CC-122 Exhibits Greater Preclinical Activity in Mantle Cell Lymphoma Than Lenalidomide through a Combination of Direct Cell-Autonomous and Increased Antibody Dependent Cell-Mediated Cytotoxicity

Introduction: CC-122 and lenalidomide (len) bind the E3 ubiquitin ligase CUL4-RBX1-DDB1-CRBN (CRL4^CRBN) resulting in degradation of the transcription factor Aiolos, leading to direct anti-lymphoma activity and T and NK cell activation. CC-122 degrades Aiolos at a faster rate and to a greater extent compared to len. Len is currently indicated for the treatment of relapsed/refractory (R/R) mantle cell lymphoma (MCL) in the United States and European Union. CC-122 is in development for DLBCL, FL and CLL in combination with the anti-CD20 monoclonal antibodies, rituximab (Rtx) and obinutuzumab (GA101). We compared the ability of len and CC-122 to effect cell autonomous activity and enhance antibody dependent cell-mediated cytotoxicity (ADCC) in pre-clinical models of MCL.

Methods: Proliferation was measured by ³H labeling. Apoptosis was measured by Annexin V/ToPro-3 flow cytometry. ADCC was measured by a 4 hour co-incubation of Rtx or GA101 labeled cells with stimulated PBMC treated with DMSO, len or CC-122 for 3 days followed by apoptosis assay. Inducible shRNA targeting luciferase or Aiolos were activated with 10 ng/ml doxycycline for 7 days followed by apoptosis assay.

Results: Lenalidomide treatment (10μΜ) for 5 days resulted in a 31% and 49% decrease in proliferation of two of the six MCL cell lines investigated, whereas CC-122 treatment (1.25μΜ) decreased proliferation in four MCL cell lines by 37-81%. There was no increase in Annexin V and ToPro-3 staining in MCL cells treated with len (0.1-10μΜ) for 7 days. By contrast, CC-122 treatment (0.1-10μΜ) reduced viability in four of six cell lines examined by 32-95%. Examination of the biochemical activity of each drug in Mino and Rec-1 cells demonstrated CC-122 (0.1-10μΜ) induced rapid Aiolos degradation at 6 hours (33-88% and 38-85%, respectively) compared to 10μΜ len (27% and 25%, respectively). In Jeko-1 cells, two distinct doxycycline inducible shRNA targeting Aiolos led to 56-97% decreased Aiolos protein expression. Furthermore, shRNA targeting Aiolos led to a 2- to 3-fold increase in apoptosis relative to shLuciferase. In ADCC assays of Z138 and Granta-519 coated with Rtx (1μg/ml) or GA101 (1μg/ml), CC-122 (10-100nM) was more potent than len (0.1-1μΜ). CC-122 treatment of PBMC increased Rtx labeled Z138 and Granta-519 apoptosis (39-59% and 36-48%, respectively) compared to len (24-35% and 33-40%, respectively), versus vehicle controls (13% and 13%, respectively). Additionally, CC-122 treatment increased GA101 mediated ADCC of Z138 and Granta-519 (60-76% and 59-67%, respectively) compared to len (49-61% and 55-63%, respectively), versus vehicle controls (35% and 41%, respectively).

Conclusions: CC-122 treatment of MCL cells resulted in considerable cell autonomous activity in in vitro proliferation and viability assays compared to len. Specific targeting of Aiolos, a substrate which is rapidly degraded by CC-122, through inducible shRNA results in increased levels of apoptosis compared to shLuciferase controls. Additionally, the combination of CC-122 with either Rtx or GA101 in in vitro co-culture ADCC assays resulted in greater apoptosis than len combined with either antibody. The combination of both enhanced cell-autonomous activity and α-CD20 mediated ADCC provide a rational combination strategy for CC-122 development in R/R MCL.