Clinical and Molecular Characterization of Disease Progression in Patients (Pts) with Low-Risk Myelofibrosis (MF) Enrolled in the MOST Study

Background: Pts with low-risk MF have a relatively good prognosis; however, few data exist to identify pts likely to experience disease progression. The Myelofibrosis and Essential Thrombocythemia Observational Study (MOST; NCT02953704) is a prospective observational study that enrolled pts with low-risk MF defined by DIPSS criteria. Here, we identify clinical and molecular characteristics of pts who progressed from low-risk to intermediate-/high-risk disease.

Methods: MOST enrollment criteria (confirmed locally and centrally) included a physician-reported MF diagnosis (primary MF, post-polycythemia vera, or post-essential thrombocythemia) and low-risk status by DIPSS (except age >65 y). Biospecimens were optionally collected every 12 months. Of 232 pts with MF enrolled, 158 met these criteria by central review, had available biospecimens, and were included in this analysis. MF progression was defined by meeting ≥1 of the following criteria during the study: hemoglobin <10 g/dL, presence of constitutional symptoms (weight loss, fever, fatigue, or sweats), platelets <100×10⁹/L, new/worsening splenomegaly, blasts >1%, white blood cell (WBC) count >25×10⁹/L, death due to disease progression, >1 red blood cell transfusion, or leukemic transformation. Uni- and multivariate logistic regressions were performed to assess risk factors of progression. RNA sequencing (RNA-seq) was performed on samples from 107 pts with available RNA specimens (42 pts with and 65 pts without progression) using Illumina NovaSeq™ X Plus (100 paired-end [PE] base reads with >50 M PE reads per sample). RNA-seq data were analyzed using differential gene expression analysis (DESeq2), fast gene set enrichment analysis (FGSEA), and cell type-specific enrichment analysis (CSEA). Driver mutation genotyping was performed on DNA if RNA was unavailable.

Results: During the study, 97/158 pts (61.4%) had evidence of MF progression from low-risk to intermediate- or high-risk disease. Baseline demographics were similar between pts with vs without disease progression. Enrollment duration was similar in pts with and without progression (median [range], 53.6 months [42-68] vs 50.7 months [42-62]). Driver mutation genotyping identified 111 pts with JAK2 V617F, 24 with CALR, 9 with MPL, and 14 triple-negative pts; of whom, 61.3%, 54.2%, 77.8%, and 64.3% had evidence of MF progression.

Univariate analysis of baseline covariates, including driver mutations, identified only WBC >11×10⁹/L at enrollment as significantly associated with increased progression risk (OR [95% CI], 2.98 [1.31-6.78]; P=0.009). All other covariates assessed (age, body mass index, time from MF diagnosis to enrollment, sex, race, driver mutation, hematocrit >0.45 L/L, and platelets >400×10⁹/L at enrollment) were not significant predictors (all P>0.05). WBC >11×10⁹/L remained significant in multivariate analyses (OR [95% CI], 2.88 [1.15-7.23]; P=0.024).

RNA-seq identified 104 upregulated and 20 downregulated genes in pts with and without progression, respectively. Of note, CD34 and MMP8 were upregulated in pts with progression. CD34 is a marker of myeloid stem/progenitor cells with a function in stem cell activity. MMP8, a collagenase largely secreted by neutrophils and monocytes, is considered an important mediator of microenvironment remodeling.

FGSEA identified 18 significantly upregulated pathways in pts with MF progression, including interferon-α and interferon-γ response (P<0.05). Using the results from differential gene expression analysis, CSEA identified spleen hematopoietic stem cells and spleen intermediate monocytes as the most differentially expressed markers at the cellular tissue level, while hematopoietic stem cells and intermediate monocytes were identified as the most differentially expressed cell types (P<0.05).

Conclusions: This analysis of prospective data from MOST found the majority of pts with low-risk MF experienced disease progression over a median ~54 months of follow-up. Multivariate regression analysis identified WBC >11×10⁹L as a significant risk factor of progression. RNA-seq analysis demonstrated differences in gene expression patterns of relevant cell types in pts with vs without MF progression. Several key genes were identified, including overexpression of CD34 and MMP8, that may provide predictive biomarker signatures of progressive disease and inform therapeutic targeting.