaVax-treated mice showed fewer pulmonary nodules versus controls. Our results demonstrate that HEBERSaVax mediates dual antitumor efficacy by simultaneously suppressing VEGF-dependent angiogenesis and promoting immune-related changes in the melanoma microenvironment. These findings support the further development of HEBERSaVax as a promising active immunotherapy for VEGF-driven advanced melanoma.