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Importance There is a lack of long-term efficacy and safety data on hereditary transthyretin amyloidosis with polyneuropathy (hATTR-PN) and on RNA interference (RNAi) therapeutics in general. This study presents the longest-term data to date on patisiran for hATTR-PN. Objective To present the long-term efficacy and safety of patisiran in adults with hATTR-PN. Design, Setting, and Participants This global open-label extension (OLE) of the APOLLO randomized clinical trial and phase 2 OLE study enrolled patients from 43 hospitals or clinical centers across 19 countries between July 2015 and August 2017, with follow-up until November 2022. Of 212 eligible patients with hATTR who completed the phase 3 APOLLO or phase 2 OLE parent studies, 211 enrolled in and 138 completed the global OLE. Intervention Patisiran, 0.3 mg/kg, intravenously once every 3 weeks for up to 5 years. Main Outcomes and Measures Outcomes evaluated at year 5 of the global OLE included disability (polyneuropathy disability [PND] score); polyneuropathy severity (Neuropathy Impairment Score [NIS]), nutritional status (modified body mass index [mBMI]), quality of life (Norfolk Quality of Life–Diabetic Neuropathy [Norfolk QOL-DN]), and Rasch-Built Overall Disability Scale (R-ODS), with no statistical hierarchy. Safety, survival probability, and mortality were also assessed. Results At the global OLE baseline, the mean (SD) age was 61.3 (12.3) years, and 156 patients (73.9%) were male. In 138 patients completing the study, PND scores remained stable or improved in 89 patients (65.0%), NISs showed a mean (SD) change of 10.9 (14.7), and mean (SD) mBMI (calculated as weight in kilograms divided by height in meters squared times serum albumin in grams per liter) increased by 46.4 (120.7) over 5 years from baseline. Norfolk QOL-DN and R-ODS scores showed mean (SD) changes of 4.1 (16.7) and –3.7 (6.2), respectively. Adverse events (AEs) leading to study withdrawal occurred in 47 patients (22.3%). Infusion-related reactions were the most common treatment-related AE (n = 34 [16.1%]). Overall, 41 patients (19.4%) died during the study. Patisiran treatment in the parent study and low familial amyloid polyneuropathy score at parent study baseline were associated with significantly improved survival. Conclusions and Relevance In the longest study of an RNAi therapeutic for any disease, patisiran treatment resulted in modest changes for patients with hATTR-PN with an acceptable safety profile. These results highlight the importance of initiating early treatment for hATTR and the potential of RNAi therapeutics in medicine. Trial Registration ClinicalTrials.gov Identifier: NCT02510261

Introduction. Non-cystic fibrosis bronchiectasis is a complex medical condition with multiple etiologies, characterized by chronic productive cough and radiologic evidence of airway lumen dilation and wall thickening. Associated exacerbations and declining lung function contribute to increasing disability and mortality. There are no data about the prevalence of non-cystic fibrosis bronchiectasis etiologies in the Colombian population.Objective. To investigate non-cystic fibrosis bronchiectasis etiology and clinical characteristics in adults evaluated in the southwest of Colombia.Materials and methods. We conducted a cross-sectional, non-interventional study. Subjects diagnosed with non-cystic fibrosis bronchiectasis were referred to by theirhealthcare providers and then enrolled between October 2018 and April 2021. Medical records and radiological studies were evaluated. Participants underwent laboratory tests, including complete blood count, serum immunoglobulin levels, and, in some cases, additional tests.Results. We included 161 subjects. The average age was 50 years old, and 59% were females. Bronchiectasis etiology was identified in 84.6% of the cases. Postinfectious(34.6%) and immune disorders (25.3%), represented by autoimmunity (13.6%) and immunodeficiency (11.7%), were the leading causes. Gender differences were noted in autoimmune (females: 18.8% versus males: 6.1%, p = 0.021) and immunodeficiency-related bronchiectasis (males: 21.2% versus females 5.2%, p = 0.002). Immunodeficiencies-associated bronchiectases were more frequent in subjects under 50 years of age, while chronic obstructive pulmonary disease-associated bronchiectases were common in subjects over 50 years of age.Discussion. The etiologies of non-cystic fibrosis bronchiectasis in Colombia are diverse, exhibiting notable differences from other global regions. Serum immunoglobulin levels and clinical immunologist consultation should be prioritized in diagnosing patients with unclear bronchiectasis etiology, particularly those with recurrent sinopulmonary infections.

Abstract Chemotherapy regimens for triple-negative breast cancer (TNBC) combine agents without knowing which agents drive response. Consequently, predictors derived from multi-agent regimens cannot be assumed to generalize to individual drugs or other regimens, motivating development of treatment-matched predictors. Here, we used TNBC patient-derived xenografts (PDXs) treated with carboplatin, docetaxel, or the combination to deconvolute drug-specific responses and identify associated molecular features. Combination treatment rarely improved upon the best single agent, with enhanced responses in only 13% of PDXs and antagonism in a comparable fraction. Proteogenomic analyses identified high cytokeratin-5 (KRT5) as a general marker of chemotherapy responsiveness and KRT5 immunohistochemistry discriminated responsive PDXs (AUROC, 0.83). To train treatment-specific predictors, we integrated these data with independent PDX and clinical cohorts with responses assessed after anthracycline-free platinum, taxane, or platinum-taxane therapy, ensuring response corresponded to the modeled treatment. Four feature selection strategies yielded largely nonoverlapping biomarker panels converging on treatment-relevant pathways. On independent test data, RNA-based predictors of complete response (CR) to platinum-based (carboplatin or cisplatin) and taxane-based (docetaxel or paclitaxel) chemotherapy achieved AUROCs of 0.80 and 0.86, respectively. For platinum-taxane regimens (carboplatin plus docetaxel or paclitaxel), proteomic-guided feature selection generated a 10-biomarker, protein- informed RNA predictor of pathologic complete response (pCR) that outperformed RNA-only feature selection and achieved an AUROC of 0.85 in an independent clinical cohort, while retaining practicality as an RNA-based assay. Treatment-matched integration of multi-omic PDX and clinical datasets provides a framework for chemotherapy-specific predictors with clinically relevant performance, supporting biomarker-guided precision selection and treatment optimization for patients with TNBC. Statement of significance Integration of multi-omic data from patient-derived xenografts with treatment-matched clinical cohorts yielded three retrospectively validated predictors of platinum, taxane, and platinum+taxane response, enabling biomarker-guided chemotherapy selection for patients with triple-negative breast cancer.

Background An increased risk of second cancers (SC), in patients with myeloproliferative neoplasms (MPN), has been suggested by a number of population-based and cancer registry studies (Blood 2011;118:6515). Such studies often included patients belonging to a broad category of MPN entities and did not always account for history of cancer prior to the diagnosis of MPN, associated comorbidities, mutations, or treatments received, in assessing risk association and prediction (Leukemia 2019;33:1996). The current study seeks to fill knowledge gaps, in this regard, in the context of a biologically more homogeneous MPN setting, vis-ẚ-vis polycythemia vera (PV) and essential thrombocythemia (ET). Methods The present study was conducted under an institutional review board approved minimum risk protocol that allowed retrospective collection and analysis of data from Mayo Clinic patient records. Diagnostic criteria were according to the International Consensus Classification (Blood 2022; 140:1200). Documented primary cancers were annotated in 17 categories, not including pre-cancerous conditions or MPN progressions. SC was defined as any cancer that developed after 3 months from the initial diagnosis of MPN. Cox proportional hazard regression model was used for univariable and multivariable analysis. Kaplan-Meier plots were used for time-to-event assessment (considering the first in case of multiple events). Results The study population included 1,968 consecutive patients (median age 60 years; 56% females) with PV (N=1,001) or ET (N=967), seen at the Mayo Clinic between 1970 and 2024. Median follow-up was 8.6 years. The incidences of arterial/venous thrombosis were14%/12% at diagnosis and 12%/10% after diagnosis. Co-morbid conditions included diabetes (12%), hypertension (52%), tobacco use (16%), and history of autoimmune/inflammatory disease (AID; 6%); hyperlipidemia was annotated in PV cases only (51%). Non-driver mutation information was available in 620 patients: TET2 (13%), ASXL1 (7%), DNMT3A (5%), SRSF2 (3%), SF3B1 (3%) and TP53 (2%). A history of another cancer was documented in 598 (30%) patients and included cancers documented before/at (N=291; 15%) or after (N=404; 20%) the diagnosis of MPN, with only the latter being referred to as “SC”. Most frequent cancer type was non-melanoma skin cancer (NMSC), accounting for 28% of cancers diagnosed before/at MPN, and 47% of SC; other SC types included lymphoid neoplasms (10%), melanoma (10%), prostate cancer (9%), and trachea/lung/bronchial cancer (8%), as the most frequent. On multivariable analysis, cancer history, before/at or after MPN diagnosis, clustered with age >60 years (OR 2.7), male sex (OR 1.5), and history of AID (OR 1.7). History of cancer prior to MPN diagnosis, NMSC or otherwise, was associated with inferior overall survival (p<0.01); by contrast, neither myelofibrosis-free (p=0.5) nor leukemia-free (p=0.8) survival was affected. Median time from MPN diagnosis to SC was 5.7 years. In multivariable analysis including all cancers, SC-free survival was adversely affected by age>60 years (HR 2.5; p<0.01), male sex (HR 1.7; p<0.01), history of previous cancer (HR 2.5, p<0.01), and AID history (HR 1.6; p<0.01); all these 4 risk factors remained significant after excluding NMSC. Interestingly, among informative cases, SRSF2 mutation also predicted SC (HR 2.6; p=0.02), even after NMSC cases were excluded (HR 3.4; p=0.02). Pre-MPN history of cancer did not affect either arterial (p=0.9) or venous thrombosis-free survival (p=0.7), in age- and sex-adjusted analysis. Conversely, after excluding post-SC thrombotic events, venous (p=0.02) but not arterial (p=0.1) events, occurring after MPN diagnosis, were predictive of SC. This association remained significant in multivariable analysis that included age, sex, and venous thrombosis history (p=0.03). Additional observations from this study, include an independent association between SC and hyperlipidemia in PV (p<0.01) and ruxolitinib monotherapy with NMSC (HR 9; p<0.01). Conclusions The risk of SC in patients with MPN is significantly higher in older patients, men, and in the presence of pre-MPN cancer history and AID. Incidental venous, but not arterial, thrombosis was associated with subsequent SC, not NMSC. The observed association of SC with SRSF2 mutations and hyperlipidemia is noted but requires further validation.

Background: PV is a myeloproliferative neoplasm that leads to excessive production of erythrocytes and increased platelet and leukocyte counts. The current standard of care is to use therapeutic phlebotomy (TP) with or without cytoreductive therapy (CRT) to treat PV. Over time, the CRT dose often needs to be increased to treat PV progression. Preliminary results from the phase 2 REVIVE study (NCT04057040) demonstrated that rusfertide was superior to placebo in achieving hematocrit (Hct) levels <45% and reducing or eliminating the need for TP in patients who were phlebotomy-dependent prior to study entry (Kremyanskaya et al. EHA2023. [Abstract LB2710]; Kremyanskaya et al. New Engl J Med 2024;390:723-35).Final data from REVIVE will be presented. Methods: In Part 1 (Weeks 1-29), subcutaneous rusfertide (starting dose: 20 mg q1w) was initiated and titrated to control Hct to <45%. In Part 2 (Weeks 29-41; blinded randomized withdrawal phase), patients were randomized to receive rusfertide or placebo. Randomized patients were eligible to participate in the open-label extension (OLE) portion of the study (Part 3), in which all patients could receive rusfertide and investigators could make CRT dose adjustments. Results: In Part 1, 70 patients were enrolled; 59 were randomized in Part 2. Fifty-eight patients continued to Part 3. As of 9 July 2024, 50 (71.4%), 37 (52.9%), and 17 (24.3%) patients received rusfertide for ≥2, ≥2.5, or ≥3 years, respectively.Overall, median (range) patient age was 57.5 years (27-77); the majority were male (70%) and had high-risk disease (55.7%). Overall, 52.9% and 47.1% of patients were treated with TP alone or TP plus CRT, respectively. Prior to enrollment, the estimated average (mean) phlebotomy rate (EAPR) in patients who enrolled on study was 8.5/year, which was reduced to <1.0/year in Part 1. In Part 2, the EAPR was <1.0/year and 6.6/year in the rusfertide and placebo groups, respectively. For patients who continued onto Part 3 and received rusfertide, the EAPR remained at <1.0/year. In Parts 1, 2, and 3, the mean (SD) weekly average rusfertide dose was 30.7 mg (12.2), 38.5 mg (21.3), and 44.9 mg (20.3), respectively. Rusfertide consistently maintained Hct <45%, including in patients receiving therapy for ≥3 years. Platelets increased following initiation of rusfertide therapy and stabilized over time; mean leukocyte counts remained stable. Prior to enrollment, iron-related parameters were consistent with systemic iron deficiency. Throughout the study, rusfertide resulted in the normalization of mean serum ferritin levels, increased mean transferrin saturation, and increased mean serum iron levels. Overall, mean corpuscular volume increased slightly over the duration of the study. Throughout the study, there was no evidence of increased markers of inflammation (eg, C-reactive protein or interleukin 6). The most common (≥20%) treatment-emergent adverse events (TEAEs) were injection site reactions (85.7%), fatigue (38.6%), COVID-19 (32.9%), pruritus (34.3%), arthralgia (30.0%), dizziness (30.0%), nausea (24.3%), headache (24.3%), and anemia (21.4%). Grade 3 TEAEs occurred in 25.7% of patients.There were no Grade 4 or 5 TEAEs. One patient developed acute myeloid leukemia after treatment discontinuation. After more than 150 patient-years of rusfertide exposure, malignancies (most of which were skin cancers) were reported in 11 patients (all had risk factors that may have contributed to the development of these malignancies). There was no obvious correlation between increasing exposure to rusfertide and malignancies reported. Six thrombotic events (5 arterial and 1 venous) occurred in 5 patients with high-risk PV. Conclusions: In REVIVE, rusfertide added to TP with or without CRT provided long-term durable control of Hct and decreased the need for TP. Patients are eligible to roll over to the phase 2 THRIVE OLE study (NCT06033586), which will continue to assess the long-term safety and efficacy of rusfertide for a total period of up to 5.8 years. The randomized phase 3 VERIFY study (NCT05210790) is ongoing and will evaluate rusfertide vs placebo in PV patients.

Background Previous studies have suggested a higher prevalence of autoimmune or inflammatory diseases (AID) in patients with myeloproliferative neoplasms (MPN) (Haematologica 2010; 95:1216); more recent studies have suggested the absence of impact on tumor genetics or disease outcome (Leukemia 2023; 37:1741). In the current study, we conducted a before and after MPN diagnosis survey, in order to examine the prevalence of AID and its impact on overall and transformation-free survival as well as the risk of thrombosis or other co-morbidities, such as cancer. Methods The current study was conducted under an institutional review board approved minimum risk protocol. Diagnostic criteria were according to the International Consensus Classification (Blood 2022; 140:1200). AID was operationally classified as occuring before or after diagnosis, with the latter requiring first diagnosis occuring ≥3 months after MPN diagnosis. Non-driver mutations were screened by next-generation sequencing (NGS) and was available for 620 patients. Cox proportional hazard regression model was used for univariate and multivariable analysis. Kaplan-Meier plots were used for time-to-event assessment, using JMP Pro 17.0.0 software SAS Institute, Cary, NC, USA. Results The study population included 1,968 consecutive patients with polycythemia vera (PV; N=1,001) or essential thrombocythemia (ET; N=967), seen at the Mayo Clinic between 1970 and 2024. Mutation information was available in 620 patients. AID was diagnosed before or after MPN diagnosis in 157 (8%) patients, including 121 (6%) before and 43 (4%) after. 134 (85%) patients had history of only one AID while 23 (15%) had ≥2 AIDs. The most prevalent diagnoses included organ-specific AID (48 cases before and 21 after MPN diagnosis), inflammatory arthritis (37 before and 6 after), connective tissue diseases (17 before and 7 after) and inflammatory dermatosis (20 before and 4 after). For each one of these four categories, thyroid disorders (21 before and 6 after), rheumatoid arthritis (22 before and 2 after), antiphospholipid syndrome (8 before and 1 after), and psoriasis (11 before and 4 after) were the most frequent, respectively. Patients with pre-MPN AID history were older (p=0.02) and were more likely to have ET vs. PV and be females (p<0.01); they were also more likely to present with venous (18% vs. 12%, p=0.04) but not arterial (p=0.2) thrombosis. Pre-MPN AID history was significantly associated with pre-MPN cancer history, with the latter including (25% vs 14%, p<0.01) or excluding (18% vs 10%, p<0.01) non melanoma skin cancers (NMSC). Pre-MPN AID history did not correlate with driver or other mutations (including TET2, ASXL1, DNMT3A), leukocyte, neutrophil, monocyte, or lymphocyte counts (p≥0.1 in all instances). Pre-MPN history of AID did not affect overall (p=0.08), myelofibrosis-free (p=0.3), leukemia-free (p=0.3), arterial thrombosis-free (p=0.9), or venous thrombosis-free (p=0.6) survival. Second cancer-free survival (SC-FS), including (p<0.01) or excluding (p<0.01) NMSC, was negatively affected by pre-MPN history of AID. Multivariate analysis confirmed the independent detrimental effect of a pre-MPN history of AID (HR 1.7, p<0.01) on SC-FS; other independent risk factors for SC-FS were older age (p<0.01), male sex (p<0.01) and pre-MPN history of cancer (p<0.01). The association between pre-MPN history of AID and second cancer was also apparent when the latter was limited to lymphoproliferative neoplasms, although with borderline significance (p=0.08), which was further confirmed by adjusting for age and sex (p=0.09). Median time from MPN diagnosis to post-MPN AID was 9 years (range 1-23), which was similar between PV and ET (p=0.5). Post-MPN development of AID was more likely in females (HR 2.3; p=0.01) and in patients with pre-MPN history of AID (HR 4.8; p<0.01) or hypertension (HR 2.1; p<0.03). These risk factors remained significant during multivariable analysis of post-MPN AID-free survival. Conclusions The current study suggests an increased risk of post-MPN AID and second cancers, in patients with pre-MPN history of AID. The study also showed a significant association between pre-MPN history of AID and venous thrombosis at time of MPN diagnosis. These observations reinforce the widely suggested role of tumor-extrinsic inflammation in exacerbating MPN-intrinsic thrombophilia and the development of cancer.

Background Conventional thrombosis risk stratification in polycythemia vera (PV) considers age > 60 years and thrombosis history for assignment to low (no risk factor) or high (at least 1 risk factor) risk category (J Clin Oncol.2011;29(6):761-70). The current study exclusively focused on low-risk PV with the following objectives: i) estimate the incidence of arterial and venous thrombosis; ii) examine the prognostic relevance of clinical and genetic risk factors for thrombosis; and iii) assess the impact of treatment on thrombosis risk. Methods Patients were retrospectively recruited from the Mayo Clinic, USA, and University of Florence, Italy, PV databases. Diagnosis was according to the International Consensus Classification criteria (Blood 2022;140:1200). Only first major arterial (AT) and unprovoked venous thrombosis (VT) were considered. Conventional statistical methods were applied (JMP Pro 17.0.0, SAS Institute, Cary, NC, USA). Results Mayo Clinic cohort A total of 346 patients with low-risk PV were considered: median age 49 years; males 54%; median hemoglobin 17.7 g/dl; median leukocyte count 10.9 x 109/L; leukocyte count >11 x 109/L 49%; median platelet count 471 x 109/L; cardiovascular (CV) risk factors 47%; hypertension 29%; hyperlipidemia 20%; smoking history 12%; diabetes mellitus 5%. Most frequent mutations, other than JAK2, were ASXL1 (13%) and TET2 (13%). Median follow-up was 13.4 years (1-52) (4,636 patient-years) during which 25 (7%) AT and 20 (6%) VT were documented. The corresponding incidence rates were 5.4 and 4.3 per 1,000 patient-years. In multivariable logistic regression analysis, baseline hemoglobin >17.5 g/dl in women or >20 g/dl in men (15% vs 4%; OR 5.1; p<0.01) and absence of leukocytosis >11 x 109/L (10% vs 4%; OR 2.9; p=0.02) were associated with increased risk of AT; in addition, diabetes mellitus (19% vs 7%; OR 3.9; p=0.06), showed borderline significance; overall incidence of AT was 20% in the presence of ≥2 risk factors (n=61), 7% in the presence of one risk factor (n=177), and 1% in the absence of all three risk factors (n=108; p<0.01). Multivariable analysis for arterial thrombosis-free survival (TFS) confirmed the predictive performance of these risk factors: hemoglobin thresholds (HR 4.4; p<0.01); absence of leukocytosis >11 x 109/L (HR 2.7; p=0.03); and diabetes (HR 2.8; p=0.09); 20-year cumulative incidence of AT was 32% in the presence of ≥2 risk factors and 7% otherwise (p<0.01). In multivariable analysis, smoking history was found to be surprisingly protective against VT (0% vs. 7%; p=0.02) while female gender was associated with a borderline higher risk (8% vs 4%; OR 2.2; p=0.09); VT incidence rates were 9% in the presence of both risk factors (n=144), 4% with one risk factor (n=172), and 0% in the absence of both risk factors (n=40; p=0.02); 20-year cumulative incidence of VT was 10% in the presence of both risk factors and 3% otherwise (p=0.03). ASXL1 and TET2 mutations did not appear to modify the risk of either VT (p=0.59) or AT (p=0.66). Treatment at diagnosis included aspirin and cytoreductive therapy in 63% and 24% of patients, respectively. Aspirin but not cytoreductive therapy was associated with a lower incidence of AT (5% vs 11%; 2.3 vs 3.0 per 1,000 patient-years; p=0.04) and VT (4% vs 10%; 1.9 vs 2.8 per 1,000 patient-years; p=0.03). Thrombosis-free survival was not influenced by documented treatment at diagnosis or at the time of event (p>0.1). University of Florence cohort A total of 284 low-risk PV patients were followed for a median of 9.1 years (1-38) with 19 (7%) AT and 24 (8%) VT events recorded; the corresponding incidence rates were 7.4 and 9.3 per 1,000 patient-years. In this Florence cohort, risk factors for AT were diabetes mellitus (HR 3.9), hyperlipidemia (HR 3.8) and advanced age, with only age remaining significant during multivariable analysis (p=0.03). Predictors of VT included hyperlipidemia (HR 4.2) and hypertension (HR 2.1). Treatment with aspirin or cytoreductive therapy did not impact thrombosis-free survival (p>0.1). Conclusion The current study confirms the infrequent occurrence of AT and VT in low-risk PV but also exposes the challenges in identifying risk factors that are reproducible in different but otherwise well-characterized patient cohorts. Discrepancies were also apparent in retrospective comparison of treatment effects on thrombosis, which reinforces the need for prospective studies.

Background Abnormal karyotype is not infrequent in polycythemia vera (PV) and has been associated with shortened survival (BJH 2018;182:437). In the current study, we utilized a large institutional database, in order to describe the spectrum and prevalence of specific cytogenetic abnormalities in PV, during different stages of the disease, and examine their clinical, molecular, and prognostic correlates, in the context of previously established risk factors. Methods After institutional review board approval, we identified 669 cytogenetically-annotated patients with PV, evaluated at the Mayo Clinic between 1973 and 2023, and met the International Consensus Classification diagnostic criteria (Arber et al. Blood 2022;140;1200). Cytogenetic reporting was in accordance with the International System for Human Cytogenomic Nomenclature. Results i) Prevalence of cytogenetic abnormalities at diagnosis and at different stages of the disease 436 of 669 (65%) patients with PV underwent cytogenetic evaluation within one year of diagnosis, 116 (17%) within 1-10 years, 47 (7%) after 10 years, and 70 (10%) at time of fibrotic or leukemic transformation. In patients with cytogenetic information obtained within 1 year of diagnosis, karyotype was normal in 369 (85%), showed loss of Y chromosome (-Y) in 15 (3%), and other abnormalities in 52 (12%). Most common abnormalities included trisomy 9 (n=11; 3%), del(20q) (n=10; 2%), trisomy 8 (n=4; 1%), other sole abnormalities (n=16; 4%), and ≥ 2 abnormalities (n=11; 3%) including complex karyotype (n=4; 0.9%). Frequency of abnormal karyotype was higher with longer disease duration; 15% within a year of diagnosis, 28% in 1-10 years, 49% over 10 years after diagnosis, and 50% at time of fibrotic or leukemic transformation (p<0.01). Sole del (20q), single abnormalities excluding 20q-, +8, +9, or -Y, and ≥ 2 abnormalities including complex karyotype were progressively acquired; respective frequencies within a year of diagnosis, over 10 years after diagnosis, and at time of fibrotic or leukemic transformation were 2%/21%/10% (del (20q), 4%/17%/14% (single abnormality excluding 20q-, +8, +9, or -Y), 3%/11%/19% (≥ 2 abnormalities), 1%/ 2%,/10% (complex karyotype) (all p-values <0.01). In addition, sole +8 displayed highest frequency at fibrotic transformation compared to within a year of diagnosis (5% vs 0.9%; p=0.04). ii) Phenotypic and Molecular Correlates 436 patients with PV (51% males; median age; 64 years) who underwent cytogenetic testing within one year of diagnosis were examined for clinical and molecular correlations. Abnormal karyotype compared to normal karyotype, correlated with older age (median: 70 vs 63 years, p<0.01), male gender (64% vs 48%, p=0.02) and lower platelet count (370 vs 465 × 109/L, p<0.01). Sole del (20q) compared to normal karyotype was associated with older age (median: 76 vs 63 years, p<0.01); sole +8 exclusively occurred in patients ≥ 60 years (p=0.03); sole +9 was associated with venous thrombosis history (45% vs 17%, p=0.03). TET2 and SRSF2 mutations were found to cluster with normal karyotype (18% vs 4%; p=0.04 and 5% vs 0%; p=0.14 in normal vs abnormal karyotype, respectively). iii) Prognostic Impact of Cytogenetic Abnormalities At a median follow-up of 7.4 years (range; 0.04-42.4 years), a total of 163 (37%) patients have died, 50 (11%) and 14 (3%) underwent fibrotic and leukemic transformation, respectively. In univariate analysis for overall survival (OS), abnormal karyotype compared to normal karyotype, was associated with inferior survival (median OS; 10.5 vs 16.3 years, p<0.01). In multivariate analysis, the adverse impact of abnormal karyotype remained significant (p=0.02; HR 2.0, 95% CI 1.2-3.3), along with age ≥60 years, leukocyte count ≥15 × 109/L, and SRSF2 mutations. Abnormal karyotype was also an independent predictor of myelofibrosis-free survival (p<0.01; HR 3.7, 95% CI 1.8-7.8), along with SRSF2 mutations. Analysis of leukemia-free survival revealed abnormal karyotype to be a significant risk factor in univariate (p<0.01), but not multivariate analysis that included SRSF2 mutation (p>0.1). Conclusions The current study provides the distribution of cytogenetic abnormalities at different stages of PV and affirms the prognostic significance of abnormal karyotype at the time of diagnosis, highlighting the importance of incorporating cytogenetic studies in the diagnostic evaluation of PV.

Background: JAK2 mutations are invariably associated with polycythemia vera (PV); more than 50% of affected patients also harbor non-JAK2 mutations, some considered prognostically relevant (BJH 2020;189:291). We queried a large single institutional database of patients with PV, in order to examine the incidence and distribution of non-JAK2 mutations, across different disease stages, in order to assess their impact on disease phenotype and prognosis, in the context of other risk factors. Methods: Mayo Clinic databases (1998-2023) were searched to identify next-generation sequencing (NGS)-annotated patients with PV, with diagnosis retrospectively fitted to meet the 2022 International Consensus Classification (ICC) criteria (Blood 2022; 140:1200). NGS was performed either through research-based targeted capture assays or a clinically ordered 42-gene panel. Conventional statistical methods were used for group comparisons and survival analyses. Results: i) Mutational frequencies in chronic vs. transformed disease phases 319 patients with PV were considered; NGS was performed at time of i) diagnosis or before fibrotic or leukemic transformation (N=270; 84.6%), ii) fibrotic progression (N=37; 11.6%), or iii) leukemic transformation (N=12; 3.8%). Mutational frequencies in newly-diagnosed/chronic phase PV were 19% for TET2, 10% for ASXL1, 4% for SRSF2, ⁓3% each for SF3B1, DNMT3A, and SH2B3, and 1-2% each for TP53 and IDH2. Significant differences in mutation distribution were apparent for TP53/SRSF2/IDH1/U2AF1 in blast (50%/25%/17%/8%) vs. fibrotic (8%/0%/0%/5%) vs. chronic (2%/4%/2%/0.4%) phase (p<0.01, <0.01, =0.03, and =0.03, respectively). ii) Phenotypic correlates of specific mutations among patients with NGS information obtained during chronic phase (N=270) Median age was 61.4 years (51% males) and incidence rates included 22% for leukocytosis ( ≥15 x 10(9)/L) and 25%/14% for arterial/venous thrombosis history. Numerically prevalent or prognostically relevant mutations were selected for comparison of clinical or laboratory characteristics: TET2, ASXL1, SRSF2, TP53, IDH2. Significant associations were apparent for ASXL1 (younger age distribution), SRSF2 (older age distribution, leukocytosis), and TP53 (leukocytosis) mutations. Median age was 66.4, 58.7, 76.8, 63.9, and 68.6 years for TET2, ASXL1, SRSF2, TP53, IDH2 mutations, respectively (p<0.01). The corresponding median leukocyte counts were 12.1, 11.4, 14.4, 16.7, and 11.5 x 10(9)/L (p<0.01). iii) Prognostic impact of mutations detected at diagnosis or chronic phase (N=270) At median follow-up of 9.5 years (range 0.1-42.4), 115 (43%) deaths, 12 (4%) leukemic transformations, and 53 (20%) fibrotic progressions were documented. In univariate analysis of mutations, overall survival (OS) was adversely affected by SRSF2 and IDH2 mutations, leukemia-free survival (LFS) by ASXL1, SRSF2, IDH2, RUNX1, and CEBPA mutations, and myelofibrosis-free survival (MFS) by SRSF2 mutation. Age-adjusted (p<0.01) multivariable analysis, including mutations and other risk variables, identified SRSF2 (HR 4.2, 95% CI 1.9-9.5), ASXL1 (HR 2.0, 1.1-3.7), IDH2 (HR 5.3, 1.8-15.3) mutations and leukocytosis ≥15 x 10(9)/L (HR 2.0, 1.3-3.1) as independent predictors of inferior OS. SRSF2 and ASXL1 mutations were also independently associated with inferior LFS (p=0.02 and 0.06, respectively) and SRSF2 (p<0.01) and karyotype (p<0.01) with inferior MFS (p<0.01). Median survival for 235 patients without any adverse mutation (ASXL1, SRSF2, or IDH2) was 17.8 years vs. 8.8 years for 35 patients with any one of the three adverse mutations (p=0.01; HR 1.8, 95% CI 1.1-2.9). The number of non-JAK2 mutations, in general, was also associated with significantly different survival outcomes: median OS was 20, 15.1, and 9.3 years for patients with non-JAK2 mutations numbering zero, one, or two or more, respectively (p<0.01) Conclusions: SRSF2 mutations in PV cluster with older and ASXL1 with younger age distribution. SRSF2 and TP53 mutational frequencies were higher in blast-phase disease. SRSF2, ASXL1, and IDH2 mutations were significantly associated with inferior OS, independent of each other and other risk factors. SRSF2 mutations were also detrimental to LFS and MFS and ASXL1 to LFS. Such information might allow further refinement of current risk models for PV, which in future studies should also account for variant allele frequency.