Abstract 3956: MYC inhibition overcomes IMiD resistance in heterogeneous multiple myeloma populations
Authors: Davis Lorraine N. , Walker Zachary J. , Ohlstrom Denis , Stevens Brett M. , Forsberg Peter A. , Mark Tomer M. , Jordan Craig T. , Sherbenou Daniel W. 
Published: June 2022

Abstract
Introduction: Multiple myeloma (MM) is a plasma cell malignancy that remains incurable, with patients enduring multiple relapses and development of drug resistance. Immunomodulatory drugs (IMiDs) are critical anti-MM agents. IMiDs act by inducing CRBN-dependent proteasomal degradation of the transcription factors IKZF1 and IKZF3, which leads to IRF4 and MYC downregulation (collectively termed the “Ikaros axis”). Although CRBN aberrations occur, whether they are a functional mechanism of resistance in patients remains unclear. Based on the importance of CRBN in IMiD response, we hypothesized that IMiD treatment fails to downregulate the Ikaros axis in IMiD-resistant MM cells.
Methods: To measure IMiD-induced Ikaros axis downregulation, we designed an intracellular flow cytometry assay that measured relative IKZF1, IKZF3, IRF4 and MYC protein levels in MM cells following ex vivo pomalidomide (Pom) treatment. We established this assay using IMiD-sensitive parental and dose-escalated Pom resistant MM1S and H929 cell lines before utilizing it in patient samples (isolated mononuclear cells). To assess the Ikaros axis in the context of MM intratumoral heterogeneity, we used mass cytometry to simultaneously characterize MM subpopulations in patient samples. Lastly, we determined ex vivo drug sensitivity in patient samples via flow cytometry.
Results: Our hypothesis was supported in MM cell lines, as sensitive parental lines showed a significant decrease in all Ikaros axis protein levels following Pom treatment, while resistant lines showed no IMiD-induced decrease in any Ikaros axis protein. However, when assessed in CD38+CD138+ cells from patient samples, Pom treatment caused a significant decrease in IKZF1, IKZF3 and IRF4 regardless of IMiD sensitivity. Mass cytometry in patient samples revealed that individual Ikaros axis proteins were differentially expressed between subpopulations. When correlating this with ex vivo Pom sensitivity of MM subpopulations, we observed that low IKZF1 and IKZF3 corresponded to Pom resistance. Interestingly, most of these resistant populations still expressed MYC. We therefore assessed whether IMiD resistant MM was MYC dependent by treating Pom resistant MM cells with MYCi975. In 88% (7/8) of patient samples tested, IMiD resistant MM cells were sensitive to MYC inhibition.
Conclusions: Our findings did not support our initial hypothesis, as IMiD-induced IKZF1 and IKZF3 degradation remains intact in IMiD resistant MM cells from patient samples. However, our data support a mechanism where the Ikaros axis no longer drives MYC expression in IMiD-resistant MM. Therefore, we propose that the critical mediator of IMiD resistance is conversion to a cell state where MYC expression is Ikaros axis independent. This suggests targeting MYC directly or via the as yet uncharacterized mechanism controlling MYC may be an effective strategy to eradicate IMiD resistant MM.
Citation Format: Lorraine N. Davis, Zachary J. Walker, Denis Ohlstrom, Brett M. Stevens, Peter A. Forsberg, Tomer M. Mark, Craig T. Jordan, Daniel W. Sherbenou. MYC inhibition overcomes IMiD resistance in heterogeneous multiple myeloma populations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3956.