Towards Biomarkers and Pathophysiology of Pulmonary Hypertension in Myeloproliferative Neoplasms
Authors: Castillo Tokumori Franco , Choi Daniel C , Kermani Pouneh , Erdos Katie , Gazda Alexander , Robichaud Nathaniel , Scandura Joseph M. 
Published: December 2024

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) &gt;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 &lt;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.