Tefferi, Ayalew

Abstract BACKGROUND: Splenectomy is performed for palliating symptoms in patients with primary myelofibrosis (PMF; as well as post essential thrombocythemia myelofibrosis/ post polycythemia myelofibrosis (Mesa et. al. Cancer2006;107(2):361–70). Pre-operative computed tomograms (CTs) of the spleen are frequently performed for estimating spleen size and anatomy, but sometimes raise concern over radiographic splenic masses. We performed a retrospective analysis of splenectomized PMF patients to determine the predictive value of CT in these patients. METHODS: Splenectomized patients, with confirmed PMF (and POST ET/PV MF), with an available pre-operative CT scan were included. Pre-operative CT scans were reviewed in a blinded fashion by a radiologist (B.B.), and histopathology was concurrently reviewed in a blinded fashion by a hematopathologist (C.Y.L.), with attention for radiographic evidence of splenic masses, splenic infarction and hemorrhage, evidence of extra-splenic masses. Additionally splenic volume was calculated by CT through an algorithm using maximal splenic length, width, and slice size (prorated ellipsoid where volume = (30+.58(XxYxZ)). Results from each review were compared and analyzed against clinical outcomes and documented splenic mass. RESULTS: Thirty-eight patients had concurrent CT and splenic pathology available underwent splenectomy for myelofibrosis (20 PMF; median age at diagnosis 64.6 years (range (46–79), 52% males). Splenic volume estimations by CT correlated well with splenic mass measurements. Splenic volume estimation by CT was 2365 CC (range 736–11355 CC), compared with a measured median splenic mass of 2000 mg (range 730–11750 mg) (bivariate analysis p<0.001). Splenic lesions were identified in a total of 33 patients (87%), 10 only by histopathology (30%), 3 only by CT (9%), and 20 by both (61%). Histopathologically 15 patients had localized hemorrhage, 13 had splenic infractions, and 8 had nodular foci of extramedullary hematopoiesis. Only 2 patients had evidence of an alternative malignancy (concurrent chronic lymphocytic leukemia (CLL, n =1) or systemic mast cell disease (n = 1). CT was very sensitive for detecting focal low density lesions (LDLs), which are roughly spherical regions of decreased contrast enhancement in the spleen, and detected the majority of macroscopic lesions. The presence of splenic infarctions histologically was associated with decreased post splenectomy survival (p=0.02; median 7.2 months vs. 15.0 months), however the sensitive (but not specific) presence of LDL lesions by CT did not have prognostic significance. The CT appearance of splenic hemorrhage was nonspecific, and since these areas of parenchymal hemorrhage were likely subacute/chronic, they could not be reliably distinguished from other LDLs. CONCLUSIONS: Pre-operative CT in patients whom undergo palliative splenectomy for myelofibrosis is a sensitive tool for determining the presence of small splenic lesions (hemorrhage, infarction and EMH). Additionally splenic volume estimation by CT, correlates well with measured splenic mass and may assist in operative planning. Splenic lesions in PMF (post ET/PV MF) by CT rarely represented an alternative malignant process, and should likely not be the sole indication for splenectomy. Splenic infarctions were associated with decreased survival after splenectomy, however CT is not always able to distinguish these lesions from EMH and hemorrhage.

Background: Myelodysplastic syndrome (MDS)/myeloproliferative neoplasm (MPN) with ring sideroblasts and thrombocytosis (MDS/MPN-RS-T) was formally included as a unique WHO-defined entity in the 2016, iteration of the classification of hematological neoplasms (under MDS/MPN overlap syndromes). Patients with MDS/MPN-RS-T have features of MDS-RS-single lineage dysplasia (SLD), persistent thrombocytosis (platelet count ≥450x109/L), with proliferation of large atypical megakaryocytes. Anemia, including red blood cell transfusion dependency (RBC-TD) is a major problem. While extensive studies have documented erythroid response rates (ERR) to erythropoiesis stimulating agents (ESA) in MDS and MDS-RS-SLD, the data in MDS/MPN-RS-T is less clear. We carried out this study to asses ERR in ESA treated patients with MDS/MPN-RS-T. Methods: After approval by the IRB, patients with 2016, WHO-defined MDS/MPN-RS-T were included in the study. The bone marrow (BM) morphology, cytogenetics and BM RS% were retrospectively reviewed. All study patients underwent next generation sequencing for 29 myeloid-relevant genes, obtained on BM mononuclear cells, at diagnosis, or at first referral, by previously described methods (Patnaik et al., BCJ 2016). Treatment details, including the type of ESA used, dose administered, side effects and reason for discontinuation were retrospectively abstracted. Erythroid responses were assessed using the 2006, international working group (IWG) MDS and the 2015, IWG MDS/MPN response criteria. Results: Forty seven patients with MDS/MPN-RS-T were identified, of which 37 (79%) patients received ESA treatment at any time point during their disease course; median age 73 years (range, 52-93), 46% male.The hemoglobin at diagnosis (HB) and baseline erythropoietin (EPO) levels were 9.1 gm/dL (range, 6.6-12.1) and 39 IU/L (range, 8-500), respectively. Ten (29%) patients were RBC-TD at baseline. None of the 37, ESA-treated patients were on concomitant cyto-reductive therapy for thrombocytosis. Twenty (54%) patients were treated with erythropoietin-a, 13 (35%) with darbepoetin, and 4 (11%) with both sequentially. Median doses were 40,000 IU weekly (range, 20,000-80,000) for erythropoietin-a and 200 mcg every 2 weeks (range, 100-500), for darbepoetin. Median treatment duration was 14 months (range, 1-173) and causes for discontinuation included; ESA failure/loss of response in 20 (54%), sustained HB rise in 1 (3%) and other causes, including adverse events in 4 (11%) patients. With regards to safety, the following events were considered potentially ESA related; treatment emergent hypertension in 2 (5%), venous thromboembolism, ischemic stroke and splenic infarction in 1 (3%) patient each, respectively. All patients with thrombotic/ischemic events had been on low dose aspirin prophylaxis. Erythroid responses by the IWG MDS and MDS/MPN response criteria were assessable in 35 (95%) patients: with 16 (46%) meeting criteria for an erythroid response, by both assessment systems. The median duration of ER was 20 months (range, 2-172). In comparison to ESA non-responders, those that responded were more likely to have baseline EPO levels ≤44 IU/L (ROC analysis; median, 28 vs 112 IU/L; p=0.0080), while there was a trend for inferior responses associated with RBC-TD (p=0.0857). Age, gender, baseline white blood cell and platelet counts, type of ESA used, BM RS %, JAK2V617F, SF3B1, DNMT3A, TET2 and ASXL1 mutational status did not impact ESA response (Table 1). On a univariate survival analysis (OS), survival was not significantly different between ESA responders vs non-responder (median OS, 76 vs 45 months; p=0.2929). After ESA failure, 7 (19%) patients received hypomethylating agents (HMA), while 5 (14%) received lenalidomide. There were no erythroid responses to HMA, while 1 of 3 (33%) assessable lenalidomide treated patients achieved a morphological complete response. Conclusion: Erythropoiesis stimulating agents are effective first line therapies for the management of anemia in patients with MDS/MPN-RS-T, with 46% achieving an erythroid response (median duration of response of 20 months). Low baseline endogenous EPO level (<44 IU/L) was the best predictor of response, with a trend towards an inferior response in the presence of RBC-TD. In the context of clonal thrombocytosis, potentially related thrombotic/ischemic events were documented in 8% of patients. Disclosures Al-Kali: Astex Pharmaceuticals, Inc.: Research Funding. Patnaik:Stem Line Pharmaceuticals.: Membership on an entity's Board of Directors or advisory committees.

Introduction: Myelodysplastic syndrome/myeloproliferative neoplasm (MDS/MPN) overlap syndromes consist of 5 distinct WHO-defined entities; namely chronic myelomonocytic leukemia (CMML), atypical chronic myeloid leukemia, BCR/ABL1- (aCML), juvenile myelomonocytic leukemia (JMML), MDS/MPN with ring sideroblasts and thrombocytosis (MDS/MPN-RS-T), and MDS/MPN, unclassifiable (MDS/MPN-U) (Arber et al., Blood 2016). With the notable exception of JMML, a bona fide RASopathy, the other entities are characterized by clinical heterogeneity and molecular diversity. Loss of function TET2 mutations (TET2MT) are common in myeloid neoplasms, especially CMML (60%), and are known leukemogenic drivers. We carried out this study to assess the TET2 mutational landscape and phenotypic correlates in patients with MDS/MPN overlap syndromes. Methods: After approval by the institutional review board, adult patients with WHO defined MDS/MPN overlap syndromes were included; with the exception of JMML. The BM morphology, cytogenetics and 2016, WHO-diagnoses were retrospectively reviewed and all patients underwent targeted next generation sequencing for 29 myeloid-relevant genes, obtained on BM mononuclear cells, at diagnosis, or at first referral, by previously described methods (Patnaik et al., BCJ 2016). Results: Five hundred and four patients were included in the study; including 387 (77%) with CMML, 48 (10%) with MDS/MPN-RS-T, 17 (3%) with aCML and 52 (10%) with MDS/MPN-U. The median age at diagnosis was 71 (range, 18-99) years, and 333 (66%) were male. TET2MT were seen in 212 (42%) patients, with the frequency of other mutations being: ASXL1 45%, SRSF2 40%, NRAS 15%, SF3B1 13%, CBL, RUNX1 and SETBP1 12% each, and JAK2 V617F 11% (Figure B). Among the MDS/MPN overlap syndromes, TET2 was more frequently mutated in CMML (49%) and aCML (47%) compared to MDS/MPN-RS-T (10%) and MDS/MPN-U (15%). The prevalence of patients with TET2MT increased with age, a finding consistent across all MDS/MPN subtypes (Figure C). Overall, 341 TET2MT were identified in 212 patients (mean 1.6 variants/patient, range 0-5): 120 (24%) had >1 TET2MT, while 113 (22%), 5 (1%) and 2 (0.4%) had 2, 3 and 5 mutations, respectively. CMML and aCML patients were more likely to have an age-independent increase in multiple TET2MT (28% and 24%), in comparison to MDS/MPN-RS-T (4%) and MDS/MPN-U (8%). TET2 MT spanned the entire coding sequence and were mostly truncating (78%, Figure A): 59 (17%) were missense, 14 (4%) involved the splice-donor/acceptor sites, 2 (0.5%) were in-frame deletions, 129 (38%) were nonsense, and 137 (40%) were frameshift mutations. Overall, the distribution of TET2MT was superimposable across CMML, aCML, and MDS/MPN-U; the only exception being the absence of splice site mutations in the latter two. One hundred and eighty-seven (55%) TET2MT were secondary to pathogenic single nucleotide variants (SNV), while the remainders were secondary to deletions (25%) and insertions (15%). Transitions comprised the most frequent type of SNV (65%), with the C:G>T:A being the most common (56%). Patients with MDS/MPN overlap syndrome and TET2MT were more likely to have additional gene mutations compared to wild type patients (mean mutation number 3.1 vs 2.1, p<0.0001), with common co-mutations being SRSF2 (51%), ASXL1 (42%), and CBL (17%). The median overall survival (OS) of the entire cohort was 29 (range, 0-170) months; 29 months for CMML, 63 months for MDS/MPN-RS-T, 14 months for aCML, and 25 months for MDS/MPN-U. On univariate analysis, OS was superior in CMML patients with TET2MT (35 months) compared to wild type cases (21 months, p<0.0001, Figure D), and in CMML patients with >1 TET2MT (41 months) in comparison to wild type (21 months, p<0.0001) and single TET2MT (29 months, p=0.0476) cases (Figure E). These observations were not seen in patients with aCML, MDS/MPN-RS-T, and MDS/MPN-U. Conclusion: Our study demonstrates that TET2MT are among the most frequent mutations in patients with MDS/MPN overlap syndromes (42%), especially CMML (49%), with an age-dependent increase in the frequency and number of TET2MT. Mutations in TET2 were found to span the entire coding sequence, with truncating mutations being more common (78%). Importantly, in CMML, TET2MT, including number of TET2MT, were associated with favorable survival outcomes. Figure Disclosures Al-Kali: Astex Pharmaceuticals, Inc.: Research Funding. Patnaik:Stem Line Pharmaceuticals.: Membership on an entity's Board of Directors or advisory committees.

Introduction: Chronic myelomonocytic leukemia (CMML), an aggressive myeloid malignancy, can be categorized into two subtypes, proliferative CMML (pCMML) and dysplastic (dCMML), based on a white blood cell (WBC) count of ≥ 13 x 109/L for the former (Arber et al. Blood 2016). While this WBC cut off is somewhat arbitrary, patients with pCMML have unique phenotypic features and a shorter survival. We carried out this study to assess the genomic, transcriptomic and epigenetic landscapes of these two CMML subtypes. Methods: Peripheral blood (PB) and bone marrow (BM) mononuclear cells (MNC) were obtained from WHO-defined CMML patients. Next generation sequencing (NGS) using a 36-gene panel was performed on 350 patients with Illumina HiSeq4000 platform with median read depth of 400X. RNA sequencing (RNA-seq) was performed on 25 patients by bulk whole transcriptome sequencing using Illumina TruSeq. DNA immunoprecipitation sequencing (DIP-seq) was performed on 18 patients using 5-methylcystocine (5mC), 5-hydroxymethylcytosine (5hmC) and bridging monoclonal antibodies with subsequent paired-end sequencing using HiSeq4000. Chromatin immunoprecipitation sequencing (ChIP-seq) was performed on 30 patients with Illumina HiSeq2500 to a depth of 25 million for histone 3 lysine 4 monomethylation (H3K4me1) and histone 3 lysine 4 trimethylation (H3K4me3) and 50 million reads for histone 3 lysine 27 trimethylation (H3K27me3) and Input per sample. Results: Five hundred and seventy-three patients with WHO defined CMML were included; median age 71 years (range 18-95 years), 67% males. 282 patients had pCMML (49%), while 291 (51%) had dCMML. As pre-defined, patients with pCMML were more likely to have higher absolute monocyte counts (p<0.0001), circulating immature myeloid cells (p<0.0001), PB blasts (p<0.0001), and higher lactate dehydrogenase levels (p=0.03). At last follow up 234 (41%) deaths and 70 (20%) leukemic transformations were documented. The median OS for pCMML vs dCMML in this cohort was 19 months vs 30 months (p<0.0001, Figure 1A) and validated in an independent Austrian cohort (p=0.02). Genomic profiling: NGS performed on 350 patients (BM MNC) revealed a higher frequency of NRAS (35 vs 17%, p=0.004), cumulative RAS pathway (NRAS, KRAS, CBL and PTPN11) (73 vs 47%, p=0.001), ASXL1 (p=0.003) and JAK2V617F (p=0.04) mutations in pCMML relative to dCMML (Figure 1B); while dCMML had a higher frequency of SF3B1 mutations (p=0.02). There were no differences in distribution of TET2 and SRSF2Transcriptomic analysis: RNA-seq was performed on PB MNC from RAS pathway mutant pCMML patients (n=12) and RAS pathway wildtype dCMML patients (n=13). Unsupervised clustering analysis resulted in appropriate segregation revealing distinct expression profiles between disease subtypes (Figure 1C). Compared to dCMML, 3729 genes were significantly differentially upregulated and 2658 genes were differentially downregulated in pCMML. Among genes most highly upregulated were mitotic checkpoint kinases including AURBK, PLK1, PLK2, PLK4 andEpigenetic profiling: ChIP-seq of PB and BM MNC from pCMML (n=18) and dCMML (n=12) patients and healthy, age-matched controls (n=10) revealed a global increase in H3K4me1, without significant differences in H3K4me3 or H3K27me3 occupancies (regardless of stratification by ASXL1 mutational status; 40% ASXL1mt in pCMML, 30% dCMML) in pCMML vs dCMML (Figure 1D). H3K4me1 occupancy was also increased in a sequence-specific manner at the transcription start sites of the aforementioned mitotic kinases (PLK1 and WEE1). DIP-seq was performed on PB MNC to assess global differences in 5-mC and 5-hmC levels, between pCMML (n=9) and dCMML (n=9), with no differences seen between the two subtypes (regardless of TET2 mutational status, 40% TET2mt in each subtype) (Figure 1E). Conclusions: Despite the somewhat arbitrary WBC distinction between pCMML and dCMML, clear phenotypic, genetic, transcriptomic, epigenetic and survival differences exist between the two subtypes, providing strong biological rationale for this distinction. pCMML patients have a higher frequency of oncogenic RAS pathway mutations, a unique transcriptomic profile enriched in mitotic check point kinases and a unique chromatin configuration with global and sequence specific enrichment in H3K4me1, with no significant global differences in 5mC, 5hmC, or H3K4me3 and H3K27me3 occupancy. Figure 1 Disclosures Geissler: AOP: Honoraria; Pfizer: Honoraria; AstraZeneca: Honoraria; Novartis: Honoraria; Celgene: Honoraria; Roche: Honoraria; Abbvie: Honoraria; Ratiopharm: Honoraria; Amgen: Honoraria. Al-Kali:Astex Pharmaceuticals, Inc.: Research Funding. Patnaik:Stem Line Pharmaceuticals.: Membership on an entity's Board of Directors or advisory committees.

Objective : Central nervous system (CNS) involvement is a known phenomenon of acute lymphoblastic leukemia. However little is known about the incidence of CNS involvement in acute myeloid leukemia (AML) as routine lumbar punctures are not done if patients are asymptomatic at diagnosis. Our practice has been to obtain lumbar puncture (LP) at or after complete remission (CR) for patients that present with any one of the following high risk features;hyperleukocytosis with WBC >10,000 or blastic crisis, extramedullary disease,monocytic differentiation. Methods: We reviewed our leukemia database at Mayo Clinic to determine the incidence of CNS involvement in patients with high risk features after due IRB approval. We also compared baseline demographic characteristics, blood counts, cytogenetic risk groups, presence of extramedullary disease, relapse rate ,transplant rate and overall survival (OS) among patients with and without CNS involvement. OS estimates were calculated by Kaplan-Meier curves and log-rank testing using JMP v.13. Results: 298 patients with AML were identified with presence of at least 1 risk factor for CNS involvement. The results of LP were only available for 126 (42%) patients. The overall incidence of CNS involvement was 12% (15/126, figure 1). Among the patients with positive disease 3 had CSF examination at diagnosis prior to CR due to the presence of CNS myeloid sarcoma (n=2) and ambiguous lineage myeloid leukemia and were excluded from further evaluation. The incidence of CSF involvement at CR1 was 9.7% (12/123). All patient with CNS positive disease received intrathecal chemotherapy with alternating cytarabine and methotrexate until they cleared their blast and additional 2-4 treatments as per treating physician's discretion .The CNS positive and negative patients were compared and did not show any statistical difference in age, gender, presence of extramedullary disease ,cytogenetic risk group, relapse rate and transplant rate as shown in table1. Molecular data was not available in most of the patients and hence was not included for analysis. The median OS was significantly lower among patients with CNS involvement compared to absence of CNS involvement but was not statistically significant likely due to small sample size (3.9 vs12.8 years (p=0.2), figure 2). The overall CNS relapse rate in patients who did not have a LP done at CR was 5.8%(10/172). Conclusions : CNS evaluation should be done in AML patients with high risk features before declaring CR. CNS disease portends overall a poor prognosis with impaired overall survival and high relapse rate. However the sample size was small and true incidence of CNS disease will need routine diagnostic LP in all high risk patients. Disclosures Patnaik: Stem Line Pharmaceuticals.: Membership on an entity's Board of Directors or advisory committees. Al-Kali:Astex Pharmaceuticals, Inc.: Research Funding.

Introduction: Isocitrate dehydrogenase 1 and 2 (IDH1/2) are metabolic enzymes in the citric acid cycle, producing alpha-ketoglutarate (αKG). Mutations in specific regions of these genes have been characterized in gliomas, AML and chronic myeloid neoplasms such as MDS and MPN. These mutations produce an oncometabolite 2-hydroxyglutarate (2HG) that disrupts the epigenetic phenotype of myeloid cells promoting oncogenesis. While this process has been characterized for arginine hot spot (R132/R140) mutations, there exist variants of unclear significance (IDHVUS), with potential pathogenic predictions using in silico assessment models. With the availability of mutant IDH inhibitors in practice, it has become important to characterize these VUS. We carried out this study to functionally interrogate potentially pathogenic IDHVUS in MDS and MPN. Methods: After institutional approval, bone marrow (BM) DNA of patients with MDS and MPN, including overlap syndromes were subjected to next generation sequencing (29-gene, including the entire coding region of IDH1/2) by previously described methods (Patnaik et al., BCJ 2016). A sub-set of patients with available plasma [IDH wild type (IDHWT), bona fide IDH mutations (IDHMT) and IDHVUS with predicted pathogenicity (≤0.01% minor allele frequency, and predicted damaging in silico scores)] were submitted for mass spectrometry (LC-MS) based detection of IDH-related metabolites; D and L isomers of 2HG, αKG, and total 2HG. The impact of variations on protein secondary structure was assessed by the GorIV prediction algorithm. Potential alterations within the tertiary structure of the proteins was evaluated by in silico mutagenesis. Results: One hundred and seventeen patients were included in the study; median age 68 years (range, 39-94), 68% male. Of these, 40 (34%) had MDS, 42 (36%) MPN, and 35 (30%) MDS/MPN overlap syndromes. Eighty six (74%) had IDHMT, including n=28 (24%) with IDH1MT (R132C/G/H/S) (11 MDS, 8 MPN, 9 MDS/MPN) and n=59 (50%) with IDH2MT (R140G/Q/W) (22 MDS, 20 MPN, 17 MDS/MPN). IDHVUS were detected in 32 (27%) patients, including 1 IDH1MT patient, and involved IDH1 in 14 (12%) cases and IDH2 in 18 (15%) cases. All IDHVUS were missense, except for a 2 amino acid insertion in IDH2, with the most frequent VUS being IDH1Y183C (7%) and IDH2T352P (7%). While there were no phenotypic differences between IDH1/2MUT and IDH1/2VUS, in comparison to patients with IDH1/2MT, those with IDH1VUS were more likely to have higher WBC counts (median 8.1 vs 3.6 x109/L, p=0.0322), higher neutrophil counts (median 3.3 vs 1.3 x109/L, p=0.0203), and lower BM blast % (median 1 vs 3%, p=0.02061). Compared to IDH1/2MT patients, those with sole IDH1/2VUS more frequently had mutations in SF3B1 (p=0.0167), while they were associated with a lower frequency of SRSF2 mutations (p=0.0002). Interestingly, TET2 mutations were detected in only 5% of patients with IDHMT and 16% of IDHVUS patients, although without reaching statistical significance. LC-MS analysis on 75 plasma samples (n=43 IDHMT, n=9 IDHVUS, n=20 IDHWT, 3 normal) revealed a significant increase in total 2HG levels in IDHMT vs IDHVUS (p=0.0089), and IDHWT (p=0.0020) cases (Figure 2A-D). While there were no significant differences in αKG levels between the groups, the D-2HG enantiomer levels were higher in IDHMT samples vs IDHVUS (p=0.0199) and IDHWT (p=0.0055). On examination of the crystal structures of the IDH genes using PDB, while we observed a direct interaction between R132/R140/R172 and the catalytic core, the IDHVUS were located away from this active center and were hence thought to be allosteric in nature (Figure 2F). These VUS however, did demonstrate predictive steric clashes in the tertiary structure context of IDH1 using the PyMol artificial mutagenesis tool. The absence of wild-type crystal structure for IDH2precluded a similar analysis for IDH2. Median overall survival (OS) of the entire cohort (Figure 1) was 36 (range, 0-190) months, with OS being inferior in IDH1/2MT patients (34 months) vs those with IDH1/2VUS (50 months, p=0.0255). Conclusion: Our study demonstrates that in spite of predictive pathogenicity of IDH1/2VUS using in silico predictive tools and crystal studies, the lack of involvement of the arginine hot spot precludes IDH neomorphic activity, as shown by normal 2HG levels. In addition, in comparison to IDHMT patients, those with IDHVUS had fewer BM blasts and a better survival. Disclosures Patnaik: Stem Line Pharmaceuticals.: Membership on an entity's Board of Directors or advisory committees.

Background MDS is a heterogeneous disease and the revised International Prognostic Scoring System (IPSS-R) is utilized in prognostication. The percentage (%) of blasts in the bone marrow is determined in the aspirate morphologically. Though the former is the standard of care the blast percentage is also reported by flow cytometry and biopsy which can many times be inconsistent. We previously presented the utilization of biopsy based blast percentage which showed meaningful prognostic groups compared to aspirate. In this study we compare the blasts as reported by the aspirate and flow cytometry in MDS-EB in calculating IPSS-R. Methods The MDS database was reviewed for cases of MDS-EB after due IRB approval at the Mayo clinic. We calculated IPSS-R scores based on the aspirate blast % (IPSS-RAsp) and flow blast% (IPSS-Rfl). The aspirate blast percentage was reported morphologically. Suboptimal aspirates were excluded from the study. The flow blast percentage was determined by immunophenotyping. The overall survival (OS) was determined by IPSS-RAsp and IPSS-RFl. OS estimates were calculated by Kaplan-Meier curves and log-rank testing using JMP v.13. Uno's concordance statistic was used to compare the 2 risk scoring systems. Results Of 1322 patients, 431 (33%) cases were identified with MDS-EB out of which 120 (29%) cases had blasts reported in the aspirate and flow. Based on aspirate MDS EB1: 54% (n=65), MDS EB2 46% (n=55). The hematological, cytogenetic and R-IPSS categories were compared between MDS-EB1 and MDS- EB 2. The blast percentage and hemoglobin levels was significantly different between MDS-EB1 and EB2 as seen in table 1, however the IPSS-R risk groups were not significantly different. The flow cytometry was concordant with aspirate in 66/120 (55%) cases. Out of the dis-concordant cases only 20% (11/54) was upstaged by flow cytometry with most of the patients being down staged as expected by the techniques used in processing the blood and hence not reliable when reported low (Figure 1). The OS outcomes based on the IPSS- R asp, IPSS-Rfl areshown in figure 2A,2B .The p value with aspirate based R-IPSS was more significant than flow cytometry based R-IPSS (p= 0.0007 vs 0.0174). We compared the two models for observed OS differences using the Uno model which was not statistically significant. (p= 0.6) Conclusions Both models did not show a difference which is likely due to the very small sample size. However flow cytometry did down stage more patients when disconcordant and may have less value in that setting. It would be ideal to compare all 3 models aspirate, biopsy and flow cytometry however we did not have enough number of patients to do the comparison. Disclosures Patnaik: Stem Line Pharmaceuticals.: Membership on an entity's Board of Directors or advisory committees. Al-Kali:Astex Pharmaceuticals, Inc.: Research Funding.

Introduction: Germline RUNX1 mutations/deletions result in a Familial Platelet Disorder with propensity to Myeloid Malignancy (FPDMM); an autosomal dominant condition characterized by thrombocytopenia, qualitative platelet defects, and myeloid clonal evolution (MDS and AML). The advent of next generation sequencing (NGS) has allowed detection of clonal cytopenias of unclear significance (CCUS) prior to bone marrow (BM) morphological changes. Somatic RUNX1 mutations (RUNX1MT) can be seen in myeloid malignances, including MDS/MPN overlap syndromes, with an indeterminate prognostic impact. We carried out this study to define the landscape of clonal evolution in FPDMM patients and to compare germline RUNX1MT with somatic mutations. Methods After IRB approval, families with RUNX1MT FPDMM (detected by germline sequencing and deletion/duplication assays) and patients with 2016, WHO-defined MDS/MPN overlap syndromes (except JMML) were included in the study. Index patients with FPDMM underwent BM biopsies and in select cases NGS at diagnosis and then as indicated for follow up, or at disease progression. All MDS/MPN overlap syndrome patients underwent a 29-gene panel NGS assay on BM samples obtained at diagnosis by previously described methods (Patnaik et al., BCJ 2016). Results Eight index patients with FPDMM were included; median age at diagnosis 58 (range, 0-71) years, 3 (38%) males (Figure A). Four (50%) had intragenic deletions involving multiple exons in 3 cases and 2 nucleotides in 1 case, 1 (13%) each had missense, frameshift and splice site mutations, while 1 (13%) patient had a whole gene deletion (Figure C). Notably, in 4 (100%) of 4 patients with large deletions, NGS testing was negative for RUNX1 abnormalities. Four (50%) index patients presented with a myeloid neoplasm; JAK2, SRSF2, ASXL1 mutant CMML, MDS/MPN-U with trisomy 8, AML with del7q and normal karyotype AML (negative NGS). Three (38%) index patients had CCUS at diagnosis with TET2 (67%) being the most common acquired mutation, and others including SRSF2, EZH2, and SF3B1. Cumulatively, myeloid clonal evolution was seen in 7 (88%) patients. Three (38%) index patients (AML-2, CMML-1) underwent allogeneic HCT, and at last follow up were in remission. Median age at myeloid clonal evolution was 61 (range, 4-71) years; median overall survival (OS) from date of myeloid clonal evolution was 12 (range, 7-53) months. For the comparative analysis, 499 patients with MDS/MPN overlap syndromes were included; 382 (77%) with CMML, 48 (10%) with MDS/MPN-RS-T (MDS/MPN with ring sideroblasts and thrombocytosis), 17 (3%) with aCML (atypical CML), and 52 (10%) with MDS-MPN-U (unclassifiable). Median age at diagnosis was 71 (range, 18-99) years, and 329 (66%) were male. RUNX1MT were detected in 57 (11%) patients; 43 (11%) with CMML, 5 (30%) with aCML, and 9 (17%) with MDS-MPN-U. No RUNX1MT were found in MDS-MPN-RS-T. Overall, 62 RUNX1MT were identified in 57 patients, with 52 (39 CMML, 4 aCML, 9 MDS-MPN-U) and 5 (4 CMML, 1 aCML) patients harboring 1 and 2 RUNX1MT, respectively. RUNX1MT spanned the entire coding sequence without specific hotspots (Figure B): 27 (44%) were missense, 2 (3%) involved the splicing sites, 5 (8%) were nonsense and 28 (45%) were frameshift. There was only one RUNX1MT (R273K), seen in both germline and somatic patients (Figures B and C). Compared to wild type patients, RUNX1MT patients with MDS/MPN overlap syndromes had significantly lower platelet counts (median 72 vs 117 x109/L, p<0.0001), without any other phenotypic differences. In the somatic RUNX1MT cohort, co-occurring mutations included ASXL1 (54%), SRSF2 (54%), TET2 (49%), and NRAS (28%), with NRAS (28% vs 13%, p=0.0028), and SRSF2 (54% vs 39%, p=0.0218) mutations clustering with RUNX1MT. Median OS of the entire cohort was 29 (range, 0-170) months. On univariate analysis, while there was a trend for an inferior OS in RUNX1MTvs wild type patients (20 vs 29 months, p=0.0523), the leukemia-free survival was clearly inferior (p=0.0175, Figure D). Conclusion RUNX1 MT FPDMM is a germline predisposition syndrome with a high rate of myeloid clonal evolution (88%), including clonal cytopenias (38%). Germline RUNX1MT have minimal overlap with somatic mutations seen in myeloid malignancies, with NGS testing often missing RUNX1 deletions. Somatic RUNX1MT in myeloid malignancies are significantly associated with thrombocytopenia and an inferior leukemia-free survival. Figure Disclosures Patnaik: Stem Line Pharmaceuticals.: Membership on an entity's Board of Directors or advisory committees.

AbstractMyeloproliferative Neoplasms (MPN) course can be complicated by thrombosis involving unusual sites as the splanchnic veins (SVT). Their management is challenging, given their composite vascular risk. We performed a retrospective, cohort study in the framework of the International Working Group for MPN Research and Treatment (IWG‐MRT), and AIRC‐Gruppo Italiano Malattie Mieloproliferative (AGIMM). A total of 518 MPN‐SVT cases were collected and compared with 1628 unselected, control MPN population, matched for disease subtype. Those with MPN‐SVT were younger (median 44 years) and enriched in females compared to controls; PV (37.1%) and ET (34.4%) were the most frequent diagnoses. JAK2V617F mutation was highly prevalent (90.2%), and 38.6% of cases had an additional hypercoagulable disorder. SVT recurrence rate was 1.6 per 100 patient‐years. Vitamin K‐antagonists (VKA) halved the incidence of recurrence (OR 0.48), unlike cytoreduction (OR 0.96), and were not associated with overall or gastrointestinal bleeding in multivariable analysis. Esophageal varices were the only independent predictor for major bleeding (OR 17.4). Among MPN‐SVT, risk of subsequent vascular events was skewed towards venous thromboses compared to controls. However, MPN‐SVT clinical course was overall benign: SVT were enriched in PMF with lower IPSS, resulting in significantly longer survival than controls; survival was not affected in PV and slightly reduced in ET. MPN‐U with SVT (n = 55) showed a particularly indolent phenotype, with no signs of disease evolution. In the to‐date largest, contemporary cohort of MPN‐SVT, VKA were confirmed effective in preventing recurrence, unlike cytoreduction, and safe; the major risk factor for bleeding was esophageal varices that therefore represent a major therapeutic target.