Moliterno, Alison R.

Abstract Background: Phlebotomy to maintain hematocrit <45% is considered one of the cornerstones for the management of polycythemia vera (PV). However, phlebotomy procedures may be inconvenient for or poorly tolerated by some patients, and often result in iron deficiency, which may be associated with additional quality-of-life burden, such as fatigue, impaired cognitive function and restless leg syndrome. The REVEAL study (ClinicalTrials.gov, NCT02252159) is being conducted to describe contemporary demographics, burden of disease, clinical management, patient-reported outcomes, and healthcare resource utilization among patients with PV in the US. This early analysis describes the patient self-reported burden of phlebotomy at the time of enrollment. Methods: REVEAL is a multicenter, noninterventional, nonrandomized, prospective, observational study of adult patients with PV who are currently under the care of a US physician. REVEAL collects physician assessments and patient-reported outcomes during a 36-month observation period. Phlebotomy-related burden is being assessed with a new 21-item phlebotomy burden questionnaire (PBQ-21) at enrollment and every 90 days thereafter; it assesses patient-reported phlebotomy frequency, practice setting, time required for phlebotomy, inconvenience, and phlebotomy-related adverse effects. Phlebotomy-related effects and bother/inconvenience (if experienced within 24 hours after a phlebotomy procedure) are graded on a 4-point scale of 1 (not at all) to 4 (extremely). Analyses of these preliminary data were descriptive. Results: At the time of data cutoff, 865 patients were enrolled (total planned enrollment, n=2000). Median age was 67 (range, 22-95) years, 55.5% were male, and 89.5% were white. Median time from PV diagnosis to study enrollment was 53 months. Overall, 748 patients completed the PBQ-21 at enrollment and 400 (53.5%) had received a phlebotomy within 3 months prior to enrollment. Mean number of phlebotomies in the past 3 months was 2.2 (SD, 1.7). Phlebotomy was most commonly performed at a doctor's office during a regular medical visit (37.8%), at an infusion center or special cancer care center (26.8%), or at a hospital (18.3%). Patients who had phlebotomy procedures within 90 days before enrollment reported a mean total treatment time of 4.2 (SD, 4.7) hours per phlebotomy procedure; for employed patients, the mean amount of work time missed was 4.6 (SD, 15.6) hours per procedure. Fatigue, bruising, dehydration, and dizziness were among the most commonly reported adverse effects of phlebotomy (Table 1). Twenty percent of patients reported that phlebotomy procedures were moderately or extremely bothersome, inconvenient (18.3%), or painful or physically uncomfortable (16.3%). Furthermore, some patients (8.8%) reported that their family and friends were moderately or severely inconvenienced by phlebotomy procedures. Conclusion: The PBQ-21 collects important and relevant patient-reported information to systematically assess the impact of phlebotomy on patients and caregivers. From the early view of REVEAL enrollment data, a considerable proportion of patients reported fatigue, bruising, dehydration, and dizziness resulting from phlebotomy procedures. Furthermore, patients reported spending a considerable amount of time (4 hours on average) per phlebotomy procedure, which is a sizeable burden for patients who are employed. In addition, some patients reported that phlebotomy procedures may be inconvenient, painful or uncomfortable, and bothersome. REVEAL provides important information for providers to understand and address when to prescribe phlebotomy for patients with PV. Disclosures Boccia: Incyte Corporation: Honoraria. Stein:Incyte Corporation: Consultancy, Membership on an entity's Board of Directors or advisory committees. Mesa:Genentech: Research Funding; Promedior: Research Funding; NS Pharma: Research Funding; CTI Biopharma: Research Funding; Gilead: Research Funding; Incyte Corporation: Research Funding; Novartis Pharmaceuticals Corporation: Consultancy; Pfizer: Research Funding. Naim:Incyte Corporation: Employment, Equity Ownership. Cordaro:Incyte Corporation: Employment, Equity Ownership. Peng:Incyte Corporation: Employment, Equity Ownership. Sun:Incyte Corporation: Employment, Equity Ownership. Parasuraman:Incyte Corporation: Employment, Equity Ownership. Moliterno:Incyte Corporation: Membership on an entity's Board of Directors or advisory committees.

Abstract Abstract 400FN2 Patients with primary and secondary myelofibrosis (MF) have a worse prognosis than those with essential thrombocytosis (ET) or polycythemia vera (PV). MF and chronic myelomonocytic leukemia (CMML) share chromosomal and genetic lesions such as ASXL1 and JAK2 mutations, but effective treatments are lacking. The aim of the study was to identify perturbed pathways in MPN and CMML that were amenable to targeted molecular therapies. In a large genomic analysis of MPN cases, we identified recurrent deletions of multiple DNA repair genes, including BRCA2 and Fanconi anemia (FANC) proteins, which have been associated with poly (ADP-ribose) polymerase (PARP) inhibitor sensitivity. Neoplastic cells from these patients are hypersensitive to PARP inhibitors in vitro and may provide a therapeutic strategy to improve treatment response for patients with advanced MPNs. Genomic screens utilizing SNP-A karyotyping were performed on primary cells from 144 patients with either ET, PV, or MF (90 were previously reported, McDevitt et al., ASH 2009). We focused on the potential functional relevance of recurrent genomic deletions, including BRCA2, by performing a GO analysis using the EASE software application (Hosack et al. 2003). This revealed statistically significant (p≤0.05) enrichment of 13 biological pathways. Notably, genes involved in cell cycle regulation and DNA repair (DR) were enriched in MF loss regions (p=0.007) as compared to regions lost in ET and PV. Several genes directly involved in DR including BRCA2, XRCC2, and several members of the FANC pathway, also critical for DNA crosslink repair, were deleted or present in copy neutral LOH regions. DNA methyltransferase inhibitors are being tested clinically in MF patients, and both cell cycle and DNA repair genes have been identified as epigenetically silenced in hematological malignancies. We therefore screened for BRCA1, BRCA2, FANC-C, L, and N promoter CpG island methylation in a series of 48 MPN samples. No methylation was identified in BRCA2 and FANC members, suggesting a potential haploinsufficiency mechanism. Methylation of BRCA1 was identified in a small number of cases and is under further study in a larger cohort to investigate the effects on BRCA1 expression, DR and sensitivity to PARP inhibitors. Based on these genetic and epigenetic findings, we then focused on a functional analysis of the homologous recombination (HR) pathway. Leukocytes from normal controls and MPN patients were exposed to ionizing radiation (IR) and stained for Rad51 foci, a marker of HR activity. Six of 15 MPN/CMML samples (40%), but none of the controls failed to form IR-induced Rad51 foci, suggesting impaired HR. Of the samples that demonstrated defects in HR [MPN-U, MF, CMML (2), MDS/MPN-U, and CML], there was no significant correlation between the MPN subtype, the JAK2 mutation status, or the specific metaphase karyotype, but the most sensitive case in our series exhibited a deletion of Ch13q that spanned the BRCA2 gene. We then examined sensitivity of MPN vs. normal myeloid progenitors to the PARP inhibitors veliparib and/or olaparib in methylcellulose (MC)-based colony forming assays. Failure to form Rad51 foci with irradiation was statistically correlated with PARP inhibitor sensitivity (p = 0.021). When an expanded series of cases were then analyzed for PARP inhibitor sensitivity using the MC colony assay, lower IC50s for both veliparib and olaparib were observed in MPN cases (N= 25, IC50S=2.69uM vs. 11.2uM, N=15 IC50S=.64uM vs. 1.27uM, respectively) relative to normal controls (p =.0003, p =.05, via Wilconon rank-sum test respectively). Two cases in this series were treated on a clinical trial combining the ABT-888 PARP inhibitor with salvage chemotherapy consisting of carboplatin and topotecan (Karp et al., ASH 2010). Both patients, one with sAML evolving out of PPV-MF, and a second with accelerating MPN-U, had failed four prior therapies but achieved complete clearance of bone marrow blasts at day 14. Interestingly, cells from this MPN-U patient (13q-) were the most sensitive to PARP inhibition in vitro. Additional accrual and correlative evaluations are underway and will be updated at the meeting. Collectively, these results demonstrate that a subset of MPN and CMML patients have intrinsic genetic and epigenetic defects affecting the FANC and HR pathways, providing pre-clinical rationale for the use of PARP inhibitors in the treatment of aggressive MPN and CMML. Disclosures: No relevant conflicts of interest to declare.

Abstract Abstract 627 Myeloproliferative diseases (MPD) are clonal hematologic disorders that present with increased numbers of functional, mature, terminally differentiated myeloid elements. Even though genetic, biochemical and functional studies have provided important insights into the pathogenesis of MPDs, the role of epigenetic changes in disease pathobiology is not well elucidated. We performed the HELP assay to study genome-wide methylation patterns in cases of Polycythemia Vera (PV), Essential Thrombocytosis (ET) and Idiopathic Myelofibrosis (IMF) and compared it with normal matched controls. The HELP assay uses differential methylation specific digestion by HpaII and MspI followed by amplification, two color labeling and hybridization to quantitatively determine individual promoter CpG methylation of 25636 loci. Analysis of 26 MPD neutrophil samples comprising 9 cases of ET, 6 cases of PV and 11 Cases of IMF was performed and compared to normal healthy controls. Unsupervised clustering based on global methylation profiles showed that IMF cases formed a distinct epigenetic cluster, while PV and ET cases were more similar to the normal controls. Further analysis of epigenetic differences between these groups showed that PV and ET samples were characterized by aberrant hypermethylation when compared to controls and had 143 genes that were uniformly hypermethylated. These genes are involved in pathways regulated by the NF-Kb and HNF-4alpha transcription factors. IMF on the other hand was characterized by both aberrantly hyper (n=162) and hypomethylated (n=95) loci when compared to controls. The pathways affected by hypomethylated genes were involved in cytokine cell signaling and MAP kinases. We subsequently validated these observations in an independent set of 8 IMF cases compared to 8 age matched controls. Methylation profiles obtained from whole blood by the HELP assay were able to clearly separate IMF cases from controls demonstrating the validity of our observations. These changes were quantified on a whole genome level by the Luminometric methylation assay (LUMA) that also revealed significantly more hypomethylation in IMF when compared to PV and ET cases (49% hypomethylation in IMF vs. 38% in PV/ET, p<0.05). Selected changes were validated by Mass Array sequencing. We next wanted to determine the epigenomic effects of Jak2V617F mutation and compared methylation profiles of MPD cases with and without the mutation. We observed that cases with Jak2 mutation had a higher number of differentially hypomethylated loci. This striking difference was seen by the LUMA assay also with 67% hypomethylation seen in mutant cases when compared to 48% in those without the mutation (p=0.02). This observation was validated in vitro in a cell line (FDCP) that expressed wither WT or mutant Jak2 kinase. We observed that expression of the mutant kinase led to global hypomethylation. This observation builds on recent data demonstrating nuclear binding of the mutant Jak2 kinase and its effects on the histone epigenetic machinery. In conclusion, we report that MPDs are characterized by various novel epigenetic alterations that affect important functional pathways and IMF is grossly epigenetically distinct from ET and PV. The Jak2 mutation also affects the methylome and leads to global hypomethylation that potentially contributes to the genomic instability and disease pathobiology. Disclosures: No relevant conflicts of interest to declare.

Abstract Background: Patients with polycythemia vera (PV) can have a pronounced symptom burden that negatively impacts their quality of life (QoL). Traditional treatment options such as phlebotomy, hydroxyurea, and interferon provide clinical benefit but frequently do not alleviate symptoms. The ongoing REVEAL study is being conducted to describe contemporary demographics, burden of disease, clinical management, patient-reported outcomes, and healthcare resource utilization among patients with PV in the US. This preliminary analysis describes the burden of PV based on patient-reported outcomes (PROs), including health-related QoL and work productivity. Methods: REVEAL is a multicenter, noninterventional, nonrandomized, prospective, observational study. Eligible patients are ≥18 years of age with a PV diagnosis and currently under active management by a physician in the US. PROs are being collected at study enrollment and every 3 months thereafter. Each of the 10 items on the Myeloproliferative Neoplasm Symptom Assessment Form (MPN-SAF) are graded from 0 (absent) to 10 (worst imaginable), with individual symptom scores ≥7 considered severe; total symptom score (TSS) ranges from 0-100. Raw scores for items on the 30-item European Organisation for Research and Treatment of Cancer Quality-of-Life Questionnaire-Core 30 (EORTC QLQ-C30) are standardized by linear transformation to 0-100. Higher scores on the EORTC QLQ-C30 indicate better functioning for global health status/QoL and functional subscales and worse severity for symptom subscales. Outcomes with the 6-item Work Productivity and Activity Impairment Specific Health Problem questionnaire (WPAI-SHP) are presented as a proportion from 0% (minimal/no impairment) to 100% (maximal impairment/productivity loss). Analyses of these preliminary data were descriptive. Results: At the time of data cutoff (June 9, 2015), REVEAL had enrolled 865 patients (total planned enrollment, n=2000), the majority were male (55.5%) and white (89.5%), with a median age of 67 (range, 22-95) years. 782 patients had completed the PRO questionnaires at the enrollment visit. A majority of patients reported ≥1 PV-related symptom. Mean (SD) MPN-SAF symptom scores were highest for fatigue (3.5 [2.7]), early satiety (2.7 [2.8]), inactivity (2.7 [2.9]), itching (2.6 [3.1]), and concentration problems (2.1 [2.6]). The mean (SD) MPN-SAF TSS (sum of 10 symptom scores) was 19.6 (16.1). The most frequently reported severe symptoms were fatigue, itching, inactivity, early satiety, night sweats, concentration problems, and bone pain (Table 1). Mean (SD) global health status/QoL reported on the EORTC QLQ-C30 was 72.8 (23.4); functional subscales were as follows: cognitive functioning, 80.3 (24.5); emotional functioning, 80.7 (24.0); physical functioning, 83.1 (20.6); role functioning, 84.3 (26.1); and social functioning, 85.0 (25.0). The most severe mean (SD) symptom subscale scores were fatigue (30.7 [28.5]), insomnia (29.5 [34.5]), pain (20.9 [29.3]), and dyspnea (17.3 [28.6]). Among patients who were employed at baseline (n=273), mean (SD) PV-related WPAI-SHP scores were as follows: absenteeism (ie, work time missed), 2.9% (10.6%); presenteeism (ie, impairment at work/reduced on-the-job effectiveness), 10.7% (20.4%); work productivity loss (ie, absenteeism plus presenteeism), 12.0% (22.2%); and activity impairment, 21.0% (28.0%). Conclusion: Preliminary descriptive data from patients currently enrolled in REVEAL suggest that patients with PV have notable impairment in QoL because of their disease; patient-reported scores on the MPN-SAF and EORTC QLQ-C30 were similar to those reported previously for patient populations with MPNs. Similarly, PV is associated with marked work productivity loss among patients who are currently employed. Subsequent analyses from REVEAL will evaluate changes over time in PROs and investigate correlations with disease progression and treatment. REVEAL can provide novel insights into questions related to the contemporary real-world QoL burden and its impact on PV-related symptoms and work productivity. Disclosures Mesa: Pfizer: Research Funding; NS Pharma: Research Funding; Gilead: Research Funding; Genentech: Research Funding; Novartis Pharmaceuticals Corporation: Consultancy; Incyte Corporation: Research Funding; CTI Biopharma: Research Funding; Promedior: Research Funding. Boccia:Incyte Corporation: Honoraria. Moliterno:Incyte Corporation: Membership on an entity's Board of Directors or advisory committees. Naim:Incyte Corporation: Employment, Equity Ownership. Cordaro:Incyte Corporation: Employment, Equity Ownership. Peng:Incyte Corporation: Employment, Equity Ownership. Sun:Incyte Corporation: Employment, Equity Ownership. Parasuraman:Incyte Corporation: Employment, Equity Ownership. Stein:Incyte Corporation: Consultancy, Membership on an entity's Board of Directors or advisory committees.

Abstract Background: Patients with polycythemia vera (PV) have an increased symptom burden (eg, fatigue, pruritus, and splenomegaly-related symptoms) and an increased risk of mortality compared with the general population, often resulting from thrombotic events, disease transformation to myelofibrosis, or secondary malignancies. There are limited data regarding PV burden and treatment patterns in a contemporary, real-world US setting. REVEAL is an ongoing observational study being conducted to describe contemporary demographics, burden of disease, clinical management, patient-reported outcomes, and healthcare resource utilization among patients with PV in the US. The objective of this assessment is to describe the clinical characteristics at enrollment among patients currently enrolled in REVEAL (enrollment from July 22, 2014 to June 9, 2015). Methods: REVEAL is a multicenter, noninterventional, nonrandomized, prospective, observational study. Eligible patients are ≥18 years of age with a PV diagnosis and currently under active management by a physician in the US. Data regarding disease and clinical characteristics, patient-reported outcomes, and health resource utilization are being collected at usual-care visits over a planned 36-month observation period. Analyses of these preliminary data were descriptive. Results: At data cutoff, 865 patients were enrolled (total planned enrollment, n=2000) from 174 sites (academic, n=27; community, n=147). Median (range) age at enrollment was 67 (22-95) years, 55.5% were male, 89.5% were white, and 45.4% had a history of smoking. Most patients (99.4%) had health insurance, including Medicare (56.0%), PPO (24.3%), or HMO (11.6%). Fifty-two percent were retired, 29.6% were employed, and 4.5% were unable to work or disabled. A total of 40.2% of patients completed a college/graduate degree, 19.9% had some college, and 32.5% completed high school or the equivalent. Documented major World Health Organization diagnostic criteria included JAK2V617F mutation in 50.9% of patients, elevated hemoglobin in 52.3%, and red cell mass in 5.5%. A bone marrow biopsy was conducted in 24.0% of patients, approximately half of whom (52.4% of the 24.0%) had trilineage myeloproliferation. Erythropoietin levels were abnormal in 19.7% of patients. Approximately half (46.6%) of patients had physician-reported PV-related symptoms at diagnosis, most frequently fatigue (25.2%) and pruritus (14.0%). At enrollment, median disease duration was 52.6 months; 42.3% of patients were diagnosed ≥5 years before enrollment. The most common comorbidities were hypertension (51.3%), obesity (14.2%), dyspnea (13.4%), myalgia (11.2%), and diabetes mellitus (11.1%). Mean hematocrit was 44.9%, hemoglobin was 14.5 g/dL, white blood cell count was 11.4×109/L, platelet count was 363.7×109/L, and absolute neutrophil count was 363.7×109/L. Twenty-one percent of patients had a history of thrombotic events, with 13.1% and 8.6% of patients experiencing ≥1 venous or arterial thromboembolic event, respectively. Of the 261 reported thrombotic events, 43.7% occurred after diagnosis of PV. The most common PV symptoms recorded at enrollment (using the Myeloproliferative Neoplasm Symptom Assessment Form) were fatigue (37.1%), pruritus (24.0%), headache (18.6%), insomnia (17.0%), dizziness (13.2%), bone pain (10.5%), and blurred vision (7.9%); 19.7% of 518 evaluable patients had palpable splenomegaly. Fewer than half of patients were receiving aspirin at enrollment (47.7%). The most common PV treatments ± aspirin were phlebotomy (PBT; 63.5%), hydroxyurea (HU; 42.2%), watchful waiting (5.3%), and ruxolitinib (3.1%); 23.0% of patients received PBT + HU. Conclusion: The REVEAL study is unique in that it captures data from both academic and community centers, and therefore includes a broader segment of the PV population typically not described in the literature. REVEAL can provide novel insights into questions related to the contemporary demographics of PV, practice patterns regarding diagnosis and therapy, and the real-world burden of PV and its impact on patients' quality of life and work productivity. Preliminary descriptive data from this early subset of patients suggest that a high proportion of patients report having PV-related symptoms at diagnosis, fewer patients are treated with aspirin than expected, and that thromboembolic event rates remain high after diagnosis. Disclosures Stein: Incyte Corporation: Consultancy, Membership on an entity's Board of Directors or advisory committees. Moliterno:Incyte Corporation: Membership on an entity's Board of Directors or advisory committees. Boccia:Incyte Corporation: Honoraria. Naim:Incyte Corporation: Employment, Equity Ownership. Cordaro:Incyte Corporation: Employment, Equity Ownership. Peng:Incyte Corporation: Employment, Equity Ownership. Sun:Incyte Corporation: Employment, Equity Ownership. Parasuraman:Incyte Corporation: Employment, Equity Ownership. Mesa:Promedior: Research Funding; Novartis Pharmaceuticals Corporation: Consultancy; NS Pharma: Research Funding; CTI Biopharma: Research Funding; Gilead: Research Funding; Pfizer: Research Funding; Incyte Corporation: Research Funding; Genentech: Research Funding.

Abstract Background:Patients with polycythemia vera (PV) often present with a broad range of clinical characteristics that may contribute to increased risks of cardiovascular (CV) morbidity and mortality, including thrombotic events (TE). Limited contemporary real-world data have been reported about the clinical burden of PV and treatment patterns in the United States. The ongoing REVEAL study collects data on disease burden, clinical management, patient-reported outcomes, and healthcare resource utilization for patients with PV in the United States. This analysis reports clinical characteristics, including underlying CV risk factors, for patients enrolled in the REVEAL study as of April 28, 2016. Methods: REVEAL is a multicenter, nonrandomized, prospective, observational study enrolling patients ≥18 years of age with a PV diagnosis who are actively managed in an academic or community setting. For this analysis, data regarding PV disease and diagnosis, clinical characteristics, and treatment patterns were collected at enrollment during usual-care visits and were based on physician assessment, electronic medical records, and local laboratory values. Ten-year CV risk factors selected for this analysis were adapted from the Framingham Heart Study for CV diseases. Results: At data cutoff, 2307 patients were available for this analysis. Mean (SD) age was 66.3 (12.2) years, 54.4% were male, 89.9% were white, 62.7% had at least some college education, and 51.1% were retired. Approximately 6% of patients had a family history of PV, primarily in parents (35.1%) and siblings (33.8%). A history of second malignancies was reported for 344 patients (14.9%). The majority of patients (84.6%) were diagnosed with PV based on an abnormal blood test alone or in combination with a bone marrow test. Among patients who were diagnosed with a mutational test (n=1078), 95.2% were diagnosed via an abnormal JAK2V617F test result. Abnormal hemoglobin (57.3%), hematocrit (55.4%), or both (47.5%) were among the most common blood values assessed for PV diagnosis. At diagnosis, 58.5% of patients were classified with high-risk PV (age ≥60 years or history of a TE); this percentage increased to 77.3% at REVEAL enrollment. The average (SD) disease duration from diagnosis to enrollment was 5.8 (6.1) years. At enrollment, 91.5% of patients were under active management for PV (phlebotomy ± aspirin, 34.0%; hydroxyurea ± aspirin, 27.0%; and phlebotomy + hydroxyurea ± aspirin, 23.2%). Underlying CV risk factors that were either diagnosed or treated in 86.0% of enrolled patients included hypertension (66.5%), history of smoking (46.2%), current smoking at enrollment (10.9%), obesity (34.2%), hyperlipidemia (27.4%), and diabetes (14.8%). At enrollment, 431 (18.7%) patients reported having ≥1 TE, including 181 patients who had a TE between PV diagnosis and enrollment. Venous and arterial TEs were reported in 11.1% and 8.6% of patients, respectively. Most commonly reported venous TEs were deep vein thrombosis (5.9%) and pulmonary embolism (2.5%); most common arterial TEs were cerebrovascular arterial thrombosis including transient ischemic attack (5.1%) and acute myocardial infarction (1.7%). Increased rates of TEs were observed among patients with hyperlipidemia (23.6%) and hypertension (21.0%; Table 1), compared with patients who did not have any risk factors (10.5%). Conclusion: A large proportion of patients in the REVEAL study had 1 or more underlying CV risks, including age, hypertension, smoking, obesity, hyperlipidemia, and diabetes, which may contribute to the risk of thrombosis. Longitudinal data from REVEAL will provide a better understanding of how these factors affect CV outcomes over time. Disclosures Stein: Incyte Corporation: Membership on an entity's Board of Directors or advisory committees, Research Funding. Naim:Incyte Corporation: Employment, Equity Ownership. Grunwald:Janssen: Research Funding; Forma Therapeutics: Research Funding; Medtronic: Equity Ownership; Alexion: Membership on an entity's Board of Directors or advisory committees; Ariad: Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding; Incyte Corporation: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding. Oh:Incyte Corporation: Membership on an entity's Board of Directors or advisory committees, Research Funding; Gilead: Membership on an entity's Board of Directors or advisory committees, Research Funding; Janssen: Research Funding; CTI: Research Funding. Paranagama:Incyte Corporation: Employment, Equity Ownership. Cordaro:Incyte Corporation: Employment, Equity Ownership. Sun:Incyte Corporation: Employment, Equity Ownership. Parasuraman:Incyte Corporation: Employment, Equity Ownership. Boccia:Celgene: Consultancy, Honoraria, Speakers Bureau; Amgen/Onyx: Consultancy, Honoraria, Speakers Bureau; Gilead: Speakers Bureau; Genentech: Consultancy, Honoraria, Speakers Bureau; Eisai: Consultancy, Honoraria, Speakers Bureau. Mesa:Ariad: Consultancy; CTI: Research Funding; Gilead: Research Funding; Galena: Consultancy; Novartis: Consultancy; Promedior: Research Funding; Celgene: Research Funding; Incyte Corporation: Research Funding.

Abstract Even though mutations in epigenetic regulators frequently occur in myeloproliferative neoplasms, their effects on the epigenome have not been well studied. Furthermore, even though primary myelofibrosis (PMF) has a markedly worse prognosis than essential thrombocytosis or polycythemia vera, the molecular distinctions between these subgroups are not well elucidated. We conducted the HELP (HpaII tiny fragment enriched by LM-PCR) assay to study genome-wide methylation in polycythemia vera, essential thrombocytosis, and PMF samples compared with healthy controls. We determined that polycythemia vera and essential thrombocytosis are characterized by aberrant promoter hypermethylation, whereas PMF is an epigenetically distinct subgroup characterized by both aberrant hyper- and hypomethylation. Aberrant hypomethylation in PMF was seen to occur in non-CpG island loci, showing further qualitative differences between the disease subgroups. The differentially methylated genes in polycythemia vera and essential thrombocytosis were involved predominantly in cell signaling pathways and were enriched for binding sites of GATA1 and other transcription factors. In contrast, aberrantly methylated genes in PMF were involved in inflammatory pathways and were enriched for NF1, LEF1, and other transcription factors. Within the PMF subgroup, cases with ASXL1 disruptions formed an epigenetically distinct subgroup with relatively increased methylation. Cases of myeloproliferative neoplasms (MPN) with TET2 mutations showed decreased levels of hydroxymethylation and distinct set of hypermethylated genes. In contrast, the JAK2V617F mutation did not drive epigenetic clustering within MPNs. Finally, the significance of aberrant methylation was shown by sensitivity of MPN-derived cell lines to decitabine. These results show epigenetic differences between PMF and polycythemia vera/essential thrombocytosis and reveal methylomic signatures of ASXL1 and TET2 mutations. Cancer Res; 73(3); 1076–85. ©2012 AACR.