TG6050, an oncolytic vaccinia virus encoding interleukin-12 and anti-CTLA-4 antibody, favors tumor regression via profound immune remodeling of the tumor microenvironment.

BACKGROUND: TG6050 was designed as an improved oncolytic vector, combining the intrinsic properties of vaccinia virus to selectively replicate in tumors with the tumor-restricted expression of recombinant immune effectors to modify the tumor immune phenotype. These properties might be of particular interest for "cold" tumors, either poorly infiltrated or infiltrated with anergic T cells. METHODS: TG6050, an oncolytic vaccinia virus encodes single-chain human interleukin-12 (hIL-12) and full-length anti-cytotoxic T-lymphocyte-associated antigen-4 (@CTLA-4) monoclonal antibody. The relevant properties of TG6050 (replication, cytopathy, transgenes expression and functionality) were extensively characterized in vitro. The biodistribution and pharmacokinetics of the viral vector, @CTLA-4 and IL-12, as well as antitumoral activities (alone or combined with immune checkpoint inhibitors) were investigated in several "hot" (highly infiltrated) and "cold" (poorly infiltrated) syngeneic murine tumor models. The mechanism of action was deciphered by monitoring both systemic and intratumoral immune responses, and by tumor transcriptome analysis. The safety of TG6050 after repeated intravenous administrations was evaluated in cynomolgus monkeys, with a focus on the level of circulating IL-12. RESULTS: Multiplication and propagation of TG6050 in tumor cells in vitro and in vivo were associated with local expression of functional IL-12 and @CTLA-4. This dual mechanism translated into a strong antitumoral activity in both "cold" and "hot" tumor models (B16F10, LLC1 or EMT6, CT26, respectively) that was further amplified when combined with anti-programmed cell death protein-1. Analysis of changes in the tumor microenvironment (TME) after treatment with TG6050 showed increases in interferon-gamma, of CD8+T cells, and of M1/M2 macrophages ratio, as well as a drastic decrease of regulatory T cells. These local modifications were observed alongside bolstering a systemic and specific antitumor adaptive immune response. In toxicology studies, TG6050 did not display any observable adverse effects in cynomolgus monkeys. CONCLUSIONS: TG6050 effectively delivers functional IL-12 and @CTLA-4 into the tumor, resulting in strong antitumor activity. The shift towards an inflamed TME correlated with a boost in systemic antitumor T cells. The solid preclinical data and favorable benefit/risk ratio paved the way for the clinical evaluation of TG6050 in metastatic non-small cell lung cancer (NCT05788926 trial in progress).

Mediation of antitumor activity by AZD4820 oncolytic vaccinia virus encoding IL-12.

Oncolytic viruses are engineered to selectively kill tumor cells and have demonstrated promising results in early-phase clinical trials. To further modulate the innate and adaptive immune system, we generated AZD4820, a vaccinia virus engineered to express interleukin-12 (IL-12), a potent cytokine involved in the activation of natural killer (NK) and T cells and the reprogramming of the tumor immune microenvironment. Testing in cultured human tumor cell lines demonstrated broad in vitro oncolytic activity and IL-12 transgene expression. A surrogate virus expressing murine IL-12 demonstrated antitumor activity in both MC38 and CT26 mouse syngeneic tumor models that responded poorly to immune checkpoint inhibition. In both models, AZD4820 significantly upregulated interferon-gamma (IFN-γ) relative to control mice treated with oncolytic vaccinia virus (VACV)-luciferase. In the CT26 study, 6 of 10 mice had a complete response after treatment with AZD4820 murine surrogate, whereas control VACV-luciferase-treated mice had 0 of 10 complete responders. AZD4820 treatment combined with anti-PD-L1 blocking antibody augmented tumor-specific T cell immunity relative to monotherapies. These findings suggest that vaccinia virus delivery of IL-12, combined with immune checkpoint blockade, elicits antitumor immunity in tumors that respond poorly to immune checkpoint inhibitors.