ABSTRACT: Rising blast-percentage or secondary (s) AML transformation (sAML) in MPNs leads to JAK inhibitor (JAKi) therapy-resistance and poor survival. Here, we demonstrate that the CDK7 inhibitor (CDK7i) SY-5609 treatment depletes phenotypically-characterized post-MPN-sAML stem/progenitor cells. In the cultured post-MPN sAML SET2 and HEL as well as patient-derived (PD) post-MPN-sAML cells, SY-5609 treatment inhibited growth and induced lethality, while sparing normal cells. RNA-Seq analysis following SY-5609 treatment demonstrated reduced mRNA expressions of MYC, MYB, CDK4/6, PIM1, and CCND1, but increased mRNA levels of CDKN1A and BCL2L1. Mass spectrometry of SY-5609-treated MPN-sAML cells also demonstrated reduced c-Myc, c-Myb, PIM1, and CDK4/6 but increased p21, caspase 9 and BAD protein levels. CRISPR-mediated CDK7 depletion also reduced viability of HEL cells. CyTOF analysis of SY-5609-treated PD, post-MPN-sAML stem/progenitor cells showed reduced c-Myc, CDK6 and PU.1, but increased protein levels of CD11b, p21 and cleaved Caspase 3. Co-treatment with SY-5609 and ruxolitinib was synergistically lethal in HEL, SET2 and PD post-MPN-sAML cells. A CRISPR screen in SET2 and HEL cells revealed BRD4, CBP and p300 as co-dependencies with SY-5609 treatment. Accordingly, co-treatment with SY-5609 and the BETi OTX015 or pelabresib or with the CBP/p300 inhibitor GNE-049 was synergistically lethal in MPN-sAML cells (including those exhibiting TP53 loss). Finally, in the HEL-Luc/GFP xenograft model, compared to each agent alone, co-treatment with SY-5609 and OTX015 reduced post-MPN-sAML burden and improved survival without inducing host toxicity. These findings demonstrate promising preclinical activity of the CDK7i-based combinations with BETi or HATi against advanced-MPNs, including post-MPN-sAML.
