Abstract Myeloproliferative neoplasms (MPN) frequently harbor activating mutations in JAK/STAT signaling genes - JAK2, CALR, or MPL ; and treatment with the JAK1/2 ATP competitive inhibitor ruxolitinib (Rux) is standard for symptomatic disease. However, emerging clinical observations suggest that Rux treatment can lead to the selective expansion of subclones with activating RAS mutations in patients with myelofibrosis. These mutations are linked to therapy resistance, disease progression, and leukemic transformation. Mechanistically, MAPK pathway activation has been associated with reduced JAK2 expression, enhanced oncogenic potential of RAS mutations, and in the escape of RAS-mutated hematopoietic cells from oncogene-induced senescence (Maslah, et al. , 2025). It has also been recently shown that in pancreatic ductal adenocarcinoma (PDAC) —an almost ubiquitously RAS-driven disease—a combination therapy using the RAS (ON) multi-selective inhibitor (RMC-7977) and the CDK4/6 inhibitor palbociclib can drive surviving tumor cells into a senescent-like state. This was seen in both transplantable and genetically engineered mouse models of PDAC (Broderick, et al. , 2025). We hypothesize that combining CDK4/6 inhibition with RAS (ON) inhibition will help overcome RAS-mediated resistance to JAK2 inhibitors, such as Rux. In order to establish a model of resistance to JAK inhibitors, we will transduce JAK2- mutant cell lines (HEL, UKE1, SET2) with an exogenous GFP- Nras Q61K-expressing vector and expose them to ruxolitinib treatment over a long period of time, with non-transduced cells as a control. We will test targeted combinations of RAS inhibition (RMC-7977), CDK4/6 inhibition (abemaciclib), and JAK inhibition (Rux) to assess the effect of these treatments on cell viability, induction of senescence and apoptosis, and cell cycle distribution. Additionally, we will use CD117+ cells from Jak2 mutant MPN mice with or without Ras mutations cultured in methylcellulose and BMEC co-cultures and treated with the above combinations. Clonogenic potential will be assessed through colony forming assays and serial replating efficiency. It is expected that Jak2Nras mutant mice will outcompete Jak2-mutant cells in these co-cultures; however, the presence of a CDK4/6 inhibitor will reverse this. These readouts will provide a mechanistic insight into how combined pathway targeting can counteract or reverse RAS-driven resistance phenotypes. Citation Format: Subyeta Chowdhury, Amritha Varshini Hanasoge Somasundara, Celina Huang, Ross Levine, Raajit Rampal. Understanding and Overcoming RAS-pathway Mediated Therapeutic Resistance and Disease Progression in Myeloproliferative Neoplasms abstract. In: Proceedings of the AACR Special Conference in Cancer Research: RAS Oncogenesis and Therapeutics; 2026 Mar 5-8; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Res 2026;86 (5Suppl₁): Abstract nr B040.
Chowdhury et al. (Thu,) studied this question.