Vegf Removal Delays The Onset Of Acquired Resistance To Target Therapy And Increases The Efficacy Of Immune Checkpoint Inhibitors In Braf-Mutated Melanoma.

The introduction of BRAF inhibitors (BRAFi) has improved response rate and overall survival of metastatic melanoma patients compared to standard chemotherapy. However, acquired drug resistance occurs in nearly all patients. The comprehension of cellular and molecular mechanisms underlying BRAFi resistance could help to identify novel actionable pathways in the treatment of BRAF-dependent tumors. VEGFA is an attractive target for combinatorial cancer therapy, and we have recently demonstrated in melanoma and CRC xenografts that targeting VEGFA enhanced the antitumor effect of BRAFi by normalizing the tumor vasculature, recruiting M1 macrophages, and inducing a remodeling of the extracellular matrix characterized by a reduction in collagen I and in cancer-associated fibroblasts (Comunanza et al., EMBO Mol Med 2017). Based on our previous proof of concept, obtained within an immunodeficient model, here we investigated the therapeutic effect of VEGFA targeting in association with PLX4720 (BRAFi) in a dedicated immune-competent model. D4M cells, a BRAFV600E-mutant melanoma murine cell line, were subcutaneously injected in syngeneic C57BL/6J mice. We demonstrated that the association of BRAFi with DC101 (antibody anti-VEGFR2) had a weak activity while we observed a synergistic antitumor effect when combined with B20 (murine anti-VEGFA neutralizing antibody). Although targeted inhibition of either BRAF or VEGFA delayed the tumor growth, only combined inhibition of both pathways resulted in the regression of initial tumor size, with an evident apoptotic effect, and delayed the onset of acquired resistance to the BRAF inhibition. Since the immune-suppressive role of VEGFA in tumors has been well characterized, we further investigated whether contrasting the VEGF effect along with simultaneous BRAF inhibition can turn into a promotion of both innate and adaptive immunity. Immune phenotype analysis revealed that the combinatorial regimen activated the host immune system, inducing the tumor infiltration of macrophages with tumor-suppressive features, NKs, CD4+ and CD8+ lymphocytes. Most of the infiltrating CD8+ lymphocytes in D4M tumors expressed high levels of PD-1 and none of the treatments significantly modulated PD-1 expression on T cells, suggesting a sustained T-cell exhaustion. We then postulated that the therapeutic effect obtained by the combinatorial VEGF blockade and BRAFi could be further enhanced by the association with an immune-checkpoint inhibitor targeting PD-1. Interestingly, we observed that the addition of anti-PD-1 blocking antibody boosted the antitumor effect and induced striking tumor volume regression in mice receiving the triplet therapy (BRAFi+anti-VEGFA+anti-PD1-1). Our findings provide biologic rationale to explore the association of immunotherapy in novel combinatorial approaches that could improve the clinical outcome exerted by oncogene-targeted therapy, and further investigation is warranted. Citation Format: Valentina Comunanza, Chiara Gigliotti, Gabriella Doronzo, Valentina Martin, Anna Gattuso, Dario Sangiolo, Federica Di Nicolantonio, Federico Bussolino. VEGF removal delays the onset of acquired resistance to target therapy and increases the efficacy of immune checkpoint inhibitors in BRAF-mutated melanoma [abstract]. In: Proceedings of the AACR Special Conference on Melanoma: From Biology to Target; 2019 Jan 15-18; Houston, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(19 Suppl):Abstract nr B17.