Dissecting the Mechanisms of Activity of SLAMF7 and the Targeting Antibody Elotuzumab in Multiple Myeloma

Abstract

BackgroundSignaling lymphocytic activation molecule family 7 (SLAMF7) is a cell surface receptor which is expressed at high levels in multiple myeloma (MM) cells and serves as a therapeutic target in this disease. Elotuzumab, a humanized monoclonal antibody targeting SLAMF7, has shown promising activity, particularly in combination with lenalidomide, in MM patients in the clinical trial setting. While the primary mechanism of action of elotuzumab in pre-clinical in vitro studies has been shown to be antibody-dependent cellular cytotoxicity (ADCC) mediated via NK-cell CD16, other possible mechanisms of activity of elotuzumab in MM are not well defined. Specifically, the downstream signaling effects of SLAMF7 have not been delineated in myeloma cells. We set out to examine the effect of stable SLAMF7 knockdown (KD) on MM tumor development in vivo and evaluated the ability of elotuzumab to exert an anti-myeloma effect within the bone marrow niche in MM mouse models.Methods:In-vitro: KD of SLAMF7 was performed in myeloma cell line MM1s using lentiviral shRNAs targeting different regions in the SLAMF7 mRNA. Specific SLAMF7 knockdown was confirmed by reduced SLAMF7 mRNA and protein expression in comparison to cells receiving scrambled control shRNA lentivirus. Survival of SLAMF7 KD cells in comparison to scrambled control cells was assessed by MTT assay. The effect of elotuzumab treatment on the survival of MM1s cells was also assessed. In vivo: Tumor Xenograft models SLAMF7 KD; MM1S-GFP-Luc+ cells (5X106) were injected intravenously (IV) into SCID-Bg mice (n=7/group) and animals underwent bioluminescent imaging (BLI) weekly following injection. Xenograft models elotuzumab; Forty eight hours after IV injection of MM1S-GFP-Luc+ cells (5X106) elotuzumab or human IgG control antibody (both 10mg/kg) was administered via intraperitoneal (IP) injection twice weekly and mice (n=7/group) were followed for tumor burden and survival outcomes. A follow up study was performed (n=5/group) using a modified form of elotuzumab (IgG1.1mAb 10mg/kg IP) harboring mutations in the IgG1 CH2 domain which decreases ADCC via decreased Fc-γ receptor binding allowing evaluation of the CD16 mediated effect in this model.ResultsSLAMF7 KD resulted in a significant reduction in the survival of MM1s cells at 24 hours as assessed by MTT assay (P = 0.001). No significant reduction in survival was noted for the MM1s myeloma cell line following elotuzumab treatment at 24 hours in the absence of effector cells. Interestingly, knockdown of SLAMF7 resulted in a reduction in tumor burden in the SCID-Bg xenograft model as assessed by weekly BLI signal (P = 0.005 at week 6) with a consequent increase in survival for mice receiving SLAMF7 KD cells. (average 49 vs. 110 days Log Rank P<0.0001).Treatment of mice that had been administered MM1s cells and subsequently treated with elotuzumab resulted in a significant reduction in tumor burden (P = 0.001 at 5 weeks) and prolonged survival (Log Rank P = 0.0001) compared to human IgG1 control antibody treated mice (n=7/group). A follow up 3 arm study comparing IgG1.1f mAb to elotuzumab or human IgG1 control antibody (n=5/group) demonstrated no significant difference in tumor burden between the group receiving IgG1.1 mAb (no CD16 binding) and human IgG1 control antibody (P =0.8 at week 3). The difference in tumor burden between IgG1.1 mAb and elotuzumab was significant (P = 0.019 at 3 weeks) indicating that the effect on tumor burden seen in these mice was indeed CD16 dependent.ConclusionsKnockdown of SLAMF7 in MM cells results in reduced tumor burden and survival in xenograft mice, indicating that SLAMF7 potentially plays a role in tumor biology in this disease. In addition elotuzumab treatment reduces tumor burden in a xenograft model of MM with defective NK cells in a CD16 dependent manner indicating that additional mechanisms beyond ADCC may be important in mediating the anti-myeloma effect of elotuzumab. DisclosuresGlavey:BMS: Consultancy, Research Funding. Robbins:Bristol-Myers Squibb: Employment; Bristol-Myers Squibb: Equity Ownership. Ghobrial:Onyx: Advisory board Other; BMS: Advisory board, Advisory board Other, Research Funding; Noxxon: Research Funding; Sanofi: Research Funding.