Introduction: Current therapies for multiple myeloma (MM), such as immunomodulatory agents, proteasome inhibitors, stem-cell transplantation, and monoclonal antibodies against tumor-associated antigens have greatly improved patient survival. However, MM remains an incurable disease as most patients will eventually relapse. Recent advances in targeted T-cell therapies have shown promise in clinical trials but the adaptive immune system may be insufficient to eradicate all MM clones. In contrast, treatments harnessing the innate immune system have been relatively underdeveloped in MM despite evidence suggesting a role of innate immunity in the efficacy of existing therapies. Innate or innate-like cells, such as NK and γδ T cells, have the potential to display strong anti-tumor activity, and strategies aimed to improve or re-direct their cytotoxicity represent a new opportunity for cancer immunotherapies and a complementary approach to existing therapies. Here we describe the preclinical characterization of CTX-8573, a novel multispecific antibody that targets B-cell maturation antigen (BCMA) on tumor cells and promotes potent cytotoxicity by NK and γδ T cells through engagement of the activating receptors NKp30 and CD16a. Method: Bispecific constructs were generated by appending two common-light chain compatible anti-NKp30 Fab fragments to the C-terminus of an anti-BCMA IgG1 antibody containing an afucosylated Fc for enhanced CD16a engagement. To test the effects of targeting NKp30 alone, variants were expressed with an aglycosylated Fc to eliminate CD16a binding. In-vitro assays were performed with primary NK or γδ T cells to determine innate-cell activation, cytokine production, proliferation, and target-cell cytotoxicity against tumor cell lines with a range of BCMA expression levels. In-vivo efficacy studies were performed in multiple humanized mouse models and pharmacokinetics and safety were evaluated in Cynomolgus monkeys. Results: CTX-8573 is highly expressed in CHO cells with minimal aggregation and displays stability, solubility, and binding to BCMA and NKp30 equivalent to the parental monoclonal antibodies. By engaging NKp30 and CD16a, CTX-8573 promotes potent cytotoxicity of BCMA expressing target cells by NK and γδ T cells with >100 fold reduced EC50 compared to the corresponding BCMA monoclonal antibody control. CTX-8573 also demonstrates robust killing of low BCMA expressing cell lines including RPMI-8226 where monoclonal BCMA antibodies lack activity. An aglycosylated variant of CTX-8573 lacking CD16a binding maintains cell killing activity, demonstrating that engagement of NKp30 alone is sufficient to promote innate cell activation and cytotoxicity, although activity is enhanced by CD16A engagement. Furthermore, CTX-8573 maintains its cytotoxic activity in presence of soluble BCMA or BCMA ligands APRIL and BAFF. CTX-8573 does not induce innate cell activation, cytokine production, or killing in the absence of BCMA expressing target cells, supporting a wide therapeutic window. Additionally, unlike daratumumab, CTX-8573 does not induce NK-cell fratricide. In-vivo, CTX-8573 demonstrates anti-tumor efficacy in multiple humanized mouse models including killing of low BCMA expressing cell lines. In Cynomolgus monkeys, CTX-8573 displays standard biphasic pharmacokinetics with a 16 day β-phase half-life and has no evidence of systemic immune activation as measured by C-reactive protein levels. Lastly, NKp30 expression is maintained on bone marrow NK cells from MM patients including the presence of a significant NKp30+CD16a- subpopulation. Conclusion: CTX-8573 represents a novel class of bispecific antibodies that promote potent tumor cell killing by NK and γδ T-cells through engagement of the activating receptors NKp30 and CD16a. CTX-8573 demonstrates strong anti-tumor efficacy in vitro and in vivo, a wide therapeutic window with no evidence of systemic toxicity, and monoclonal-like pharmacokinetics and manufacturability. Together, these data highlight the potential of CTX-8573 as a novel treatment for MM either alone or as a complement to existing therapies. Disclosures Watkins-Yoon: Compass therapeutics LLC: Employment, Equity Ownership. Guzman:Compass therapeutics LLC: Employment, Equity Ownership. Oliphant:Compass therapeutics LLC: Employment, Equity Ownership. Haserlat:Compass therapeutics LLC: Employment, Equity Ownership. Leung:Compass therapeutics LLC: Employment, Equity Ownership. Chottin:University of Louisiana at Lafayette: Employment. Ophir:Compass therapeutics LLC: Employment, Equity Ownership. Vekeria:Compass therapeutics LLC: Employment, Equity Ownership. Nanjappa:Compass therapeutics LLC: Employment, Equity Ownership. Markrush:Compass therapeutics LLC: Employment, Equity Ownership. McConaughy:Compass therapeutics LLC: Employment, Equity Ownership. Wang:Compass therapeutics LLC: Employment, Equity Ownership. Schilling:Compass therapeutics LLC: Employment, Equity Ownership. Kim:Compass therapeutics LLC: Employment, Equity Ownership. Wu:Compass Therapeutics LLC: Employment, Equity Ownership. Liu:Compass therapeutics LLC: Employment, Equity Ownership. Rogers:University of Louisiana at Lafayette: Employment. Villinger:University of Louisiana at Lafayette: Employment. Gong:Compass therapeutics LLC: Employment, Equity Ownership. Hamilton:Compass therapeutics LLC: Employment, Equity Ownership. Bobrowicz:Compass therapeutics LLC: Employment, Equity Ownership. Schuetz:Compass therapeutics LLC: Employment, Equity Ownership. Schmidt:Compass therapeutics LLC: Employment, Equity Ownership. Draghi:Compass therapeutics LLC: Employment, Equity Ownership.
Blockade of inhibitory checkpoint pathways, such as PD-1/PD-L1 and CTLA-4, has provided significant benefit to subsets of patients and changed the cancer therapy landscape. These checkpoint inhibitors promote adaptive T cell-mediated anti-tumor immunity while signaling regulatory protein alpha (SIRPα)/CD47 axis represents an innate specific checkpoint that has become an attractive target for immunotherapy. CD47 is expressed by virtually all cells, providing an anti-phagocytic “don’t eat me” signal through its interaction with the inhibitory immune receptor SIRPα which is expressed on myeloid cells. Blockade of the CD47-SIRPα interaction has been shown to promote phagocytosis and antigen presentation. Bispecific antibodies are emerging as an attractive therapeutic modality capable of simultaneously engaging two targets with a single drug to improve efficacy. Here we describe the discovery and characterization of a series of fully human, tetravalent common light chain SIRPαxPD-L1 bispecific antibodies. CTX-5861 was identified based on epitope binning and functional studies. The CD47 binding domain of SIRPα is known to be highly polymorphic, with allelic variants differing by as many as 13 amino acids. CTX-5861 bound to all tested polymorphic variants with higher specificity towards SIRPα expressed on myeloid cells as opposed SIRPγ expressed on T cells. In vitro studies showed that, CTX-5861 enhanced antibody-dependent cell mediated phagocytosis by macrophages and increased antigen presentation by dendritic cells. In vivo, CTX-5861 combination with tumor targeting antibodies significantly reduced tumor burden in the B16F10 lung metastasis and subcutaneous MC38 models without inducing hematological toxicities commonly associated with anti-CD47 antibodies. In the MC38 model, the addition of either PD-L1 blockade or CD137 agonism to the combination regimen led to curative anti-tumor activity in 40-60% of the mice. Quadruple combination of tumor targeting antibody, CTX-5861, PD-L1 blockade, and CD137 agonism achieved a 100% cure rate. These results suggest that SIRPα blockade, as mediated by CTX-5861, could be of central importance in rationally-designed, multi-agent immunotherapy combinations. Through its ability to enhance tumor antigen processing and presentation without incurring hematological toxicity, CTX-5861 represents a highly promising novel agent for cancer immunotherapy. Note: This abstract was not presented at the meeting. Citation Format: Ugur Eskiocak, Wilson Guzman, Thomas Daly, Allison Nelson, Pearl Bakhru, Jason Lajoie, Sara Haserlat, Samantha Ottinger, Lucy Liu, Amanda Oliphant, Alan Leung, Girish Hemashettar, Rachel Rennard, Benjamin Wolf, Monia Draghi, Michael Schmidt, Robert Tighe. CTX-5861 mediated SIRPα blockade combines with tumor targeting antibodies, checkpoint blockade and/or CD137 agonism to elicit curative anti-tumor activity in syngeneic mouse models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3239.
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