S192: SINGLE-CELL RNA PROFILING OF MYELOFIBROSIS PATIENTS REVEALS PELABRESIB-INDUCED DECREASE OF MEGAKARYOCYTIC PROGENITORS AND NORMALIZATION OF CD4+ T CELLS IN PERIPHERAL BLOOD

Abstract

Background: Abnormal differentiation of the megakaryocytic (MK) lineage in conjunction with overproduction of proinflammatory cytokines (Ck), resulting in bone marrow fibrosis, anemia, extramedullary hematopoiesis and often hepatosplenomegaly, are the main biological and clinical characteristics of myelofibrosis (MF). Bromodomain and extraterminal domain (BET) proteins play a key role in the regulation of neoplastic myeloproliferation and proinflammatory Ck production. Pelabresib (CPI-0610) is a potent, selective BET inhibitor under investigation in MF patients (pts) as monotherapy and in combination with the Janus kinase inhibitor (JAKi) ruxolitinib (RUX) in the ongoing MANIFEST Phase 2 study (NCT02158858). Arm 1: pelabresib as monotherapy in RUX-intolerant, ineligible or refractory MF pts; Arm 2: pelabresib in combination with RUX as an ‘add-on’ in MF pts with suboptimal response to RUX; Arm 3: pelabresib combination therapy with RUX in JAKi-naïve MF pts. Pelabresib as monotherapy and in combination with RUX improved spleen volume reduction, symptoms and hemoglobin levels (Talpaz M, ASH 2020). The effects of pelabresib on hematopoietic stem/progenitor cells (HSPCs) and immune cell populations in MF pts are presented here. Aims: We aimed to characterize the cellular composition and transcriptional alterations occurring in peripheral blood (PB) obtained from MF pts enrolled in the MANIFEST trial. Methods: We performed single-cell RNA sequencing on 234,904 CD34+ HSPCs and 135,970 CD34- mature PB cells (Figure). Cells were obtained from a random pool of 20 pts (Arm 1 n=5, Arm 2 n=8, Arm 3 n=7), which included a baseline sample (BL) and samples collected during treatment (range: C3 through C12) for each pt. Mobilized PB cells from healthy donors (HD) were used as a control (n=11). Results: Analysis of CD34+ HSPCs at BL demonstrated a statistically significant increase of MK, neutrophilic and erythroid progenitors in MF pts compared with HDs and decreased numbers of myeloid and B cell lineage progenitors. Pelabresib as monotherapy and in combination with RUX led to a significant reduction of MK-, neutrophilic and erythroid progenitors as compared with BL in all pts analyzed. Analysis of CD34- cells from MF pts identified a significantly lower proportion of CD4+ T cells and increased numbers of erythroid cells at BL compared with HD. Individual pts also exhibited reduction in natural killer cells and CD16+ monocytes as well as elevated MK lineage cells. Pelabresib as monotherapy and in combination with RUX increased the proportion of CD4+ T cells, and more importantly, reduced MK lineage cells compared with BL in both treatment-naïve and RUX relapsed/refractory (r/r) pts. In MF pts at BL, a larger spleen volume was observed in pts with lower numbers of CD4+ T cells and increased numbers of MK- and myeloid CSF3R+ cells. Updated results of the cell composition analysis and transcriptional reprogramming during pelabresib treatment, as well as correlation with effects on cytokine plasma levels and clinical efficacy data, will be presented. Image:Summary/Conclusion: The single-cell profiling of a subset of MF pts enrolled in the MANIFEST study suggests that pelabresib alone and in combination with RUX induces an improvement of the myeloid-lymphoid imbalance in both JAKi-naïve and r/r MF pts. The observed effects of pelabresib on HSPCs from MF pts confirm the ex vivo activity on erythroid and MK differentiation of stem cells (Keller P, EHA 2021; Verstovsek S, ASH 2021) and implies a potential disease-modifying effect warranting further investigation.