Comprehensive preclinical studies of BT-001: An oncolytic vaccinia virus armed with Tregdepleting @CTLA4 and GM-CSF

Abstract Background Immune checkpoint blockade (ICB) is a clinically proven concept to treat cancer. Still, a majority of cancer patients including those with poorly immune infiltrated “cold” tumors are resistant to currently available ICB therapies. Oncolytic viruses are natural immunomodulators able to turn “cold” tumors “hot” and, for some of them, to deliver therapeutic payloads intratumorally. Payloads of choice are those with tolerability and/or pharmacokinetic/biodistribution issues, such as @CTLA4 antibodies and GM-CSF respectively. Here, Transgene and BioInvent present a preclinical characterization of a highly efficacious and potentially safe strategy to target CTLA-4 and GM-CSF using vectorization in an oncolytic vaccinia virus. Methods A novel human IgG1 CTLA-4 antibody (4-E03) was identified using function-first screening for mAbs and targets associated with checkpoint blocking and superior Treg depleting activity. A tumor-selective oncolytic Vaccinia vector was then engineered to encode this novel αCTLA-4 as full-length antibody (i.e. distinct transgenes for light and heavy chains) and GM-CSF (VVGM-αhCTLA4, BT-001). Viruses encoding matching mouse surrogate payloads were additionally generated, enabling proof-of-concept studies in syngeneic immune competent mouse tumor models. Results Our studies demonstrate that two chains cloning approach allowed to express @CTLA4 at a high level in numerous infected human tumor cell lines without affecting replication, oncolytic activity or genetic stability of the virus. Intratumoral (i.t.) administration of VVGM-αCTLA4 achieved tumor-restricted CTLA-4 receptor saturation at least for 10 days, Treg-depletion antigen cross-presentation, stronger systemic expansion of tumor-specific CD8+ T cells and antitumor immunity compared with systemic αCTLA-4 antibody or unarmed virus therapies. Remarkably, in a clinically relevant mouse model resistant to systemic immune checkpoint blockade, i.t. VVGM-αCTLA4 synergized with αPD-1 to reject “cold” tumors. Conclusion Our findings demonstrate in vivo proof-of-concept for tumor delivery by Vaccinia virus of GM-CSF and strongly Treg-depleting, immune checkpoint blocking, vectorized αCTLA-4. This vector-based delivery is a highly effective and safe strategy to target CTLA-4 which overcomes current limitations of approved αCTLA-4 regimens. A clinical trial evaluating i.t. VVGM-αhCTLA4 (BT-001) alone and in combination with αPD-1 in metastatic or advanced solid tumors has been initiated. Citation Format: Jean-Baptiste Marchand, Monika Semmrich, Christelle Ziller, Matilda Rehn, Laetitia Fend, Petra Holmkvist, Nathalie Silvestre, Carolin Svensson, Kleinpeter Patricia, Jules Deforges, Fred Junghus, Linda Mårtensson, Johann Foloppe, Ingrid Teige, Björn Frendéus, Eric Quéméneur. Comprehensive preclinical studies of BT-001: An oncolytic vaccinia virus armed with Treg-depleting @CTLA4 and GM-CSF [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3567.