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Background Clonal cytopenia of undetermined significance (CCUS) is a newly described entity with a high probability to progress into myeloid disorders upon follow-up (Malcovati et al, Blood 2017). Little data is known on how to effectively treat patients with CCUS. We hereby describe a single institution experience in managing symptomatic CCUS patients. Methods Patients who had CCUS and underwent active treatment were identified after receiving IRB approval. CCUS diagnosis was rendered if patients had non-diagnostic bone marrow biopsy evaluation combined with evidence of pathogenic myeloid somatic mutation using our institution next generation sequencing (NGS) panel (OncoHeme, Mayo Clinic). "Symptomatic patients" is determined by patients who need therapy beyond blood transfusion. Treatment type is based on physicians' choice. Clinical and lab results were collected. Response and progression were graded based on the MDS IWG 2006 criteria. The date of treatment initiation was used for overall survival (OS) and progression (either to myeloid neoplasm or worsening cytopenias) free survival (PFS) calculation. Stata 14.1(StatCorp) was used for data analysis. Results 1) Cohort characteristics: Between 2015 and 2020, 24 patients met inclusion criteria with median age of 72 years (range: 24-87). Twenty (83%) were men. 15 (63%) were white, 1 (4%) African American, 1 (4%) Native American and 7 unknown (33%). Eighty percent had ECOG PS≤1. Four (17%) patients had prior other hematology disorders and 3 (13%) had solid tumor. Two (8%) patients had previous radiation therapy, 2 (8%) received chemotherapy and 1 (4%) received allogeneic stem cell transplantation. At the time of treatment, the median of hemoglobin was 8.9 (range 6.8-13.7) g/dL, white blood cell 2.9 (range 1.4-12.3) *109/L, and platelet 76 (range 8-407) *109/L. Red blood cell (RBC) and platelet transfusion dependency rate were 54% and 29%, respectively. We identified 23 different mutations. Most common mutations occurred in SRSF2 (N=5, 21%), TET2 (N=5, 21%), U2AF1 (N=5, 21%), and TP53 (N=4, 17%). Ten (42%) patients had 1 mutation, and 14 (58%) had ≥2 mutations. Treatment included hypomethylating agents (HMA) (N=10, 42%), growth factors (N=11, 46%), steroid (N=4, 17%), testosterone (N=2, 8%), cyclosporine (N=2, 8%), rituximab (N=1, 4%), immunoglobulin (N=1, 4%), vitamin B12 and iron (N=1, 4%). Seven (29%) patients received ≥2 treatments. Median follow up duration was 17.5 months (range 8.9-26.1). Median time from the diagnosis to treatment was 2.1 months (range 0-26.8). 2) Molecular correlation with response The overall response rate (RR) was 50% (hematological or symptomatic improvement). Most responders (67%) were treated with HMA (p=0.013). HMA treatment effect was not significantly associated with DNA methylation genes (DNMT3A, IDH1, IDH2 and TET2, all p>0.05). The RR was 100% for the three patients with IDH1 mutation (treated with HMA, steroid and erythropoietin stimulating agent (ESA), respectively). The RR was 100% for the 3 patients with ZRSR2 mutation (2 HMA, 1 testosterone). The RR was 0% for 3 patients with SF3B1 mutation (1 testosterone and ESA, 1 vitamin B12 and iron, and 1 HMA). The number of mutations had no impact on RR (p=0.41). Table demonstrates the RR for each treatment. 3) Survival and progression outcome At the last day of follow up, RBC and platelet transfusion dependency rate were 38% and 25%, respectively. The median PFS was 16.9 months (95% CI: 7.5-65.3). Six patients (25%) progressed to myeloid malignancy with 4 myelodysplastic syndrome (MDS) and 2 acute myeloid leukemia (AML). Five patients progressed with worsening cytopenias. Among the 4 MDS patients, they had SF3B1, TET2, and ASXL1 mutations at the diagnosis of CCUS. Within the 2 AML pts, 1 had RUNX1 mutation, 1 had BCOR, DNMT3A, PHF6, RUNX1, SF3B1, STAG2, and TET2 mutations at the diagnosis of CCUS. The median OS was not reached with an estimated 2 year OS 77%. Three (13%) patients progressed with cytopenia and died. Their NGS at CCUS diagnosis showed ASXL1, TP53 and U2AF1 mutations, respectively. Conclusion Patients with CCUS responded to available treatments used for myeloid diseases. HMA were effective in this cohort. Mutation profile may have an impact on treatment response. Studies of larger cohorts are needed to validate our findings. Additionally, development of response and progression criteria would be helpful for such patients for reporting treatment outcomes. Disclosures Shah: Dren Bio: Consultancy.

Introduction: Myeloproliferative neoplasms (MPN), including primary myelofibrosis (PMF), essential thrombocythemia (ET) and polycythemia vera (PV), have a propensity to evolve into blast phase myeloproliferative neoplasm (BP-MPN) with a 20-year incidence rate of 9.3 %, 3.9% and 2.6%, respectively. (Szuber et al., 2019)Treatment options for BP-MPN are limited and the prognosis of these patients is dismal with a median survival of only 3.6 months and 5-year survival rate of <5%.(Tefferi et al., 2018) Considering the recent reported data on the efficacy of venetoclax when combined with hypomethylating agents (HMA) in acute myeloid leukemia (AML) in both relapsed/refractory and newly diagnosed unfit patients, we have extended such combination therapy for patients with BP-MPN. Methods: We retrospectively analyzed 14 consecutive BP-MPN patients who received venetoclax plus HMA therapy between August 2018 and June 2020. We collected data regarding clinical characteristics of chronic phase MPN and BP-MPN, cytogenetic and leukemia mutation profile, efficacy and outcome. Oral venetoclax was administered in combination with azacitidine 75 mg/m2 days 1-7 (5 patients) or decitabine 20 mg/m2 days 1-5 (9 patients). Venetoclax dose was adjusted based on drug interactions particularly with azole antifungal prophylaxis. Diagnostic, risk and response assignments were according to the 2017 European LeukemiaNet (ELN) criteria.(Döhner et al., 2017) Minimal residual disease (MRD) assessment by flow cytometry, karyotype or next-generation sequencing (NGS) was performed in a subset of patients. Results: Patient characteristics at time of leukemic transformation, treatment details, response rates and overall outcome are shown in Table 1. Median age of patients was 67 years (range 48-81) with poor-risk cytogenetics in 69% of patients. JAK2 was mutated in 10 patients (71%) and CALR in 2 (14%); other mutations included TP53 in 5 patients (36%), TET2 in 4 (29%), KRAS in 3 (21%), IDH1/2 in 3 (21%), ASXL1 in 2 (14%) and U2AF1 in 2 (14%). Eight patients (57%) received venetoclax and HMA combination therapy upfront for their BP-MPN, 2 patients (14%) had failed HMA therapy previously and one patient had prior allogeneic hematopoietic stem cell transplant (AHSCT). Two patients (14%) presented with myeloid sarcoma; one of these two patients documented partial resolution of the extramedullary tumor by imaging studies, after treatment with venetoclax plus HMA. Among the remaining 12 patients, overall response rate (ORR) was 42% (n=5) and included complete remission (CR) in 3 patients (25%) and partial remission (PR) in another 2 (17%). The best response to therapy was seen after a median of 1 month (range, 1-2). Among 3 CR responders, 2 (66.6%) had minimal residual disease negative by NGS and not evidence of preceding MPN, then they successfully transitioned to AHSCT, while the third CR patient at the time of best response showed persistence of the TP53 mutation. Additionally, one PR patient subsequently relapsed and received salvage chemotherapy followed by AHSCT. (Table 2) Although data is limited by the small cohort and short follow up when the outcome of patients treated with venetoclax in combination with HMA were compared to Mayo Clinic's historical control of patients with BP-MPN treated with HMA alone (n=26) or intensive chemotherapy (n=69), there was higher CR rate in patients treated with venetoclax and HMA (25%) compared to those receiving HMA alone (4%; p=0.048) but not to those receiving intensive chemotherapy (35%; p<0.0001). Moreover, the intensive chemotherapy cohort showed 24% of CR with incomplete hematologic recovery (CRi) not seen in patients receiving HMA alone or HMA with venetoclax (Figure1). Conclusions: The relatively high rate of complete response observed in our patients with BP-MPN were similar to those reported in a pivotal study of elderly unfit AML patients treated upfront with venetoclax + HMA with overall response rates of 68%.(DiNardo et al., 2019) Furthermore, responders included patients with adverse molecular risk factors who usually respond poorly to conventional chemotherapy (i.e. TP53 mutated patients). Our observations provide preliminary evidence for the potential efficacy of venetoclax and HMA combination therapy in BP-MPN with the goal of achieving CR/CRi followed by consolidative AHSCT wherever possible to provide durable remission and meaningful survival benefit. Disclosures Foran: Agios: Honoraria, Research Funding; Trillium: Research Funding; Takeda: Research Funding; Kura Oncology: Research Funding; Aptose: Research Funding; Aprea: Research Funding; Actinium: Research Funding; Boehringer Ingelheim: Research Funding; Abbvie: Research Funding; BMS: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees; Servier: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Revolution Medicine: Consultancy; Xencor: Research Funding; H3Biosciences: Research Funding.

BACKGROUND: MF is characterized by anemia, splenomegaly, debilitating constitutional symptoms, and shortened survival. Fedratinib is an oral JAK2 inhibitor approved in the United States for treatment (Tx) of adult patients (pts) with intermediate (INT)-2 or high-risk MF. The randomized, placebo (PBO)-controlled, phase III JAKARTA trial evaluated clinical outcomes with fedratinib in pts with JAK-inhibitor-naïve MF. Initial findings from the JAKARTA study have been reported (Pardanani, JAMA Oncol, 2015). JAKARTA data were recently reanalyzed in preparation for regulatory review. OBJECTIVE: Evaluate the clinical efficacy and safety with fedratinib 400 mg/day in updated analyses of the JAKARTA study. METHODS: JAKARTA was an international, randomized, double-blind, PBO-controlled study that enrolled pts aged ≥ 18 years with INT-2 or high-risk primary, post-PV, or post-ET MF. Pts were randomly assigned 1:1:1 to fedratinib 400 mg, fedratinib 500 mg, or PBO, administered orally once-daily for ≥ 6 consecutive 4-week cycles ("randomized Tx period"). After completing 6 Tx cycles (or earlier in the case of disease progression), pts randomized to PBO could crossover to fedratinib. The primary endpoint was the spleen volume response rate (SVRR), the proportion of pts who achieved a ≥ 35% spleen volume reduction from baseline (BL) confirmed by MRI/CT at end of cycle 6 (EOC6), confirmed by follow-up scan 4 weeks later. Secondary endpoints included SVRR without 4-week confirmation, durability of spleen response, and symptom response rate (proportion of pts with a ≥ 50% reduction from BL in total symptom score [TSS] on the modified Myelofibrosis Symptom Assessment Form [MFSAF]) at EOC6. An exploratory analysis evaluated proportions of pts with ≥ 50% reductions from BL at EOC6 in individual MFSAF symptom scores: night sweats, pruritus, abdominal discomfort, early satiety, pain under ribs on left side, and bone or muscle pain. These analyses are limited to pts randomized to fedratinib 400 mg (the approved starting dose of fedratinib) or to PBO. Efficacy analyses were performed for the intention-to-treat (ITT) population and safety was assessed in all pts who received ≥ 1 dose of study drug. RESULTS: 96 pts were randomized to fedratinib 400 mg and 96 pts to PBO. In the fedratinib and PBO arms, median ages at BL were 63 (range 39-86) and 66 (27-85) years, respectively, 65% and 60% of pts had primary MF, 76% and 73% had constitutional symptoms, median spleen volumes were 2652 (316-6430) and 2660 (662-7911) mL, and median TSS were 15.3 (0-57) and 12.4 (0-53). Median durations of exposure were 15.5 months in the fedratinib 400 mg arm and 6 months in the PBO arm. With a confirmation scan 4 weeks after EOC6, SVRR was significantly higher in the fedratinib arm (37% [95%CI 27%, 46%]) vs. the PBO arm (1% [0%, 3%]) (P < 0.0001). Without the confirmation scan, SVRR at EOC6 was 47% (95%CI 37%, 57%) in the fedratinib 400 mg arm and 1% (0%, 3%) in the PBO arm (P < 0.0001; Figure). Estimated median duration of spleen response with fedratinib was 18.2 months. Symptom response rate at EOC6 for evaluable pts (ie, with TSS data available at BL and EOC6) was 40% (36/89) with fedratinib and 9% (7/81) with PBO (P < 0.0001). Fedratinib was associated with higher response rates vs. PBO in all 6 MFSAF symptoms: abdominal discomfort (41% vs. 15%, respectively), night sweats (58% vs. 22%), bone or muscle pain (33% vs. 14%), early satiety (51% vs. 18%), pruritus (39% vs. 20%), and pain under the ribs on the left side (42% vs. 24%). The most common Tx-emergent adverse events (TEAEs) during the randomized Tx period were grade 1-2 gastrointestinal events (Table). New or worsening grade 3 anemia and grade ≥ 3 thrombocytopenia laboratory abnormalities were reported in 34% and 12% of fedratinib-treated pts, most (75%) occurring within 3-4 months of Tx initiation. Compared with PBO, fedratinib-treated pts more frequently experienced increased serum creatinine, amylase, and lipase; these were mainly grade 1-2 in severity. TEAEs led to Tx discontinuation for 14% of fedratinib pts and 8% of PBO pts. CONCLUSIONS: Fedratinib 400 mg/day significantly reduced splenomegaly and symptom burden in pts with JAK-inhibitor-naïve MF. Relatively few pts discontinued fedratinib therapy due to TEAEs, suggesting these events are manageable. These data corroborate results initially reported for the JAKARTA trial, and further support the efficacy and safety of fedratinib in this pt population. Disclosures Masszi: AbbVie: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Janssen-Cilag: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees. Drummond:Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Gilead: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Astellas: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Blueprint Medicine Corporation: Research Funding; Jazz: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Jourdan:Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Abbvie: Membership on an entity's Board of Directors or advisory committees. Vannucchi:Blueprint: Membership on an entity's Board of Directors or advisory committees; Celgene/BMS: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; AbbVie: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Incyte: Membership on an entity's Board of Directors or advisory committees. Ribrag:AZD: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other; Infinity: Honoraria, Membership on an entity's Board of Directors or advisory committees; Roche: Honoraria, Membership on an entity's Board of Directors or advisory committees; Incyte: Honoraria, Membership on an entity's Board of Directors or advisory committees; MSD: Honoraria, Membership on an entity's Board of Directors or advisory committees; BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees; gilead: Honoraria, Membership on an entity's Board of Directors or advisory committees; nanostring: Honoraria, Membership on an entity's Board of Directors or advisory committees; EPZ: Honoraria, Membership on an entity's Board of Directors or advisory committees; epizyme (EPZ): Research Funding; argenX: Research Funding; pharmamar: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; servier: Consultancy; gustave roussy comprehensive cancer center: Current Employment. Rambaldi:Abbvie: Honoraria; Jazz: Honoraria; Astellas: Honoraria; Omeros: Honoraria; Novartis: Honoraria; Gilead: Honoraria; Bristol-Myers Squibb: Honoraria; Sanofi: Honoraria; Amgen: Honoraria; Pfizer: Honoraria; Roche: Honoraria; MSD: Honoraria. Rose:Bristol-Myers Squibb Company: Current Employment, Current equity holder in publicly-traded company. Zhang:Bristol-Myers Squibb Company: Current Employment, Current equity holder in publicly-traded company. Harrison:Promedior: Honoraria; Roche: Honoraria; Sierra Oncology: Honoraria; Novartis: Honoraria, Research Funding, Speakers Bureau; Celgene: Honoraria, Research Funding, Speakers Bureau; Shire: Honoraria, Speakers Bureau; Incyte Corporation: Speakers Bureau; Gilead Sciences: Honoraria, Speakers Bureau; Janssen: Speakers Bureau; AOP Orphan Pharmaceuticals: Honoraria; CTI Biopharma Corp: Honoraria, Speakers Bureau.

Background: Chronic myelomonocytic leukemia (CMML) is a rare chronic myeloid malignancy with a poor prognosis. There are few epidemiologic studies on CMML. We studied population-based outcomes in CMML using the Surveillance, Epidemiology, and End Results (SEER) and the National Cancer Database (NCDB). Methods: The SEER 18 registries and NCDB Participant User File were used to identify patients with ICD-O-3 diagnosis code 9945/3 from 2004-2015. Incidence was identified using SEER and age-adjusted to the U.S. 2000 standard population. Causes of death were obtained and CMML-related death was defined as death from any myeloid disorder (CMML, acute monocytic leukemia, acute myeloid leukemia [AML], chronic myeloid leukemia, other myeloid/monocytic leukemia, and aleukemic, subleukemic, and NOS) to avoid misattribution. Relative survival (RS) was defined as the ratio of the proportion of observed survivors in a cohort of CMML patients to the proportion of expected survivors in a comparable set of individuals that did not have CMML, adjusting for the general survival of the US population for race, sex, age, and time when the diagnosis was established. Time to AML was calculated using the left-truncated life tables session and a 3 month latency period was used to prevent misattribution. Per the NCDB's system on classifying treatment, hydroxyurea, decitabine, and azacitidine are considered chemotherapy. The Kaplan-Meier method was used to calculate overall survival (OS), and Cox regression model was used to identify predictors of survival. Variables significant in univariate analysis (age, Charlson Deyo score [CDS], insurance type, and treatment facility type) were included in a multivariate analysis. Statistical analyses were performed using SAS version 9.0. Results: In the SEER database (n=4437), the median age at diagnosis was 76 years (interquartile range [IQR] 68-83) and 2,783 patients (63%) were male. Incidence rates (1 case/1,000,000 individuals) were as follows: overall 4.4, male 6.6, female 2.9, Non-Hispanic White 4.9, and Non-Hispanic Black 3.3. The incidence did not significantly change from 2004 to 2015. With a median follow up of 5.8 years (95% CI: 5.5-6.3), the median OS was 1.3 years (95% CI: 1.3-1.4). 3635 patients (82%) died. Among those who died, 2016 patients (55%) deaths were CMML-related. When comparing CMML patients to the US general population, 3,156 patients were matched to the expected survival tables. In the general U.S. population, the expected survival for 12, 24, 36, 48 and 60 months was 95.6%, 91.3%, 87.1%, 83.1%, and 79.2%, respectively. In contrast, the RS for patients with CMML at the same time points was 63.6%, 46.2%, 35.2%, 28.8%, and 23.1% (Figure 1). 229 patients (5.2%) progressed to AML. The median time to AML was 1.2 years (IQR 0.6-2.3). In NCDB (n=6403), the median age at diagnosis was 74 years (IQR 66-82). With the median follow up of 6 years (95% CI 3.5-9), the median OS was 1.3 years (95% CI 0.4-3.3). The distribution of CDS score 0, 1, and >1 was 4518 (71%), 1193 (18%), and 692 (11%) respectively. 4687 (77%) had government insurance, while 1399 (23%) had private insurance. 3750 (60%) were treated at a non-academic center. The year of diagnosis was associated with improved OS (HR 0.99, 95% CI: 0.98-0.99). Patients who were diagnosed in 2012-2015 had improved OS compared to patients diagnosed in 2004-2007 (HR 0.89, 95% CI 0.83-0.95) with OS at 1- and 5- years being 60% and 16% (2012-2015) vs. 56% and 17% (2008-2011) vs. 53% and 15% (2004-2007) respectively. 3029 (48%) patients received chemotherapy, while 270 (4%) patients received stem cell transplant as the first-line therapy. The median time to chemotherapy from diagnosis was 19 days (IQR 6-43). OS at 1-, 5-, and 10-years was 57%, 18%, and 4% (no chemotherapy), 56%, 14%, and 7% (received chemotherapy), 79%, 44%, and 38% (received stem cell transplant), and 56%, 15%, 5% (no stem cell transplant). OS curves are shown in Figure 2. Factors independently predicting inferior OS were age ≥ 65 years at the time of diagnosis (p<0.001), CDS ≥ 1 (p<0.001), government insurance (p<0.001), and treatment at a non-academic center (p<0.001) (Table). Conclusion: Despite an improvement in survival over the years, CMML continues to be a cause of significant morbidity and mortality. Older age, CDS ≥ 1, government insurance, and treatment at non-academic facilities were predictive of inferior survival. Disclosures Shah: Dren Bio: Consultancy.

Nearly 20% of patients with myelofibrosis progress to blast phase disease; an aggressive form of acute myeloid leukemia. Although previous studies have implicated loss of TP53 or JARID2 in progression, by and large the genetic events that lead to conversion to blast phase remain unknown. To identify genes whose loss drives progression, we performed a focused CRISPR/Cas9 screen in which murine Jak2V617F bone marrow cells expressing Cas9 were transduced with two separate sgRNA libraries of known tumor suppressor genes and subjected to colony replating assays. Transduction of one of the two libraries led to serial replating and enhanced self-renewal of the Jak2V617F cells. Subsequent DNA sequencing revealed enrichment of all four guides targeting STK11, the gene that encodes LKB1 which regulates a number of key cellular pathways including energy utilization by activation of AMPK. To confirm that loss of Stk11 is the event that leads to increased clonogenicity, we collected cells from Jak2V617F/Vav-Cre+ and control Vav-Cre+ mice and induced Stk11 knockout by electroporating Cas9-Stk11 sgRNA ribonucleoprotein complexes. Consistent with the screening results, only Jak2V617F Vav-Cre+ cells with Cas9-Stk11 sgRNA showed serial replating. To determine whether Stk11 is required for growth of cells with a different driver of enhanced JAK/STAT signaling, we doubly transduced Stk11 homozygous floxed bone marrow cells with MPLW515L-mCherry and Cre-GFP to delete Stk11. As expected, cells with both MPLW515L and Cre recombinase showed enhanced self-renewal, while singly infected control cells failed to replate. These results demonstrate that activation of JAK/STAT signaling can overcome the requirement for Stk11 in normal hematopoiesis and suggest that STK11 loss may be a strong driver of malignant transformation in combination with enhanced JAK-STAT signaling. We next investigated the mechanism by which loss of STK11 cooperates with enhanced JAK/STAT signaling to promote leukemia. RNA-sequencing of wild-type, Stk11+/+/ MPLW515L, and Stk11-/-/MPLW515L hematopoietic cells revealed enrichment of a number of pathways related to hypoxia, oxidative phosphorylation and mitochondrial translation in cells lacking LKB1. Western blot assays confirmed activation of mTOR signaling as well as HIF1a stabilization and pathway activation, both of which have been reported to lie downstream of LKB1 loss. We also performed a number of studies to determine the relevance of reduced LKB1 expression to leukemic progression. First, we induced deletion of Stk11 in mice that were transplanted with HSPCs expressing MPLW515L after development of the MPN phenotype. Loss of Stk11 caused a rapid lethality that was associated with enhanced bone marrow fibrosis and osteosclerosis. We also observed accumulation of leukemic blasts in small clusters consistent with AML transformation arising in the spent phase MPN. Additionally, we deleted STK11 by CRISPR/Cas9 in primary MPN patient samples and monitored their engraftment in immunocompromised mice. We observed enhanced engraftment and increased reticulin fibrosis and osteosclerosis in mice that received the STK11 edited cells compared to those with non-targeted sgRNA. Third, we compared the expression of STK11 in paired blast and chronic phase myelofibrosis patient samples by RT-PCR. Consistent with the hypothesis that loss of STK11 facilitates leukemia, we found that its expression was decreased by more than 50% in five of seven paired post-MPN AML patient samples, with two having STK11 levels below 20%. We further validated downregulation of LKB1 by immunohistochemistry on paired chronic and blast phase MPN specimens and observed little staining in the blast phase specimens. Finally, to further show that the mechanism of in vitro enhanced self-renewal is related to leukemia progression, we stained the paired marrows for HIF1a and saw a dramatic increase in staining at the AML phase. We also analyzed RNA-seq data of paired chronic versus blast phase MPNs specimens and observed that there is a strong congruence of enriched pathways that are associated with the in vitro mouse HSPC phenotype and the human blast phase progression, such as oxidative phosphorylation and hypoxia. Together, our study demonstrates that loss of LKB1/STK11 promotes transformation of cells with activated JAK/STAT signaling and that STK11 is a prominent candidate tumor suppressor gene in post-MPN AML. Disclosures Gurbuxani: UpToDate: Honoraria. Hoffman:Dompe: Research Funding; Protagonist: Consultancy; Abbvie: Membership on an entity's Board of Directors or advisory committees; Forbius: Consultancy; Novartis: Membership on an entity's Board of Directors or advisory committees. Levine:Astellas: Consultancy; Amgen: Honoraria; Gilead: Honoraria; Qiagen: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Morphosys: Consultancy; Novartis: Consultancy; Prelude Therapeutics: Research Funding; Loxo: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Imago: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; C4 Therapeutics: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Isoplexis: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Honoraria, Research Funding; Roche: Consultancy, Honoraria, Research Funding; Lilly: Consultancy, Honoraria; Janssen: Consultancy. Rampal:Galecto: Consultancy; Incyte: Consultancy, Research Funding; Constellation: Research Funding; Stemline: Consultancy, Research Funding; Celgene: Consultancy; Jazz Pharmaceuticals: Consultancy; CTI Biopharma: Consultancy; Abbvie: Consultancy; Pharmaessentia: Consultancy; Promedior: Consultancy; Blueprint: Consultancy.

Abstract Disease overview Polycythemia vera (PV) and essential thrombocythemia (ET) are myeloproliferative neoplasms (MPN) respectively characterized by clonal erythrocytosis and thrombocytosis; other disease features include leukocytosis, splenomegaly, thrombosis, bleeding, microcirculatory symptoms, pruritus and risk of leukemic or fibrotic transformation. Diagnosis Bone marrow morphology remains the cornerstone of diagnosis. In addition, the presence of JAK2 mutation is expected in PV while approximately 90% of patients with ET express mutually exclusive JAK2 , CALR or MPL mutations (so called driver mutations). In ET, it is most important to exclude the possibility of prefibrotic myelofibrosis. Survival Median survivals are approximately 15 years for PV and 18 years for ET; the corresponding values for patients age 40 or younger were 37 and 35 years. Certain mutations (mostly spliceosome) and abnormal karyotype might compromise survival in PV and ET. Life‐expectancy in ET is inferior to the control population. Driver mutations have not been shown to affect survival in ET but risk of thrombosis is higher in JAK2 mutated cases. Leukemic transformation rates at 10 years are estimated at <1% for ET and 3% for PV. Thrombosis risk In PV, two risk categories are considered: high (age > 60 years or thrombosis history present) and low (absence of both risk factors). In ET, four risk categories are considered: very low (age ≤ 60 years, no thrombosis history, JAK2 wild‐type), low (same as very low but JAK2 mutation present), intermediate (age > 60 years, no thrombosis history, JAK2 wild‐type) and high (thrombosis history present or age > 60 years with JAK2 mutation). Risk‐adapted therapy The main goal of therapy in both PV and ET is to prevent thrombohemorrhagic complications. All patients with PV require phlebotomy to keep hematocrit below 45% and once‐daily or twice‐daily aspirin (81 mg), in the absence of contraindications. Very low risk ET might not require therapy while aspirin therapy is advised for low risk disease. Cytoreductive therapy is recommended for high‐risk ET and PV, but it is not mandatory for intermediate‐risk ET. First‐line drug of choice for cytoreductive therapy, in both ET and PV, is hydroxyurea and second‐line drugs of choice are interferon‐α and busulfan. We do not recommend treatment with ruxolutinib in PV, unless in the presence of severe and protracted pruritus or marked splenomegaly that is not responding to the aforementioned drugs. New treatment directions Controlled studies are needed to confirm the clinical outcome value of twice‐daily vs once‐daily aspirin dosing and the therapeutic role of pegylated interferons and direct oral anticoagulants.

Abstract Introduction: Primary myelofibrosis (MF) is a chronic clonal malignancy which is compounded by the presence of inflammation, bone marrow fibrosis/scarring and risk of transformation to acute myeloid leukemia (AML). Ruxolitinib (Rux), a JAK2 inhibitor, is beneficial for reduction of symptoms and splenomegaly, however, significant myelosuppression is also a toxic side effect of this treatment, limiting optimal drug dosage in many patients. 9-ING-41 is a small molecule GSK-3β inhibitor which has significant clinical activity in patients with advanced malignancies and causes no myelosuppression. We thus assessed the effect of 9-ING-41in MF alone or in combination with Rux. We performed ex-vivo colony assays with primary cells from cases with untreated MF along with normal bone marrow (BM). The experiments were designed to assess the effect of drug on the ability of MF stem/progenitor cells to proliferate/differentiate by evaluating the number, size, and the morphology of colonies, both with and without treatment. Methods: Primary peripheral blood mononuclear cells from MF patients and BM from controls were plated in methylcellulose in duplicate, containing cytokines promoting hematopoietic growth, in the presence of either no drug, Rux only (0.05µM), 9-ING-41 only (range: 0.125µM - 10µM), or a combination of 9-ING-41 (range: 0.125µM - 10µM) and Rux (0.05uM). Colonies were counted ten days later, and colony growth frequency, distribution, and morphology were calculated. Results: We observed an increasing presence of large primitive granulocytic/erythroid/macrophage/monocyte (GEMM) colonies proportional to no drug treatment and linear to increasing concentrations of 9-ING-41 in both MF cases relative to other colony types and normal BM, suggestive of a selective primitive proliferative and/or differentiation effect of GSK-3β inhibition on MF hematopoietic progenitor growth. We did not observe this effect in the combinatory treatments (9-ING-41 + Rux), and there was no significant difference on MF colony number or size in colonies grown without treatment or with Rux alone (Figure 1 A/C/E). However, the morphology of the colonies within different treatments was remarkable. In both MF cases, either without treatment or addition of Rux only (0.05µM), the colonies appeared similar to each other -irregular and disorganized relative to normal bone marrow (Figure 1, B/D/F). In the presence of 9-ING-41 alone, we observed morphologically very large, fully differentiated primitive colonies in both MF cases. However, the most striking observation was the comparison of colonies after combination of both inhibitors. These colonies appeared normal and healthy-looking -discreet, round size and similar in morphology to normal bone marrow colonies. Additional studies are currently underway to look at the signaling consequences, including proliferation, differentiation, and cell cycle effect. Our preliminary results suggest this same mechanism might be occurring in MF cases and could explain the selective presence of large differentiated primitive colonies in MF vs. normal BM in the presence of 9-ING-41. Combining 9-ING-41 with Rux may reduce the dose needed for optimal therapeutic response and/or reverse myelosuppression. A Phase 2 study of 9-ING-41 in patients with advanced MF will be conducted. Citation Format: Terra L. Lasho, Francis Giles, Christy Finke, Naseema Gangat, Ayalew Tefferi. 9-ING-41, a GSK-3β-selective small molecule inhibitor, in combination with ruxolitinib in JAK2617F primary myelofibrosis [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5217.

7083 Background: Patients with Myelofibrosis (MF) suffer from significant fatigue, constitutional symptoms and splenomegaly (Mesa et. al. Cancer 2007) not improved by current therapy. The 6MWT measures functional status in cardio-pulmonary clinical trials. We undertook a prospective feasibility study of the 6MWT in MF patients for use in therapeutic clinical trials. Methods: MF patients (able to walk, normal vitals, not on a JAK2 inhibitor) were eligible. Baseline features and the MF symptom assessment form (MFSAF: Mesa et. al. Blood 2008) was administered. The 6MWT was administered in standardized fashion (American Thoracic Society: observed laps of a 30 - 35 meter long course, indoors, level, without encouragement), and then repeated for further validation. Results: Safety of the 6MWT: Fifty five MF patients were enrolled with only 2 patients (3%) stopping the 6MWT early secondary to fatigue. There were no adverse events attributable to undergoing the 6MWT in test subjects. Results of the 6MWT: Current results of our ongoing study demonstrate compared to age matched healthy volunteer controls, males with MF walk approximately 90 meters less and females with MF walk approximately 60 meters less (Table). Correlation of 6MWT Results to Patient Reported Symptoms (MFSAF): Results from the MFSAF demonstrated the fatigue (as assessed by the brief fatigue inventory (Medoza et. al. Cancer 1999) embedded in the MFSAF) showed patients with a higher BFI (i.e., more fatigued) had more impairment in the 6MWT than those with low BFI scores. Symptomatic burden as assessed by the MFSAF were typical for MF, correlation of individual symptom severity with 6MWT will require a larger trial for adequate power. Conclusions: The 6MWT can be safely performed in un-selected MF patients, and demonstrates a functional deficit compared to published controls. Validation of the ability of the 6MWT to measure functional improvements in MF patients as a response to novel therapy trials is planned. [Table: see text] No significant financial relationships to disclose.

AbstractVenetoclax and hypomethylating agent (HMA) combination therapy is FDA‐approved for elderly or unfit acute myeloid leukemia (AML) patients unable to withstand intensive chemotherapy. The primary objective of the current study was to impart our institutional experience with the above regimen, outlining response, survival outcomes, and its determinants amongst 86 treatment‐ naïve and relapsed/refractory AML patients. A total of 44 treatment‐naïve AML patients, median age 73.5 years, enriched with secondary, therapy related and ELN adverse risk disease (n = 27) were studied. The CR/CRi rates of 50% (22 of 44 patients) were superior to 23% in a matched AML cohort treated with HMA alone (P = .005). Response rates were similar with TP53, FLT3, NPM1 and IDH mutations (P = .31). Moreover, CEPBA mutations (P = .03) and neutropenia (P = .05) emerged as predictors of complete response. Survivalwas prolonged in patients achieving CR/CRi (17 vs 3 months without CR/CRi, P < .001; conversely adverse ELN risk portended inferior survival. Amongst 42 relapsed/refractory AML patients, half received ≥2 prior therapies excluding transplant, and 15 (35.7%) had received HMA. A group of 14 patients (33.3%) attained CR/CRi; age > 65 years, AML with myelodysplasia, JAK2, DNMT3A, and BCOR mutations predicted complete response. Survival distinctions were based on CR/CRi (median survival 15 vs 3 months with/without CR/CRi; P < .001), and TP53 mutation status (P = .04). In summary, we corroborate existing reports demonstrating superior response and prolonged survival with venetoclax and HMA in treatment ‐naïve and relapsed/refractory AML patients regardless of genotype. Additionally, we identify unique predictors of response to therapy which require validation.