Dual inhibition of DNMTs and EZH2 can overcome both intrinsic and acquired resistance of myeloma cells to IMiDs in a cereblon‐independent manner

Abstract

Thalidomide and its derivatives, lenalidomide and pomalidomide (also known as IMiDs), have significantly changed the treatment landscape of multiple myeloma, and the recent discovery of cereblon (CRBN) as their direct biological target has led to a deeper understanding of their complex mechanism of action. In an effort to comprehend the precise mechanisms behind the development of IMiD resistance and examine whether it is potentially reversible, we established lenalidomide‐resistant (‐LR) and pomalidomide‐resistant (‐PR) human myeloma cell lines from two IMiD‐sensitive cell lines, OPM2 and NCI‐H929, by continuous culture in the presence of lenalidomide or pomalidomide for 4–6 months, until acquirement of stable resistance. By assessing genome‐wide DNA methylation and chromatin accessibility in these cell lines, we found that acquired IMiD resistance is associated with an increase in genome‐wide DNA methylation and an even greater reduction in chromatin accessibility. Transcriptome analysis confirmed that resistant cell lines are mainly characterized by a reduction in gene expression, identifying SMAD3 as a commonly downregulated gene in IMiD‐resistant cell lines. Moreover, we show that these changes are potentially reversible, as combination of 5‐azacytidine and EPZ‐6438 not only restored the observed accessibility changes and the expression of SMAD3, but also resensitized the resistant cells to both lenalidomide and pomalidomide. Interestingly, the resensitization process was independent of CRBN. Our data suggest that simultaneous inhibition of DNA methyl transferases and EZH2 leads to an extensive epigenetic reprogramming which allows myeloma cells to (re)gain sensitivity to IMiDs.

Expression, mutation, and methylation of cereblon‐pathway genes at pre‐ and post‐lenalidomide treatment in multiple myeloma

“…Within the 22 paired samples examined, although a fraction of patients showed increased CRBN expression, there was a significant reduction in CRBN expression after Ld therapy. A reduction of CRBN could facilitate the non‐CRBN‐dependent process of proliferation/survival in MM cells and contribute to the poor sensitivity to IMiDs.…”

“…As there was significantly reduced CRBN expression in the post‐Ld samples, we explored the methylation status of the CRBN promoter; there was no hypermethylation in the promotor in the clinical samples. This is consistent with the observations by Dimopoulos et al, who showed no involvement of promoter methylation in the regulation of CRBN expression in Len‐resistant MM cell lines. However, the mechanism by which CRBN expression is downregulated remains to be understood.…”

Epigenetic plasticity potentiates a rapid cyclical shift to and from an aggressive cancer phenotype

“…To test whether epigenetic alterations are involved in the interconversion between SP and NSP cells, we performed Acce SssI ble assay to measure DNA methylation and chromatin accessibility in these two subpopulations. The Acce SssI ble assay was developed in our group and has been used successfully to identify potential epigenetic driver genes in renal cell carcinoma as well as epigenetic changes after epigenetic therapy . Acce SssI ble assay couples the analysis of chromatin accessibility and endogenous DNA methylation using the Infinium MethylationEPIC BeadChip array platform, requires very little input material, is cost‐effective, and uses simple and straightforward computational techniques for analysis …”

“…The AcceSssIble assay was developed in our group 24 and has been used successfully to identify potential epigenetic driver genes in renal cell carcinoma 22 as well as epigenetic changes after epigenetic therapy. 23,24,29 AcceSssIble assay couples the analysis of chromatin accessibility and endogenous DNA methylation using the Infinium MethylationEPIC BeadChip array platform, requires very little input material, is cost-effective, and uses simple and straightforward computational techniques for analysis. 22,24 Differences in global DNA methylation and chromatin accessibility levels between SP and NSP cells of J82 were calculated and plotted, and six groups (a-f ) of loci with differential methylation and/or accessibility were designated ( Figs.…”

PGC1β regulates multiple myeloma tumor growth through LDHA‐mediated glycolytic metabolism

“…However, MM remains largely incurable due to inevitable relapse and development of chemoresistance (Teoh et al ., 2014). Development of novel therapeutic approaches and targeting of abnormal cancer metabolism in molecular and cellular heterogeneity of MM may provide us with new strategies for overcoming this disease (Dalva‐Aydemir et al ., 2015; Dimopoulos et al ., 2018; Naymagon and Abdul‐Hay, 2016). …”