Project description:BackgroundInterleukin-18 (IL-18), or interferon (IFN)-γ-inducing factor, potentiates T helper 1 and natural killer cell activation as well as CD8+ T-cell proliferation. Recombinant IL-18 has displayed limited clinical efficacy in part due to the expression of the decoy receptor, IL-18 binding protein (IL-18BP). A series of IL-18 variants that are devoid of IL-18BP binding, termed DR18 (decoy-resistant IL-18), was developed via directed evolution. We tested DR18 using oncolytic adenovirus (oAd) as a platform for delivery in syngeneic mouse tumor models.MethodsoAd harboring wild-type IL-18 or DR18 (oAdDR18) was constructed by inserting IL-18 mutant into modified oAd backbone with Ad5/3 chimeric fiber. The delivery effect and IFN-γ induction were determined by ELISA. The antitumor efficiency of oAdDR18 was tested in CT26, B16BL6 and 4T1 tumor-bearing mice, or athymic nude mice and compared with recombinant DR18 protein (rDR18). 4T1 lung metastasis model was used to evaluate the antitumor efficiency of local and distant tumors. Antitumor memory and synergistic effect with an anti-programmed cell death protein-1 (PD-1) antibody was evaluated. The phenotypes of the immune cells in tumor microenvironment were analyzed by flow cytometry and immunohistochemistry.ResultsMice received oAdDR18 maintained stable production of IL-18 and IFN-γ compared with those received rDR18. Intratumoral delivery of oAdDR18 significantly reduced tumor growth across several tumor models, but not in the athymic nude mouse model. Mice that had tumor remission showed antitumor memory. The antitumor effect was associated with intratumor infiltration of CD4+ and CD8+ T cells. DR18 delivered by oAd demonstrated long-lasting and enhanced antitumor activities against local and distant tumors compared with that received rDR18 or wild-type IL-18 delivered by oAd (oAdwtIL-18). oAdDR18 treatment also reduced 4T1 lung metastasis. In addition, combination of this virotherapy with immune checkpoint inhibitors (ICIs)like the anti-PD-1 antibody further enhanced the antitumor activity as compared with respective monotherapy.ConclusionsoAdDR18 demonstrates enhanced antitumor activities through the induction of stronger local and system immunities and modulation of the tumor microenvironment compared with those of oAdwtIL-18 and rDR18. A combination of oncolytic virotherapy with cytokine engineering would lead to cytokine-based therapeutics for cancer and other diseases.

Project description:Oncolytic vaccinia viruses (VVs) are potent stimulators of the immune system and induce immune-mediated tumor clearance and long-term surveillance against tumor recurrence. As such they are ideal treatment modalities for solid tumors including lung cancer. Here, we investigated the use of VVL-m12, a next-generation, genetically modified, interleukin-12 (IL-12)-armed VV, as a new therapeutic strategy to treat murine models of lung cancer and as a mechanism of increasing lung cancer sensitivity to antibody against programmed cell death protein 1 (α-PD1) therapy. The cytotoxicity and replication of VVL-m12, VVL-h12 and control VVs were assessed in lung cancer cell lines. Subcutaneous lung cancer mouse models were established to investigate the anti-tumor activity of the viruses after intratumoral delivery in an immunocompetent disease model. Synergy with α-PD1 or a VV armed with soluble PD-1 (VV-sPD1) was investigated and functional mechanisms behind efficacy probed. Tumor-targeted VVL-m12 replicated to high levels, was cytotoxic in lung cancer cell lines. VVL-m12 demonstrated superior antitumor efficacy in subcutaneous lung cancer models compared with other VVs examined. Importantly, rational combination of VVL-m12 and PD-1 blockade worked synergistically to significantly enhance survival of animals and safely cured lung cancer with no evidence of recurrence. VVL-m12 therapy induced increased intratumoral infiltration of CD4+ and CD8+ T cells and was able to clear tumor at early time points via increased induction and infiltration of effector T cells and central memory T cells (TCM). In addition, VVL-m12 increased dendritic cell activation, induced polarization of M2 macrophages towards an M1 phenotype, and inhibited tumor angiogenesis in vivo. These results demonstrate that VVL-12 has strong potential as a safe and effective antitumor therapeutic for lung cancer. Importantly, VVL-12 can sensitize lung cancers to α-PD1 antibody therapy, and the combined regime creates a highly effective treatment option for patients.

Project description:BackgroundOncolytic virotherapy with vaccinia virus (VV) can lead to effective anti-tumor immunity by turning "cold" tumors into "hot" tumors. However, its therapeutic potential is affected by the tumor's local immunosuppressive tumor microenvironment (TME). Therefore, it is necessary to explore the use of immune checkpoint inhibitors to arm oncolytic VVs to enhance their anti-tumor efficacy.MethodsA novel recombinant oncolytic VV, VV-α-TIGIT, which encoded a fully monoclonal antibody against T-cell immunoglobulin and ITIM domain (TIGIT) was generated by homologous recombination with a shuttle plasmid. The anti-tumor efficacy of the VV-α-TIGIT was investigated in several subcutaneous and ascites tumor models.FindingsThe functional α-TIGIT was sufficiently produced and secreted by tumor cells infected with VV-α-TIGIT, which effectively replicated in tumor cells leading to significant oncolysis. Intratumoral injection of VV-α-TIGIT improved anti-tumor efficacy in several murine subcutaneous tumor models compared to VV-Control (without α-TIGIT insertion). Intraperitoneal injection of VV-α-TIGIT achieved approximately 70% of complete tumor regression in an ascites tumor model. At the same time, treatment with VV-α-TIGIT significantly increased the recruitment and activation of T cells in TME. Moreover, the in vivo anti-tumor activity of VV-α-TIGIT was largely dependent on CD8+ T cell-mediated immunity. Finally, the tumor-bearing mice cured of VV-α-TIGIT treatment resisted rechallenge with the same tumor cells, suggesting a long-term persistence of tumor-specific immunological memory.InterpretationThe recombinant oncolytic virus VV-α-TIGIT successfully combines the advantages of oncolytic virotherapy and intratumorally expression of immune checkpoint inhibitor against TIGIT. This novel strategy can provide information on the optimal design of novel antibody-armed oncolytic viruses for cancer immunotherapy.FundingThis work was supported by the National Natural Science Foundation of China (81773255, 81472820, and 81700037), the Science and Technology Innovation Foundation of Nanjing University (14913414), and the Natural Science Foundation of Jiangsu Province of China (BK20171098).

Project description:By their selective infection or replication into tumors, oncolytic viruses trigger profound modifications of tumor immune microenvironment with a massive infiltration and activation of T cells. This tumor warming up can be further amplified by the local delivery of powerful immunomodulatory molecules encoded into virus’ genome. Those properties are particularly of interest for tumor poorly infiltrated or infiltrated with anergic T cells to reverse patient’s unresponsiveness to immune checkpoint inhibitors (ICI). TG6050 is a new oncolytic vaccinia virus encoding single-chain human Interleukin-12 (hIL-12) and full length anti-cytotoxic T-lymphocyte-associated antigen-4 (@CTLA-4) monoclonal antibody. Replication, oncolysis, selectivity, transgenes expression and functionality of TG6050 were characterized in vitro. Virus and IL-12 pharmacokinetics, antitumoral activities (alone or combined with ICI), and mechanism of action (MOA) were investigated in “hot” (highly infiltrated) and “cold” (poorly infiltrated) syngeneic murine tumor models. MOA was deciphered by monitoring both systemic and tumor immune responses, and tumor transcriptome analyses. Finally, TG6050 safety after repeated intravenous administrations was evaluated in cynomolgus monkeys and special attention was paid to the level of circulating IL-12 potentially associated with toxic effects. In vitro or in vivo replication of TG6050 in tumor cells allows local expression of functional IL-12 and @CTLA-4. The expression of transgenes, together with the virus replication, translated into an antitumoral activity in both “cold and “hot” tumor murine models that was further amplified by combination with ICI. Analysis of tumor microenvironment (TME) after TG6050 treatment showed an increase of interferon gamma, CD8+ T cells, M1/M2 macrophages ratio and a drastic decrease of regulatory T cells. Those local modifications were observed together with an enhanced systemic and specific antitumor adaptive immune response. In toxicology studies, the virus did not produce any observable adverse effects in cynomolgus monkeys. Conclusions: TG6050 allows the delivery of functional IL-12 and @CTLA-4 into tumor resulting in an excellent antitumor activity along with a shift towards an inflamed TME, and a boost of the systemic antitumor T-cells. Toxicological studies in macaques demonstrated the safety of TG6050. Together these results paved the way for clinical evaluation of TG6050 in metastatic non-small cell lung cancer (NCT05788926).

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

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

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