Pacritinib Effectively Inhibits Pro-Survival Signaling and Overcomes BTKCys481Ser-Mediated Resistance in MYD88-Mutated B-Cell Malignancies
Authors: Liu Shirong , Kofides Amanda , Liu Xia , Sun Hao , Canning Alexa G , Tsakmaklis Nickolas , Guerrera Maria Luisa , Patterson Christopher J , Hunter Zachary R , Meid Kirsten , Gokhale Prafulla C. , Hatcher John M , Castillo Jorge J. , Sarosiek Shayna R , Treon Steven P. 
Published: December 2024

Background: Somatic mutations in MYD88 are prevalent in various B-cell malignancies, including Waldenström Macroglobulinemia (WM; 95-97%), primary CNS lymphoma (70-80%), ABC DLBCL (40%), marginal zone lymphoma (5-10%), and CLL (5-15%). These mutations drive oncogenic pro-survival signaling through cascades involving HCK-directed BTK, ERK, SYK, and IRAK1/IRAK4 activation. HCK, an SRC family member, is transcriptionally upregulated by mutated MYD88 and activated via IL-6/IL-6R/gp130/JAK2 signaling. Although BTK-inhibitors are effective against MYD88-mutated B-cell malignancies, complete responses are rare and resistance often arises due to mutations in BTKCys481. Pacritinib, an FDA-approved kinase inhibitor for myelofibrosis, targets crucial signaling pathways associated with mutated MYD88, including IRAK1, JAK2, and SRC (a homolog of HCK), potentially offering a new therapeutic option for MYD88-mutated malignancies.

Methods: We conducted comparative studies to evaluate the effects of pacritinib versus the covalent BTK-inhibitors ibrutinib and zanubrutinib on pro-survival signaling, proliferation, and survival in MYD88-mutated cell lines and primary MYD88-mutated WM cells. Additionally, we assessed pacritinib impact on BTKCys481Ser-expressing, covalent BTK-inhibitor-resistant WM and ABC DLBCL lymphoma cell models. Efficacy was also evaluated in a BTKCys481Ser TMD8 cell xenografted murine model.

Results: Pacritinib induced higher levels of apoptosis in MYD88-mutated WM (BCWM.1) and ABC DLBCL (TMD8) cells, as well as in primary (CD19+) MYD88-mutated WM cells versus either ibrutinib or zanubrutinib. The apoptosis-inducing activity was dose-dependent, with significant apoptosis observed at 0.5 µM, which was well within pharmacologically achievable concentrations of pacritinib. Pacritinib suppressed JAK2 and IRAK1 activation and robustly inhibited p-HCK (Y411) and its downstream partner p-BTK (Y223) in MYD88-mutated BCWM.1 and TMD8 cells. Additionally, synergy in inhibiting cell growth was observed between pacritinib and covalent BTK inhibitors, and the BCL-2 inhibitor venetoclax in MYD88-mutated cell lines and primary WM cells. Pacritinib, alone or combined with venetoclax, induced high levels of apoptosis in BTKCys481Ser-expressing, BTK inhibitor-resistant MYD88-mutated WM and ABC DLBCL lymphoma cells. In a BTKCys481Ser TMD8 cell xenograft murine model, pacritinib treatment significantly suppressed tumor growth versus vehicle control or ibrutinib.

Conclusions: Pacritinib effectively targets mutated MYD88 pro-survival signaling pathways and shows superior apoptotic activity in MYD88-mutated WM and ABC DLBCL cells over covalent BTK-inhibitors. Pacritinib also showed robust synergistic interactions with BTK and BCL-2 inhibitors and overcame covalent BTK-inhibitor resistance associated with mutated BTKCys481Ser both in vitro and in vivo in TMD8 Cys 481 mutated xenograft murine model. Our studies provide a framework for the investigation of pacritinib in MYD88-mutated lymphomas. Based on these findings, a phase II clinical trial investigating pacritinib in relapsed/refractory WM is being initiated.