C019 | INIBITION OF PI3KΔ BY ROGINOLISIB (IOA-244) AS A POTENTIAL THERAPEUTIC STRATEGY FOR MYELOFIBROSIS

Due to its clonal stem cell origins and the resulting dysregulation of the JAK/STAT pathway, myelofibrosis (MF) treatment has undergone a paradigm shift with the development of JAK-inhibitors. Other pathways that regulate cell growth, survival, proliferation, and metabolism, such as the PI3K/AKT/mTOR pathway, have been shown to be implicated as well. Owing to the toxicities of current PI3K inhibitors, isoform-selective PI3K inhibitors are being developed. This study reports the investigation of Roginolisib (IOA-244), a first-in-class non-ATP-competitive PI3Kδ inhibitor, for the treatment of MF, both alone and in combination with Ruxolitinib (Ruxo). First, we evaluated PI3Kδ isoform expression in CD34+ cells from cord blood (CB, as controls) and peripheral blood of both naive- or under-Ruxo treatment MF patients using western blot analysis (n=2 per group). The PI3Kδ isoform was found to be expressed in all patient samples, regardless of Ruxo treatment status. In contrast, it was not detected in the CB samples. Next, MF CD34+ cells were used in clonogenic assays with increasing doses of IOA-244 (0, 0.5, 1, and 3 µM), alone or in combination with Ruxo, to evaluate the growth of erythroid (BFU-E) and granulocyte-macrophage (CFU-GM) progenitor cells. IOA-244 significantly reduces colony formation, even though it appears to be 2.7-fold more effective against CFU-GM (IC50, 0.79±0.38 µM) than BFU-E (IC50, 2.12±0.03 µM). Finally, we aimed to ascertain the synergistic inhibition of BFU-E and CFU-GM by suboptimal doses of IOA-244 (0.5-1 µM) and Ruxo (50-100 nM). CFU-GM colony formation was reduced to 73%±7.7%, 81%±1.9%, 81%±2%, and 86.7%±1%, respectively, for the combination of 0.5 µM IOA-244+50 nM Ruxo, 0.5 µM IOA-244+100 nM Ruxo, 1 µM IOA-244+50 nM Ruxo, and 1 µM IOA-244+100 nM Ruxo. However, also BFU-E colony formation was reduced to 65%±12%, 75%±9.5%, 69%±13.4%, and 81.9%±3.3%, respectively, using the same drug combination. Drug combination effects were analyzed using the Chou-Talalay method to calculate the combination index (CI), where CI<1 indicates synergy. The aforementioned combinations all resulted in synergistic activity, with CI values of 0.25, 0.25, 0.5 and 0.5 for CFU-GM and 0.36, 0.51, 0.37 and 0.5 for BFU-E. In summary, our findings demonstrate efficacy of IOA-244 in MF CD34+ cells; furthermore, the concurrent inhibition of PI3Kδ and JAK2 pathways by IOA-244 and Ruxo leads to enhanced activity in in-vitro MF models.