Sulai, Nanna

Abstract Abstract 3803 Background: Ring sideroblasts (RS) represent abnormal mitochondrial iron deposits that are commonly present in myelodysplastic syndromes (MDS). The presence of ≥15% RS, by definition (World Health Organization-WHO) is necessary for a diagnosis of a MDS-RS; with refractory anemia with ring sideroblasts (RARS) being a specific morphologic category. RS can also be seen with other morphologic categories such as; refractory cytopenias with multilineage dysplasia (RCMD), MDS-unclassifiable (MDS-U) and refractory anemia with excess blasts (RAEB-1/2). Amongst these categories, RARS is generally believed to have the best survival rate with the lowest risk of leukemic transformation and this has been attributed to the absence of associated dysplasia in more than one lineage. However, it has not been systematically shown whether or not the exact percentage of RS provides additional prognostic information in the context of an accurate morphologic categorization and risk stratification by karyotype or the revised IPSS (IPSS-R). Methods: 200 patients with primary MDS without excess blasts and ≥1% RS were seen at the Mayo Clinic from 1997 through 2007. All patients underwent bone marrow (BM) examination and cytogenetic evaluation at diagnosis and the pathology slides, including iron stains, were centrally re-reviewed to accurately quantify bone marrow (BM) RS percentage and to confirm WHO morphologic categories. Molecular profiling included analysis for IDH1, IDH2, JAK2 and MPL mutations. For the purposes of this study patients were divided into 4 categories based on the RS percentage; <5%, 5–14%, 15–50% and >50%. In addition, each patient was assigned IPSS and IPSS-R prognostic scores at diagnosis and risk stratified by karyotype according to the IPSS-R cytogenetic risk categories. Cox proportional regression method was used for multivariable analysis. Results: Of the overall 200 study patients, 140 (70%) were male and median age was 71 years (range, 17–90 years). At presentation, 34 (17%) patients were red cell transfusion-dependent. There were 56 (28%) patients with <5% RS (RCMD-52, MDS-U-14), 32 (16%) with 5–14% RS (RCMD-29, MDS-U-3), 79 (39%) with 15–50% RS (RARS-43, RCMD-29, MDS-U-2) and 33 (16%) with >50% RS (RARS-13, RCMD-17, MDS-U-3). Three patients (2%) were IDH1 mutant (RARS-1, RCMD-2), 17 (8%) IDH2 mutant (RARS-1, RCMD-14, MDS-U-2), 5 JAK2 V617F (3%) mutant (RARS-2, RCMD-3) and one patient with RCMD had the MPL W515L mutation. At a median follow-up of 33 months, 156 (73%) deaths and 24 (12%) leukemic transformations were documented. Median survivals were 63 months for MDS with >50% RS, 43 months for MDS with 15–50% RS, 35 months for MDS with 5–14% RS, and 14 months for MDS with <5% RS (p=0.005). In univariate analysis, additional risk factors included decreased hemoglobin, decreased platelet count, increased circulating blasts, WHO morphologic categories, red cell transfusion need, IPSS, IPSS-R and cytogenetic risk categories per IPSS-R. On a multivariable analysis only the IPSS-R prognostic score (p<0.0001) and the WHO morphologic categories (p=0.02 for RARS) retained significance. Univariate analysis disclosed significantly inferior leukemia-free survival (LFS) in patients with RCMD, thrombocytopenia, increased circulating blasts, and poor IPSS and IPSS-R prognostic scores. On multivariable analysis once again only the IPSS-R prognostic scores (p=0.02) and WHO morphologic categories (p=0.02) retained their significance. In other words, RS percentage, independent of WHO classification, as a continuous or categorical variable (i.e.: <5%, 5–14%, 15–50% & >50%) did not affect either overall survival or LFS. Conclusions: In MDS without excess blasts, once an accurate WHO morphologic categorization is made based on the presence or absence of multi-lineage dysplasia, there is no additional prognostic value for quantifying bone marrow ring sideroblasts. WHO classification and IPSS-R prognostic scores remain the most important factors in assessing patients with MDS. Disclosures: No relevant conflicts of interest to declare.

Abstract Abstract 3850 Background: We have recently shown that primary myelofibrosis (PMF) was associated with increased plasma levels of IL-1b/IL-1RA/IL-2R/IL-6/IL-8/IL-10/IL-12/IL-13/IL-15/TNF-α/G-CSF/INF-α/MIP-1α/MIP-1b/HGF/IP-10/MIG/MCP-1/VEGF in PMF (J Clin Oncol 2011; 29: 1356). In addition, increased levels of IL-8, IL-2R, IL-12, IL-15 and IP-10 were independently associated with inferior survival. In the current study, we focused on the plasma cytokine profile in polycythemia vera (PV) and examined how it compares with PMF or in a cohort of PV patients whose samples were collected during follow-up. We also looked for phenotypic and prognostic correlates. Methods: Study inclusion for patients with PV required availability of archived plasma within one year of diagnosis. Samples from PV patients seen at different points post-diagnosis were also collected, in order to compare their cytokine profile with that of patients whose samples were collected within one year of diagnosis. Standard procedures were followed to centrifuge peripheral blood samples at 4°C and store aliquots at −80°C. Concentrations of 30 plasma cytokines/chemokines were analyzed in duplicates using Multiplex Bead-based Luminex Technology (Invitrogen, Carlsbad, CA, USA): IL-1β, IL-1RA, IL-2, IL-2R, IL-4, IL-5, IL-6, IL-7, IL-8, IL-10, IL-12, IL-13, IL-15, IL-17, EGF, eotaxin, FGF-b, GM-CSF, G-CSF, HGF, IFN-α, IFN-γ, IP-10, MCP-1, MIG, MIP-1α, MIP-1β, RANTES, TNF-α, and VEGF. Measurements were performed on a Luminex 200 analyzer (Luminex Corporation, Austin, TX, USA) and resulting data were evaluated using STarStation Software Version 2.3 (Luminex Corporation, Austin, TX, USA). Results I: Plasma cytokines in PV (n=36 for samples collected within one year of diagnosis and n=40 for samples collected beyond that time point) vs. PMF (n=92) vs. normal controls (n=35) There was no significant difference between the three groups in the plasma levels of the following cytokines: IL-1β, IL-2, IL-4, EGF, and IL-17. PV and normal controls had similar levels of IL-2R, IL-5, FGF-b, and TNF-α; all four cytokines were elevated in PMF. The levels of the following cytokines were equally increased in PV and PMF: IL-1RA, IL-7, HGF, MIG, and VEGF. The following cytokines were also elevated in both PMF and PV but predominantly so in PV (MIP-1α) or PMF (IL-6, IL-8, IL-12, IL-13, IL-15, MIP-1β, IP-10, and MCP-1). Eotaxin and GM-CSF were elevated in PV but not PMF. Compared to normal controls, significantly lower levels were noted for IL-10 in PV and IFN-α and IFN-γ for PMF. IFN-α and IFN-γ levels were both elevated in PV. Comparison of PV samples collected within one year of diagnosis (n =36) vs. those collected beyond that time point (n =40) showed significantly increased levels of IL-8 (p=0.0005) and borderline significant increases in IP-10 (p=0.09) and IL-2R (p=0.09). Results II: Clinical correlates of plasma cytokines in PV Among the above-listed cytokines that were abnormally increased or decreased in PV, significant correlations were seen with sex (IP-10 and GM-CSF), age (IP-10, eotaxin), platelet count (MIG), hemoglobin level (IL-6), arterial thrombosis (IFN-γ, IL-1RA, VEGF, MIP-1α), and abnormal karyotype (IL-13, MIP-1α). None of the cytokines correlated with leukocyte count, venous thrombosis, pruritus or microvascular symptoms. The following, treated as continuous variables, were associated with inferior survival: IL-7, HGF, IFN-α, MCP-1, GM-CSF and IL-10 (p<0.05 for all). The number of events and sample size were too small to perform multivariable analysis. Conclusions: Plasma cytokine levels in PV are markedly abnormal and show both similarities and differences with PMF; the similarities became more pronounced in patients whose samples were examined at later stages of their disease. The current study suggests prognostic implications that warrant validation in a larger number of patients. Disclosures: No relevant conflicts of interest to declare.

Abstract Abstract 1751 Because IDH mutations are frequent in blast-phase myeloproliferative neoplasms (MPN), they might contribute to leukemic transformation. We examined this possibility in 301 consecutive patients with chronic-phase primary myelofibrosis (PMF). All study patients were fully characterized for karyotype, JAK2 and MPL mutational status, and Dynamic International Prognostic Scoring System-plus (DIPSS-plus) risk status. DNA from bone marrow or peripheral blood was used to screen for IDH1 and IDH2 mutations, by direct sequencing and/or high resolution melting (HRM) assay. Mutant IDH was detected in 12 patients (4%): seven IDH2 (five R140Q, one R140W and one R172G) and five IDH1 (three R132S and two R132C). MPL exon 10 was mutated in 18 patients (6.3%) and constituted W515L in 14 patients, W515K in 3 and a frameshift mutation in 1 patient. JAK2V617F was detected in 169 (56%) patients. Six patients displayed both JAK2V617F and IDH mutations (IDH2R140Q in 2 patients, IDH2R140W in 1 and IDH1R132S in 3); JAK2V617F allele burden was 1%, 7%, 22%, 27%, 30% and 96%, respectively. One patient displayed both IDHR140Q and MPLW515R. One-hundred and seven (36%) patients were negative for all three mutations. The 12 IDH-mutated patients were clinically compared to patients belonging to the three other molecular subgroups: mutated for JAK2 only (n=164), mutated for MPL only (n=18) and unmutated for all three (n=107). The four molecular subgroups were remarkably similar in their phenotype with few exceptions; IDH-mutated patients were significantly older than those with no mutations (p=0.04) whereas age distribution was similar between patients with mutant IDH, MPL or JAK2. In univariate analysis, overall survival (OS) for IDH-mutated patients was significantly shorter than those for JAK2-mutated (p=0.03), MPL-mutated (p=0.047) or unmutated (p=0.0009) patients. IDH-mutated patients also showed significantly shorter leukemia-free survival (LFS), compared to those with mutant JAK2 (p=0.0008), mutant MPL (p=0.02) or no mutations (p=0.001). After accounting for age, the presence of mutant IDH remained a significant disadvantage for both OS (p=0.04) and LFS (p=0.005). Multivariable analysis of OS that included risk categorization per DIPSS-plus confirmed the independent prognostic relevance of mutant IDH (p=0.03): HR for patients with no mutations =0.39, 95% CI 0.2–0.75; HR for JAK2-mutated patients =0.50, 95% CI 0.27–0.95; HR for MPL-mutated patients =0.53, 95% CI 0.23–1.2. A similar analysis for LFS that included risk factors for LT (i.e. unfavorable karyotype and platelet count <100 × 109/L) as covariates also confirmed the prognostic relevance of mutant IDH (p=0.003): HR for patients with no mutations =0.16; 95% CI 0.06–0.46; HR for JAK2-mutated patients =0.18; 95% CI 0.06–0.48; HR for MPL-mutated patients =0.09; 95% CI 0.01–0.76). Further analysis disclosed that the negative OS and LFS effect of mutant IDH was most pronounced in the presence (p=0.0002 and <0.0001, respectively) as opposed to the absence (p=0.34 and 0.64, respectively) of concomitant JAK2V617F expression. Analysis of paired samples obtained during the chronic and blast phases of the disease was possible in 2 IDH-mutated patients and showed the presence of both IDH and JAK2 mutations at both time points. Our observations suggest that IDH mutations in PMF are independent predictors of leukemic transformation and raise the possibility of leukemogenic collaboration with JAK2V617F. Disclosures: No relevant conflicts of interest to declare.

Abstract Abstract 2851 Background: It is widely appreciated that total cholesterol (TC) and triglyceride (TG) levels are decreased in patients with primary myelofibrosis (PMF). However, a systematic study looking into the serum lipid profile in PMF and its phenotypic and prognostic relevance is lacking. Methods: An updated Mayo Clinic database of 1008 karyotypically- and DIPSS-plus-annotated patients with PMF was used to identify study patients in whom serum lipid and lipoprotein levels were available at time of referral. Overall (OS) and leukemia-free (LFS) survivals were calculated from the time of referral, which was also the time of serum lipid analysis. Results: A total of 207 patients met the above stipulated criteria and had measurement of TC (n=207), TG (n=204), HDL cholesterol (HDL-C) (n=187), LDL cholesterol (LDL-C) (n=176) and non-HDL cholesterol (N-HDL-C) (n=184). The corresponding median (range) values (mg/dL) were 132 (57–296), 147 (42–742), 31 (8–96), 67 (11–186) and 104 (7–249). TC was significantly (P<0.05) lower in the presence of male sex, older age, red cell transfusion need, hemoglobin <10 g/dL, marked splenomegaly, circulating blasts, constitutional symptoms, JAK2 V617F, and ASXL1 mutations. TG was significantly (p<0.05) lower in the presence of JAK2 V617F, red cell transfusion need, hemoglobin <10 g/dL and thrombocytosis. DIPSS-plus risk categories showed significant differences in their TC (p<0.0001), HDL-C (p=0.004), LDL-C (p=0.004), and N-HDL-C (p=0.009) profiles with higher levels seen in lower risk patients; there were no correlations with TG levels. In univariate analysis, lower levels of TC, HDL-C, LDL-C and N-HDL-C were all associated with shortened survival (P<0.01 for each association) whereas survival was not affected by TG level (p=0.42). Multivariable analysis in the context of DIPSS-plus confirmed the independent prognostic relevance of both TC (p=0.05) and HDL-C (p=0.02) for OS. A similar analysis disclosed no correlation with LFS. Exclusion of patients receiving statin or other anti-lipid treatment did not affect the overall results. Patients in the lower quartiles for either TC (approximately <100 mg/dL) or HDL-C (approximately <25 mg/L) compared to those with higher levels displayed significantly shorter survival (Figure; RR 2.4, 95% CI 1.6–3.7; p<0.0001) that was not accounted for by DIPSS-plus (RR 1.6, 95% CI 1.1–2.5; p=0.03). Conclusions: Decreased levels of total or HDL cholesterol in PMF cluster with adverse disease features and carry a DIPSS-plus independent adverse prognostic value. Measurement of TC and HDL-C in PMF might be useful in monitoring disease progression and treatment response. Disclosures: No relevant conflicts of interest to declare.

Abstract The presence of ≥ 15% bone marrow (BM) ring sideroblasts (RS) and < 5% blasts is required for a diagnosis of refractory anemia with ring sideroblasts. We examined the phenotypic and prognostic relevance of this “15%” RS threshold in 200 patients with myelodysplastic syndromes (MDS) without excess blasts and with ≥ 1% RS. The impact of RS% was assessed both as a continuous and categorical variable: < 5% (n = 56), 5%-14% (n = 32), 15%-50% (n = 79), and > 50% (n = 33). RS% correlated (P < .05) directly with age, platelet count, transfusion dependency, BM cellularity, and mutant SF3B1 and inversely with hemoglobin level, multilineage dysplasia, and high-risk karyotype; but did not correlate with IDH mutations. At a median follow-up of 33 months, 156 (73%) deaths and 24 (12%) leukemic transformations were documented. Neither univariate nor multivariable analysis showed significant effect for RS% on overall or leukemia-free survival, suggesting the limited prognostic value of quantifying BM RS in MDS.