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Background: Myelodysplastic syndrome (MDS) is a clonal hematopoietic neoplasm that results in bone marrow failure and frequently leads to acute myeloid leukemia (AML). Hypomethylating agents (HMA) are the only FDA-approved treatment for MDS and among few options for chemotherapy-ineligible patients with AML. There are limited options for patients in whom HMA therapy fails and who are not candidates for allogeneic stem cell transplant, so therapies that complement or restore sensitivity to HMAs are needed. Studies have shown increases in bone marrow microvessel density (MVD) and angiogenic markers in patients with MDS and AML. A decrease in MVD has been shown to correlate with response to hypomethylating agents in MDS. A receptor-ligand interaction, comprised of receptor EphB4 and membrane localized ligand EphrinB2, mediates angiogenesis in normal tissue and appears to be a target unique to many cancer types. We have previously shown EphB4 to be highly expressed and a driver of leukemic cell survival in a subset of AML patients. Our group has developed a human fusion protein, sEphB4-HSA, that blocks bidirectional signaling induced by EphB4-EphrinB2 interaction to inhibit tumor cell proliferation and angiogenesis. In phase I clinical trials of sEphB4-HSA in various tumor types, there were no myelosuppressive effects and minimal toxicity. Given its safety in phase I and potential to inhibit leukemic cell proliferation and angiogenesis, we proposed a pilot trial to evaluate the safety of sEphB4-HSA in combination with HMAs in MDS and AML patients who have failed treatment with HMAs. Methods: This pilot study was designed to enroll 6 patients with relapsed/refractory intermediate or high-risk MDS and 6 patients with AML refractory to or relapsed to HMA treatment and who are deemed unfit for chemotherapy. Treatment consisted of sEphB4-HSA 15 mg/kg IV every 2 weeks in combination with the FDA-approved HMA most recently or currently being used for treatment (decitabine 20mg/m2 IV/1hr on days 1 to 5 every 28 days or azacitidine 75mg/m2 SC or IV on days 1 to 7 every 28 days). Patients were treated for as long as they were receiving clinical benefit up to 12 months. The primary endpoint was toxicity and tolerability of sEphB4-HSA in combination with HMA. Toxicity was assessed and graded after each cycle according to the CTCAE version 4. Tolerability was defined as the ability to complete two cycles of treatment without the occurrence of dose-limiting toxicity. A secondary efficacy endpoint was to assess best overall response, based on the IWG Working Group Criteria for MDS and AML, during the first two cycles of treatment. Enrollment was stopped after 7 patients due to expiration of funding. Results: Three patients with intermediate-risk MDS were treated for a median duration of 6 cycles (2-12) and 4 patients with AML were treated for 2 cycles. Median age was 75.5 years (67.9-84.8) and 57.1% were male. HMA included azacitadine in 6 patients and decitabine in one patient. There were no dose-limiting toxicities. There were 8 grade 3/4 events attributed to HMA, which included: neutropenia (2), thrombocytopenia (3), and leukopenia (3). There were 3 grade 3/4 events attributed to sEphB4, which included: febrile neutropenia (1), leukopenia (1), and hypertension (1). Of the MDS patients, 2 had stable disease, 1 patient after 2 cycles and 1 patient after 4 cycles. One patient achieved a hematologic improvement-erythroid after 6 cycles. AML patients had no disease response. Reasons for treatment discontinuation were death (1), disease progression (2), patient's decision (1), physician's decision (2), and hospice (1). Notably, a comparison of bone marrow biopsies at baseline and after 8 weeks of treatment demonstrated a decrease in MVD (Figure 1). Discussion: This pilot study found sEphB4 in combination with HMAs to be tolerable with no significant toxicity beyond that expected with HMA therapy and associated with potential clinical benefit in MDS patients. Improvement in abnormal bone marrow MVD may indicate a potential for sEphB4-HSA plus HMA therapy to alter the malignant microenvironment in MDS/AML. Disclosures No relevant conflicts of interest to declare.

Background: Novel coronavirus infection (SARS CoV-2 or COVID-19) is associated with a high risk of thrombotic complications, including macro- and micro-thrombi in major organs, leading to increased morbidity and mortality. Anticoagulant use, mainly heparin, which has both anticoagulant and anti-inflammatory properties, has been suggested as potentially beneficial. However, the optimal dose of anticoagulant for patients with COVID-19 is unknown. Establishing the optimal thromboprophylaxis strategy and determining the role of biomarkers for patient risk stratification may help to improve outcomes in COVID-19. Methods: This single-center retrospective cohort study is part of an ongoing Quality Improvement project on the use of an anti-factor Xa-driven heparin protocol, which includes a low-dose intravenous (IV) unfractionated heparin (UFH) option, being conducted at our medical center. Data on the type, dose, and indication for anticoagulation as well as outcomes including thrombosis, bleeding and survival was collected for inpatients diagnosed with COVID-19 between mid-March and June 15, 2020. To address COVID-coagulopathy we developed a d-dimer-based anticoagulation protocol for patients with COVID-19 (Figure 1). We recorded anticoagulant use as either standard prophylactic, escalated prophylactic (low-dose intravenous unfractionated heparin titrated to achieve an anti-factor Xa level of 0.1-0.3 anti-Xa units or enoxaparin 0.5mg/kg subcutaneously every 12 hours) or standard therapeutic dose used during the hospitalization. The primary endpoints assessed were ISTH-defined major and clinically relevant non-major bleeding (CRNMB) events and survival. Secondary endpoints included incidence of breakthrough thrombosis and duration of hospitalization. Results: A total of 263 patients with COVID-19 were reviewed. Of these, 68.44% of patients received prophylactic, 12.55% escalated prophylactic and 19.01% therapeutic dosage. Of total, 129 (49%) were receiving ICU level of care. No major bleeding events were observed. The incidence of CRNMB was 4.56% in the whole cohort, which did not differ significantly between the escalated prophylactic and therapeutic groups (12% and 12.12%, respectively). Patients treated with standard prophylaxis had less CRNMB (1.11%), but this was not statistically significant in a multivariate analysis that included other confounding factors such as age, sex, ethnicity, BMI, comorbidity, HASBLED bleeding risk, and sepsis induced coagulopathy score (SICS). The mortality rate was 12.6% in the whole cohort (7.22%, 21.21% and 26% in prophylactic, escalated prophylactic, and therapeutic dosage, respectively). Factors significantly associated with increased mortality included age and ICU level of care (HR 1.10, 95%CI [1.05, 1.15] and HR 20.42, 95%CI [2.84, 146.72], respectively). The use of therapeutic dose heparin and high-flow nasal cannula demonstrate a survival benefit in multivariate analysis (HR 0.13, 95%CI [0.04,0.44] and HR 0.23, 95%CI [0.07, 0.72], respectively; Figure 2). Breakthrough thrombosis occurred in 7 (2.66%) patients; 1 (0.56%), 1 (3.03%) and 5 (10%)) in prophylactic, escalated prophylactic and therapeutic dosage, respectively but very few diagnostic tests were performed during this time period. Duration of hospitalization was significantly longer in the therapeutic dose group when compared to escalated prophylaxis and standard prophylactic groups. Conclusion: In this cohort of inpatients with COVID-19, there were no major bleeding events related to any dose of heparin or LMWH prophylaxis. By multivariate analysis, implementation of a d-dimer-titrated anticoagulation strategy was not associated with increased CRNMB. Therapeutic dose heparin based on a d-dimer-driven anticoagulation protocol was associated with a survival benefit in COVID19-infected patients. Limitations of this study include the retrospective observational nature and a lack of a uniform diagnostic protocol for patients with suspected VTE. Although no significant difference in bleeding events were observed in our study subgroups, randomized clinical trials are necessary to determine optimal thromboprophylaxis strategy in the COVID-19 population. Disclosures No relevant conflicts of interest to declare.

Background: Idiopathic erythrocytosis (IE) is an entity characterized by a persistently elevated hemoglobin, variable erythropoietin (EPO) level, absence of janus kinase 2 (JAK2) mutations suggestive of polycythemia vera (PV) and no identifiable secondary cause. Previous studies have compared IE to PV, showing a lower incidence of venous thrombosis and leukemic transformation in IE but similar incidence of arterial events. PV is known to be associated with constitutional symptoms and splenomegaly, while hereditary erythrocytosis can be associated with recurrent headaches and fatigue. A comprehensive assessment of clinical features including symptoms in IE has not been performed. Methods: Patients signed informed consent to participate in an observational study approved by the Institutional Review Board. Enrollment criteria included: age 18 years or older; hemoglobin level greater than 16 g/dL on two occasions at least 3 months apart or greater than 15 g/dL if undergoing phlebotomy; negative testing for JAK2 mutations; and negative work up for secondary causes of erythrocytosis. Baseline assessment included history and physical exam, vital signs, pulse oximetry, and body mass index. Baseline laboratory exams included a complete blood cell count, complete metabolic panel, C-reactive protein, iron panel, ferritin, hemoglobin A1C, erythropoietin level. Abdominal ultrasound was performed to evaluate for splenomegaly. The Myeloproliferative Symptom Assessment Form (MPN-SAF) was used to assess for the presence and severity of a broad range of symptoms that may be expected to occur in patients with IE. The MPN-SAF was administered at baseline and every 6 months thereafter. Results: 35 patients had data available for analysis. Patient characteristics are shown in Table 1. The most prevalent co-morbid conditions were those known to be associated with cardiovascular disease risk and metabolic syndrome, including hepatic steatosis identified on abdominal ultrasound in 63% of patients. Three (8.6%) patients had a history of venous or arterial thrombosis. Two (5.8%) patients had a history of lymphoma (NK/T-cell and Hodgkin). Three patients (8.6%) had a first-degree relative with a myeloproliferative neoplasm (chronic myelomonocytic leukemia, essential thrombocytosis and polycythemia vera) and one patient had a son with IE and history of stroke. 16 (46%) patients were taking aspirin and 11 (31%) had undergone phlebotomy within 3 months of study enrollment. Patients reported the following symptoms on the MPN-SAF at baseline: fatigue (77%), early satiety (57%), difficulty sleeping (57%), numbness/tingling (51%), headaches (49%), concentration problems (40%), itching (40%), bone pain (37%), night sweats (37%), depression (37%), abdominal pain (37%), abdominal discomfort (37%), inactivity (37%), problems with sexual desire/function (34%), dizziness/lightheadedness (31%), cough (26%), fever (17%), and unintentional weight loss (17%). Fatigue carried the highest average symptom intensity (3.77, SD 3.17). Discussion: In this study, we describe the clinical features associated with IE in a multiracial cohort. Patients with IE are frequently symptomatic and have a high incidence of hepatic steatosis by ultrasound. Disclosures No relevant conflicts of interest to declare.

Introduction: Idiopathic erythrocytosis (IE) is characterized by a persistently elevated hemoglobin, equivocal erythropoietin (EPO) levels, absence of janus kinase 2 (JAK2) mutations suggestive of polycythemia vera (PV) and no secondary cause. One study used a targeted 21 gene next-generation sequencing panel and identified novel variants in known erythrocytosis-related genes as well as novel genes associated with the oxygen-sensing pathway. However, expanded sequencing of blood and matched tissue samples in a large ethnically diverse group of IE patients has not been performed. Methods: All patients signed informed consent to participate in an observational study approved by the Institutional Review Board; they provide blood and buccal mucosa samples at study entry and at 24-month follow-up. Patients were enrolled if JAK2 testing and a complete work up for secondary causes was negative. They were required to have hemoglobin levels greater than 16 g/dL on two occasions or greater than 15 g/dL if undergoing phlebotomy. Our initial sequencing of 20 IE patients was performed utilizing high resolution whole-exome sequencing of circulating blood samples (disease) at a mean coverage of 390x and matched normal (buccal) samples at mean coverage of 300x. To stratify samples by genetic ancestry, we performed a population stratification principle component analysis (PCA) and STRUCTURE using Ancestry Informative Markers derived from 1K Genome Phase1_v3 Exome database. The primary in-silico analysis was performed on the baseline samples from treatment-naïve patients. The whole-exome data was generated in accordance to GATK's best practices with same filters applied as described by Exome Aggregation Consortium. The additional downstream in-silico paired analysis was performed using MutSig2.0 (Mutation Significance) algorithm to determine significant mutations and GISTIC (The Genomic Identification of Significant Targets in Cancer) to identify the significant copy number events, IPA (Ingenuity Pathway Analysis) to determine pathways along with other computational . Results: Median age at baseline was 52 years (range 35-71). Six patients (30%) were female and 14 (70%) were male. Median values and ranges for laboratory parameters at baseline were as follows: WBC 6.6 x 109/L (5-9.7), Hgb 17 g/dL (15.5-19.8), Plt 218 x 109/L (86-374), and EPO level 9.8 IU/L (2-14.3). Three patients had a personal history of malignancy, including 2 with lymphoma. Two patients had a family history of myeloid malignancy (chronic myeloid leukemia and PV). Our ancestry analysis of initial 20 patients with IE identified 6 patients with high European percent ancestry (EUR), 1 patient with high Asian percent ancestry (EAS) and 13 patients with high percent Ad Mixed ancestry (AMR). In our cohort, 60% (12/20) of patients had been also diagnosed with a liver disorder (11 with fatty liver, 1 with cirrhosis) that was not significantly different across populations. We identified, on average, 42 non-silent somatic mutations (not present in the buccal samples) in whole blood across our cohort with no statistical difference (p=0.671) in mutation burden between ancestry groups or between patients with and without liver disease. Age, gender, and ethnicity were not associated with mutation burden. Utilizing MutSig algorithm, we identified a novel candidate gene, CHAF1A, with high mutation prevalence of 30% in patients with IE. Further analysis of mutation landscape identified somatic nonsilent mutations in 25 known oncogenes which were present in at least 10% of patients. Our mutation signatures in IE identified a significant association with failure of double stranded DNA repair. Only one patient had a mutation in TET2. Further analysis of copy number indicated copy number loss in genes such as SETD3 and GSH associated with chromatin assembly which may suggest alterations in chromatin assembly and changes in the epigenome. Our analysis also identified a high number of 9p and 13q gains in patients with IE. Conclusion: In this study, we utilized high-resolution next generation sequencing in association with comprehensive clinical annotation to determine potential molecular drivers of IE in a multi-ethnic population. We identified somatic mutations in a subset of patients which may represent clonal hematopoiesis. Long term follow up of outcomes in this cohort may clarify the significance of these mutations in the pathogenesis of IE. Disclosures No relevant conflicts of interest to declare.