Targeted CD47 checkpoint blockade using a mesothelin-directed antibody construct for enhanced solid tumor-specific immunotherapy
Anna Reischer, Alexandra Leutbecher, Björn Hiller, Enrico D. Perini, Kieron White, Alejandra Hernández-Cáceres, Alexandra Schele, Benjamin Tast, Lisa Rohrbacher, Lis Winter, Bastian Czogalla, Sven Mahner, Heinrich Flaswinkel, Heinrich Leonhardt, Lorenza Wyder, Christian Wichmann, Denis Maenner, Fabian Trillsch, Mirjana Kessler, Karl‐Peter Hopfner, Nadja C. Fenn, Marion Subklewe
Abstract
Abstract The immune checkpoint CD47 is highly upregulated in several cancers as an innate immune escape mechanism. CD47 delivers a “don’t eat me” signal to its co-receptor signal regulatory protein α (SIRPα), thereby inhibiting phagocytosis. Blocking the CD47–SIRPα axis is a promising immunotherapeutic strategy against cancer. However, early trial data has demonstrated on-target off-leukemia toxicity. In addition, the ubiquitous expression pattern of CD47 might contribute to an antigen sink. In this study, we combined low-affinity CD47 checkpoint blockade and specific tumor targeting in a multivalent and multifunctional antibody construct to prevent CD47-related toxicities. First, we established a local inhibitory checkpoint monoclonal antibody (LicMAb) by fusing two N -terminal extracellular domains of SIRPα to a full-length anti-human mesothelin (MSLN)-IgG1 antibody, a well-described tumor-associated antigen in epithelial ovarian cancer (EOC) and pancreatic ductal adenocarcinoma (PDAC). Next, we evaluated the SIRPα-αMSLN LicMAb for mediating a tumor-restricted immune response as observed by antibody-dependent cellular cytotoxicity (ADCC) and phagocytosis (ADCP). Our data validates CD47 and MSLN as highly upregulated targets expressed on various solid cancer entities, particularly EOC. We show tumor-specific binding and CD47 blocking by the SIRPα-αMSLN LicMAb even in the presence of healthy CD47-expressing cells. Furthermore, the LicMAb induces NK-cell-mediated cytotoxicity and improves phagocytosis of EOC and PDAC tumor cells. Moreover, cell death in EOC-derived organoids was specifically LicMAb-driven. Hence, the SIRPα-αMSLN LicMAb combines a tumor-restricted blockade of the CD47–SIRPα axis with a specific antitumor response while preventing on-target off-tumor toxicities. Our data supports the multifunctional SIRPα-αMSLN LicMAb as a promising approach to treating solid tumors. Graphical abstract The local inhibitory checkpoint monoclonal antibody (LicMAb) binds mesothelin (MSLN) with high affinity and simultaneously blocks CD47 on MSLN-expressing tumor cells to inhibit the “don’t eat me” signal. CD47 is blocked by the fused extracellular SIRPα domain that intrinsically has a low affinity. Furthermore, the SIRPα-αMSLN LicMAb is based on a human IgG1 backbone to provide an Fc receptor (FcR)-activating stimulus to enable direct NK-cell-mediated killing by granzyme B (GrzB) and perforin secretion, and an additional pro-phagocytic signal to phagocytic cells, such as macrophages (MØ). This leads to tumor-restricted antibody-dependent cellular cytotoxicity (ADCC) and antibody-dependent cellular phagocytosis (ADCP) of cancer cells. This scheme was created with BioRender (BioRender.com/g77u465).