Abstract A076: DSP107—a novel SIRPα-4-1BBL dual signaling protein (DSP) for cancer immunotherapy

Abstract

Background: The (re)activation of anticancer innate and adaptive immunity is at the forefront of developments in cancer therapy. Here, we report on a new immunotherapeutic fusion protein, termed Dual Signaling Protein 107 (DSP107). DSP107 was designed to combine activation of innate and adaptive immunity, by both blocking CD47/SIRPα interaction and activating 4-1BB. CD47 is overexpressed on cancer cells and upon binding to SIRPα on phagocytes transmits a “don’t eat me” signal, thereby suppressing innate immunity. 4-1BB is a costimulatory receptor that is transiently upregulated on tumor-infiltrating T-cells and is considered a surrogate marker for the tumor-reactive T-cell population. Activation of 4-1BB using its ligand or by agonistic antibodies reactivates anti-cancer T-cell immunity. In DSP107, the extracellular domains of SIRPα and 41BBL have been fused, yielding a dual function protein. DSP107 is produced as a homotrimer due to the trimerization property of 41BBL, an essential element for activating the 41BB receptor, a member of the TNF super-family of receptors. DSP107 was designed to bind to CD47 on tumor cells, thereby removing the inhibitory signal delivered to phagocytes. Simultaneously, CD47-mediated surface immobilization of DSP107 enables delivery of the 41BBL-4-1BB costimulatory signal to tumor localized T-cells. This dual immunomodulatory effect of DSP107 is designed to unleash both innate and adaptive immune responses targeted to the tumor site. Methods and Results: Trimeric DSP107 was successfully produced in a mammalian expression system. Both sides of DSP107 bound their cognate counterparts in kinetic Blitz binding assays and on human tumor and immune cell surfaces. The binding affinity of DSP107 was 1.6 nM for human CD47 and 0.69 nM for human 4-1BB as determined using BIAcore analysis. DSP107 blocked the interaction of SIRPα with CD47 in an ELISA-based competition assay (EC50 of 0.03 nM). DSP107 induced granulocyte- and macrophage-mediated phagocytosis of several lymphoma, leukemia and carcinoma cell lines in vitro. Further, DSP107 treatment triggered phagocytosis of primary AML cells by autologous macrophages. Co-treatment with DSP107 and therapeutic tumor-targeting antibodies, i.e., rituximab or cetuximab, resulted in enhanced phagocytosis of lymphoma or carcinoma cells, respectively. In a reporter assay measuring IL-8 secretion upon binding to/activation of 4-1BB, DSP107 activated 4-1BB signaling only in the presence of CD47-expressing cells. Further, DSP107 augmented the activation of purified T-cells activated by suboptimal concentrations of αCD3 + IL2 or αCD3/αCD28 Dynabeads in CD47 coated plates, as measured by percentage of CD25 expressing cells (up to 3-fold). When PBMCs were co-cultured with or without CD47-expressing cancer cells and stimulated with suboptimal concentrations of αCD3 + IL2, DSP107 treatment resulted in increased secretion of IFNg up to 2-fold), and increased T-cell proliferation (up to 2-fold). Conclusions: Here we demonstrate the feasibility and functional activity of DSP107, a novel therapeutic protein that combines activation of both the innate and adaptive immune system. Dual targeting, by the two functional sides of DSP107, offers multiple functionalities that act simultaneously and may result in a synergistic effect. DSP107 is now in IND-enabling studies and CMC development. The DSP platform can be designed for selective tumor site or microenvironment targeting and is adaptable to most checkpoint targets. Citation Format: Yosi M. Gozlan, Susan Hilgendorf, Alexandra Aronin, Yehudith Sagiv, Liat Ben-gigi-Tamir, Shira Amsili, Ami Tamir, Iris Pecker, Shirley Greenwald, Ayelet Chajut, Adam Foley-Comer, Yaron Pereg, Amnon Peled, Michal Dranitzki-Elhalel, Edwin Bremer. DSP107—a novel SIRPα-4-1BBL dual signaling protein (DSP) for cancer immunotherapy [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A076.