Peripheral blood plasma proteomics reveals dysregulated cholesterol transport and complement pathways in myeloproliferative neoplasms

Abstract Myeloproliferative neoplasms (MPNs) are rare, age-related hematologic malignancies characterized by the clonal expansion of myeloid lineages. The Philadelphia chromosome-negative (Ph-) MPNs include essential thrombocythemia (ET), polycythemia vera (PV), and myelofibrosis (PMF), Ph+ MPNs include chronic myeloid leukemia (CML). While genetic drivers of MPNs are increasingly understood, systemic biological changes linked to aging and MPN pathogenesis remain unclear. To investigate proteomic alterations associated with MPNs and aging, we performed liquid chromatography/mass spectrometry (LC/MS/MS) analysis of peripheral blood plasma procured from eleven MPN patients at diagnosis: three with ET, four with MF, and four with CML, as well as twelve healthy donors (with no cardiovascular disease), stratified into “young” (≤30 years, n=6) and “old” (≥40 years, n=6) groups. MPN patient samples were obtained from Rhode Island Hospital, and age and sex-matched healthy donor plasmas were sourced from the Rhode Island Blood Bank. All plasma samples were subjected to albumin depletion followed by LC/MS/MS analysis. Proteomic profiling identified 1,224 unique plasma proteins across all sample types. Hierarchical clustering and principal component analysis of protein abundance profiles showed clear separation between MPN and healthy plasma samples, as well as age-based separation within healthy donors. Set intersection analysis showed 851 proteins commonly identified in all groups, 43 proteins unique to MPNs, and 3 proteins unique to healthy donors. Volcano plots highlighted markedly increased protein abundance in MPN samples compared to healthy controls. In healthy, aged samples, we observed, consistent with immunosenescence and pro-thrombotic states in aging, an upregulation of complement cascade activators (e.g., C1QA, C1QB) and coagulation proteins (e.g., PROS1) and a reduction in immunoglobulins and T-cell signaling components (e.g., IGHM, IGHV3-7, SFTPD, PTPN6). MPN plasma proteomes showed significant alterations in pathways involving coagulation, complement activation, innate immunity, and cholesterol transport. CML and MF samples displayed similar proteomic patterns, including upregulation of complement activators (e.g., C1QC), downregulation of complement inhibitors (e.g., C4BPA, C4BPB, CLU), and loss of apolipoproteins (e.g., APOA1, APOB, APOM). ET samples showed predominant dysregulation of proteins involved in platelet activation (e.g., PF4, VWF, GP1BA), consistent with the clinical phenotype. Functional interaction network analysis revealed that dysregulated cholesterol transport proteins, particularly apolipoproteins, were connected to dysregulated complement cascade regulators. Cholesterol transport and complement clusters were prominently linked through clusterin - CLU, which is a complement inhibitor known to associate with apolipoproteins in plasma. Correlation analysis across all samples showed inverse relationships between apolipoproteins abundance (APOE, APOB, APOC3) and complement activators (C1S, C1QB, C4A), and positive correlations with between apolipoproteins abundance and complement inhibitors (CFI, C4BPA, C4BPB). These findings support a link between metabolic and immune dysregulation in MPNs, specifically implicating cholesterol transport deficiency in the overactivation of the complement system. This proteomic signature may help explain aspects of the chronic inflammatory environment observed in both aging and myeloid malignancies and its link to metabolism. Metabolic abnormalities have been implicated in MPN pathophysiology and prognosis. Hypocholesterolemia, cachexia, and dyslipidemia correlate with reduced survival in MPN patients, statin use is associated with improved outcomes, and metformin use is linked to reduced MPN incidence. Our results suggest that disrupted cholesterol efflux and apolipoproteins deficiency may exacerbate complement activation driving inflammation and thrombosis. Notably, cholesterol uptake by macrophages enhances complement activation and atherogenesis, further supporting the pathobiological relevance of this axis. Our plasma proteomic analysis reveals a mechanistic link between impaired cholesterol transport and complement overactivation in MPNs, highlighting the metabolic-immune axis as a potential therapeutic target in myeloid malignancies and aging.