Relapse remains the leading cause of treatment failure following allogeneic hematopoietic stem cell transplantation (alloHCT) for high-risk myeloid malignancies. Epigenetic dysregulation contributes to leukemogenesis and therapeutic resistance. To investigate whether pre-transplant epigenetic priming with azacitidine enhances chemosensitivity and improves alloHCT outcomes. We conducted an open-label, prospective phase II study evaluating azacitidine incorporated into reduced-intensity conditioning in patients with acute myeloid leukemia (AML) or myelodysplastic syndrome (MDS) undergoing alloHCT from matched related or unrelated donors. The primary endpoints were overall survival (OS) and progression-free survival (PFS). Secondary endpoints included relapse incidence, non-relapse mortality (NRM), and pharmacodynamic evaluation of DNA methylation changes in bone marrow CD34+ cells. Thirty-nine patients were enrolled. Most patients had AML (85%), 62% with adverse-risk AML or high/very high-risk MDS, and 23% had TP53 mutations. Neutrophil and platelet engraftment occurred in 97% of patients at median of 13 and 16 days, respectively. One-year OS and PFS were 64% and 54%, respectively, with NRM of 15% and relapse incidence of 31%. Pharmacodynamic analyses demonstrated azacitidine-induced DNA hypomethylation in bone marrow CD34+ cells, which correlated with improved survival. Higher baseline methylation levels were also associated with superior survival. Azacitidine priming prior to alloHCT is feasible and demonstrates encouraging survival outcomes in high-risk myeloid malignancies. Epigenetic priming induces measurable biologic reduction of DNA methylation that is plausibly linked to improved relapse-free survival. These findings support further evaluation in randomized studies and suggest DNA methylation may serve as a predictive biomarker of response.

Pulmonary Hypertension (PH) is a known complication of myeloproliferative neoplasms (MPN) with estimated prevalence as high as 50%. Patients with Polycythemia Vera (PV) report a wide spectrum of symptoms that significantly overlap with those reported by patients with PH. Yet, it is not known how PH affects outcomes and survival in patients with PV. To address this gap, we investigated the impact of echocardiogram (ECHO) PH risk on survival of patients with PV from our large single center cohort. Of 637 patients with PV, 134 had at least one ECHO and were included for analysis. Overall survival (OS) did not differ between patients who had or did not have ECHO. PH risk was established based on tricuspid regurgitation jet velocity. Kaplan-Meier analysis showed that high PH risk is associated with shortened survival compared to mild PH risk (median survival 1.7 vs 3.7 years) or normal PH risk (median survival not yet reached). Cox Proportional Hazard Models found high PH risk was associated with >3-fold increased risk of death, independent of age and thrombosis history. Logistic regression identified age (odds ratio 6.9) and duration of PV diagnosis (odds ratio 5.4) as significant risks for PH. Based upon these results and receiver operator characteristic optimization, we recommend echocardiographic screening for patients with PV older than 70 years or with duration of PV longer than 8 years. Further studies inclusive of invasive hemodynamics, advanced CV imaging, and MPN-associated biomarkers are needed to best characterize this Group 5 PH patient population for therapeutic interventions.

Background: Janus Kinase (JAK) inhibitors are the current standard of care for patients (pts) with myelofibrosis (MF). However, many pts may not achieve spleen volume reduction (SVR) or total symptom score (TSS) response after frontline treatment, and most pts with relapsed/refractory (R/R) MF lack adequate responses. Momelotinib (MMB), a recently approved JAK/ACVR1 inhibitor for MF pts with anemia, showed symptom and spleen responses in about 25% pts in R/R setting. Combination strategies of JAK inhibitor and agent with unique mechanism of action and minimal overlapping toxicities (e.g. cytopenias) are needed to improve response rates in MF. PIM1 expression is upregulated in MF CD34 cells. In preclinical models, PIM1 knockout (KO) was shown to prevent MF progression without affecting PLT counts, whereas pan-PIM KO caused thrombocytopenia (TCP). Nuvisertib (NUVI, TP-3654), an oral investigational highly selective PIM1 kinase inhibitor, alone and in combination with ruxolitinib (RUX) showed spleen size reduction and bone marrow (BM) fibrosis improvement in JAK2V617F and MPLW515L MF mouse models. Preliminary data from the ongoing Phase 1/2 study in R/R MF pts with PLT count ≥25 x 109/L showed that NUVI monotherapy was well tolerated with limited myelosuppression, and clinical activity including SVR and TSS responses strongly correlating with cytokines modulation, and hemoglobin (Hgb), PLT, and BM improvement. Preclinical and monotherapy clinical data support the development of NUVI + MMB combo in MF. Methods: The global Phase 1/2 study evaluates the safety and efficacy of NUVI + MMB combo in pts with MF (NCT04176198, Arm 3). Key eligibility criteria include primary or secondary MF, previously treated with JAK inhibitor, DIPSS intermediate or high-risk MF, Hgb <10 g/dL, PLT ≥50 x 109/L, splenomegaly (≥450 cm3 by imaging), and ≥2 measurable symptoms with each score ≥3 or a total average score of ≥10 per MFSAF v4. The study aims to identify the RP2D of NUVI when given with MMB, and to assess the safety, clinical activity (SVR, TSS improvement), and PK and PD markers (cytokine, BM fibrosis etc.). Results: Here we present the first ever combination data of MMB in MF. As of 29 May 2025, total 18 pts enrolled in 4 dose levels of NUVI BID at 240 mg (n=4), 360 mg (n=8), 480 mg (n=5) and 720 mg (N=1) + MMB 200 mg QD using the BLRM dose escalation. At baseline, median age 75 years (range 51, 82); TSS 29 (9, 37); spleen volume 1370 cm3 (614, 4250); Hgb 9.1 g/dL (7.9, 10.1; 50% pts required transfusion); and PLT 196 x 109/L (81, 601). All pts received prior JAK inhibitor, and 53% pts had high molecular risk mutation. Median treatment duration of NUVI + MMB combo was 21 weeks (1, 30), and 13 of 18 (72%) pts were on treatment. One DLT of Grade 4 TCP without any bleeding occurred in NUVI 360 mg BID + MMB 200 mg QD dose. Treatment-related adverse events (TRAEs) occurring in ≥20% of pts were diarrhea, nausea, and TCP. Grade ≥3 TRAE occurring in ≥2 pts included TCP (n=2; 1 pt had baseline TCP). Mean Hgb and PLT remained stable throughout the 24-week treatment. Emerging NUVI + MMB combo safety data was generally consistent with NUVI monotherapy data. 5 pts in the NUVI 360 mg BID + MMB 200 mg QD dose completed ≥24 weeks of treatment and were considered efficacy evaluable. TSS improvement at WK24 was observed in all 5 patients (median change -65%, range -38% to -72%); 3 of 5 (60%) pts showed ≥50% TSS reduction. In addition, absolute reduction was observed in all 7 symptom parameters, including >50% reduction in mean fatigue score at WK24. 2 of 5 (40%) pts showed ≥25% SVR at WK24. Decreased EN-RAGE and increased adiponectin were observed in all 5 pts, consistent with NUVI monotherapy findings where modulation of these cytokines strongly correlated with TSS50, individual symptoms and SVR25 responses. Anemia improvement was observed in 2 of 5 (40%) pts during 24 weeks of treatment: 1 pt showed Hgb response (defined as mean ≥1.0 g/dL increase for ≥12 weeks without transfusion), and 1 pt achieved a >50% reduction in transfusions. Dose escalation is ongoing, and updated data will be presented.Conclusions: NUVI + MMB combo appeared to be well tolerated. Preliminary data showed early clinical activity including 60% TSS50 response and absolute symptom improvement, 40% SVR25 response, cytokine modulation and anemia improvement in R/R MF pts with anemia. Preliminary data supports further development of NUVI + MMB combo for pts with MF.

Background:In chronic myeloid leukemia (CML), absolute BCR-ABL molecular thresholds serve as validated milestones to predict long-term clinical benefit. In polycythemia vera (PV), JAK2V617F variant allele frequency (VAF) is routinely measured and reflects clonal burden, but lacks similar standing as a therapeutic endpoint. In PV, VAF>50% is associated with increased risk of events1, but current ELN guidelines limit molecular response assessment to patients with baseline VAF >20%, and emphasize a relative reduction in VAF. However, relative change is inherently ambiguous (e.g., 80% to 40% and 8% to 4% represent the same fold reduction but vastly different disease states)2,3, and fails to capture the biological significance of achieving or maintaining low VAF. We hypothesized that achieving an absolute JAK2 VAF <20% at any point during the disease course would be associated with prolonged event-free survival (EFS), independent of starting VAF or treatment received.Methods:We analyzed 1058 samples from 214 patients with PV and serial JAK2V617F VAF measurements, linked to clinical outcomes, including myelofibrosis transformation, thrombosis, and death. The cohort was followed longitudinally with a median of 12.0 years. VAF values were categorized as ≤20% or >20%. Time-varying Cox proportional hazards models were used to evaluate associations between achieving VAF <20% at any time point (regardless of baseline) and time to MPN complication (EFS), with adjustments for age >60 at diagnosis and thrombotic history. Additional models explored interactions with baseline VAF category, age at time of first measured VAF, age at time of lowest VAF, and treatment exposure.Results:Achieving JAK2V617F VAF <20% at any time point was associated with significantly prolonged EFS (HR 0.40, 95% CI 0.20–0.77, p=0.006), with consistent results after adjusting for baseline VAF, age, and history of thrombosis. The benefit was more pronounced in patients who started with VAF >20% and subsequently declined (median EFS 18.3, p = 0.04), though patients who started and remained below 20% already had a better EFS (median EFS 27.1). Considering that VAF tends to increase by ~1% per year4, maintenance of low VAF defies this biology and is associated with improved outcomes. In contrast, patients who remained >20% or who rose >20% tended to experience earlier events, including myelofibrosis transformation (HR 2.54, p = 0.052). The association was especially strong for myelofibrosis-free survival (HR 0.31, 95% CI 0.12-0.77, p=0.012), suggesting that sustained molecular control may influence disease progression. While being older age at diagnosis (>60) remained a risk factor, the protective effect of achieving VAF <20% persisted across age strata. Unlike MFS, thrombosis-free survival was not linked to this JAK2-mutation milestone.Conclusions:Achieving JAK2V617F VAF <20%—whether through disease biology or therapeutic intervention—is independently associated with prolonged EFS in PV. This absolute molecular threshold avoids the ambiguity of relative change and applies to all patients, not just those with high baseline burden. These findings support the adoption of JAK2 VAF <20% as a molecular response milestone in PV clinical trials, analogous to milestone-based frameworks in CML. Prospective studies are needed to validate this threshold and determine its utility as a surrogate endpoint for disease-modifying activity. References:Guglielmelli P, Loscocco GG, Mannarelli C, et al. JAK2V617F variant allele frequency> 50% identifies patients with polycythemia vera at high risk for venous thrombosis. Blood Cancer J. 2021;11(12):1-9.Harrison CN, Nangalia J, Boucher R, et al. Ruxolitinib Versus Best Available Therapy for Polycythemia Vera Intolerant or Resistant to Hydroxycarbamide in a Randomized Trial. Journal of Clinical Oncology. 2023;41(19):3534-3544.Kiladjian J, Klade C, Georgiev P, et al. Event-free survival in early polycythemia vera patients correlates with molecular response to ropeginterferon alfa-2b or hydroxyurea/best available therapy (PROUD-PV/CONTINUATION-PV). Hemasphere. 2025;9(5).Kiladjian JJ, Klade C, Georgiev P, et al. Long-term outcomes of polycythemia vera patients treated with ropeginterferon Alfa-2b. Leukemia. 2022;36(5):1408-1411.

Current prognostic models for polycythemia vera (PV)—including ELN (Barbui et al. Leukemia, 2018), MIPSS-PV (Tefferi et al. Br J Haematol, 2020) and the Myeloproliferative Neoplasm Personalized Risk Calculator (MPN-PRC) (Grinfeld et al. NEJM, 2018)—were all developed using diagnostic (dx) data to guide initial treatment. In practice, however, they are often reapplied during follow-up to inform ongoing care. Dynamic application has not been validated and may misrepresent evolving risk. We evaluated model performance over time in a real-world PV cohort. We analyzed 475 patients (pts) with PV treated at our institution (median age at dx 56 years [yrs], median follow-up 11 yrs) (Abu-Zeinah et al. Leukemia, 2021). Of 225 pts with complete genomic data, 53% had ≥1 co-occurring mutation, and 19% had an abnormal karyotype. Outcomes were estimated using Kaplan-Meier (KM) methods, and multivariable Cox proportional-hazards models were used to define associations. The ELN model, based on age ≥60 or thrombosis history (hx), stratifies thrombosis-free survival (TFS) at dx (p=0.00037 in our cohort). We reapplied ELN every 5 yrs post-dx, tracking risk shifts, new thrombotic events, and hazard ratios (HRs). Most pts shifted into the high-risk group due to aging. By 5 yrs, ELN parameters lost prognostic significance (HRs p>0.05), and 10-year TFS converged across risk groups. These findings highlight the importance of using models within the temporal scope for which they were developed. MIPSS-PV stratified overall survival (OS) at dx (p<0.0001), but in our cohort, age dominated mortality risk (age >67 yrs HR 6.3 [1.6–5.6], p<0.001) while WBC ≥15, SRSF2 mutations, and thrombosis hx were not significant. When reapplied every 5 yrs, MIPSS-PV progressively reclassified pts into higher risk groups, yet 10-yr OS converged across strata. This identifies a disconnect between static risk frameworks and dynamic disease biology. Models must be applied for their intended outcomes. ELN was developed for TFS, not OS or myelofibrosis-free survival (MFS); MIPSS-PV for OS, not other outcomes. In our cohort, ELN modestly stratified OS and MFS at dx, but this was entirely driven by age (OS: age ≥60 HR 8.9 [5.7–14], p<0.001; MFS: HR 2.1 [1.3–3.4], p=0.002). MIPSS-PV stratified TFS and MFS weakly via age alone and performance declined over time. These findings illustrate the risk of applying models beyond their designed scope and the dominant role of age in predicting OS. To evaluate the MPN-PRC in PV, clinical/genomic data were uploaded to the online calculator (Grinfeld et al. https://www.sanger.ac.uk/tool/progmod/progmod/) to generate pt-specific predictions for event-free survival (EFS) and OS, MFS, and leukemia-free survival (LFS) at 5-, 10-, and 20-yrs post-dx. Actual outcomes were estimated using KM methods and compared to predictions using truncated c-indexes (Uno et al. Stat Med, 2011; Shi, Github, https://github.com/YushuShi/correctedC)with 1000 bootstraps. The MPN-PRC stratified OS well at all timepoints (c-index ≥0.80). MFS prediction was strong at 5 yrs (c-index 0.91) but deteriorated at 10/20 yrs (c-index ~0.5). LFS prediction was poor (c-index <0.7), likely due to low leukemic transformation in our cohort (10-yr LFS = 99%). EFS was consistently overestimated (r = 0.43). To identify the variables driving MPN-PRC predictions, we systematically varied age, sex, and blood counts across observed ranges, adding individual co-mutations. Outputs were bootstrapped 10,000 times. Age dominated OS predictions; high WBC and TP53 and SRSF2 mutations modestly increased 10/20 yr post-dx risk. For MFS, SRSF2 had the strongest effect, with high PLT adding mild risk. For LFS, TP53 and SRSF2 most strongly increased predicted AML risk. We conclude: 1. Prognostic models should be used within scope: ELN, MIPSS-PV, and MPN-PRC stratify PV risk effectively at dx but reapplying them beyond 5 yrs fails to reflect evolving risk. These tools remain valuable when used for their validated outcomes and timeframes. Overextending scope can mislead clinical decisions. 2. Age dominates OS prediction: Age is immutable and is the strongest predictor of OS. Any new prognostic variable must be benchmarked against an age-only model to establish added value. 3. Future models must be dynamic: Therapies and life events reshape disease trajectories. PV risk assessment must move from static snapshots to time-sensitive, adaptive frameworks that reflect the continuous nature of PV care and risks.

Resistance to cyclin-dependent kinase 4/6 (CDK4/CDK6) inhibitors leads to treatment failure and disease progression in women with hormone receptor+HER2− (HR+HER2−) breast cancer (BC). We delineated a hypoxia-sensitive, CCL2-dependent pathway recruiting interleukin-17A (IL-17A)-secreting γδ T cells to mouse HR+HER2− BCs following CDK4/CDK6 inhibition, resulting in repolarization of tumor-associated macrophages (TAMs) toward an immunosuppressive CX3CR1+ phenotype associated with resistance. Increased IL-17A signaling and intratumoral γδ T cell abundance positively correlated with advanced grade and/or reduced survival in two cohorts of individuals with HR+HER2− BC. Circulating γδ T cells and plasma CCL2 levels negatively correlated with progression in an independent series of individuals with HR+HER2− BC receiving CDK4/CDK6 inhibitors. Intratumoral γδ T cells were increased in post- versus pretreatment biopsies from individuals with HR+HER2− BC relapsing on CDK4/CDK6 inhibitors. CX3CR1+ TAMs had negative prognostic impact in women with HR+HER2− BC receiving neoadjuvant PD-1 blockage and radiotherapy. Thus, γδ T cells and CX3XR1+ TAMs may favor resistance to CDK4/CDK6 inhibitors in individuals with HR+HER2− BC. Petroni et al. report that the infiltration of IL-17A-secreting γδ T cells in the tumor microenvironment coupled with the accumulation of immunosuppressive macrophages is associated with resistance to CDK4/CDK6 inhibition in HR+HER2− breast cancer.

Improving outcomes for patients with myeloproliferative neoplasms (MPNs) requires therapies that prolong event-free survival (EFS). MPNs' pathology is driven by the emergence, expansion, and persistence of hematopoietic clones carrying driver and co-occurring mutations that confer a competitive fitness advantage and alter lineage bias. The mutated stem and progenitor cells (HSPCs) sustain abnormal blood cell counts and drive thrombosis, disease progression, and leukemic transformation. Therefore, reducing clonal fitness—particularly in lineages responsible for pathology—is a critical therapeutic path towards prolonging EFS. While JAK2V617F is known to promote stem cell expansion and myeloid bias, its lineage-specific fitness effects due to allele burden, co-occurring mutations (present in >50% of cases), and variants of unknown significance (VUS) remain poorly defined. To address this, we developed a single-cell pipeline integrating immunophenotype and genotype to quantify mutation-specific clonal fitness across hematopoiesis using primary patient samples.MethodsWe adapted the MissionBio Tapestri platform to develop an automated, single-cell genotyping and immunophenotyping workflow optimized for primary MPN blood samples. The original MissionBio system uses 312 amplicons to cover 45 genes (Myeloid DNA panel) and 45 oligo-conjugated antibodies (Biolegend TotalSeq-D). We designed a custom DNA panel of 114 amplicons that captures 100% of mutations associated with MPNs observed in >9,700 samples from 1,243 patients at Weill Cornell Medicine. We designed an antibody panel with 29 antibodies, which identified all major immunophenotypic populations. Peripheral blood was re-proportioned using CD34 enrichment to ensure adequate representation of HSPCs and mature leukocytes in a single-tube cell suspension. Cells were labeled with barcoded antibodies and genotyped using the Tapestri instrument for single-cell emulsification. Sequenced DNA and protein libraries were analyzed using custom Python-based tools. The Mosaic library enabled single-cell variant calling and clone assignment. We trained a machine learning model on > 80,000 cells to assign cell identity based on antibody expression. Genotype-immunophenotype integration enabled clone identification and lineage tracking. Fitness across each hematopoietic lineage was defined as relative enrichment compared to wild-type counterparts for each clone.ResultsFitness of JAK2V617F clones depended on zygosity, co-occurrent mutations, and differentiation trajectory. Both monoallelic and biallelic JAK2V617F clones favored myeloid over lymphoid differentiation but differed in the magnitude of fitness and lineage preference. Myeloid fate preference varied between samples with JAK2V617F clones from some patients exhibiting stronger granulocytic vs erythroid differentiation and vice versa, suggesting that additional factors shape hematopoietic fitness. For instance, in a sample with JAK2V617F and a co-occurring DNMT3A R882H mutation, we identified six main clones. When comparing the JAK2V617F monoallelic clones, fitness toward the neutrophil lineage was more pronounced in the double-mutated clones (log2FC 1.94) and even higher in the double-mutated clone with DNMT3A loss of heterozygosity (log2FC 2.64). On the other hand, erythroid differentiation was primarily increased only in the JAK2V617F biallelic clone (log2FC 1.91). Monoallelic JAK2V617F was strongly depleted and biallelic clones were virtually absent in lymphoid populations, consistent with findings from orthogonal approaches. (Choi et al, Leukemia 2024) The pipeline also enabled in vivo assessment of several VUS, some of which showed measurable lineage-specific fitness effects, suggesting pathogenicity. Validation using paired flow cytometry and digital PCR analysis as previously reported (Abu-Zeinah et al, Blood Advances 2022) showed very high concordance, confirming the accuracy of the approach.ConclusionThis patient-centered, single-cell pipeline enables in vivo quantification of mutation-specific clonal fitness and differentiation bias in human hematopoiesis. This framework offers a scalable, biologically meaningful approach to dissect clonal architecture, functionally assess VUS, and guide therapy based on clone-level fitness. Longitudinal analysis of readily available blood samples promises to inform how specific treatments reshape MPN clonal dynamics and influence MPN event risk.

Background: The optimal treatment of early primary myelofibrosis (PMF) – prefibrotic PMF (prePMF) or low/intermediate-1 (int-1) risk overt PMF (oPMF) – is not well established. Most clinical trials have focused on higher risk oPMF patients (pts) but efforts are also needed in identifying treatments to prevent progression of early PMF and improve overall survival (OS). Interferon alfa (IFN) is one of few therapies that has long been investigated for early PMF treatment (Silver, Semin Hematol,1990). Studies of IFN thus far have demonstrated tolerability and promising efficacy, including in hematologic, molecular, and marrow responses (Gill et al. Blood,2023; Silver et al. Blood,2011). However, IFN has not yet been compared to other treatments or observation alone in early PMF. It also remains unknown if IFN improves OS or progression-free survival (PFS). In this comparative cohort study, we report the long-term outcomes associated with IFN use in early PMF. Methods: Medical records were queried and manually reviewed for pts with the diagnosis (dx) of prePMF and oPMF as per 2022 WHO and ICC criteria. The cohort was limited to pts with prePMF or low or int-1 risk oPMF by the Dynamic International Prognostic Scoring System Plus (DIPSS+). Demographic, clinical, lab, treatment, and outcome data were collected. Overall response rate (ORR) included hematologic, anemia, or spleen response as defined by the IWG-MRT 2013 response criteria. Pts that received ≥90 days of IFN therapy were assigned to the IFN arm and all others to the control group (NoIFN). Progression was defined as prePMF to oPMF advancement, accelerated or blast phase transformation, or PMF requiring bone marrow transplantation. OS and PFS were estimated using Kaplan-Meier methods. Cox proportional-hazards models were used for univariable and multivariable analysis (MVA) of mortality and progression risk. Results: We identified 92 pts with early PMF (median age, 59 years [yrs] at dx), of whom 46 (50%) received IFN. The driver mutation was JAK2 in 53(58%), CALR in 26 (28%), MPL in8 (9%), and triple negative in 4 (4%). There were no statistically significant differences in age, sex, race, driver mutation, blood counts, and DIPSS+ between groups, though the IFN group tended to be younger (56 vs 61 yrs, p=0.07) and included more pts with oPMF (30 [65%] vs 22 [48%], p=0.09) and int-1 risk (15 [33%] vs 9 [20%], p=0.2). Median time from dx to IFN initiation was 0.6 yrs; median IFN duration was 3 yrs. In the NoIFN group, 17 (37%) were on observation only, and others received one or more cytoreductive therapy including ruxolitinib (26%), hydroxyurea (28%), or other (30%). Median follow-up duration was similar in both groups (8.5 yrs IFN vs 10.4 yrs non-IFN, p=0.5). ORR with IFN was 70%; median response duration of 2.4 yrs. Hematologic response was achieved in 31 pts, anemia response from transfusion-independent baseline in 1, and spleen response by palpation in 3. Median OS was longer in the IFN group (18 vs 14 yrs, p<0.01), with 5-, 10-, and 15-yr OS rates of 100%, 100%, 93% vs 97%, 72%, 40% for IFN vs NoIFN, respectively. The OS advantage with IFN was confirmed in MVA: time on IFN (per yr) was associated with a significantly lower mortality risk (HR 0.85, CI 0.77-0.95, p=0.003), independent of dx age and DIPSS+ features. 6 IFN pts died vs 24 NoIFN pts, of whom 4 and 15, respectively, were disease-related (due to progression in 3/4 and 6/15, respectively, and non-progression events in others). Median PFS, however, was not significantly different (12 yrs IFN vs 14 yrs NoIFN, p=0.86). Conclusion: In our retrospective analysis of 92 pts with early PMF, treatment with IFN yielded superior OS. MVA confirmed that this survival benefit was independent of age and DIPSS+ lab parameters, supporting the use of IFN for early PMF. Randomized controlled trials are needed to validate and further assess the advantages of IFN on both short and long-term outcomes. The upcoming HOPE-PMF phase 3 trial evaluating ropeginterferon alfa-2b vs observation in early PMF pts (Abu-Zeinah et al. Ann Hematol,2024) will provide essential data on the effects of IFN in a multi-center, randomized, double-blinded, prospective design. Such studies are critical in improving the treatment paradigm and bettering outcomes for the often-overlooked early PMF population.

Janus kinase (JAK) inhibitors provide limited depth and durability of response in myelofibrosis. We evaluated pelabresib—a bromodomain and extraterminal domain (BET) inhibitor—plus ruxolitinib (a JAK inhibitor) compared with placebo plus ruxolitinib as first-line therapy. In this phase 3 study (MANIFEST-2), JAK inhibitor-naive patients with myelofibrosis were randomized 1:1 to pelabresib 125 mg once daily (QD; 50–175 mg QD permitted) for 14 days followed by a 7-day break (21-day cycle), or to placebo in combination with ruxolitinib 10 or 15 mg twice daily (BID; 5 mg QD–25 mg BID permitted). Primary endpoint was reduction in spleen volume of ≥35% from baseline at week 24. Key secondary endpoints were absolute change in total symptom score (TSS) and TSS50 response (≥50% reduction in TSS from baseline at week 24). The primary endpoint was met in 65.9% of patients randomized to pelabresib–ruxolitinib (n = 214) versus 35.2% to placebo–ruxolitinib (n = 216) (difference, 30.4%; 95% confidence interval (CI), 21.6, 39.3; P < 0.001). Absolute change in TSS was −15.99 versus −14.05 (difference, −1.94; 95% CI, −3.92, 0.04; P = 0.0545) and TSS50 was achieved in 52.3% versus 46.3% (difference, 6.0%; 95 CI, −3.5, 15.5) with pelabresib–ruxolitinib versus placebo–ruxolitinib. Exploratory analyses of proinflammatory cytokine amounts and bone marrow morphology showed greater improvement with the combination. Thrombocytopenia and anemia were the most common treatment-emergent adverse events, occurring in 52.8% (13.2% grade ≥3) versus 37.4% (6.1% grade ≥3) and 44.8% (23.1% grade ≥3) versus 55.1% (36.5% grade ≥3), respectively. Pelabresib in combination with ruxolitinib is well tolerated, improves signs of underlying myelofibrosis pathobiology and provides substantial clinical benefit over standard-of-care JAK inhibitor monotherapy. ClinicalTrials.gov identifier: NCT04603495 . In a randomized phase 3 trial, the combination of the BET inhibitor pelabresib with the JAK inhibitor ruxolitinib resulted in a significantly higher spleen volume reduction from baseline versus placebo with ruxolitinib in patients with JAK inhibitor-naive myelofibrosis.

ABSTRACT:DNMT3A mutations in patients with polycythemia vera were heterogeneous and not enriched in interferon alfa-treated patients. DNMT3A mutations had no detectable impact on the hematologic response, molecular response, or survival outcomes.

Assessing treatment response in patients with myeloproliferative neoplasms is difficult because data components exist in unstructured bone marrow pathology (hematopathology) reports, which require specialized, manual annotation, and interpretation. Although natural language processing (NLP) has been successfully implemented for the extraction of features from solid tumor reports, little is known about its application to hematopathology.An open-source NLP framework called Leo was implemented to parse document segments and extract concept phrases utilized for assessing responses in myeloproliferative neoplasms. A reference standard was generated through the manual review of hematopathology notes.Compared with a reference standard (n = 300 reports), our NLP method extracted features such as aspirate myeloblasts (F1 = 98%) and biopsy reticulin fibrosis (F1 = 93%) with high accuracy. However, other values, such as myeloblasts from the biopsy (F1 = 6%) and via flow cytometry (F1 = 8%), were affected by sparsity representative of reporting conventions. The four features with the highest clinical importance were extracted with F1 scores exceeding 90%. Whereas manual annotation of 300 reports required 30 hours of staff effort, automated NLP required 3.5 hours of runtime for 34,301 reports.To the best of our knowledge, this is among the first studies to demonstrate the application of NLP to hematopathology for clinical feature extraction. The approach may inform efforts at other institutions, and the code is available at https://github.com/wcmc-research-informatics/BmrExtractor.

Background: PIM1 is overexpressed in hematologic malignancies, including myelofibrosis (MF), contributing to disease progression by modulating cytokine-driven pathways such as PI3K/AKT and JAK/STAT. Elevated pro-inflammatory cytokines are a hallmark of MF and are closely linked to symptom burden and poor prognosis. Preclinically, PIM1 knockout was shown to prevent MF progression without affecting the platelet (PLT) counts, whereas pan-PIM knockout caused thrombocytopenia (TCP). Nuvisertib (TP-3654), an oral, investigational, highly selective PIM1 kinase inhibitor, alone or in combination with JAK inhibitor ruxolitinib has demonstrated reduction in spleen size, bone marrow (BM) fibrosis, and expression of cytokine response genes in JAK2V617F and MPLW515L MF mouse models. Methods: The ongoing global phase 1/2 study evaluates the safety and efficacy of nuvisertib monotherapy in patients (pts) with MF (NCT04176198, Arm 1). Study population includes primary or secondary MF, previously treated with or ineligible for JAK inhibitor, DIPSS intermediate or high-risk MF, PLT ≥25 x 109/L, splenomegaly (≥450 cm3 by imaging), and ≥2 measurable symptoms per MFSAF v4. The study aims to identify the RP2D of nuvisertib monotherapy and assess the safety, clinical activity (spleen volume reduction [SVR], total symptom score [TSS] improvement), and PK and PD markers (cytokine, BM fibrosis etc.). Results: As of 29 May 2025, total 77 pts enrolled in 5 dose levels of nuvisertib from 480 mg QD to 720 mg BID. At baseline, median age 71 years (49, 85); spleen volume 1988 cm3 (270, 7718); TSS 23 (4, 62); hemoglobin (Hgb) 9.7 g/dL (5.6, 17.2; 52% pts were <10 g/dL; 39% pts required transfusion); PLT 96 x 109/L (24, 816; 51% pts were <100 x 109/L). 75% pts were DIPSS Int-2 or high risk; 41% pts had high molecular risk mutation; and 30% pts received ≥2 prior JAK inhibitors. Median nuvisertib treatment duration was 22 weeks (2, 197), and 16 (21%) pts on active treatment. No DLT occurred. Treatment-related adverse events (TRAEs) occurring in ≥20% of pts were primarily grade 1/2 diarrhea, nausea, and vomiting. Grade ≥3 TRAE occurring in ≥3 pts included TCP (n=8, 7 of 8 pts had baseline TCP). Mean Hgb and PLT remained stable throughout the 24-week treatment. In pts treated with 720 mg BID dose for ≥12 weeks, ≥25% SVR was observed in 4 of 20 pts (20% SVR25 response) and ≥50% reduction in TSS in 9 of 20 pts (44% TSS50 response) at any time. Absolute improvement in symptoms was seen across all 7 parameters. A strong correlation (p<0.001) between cytokine modulation (e.g. ↓ENRAGE, ↓MIP1β, ↓PAI-1, ↓IL-1Ra, and ↑adiponectin) and SVR25, TSS50, and individual symptom improvement were observed. In pts with baseline Hgb <10 g/dL, 6 of 26 (23%) pts showed Hgb response [mean ≥1.0 g/dL Hgb increase for ≥12 weeks without transfusion, including 3 pts with ≥1.5 g/dL Hgb increase]. Hgb responses were also observed in pts with baseline Hgb >10 g/dL. In pts with baseline PLT <100 × 109/L, 8 of 30 (26.7%) pts showed PLT response [≥30×109/L increase maintained ≥4 weeks], and PLT recovered to ≥100 × 109/L in all responders. Modulation of circulating biomarkers was observed in PLT responder pts including increased TN-C, a protein reported to be involved with PLT recovery, and decreased VCAM-1, a marker involved in PLT endothelial cell adhesion and chronic inflammation. 13 of 34 (38.2%) evaluable pts (assessments at baseline and every 24 weeks) showed ≥1 grade reduction in BM fibrosis which correlated with cytokine reduction (e.g. ↓MIP1β, ↓TNFR1); and 11 of 13 pts also showed at least one of SVR25, TSS50, Hgb or PLT responses (5 pts showed dual Hgb and PLT responses). The 1-year overall survival rate following nuvisertib treatment in this heavily pretreated pts with relapsed/refractory (R/R) MF was 81% which also correlated with cytokine modulation (e.g. ↓MIP1β, ↓TNFR1, and ↑FVII). Conclusions: Nuvisertib monotherapy appeared well tolerated with no DLTs. Preliminary data in pts with R/R MF showed that nuvisertib treatment leads to significant modulation of cytokine profiles, demonstrating a strong correlation with clinical responses, including SVR25 and TSS50 responses, and improvements in Hgb, PLT and BM fibrosis, suggesting that selective PIM1 inhibition may offer disease-modification with limited hematologic toxicity. Emerging data supports ongoing clinical development of nuvisertib in combination with ruxolitinib and momelotinib (NCT04176198, Arms 2 and 3, respectively).

Background: Rusfertide is a weekly, self-administered, subcutaneously injected first-in-class hepcidin mimetic. Previously, we presented data from Part 1a (Weeks [Wks] 0-32) of the phase 3 VERIFY study (NCT05210790) showing that rusfertide added to current standard-of-care (SOC) therapy for polycythemia vera (PV) significantly reduced the mean number of phlebotomies (PHLs), improved hematocrit (Hct) control, and improved patient-reported outcomes addressing fatigue and disease-related symptoms (Kuykendall AT, et al. J Clin Oncol. 2025;43(17_suppl.):LBA3). Herein, we present results from Part 1b (Wks 32-52) to assess the durability of response in patients (pts) who continued rusfertide and the efficacy of rusfertide in pts who crossed over from PBO. Methods: In VERIFY Part 1a (double-blind period), pts requiring frequent PHL (≥3 PHLs in 28 wks or ≥5 in 52 wks prior to randomization) were randomized 1:1 to receive PBO or rusfertide once weekly in addition to current SOC therapy, including cytoreductive therapy (CRT). After completing Part 1a, all pts (including those on PBO) were eligible to receive rusfertide in Part 1b; pts completing Part 1b were eligible to receive rusfertide in Part 2 (long-term extension period; Wks 52-156). Pts randomized to rusfertide in Part 1a continued to receive the same dose at the beginning of Part 1b unless dose adjustment was needed. Pts randomized to PBO received a starting dose of 20 mg of rusfertide. Part 1b evaluated the proportion of rusfertide-treated pts who achieved absence of PHL eligibility (ie, durable response from Wks 0-52), median time to first Hct ≥45%, and median time to first PHL. Results: Of 293 pts randomized to receive rusfertide (n=147) or PBO (n=146), 274 (94%) began Part 1b, and 254 (87%) remain on study and are receiving open-label rusfertide in Part 2. Safety data are reported for the safety analysis set (n=291; 145 and 146 in the rusfertide and PBO groups, respectively). Median (range) rusfertide exposure was 61 (2, 133) wks. The proportion of pts who continued to have an absence of PHL eligibility between Wks 0-52 (ie, pts initially randomized to rusfertide with durable response through the end of Part 1b) was 61.9% (n=91/147). In pts originally randomized to PBO who crossed over to rusfertide at Wk 32, the proportion of pts who achieved absence of PHL eligibility was 32.9% (n=48/146) and 78.0% (n=110/141) in Parts 1a (Wks 20-32) and 1b (Wks 40-52), respectively. During Part 1a (Wks 0-32), median (95% confidence interval [CI]) time to first PHL was not reached (NR) and 16.1 wks (12.7, 20.1) in the rusfertide and PBO groups, respectively. During Part 1b (Wks 32-52), median time to first PHL was NR in the rusfertide group and those who crossed over from PBO to rusfertide at Wk 32. In the rusfertide group, median time to first Hct ≥45% was NR during Part 1b. Median (95% CI) time to first Hct ≥45% was 21.1 wks (20.1, NR) in pts who crossed over from PBO to rusfertide at Wk 32. Mean Hct remained <43% throughout Part 1b in pts who continued rusfertide and those who switched from PBO to rusfertide. During Part 1b, numerical improvements (ie, change from baseline) in the PROMIS Fatigue SF-8a and MFSAF TSS7 were durable in the rusfertide group. Overall, the most common treatment-emergent adverse events (AEs) in rusfertide-treated pts (n=285) were injection site reactions (47.4%), anemia (25.6%), and fatigue (19.6%); most were grade 1 or 2. In Part 1a, fatigue occurred in 15.8% and 15.9% of pts in the PBO and rusfertide groups, respectively, and in Part 1b, the incidence was similar between pts originally randomized to both PBO and rusfertide (9.3% vs 9%, respectively). Serious AEs occurred in 8.1% of rusfertide-treated pts. In Part 1a of this ongoing study, 3 (2.1%) and 8 (5.5%) pts in the rusfertide and PBO groups had a non-PV malignancy; during Part 1b, non-PV malignancies were reported in 6 (2.2%) pts. Two pts (rusfertide group) had TEs (1 pt each in Parts 1a and 1b). Conclusions: Rusfertide met the primary and all 4 key secondary endpoints in VERIFY and continued to provide durable, sustained control of Hct <45% and a relative absence of PHL up to Wk 52. This benefit was maintained across all subgroups, including age, PV risk category, and ongoing CRT. Similar findings were also observed after pts crossed over from PBO to rusfertide. Rusfertide's safety profile was consistent with prior observations.Supported by: Protagonist Therapeutics, Inc.

Background: Polycythemia vera (PV) is a myeloproliferative neoplasm (MPN) characterized by erythrocytosis that is often accompanied by fatigue, pruritus, problems with concentration, and other symptoms. At times, these symptoms can be debilitating. Rusfertide is a first-in-class hepcidin mimetic peptide that controls erythrocytosis. In the ongoing randomized phase 3 VERIFY study (NCT05210790), compared to placebo (PBO) plus current standard of care (SOC), rusfertide added to SOC for PV met its primary endpoint, 2 key secondary patient-reported outcome (PRO) endpoints, and 2 additional key secondary endpoints at Week 32. Here, we present additional data from the 2 key secondary endpoints in VERIFY Part 1a that relied on the PROMIS Fatigue Short Form (SF)-8a and MFSAF Total Symptom Score 7 (TSS7) PROs and PRO-focused exploratory endpoints. Methods: In the VERIFY Part 1a double-blind period (Weeks 0-32), patients (pts) requiring frequent phlebotomy with or without cytoreductive therapy (CRT) to achieve and maintain hematocrit <45% received once-weekly rusfertide or PBO added to pts' current SOC. Two key alpha-controlled secondary endpoints evaluated mean change from baseline (BL) to end of VERIFY Part 1a (Week 32) in the PROMIS Fatigue SF-8a T-score and the MFSAF TSS7 (includes fatigue, night sweats, itch, abdominal discomfort, pain under left ribs, early satiety, and bone pain). Exploratory endpoints included mean change from BL at Week 32 in pts who were symptomatic at BL (ie, pts with moderate or severe symptoms at BL; severity determined using Patient Global Impression of Severity [PGI-S] groupings) in the PROMIS Fatigue SF-8a T-score, the MFSAF TSS4 (ie, PV-relevant symptoms, eg, fatigue, night sweats, itch, and abdominal discomfort), and the single item measuring problems with concentration from the MPN Symptom Assessment Form (MPN-SAF). Negative numbers indicate improvement in all these measures. Results: Pts (N=293; 73.0% male; median age, 57 yrs) received rusfertide (n=147) or PBO (n=146) with or without CRT. At BL, mean (standard deviation [SD]) PROMIS Fatigue SF-8a T-Score was similar in the rusfertide and PBO groups (52.5 [11.7] and 51.2 [10.1], respectively). Mean (SD) change from BL to Week 32 in the PROMIS Fatigue SF-8a T-Score was -3.9 (8.2) and -1.1 (8.1) in the rusfertide and PBO groups, respectively (least-squares means [LSM] difference (standard error [SE]), -1.98 [0.88]; p=0.025). At BL, mean (SD) MFSAF TSS7 was 10.7 (11.4) and 9.5 (10.2) in the rusfertide and PBO groups, respectively. Mean (SD) change from BL to Week 32 in the MFSAF TSS7 was -2.96 (8.0) and -0.06 (5.7) in the rusfertide and PBO groups, respectively (LSM difference [SE], -1.87 [0.82]; p=0.024). In pts who were symptomatic at BL, mean (SD) change from BL to Week 32 was a) -9.8 (8.8) and -5.2 (8.3) for the PROMIS Fatigue SF-8a T-score in the rusfertide and PBO groups, respectively (LSM difference [SE], -3.83 [1.92]; nominal p=0.0496), and b) was -6.4 (5.8) and -1.8 (5.7) for MFSAF TSS4 in the rusfertide and PBO groups, respectively (LSM difference [SE], -4.54 [1.33]; nominal p=0.001). In the rusfertide group, a greater number of pts improved by ≥1 severity category (eg, improvement from “severe” to “moderate”; improvement from “moderate” to “mild”) in the PROMIS Fatigue SF-8a and MFSAF TSS7 vs PBO. In pts who were symptomatic at BL, mean (SD) change from BL to Week 32 in the MPN-SAF problems with concentration item was -2.5 (1.9) and -1.9 (2.2) in the rusfertide and PBO groups, respectively (LSM difference [SE], -1.08 [0.48]; nominal p=0.027). Conclusions: In VERIFY, the first phase 3 study in PV to prospectively investigate PROs as key secondary endpoints, rusfertide led to statistically significant improvements vs PBO in patient-reported symptoms. Rusfertide significantly reduced fatigue and overall symptom burden (PROMIS Fatigue SF-8a and MFSAF TSS7 scores) and resulted in significant improvements from BL (p<0.05) in pts with moderate-to-severe symptoms at BL in core PV symptoms (MFSAF TSS4) and problems with concentration (MPN-SAF) vs PBO. In the rusfertide group, more pts improved by ≥1 severity category in the PROMIS Fatigue SF-8a and MFSAF TSS7 vs PBO. Overall, these results confirm the robustness of rusfertide's clinical benefit in PV, particularly in pts with moderate or severe symptoms at BL. Support: Protagonist Therapeutics, Inc.

Chronic systemic inflammation is a key driver of polycythemia vera (PV) progression, but the immunomodulatory effects of current treatments remain poorly defined. The neutrophil-to-lymphocyte ratio (NLR) is an accessible biomarker of systemic inflammation proven in other contexts, but its role in monitoring PV disease activity has not been established. Using data from three of the largest PV clinical trials, we evaluated the effects of PV therapies on NLR and its relationship with molecular response and clinical outcomes. In 404 hematocrit-controlled patients from the ECLAP study, hydroxyurea (HU) failed to significantly lower NLR (p = 0.11) due to the parallel declines in ANC and ALC. Neither leukocyte counts nor NLR were significantly reduced by phlebotomy in ECLAP patients treated without cytoreductive therapy. In contrast, the Low-PV study showed that while phlebotomy tended to increase NLR, low-dose ropeginterferon alfa-2b (Ropeg) significantly reduced NLR (-18.2% and -36.3% in patients with low and high baseline NLR, respectively) by suppressing ANC rather than lymphocytes. NLR reduction correlated with the primary Low-PV endpoint (p = 0.021) and reduction of JAK2 variant allele frequency (VAF) [1]. The PROUD-PV/CONTINUATION-PV study confirmed the superior effect of Ropeg over HU, with a significantly greater NLR reduction at 60 months (-56.5% versus -33.6%, respectively, p = 0.019) in patients with high baseline NLR. Moreover, NLR reduction was associated with decreased JAK2V617F VAF (p < 0.0001) and improved event-free survival (p = 0.010). These findings identify NLR as a dynamic biomarker of treatment response and prognosis in PV and support its incorporation into routine monitoring.

Background: Risk stratification algorithms for polycythemia vera (PV) were developed to predict thrombosis risk using immutable factors of age and thrombotic history. While current PV care standards have greatly reduced thrombotic complications, the long-term risks of progression and mortality persist. Therefore, dynamic prognostic models for PV are needed to better predict outcomes including overall, myelofibrosis-free and leukemia-free survival (OS/MFS/LFS). Grinfeld et. al1 developed the myeloproliferative neoplasms (MPN) Personalized Risk Calculator (MPN-PRC) to stratify risk using genomic profiles and select laboratory and clinical factors. This model has not been further validated. We used data from our richly annotated cohort of patients (pts) with PV to assess the performance of the MPN-PRC in predicting OS/MFS/LFS. Methods: Model parameters including age, sex, thrombotic history, splenomegaly, white blood cell count (WBC), platelet count (PLT), hemoglobin (HGB), karyotype and genomic data were collected from our MPN research data repository, as previously described2. Pt data was uploaded to the online risk calculator3, and the outcome predictions were compiled. We computed truncated c-indexes with confidence intervals (CI) based on 1000 bootstraps for each type of outcome, treating events from other outcomes as censoring4,5. Kaplan-Meier methods were used to estimate OS, MFS, LFS and event-free survival (EFS) with PV events defined as MF progression, leukemic transformation or death, as done for the MPN-PRC. Cox proportional hazard models were used for multivariable analysis (MVA) of MF progression and mortality risk. Results: Prognosis was predicted for 475 PV pts. Median age was 56 years (yrs), 49% were female, and 15% had a history of thrombosis. Median follow-up duration was 11 yrs. Complete genomic data and karyotype was available in 225 (47%) and 232 (49%) pts, respectively. At genomic sampling, median blood counts were the following: WBC 10 x103/µL; PLT 427 x103/µL; and HGB 14.5 g/dL. Median OS was 27.2 yrs. The model effectively predicted OS across all considered timepoints with computed c-indexes of ≥0.8 and a significant bootstrap 95% CI (5-yr 0.84, CI 0.77-0.90; 10-yr 0.85, CI 0.80-0.91; 20-yr 0.81, CI 0.59-0.98). As expected, age was a strong predictor of OS at all time points. In addition, TP53 mutation was associated with lower 5 yr and 10 yr OS (p=0.005, p=0.019) in univariate models but was not significant in MVA including age. A higher 20-yr mortality was associated with older age (p<0.001), PHF6 mutation (p=0.013), chromosome 5q deletion (p=0.011) and splenomegaly (p=0.045), but only age and PHF6 status remained significant in MVA. Median MFS was 28.6 yrs, with 92 (19%) pts progressing to MF. MFS prediction was excellent at the 5-yr timepoint (c-index 0.91, CI 0.89-0.94), but poor thereafter (c-index at 10-yr 0.49, CI 0.32-0.62; at 20-yr 0.56, CI 0.34-0.81, not significant). Pts who progressed to MF within 5 yrs were older (71 vs 56, p=0.085) and had lower PLT (273 vs 432 x103/µL, p=0.035). We identified no significant imbalance in genomic profiles between those that progressed early vs those that did not. Median LFS was not reached and prediction of LFS was weak (c-index 5-yr 0.68, CI 0.48-0.88; 10-yr 0.72, CI 0.53-0.89; 20-yr 0.65, CI 0.58-0.93). This poor performance may be due to the long 10-yr LFS of 99% in our cohort. Actual median EFS was 20 yrs whereas the model predicted 24 yr median EFS. Including all events occurring within the 25 yr period considered by the MPN-PRC, correlation between predicted and actual EFS was poor (correlation coefficient 0.43). Conclusion: The MPN-PRC performed well in predicting OS and early onset MF but did not reliably predict long-term MFS or LFS. Predictions were primarily driven by age, with genomic profiles adding limited value. The model overestimated median EFS in our PV cohort, perhaps owing to the MPN-PRC model having been trained on a cohort predominated by pts with ET. These findings confirm the model's utility in clinical practice for short-term, personalized OS and MFS prognosis for pts with PV, while indicating a need for refinement in long-term predictions to aid clinical decision-making. References 1. Grinfeld et al. NEJM, 2018. 2. Abu-Zeinah et al. Leukemia, 2021. 3. Grinfeld et al. https://www.sanger.ac.uk/tool/progmod/progmod/ 4. Uno et al. Stat Med, 2011. 5. Shi, Y. https://github.com/YushuShi/correctedC

Introduction Pulmonary hypertension (PH) is a morbid disease found in up to half of patients with myeloproliferative neoplasms (MPNs), with an estimated prevalence of PH of ~29%. 1 Recent studies indicate that all-cause mortality is higher in patients with MPNs diagnosed with PH.2 We found that a quarter of patients with polycythemia vera (PV) have PH, which independently shortens their survival 3-fold (See ASH Abstract ID 210231). Patients with PV and advanced PH have a median survival of under a year. To identify patients at risk for PH and to better understand the pathophysiology of MPN-related PH (WHO Group 5 PH), we performed cytokine profiling of 92 patients treated at the Weill Cornell Medicine Silver MPN Center. Methods Clinical data, including echocardiography (echo), were extracted from our deeply annotated MPN Research Data Repository. PH was defined by mean pulmonary artery pressure (mPAP) >35 mmHg or tricuspid regurgitation jet velocity (TRV) ≥2.8m/s. Fresh peripheral blood samples were collected from 92 patients with MPNs, and serum was collected and cryopreserved for batch analysis. Samples were analyzed at Nomic Bio using the nELISA platform. This new multiplexing platform isolates target-specific antibody pairs using color-coded beads, preventing reagent cross-reactivity. Fluorescent oligonucleotides label correctly formed complexes, which were detected by high-throughput spectral flow cytometry. This nELISA platform allowed us to analyze 267 cytokines per sample with a sensitivity of ~0.1 pg/ml. Fifty patients were screened by echo for PH: 10 had PH (2 mild and 8 advanced), and the remainder have not yet been screened by echo. Analysis of scaled normalized data was done in R to identify differentially detected cytokine levels in patients with and without PH. Results In the PH group, nine cytokines were significantly enriched (log2 fold change ≥ 1 & p-value <0.05): APRIL (56,000 vs 3,800 pg/ml), BMP4 (26 vs 8 pg/ml), CCL13 (340 vs 140 pg/ml), CCL17 (240 vs 110 pg/ml), CCL2 (360 vs 180 pg/ml), FGF-4 (87 vs 40 pg/ml), GM-CSF (59 vs 24 pg/ml), IL-22 BP (16,000 vs 7,800 pg/ml), IL-35 (6,000 vs 2,500 pg/ml), and IL-9 (4,400 vs 1,300 pg/ml). These cytokines include members of FGF, TNF and TGFβ superfamilies known for their pathogenic roles in PH. Mutations in BMP receptor II mutations and downstream TGFβ superfamily signaling mediators such as SMAD1/SMAD9/ALK1/ENG are the most common genes mutated in familial PH. We also identified cytokines linked to MPN hematopoiesis, including those involved in monocyte (GM-CSF, CCL2, and CCL13) and megakaryocyte biology (FGF-4). Interestingly, several anti-inflammatory cytokines (IL-9, IL-22 BP, and IL35) that decrease endothelial proliferation in the pulmonary vasculature were elevated in patients with PH, potentially indicating compensatory mechanisms.3 Across all patients, detection of BMP4, FGF-4, GM-CSF, IL-22 BP and IL-9 was bimodal, suggesting these cytokines may help identify patients with MPN and concomitant PH. Among the 42 patients in our cohort who had not undergone echo screening, we identified 35 with a cytokine signature suggestive of PH. We will report the results of their screening echo. Conclusions We found several proinflammatory and chemotactic cytokines enriched in patients with MPN and PH. These cytokines may help identify patients with MPN at risk of PH. The identified mitogenic and chemotactic cytokines identified highlight the need to understand the role of enriched myeloid cells in MPN-related PH. Ferrari A, Scandura J, Masciulli A, Krichevsky S, Gavazzi A, Barbui T. Prevalence and risk factors for Pulmonary Hypertension associated with chronic Myeloproliferative Neoplasms. Eur J Haematol. 2021;106(2):250-259. Kim J, Krichevsky S, Xie L, et al. Incremental Utility of Right Ventricular Dysfunction in Patients With Myeloproliferative Neoplasm-Associated Pulmonary Hypertension. Journal of the American Society of Echocardiography. 2019;32(12):1574-1585. Mamazhakypov A, Viswanathan G, Lawrie A, Schermuly RT, Rajagopal S. The role of chemokines and chemokine receptors in pulmonary arterial hypertension. Br J Pharmacol. 2021;178(1):72-89.