Gangat, Naseema

Abstract Background In myelodysplastic syndromes (MDS), we have previously shown the adverse prognostic effect of ALC <1.2 x 10(9)/L (Nisha et al. Am J Hematol 2010;85:160). In the current larger study of 889 patients with MDS, we sought to validate and expand on our previous observations and also examine the effect of lymphocyte-to-monocyte ratio (LMR). Methods We retrospectively recruited 889 consecutive patients with primary MDS who were untreated at the time of referral to our institution, and in whom ALC and absolute monocyte count (AMC) at time of referral was documented. The diagnosis of MDS and leukemic transformation (LT) was according to WHO criteria (Blood. 2009;114:937). Complete follow-up information was updated in January 2015. Comparison of laboratory and clinical features was made between patients with subnormal and normal ALC; patients with above normal ALC were considered separately. Overall and leukemia-free survivals of patients with normal ALC was compared to those with either subnormal ALC or, in order to validate our previous observation, with ALC <1.2 x 10(9)/L. Results Patient characteristics: Median (range) values for the 889 study patients (69% males) included: age 72 (18-98), hemoglobin 9.6 g/dL (5.4-15.7), leukocyte count 3.4 x 10(9)/L (0.4-35), AMC 0.2 x 10(9)/L (0-1.7), platelet count 106 x 10(9)/L (2-1804), circulating blasts 0% (0-18) and bone marrow blasts 3% (0-19). Transfusion need was documented in 33% of patients and abnormal karyotype in 49%. Risk stratification by the revised international prognostic scoring system (IPSS-R) was very high in 10%, high in 16%, intermediate in 20%, low in 36% and very low in 16%. The median (range) ALC for the entire study population of 889 patients was 1.16 x 10(9)/L (0.02-8.9). The normal range of ALC at our institution was 0.9-2.9 x 10(9)/L; the number of patients with subnormal, normal and above normal ALC was 261 (29%), 598 (67%) and 30 (4%), respectively. In addition, 442 (50%) and 417 (47%) patients had ALC below 1.2 x 10(9)/L and ALC between 1.2 and the upper limit of normal, respectively. After a median follow-up of 27 months, 712 (80%) deaths and 116 (13%) leukemic transformations were documented. Comparison of patients stratified by absolute lymphocyte count Compared to patients with normal ALC, patients with subnormal ALC displayed lower hemoglobin (p=0.002), higher red blood cell transfusion need (p=0.0003), lower leukocyte count (p<0.0001), lower monocyte count (p=0.002) and lower platelet count (p<0.0001), whereas borderline association was also apparent for older age (p=0.06). In univariate analysis, survival was adversely affected by lower ALC, treated as a continuous variable (p=0.01). Similarly, when compared to patients with normal ALC, patients with subnormal ALC (p=0.001; HR 1.3, 95% CI 1.1-1.5) and those with ALC <1.2 x 10(9)/L (p=0.0002; HR 1.3, 95% CI 1.2-1.6) experienced significantly shortened survival. There was no difference in survival between patients with above normal ALC and those with either subnormal (p=0.58) or normal (p=0.41) ALC; accordingly, patients with above normal ALC were excluded from subsequent analysis. In multivariable analysis, the adverse effect of subnormal ALC was shown to be independent of age, gender, anemia, thrombocytopenia, neutropenia , circulating blast percentage, bone marrow blast percentage or cytogenetic risk stratification by and IPSS; the same held true for ALC <1.2 x 10(9)/L. The association with IPSS-R was of borderline significance for both ALC <1.2 x 10(9)/L (p=0.06) and subnormal ALC (p=0.1). The adverse effect of ALC was most apparent in patients with very low or low risk disease per IPSS-R (p=0.0007 for subnormal ALC and 0.0003 for ALC <1.2 x 10(9)/L), compared to those with higher risk status (p=0.67 and 0.7, respectively). Neither subnormal ALC (p=0.4) nor ALC <1.2 x 10(9)/L (p=0.1) affected leukemia free survival. Analysis using LMR in place of ALC showed similar but not necessarily superior results. Conclusions Compared to those with normal ALC, MDS patients with either subnormal ALC or with ALC below 1.2 x 10(9)/L experienced shorter overall, but not leukemia-free, survival; this adverse effect was most apparent in lower risk patients and was independent of most of the currently established risk factors for survival in MDS. The observations from the current study suggest the possible therapeutic value of augmenting host immunity in lower risk MDS. Disclosures Al-Kali: Onconova Therapeutics, Inc.: Research Funding; Celgene: Research Funding.

Abstract Background: Myelodysplastic syndromes (MDS) are rare hematological neoplasms that are more typically seen in elderly patients. Young patients (< 50 years old) have been reported to comprise between 3-6% in the Surveillance, Epidemiology and End Results (SEER) (Ma X et al, Cancer 2007; Rollison R et al, Blood 2008) with a better overall survival (OS). We hereby report the characteristics of young MDS patients with long survival follow-up. Methods: A total of 1012 MDS patients' data from Oct 1993 to Dec 2015 at Mayo Clinic were reviewed after appropriate IRB approval was obtained. All cases had their bone marrow slides reviewed at our institution. Patients youngers than 50 years old (yMDS) were included as cohort 1, while the rest was included as control group (cohort 2). Prognostic factors were analyzed by univariate and multivariate analyses. Survival estimates were calculated using Kaplan-Meier curves and univariate and multivariate analyses was based on log-rank testing using JMP software version 10. Results: Characteristics: We identified 68 (7%) yMDS patients with a median age of 42 years (range, 18-49). Female gender was more common in yMDS (43% vs 31%, p= 0.05). Upon comparison between cohort 1 and 2, only platelets were significantly lower in yMDS (61 vs 102, p <0.0001), but not white blood cells, hemoglobin or degree of marrow fibrosis. MDS subtyping according to World Health Organization 2016 showed single lineage dysplasia in 15% vs 2%, multilineage dysplasia 24% vs 36% , ring sideroblasts 9% vs 15%, isolated del (5q) 1% vs 3%, excess blasts 44% vs 31%, and unclassifiable in 6% vs 5%. As expected, therapy related MDS (t-MDS) was more frequent in yMDS (33% vs 17%, p= 0.005). Transformation to acute myeloid leukemia (AML) was also more frequent in yMDS (28 % vs 11%, p= 0.0004). Compared to cohort 2, yMDS IPSS-R scores were very high, high, intermediate, low, and very low in 24% vs 13%, 29% vs 16%, 21% vs 20%, 19% vs 35%, and 7% vs 17%, respectively. Allogenic hematopoietic cell transplantation (HCT) was more frequent in yMDS (42% vs 4%, p< 0.0001). Survival outcome: Median OS was longer for cohort 1 vs cohort 2 but did not reach statistical significance (43 vs 21 months, p= 0.1). Median progression free survival (PFS) was shorter for cohort 1 vs cohort 2 but did not reach statistical significance (8 vs 12 months, p= 0.3). Median OS for cohort 1 based on R-IPSS was 44, 105, 40, 18, and 12 months for very low, low, intermediate, high and high risk groups, respectively (p= 0.09). Median OS was shorter in t-MDS vs de novo MDS in cohort 1 (13 vs 47 months, p = 0.04). Young patients who had transformed to AML had a worse median OS (18 vs 93 months, p=0.001). On multivariate analysis neither t-MDS nor R-IPSS had a statistically significant impact on OS. Conclusions: MDS is rarely diagnosed under the age of 50. IPSS-R was less powerful in detecting differences between its risk groups for this patient population, although more than half of the patients with yMDS had either high or very high risk. Among this cohort of yMDS patients, there was a significantly higher proportion with therapy-related myeloid neoplasms compared to older patients (one third of patients), with subsequent higher rates of transformation to AML and higher allogeneic HCT. In our study, we did not find an improved OS for yMDS patients compared to older patients. Disclosures Al-Kali: Celgene: Research Funding; Onconova Therapeutics, Inc.: Research Funding.

Abstract Background: Approximately 1,000 acute promyelocytic leukemia (APL) patients are diagnosed in the US annually. APL has the highest cure and early mortality rates of all acute leukemias. The availability of all trans-retinoic acid (ATRA) in 1995 and arsenic trioxide in 2000 has further improved the overall survival (OS). However, whether there is uniform improvement in OS across demographic subgroups and if sooner time-to-treatment (TTT) reduces early mortality are unknown. We used the National Cancer Data Base (NCDB), a nationwide oncology registry covering 70% of the US cancer population to answer these questions. Methods: We identified patients with APL diagnosed from 1998-2011 using ICD-O-3 code 9866. Survival functions were estimated using the Kaplan-Meier method and 30-day mortality was estimated as 1-survival at 30 days. We compared OS according to demographic subgroups (gender, age [18-40, 41-64, and >65 years], race [White, Black, Native American, Asian/Pacific Islander, and other], and ethnicity) across years of diagnosis (Period [P] 1: 1998-2001, P2: 2002-2006, and P3: 2007-2011). For 30-day mortality, we compared the rates according to demographic subgroups and time periods and determined the effect of TTT (days from APL diagnosis to ATRA and/or chemotherapy). For this analysis we included only patients treated within 30 days of diagnosis and those diagnosed from 2003 to 2011. TTT was grouped as 0, 1, 2, or >3 days. We used mixed-effects logistic regression model to obtain odds ratio comparing treatment delays. Our model accounted for multiple subjects per facility with a random intercept and was adjusted for gender, Charlson-Deyo score, and year of diagnosis. Results: Characteristics: There were 8,084 APL patients in our study. The median age at diagnosis was 51 (24% were older than 65) years and 50% were females. White race (85%) was the most common, followed by black (11%) and Asian (3%). Twelve percent were Hispanic in origin. Slightly more patients (54.8%) were treated in academic centers. Thirty seven percent had either Medicaid or Medicare insurance. OS: Race had no impact on OS, although Asians had a trend towards improved OS (P=0.06). Both females and young age affected OS favorably (P <0.0001). In general, OS improved over the three time periods (P=0.002). Across Periods 1-3, we observed significant improvement in OS among the males (P=0.03), 18-40 (P=0.035) and 41-64 (P=0.005) age groups, Whites (P=0.012), and non-Spanish/Hispanics (P=0.002), but not among their counterparts. 30-day mortality rate: Age above 65 years was associated with highest 30-day mortality (33%). Similarly male gender had statistically significant higher 30-day mortality (shown in the Figure). There was no statistically significant difference among Periods 1-3. As expected, untreated patients had an increased risk of death versus treated (HR=1.9, confidence interval 1.7-2.1). TTT: A third (33.5%) of the patients was treated within two days of APL diagnosis. TTT was not independently associated with lower 30-day mortality rate (shown in the Table). Presence of disseminated intravascular coagulopathy (DIC) data was not available to analyze. Conclusions: From 1998-2011, the OS of APL continued to improve but not uniformly across demographic subgroups. In contrast, the 30-day mortality did not improve in the same time period. Both Age above 65 and male gender predicted worse 30-day mortality and OS. Our study did not find a favorable association between shorter TTT and lower 30-day mortality rate. Our study was not able to study these findings among different risk groups. Disclosures Al-Kali: Celgene: Research Funding. Tibes:TetraLogic Pharmaceuticals: Research Funding.

Abstract Background Increased serum levels of lactate dehydrogenase (LDH) typically accompany primary myelofibrosis (PMF) and might be linked to increased cell turnover from clonal myeloproliferation, including leukocytosis, low grade hemolysis and extra-medullary hematopoiesis in the liver. Despite the fact that serum LDH has been extensively utilized as a prognostic marker in both lymphoid neoplasms and solid tumors, there are limited studies in PMF that have systematically examined the clinical significance of the degree of serum LDH elevation in PMF. Methods Study patients fulfilled the 2016 WHO criteria for the diagnosis of PMF, including both pre-PMF and overt PMF (Blood. 2016;127:2391). Additional selection criteria included the availability of serum LDH at time of referral. Marked elevation of serum LDH was defined as a value of ≥1000 U/L (i.e. over 4-fold increase from the upper limit of the normal range for our institution, which was 122-222 U/L), based on preliminary analysis of the threshold for prognostic effect. Targeted next generation sequencing was used to screen for prognostically-relevant mutations (Blood 2015126:354). Statistical analyses considered clinical and laboratory parameters obtained at time of initial referral to the Mayo Clinic. Results LDH values in overt PMF versus pre-PMF: The entire study population consisted of 357 patients, including 311 with overt PMF and 46 with pre-PMF. The median serum LDH level was 514 U/L (range 136-2263): overt PMF 532 U/L (range 136-2263); pre-PMF 401 U/L (range 180-1237) (p=0.0003). Accordingly, in order to avoid the confounding effect of the difference in serum LDH level between overt PMF and pre-PMF, and considering the small number of patients with pre-PMF, further analysis was limited to the 311 patients with overt PMF. Patient characteristics: The 311 patients (median age 64 years; 66% males) with overt PMF included 205 (66%) JAK2, 49 (16%) type 1/like CALR, 13 (4%) type 2/like CALR, 16 (5%) MPL mutated cases and 28 (9%) were "triple-negative". DIPSS-plus risk distribution was 31% high, 43% intermediate-2, 15% intermediate-1 and 12% low; 30% displayed red cell transfusion-dependency and 37% abnormal karyotype, including 14% with unfavorable karyotype. 184 patients were screened for ASXL1 mutations with 42% mutated and 205 for SRSF2 mutations with 16% mutated. Clinical correlates of markedly elevated LDH (≥1000 U/L): Among all 311 study patients with overt PMF, 37 (12%) displayed marked elevation of LDH (≥1000/L). Patients with marked elevation of LDH displayed significantly higher leukocyte count (p=0.005; R-squared = 0.05), circulating blast percentage (p=0.03; R-squared = 0.07) and SRSF2 mutational frequency (44% vs 12%; p<0.0001). Survival analysis: After a median follow up of 3 years, 199 (64%) deaths and 31 (10%) leukemic transformations were documented. In univariate analysis, increased serum LDH level was associated with inferior survival, both as a continuous variable (p=0.002) and as a categorical variable with the cutoff level of 1000 U/L (HR 2.02, 95% CI 1.3-3.1; p=0.001); the survival effect LDH ≥1000/L was independent of DIPSS-plus (HR 1.6, 95% CI 1.1-2.5). Other variables that were significantly associated with shortened survival, on univariate analysis, included all 8 DIPSS-plus variables (p≤0.01 in all instances), absence of CALR type 1/type 1-like (p<0.0001) and presence of ASXL1 (p<0.0001) or SRSF2 (p=0.0006) mutations. In multivariable analysis that included only genetic risk factors, serum LDH retained its significance (HR 2.2, 95% CI 1.3-3.6), along with absence of CALR type 1/type 1-like, or presence of ASXL1 or SRSF2 mutations or unfavorable karyotype. In multivariable analysis that included only clinical variables, serum LDH ≥1000/L was again independently predictive of poor survival (HR 1.7, 95% CI 1.1-2.6), along with age >65 years, hemoglobin <10 g/dL, platelets <100 x 10(9)/L, leukocyte count >25 x 10(9)/L and constitutional symptoms. Patients with marked LDH elevation were also more likely to undergo leukemic transformation (HR 3.1, 95% CI 1.2-7.6). Conclusion The current study suggests that marked elevation of serum LDH in PMF indicates aggressive tumor biology that is currently not accounted for by established clinical or genetic risk factors; the low R-squared values seen in relation to leukocyte count and circulating blasts are consistent with this assumption. Disclosures No relevant conflicts of interest to declare.

Abstract Background: Acute erythroleukemia (AML-M6) is an aggressive myeloid leukemia, with poor outcomes (median survival < 12months). In 2008, the WHO further sub-classified this disease into two subgroups- erythroid leukemia and pure erythroid leukemia, based on the number of erythroblasts and myeloblasts. We carried out this study to evaluate differences in presenting features and outcomes between erythroid leukemia and pure erythroid leukemia. Methods: After due IRB approval, out of 923 AML, 43(4.6%) pts who met the 2008 WHO criteria for acute erythroleukemia (2000-2014) were identified, and clinical characteristics retrospectively reviewed. A diagnosis of erythroid leukemia (M6a) was considered if > 50% of the total nucleate BM cells were of erythroid lineage and > 20% of non-erythroid cells were myeloblasts. Pure erythroid leukemia (M6b) was diagnosed in pts who presented with > 80% of BM nucleate cells consisting of immature erythroid precursors. Cytogenetic risk stratification was based on NCCN AML guidelines. Statistical comparisons were performed with the χ2 test for categorical variables and the Wilcoxon test for continuous variables. Univariate and multivariate analyses were performed with the Cox logistical regression. Survival analysis was based on the Kaplan-Meier method and log rank test. For all analyses, P ≤ 0.05 was considered statistically significant. Results: The median age at diagnosis was 65 years (27-90) and 34(79%) were male. 30 (69%) had M6a and 13 (30%) had M6b. 27 (62%) had de novo AML M6, whereas 16 (37%) had secondary AML (9 underlying MDS and 7 therapy related).The median time to leukemic transformation for pts with underlying MDS was 8 months (2-83). Median follow up in our cohort was 7.2 months (2-157) at which time 13 (30%) deaths were documented. A. Patient characteristics: At time of diagnosis median laboratory values included; Hemoglobin 8.5 g/dL (4.6-11), WBC 2x109 (0.7-11.1), ANC 0.47x109 (0.01-4.07), Platelet 33x 109 (7-177), peripheral blast % 2 (0-35), and bone marrow blast % 11 (0-84). Cytogenetic information was available in 38 pts; of which 6(16%) were diploid, 21(55%) were complex/monosmal and 1(3%) was complex/non monosomal. Intermediate risk cytogenetic was observed in 15 pts (39%), and poor risk cytogenetic was identified in 23(61%). Comparing M6a and M6b, we observed that pts with M6a tend to have more leukopenia and neutropenia and higher marrow blast percentage compared to M6b (Table 1). None of the patients experienced extramedullary disease or CNS involvement. B. Outcome and prognostics: We did observe a trend towards worse survival in pts with M6b, 7 months vs 12.9 months with M6a (p= 0.08) (Figure 1). There was no difference in survival in acute erythroleukemia compared to other acute myeloid leukemias, 10.1 vs 14 months (p= 0.28), respectively (Figure 2). In univariate analysis, WBC, ANC, and cytogenetic risk impacted survival (Table 2), however only cytogenetic risks retained significance in the multivariate model; Intermediate risk HR 0.18 (0.05-0.51; p = 0.0009) and poor risk HR 5.4 (1.9-17.6; p = 0.0009). Conclusion: Acute Erythroid Leukemia is a rare leukemia with ~5% incidence in our cohort. We observed that regardless of the percentage of bone marrow erythroblasts and myeloblasts, survival in patients with Acute Erythroleukemia are predominantly impacted by cytogenetic risk stratification. This raises a question on the clinical relevance of the sub-categorization of acute erythroleukemia into erythroid leukemia and pure erythroid leukemia. Larger studies are needed to validate our results. Figure 1. Figure 1. Disclosures Al-Kali: Celgene: Research Funding.

Abstract Background: Ring sideroblasts (RS) are erythroid precursors with perinuclear mitochondrial iron accumulation seen in clonal disorders, such as myelodysplastic syndromes (MDS), and non-clonal conditions, including copper deficiency, lead poisoning, and congenital sideroblastic anemias (CSAs). CSAs involve a broad clinical and pathological spectrum related to mutations in ALAS2, ABCB7, etc. or mitochondrial aberrations (Pearson Syndrome; Myopathy, Lactic Acidosis and Sideroblastic Anemia (MLASA); etc.). MDS-RS is often associated with SF3B1 mutations, which have not been reported in other SAs. This study sought to further describe the characteristics and outcomes of patients with CSAs. Methods: After due IRB approval, we retrospectively identified all patients with a CSA diagnosis seen at our institution from 1990-2014. All bone marrow (BM) slides were re-reviewed to confirm the presence of RS. Secondary causes of SA were excluded. Genetic testing on the BM specimen was carried out when possible through research collaborations with the Boston Children's Hospital. Data was retrospectively extracted and is reported in Table 1. Data was analyzed based on the presence or absence of mitochondrial inheritance. Results: Seventeen patients with CSA were identified, 3 (17.6%) with mitochondrial inheritance. In the non-mitochondrial inheritance group, the median age at diagnosis was 29 years (range, 1-61 years). Six (42.9%) were males. At a median follow-up of 119 months (range, 1-401 months), 1 (7.1%) death was documented. The median overall survival (OS) has not been reached. Six of 14 (42.9%) underwent genetic testing, and a mutation was identified in 4 (ALAS -2, FECH -1, and a novel mutation, currently being validated). Six of 6 (100%) samples analyzed for SF3B1 mutations were negative. Median laboratory values at diagnosis included hemoglobin 9.5 g/dL, MCV 85.8, WBC 6.4, and platelets 297,000. Three (21.4%) patients had splenomegaly. Nine (64.3%) had iron overload; 3 (33%) were treated with iron chelation therapy, with only 1 documented response (≥ 500 µg/L reduction in serum ferritin). Twelve (85.7%) were treated with pyridoxine. Outcomes of pyridoxine treatment are described in Table 1. Three of 14 (21.4%) received red blood cell transfusions, and 3 (21.4%) received erythropoiesis stimulating agents (ESA). One (7.1%) patient underwent allogeneic stem cell transplant. Three patients had SAs of mitochondrial inheritance; aberrations were identified in 2 (YARS2 and a 4 kilobase mitochondrial deletion). In this group, the median age at diagnosis was 2 years (range, 1-13 years), and all were males. At a median follow-up of 173 months (range, 119-329 months), 2 (66.7%) deaths were documented. Median laboratory values at diagnosis included hemoglobin 9.0 g/dL, MCV 97.0, WBC 5.2, and platelets 175,000. Conclusions: Congenital SAs are rare, and the majority are of non-mitochondrial inheritance. The molecular basis for disease can be ascertained in less than 50% of patients. Unlike MDS with RS, CSAs are not characterized by the SF3B1 mutation, which may be used to establish clonality. With appropriate supportive care measures, survival for non-mitochondrially inherited CSA remains favorable. Table 1. Demographics and Outcomes for 17 Patients with CSA Age at Diagnosis (Years) Gender Molecular Mutation Additional Features Pyridoxine Response Iron Overload Survival (Months) Outcomes Non-Mitochondrial Inheritance 5 M R452H in ALAS2 N Y2 Y 173 Alive3 3 M Mutation being validated Splenomegaly Y2 Y 254 Alive3 6 M Unknown, PTPN11, ALAS 2, ABCB7 negative Splenomegaly N Y 137 Alive, status-post transplant 29 F Unknown1 N N/A N 131 Alive3 41 F Unknown1 N Y2 N 83 Alive4 40 M Promoter mutation in ALAS2 Splenomegaly N N 106 Alive4 31 F Unknown1 N N Y 401 Deceased 54 M Unknown1 N Lost to follow-up Y 1 Alive3 3 M FECH N N/A N 189 Alive3 1 F SCAD -VUS N Lost to follow-up Y 13 Alive3 29 F Unknown1 N N Y 205 Alive3 58 F Unknown1 N N Y 47 Alive, unknown transfusion status 61 F Unknown1 N Y2 Y 58 Alive3 23 F Unknown1 Alpha-thalassemia-2-trait Y2 N 37 Alive3 Mitochondrial Inheritance 13 M YARS2 MLASA N Y 329 Alive4 2 M Unknown, mitochondrial deletions and ABCB7 negative Myopathy, Ataxia N/A N 173 Deceased 1 M 4 kilobase mitochondrial deletion Pearson Syndrome, B cell lymphoma Y2 N 119 Deceased 1No molecular testing performed 2Sustained hemoglobin response of ≥ 1 g/dL over baseline for ≥ 12 weeks 3Transfusion-independent 4Transfusion-dependent Disclosures Steensma: Incyte: Consultancy; Onconova: Consultancy; Amgen: Consultancy; Celgene: Consultancy.

Abstract Background : Polycythemia Vera (PV) is associated with JAK2 mutations. In essential thrombocythemia (ET), ̴ 85% of patients harbor one of 3 "driver" mutations, with frequencies of ̴ 58%, 23% and 4%, for JAK2, CALR and MPL, respectively; ̴ 15% are wild type for all three mutations and are referred to as "triple negative". We applied next-generation sequencing (NGS) with a 27-gene panel of myeloid malignancy-relevant genes, in order to describe the prevalence of "non-driver" mutations and their prognostic relevance in PV and ET. Methods: Targeted capture assays were carried out on bone marrow or whole blood DNA for the following genes: TET2, DNMT3A, IDH1, IDH2, ASXL1, EZH2, SUZ12, SRSF2, SF3B1, ZRSR2, U2AF1, PTPN11, Tp53, SH2B3, RUNX1, CBL, NRAS, JAK2, CSF3R, FLT3, KIT, CALR, MPL, NPM1, CEBPA, IKZF, and SETBP1.Paired-end indexed libraries were prepared using the NEBNext Ultra Library prep protocol (NEB, Ipswich, MA /Agilent Technologies Inc, Santa Clara, CA). Capture libraries were assembled according to Nimblegen standard library protocol (Roche Nimblegen, Inc, Basel, Switzerland). Base-calling was performed using Illumina's RTA version 1.17.21.3. Genesifter® software (PerkinElmer, Danvers, Massachusetts) was utilized to analyze targeted sequence data. Nucleotide variants were called using the Genome Analysis Toolkit (GATK -Broad Institute, Cambridge, MA). Specific variants were deemed as mutations if they are associated with a hematologic malignancy (COSMIC database), or if they have not been associated with a dbSNP. Results: 314 patients with PV (n =133; median age 64 years, 53% females) or ET (n =181; median age 58 years, 59% females) were evaluated. Median follow-up was 9.8 years for PV and 9.2 years for ET. During this time 59 (44%) deaths, 15 (11%) fibrotic and 5 (4%) leukemic transformations were documented for PV and the corresponding percentages for ET were approximately 33%, 8% and 2%. Driver mutation distribution was 98% JAK2 for PV and 52% JAK2, 24% CALR, 20% triple-negative and 4% MPL for ET. Polycythemia Vera Mutations other than JAK2, CALR or MPL were seen in 58 (44%) patients; 29% harbored one, 14% two and one 3 mutations. None of 3 JAK2 -unmutated cases expressed non-driver mutations. Mutational frequencies were 18% TET2, 11% ASXL1, 5% SH2B3, 3% SF3B1 and ≤2% SETBP1, IDH2, DNMT3A, CEBPA, CSF3R and SUZ12, SRSF2, ZRSR2, TP53, CBL, NRAS, RUNX1, KIT, PTPN11 and FLT3. "Number of mutations" significantly affected overall (OS; Figure 1) and myelofibrosis-free (MFS) survival; respective HR (95% CI) were 2.6 (1.3-5.2) and 1.7 (0.97-3.1) and 13.7 (2.9-63.4) and 5.1 (1.5-17.6) for ≥2 mutations and one mutation, respectively. OS was also adversely affected by SRSF2 (p=0.006) and RUNX1 (p=0.04) and borderline affected by TET2 (p=0.06), IDH2 (p=0.08), ASXL1 (p=0.19) and SF3B1 (p=0.19) mutations. In multivariable analysis, SRSF2 and RUNX1 retained significance whereas the others remained borderline significant. In both univariate and multivariable analyses, leukemia-free survival (LFS) was adversely affected by IDH2 and RUNX1 mutations. In univariate analysis, ASXL1, IDH2, RUNX1 and KIT mutations predicted fibrotic progression, whereas SETBP1 was of borderline significance (p=0.07); all, including SETBP1, were significant during multivariable analysis. Essential thrombocythemia Mutations other than JAK2, CALR or MPL were seen in 83 (46%) patients; 35% harbored one, 7% two and 4% three mutations; prevalence in JAK2, CALR, MPL mutated and triple-negative cases was 52%, 43%, 43% and 35%, respectively (p=0.52). Mutational frequencies were: 13% TET2, 11% ASXL1, 6% DNMT3A, 5% SF3B1, 4% CEBPA, 2% TP53, SH2B3, EZH2 and CSF3R and <2% for SETBP1, IDH2, SRSF2, ZRSR2, CBL, NRAS, RUNX1, U2AF1, KIT, PTPN11 and FLT3. "Number of mutations" significantly affected OS (Figure 2) but not MFS or LFS; HR (95% CI) for OS were 6.6 (2.5-17.8) for 3 mutations and 2.2 (1.3-3.9) for one or two mutations. In univariate analysis, survival was adversely affected by CBL, EZH2, SF3B1, SRSF2 and IDH2 mutations. In multivariable analysis, EZH2 and SF3B1 remained significant. Univariate analysis identified SETBP1 and SF3B1 mutations as risk factors for fibrotic progression and EZH2, TP53 and CSF3R mutations for leukemic transformation. Conclusions: "Non-driver" mutations occur in more than 40% of patients with PV or ET; the number of such mutations and presence of certain specific ones likely predict OS, MFS or LFS. Figure 1. Figure 1. Figure 2. Figure 2. Disclosures Pardanani: Stemline: Research Funding.

Abstract Background : In primary myelofibrosis (PMF), ̴ 88% of patients harbor one of three "driver" mutations, with mutational frequencies of approximately 60%, 22% and 6% for JAK2, CALR and MPL, respectively. Other "non-driver" mutations have also been described in PMF and some of them and their number have been associated with inferior survival (Leukemia. 2014;28:1804). We applied next generation sequencing (NGS) with a broader panel of MPN-relevant genes, in order to identify additional mutations of prognostic relevance as well as obtain additional information regarding the prognostic value of 'number of mutations'. Methods: Targeted capture assays were carried out on bone marrow or whole blood DNA specimens obtained at time of referral for the following genes: TET2, DNMT3A, IDH1, IDH2, ASXL1, EZH2, SUZ12, SRSF2, SF3B1, ZRSR2, U2AF1, PTPN11, Tp53, SH2B3, RUNX1, CBL, NRAS, JAK2, CSF3R, FLT3, KIT, CALR, MPL, NPM1, CEBPA, IKZF, and SETBP1. Paired-end indexed libraries were prepared from individual patient DNA using the NEB Next Ultra Library prep protocol on the Agilent Bravo liquid handler (NEB, Ipswich, MA/Agilent Technologies Inc, Santa Clara, CA). Capture libraries were assembled according to Nimblegen standard library protocol (Roche Nimblegen, Inc, Basel, Switzerland). Base-calling was performed using Illumina's RTA version 1.17.21.3. Genesifter® software was utilized (PerkinElmer, Danvers, Massachusetts) to analyze targeted sequence data. Nucleotide variants were called using the Genome Analysis Toolkit (GATK-Broad Institute, Cambridge, MA). Specific variants were deemed as mutations if they are associated with a hematologic malignancy (as identified by COSMIC database), or if they have not been associated with a dbSNP. Results: 180 PMF patients were evaluated (median age 63 years; 65% males). DIPSS-plus risk distribution was 32% high, 38% intermediate-2, 17% intermediate-1 and 13% low. Driver mutation distribution was 62% JAK2, 22% CALR, 9% triple-negative and 7% MPL. Karyotype was abnormal in 41% of patients and unfavorable in 12%. Mutations other than JAK2, CALR or MPL (i.e. "non-driver" mutations) were seen in 150 (83%) patients including 88% of "triple-negative" cases. 62 (34%) patients harbored one, 55 (31%) two, 16 (9%) three and 17 (10%) four or more. Mutational frequencies were: ASXL1 36%, TET2 18%, SRSF2 17%, U2AF1 17%, ZRSR2 11%, SF3B1 10%, DNMT3A 9%, CEBPA (9%), Tp53 7%, SETBP1 6%, CBL 5%, IDH1/2 5%, SH2B3 4%, CSF3R 4%, NRAS 4%, RUNX1 3% and ≤2% for SUZ12, KIT, PTPN11, NPM1 and EZH2. DIPSS-plus high/intermediate-2 risk patients displayed higher number of mutations (p=0.0004) and higher mutational frequencies for ASXL1 (p=0.02), SRSF2 (p=0.004) and CBL (p=0.02). Associations noted included JAK2 with U2AF1 (p=0.03), unfavorable karyotype with CBL (p=0.01) and normal karyotype with ZRSR2 mutations (p=0.04). At a median follow-up of 4 years, 111 (62%) deaths were documented. For examination of impact on survival, we considered 'number of mutations' and specific mutations with >2% frequency. Accordingly, in univariate analysis, survival was adversely affected by 'number of mutations' (Figure 1) and presence of ASXL1, SRSF2, IDH1/2, U2AF1, RUNX1 and CBL mutations. For multivariable analysis, we considered three categories (zero, 1-3 and ≥4) for number of mutations based on the results from univariate analysis (Figure 1); the results showed ≥4 mutations, 1-3 mutations, RUNX1, CBL, ASXL1 and SRSF2 mutations were independently associated with shortened survival; the respective HR (95% CI) were 4 (1.4-11.1), 3 (1.3-6.8), 2.9 (1.1-8.1), 2.8 (1.3-6.3), 1.8 (1.2-2.7) AND 1.7 (1.03-2.7). When the multivariable analysis was repeated including only the 150 patients with at least one non-driver mutation, the 'number of mutations' was no longer significant (p=0.35) but ASXL1, CBL, RUNX1 and SRSF2 mutations retained their significance. The prognostic relevance of ASXL1 and CBL continued to be apparent even after the addition of DIPSS-plus and driver mutation profile to the multivariable model. Conclusions: Mutations other than JAK2, CALR or MPL occur in more than 80% of patients with PMF, including those with "triple-negative" driver mutational status. The absence of such mutations is independently favorable for survival while the prognostic effect of their presence is influenced by ASXL1, CBL, RUNX1 and SRSF2 mutations. Figure 1. Figure 1. Disclosures Pardanani: Stemline: Research Funding.

Abstract Background:Morphologic complete remission (CR) is the gold standard for assessing response in acute myelogenous leukemia (AML). CR with incomplete count recovery (CRi) and CR with incomplete platelet recovery (CRp) are associated with inferior overall survival post induction therapy (Walter et al, JCO 2010 and Chen et al, JCO 2015). Furthermore, residual cytogenetic disease is associated with a higher relapse compared with patients with cytogenetic CR (CCR; Chen et al, JCO 2011). There is a paucity of literature regarding the impact of CRi, CRp and CCR prior to allogeneic transplantation. Aim:To study the impact of CRi, CRp and CCR and their impact on relapse incidence (RI), non-relapse mortality (NRM) and overall survival (OS) in reduced intensity allogeneic stem cell transplantation (RIC-SCT). Methods:After due IRB approval, patients (pts) with acute myelogenous leukemia (AML) or myelodysplastic syndrome (MDS) who received RIC-SCT at our institution between 2008-2014 were identified. All clinical and pathologic data were retrospectively extracted. CR, CRi, CRp and CCR were defined in accordance to the international working group definition. Categorical and continuous variables were analyzed using Pearson's chi-squared and Wilcoxon/Kruskal-Wallis, respectively. Survival estimates were calculated using the kaplan-meier and log-rank test. RI was estimated on a competing event analysis using Grey's model. Cox proportional regression was used to compute hazard ratios (HR). Results: A. Baseline characteristics: A total of 134 patients were identified. A total of 70 (52%) were in CR, 40 (30%) in CRi and 24 (18%) in CRp. 129 patients had evaluable cytogenetic data pre-SCT; 81 (63%) were in CCR and 48 (37%) had persistent cytogenetic disease. Baseline patient, disease and transplant characteristics stratified by peripheral count recovery pre-SCT are shown (Table 1). Significantly more patients in CR had higher KPS, higher proportion AML diagnosis, more likely to be in first complete remission (CR1), higher rate of cytogenetic CR and a higher proportion of minor ABO mismatched donor. No other significant variables were identified between the three cohorts. B. Transplant outcomes: Median follow up of the entire cohort was 41.9 months and the 2-yr OS, RI and NRM were 61.7%, 25.7% and 21.9%. Patients in CR, CRi and CRp had a similar 2-year rates of RI (26.3%, 18.4% and 28.9%, respectively; p = 0.66), 2-year NRM (20.5%, 38.8% and 22.1%, respectively; p = 0.18) and 2-year OS (61.8%, 55.1% and 58.6%, respectively; p = 0.82). RI was significantly lower for patients in CCR pre-SCT with HR 0.25 (95% CI 0.11-0.51 p = 0.0001) without a difference in overall survival HR 0.68 (0.41-1.15; p = 0.14). However, we observed a similar RI in CR, CRi and CRp patients who had persistent cytogenetic disease pre-SCT (p = 0.55). The outcome remained similar between strata after excluding MDS patients (data not shown). Conclusion: We observed equivalent post-transplant outcomes between CR, CRi and CRp despite the presence of persistent cytogenetic disease. This suggests that RIC SCT had a role in mitigating the negative prognosis associated incomplete hematologic recovery. These findings have implications as physicians may not need to postpone SCT to await count recovery. This important question should be further validated in a larger cohort of patients. Figure 1. Figure 1. Figure 2. Figure 2. Disclosures Al-Kali: Novartis: Research Funding.

Abstract Background: Severe myelosuppression (MS) and bone marrow failure (BMF) are rare but well characterized adverse effect of temozolomide (TMZ). The existing literature is limited to case reports and data reported to regulatory agencies, which usually describe very severe cases and lack long term and detailed follow-up. Methods: We identified cases of myelosuppression attributed to TMZ seen at the Mayo clinic, from 1998-2014. We also performed a literature (PubMed) search to identify published cases of TMZ induced MS and BMF. TMZ induced MS was defined as a decrease in at least 2 cell lines below normal that was clinically attributable to TMZ and persisted for > 28 days after the last TMZ dose. Severe MS was defined as bone marrow cellularity of <25% or at least 2 of: a) reticulocyte count <40,000/µL b) absolute neutrophil count (ANC)<500/ µL c) platelets <20,000/µL. Very severe MS was defined by an ANC <200/ µL. Results: A total of 15 Mayo patients and 10 published cases were identified. The baseline characteristics of patients are shown in table 1. The median interval between initiation of TMZ and diagnosis of MS was 48 days (range 26-121). The hematologic outcomes of the group are shown in table 2. Seventeen patients developed neutropenic fever, 4 had severe sepsis, and 6 experienced bleeding complications. Kaplan-Meier recovery curves for each cell line are shown in the figure. Myelosuppression was fatal in 3 cases. Most patients recovered their ANC (median-62 days, range 28-180). Persistent thrombocytopenia was seen in 19 patients, without evidence for clonal evolution or myelodysplasia. Four patients remain dependent on growth factor support (G-CSF and thrombopoietin agonists). Conclusion: MS is usually a non-fatal complication of TMZ. Most cell lines recover spontaneously and adequately with the exception of platelets, which can remain significantly suppressed for prolonged periods. Infectious complications are very common. Most patients respond to supportive care, including the use of growth factors. Figure 3 Figure 3. Disclosures No relevant conflicts of interest to declare.