Abstract 7225: Combination JAK1/2 and CDK8/19 inhibition demonstrates enhanced efficacy in myeloproliferative neoplasms
Authors: Zaroogian Zachary , Adamczyk Elżbieta , Moszyńska Adrianna , Obacz Marta , Pakulska Urszula , Durham Benjamin , Mazan Milena , Mowla Shoron , Wnęk Katarzyna , Somasundara Amritha Varshini Hanasoge , Barczuk Beata , Gołas Aniela , Levine Ross , Rzymski Tomasz , Rampal Raajit 
Published: March 2024

Abstract
Myeloproliferative neoplasms (MPNs) are characterized by aberrant activation of the JAK-STAT pathway. Ruxolitinib (RUX) is a JAK1/2 inhibitor used to treat myelofibrosis (MF) and hydroxyurea-resistant/intolerant polycythemia vera. While RUX has demonstrated important clinical benefits, a large proportion of patients have persistent disease manifestations despite therapy. CDK8 regulates phosphorylation of STAT proteins in a RUX-independent manner; thus, CDK8 inhibition may alter downstream STAT target gene expression. RVU120 is a highly selective and potent CDK8/19 inhibitor that inhibits STAT1/5 phosphorylation. We hypothesized that the combination of RUX and RVU120 would act in a cooperative manner to attenuate JAK-STAT signaling and reduce MPN phenotypes in vivo.
Using JAK2V617F mutant leukemic cell lines, we identified dose-responsive changes in viability with RUX and RVU120 in all cell lines. Pharmacodynamic analysis validated that STAT5 protein phosphorylation was decreased by RUX/RVU120, although at different phosphorylation sites by RUX (pTYR694) and RVU120 (pSER726/731); exposure to RUX+RVU120 inhibited phosphorylation at both sites. Notably, analysis of combinatorial effects of RUX+RVU120 on viability demonstrated drug synergies (combinatorial index &lt;1) in RUX-naïve and resistant SET-2 cells. Nascent RNA expression data exploring JAK-STAT and mediator-related pathways of pathogenesis will be presented at the meeting.
For in vivo studies, we used two previously-described murine models of MF and PV. Briefly, retrovirally-transfected (MSCV-MPLW515L-IRES-GFP) or Jak2V617F-conditional knock-in bone marrow (BM) was intravenously injected into lethally-irradiated congenic recipients. Mice were randomized and treated for several weeks with vehicle (VEH), single-agent RUX/RVU120, or RUX+RVU120 arms. Spleen weights between VEH and RUX/RVU120 treated mice at time of terminal sacrifice were significantly reduced across both models (p⇐0.009). In MPLW515L mice, WBCs and PB GFP% were significantly reduced (p&lt;0.002) and histopathology demonstrated near-elimination of BM reticulin fibrosis and improved trilineage hematopoiesis between RUX+RVU120 and RUX arms. Data on overall survival, cytokine profiles, and the impact of RVU120 on dynamic gene expression will be presented at the meeting.
The combination of RVU120 and RUX demonstrated biochemical synergy and differential inhibition of STAT5 phosphorylation in vitro. Further, in vivo treatment with RVU120/RUX+RVU120 resulted in significant reductions of disease manifestation (splenomegaly, WBC, fibrosis scoring, hematopoiesis) when compared to VEH/RUX. These data nominate JAK1/2 and CDK8/19 inhibition as a potential novel therapeutic strategy in MPNs; further work on nascent RNA expression and cytokine profiles will potentially elucidate additional mechanisms of action.
Citation Format: Zachary Zaroogian, Elżbieta Adamczyk, Adrianna Moszyńska, Marta Obacz, Urszula Pakulska, Benjamin Durham, Milena Mazan, Shoron Mowla, Katarzyna Wnęk, Amritha Varshini Hanasoge Somasundara, Beata Barczuk, Aniela Gołas, Ross Levine, Tomasz Rzymski, Raajit Rampal. Combination JAK1/2 and CDK8/19 inhibition demonstrates enhanced efficacy in myeloproliferative neoplasms [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7225.