ALBERT

Description: Introduction: MDS encompasses a heterogeneous group of clonal neoplastic hematopoietic stem cell diseases characterized by ineffective hematopoiesis and frequent progression to acute myeloid leukemia (AML). While the advent of hypomethylating agent (HMA) therapy has modestly improved survival in higher-risk MDS, outcomes following HMA failure are dismal. The hedgehog (HH) signaling pathway is an important regulator of hematopoietic cell survival and differentiation, with upregulation of HH target genes and signaling proteins a frequent occurrence in MDS and AML. PF-04449913 (PF-04) is an orally bioavailable inhibitor of Smoothened (SMO), which demonstrated antileukemic activity in a prior phase 1 study. We performed a phase 2 study of PF-004449913 in patients with MDS following HMA failure. Objectives: 1) primary: to estimate overall IWG-2006 response rate (CR + PR + HI + marrow CR) to PF-04449913 in patients with relapsed/refractory MDS and CMML; 2) secondary: to assess safety , overall survival, time to AML transformation, and effect of PF-04 on HH target gene expression. Methods: This was a single center, open-label phase 2 study. Key eligibility criteria included: 1) patients age ≥ 18 with MDS, CMML, or AML (20-29% blasts) following failure of HMA; 2) ECOG 0-2; 3) Adequate kidney and hepatic function. Treatment consisted of PF-04 at 100 mg/day in 4-week cycles for 4 cycles, with continuation allowed for achievement of stable disease or better. Dose increase to 200 mg/day was permitted after cycle 2 for patients with stable disease. Bone marrow biopsy for response assessment occurred after cycle 2, cycle 4, and every 4th cycle thereafter. For correlative studies, total cellular RNA was isolated from ficolled mononuclear cells in baseline marrow samples, and qPCR performed for expression of GLI1, PTCH1, and SMO). Results: Thirty five patients were enrolled, 24M and 11F. Median age was 75 years (range 55-88). A majority of patients had higher-risk (54%) vs lower-risk (43%) MDS by IPSS at time of enrollment. A total of thirty four patients (97%) had received prior azacitidine for a median of 9 cycles (range 1-88), while 6 patients had received decitabine for a median of 4 cycles (5 of whom had also received azacitidine). The median number of cycles of PF-04 received was 3 (range 1-11). All 35 patients were evaluable for response. Two of 35 patients (6%) achieved response per MDS IWG 2006 criteria, including one patient with HI-N/HI-P and another with marrow CR and HI-N. Nineteen additional patients (54%) had stable disease following cycle 2 (8 weeks). With a median follow-up of 10.1 months, the median survival for all treated patients was 10.2 months (95% CI 6.8-13.6 mo)(Figure 1). Median survival for patients with low/INT-1 risk MDS was longer than for INT-2/high risk patients (22.4 mo vs 7.5 mo, p=0.013). Six patients (17%) died while on-treatment, but none of the deaths were considered related to PF-04. The most common treatment-emergent adverse events included myalgia/muscle cramps (65%), dysgeusia (60%), nausea (31%), anorexia (31%), oral mucositis (25%), and AST elevation (22%). The vast majority of these events were grade 1/2. Except for infection (11%), there were no treatment-emergent grade 3/4 events in 〉 10% of patients. Only 5 patients (14%) required dose reduction to 50 mg/day due to intolerance. In the pharmacodynamics analysis, responding patients had higher baseline expression of SMO, PTC ,and GLI1 than non-responders, while non-responders had higher post-treatment increases in these transcripts compared with responders. Conclusions: PF-04 was well tolerated in patients with advanced, HMA refractory MDS, but had limited efficacy in this unselected, heavily pre-treated patient population. The pharmacodynamic analysis suggests predictive potential of HH pathway gene expression for response warranting further investigation. Figure 1 Figure 1. Disclosures Komrokji: Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Consultancy, Speakers Bureau; Incyte: Consultancy. Sweet:Novartis: Consultancy, Speakers Bureau; Ariad: Consultancy, Speakers Bureau; Pfizer: Speakers Bureau; Incyte Corporation: Research Funding; Karyopharm: Honoraria, Research Funding.

Description: Background: We reported that ineffective hematopoiesis in MDS results from caspase-1-dependent, pyroptotic cell death. Pyroptosis, which culminates in cytolysis, is initiated by the NOD-like receptor NLRP3, a redox-sensitive, cytosolic sensor of danger signals. Upon activation, NLRP3 recruits the ASC (apoptosis-associated speck-like protein containing a CARD (caspase activation and recruitment domain)) adapter protein that polymerizes to create large cytoplasmic filaments. Exposure of the ASCCARDdomain on the filament surface fosters docking of pro-caspase-1 that initiates processing of pro-IL1β and -IL18. Large cytoplasmic aggregates formed by filament clusters are known as ASC specks. Upon cytolysis, ASC specks are released into the extracellular space where they retain catalytic activity and propagate inflammation. Given the magnitude of pyroptosis in the bone marrow (BM) and maturing cells, we hypothesized that detection of ASC specks in peripheral blood (PB) plasma may serve as a biologically-rational MDS biomarker. Methods: We optimized a flow cytometry assay for detection of extracellular ASC specks averaging 1µm in size. Following protein quantitation, 300µg was aliquoted from each donor and stained with rabbit-anti-ASC 1o-Ab at a 1:1500 dilution for 60' at 37°C. 1:1500 dilution 2o-Ab was added and incubated for 30' at 37°C. Sample acquisitions were made on a BD FACSCalibur flow cytometer. PB plasma samples were obtained from normal donors (ND, n=40) and patients with MDS (n=220), de novo AML (n=16), 2o AML (n=26), CMML (n=20), CLL (n=50), CML (n=52), ALL (n=7), ET (n=20), PV (n=20), myeloma (n=20) and patients with Type-2 diabetes (T2D, n=25) on IRB-approved protocols, providing 〉90% power to detect significant differences between diseases. The diagnostic biomarker is defined as % of PB plasma ASC specks adjusted for glucose. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) were used to evaluate classification accuracy of the biomarker at varied cutoff points. Results: The % PB-ASC specks was significantly increased in lower-risk (LR) MDS vs. ND (n=196, 27.7±1.5 vs. 7.2±0.7, respectively; p=2.8x10-32). No significant difference was observed between ND and higher-risk (HR) MDS (12.4±3.0), however, % of PB-ASC specks was significantly increased in LR (n=196) vs. HR (n=16) MDS (p= 2.0x10-5). Because hyperglycemia stimulates NLRP3 inflammasome assembly and activation, we measured plasma glucose by colorimetric assay to adjust for this potentially confounding variable. % PB-ASC specks were normalized to plasma glucose concentration and corrected for volume. Glucose-adjusted speck % in LR-MDS remained significantly higher vs. ND (1.2±0.2 and 0.02±3.0x10-3, respectively; p=3.7x10-6). Notably, the corrected % PB specks was not significantly greater in HR-MDS (0.6±0.5) vs. ND or between LR-MDS vs. HR-MDS, although the HR-MDS sample size is small. Importantly, LR-MDS samples (1.2±0.2) displayed significantly greater corrected % ASC specks compared to de novo AML (0.3±0.2, p=2.3x10-3), 2o-AML (0.16±0.05, p=5.5x10-5), CMML (0.03±0.01; p=4.4x10-6), CLL (0.03±4.4x10-3, p=4.4x10-6), CML (0.04±0.01; p=5.2x10-6), ALL (0.2±0.09, p=9.9x10-5), ET (0.17±0.07, p=8.7x10-5), PV (0.12±0.06, p=3.8x10-5), myeloma (0.14±0.04, p=3.8x10-5) and PB from non-cancer patients with T2D (p=4.8x10-6), suggesting specificity for MDS (see Figure). A cut off of 0.053 was selected to minimize total misclassification error (false positive + false negative). With this cutoff, the biomarker achieves 99% sensitivity and 81% specificity in classifying MDS from ND. The corresponding PPV and NPV are both 95%. In a preliminary cohort of lenalidomide-treated LR-MDS patients, PB-ASC specks decreased a mean of 48% (range, 3-69%) at week 16, suggesting that specks may serve as a biomarker index of ineffective hematopoiesis. Conclusion: ASC specks are readily quantified by flow cytometry and profoundly increased in PB plasma of MDS patients compared to ND and other hematologic malignancies. ASC specks are a sensitive and specific diagnostic biomarker for MDS that may also serve as an index of disease activity and emerging treatment response. Disclosures Pinilla-Ibarz: Novartis: Consultancy; Janssen: Consultancy, Honoraria; Abbvie: Consultancy, Speakers Bureau; Pharmacyclics: Consultancy, Speakers Bureau; Novartis: Consultancy; Gilead: Consultancy, Speakers Bureau; Gilead: Consultancy, Speakers Bureau. Komrokji:Novartis: Consultancy, Speakers Bureau; Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding.

Description: BACKGROUND: Venous thromboembolism is a well-known complication of malignancy. The Khorana Risk Score (KRS) utilizes the type of cancer, blood counts, and body mass index to predict risk of thrombosis in cancer patients. Patients with a KRS of greater than 3 are considered high risk for developing blood clots and may benefit from prophylactic anticoagulation. We wanted to determine if the use of the KRS would have led to prophylactic anticoagulation (AC) in these cancer patients prior to their thrombotic events. METHODS: This study utilized a group of 286 patients treated at Moffitt Cancer Center between May 1, 2010 and June 30, 2015 with a wide variety of cancer diagnoses with known venous thromboembolic events (VTE) (pulmonary embolism or deep venous thrombosis). We aimed to validate the KRS by retrospective analysis to determine if the KRS would have predicted VTE in these patients on the day of their venous thromboembolic event. The demographics are summarized using descriptive statistics. Wilcoxon rank-sum test is used for testing the difference in continuous variables. Fisher exact test is used for testing the difference in categorical variables. RESULTS: We found that the majority of these cancer patients with known VTE (89%) would have been classified as low risk of having VTE. Only 11% would have been classified as high risk to where prophylactic anticoagulation might have been indicated. CONCLUSIONS: We found that in a diverse group of cancer patients who had thrombotic events, the KRS did not appropriately stratify patients at the highest risk of VTE. The majority of patients with VTE were actually found to be low risk which would not have led to improved awareness or medication prophylaxis. Improved risk stratification methods are needed for cancer patients to implement an effective prophylactic strategy. Table Table. Disclosures No relevant conflicts of interest to declare.

Description: Introduction: Fludarabine (Flu)-based conditioning regimens, combined with either melphalan (Mel) or intravenous busulfan (Bu), are often utilized in patients undergoing allogeneic hematopoietic cell transplantation (HCT) for acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS). While recent data supports that a myeloablative conditioning (MAC) strategy improves survival in patients who are able to tolerate this approach, the optimal regimen for patients elderly or unfit for MAC remains unclear. A single-center retrospective analysis was performed to evaluate outcomes in patients with AML or MDS deemed eligible only for reduced intensity regimens. Methods: Patients with AML and MDS who underwent peripheral blood mobilized allogeneic HCT at a single institution between January 2008 and December 2014 were identified. Of those, patients who received conditioning with either once-daily Bu (75-95 mg/m2) targeted to an area under the curve of 3500 µM*L/min and Flu 160 mg/m2 (Flu/Bu 3500) or Mel 140 mg/m2 plus Flu 120-160 mg/m2 (Flu/Mel) were evaluated and compared. All disease statuses (complete remission (CR), second CR, or with active disease) at time of HCT were included. Regimen toxicities, cumulative incidences of acute and chronic graft-versus-host disease (GVHD), non-relapse mortality (NRM), cumulative incidence of relapse (CIR), progression free survival (PFS), and overall survival (OS) were evaluated. Results: We identified 150 consecutive patients who received Flu/Bu 3500 (n=81) or Flu/Mel (n=69). Regimens were selected at physicians' discretion. The median ages for Flu/Bu 3500 and Flu/Mel were similar (66.9 (range, 48.2-75.9) and 65.8 (40.9-75.2) (p=0.14)). No differences were detected between the two groups with regard to recipient and donor gender, primary disease, donor type, recipient and donor cytomegalovirus (CMV) status, and Karnofsky performance status. Additionally, there were no differences between either groups in disease specific prognostic variables such as Disease Risk Index (Armand et al.), blast percentage at time of HCT, International Prognostic Scoring System (IPSS) and revised IPSS scoring in MDS patients, or AML cytogenetics at time of HCT. Patients receiving Flu/Mel had higher HCT-specific comorbidity index (HCT-CI, 〉3) when compared to Flu/Bu 3500 prior to transplantation (66.7% versus 46.9%, p=0.02). OS was similar between both arms (p=0.1) as was NRM (Hazard ratio (HR) 0.61 (CI, 0.34 - 1.1, p=0.1)) and PFS, HR 1.26 (CI, 0.83 - 1.94, p=0.2). However, CIR at 2 years post-allograft was significantly higher in the Flu/Bu 3500 arm, HR 2.54 (CI, 1.4 - 4.6, p=0.002) in comparison to Flu/Mel regimen (Figure 1). There was no difference detected in the cumulative incidences of either grades 2-4 acute GVHD, HR 1.00 (CI, 0.66 - 1.55, p=1.0), or grades 3-4 acute GVHD, HR 0.66 (CI, 0.27 - 1.57, p=0.3) between either conditioning regimen. The cumulative incidence of chronic GVHD was also similar, HR 1.27 (CI, 0.84 - 1.92, p=0.2). With regard to toxicity, diarrhea occurred more frequently in the Flu/Mel arm (p=0.0003) in the first 20 days following transplant. However, in the same period, mucositis occurred more frequently in the Flu/Bu 3500 arm (p=0.005). No differences were noted between the arms when assessing incidence of sinusoidal obstructive syndrome, diffuse alveolar hemorrhage, or thrombotic microangiopathy up to 90 days. Amongst patients receiving Flu/Mel, the most common cause of death was pneumonia or pulmonary failure (n=7) whereas the most common cause of death in the Flu/Bu 3500 arm was disease related (n=31). Conclusion: Flu/Mel resulted in a lower CIR at 2 years post-HCT compared to patients receiving Flu/Bu 3500 conditioning. Regarding toxicity, Flu/Mel produced more diarrhea but significantly less mucositis in comparison to Flu/Bu 3500; otherwise toxicity was comparable. Though there were no differences in OS and NRM between the two conditioning regimens, we speculate that the impact of the higher HCT-CI in the Flu/Mel arm may have contributed negatively to the lack of benefit in NRM and OS. Disclosures Locke: Kite: Membership on an entity's Board of Directors or advisory committees. Nishihori:Signal Genetics: Research Funding; Novartis: Research Funding. Fernandez:Chimerix: Honoraria; Otsuka: Honoraria; Sanofi: Speakers Bureau.