MAP4K2 Silencing Overcomes IMiDs-Resistance in Multiple Myeloma

Recently, mitogen-activated protein kinase kinase kinase kinase 2 (MAP4K2) has emerged as an important key regulator of the stress-activated MAPK core signaling pathways. MAP4K2, also called Germinal Center Kinase (GCK), is predominantly and highly expressed in the germinal center of B cells. Recently we have shown that MAP4K2 knockdown in K- or N-RAS mutated MM cells induces MM cell growth inhibition, associated with the downregulation of critical transcriptional factors including IKZF1/3, BCL-6, and c-MYC proteins (Li et al. Blood 2021). Importantly, MAP4K2 silencing induces IKZF1 protein degradation without affecting IKZF1 mRNA level. Furthermore, IMiDs-resistant K-RAS ^Mut MM cells are sensitive to MAP4K2 inhibition induced IKZF1 degradation and cell growth suppression, suggesting that MAP4K2 inhibition overcomes IMiDs resistances in MM.

To further validate MAP4K2 inhibition as a novel strategy to overcome IMiDs-resistance, we generated lenalidomide-resistant human myeloma cell lines. In this model, MM1S-LEN ^RES cells showed significantly decreased expression of CRBN protein compared to the parent cells. Accordingly, upon lenalidomide treatment, CRBN-mediated down-regulation of IKZF1, c-MYC, IKZF3 and IRF4 were abrogated in the MM1S-LEN ^RES cells. As expected, MM1S-LEN ^RES cells were resistant to lenalidomide induced growth inhibition in cell proliferation assay. In contrast, MAP4K2 inhibition using TL4-12 potently induced IKZF1, c-MYC, and IRF4 downregulation as well as cell proliferation inhibition, demonstrating that MAP4K2 regulates IKZF1 and cell growth independently of CRBN. These results indicate that MAP4K2 is a novel therapeutic target to overcome IMiDs-resistance MM.

Iberdomide (CC-220) is an orally available IMiD® compound under development for the treatment of relapsed/refractory multiple myeloma. Previous biochemical and structural studies demonstrated that Iberdomide binds to cereblon with a higher affinity than lenalidomide or pomalidomide. Here, we evaluated the combination effects of Iberdomide with MAP4K2 silencing in MM. Tet-on sh-MAP4K2 lentivirus were introduced into RAS ^Mut MM cells to establish the inducible MAP4K2 knockdown cells upon doxycycline treatment. To address the combination effects, Tet-on sh-MAP4K2 RAS ^Mut MM cells were treated with doxycycline and different dosages of iberdomide. We found that MAP4K2 silencing strongly increased iberdomide-induced apoptosis (iberdomide alone vs. with MAP4K2 KD: 32% vs 92). Similar, in western blot assays, MAP4K2 silencing combined with Iberdomide significantly enhanced downregulation of IKZF1, c-MYC, and IRF4 compared to the iberdomide treatment alone. These data suggest that combination of iberdomide and MAP4K2 inhibition have synergetic anti-MM effects.

Taken together, our findings demonstrate that MAP4K2 is a novel therapeutic target to bypass IMiDs resistance in RAS mutated MM. Combination of MAP4K2 inhibition with Iberdomide results in synergetic anti-cancer effects in MM, therefore could be a potent novel therapeutic regimen for patients with relapsed/refractory multiple myeloma.