ALBERT

Description: Bone marrow (BM) fibrosis is a key pathomorphologic feature of patients (pts) with primary myelofibrosis (PMF) and the fibrotic phases of essential thrombocythemia (post-ET MF) and polycythemia vera (post-PV MF). The degree of BM fibrosis appears to correlate with survival. Indeed worse survival has been associated with increased BM fibrosis. The BM stromal microenvironment is important in the pathogenesis of BM fibrosis. Cellular components (fibroblasts, macrophages, endothelial cells, adipocytes), structural fibrils (collagen, reticulin) and extracellular matrix components are all forming elements of the BM stroma. Increased stromal fibrosis has been linked to abnormalities in the number/ function of megakaryocytes and platelets in hematologic diseases. Several cytokines like Platelet Derived Growth Factor (PDGF) and Transforming Growth Factor-Beta (TGF-b) have been also linked to the pathophysiology of BM fibrosis. PDGF has been shown to increase fibroblast growth in megakaryocytes and platelets although increased PDGF did not correlate with increased production of either reticulin or collagenous fibrosis. Moreover, PMF pts have increased TGF-b levels in platelets, megakaryocytes, and monocytes. Nitric Oxide (NO) is a ubiquitous gas important in physiologic processes particularly vasodilatation. Dysregulation of NO levels has been implicated in pulmonary hypertension (PH), hemoglobinopathies, and cardiovascular diseases. In Peyronie’s disease, a localized fibrosis of the penile tunica albuginea, increased NO production by expression of iNOS decreases collagen deposition by neutralization of profibrotic reactive oxygen species and decreased myofibroblast formation. Aside from its role in maintaining normal vascular tone, NO also plays a role in fibroblast formation and collagen biosynthesis. We previously reported that ruxolitinib, a JAK1/2 inhibitor restores NO levels leading to improvement of PH in MF pts (Tabarroki et al., Leukemia 2014). We now hypothesize that plasma/serum NO level is a key regulator of BM fibrosis in MF and that ruxolitinib treatment (Tx) leads to improvement of BM fibrosis by NO modulation. Using a Sievers 280i NO analyzer we measured the plasma/serum NO level of a large cohort (n=75) of pts with myeloid and myeloproliferative neoplasms (MPN) [MDS, RARS/RCMD=8; MPN, ET=8, PV=8, MF=24, Mastocytosis=7; MDS/MPN, CMML=11, MDS/MPN-U, RARS-T=9]. Healthy subjects (n=10) were used as a control. MPN pts had low NO (nM) levels among the pts studied with the lowest level found in MF pts: MF=30.31±11.8, PV=39.0±16.1, ET=36±20.3, RARS=74.6±41.7 (P=.01), CMML=84.4±89.2 (P=.04), RCMD=163.4±103.8 (P

Description: Disease type and status at the time of allogeneic hematopoietic stem cell transplantation (HSCT) dominantly influence HSCT outcome. It is therefore important to stratify patients by disease risk in any retrospective or prospective transplantation study that enrolls patients across multiple disease types or status. We previously proposed a Disease Risk Index for this purpose, based on a retrospective study of patients transplanted at 2 institutions (Armand et al, Blood 2012;120:905). Here we present the results of a study designed to validate and refine the DRI in a larger multicenter population. We included 13,131 adult patients who underwent HSCT for hematologic malignancies, excluding very rare diseases, between 2008 and 2010 and were reported to the CIBMTR. Their median age was 52 (range, 18-80) years. The cohort included a broad representation of diseases, disease status, donor types, and graft sources. 53% of patients were conditioned with a myeloablative regimen. The median follow-up for survivors was 24 months. The original DRI stratified patients into 4 groups with 2y OS of 64% in the low-risk, 51% in the intermediate, 34% in the high, and 24% in very high risk group (p

Description: Although outcomes after allogeneic hematopoietic cell transplantation (alloHCT) for AML have improved, this has mainly been attributed to a reduction in transplant-related mortality rather than reduced leukemia relapse. NK cell alloreactivity is regulated by inhibitory and activating signals mediated through cell-surface receptors including the killer immunoglobulin-like receptors (KIRs). Group A and B KIR haplotypes have distinct centromeric (Cen) and telomeric (Tel) gene-content motifs and donor Cen group B KIR haplotypes have been reported to be associated with decreased relapse and improved survival in AML patients undergoing unrelated donor alloHCT. We hypothesized that donor KIR genotype may also be predictive of outcomes after matched related donor (MRD) alloHCT. We evaluated 93 AML patients in CR1/CR2 who underwent T-cell replete alloHCT using HLA- matched related donors at our institution from 1/2000-3/2013. Sixty-six had myeloablative conditioning (MAC) that that was busulfan/cyclophosphamide-based and 27 had reduced-intensity conditioning (RIC) with fludarabine/total body irradiation or busulfan/fludarabine. Donors were KIR genotyped to assign haplotypes A/A vs. B/X and the distinctive Cen and Tel gene-content motifs of group A and B KIR haplotypes according to the presence or absence of one or more B haplotype-defining KIR genes. KIR B–content score for each KIR genotype was defined as the number of Cen and Tel gene-content motifs containing B haplotype–defining genes (range, 0-4). As compared to those with haplotypes B/X (n=40; B content scores of 1-4) those with haplotype A/A (n=25; B content score of 0) undergoing MAC had significantly lower 100-day, 6-, 12- and 24-month non-relapse mortality (NRM) (8% vs. 0%, 13% vs. 0%, 15% vs. 0%, 25% vs. 0%, respectively, Figure 1) which was confirmed on multivariable analysis (HR 9.19, p=0.03). There were no differences between these groups regarding patient and transplant-related characteristics, or for acute or chronic GVHD, relapse, or survival. The causes of death in the group with haplotypes B/X were most commonly attributed to infection and then GVHD. However, within the group with B/X haplotypes, the B motif content score (1-4) was not associated with significant differences in NRM (HR 0.79, p=0.56). No difference in outcomes was observed for those undergoing RIC. The number of donor activating KIR genes (2SD1, 2DS2, 2DS3, 2DS4, 2DS5, and 3DS1) was then assessed. As compared to those with 3-6 activating KIR genes (n=20) those with 0-2 (n=41) undergoing MAC had significantly lower 100-day, 6-, 12- and 24-month non-relapse mortality (NRM) (15% vs. 0%, 15% vs. 5%, 15% vs. 5%, 29% vs. 8%, respectively, Figure 2) which was confirmed on multivariable analysis (HR 4.07, p=0.01). There were no differences in other post-transplant outcomes when comparing these groups or when considering those undergoing RIC. An increase of 1 donor activating KIR also was highly associated with NRM (HR 1.37, p=0.008). Overall, these results suggest that in the MRD MAC alloHCT setting donor KIR genotype may be predictive of increased NRM risk, particularly for those with B/X haplotypes and greater numbers donor activating KIRs. No comparable effects were observed in the RIC setting. Future strategies to further enhance immune reconstitution post-transplant may be appropriate to pursue for these higher risk patients. These results may have potential implications to improve donor selection for those AML patients with multiple HLA-matched related donors and need to be validated in larger cohorts. Figure 1 Figure 1. Figure 2 Figure 2. Disclosures No relevant conflicts of interest to declare.

Description: Background: Patients (pts) with sAML are evaluated together in large AML trials regardless of whether their sAML arises from antecedent MDS vs. from MPD vs. t-AML. Prognostic factors and outcomes may differ among these subgroups, and a prognostic scoring system would be helpful in determining which patients would benefit from induction chemotherapy. Methods: We conducted a retrospective review of all pts with newly diagnosed, pathologically-confirmed AML at Cleveland Clinic between 1997 and 2007 to identify sAML pts treated with cytarabine-based induction chemotherapy. Data on known prognostic factors (age, white blood cell count (WBC) at diagnosis, cytogenetic risk groups (as defined by CALGB 8461), and AML etiology) were collected as baseline characteristics and controlled for in stepwise multivariable analyses. Complete response (CR) and overall survival (OS) were analyzed. A prognostic scoring system for OS was developed based on the number of poor prognostic features present, derived from significant multivariable factors. Pts received 1 point for adverse cytogenetics, 1 point for having 1-10% peripheral blasts, and 1 point for AML arising from MDS or MPD. Pts with 0 points were favorable, 1 point intermediate, 2 or more points unfavorable. Results: Of 584 AML pts identified, 361 were treated with remission induction therapy, of whom 90 had AML arising from MDS, MPD, or t-AML. Thirty-nine (43%) had antecedent MDS, 21 (23%) an MPD, and 30 (33%) had t-AML, and 47% were female. Pts with AML arising from MDS were older at AML diagnosis (median of 67 years) vs. from MPD (61 years) and t-AML (60 years) (p=.02) but a shorter time from antecedent diagnosis/event (7 months, vs. 47 and 37 months, respectively (p

Description: Abstract 4481 Patients undergoing hematopoietic cell transplantation (HCT) require central venous access during treatment, predisposing this inherently susceptible population to infection. Central line-associated blood stream infection (CLABSI) is defined by the National Healthcare Safety Network as a primary bloodstream infection (BSI) in a patient with a central line within the 48-hour period before the development of the BSI. CLABSI surveillance is being increasingly used as an objective measure of quality of care delivered at individual hospitals. The Centers for Disease Control and Prevention have developed guidelines for the insertion, surveillance, and timely removal of these lines to prevent CLABSI, of which approximately 10% are fatal, and the Centers for Medicare & Medicaid will adjust reimbursement for CLABSI. The incidence, risk factors, and impact on survival of CLABSI in acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS) patients undergoing HCT has not been reported. AML or MDS patients undergoing HCT between August 2009 and December 2011 were identified from the Cleveland Clinic Unified Transplant Database, and occurrence of CLABSI was determined from the infection control database. Variables analyzed included occurrence of CLABSI, as well as patient demographics, disease type, prior treatment, HCT comorbidity index, transplant type/HLA-match, CD34+ count, and time to neutrophil recovery (absolute neutrophil count 〉500). CLABSI incidence was estimated using Kaplan-Meier method, and univariable and multivariable risk factors were identified by Cox proportional hazards analyses. Of the 73 patients identified, 48 were male; 68 were Caucasian; 44 had AML, and 29 MDS. The median age at transplant was 52 (range 16–70), and 39 had a low to intermediate HCT comorbidity index (0–2), while 34 had a high index (≥3). Patients received a median of 2 prior chemotherapy regimens (range 0–6), 3 had prior radiation, and 6 had prior transplant. Preparative regimen was myeloablative (n=54) or reduced-intensity (n=19); 34 received bone marrow (BM), 24 peripheral stem cells (PSC), and 15 cord blood cells (CBC). The median CD34+ count was 2.42 × 106/kg and median time to neutrophil recovery was 14 days (range 6–24) with BM/PSC compared to 28 days with CBC (range 19–77). Among these 73 patients, 23 (31.5%) developed CLABSI, of whom 16 (69.6%) died. The majority (16/23) of CLABSI occurred within 14 days (median 9 days, range 2–211 days) from HCT (Figure 1), but timing of CLABSI was highly associated with cell source: median of 5 days (range 2–12 days) for CBC and 78 days (range 7–211 days) for BM/PSC (p

Description: Key Points For patients with acute myelogenous leukemia, post-transplant survival is not determined by donor source (unrelated vs related). However, for patients with myelodysplastic syndromes, donor source remains an important determinant of post-transplantation outcomes.

Description: Myelodysplastic syndromes (MDS) are a heterogeneous group of blood cancers characterized by bone marrow (BM) failure, peripheral blood cytopenias, dysplasia, chromosomal abnormalities and an increased risk for transformation to acute myeloid leukemia (AML). Patients (pts) with higher risk disease are primarily treated with pharmacologic treatments like hypomethylating therapy (HMT) (5-azacytidine and decitabine). 5-azacytidine (AZA) and decitabine (DAC) can result in overall response rates of 36% with a median duration of response of 15 months and 17-21% with a median duration of response of 10 months, respectively. Pts refractory to HMT have poor outcomes with a median overall survival of ∼4 months. Spliceosome gene mutations are frequently found in certain subtypes of MDS specifically SF3B1 (∼28%), U2AF1 (6-12%) and SRSF2 (6-12%). The prognostic value of spliceosome mutations in different MDS subtypes has been largely investigated while the impact of these mutations on treatment response is still unknown. We aim to investigate the frequency of three commonly mutated spliceosome genes (SF3B1, U2AF1, and SRSF2) in pts who failed HMT in order to define mutational frequency and evaluate the feasibility of targeted therapy with next generation spliceosome inhibitors. We screened a cohort of 120 pts (MDS, 70; MDS/MPN, 33; MDS/sAML, 17; median age: 69; male/female: 85/35) that underwent HMT (AZA: 58; DAC: 21; AZA/DAC: 7; AZA/REV: 25; DAC/REV: 4; AZA/DAC/REV: 5). Forty-eight percent of pts failed HMT therapy as refractory or relapse. We performed Sanger sequencing on BM/peripheral blood DNA for known pathways involved in MDS pathogenesis including methylation (TET2, DNMT3A, IDH1/2), histone (ASXL1, UTX, EZH2), signaling (CBL, N/KRAS), transcription (RUNX1, TP53, JAK2), and RNA splicing (SF3B1, U2AF1, SRSF2). Data analysis was available for 90 pts. We detected a total of 131 mutations in different pathways. In total, spliceosome mutations were observed in 28/90 (31%) of pts. When we analyzed the presence of the mutations in relation to the rate of response, we found that pts who failed HMT have frequent spliceosome mutations: 17/58 (29%). We have reported that molecular mutations in TET2 and DNMT3A can predict response to treatment to HMT (Traina F, Blood (ASH Annual Meeting Abstracts), Nov 2011; 118: 461). Indeed, the frequency of mutations in methylation genes was lower in the group of pts who failed HMT (11/58; 18.9%) compared to pts who achieved hematological response (11/32; 34%). Spliceosome inhibitors have been proposed for targetted therapy in MDS. The presence of spliceosome mutations in pts who failed HMT can open a new era of investigation leading to the possibility of using spliceosome inhibitors in pts who fail conventional therapy. We performed RNA-sequencing analysis on BM cells of pts who failed HMT compared to pts who achieved hematological response (n=2 vs 2) in order to define any specific gene signature explaining the differences in response to HMT. We performed differential gene expression testing on 11,459 expressed genes. In total, 158 genes were differentially expressed at FDR 〈 .2 in responders compared to not responders. We identified several interesting genes involved in tumorigenesis and epigenetic regulation such as YPEL3, and ST14, which were up-regulated in responders vs not responders (FC: 4 and 7.5; P

Description: Abstract 461 Aberrant DNA methylation is a hallmark of myelodysplastic syndromes (MDS), MDS/myeloproliferative neoplasms (MDS/MPN) and secondary acute myeloid leukemia (sAML). It provides a rationale for treating these malignancies with hypomethylating agents like 5-azacitidine (AZA) and decitabine (DAC). However, treatment outcomes remain limited and heavily weighed on morphologic/cytogenetic results. The discovery of novel mutations has provided important insight into the pathogenesis of MDS and related disorders. Genes implicated in epigenetic regulation, including DNMT3A, TET2, IDH1/IDH2, EZH2, ASXL1 and UTX have been found mutated in MDS, while others have also been implicated in MDS pathogenesis. There is limited data on the predictive value of these genetic defects for treatment response and disease outcome. We hypothesized that these defects are important biomarkers predictive of response to hypomethylating agents. We studied 88 patients with MDS (RCUD=2, RARS=6, RCMD=11, MDS-U=3, RAEB-1/2=29, CMML1/2=16, MDS/MPN-U=5, RARS-T=5, AML from MDS=11) who received hypomethylating agents (AZA=53, DAC=24, both=11). The median number of cycles was 7 [range 1–35], median age was 69 years (range 42–82) and median follow-up was 18 months (range 0–76). Responses were scored according to IWG criteria. DNMT3A, TET2, IDH1/2, EZH2, ASXL1, UTX, KRAS, NRAS, CBL, RUNX1, TP53 and SF3B1 were sequenced using standard techniques. Categorical variables were analyzed using Chi-square statistics. Overall survival (OS) was analyzed using Kaplan-Meier; p-values ≤ 0.05 were considered statistically significant. Mutated patients were older than wild type (WT) cases (72 vs. 68 years, p=.01) but were well matched for marrow blast %, cytogenetic risk group and cycles of hypomethylating agents received. We found mutations in 40/88 (45%) patients. Mutations were most frequent in SF3B1 (6/11; 55%), ASXL1 (13/50; 26%), TET2 (18/88; 20%), KRAS (3/34; 9%), and DNMT3A (7/88; 8%). Less common were mutations in EZH2 (2/43; 5%), TP53 (1/23; 4%), IDH1 (4/88; 5%), IDH2 (3/88; 3%), and UTX (1/36;3%). No mutations were found in CBL, NRAS or RUNX1. Based on single mutations, overall response rate (ORR) was higher in mutated vs WT patients for DNMT3A (6/7 [86%] vs 33/81 [41%]; p=.02), ASXL1 (11/13 [85%] vs 14/37 [38%]; p=.003), and TET2 (12/18 [67%] vs. 27/70 [39%]; p=.03). All heterozygous DNMT3A mutants responded to hypomethylating agents. Differences remained significant when stratified to AZA treatment alone for DNMT3A (6/7 [86%] vs 21/56 [38%]; p=.01) and ASXL1 (9/11 [82%] vs 12/29 [41%]; p=.02) but not TET2 (6/10 [60%] vs 21/53 [40%]; p=0.22). The predictive value of combined mutations were analyzed for DNMT3A, TET2 and/or IDH1/2, showing better response to hypomethylating therapy in patients who had a mutation; ORR (mutated: 18/28 (64%) vs WT: 21/60 (35%); p=.01). This difference remained significant in patients receiving only AZA (n=53); ORR was 11/18 (61%) in mutant and 11/35 (31%) in WT patients (p=.03). No differences in ORR were noted for KRAS, EZH2 and IDH1/2 mutant and WT patients. No SF3B1 mutants responded to treatment while both patients with UTX and TP53 mutations responded. The frequency of AML evolution was also analyzed and showed no difference between mutant and WT cases for TET2 (7/18 [39%] vs 22/70 [31%];p=.52), ASXL1 (4/10 [40%] vs 11/35 [31%]; p=.61), and DNMT3A (3/7 [43%] vs 26/81 [32%];p=.56). No differences in OS and progression free survival (PFS) were noted between responders and non-responders to hypomethylating therapy (28 vs 17 mos, p=.25; 16 vs 8 mos, p=.54). Comparison of survival outcomes for mutant and WT patients showed no significant difference for DNMT3A (OS: 30 vs 21 mos, p=0.43; PFS: 20 vs 11, p=.53), ASXL1 (OS: 28 vs 22, p=.68; PFS: 16 vs 10, p=.88), and TET2 (OS: 30 vs 20 mos, p=.30). PFS was better in TET2 mutants compared to WT (19 vs 9, p=.03). No survival differences were noted between mutant and WT cases who responded to hypomethylating agents for DNMT3A (OS: 25 vs 28,p=.84; PFS: 14 vs 16, p=.78), ASXL1 (OS: 10 vs 18, p=.48; PFS: 10 vs 6, p=.76) TET2 (OS: 27 vs 16, p=.79; PFS: 18 vs10, p=.19). In conclusion, DNMT3A, ASXL1 and TET2 mutations were independently associated with a better response to hypomethylating drugs. Moreover, combined mutations in DNMT3A/TET2/IDH1/IDH2 may influence the response to hypomethylating agents, especially AZA supporting its role as a predictive biomarker in MDS treatment. Disclosures: Maciejewski: Celgene and Eisai, NIH, AA&MDS Foundation: Research Funding. Tiu:MDS Foundation Young Investigator Award: Research Funding.

Description: Abstract 2015 Allogeneic hematopoietic stem cell transplantation (AlloSCT) remains the only curative option for MDS. Several retrospective studies evaluated the impact of various prognostic factors (i.e. cytogenetic risk group, WHO classification, ferritin level etc.) on post-transplant outcomes of pts with MDS, however comprehensive analyses including a cytogenetic abnormalities detected by SNP array (SNP-A) karyotyping method have not been performed. We have analyzed prognostic factors of post-AlloSCT outcomes among 74 pts with MDS (2000–2010) including the predictive value of SNP-A abnormalities. Cox proportional hazards analysis was used to identify univariable prognostic factors for acute GVHD (aGVHD), chronic GVHD (cGVHD), disease relapse, relapse free (RFS) and overall survival (OS). Multivariable prognostic factors were identified by stepwise Cox proportional hazards analysis. The median time from MDS diagnosis to transplant for all pts was 6 mos (range, 0.2– 141 mos). The median age at transplant was 51 yrs; 32% of the pts had a hematopoietic cell transplant co-morbidity index (HCT-CI) score ≥ 3; 69% had ≥1 prior chemotherapies; and only 30% were in remission prior to their transplant. 27 pts (37%) had RAEB-2, 11 (15%) had RAEB-1, and 9 (12%) had treatment-related MDS. 42 pts (58%) belonged to an intermediate-2 or higher IPSS risk category. 23 pts (31%) had adverse karyotype (complex or monosomy 7) detected by metaphase cytogenetics (MC). SNP abnormalities were identified in 58% of patients; 79% of all patients with SNP abnormalities had lesions not previously detected by traditional cytogenetic techniques. Median pre-transplant ferritin level was 1127 (range, 9–5201). 73% of the pts received myeloablative conditioning. In 61% of cases stem cells were harvested from the bone marrow. Matched related donors accounted for half of the cases. Twelve pts (16%) died within 100 days of transplant and 39 pts (53%) within the median follow up of 36 mos (range, 5–114). MDS relapse occurred in 22 pts (30%). The rates of grade II-IV aGVHD and extensive cGVHD were 49% and 24% respectively. Disease relapse was the most common cause of death (31%) followed by aGVHD (18%) and cGVHD (13%). In univariate analysis, aGVHD was associated with myeloablative (p

Description: Abstract 2356 Introduction: Allogeneic stem cell transplantation (SCT) is commonly offered to patients with acute myeloid leukemia (AML) over the age of 55 with good performance status. For this patient population, potential sibling donors are necessarily older and have more comorbidities than siblings of younger patients. It is not known whether the advanced donor age of older patients outweighs the beneficial effect of having a sibling donor. To address this question, we retrospectively analyzed data from 62 consecutive patients who received allogeneic SCT at the Cleveland Clinic from 1990–2009. Methods: Inclusion criteria were: age ≥ 55 years, diagnosis of AML, and history of myeloablative or nonmyeloablative allogeneic SCT from related or unrelated adult donors. 31 patients underwent transplantation from matched related donors (30 siblings, 1 cousin). 31 patients received unrelated donor transplants. The median age was 59 for patients with related and unrelated donors (P=0.89). There was an equal percentage of males (64.5%) in both groups (P=1.0). There was an equivalent distribution hematopoietic cell transplant comorbidity index scores of low, intermediate and high for patients with related and unrelated donors (38.7%, 37.1% and 24.2% vs. 38.7%, 32.3%, and 29.0% P = 0.65). 45.2% of patients with related donors underwent myeloablative conditioning vs. 54.8% for patients with unrelated donors (P =0.61). Results: Sibling donors were significantly older than unrelated donors [median 59 years (range 41–75) vs. 36 (range 24–58), P 〈 0.001]. The incidence of acute and chronic graft-versus-host disease (GVHD) was similar in both groups. At five years, the cumulative incidence of relapse (43.2% vs. 38.7%, P = 0.88), non-relapse mortality (48.1% vs. 48.8%, P =0.91) and overall survival (17.2% vs. 24.1%, P =0.88) were similar for recipients of matched related and unrelated donor transplants. Older donor age was not predictive of death in univariate or multivariate analysis. Kaplan-Meier estimates of overall survival for recipients of related and unrelated donor transplants are shown. Conclusion: Patients with AML 55 years or older who underwent related donor transplantation had significantly older donors but equivalent survival when compared to patient who underwent unrelated donor transplantation. Advanced donor age should not be a contraindication to allogeneic SCT for older patients with AML. Disclosures: No relevant conflicts of interest to declare.