Fauble, Veena

Abstract The lysine deacetylase inhibitor suberoylanilide acid (SAHA) exhibits promising but limited activity in the treatment of acute myeloid leukemia (AML). To identify potential targets for rational combination therapies that increase the efficacy of SAHA in AML, we performed a functional RNA-interference (RNAi) drug modifier screen to identify genes that, when inhibited, potentiate or antagonize the in vitro anti-leukemic activity of SAHA. A total of 901 kinase, phosphatase and closely associated signaling genes were silenced in TF-1, HEL and THP-1 cells, with four different siRNA sequences per gene, both alone and in combination with SAHA treatment. Screen hit lists for each cell line were over-laid on an integrated functional relationship network. A community detection algorithm was then applied to this sub-network and siRNA sensitive modules were identified. Each module represents a highly connected set of genes in the integrated network. To identify pathways represented by each module, enrichment was evaluated using the National Cancer Institute (NCI) Protein Interaction Database (PID) pathways. Both cell line-specific and universal sensitizing targets, grouped into a small number of pathways, emerged from these screens. Seven pharmacologic inhibitors interfering with these pathways have been assessed in a panel of four or five AML cell lines, including SET-2, TF-1, HEL, THP-1, and OCI-AML3. Of the seven tested inhibitors targeting SAHA sensitizing networks, one combination exhibiting universal SAHA sensitization in all AML cell lines tested was further assessed in drug dose response assays using ex vivo cultures of bone marrow mononuclear cell populations derived from patients with various myeloid malignancies. The combination demonstrated “one-sided” SAHA EC50 fold-enhancement, as well as strong synergy by “two-sided” CalcuSyn combination index determination in all ex vivo specimens examined to date (N=6), including polycythemia vera (PV), chronic myelomonocytic leukemia (CMML), myelodysplastic syndrome/myeloproliferative neoplasm (MDS/MPN) overlap syndrome, MDS transformed to AML, and de novo AML samples. Thus, this first reported large-scale SAHA RNAi modifier screen in AML has identified a specific pathway/target whose inhibition broadly synergizes with HDAC inhibition in myeloid malignancies. A novel compound putatively targeting this respective pathway/target is currently in early clinical development and has shown limited single-agent activity. With the full dataset presented, we will propose a novel rational combination to enhance the activity of HDAC inhibition in AML and other myeloid malignancies that could be translated into design of a clinical trial. Disclosures: No relevant conflicts of interest to declare.

Abstract Introduction: Most myelofibrosis (MF) patients develop anemia during evolution of the disease process predicting decreased survival. Previous studies exploring the effect of danazol for the treatment of anemia in MF demonstrate responses in anemia. In prior randomized-controlled studies, ruxolitinib demonstrated improvements in splenomegaly, symptom burden, and even survival yet improvements in cytopenias were uncommon. We designed a phase II multicenter pilot study to evaluate the efficacy and tolerability of combination therapy with ruxolitinib and danazol in MF patients with anemia. Methods: This was a Simon optimum two-stage phase II trial. Eligible subjects received ruxolitinib, a JAK 1/JAK2 inhibitor, at 10mg (plat >75 x10x9) BID or 5mg (plat <75 x10 x9) BID) representing initial dose reduction for JAK inhibitor naïve patients to minimize cytopenias, in combination with tapered danazol up to 200mg orally TID. Dose escalation was allowed after completion of 28 days for lack of response or for disease progression. Pts without progression continued for 6 cycles at 56 days each. Treatment modifications were allowed for either medication based on emergent adverse events (AE). Treatment responses were evaluated every cycle by IWG-MRT criteria (Blood 2006). Patient reported outcome questionnaires (MPN-SAF, EORTC QLQ-C30) were administered at baseline, prior to treatment cycles, and at study discontinuation. Results: Patients: Fourteen pts enrolled (9 Mayo Arizona, 5 Mount Sinai). Median (Med) age 70.5 (range 43-78), M:F ratio 1.8:1. Ten primary MF, 2 post-essential thrombocythemia MF, and 2 post-polycythemia vera MF. JAK2 V617F positive in 42.9%. DIPSS Low in 1 pt, Int-1 in 1pt, Int-2 in 8 pts, High in 2 pts, and unknown in 2pts. Transfusion dependence in 35.7% at baseline. Med baseline hemoglobin (hgb) was 9.0 g/dL (R: 8.3 - 12.4), platelet (plt)157 x109/L (R:143 - 520). Thirteen pts received prior therapy, with 9 pts (64.2%) on a JAK inhibitor within 3 months of study entry. Tolerability: Nine pts (64.2%) ended active treatment, due to progression of disease in 2 pts (22.2%), adverse event (AE) in 2, pt preference in 2, stem cell transplant in 1, unrelated death in 1, and comorbidity in 1. Med duration of therapy: 91 days (range 24-287). AEs regardless of attrition included: hematologic grade 3 or >: anemia in 7 pts (50%), neutropenia in 2 (14.3%), leukopenia in 1 (7.1%), and thrombocytopenia in 2 (14.3%). Non-hematologic grade 3 or >: electrolyte abnormalities in 3 pts (21.4%), infection in 2 (14.3%), edema in 1(7.1%), hypertension in 1 (7.1%), and intracranial hemorrhage in 1 (7.1%). Efficacy: Of the 5 JAK inhibitor naïve pts, 4 had stable or increasing hgb levels with therapy despite new ruxolitinib therapy. Of the 9 pts on prior JAK inhibitor (5 ruxolitinib, 4 other), 5 (55.5%) and 7 (77.7%) patients had stable or increasing hgb or plt levels, respectively. See Table #1. Four pts (28.6%) had at least 50% improvement from baseline on MPN-SAF TSS (95% CI 8.4%-58.1%). Overall treatment response: (IWG-MRT) stable disease (SD) in 10 pts (71.4%), clinical improvement (CI) in 3 (21.4%) all of which were spleen responses, and progressive disease (PD) in 1 (7.1%). Trial was halted secondary to lack of anemia response. Table 1. Patient Specific Response Patient IWG-MRT Response: Study Cycles (N): Prior therapy (within 3 mo) Hemoglobin Response*: (initial, final g/dL) Platelet Response*: (initial, final 10(9)/L) Ruxolitinib 1 CI-Spleen 5 D (8.8, 7.6) S (221,212) 2 SD 4 D (10.3,7.9) S (54,46) 3 SD 2 S (8.8,8.9) S (82,101) 4 SD 3 I (8.9,9.8) S (441,458) 5 SD 3 S (9.6, 9.5) I (161, 231) JAK Inhibitor: 6 SD 5 Momelotinib S (9.0, 9.2) S (132,113) 7 SD 1 NS-018 D (8.3, 7.1) D (120,43) 8 SD 3 NS-018 D (10.1,9.0) D (138,111) 9 SD 2 LY2784544 S (8.7,9.1) S (56,21) Other: 10 CI-Spleen 9 Procrit I (10.5,11.9) S (153,198) 11 CI-Spleen 7 None I (8.9,10.2) D (338,133) 12 PD 1 Hydroxyurea S (12.4, 12.2) D (310, 82) 13 SD 1 Procrit D (10.1, 8.6) D (192, 71) 14 SD 1 Hydroxyurea S (9.1,8.9) D (346,119 *S=stable (Hgb +/- 0.5g/dL, Plt +/- 50x109, D=decreasing, I=increasing. Conclusions: Although overall well tolerated, the addition of danazol to ruxolitinib therapy has modest incremental efficacy by IWG-MRT criteria. Clinical improvement was observed in 21.4% of treated patients, however responses were limited to spleen reduction. The degree to which danazol may stabilize expected cytopenias is interesting and may require further investigation. Disclosures Hoffman: Promedior: Research Funding; Geron: Consultancy, Membership on an entity's Board of Directors or advisory committees; All Cells, LLC: Consultancy, Membership on an entity's Board of Directors or advisory committees. Mesa:Gilead: Research Funding; Genentech: Research Funding; Novartis Pharmaceuticals Corporation: Consultancy; CTI Biopharma: Research Funding; Pfizer: Research Funding; Promedior: Research Funding; NS Pharma: Research Funding; Incyte Corporation: Research Funding.

Abstract Abstract 2850 Myelofibrosis (MF), both primary myelofibrosis (PMF) and post essential thrombocytosis/polycythemia vera myelofibrosis (post ET/PV-MF), are chronic myeloproliferative neoplasms characterized by a progressive clinical course that leads to shortened survival. The heterogeneous nature of MF lends itself to a variable clinical course that commonly includes hepatosplenomegaly, constitutional symptoms, and progressive cytopenias. Medical therapy for myelofibrosis has been effective in palliation of common symptoms related to anemia, splenomegaly but allogeneic stem cell transplantation (ASCT) remains the only potentially curative therapeutic modality. The timing of ASCT, choice of conditioning regimen, patient selection and the impact of co-morbidities are critical to optimal use of this modality. We have retrospectively analyzed our experience for patients undergoing ASCT for primary and post ET/PV myelofibrosis at Mayo Clinic across all 3 sites: Scottsdale Arizona; Rochester, Minnesota; and Jacksonville, Florida. Methods: We conducted a retrospective review of all patients from 1992 to 2012 with PMF or post ET/PV-MF who underwent an ASCT at Mayo Clinic. This retrospective review included a waiver of informed consent and was approved by the Mayo Clinic Institutional Review Board. Patients that transformed to the MPN blast phase and individuals who underwent a second transplant for either relapse or graft failure were also included. Overall survival was estimated using Kaplan-Meier. Associations between prognostic factors and overall survival were assessed using Cox regression. Results: Baseline Patient Characteristics: Forty-eight patients (29 male) with a median age of 57 yrs (range 31–73) underwent ASCT. DIPSS-plus risk score was low in 2, intermediate 1 in 2, intermediate 2 in 9 and high in 35 patients. Disease type was PMF in 24, PV-MF in 10, ET-MF in 10, and MPN/MDS overlap in 4 pts. Eight patients evolved to the MPN-blast phase prior to transplant. JAK-2 mutation was positive in 27 patients and negative in 12 patients with JAK-2 status unknown in 9 patients. The median time from PV/ET to MF was 136 months (83–189 months) and from MF diagnosis to transplant was 59 months (3–144 months). All but 2 patients were red cell transfusion dependent prior to transplant (96%). Eight patients underwent splenectomy prior to transplant. Transplantation Characteristics: Patients who underwent ASCT received either myeloablative (MA, 13 pts) or reduced-intensity conditioning (RIC, 35 pts) regimen. The graft was from an unrelated donor in 22 pts and related donor in 26 pts with 44 being matched and 4 being mismatched. The myeloablative regimens included Bu/Cy, TBI/Cy, and Bu/Flu. The RIC regimens included flu/mel, Bu/Flu, FBM, and TBI/Flu. GVHD prophylaxis was with tacrolimus/MTX, CSA/MTX, and tacrolimus/MMF. Nineteen patients received ATG. Infectious disease prophylaxis, CMV monitoring, and additional supportive care measures were according to institutional guidelines. A majority of patients received peripheral blood stem cells with only 1 patient receiving bone marrow as the source of stem cells. Outcomes: The median time to neutrophil engraftment was 17.2 days. Two patients received a second transplant for relapsed disease. The incidence of graft failure was 10%. Graft versus host disease both acute and chronic occurred in 33 patients and 15 patients respectively. Day 100 overall survival was 92% and 2 year survival was 62% (95% CI 45–75%). There were no statistically significant associations between individual prognostic factors (DIPSS-plus classification, age, and HLA match) and survival. Conclusions: The Mayo Clinic national experience of ASCT for MF across our geographically diverse Cancer Center is encouraging. Our centers have transplanted 48 patients over the past 10 years with very favorable outcomes. Our results from a multi center practice are consistent with regards to survival, graft versus host disease, and types of transplants being performed compared to currently available published data from large single location transplant centers. This data helps confirm the prevailing knowledge that ASCT is a useful treatment. Disclosures: Mesa: Incyte: Research Funding; Lilly: Research Funding; Sanofi: Research Funding; NS Pharma: Research Funding; YM Bioscience: Research Funding.

Abstract Background: Donor cell chimerisms are tested at frequent intervals after allogeneic hematopoietic stem cell transplantation (alloHSCT). However, the predictive value of this testing in patients with myelofibrosis (MF) is not well-defined. In this study, our primary objective is to evaluate the correlation of day 30 and day 100 donor cell chimerism and overall survival (OS) in patients (pts) who undergo alloHSCT for MF. Methods: Charts were reviewed for pts who underwent alloHSCT for MF at Mayo Clinic in Arizona, Minnesota and Florida between January 2005 and December 2015. Patients who died within 30 days of alloHSCT were excluded. Chimerism studies were evaluated in the peripheral blood for CD3 and CD33 fractions at day +30 (D30) and day +100 (D100). Cut-off value of 100% and less than 100% was used because of limited variability below 100%. Standard statistical methods were used to compare continuous and categorical variables. The Kaplan-Meier method was used to provide survival curves. Results: Baseline characteristics: Sixty three consecutive patients who underwent alloHSCT for MF since January 2005 until December 2015, and had D30 and/or D100 chimerism studies available were included in the study. Of these, 24 (38%) were female. Median age at diagnosis was 56 years (range, 19 - 73 years) and median age at AHSCT was 58 years (range, 19 - 73 years). Dynamic International Prognostic Scoring System-Plus (DIPSS-Plus) risk category at the time of alloHSCT was high in 31 (49%), intermediate-2 in 30 (48%) and intermediate-1 in 2 (3%) pts. JAK2 V617F mutation status was available in 61 pts and was positive in 35 (57%) pts. Nineteen (30%) pts had been treated with a JAK2 inhibitor prior to alloHSCT. Fifty-two (83%) pts had at least grade-2 fibrosis in bone marrow at the time of HCT. Transplantation: Median time from diagnosis to AHSCT was 12.9 months (range, 2.3 - 248.2 months). Fifty-five (87%) received a reduced intensity conditioning (RIC) regimen while 8 (13%) received myeloablative conditioning regimen. Of these 55 pts, RIC regimens used were busulfan fludarabine based regimen in 15 (27%), fludarabine melphalan based regimen in 18 (33%) and fludarabine melphalan carmustine in 22 (40%) pts. Donor source was matched related in 30 (48 %), matched unrelated in 28 (44%), single-antigen mismatched related in 4 (6%) and mismatched unrelated in 1 (2%). Peripheral blood stem cells were used in all AHSCTs. Outcomes: Median follow-up was 24.4 months (range, 1.5 - 83.6 months). Amongst pts who had <100% donor chimerism at D30 in CD33 fraction, 2 had 90%, 1 had 80% and 2 had zero% donor chimerism. At D100, 1 pt had 95%, 1 had 90%, 1 had 60%, 2 had 5% and 1 had zero% donor chimerism. Eighteen out of 57 (32%) pts who had 100% donor chimerism in CD33 fraction at D30 had died at the time of analysis compared to 4 out of 5 (80%) patients with less than 100% (p=0.03). Eight out of 38 pts (21%) who had 100% donor chimerism in CD33 fraction at D100 died versus 3 out of 6 (50%) with less than 100% (p=0.12). There was no significant survival effect of CD3 chimerism (p values=0.68, 0.59). Of the 23 deaths that had occurred at the time of analysis, cause of death was disease progression or relapse in 8 (35%), graft versus host disease in 6 (26%), transplant related (sepsis / engraftment failure) in 5 (22%) and unrelated causes in 4 (17%). Survival analysis: As depicted in the Kaplan Meier curve (figure 1), median overall survival (OS) for pts with 100% donor chimerism in CD33 fraction at day 30 was not reached while for <100% donor chimerism was 16 months (p=0.04). Survival at 3 years for patients with 100% CD33 donor chimerism at D30 was 65% while it was 40% for pts with less than 100%. Also, survival at 3 years for patients with 100% CD33 donor chimerism at day 100 was 75% compared to 50% for patients with less than 100%. Conclusions: In our study, 100% donor chimerism in CD33 fraction is associated with improved survival after alloHSCT for MF compared to pts with less than 100%. Further understanding regarding the reason for this relationship is critical. Larger studies to replicate these results would be imperative to validate these findings. Disclosures Mesa: Incyte: Research Funding; Ariad: Consultancy; Promedior: Research Funding; Novartis: Consultancy; Celgene: Research Funding; Gilead: Research Funding; CTI Biopharma: Research Funding; Galena: Consultancy.

Abstract Background: Myeloproliferative neoplasms (MPN) including polycythemia vera (PV), essential thrombocythemia (ET), and myelofibrosis (MF) are clonal hemopathies characterized by burdensome symptom profiles and impaired quality of life. Few studies have evaluated patient-reported outcomes during treatment with non-experimental pharmacological regimens. Aims: The Myeloproliferative Neoplasm Quality of Life (MPN-QOL) Study Group aims to objectively quantify MPN symptom severity, frequency and quality of life at baseline and throughout treatment with non-experimental therapies utilizing the Myeloproliferative Neoplasm Symptom Assessment Form - Total Symptom Score (MPN-SAF TSS; JCO 2012). In this abstract, we provide updated results for the prospective international cohort trial currently in active enrollment: the MPN Experimental Assessment of Symptoms by Utilizing Repetitive Evaluation (MEASURE) trial. Methods: This study aims to recruit 180 international ET, PV, and MF (including primary MF and post-ET or post-PV MF) patients receiving non-experimental medical therapy and/or phlebotomy. Patients complete the MPN-SAF for seven consecutive days at enrollment and repeat the survey for an additional seven consecutive days between 90 days and six months. Patients also complete the European Organisation for Research and Treatment of Cancer (EORTC) and M.D. Anderson Symptom Inventory (MDASI) instruments at enrollment and on the first day of the second assessment. At visits, physicians acquired demographic, laboratory, physical examination, and radiographic data. Descriptive statistics were used to summarize data. Results: Clinical Data The MEASURE trial opened for enrollment in 2012 and remains in recruitment phase with 15 participating international sites. To date, 39 patients have been enrolled and 25 have completed both study visits. Participants include ET (28%), PV (24%), and MF (48%; 50% primary MF, 8% post-ET, 42% post-PV) patients. The majority of patients are male (64%) and of expected age (mean 69.3, range 39-89) for the disorders. Seventeen percent had prior thrombosis, 9% required red blood cell transfusion, and none reported prior splenectomy or hemorrhage. Mean hematologic measures included hemoglobin 13.2 g/dL, WBC count 11.4 x109/L, ANC 8.5 x109/L, and platelets 514 x109/L. Therapies received prior to enrollment included aspirin (n=16), hydroxyurea (n=11), phlebotomy (n=8), warfarin/clopidogrel/anticoagulation (n=8), erythropoietin (n=2), and interferon (n=1). The most common current MPN therapies were hydroxyurea (n=9), aspirin (n=9), interferon (n=4), and phlebotomy (n=2). Symptom Assessment In comparing MPN-SAF TSS mean symptom scores, all symptoms except bony pain improved between the first and second visits, including fatigue, early satiety, abdominal discomfort, inactivity, concentration, night sweats, itching, fever, weight loss, and overall quality of life (Figure1). Total MPN-SAF TSS scores improved from a mean of 32.3 to 25.9. On the EORTC, mean scores for physical, role, emotional, and social functioning improved from the first to the second visit (Figure 2). Cognitive functioning showed a slight decline. Global health status measure improved from 60.2 to 72.9. On the MDASI, symptom severity scores decreased from 3.6 to 2.8 from the first to second visit (Figure 3). Symptom distress measure decreased from 4.1 to 3.0. Discussion: Interim results from the MEASURE trial demonstrate that standard, non-experimental treatment regimens offer improvement in quality of life-related symptoms on multiple patient-reported survey instruments including the MPN-SAF TSS, EORTC QLQ-C30, and MDASI. Updated data including symptom correlations and mutational analysis to be presented at the 2016 ASH conference. Disclosures Ross: Novartis Pharmaceuticals: Honoraria, Research Funding; BMS: Honoraria. Radia:Novartis: Honoraria; Pfizer: Honoraria. McMullin:Novartis: Honoraria, Speakers Bureau. Cargo:Novartis: Honoraria; Celgene: Honoraria, Research Funding. Sekhar:Novartis: Research Funding. Mesa:Gilead: Research Funding; CTI Biopharma: Research Funding; Galena: Consultancy; Ariad: Consultancy; Incyte: Research Funding; Novartis: Consultancy; Celgene: Research Funding; Promedior: Research Funding.

Abstract Background: Myelofibrosis (MF) is a myeloproliferative neoplasm that is characterized by significant scar tissue and fibrosis in the bone marrow, enlarged spleen and/or liver from extramedullary hematopoiesis, and may include significant constitutional symptoms such as bone pain, night sweats, pruritis, and cachexia. The only curative therapy is allogeneic stem cell transplant. Although the symptom burden has been explored in the literature, the impact of hematopoietic stem cell transplant (HCST) on QoL in patients with MF has not been evaluated. We sought to longitudinally describe QoL in patients undergoing HCST for MF. Methods: We prospectively followed patients undergoing HSCT for MF. We assessed symptoms, functioning, and QoL using the FACT-BMT and MPN-SAF total symptom score (TSS) pre-transplant and at day 30, day 100 and one year post-transplant. Scores at the post-transplant time points were compared with baseline scores by paired t-tests. Pearson correlations between FACT-BMT and MPN-SAF TSS questionnaires were also computed. Results: 16 patients were enrolled [median age 64.0 (49-69) years; 13 (81%) male; 13 (81%) Caucasian], two did not have day 30 data as they died prior to then or did not go to transplant. Fourteen patients had day 30 information, 11 had day 100 information, and only 4 had one year information. Of the 14 who had day 30 information, 6 patients died within the first year, two from treatment related mortality and four from relapse. One patient had intermediate-1 risk, the remainder of the patients were intermediate-2 or high risk. All patients had RIC conditioning. Mean MPN-SAF TSS score was 28.1 (SD=14.2) and FACT-BMT total score was 99.8 (SD=17.4) at baseline. FACT-BMT and MPN-SAF TSS at baseline were inversely correlated; lower symptom score was associated with higher QoL (r=-0.62; p=0.01). FACT-BMT at day 30 was lower (mean change: -12.5, SD=16.7; p=0.03). Two MF-specific symptoms showed improvement that reached statistical significance compared to baseline: night sweats mean improvement day 30, 2.5 (SD=3.1; p=0.01) and mean improvement day 100, 1.7 (SD=2.6; p=0.05, Figure 1); headache mean improvement day 100, 1.5 (SD=1.9) p=0.02. In general, scores showed a worsening at day 30, improvement at day 100 and stability at one year. The MPN-SAF TSS worsened at day 30 (6 points) and improved by day 100 (4.5 points). Changes that showed improvement at day 100 include Brief Fatigue Inventory (BFI) with a mean improvement of 1.2 points and concentration (1 point). Of the four surveys that were collected at one year, a modest decline was noted in BFI (1.5 points), inactivity (1.5 points) and cough (3 points). However improvements were noted in night sweats (2.25 points), abdominal discomfort (1 point), insomnia (1.75 points), bone pain (1 point). Discussion: This is the first study to evaluate serially the QoL and symptom burden of patients who underwent a transplant for MF. A decline in QoL in the first 30 days was observed, with modest improvement at day 100. Few surveys have been completed at 1 year to date in this ongoing study. Collection of surveys past one year may be more informative regarding long-term impact of transplant on quality of life. Figure 1 Figure 1. Disclosures Mesa: Gilead: Research Funding; Novartis: Consultancy; Ariad: Consultancy; CTI Biopharma: Research Funding; Galena: Consultancy; Celgene: Research Funding; Promedior: Research Funding; Incyte: Research Funding.

Abstract Hypomethylating agents have changed treatment and outcomes in MDS and are active in AML patients. However a significant number of patients do not respond to 5-Azacytidine or Decitabine and there is no clinically validated assay that predicts response to 5-Azacytidine (5-Aza) or Decitabine (DAC). Clinical parameters and recently TET2 mutations have been identified as genetic predictors of response to 5-Aza. Several recent papers report on the possible predictive value of anti-apoptotic proteins, for example BCL2L10. In previous genome-scale RNA-interference (RNAi) screens anti-apoptotic BCL-2 family members, most potently BCL-XL, were identified as top targets whose down-regulation sensitize to 5-Aza. Results were confirmed with the BCL-XL and BCL-2 inhibitor ABT-737, as well as the BCL-2-specific inhibitor ABT-199, both of which sensitized myeloid cells to 5-Aza. Although it is expected that interfering with anti-apoptotic genes would sensitize to hypomethylating similar to cytotoxics, few of the other >900 genes from our RNAi sensitizer screens, including few of the 571 kinases, sensitized to 5-Aza as potently as inhibition of BCL-XL. Thus BCL-2 family members constitute an especially potent context to modulate the activity of 5-Aza. To assess if protein expression of the top targets BCL-XL, BCL-2 or MCL-1 correlate with and may explain a preferential sensitization, 577 primary AML samples were examined by Reverse Phase Protein Array (RPPA). There was substantial overlap of expression across and within FAB subgroups and cytogenetics, and expression of neither of these genes/proteins could explain the observed effects. This may be expected due to functional redundancies amongst the different anti-apoptotic BCL-2 family members, as well as distinct, yet overlapping binding affinities between anti- and pro-apoptotic BCL-2 proteins and BH3 peptides. Thus, we next aimed to functionally interrogate the overall balance of pro- and anti-apoptotic BCL-2 family members, also described as “primedness” by BH3 profiling. This assay uses peptides derived from pro-apoptotic BH3-only proteins to determine the capacity of cells to initiate apoptosis in response to pro-apoptotic BH3 molecules [anti-apoptotic BCL-2 molecules like BCL-XL and BCL-2 bind and inhibit BH3-only molecules]. This assay is a broad functional readout that incorporates many parameters including the consequences of varying BCL2L10 expression. To establish a proof-of-concept that the functional interrogation of specific apoptotic thresholds may be an optimal readout for 5-Aza response, we first assessed a broad panel of 13 AML-derived cell lines by BH3 profiling and in parallel 5-Aza drug dose response experiments. Several BH3 profiling metrics, including NOXA plus BIM, significantly correlated with 5-Aza sensitivity in vitro (Figure 1). Next, to correlate actual clinical responses to 5-Aza with BH3 profiling, specimens from 28 AML, MDS and MDS/MPN overlap patients treated with 5-Aza or DAC-based regimens and for whom clinical outcome was available, were assayed in BH3 assays. In the clinic, the best BH3 metrics were combined values from NOXA and BIM peptides similar to cell lines. NOXA plus BIM discriminated clinical responses defined as achieving any response (sensitive) versus a patient being resistance/refractory to 5-Aza/DAC based regimens with statistical significance (Mann-Whitney two-tailed p = 0.001). This was true for the entire group of patients (5-Aza- and DAC- regimens) with a Receiver Operating Characteristic (ROC) of an AUC=0.875, with a further increase for patients treated only with 5-Aza based regimens with an AUC=0.95.Figure 1BH3 metrics distinguish cell lines with higher EC 50 (> 2uM) from those with lower EC50 (generally < 1uM).Figure 1. BH3 metrics distinguish cell lines with higher EC 50 (> 2uM) from those with lower EC50 (generally < 1uM). In conclusion, due to significant expression overlap and functional redundancies of the BCL-2 family proteins, expression does not correlate with 5-Aza clinical or in vitro responses. However, BH3 profiling as a general functional readout of apoptotic primedness, significantly discriminated responses in patients as well as in vitro. Thus, BH3 profiling incorporates the entirety of pro- and anti-apoptotic molecules (including BCL2L10) and is promising to predict responses to hypomethylating agents. We propose to prospectively validate BH3 profiling as a predictive biomarker assay for 5-Aza or DAC based regimens. Disclosures: Pierceall: Eutropics: Employment, Equity Ownership. Lena:Eutropics: Employment, Equity Ownership. Cardone:Eutropics: Employment, Equity Ownership. Elashoff:Eutropics: Employment. Blake:Noel Blake: Employment, Equity Ownership.

Abstract Introduction Myelofibrosis (MF) and Post PV/ET MF represents a group of debilitating hematological disorders in which quality of life (QOL) is severely compromised in most patients as a result of persistent constitutional symptoms, progressive cytopenias, and splenomegaly. Conventional therapeutic modalities have largely focused on symptoms palliation rather than cure. Allogeneic stem cell transplantion (ASCT) remains the only potential curative option for intermediate and high risk disease. Till now, outcome data for MF have historically focused on survival benefit. Until now, few studies have evaluated the QOL, financial burden, and symptom response in MF patients undergoing treatment including ASCT. The development of the Myeloproliferative Symptom Assessment Form Total Symptom (MPN-SAF TSS) score in 2012 allows us to objectively quantify with a standardized tool these crucial aspects of patients care. The Myeloproliferative Neoplasm Quality of Life (MPN-QOL) Study Group aims to objectively quantify symptomatic response to standard available treatments by utilizing the MPN-SAF TSS. We introduce two MPN-QOL prospective trials currently in active enrollment: The MPN Experimental Assessment of Symptoms by Utilizing Repetitive Evaluation (MEASURE) Trial and the Symptoms Yielded in Myelofibrosis Patients after Transplant as Objectified by MPN-SAF TSS (SYMPTOMS) Trial. Our intention with these studies is to quantify the QOL and symptom burden of PMF and post PV/ET MF patients undergoing treatment including ASCT. Methods The MEASURES trial is a prospective questionnaire based study evaluating the responsiveness of the MPN-SAF TSS in detecting symptomatic changes in target symptoms for an anticipated 180 ET, PV and MF (including primary MF, post-ET and post PV MF) patients receiving non-experimental medical therapy (aspirin, hydroxyurea, anagrelide, interferon, busulfan, melphalan, cladribine, thalidomide, lenalidomide, prednisone, danazol, ruxolitinib) and/or phlebotomy. Patients complete the MPN-SAF for seven consecutive days at the time of enrollment and repeat the survey for an additional seven consecutive days between 90 days and six months. Patients also complete the MDASI, EORTC and Global Impression of Change Items on the first day of the second assessment. Physicians acquire demographic, laboratory, physical examination and radiographic data, along with serial response assessments. In parallel, the SYMPTOMS trial is prospective questionnaire based study evaluating the QOL of patients undergoing ASCT utilizing MPN-SAF TSS, the FACT-BMT, Global Impression of Change, and a financial questionnaire. Patients will be evaluated at various time points pre-transplant, day 30, day 100 and 1 year post-transplant. Participants (N=110) will be prospectively enrolled from Mayo Clinic Arizona (MCA) with other centers to soon join. To date we have enrolled 5 patients from MCA on the SYMPTOMS trial and 32 patients on the MEASURES trial. Results Both trials began open enrollment in the summer of 2012 and remain in recruitment phase. Our updated preliminary data will be presented at the ASH 2013 meeting Conclusion Myeloproliferative Neoplasms have been associated with debilitating symptom profile that can significantly impair the QOL. We recognize the burden of this disease and treatment and have therefore initiated two ongoing parallel prospective trials with goals to quantify the QOL and symptom burden of MPN patients. By quantifying degree of burden and impairment in QOL in a standardized format in MPN patients undergoing a range of treatments, we will in the future be better able to inform our patients of the likely benefits and toxicities of the available treatment options. Disclosures: Birgegard: Vifor Pharma: Honoraria.

Abstract Abstract 1954 Introduction: Vitamin D (VD) deficiency can cause osteomalacia, aching bone pain, muscle weakness, fatigue, increase risk of fracture and precipitate or exacerbate osteopenia/osteoporosis. Allogeneic hematopoietic stem cell transplant (HCT) patients are susceptible to low VD level secondary to poor oral intake, decreased exposure to sunlight and treatment related malabsorption. VD level has been correlated to cancer incidence and VD metabolites have been used in the treatment of myeloid leukemia. VD shows promising immunomodulatory properties and correction of low VD level may mitigate manifestations of graft versus host disease (GVHD). Reports have suggested that low VD level appears to increase the incidence of GVHD. We hypothesize that there is a relationship between low VD level and morbidity (specifically incidence of acute GVHD), mortality and relapse incidence after HCT. We therefore studied VD levels pre- and post-HCT to determine if VD level impacts these outcomes. Patients and Methods: 241 patients underwent myeloblative or non-myeloablative HCT between January 1, 2009 and January 31, 2011 at our institution. Baseline demographic data, disease characteristics, transplant variables and outcomes data were obtained from the transplant database. These data were supplemented by retrospective chart review for pre- and post-transplant VD level. Categories for VD level included normal (>30 ng/ml) or abnormal (<30 ng/ml). Results: 131 (54%) of patients who underwent myeloblative or non-myeloablative HCT had their VD level evaluated either pre-transplant, post-transplant or both. Pre-HCT 57 (56%) patients, 100 days post-HCT 55 (59%) patients and 365 days post-HCT 12 (32%) patients had a low VD level. 51 (38%) of patients had a reduced intensity HCT and 80 patients (62%) had an ablative conditioning regimen. 11 (8.4%) patients had acute GVHD. Comparison of VD level among those with acute GVHD did show a higher incidence of acute GVHD between those with a low or normal VD level (HR=3.14, 95% CI: 0.35–28.33) however this association was not statistically significant (p=0.3079). Survival analysis in those with a low VD level pre-HCT showed there was not a higher risk of mortality (HR=1.14, 95% CI: 0.18–7.38) after adjusting for post- VD level, and this association was not statistically significant (p=0.8921). Survival analysis in those with a low VD level post-HCT did show a higher risk of mortality (HR=2.59, 95% CI: 0.26–25) after adjusting for pre-HCT VD level, however, this association was not statistically significant (p=0.4155). The relationship between VD level and relapse at 1 year post-HCT was not able to be examined because the VD level for patients who relapsed was not available. Conclusions: Half of patients undergoing HCT had VD testing pre- or post- HCT. Just over half of the patients tested had a low VD level pre- and 100 days post-HCT. It is notable that 365 days post HCT the number of patients with low VD level had decreased. This could be attributable to less time in the hospital thus increasing sun exposure, increasing performance status allowing better ingestion and absorption of VD in the gastrointestinal tract or proper supplementation of a low level noted previously. There was no significant difference in incidence of acute GVHD by VD level though there was a trend for increased risk. Pre- and post-HCT VD level did not significantly impact mortality but there was a trend toward higher risk in those with a low VD level post-HCT. This is the first study, to our knowledge, to evaluate the impact of VD level on mortality post-HCT. Our study confirms that many patients have a low VD level pre-and post-HCT. These findings, especially the trend towards a higher mortality risk and higher incidence of acute GVHD in those who have a low VD level post-HCT, warrant further prospective investigation. VD supplementation may be a low cost, easy to implement addition to routine post HCT care that might reduce HCT associated mortality. Disclosures: Reeder: Celgene: Research Funding.