Pardanani, Animesh

Abstract Background A recent study from the International Working Group on Myeloproliferative Neoplasms Research and Treatment (IWG-MRT) identified prior arterial events and hypertension as risk factors for subsequent arterial thrombosis and prior venous events and age 65 years or older as risk factors for venous thrombosis in polycythemia vera (PV) (Blood 2014;124:3021). In the current single center study, we sought to validate the aforementioned findings from the IWG-MRT and look into further details regarding arterial versus venous thrombosis in PV. Methods Study patients were selected from our institutional database of myeloproliferative neoplasms (MPN) and fulfilled the 2008 World Health Organization (WHO) criteria for the diagnosis of PV (Blood. 2009;114:937). Screening for the two most frequent mutations other than JAK2 (TET2 and ASXL1) were performed according to conventional methods (Leukemia. 2014;28:2206). Statistical analyses considered clinical and laboratory parameters obtained at time of diagnosis. Results Patient characteristics: Analysis was conducted on 587 patients (median age 60 years; 48% males). ASXL1 and TET2 mutational status was available in 133 patients. Among informative cases, 31% had palpable splenomegaly, 34% microcirculatory symptoms, 30% pruritus, 8% erythromelalgia, 42% hypertension, 9% diabetes and 11% were active tobacco users. TET2 and ASXL1 mutations were documented in 18% and 11%, respectively. During follow-up, 224 (38%) patients died and median follow-up for living patients was 109 months. Median survival was 16 years and leukemic or fibrotic transformations were documented in 4% and 14%, respectively. Thrombotic events: A total of 235 (40%) patients experienced at least one thrombotic event before or after diagnosis and included 153 (26%) arterial and 104 (18%) venous events. Thrombotic events were recorded before or at time of diagnosis in 146 (25%) patients and included 97 (17%) arterial and 55 (9%) venous events. Thrombotic events after diagnosis were recorded in 128 (22%) patients and included 82 (14%) arterial and 57 (10%) venous events. Associations for arterial, venous or all thrombotic events occurring at any time before or after time of diagnosis: The occurrence of any thrombotic event before or after diagnosis correlated with active tobacco use (p=0.04), diabetes history (p=0.03), hyperlipidemia (p=0.04) and major hemorrhage (p=0.03). The occurrence of any arterial event before or after diagnosis correlated with advanced age (p=0.006), diabetes history (p=0.005), hypertension history (p=0.004) and hyperlipidemia (p<0.0001). The occurrence of any venous event before or after diagnosis correlated with female sex (p=0.008), younger age (p<0.0001), lower platelet count (p=0.0014) and palpable splenomegaly (p=0.006). TET2 or ASXL1 mutations did not correlate with overall, arterial or venous thrombosis. Risk factors for arterial versus venous events after time of diagnosis: In univariate/multivariable analysis, thrombosis-free survival (TFS) was adversely affected by prior vascular events (p=0.002/0.03 multivariable) and increased leukocyte count (p=0.03/0.03); arterial thrombosis-free survival was adversely affected by prior arterial events (p<0.0001/P<0.0001), hyperlipidemia (p=0.001/0.03), hypertension (p=0.02/NS) and advanced age (p=0.02/NS); venous thrombosis-free survival was adversely affected by prior venous events (p=0.04/0.05), increased leukocyte count (p=0.0009/0.002), and younger age (p=0.02/NS). In addition, major hemorrhage at diagnosis was associated subsequent venous thrombosis in both univariate and multivariable analysis. TET2 or ASXL1 mutations did not affect the risk of overall, arterial or venous thrombosis. Conclusions In contemporarily managed patients with PV, prior vascular events predict future events for both arterial and venous thrombosis; additional risk factors included hyperlipidemia for arterial and increased leukocyte count for venous thrombosis. The current study also identifies association of arterial thrombosis in general with older age and history of hypertension, diabetes or hyperlipidemia whereas venous thrombosis was associated with younger age, female sex and palpable splenomegaly. Additional findings included association of major hemorrhage with thrombosis and higher platelet count with lower risk of venous thrombosis. Disclosures No relevant conflicts of interest to declare.

Abstract Background The International Working Group for Myeloproliferative Neoplasms (MPN) Research and Treatment (IWG-MRT) previously published on 1104 patients with "essential thrombocythemia (ET)" and identified 180 (16%) misdiagnosed cases with early/prefibrotic primary myelofibrosis (PMF); the latter diagnosis was associated with inferior overall (OS), leukemia-free (LFS) and myelofibrosis-free (MFFS) survival (J Clin Oncol. 2011; 29:3179). Among the 891 patients with "true" World Health Organization (WHO)-defined ET, risk factors for OS included age ≥60 years, leukocyte count ≥11 x 10(9)/L and history of thrombosis (Blood2012120:1197). In the current study, we explored the possibility that increased serum lactate dehydrogenase (LDH) level or mild grade-1 reticulin fibrosis might identify occult cases of early/prefibrotic PMF in otherwise WHO-defined ET and contribute to disease prognostication. Methods Study patients were selected from the Mayo Clinic institutional database of MPN based on their meeting the 2008 WHO criteria for diagnosis of ET (Blood. 2009;114:937). The degree of bone marrow (BM) reticulin fibrosis was based on "real life" BM reports from Mayo Clinic hematopathologists and often in accordance with the European consensus scoring system (Haematologica 2005;90:1128). Serum LDH levels were obtained at time of diagnosis or within 12 months of diagnosis, in the absence of any treatment. Information on the two most frequent non-driver mutations seen in ET (i.e. TET2 and ASXL1) was available in a subset of the patients (Blood 2015 126:354). Statistical analyses considered clinical and laboratory parameters obtained at time of diagnosis Results Patient characteristics: 183 patients (57% females) were evaluable for BM reticulin grading; median (range) levels were for age 59 years (15-88), hemoglobin 14 g/dL (9.1-17.1), platelets 812 x 10(9)/L (454-3300) and leukocyte count 8.5 x 10(9)/L (3.1-31.5). Palpable splenomegaly was reported in 27 (15%) patients and 12% displayed leukoerythroblastosis (LES). Driver mutational status was JAK2 in 72%, CALR 17%, MPL 2% and triple-negative in 9%. Thrombosis history at diagnosis was obtained from 13% of the patients and 12% experienced the same post diagnosis. Risk stratification according to the International Prognostic Scoring System for ET (IPSET) was 19% high, 43% intermediate and 38% low (Blood2012120:1197). BM reticulin fibrosis was graded "0" in 113 (62%) patients and "1" in 70 (38%). Serum LDH information was available in 110 patients (median194 U/L; range 135-669) and was increased in 31 (28%) patients. Mutational status for TET2 and ASXL1was available in 76 patients with mutational frequencies of 8% each. Clinical and laboratory correlates for mild reticulin fibrosis and serum LDH: We found no significant correlation between mild reticulin fibrosis (grade-1) and a number of clinical parameters, including age, sex, hemoglobin level, leukocyte count, platelet count, driver mutational status, LES, presence of palpable splenomegaly, thrombosis history or serum LDH (p=0.36). In contrast, increased serum LDH was associated with increased leukocyte count (p=0.002), increased platelet count (p<0.001), presence of palpable splenomegaly (p=0.03) and higher IPSET score (p=0.002); there were no significant correlations with age, sex, hemoglobin level, LES, thrombosis history, driver mutational status or TET2/ASXL1mutations. Survival analysis: After a median follow-up of 91 months, 42 (23%) deaths, 2 (1%) leukemic transformations, 7 (4%) fibrotic progressions and 22 (12%) thrombotic events were documented. In univariate analysis, risk factors for OS included age ≥60 years (p=0.002; HR 10.2, 95% CI 2.3-44.6), male sex (p=0.02; HR 3.2, 95% CI 1.2-8.2), leukocyte count ≥15 x 10(9)/L (p=0.007; HR 4.7, 95% CI 1.5-14.6), and increased serum LDH (p=0.004; HR 3.7, 95% CI 1.5-9.1), but not BM reticulin fibrosis (p=0.17). In multivariable analysis, age, sex and LDH remained significant; serum LDH also remained significant for OS, in the context of IPSET, whereas it did not appear to influence thrombosis-free survival (p=0.58), LFS (p=0.84) or MFFS (p=0.34). Conclusions The current study suggests that serum LDH might supersede leukocyte count as an independent risk factor for overall survival in ET; in contrast, mild reticulin fibrosis was irrelevant in terms of both prognostic value and disease phenotype. Disclosures No relevant conflicts of interest to declare.

Abstract Background In polycythemia vera (PV), abnormal karyotype at diagnosis has been reported in 10% to 20% of patients and some studies have suggested an association with inferior survival (Leukemia2013;27:1874) . In the current study, we examined the prognostic contribution of abnormal karyotype, in general, and specific abnormalities, in particular, in newly diagnosed PV. Methods Study patients were selected from our institutional database of myeloproliferative neoplasms (MPN) and fulfilled the 2008 World Health Organization (WHO) criteria for diagnosis of PV (Blood. 2009;114:937). Cytogenetic analysis and reporting was done according to the International System for Human Cytogenetic Nomenclature (Cytogenetic and Genome Research2013;141:1-6). Assignment as "unfavorable karyotype" was according to criteria established for PMF: favorable = normal + favorable abnormalities; unfavorable = unfavorable abnormalities (Leukemia. 2011;25:82). Screening for the two most frequent mutations in PV, other than JAK2 (i.e. TET2 and ASXL1), were performed according to conventional methods (Leukemia. 2014;28:2206). Statistical analyses considered clinical and laboratory parameters obtained at time of diagnosis. Results Patient characteristics: Median (range) values for the 239 study patients (53% females) included: age 62 (17-94), leukocyte count 11.7 x 10(9)/L (4.3-59.3) and platelet count 479 x 10(9)/L (37-2747). Palpable splenomegaly was present in 26% of the patients, pruritus in 27%, erythromelalgia in in 8%, hypertension in 47%, diabetes in 10% and hyperlipidemia in 28%. Thrombosis history at diagnosis was documented in 33% of the patients and 20% experienced the same after diagnosis. 73% of the patients were "high risk" by conventional risk stratification. Mutation screening for TET2 and ASXL1 was performed in 80 patients and mutational frequencies were 19% and 11% respectively. All study patients provided cytogenetic information, which was abnormal in 46 (19%) patients. The most frequent abnormalities were isolated +9 (n=11; 24% of abnormal karyotype and considered favorable), isolated del(20q) (n=8; 17% of abnormal karyotype and considered favorable), isolated loss of Y chromosome (n=7; 15% of abnormal karyotype and considered favorable) and isolated +8 (n=5; 11% of abnormal karyotype and considered unfavorable). A total of 9 (20% of abnormal karyotype) patients displayed unfavorable karyotype that included +8 in 7 patients, del(11q) in one patient and +20 in one patient. After a median follow-up of 83 months, 70 (29%) deaths, 48 (20%) thrombotic events, 20 (8%) fibrotic progressions and 7 (3%) leukemic transformations were documented. Comparison of patients with and without cytogenetic abnormalities Patients with abnormal cytogenetics were older (p=0.048), displayed lower platelet count (p=0.005) and were more likely to be high risk (p=0.02); there was no significant correlation with TET2 or ASXL1 mutation. In univariate analysis, patients with abnormal cytogenetics displayed inferior leukemia-free (p=0.007; HR 10.6, 95% CI 1.9-58.7), myelofibrosis-free (p<0.0001; HR 7.7, 95% CI 3.1-19.3) and overall (p=0.13; HR 1.6, 95% CI 0.9-2.8) survival. Furthermore, the difference in overall survival became significant when cytogenetic abnormalities were classified into unfavorable (p=0.006) and favorable (p=0.6) categories. On the other hand, inferior leukemia-free and myelofibrosis-free survival was noted in both patients with favorable and unfavorable cytogenetic abnormalities, when compared to normal karyotype. During multivariable analysis that included age and leukocytosis (≥15 x 10 (9)/L) as covariates, the adverse effect of abnormal cytogenetics on leukemia-free (p=0.009) or myelofibrosis-free (p<0.0001) survival and that of unfavorable karyotype on overall (p=0.05) survival were shown to be independent. Finally, patients with abnormal cytogenetics were less likely to experience thrombosis after diagnosis (p=0.04; HR 0.3, 95% CI 0.09-0.97), an effect that was independent of both age and thrombosis history. Conclusions Cytogenetic abnormalities in PV confer an independent adverse prognostic effect on overall, leukemia-free and myelofibrosis-free survival, but not thrombosis-free survival; the adverse effect on leukemia-free and myelofibrosis-free survival was seen with both favorable and unfavorable cytogenetic abnormalities. Disclosures No relevant conflicts of interest to declare.

Abstract Background The International Working Group for Myeloproliferative Neoplasms (MPN) Research and Treatment (IWG-MRT) has previously identified older age, leukocytosis, venous thrombosis and abnormal karyotype as risk factors for overall survival (OS) in polycythemia vera (PV), and older age, abnormal karyotype and leukocytes ≥15 × 109/l as risk factors for leukemia-free survival (LFS) (Leukemia2013;27:1874). Serum lactate dehydrogenase (LDH) is a surrogate quantitative measure of cell turnover and tumor burden. We hypothesized that, compared to leukocyte count, serum LDH might be a biologically more accurate indicator of tumor aggression and sought to examine its potential utility as a prognostic biomarker in PV. Methods Study patients were selected from our institutional database of MPN based on their meeting the 2008 or 2016 World Health Organization criteria for diagnosis of PV (Blood. 2009;114:937; Blood 2016 127:2391) and availability of serum LDH level, obtained within six months of diagnosis. Information on karyotype and targeted next-generation sequencing was available in a subset of the patients (Blood 2015 126:354). Statistical analyses considered clinical and laboratory parameters obtained at time of diagnosis. Age and leukocyte count prognostic categories were based on previously published IWG-MRT study (Leukemia2013;27:1874). Results Patient characteristics: A total of 216 patients (50% females) met the above-outlined criteria: median (10th-90th percentile) levels were for age 63 years (38-81), hemoglobin 18 g/dL (16.6-20.7), platelets 467 x 10(9)/L (237-833), leukocyte count 11.7 x 10(9)/L (7.2-20.3) and spleen size 0 cm (0-2). Pruritus was documented in 28%, microcirculatory symptoms in 31%, erythromelalgia in 6%, hypertension in 45%, diabetes in 7%, active tobacco use in 13%, hyperlipidemia in 25%, palpable spleen in 26% and abnormal karyotype in 15% of informative cases. Grading for bone marrow reticulin fibrosis was documented for 144 patients: 50% grade "0", 42% grade "1", 7% grade "2" and 1 grade "3". Thrombosis history at diagnosis was documented in 26% of patients and occurred after diagnosis in 19%. Mutation analysis was available in 72 patients and revealed TET2 mutations in 18 (25%), ASXL1 in 5 (7%), SRSF2 in 2 (3%) and IDH2 in one (1.4%). Serum LDH levels and correlates: Median serum LDH level was 226 U/L (range 71-1008); the upper normal limit (UNL) for Mayo Clinic was 222 U/L. Increased serum LDH was recorded in 110 (51%) patients that included 26 (12%) patients with LDH ≥1.5 x UNL. A significant correlation was demonstrated between increased serum LDH and older age (p=0.007), female sex (p=0.04), leukocytosis (p=0.01), venous thrombosis history (p=0.005) and, interestingly, absence of active tobacco use (p=0.003); no correlation was noted for bone marrow reticulin fibrosis, presence of palpable splenomegaly, abnormal karyotype or ASXL1 mutation. Survival analysis: After a median follow-up of 77 months, 82 (38%) deaths, 8 (3.7%) leukemic transformations, 13 (6%) fibrotic progressions and 41 (19%) thrombotic events were documented. In univariate analysis, the following adversely affected OS: advanced age (P<0.0001), increased LDH as a continuous variable (p=0.0006), LDH ≥1.5 x UNL (p<0.0001), leukocyte count ≥15 x 10(9)/L (p=0.02) and arterial thrombosis history (p=0.001). In multivariable analysis, only age and LDH remained significant; multivariable HR (95% CI) were 5.4 (2.8-10.4) for age ≥67 years, 3.2 (1.8-6.0) for LDH ≥1.5 x UNL and 2.4 (1.2-4.7) for age 57 to 66 years. In the subset of patients with available information on karyotype and non-JAK2 mutations (n=72), advanced age, LDH ≥1.5 x UNL and ASXL1 mutations remained significantly associated with inferior survival. Serum LDH also predicted leukemic transformation (HR 17.8, 95% CI 4.1-78) and fibrotic progression (HR 11.6, 95% CI 25-53.7); in multivariable analysis, only serum LDH ≥1.5 x UNL (HR 22.3, 95% CI 2.1-234.6) and abnormal karyotype (HR 9.5, 95% CI 1.2-74.7) predicted leukemic transformation and only the former predicted fibrotic progression. Conclusions Serum LDH is a simple and inexpensive biomarker that might supersede leukocyte count as an independent predictor of overall, leukemia-free and myelofibrosis-free survival in PV; advanced age and ASXL1 mutations also retained significance for OS and abnormal karyotype for LFS. Disclosures No relevant conflicts of interest to declare.

Abstract Background: We sought to retrospectively analyze the clinical differences, at diagnosis and throughout the disease course, between patients with primary myeloifbrosis whom undergo blastic transformation (PMF-BP) and those whom expire from PMF complications without undergoing transformation. Methods: An international collaborative database of patients with PMF which progressed to PMF BP, and a control group of individuals with PMF who expired secondary to PMF was created. Data regarding clinical course, bone marrow, karyotypic, quantitative JAK2V617F analysis (when able), laboratory values, and therapy at the diagnosis of PMF, PMF BP and up to 4 return visits (R1-4) in between these milestones was abstracted. Results: COMPARISON AT DIAGNOSIS OF PMF: 136 cases of PMF who eventually underwent PMF-BP were included, with a control group of 42 PMF patients, were analyzed. Both groups of patients had similar demographic (median age at diagnosis 59 years (range 23–83)/ 59.1 (15–93); and disease parameters (median hemoglobin 10 g/dL (range 2–15.7) /9.8 (4.9–14.6); median platelet count 181 x 109/L (range 7–1400)/168 (16–1916)), and Lille PMF prognostic scores (p = n.s.) for PMF-BP vs. PMF control group respectively. Additionally, blasts (peripheral blood /marrow) were a median of 0% (range 0–18%)/ 2% (range 0–16%) for the study group versus 0.25% (range 0–5%)/ 3% (range 0–4%) (p = n.s. for both) for the control group, respectively. Both groups had similar rates of requiring therapy (86%; 93%) and survival from diagnosis (median 34 months (range 2–441); 29 months (1–236)) (p = n.s.). However, lactate dehydrogenase (LDH) (although high in both groups) was higher at diagnosis (median 802 IU/L (range 83–10353) vs. 416 IU/L (124–2197)) (p=0.04), as well as an abnormal marrow karyotype (85% vs. 25%; p<0.001) amongst the PMF BP group. SUBSEQUENT CLINICAL COURSE: Analyzing the clinical evolution between groups demonstrated persistently higher LDH and worsening thrombocytopenia in the PMF-BP cohort (see figure). Decreasing Platelets Approaching PMF BP Decreasing Platelets Approaching PMF BP Peripheral blood blasts increased in both groups, but values above 10% were unique to the PMF BP group. Additionally marrow karyotype displayed clonal evolution developed in 56% of the PMF-BP patients as opposed to 14% of controls (p<0.001). Initial and serial quantitative JAK2V617F mutation analysis (available from 20 /19 patients from the PMF-BP and control group at diagnosis, with 11 / 12 serial samples) showed no difference between the groups, nor a pattern of mutation burden elevation with transformation to leukemia in the study group. Conclusions: Increasing LDH, peripheral blood blast percentages >10%, and clonal evolution are features commonly seen in PMF patients whom evolve to blast phase and may be a harbinger of movement towards PMF BP.

Abstract Background The impact of JAK inhibitor (JAKi) therapy on the natural history of myelofibrosis remains unclear. While a Phase 3 study showed that overall survival of ruxolitinib-treated intermediate-2 or high-risk myelofibrosis patients was superior as compared to placebo-treated patients, it remains unclear whether such an advantage would persist in patients receiving standard therapy, and whether this result is generalizable to other JAK inhibitors. We conducted a sponsor-independent, single-center retrospective analysis of overall and leukemia-free survival after long-term follow up in patients receiving JAKi treatment with comparison to two control populations: (i) a contemporaneously diagnosed (2005 onwards) primary myelofibrosis (PMF) patient cohort receiving standard therapy (JAKi and pomalidomide naïve); and (ii) a pomalidomide treated patient cohort (JAKi-naïve). Methods Baseline clinical, cytogenetic and molecular data were collected at the onset of study treatment with the first JAKi or pomalidomide, or at the time of first referral for patients receiving standard therapy. Information regarding bone marrow karyotype and DIPSS-plus risk status was available for all patients. Information on survival and leukemic transformation was updated in July 2013. Results The 3 patient groups were: (i) JAKi-treated: n=157, median age 65 years, range 34-89. DIPSS-plus risk: Low/Int-1 20 (13%), Int-2 66 (42%), and High 71 (45%). Median follow up from first JAKi treatment was 30 months (range 1-67 months); during this period, 72 deaths (46%) and 14 (9%) leukemic transformations were documented. The first JAKi was ruxolitinib in 51 patients (33%), momelotinib (CYT387) in 79 (50%), and fedratinib (SAR302503) in 27 (17%). The proportion of patient deaths and leukemic transformation (median follow up, range) per specific JAKi was: ruxolitinib 57% and 12% (40 months, 2-67), momelotinib 43% and 9% (29 months, 1-42), and fedratinib 33% and 4% (21 months, 2-53), respectively. (ii) Control#1 (Pomalidomide-treated): n=69, median age 67 years, range 36-87. DIPSS-plus risk: Low/Int-1 0 (0%), Int-2 28 (41%), and High 41 (59%). Median follow up was 25 months (range 2-70 months); during this period, 47 deaths (68%) and 7 (10%) leukemic transformations were documented. (iii) Control# 2 (Standard therapy): n=319, median age 66 years, range 22-90. DIPSS-plus risk: Low/Int-1 98 (31%), Int-2 122 (38%), and High 99 (31%). Median follow up from time of referral was 18 months (range 1-92 months); during this period, 101 deaths (32%) and 18 (6%) leukemic transformations were documented. For patients with Int-2 or high-risk disease (n=221), 92 deaths (42%) and 13 (6%) leukemic transformations were recorded (median follow up 16 months, range 1-92). The survival analysis focused on DIPSS-plus Int-2 and high-risk patients for all groups, reflecting the population predominantly selected for treatment on JAKi clinical trials. There was no significant difference in the median overall survival of myelofibrosis patients treated with JAKi (39 months) versus standard therapy (35 months) versus pomalidomide (26 months) (p=0.12) (Figure). The 5-year survival rates were 31%, 33% and 26%, respectively. Similarly, the median overall survival of patients treated with ruxolitinib (35 months), momelotinib (37 months) or fedratinib (42 months) was not significantly different (p=0.4). Baseline DIPSS-plus status (ie, Int-2 vs. high-risk) effectively stratified JAKi treated patients in terms of overall survival (median survival 31 vs. 60 months; p=0.0002). There was no significant difference in leukemia-free survival when comparing JAKi-treated patients versus standard therapy verus pomalidomide-treated patients (p=0.6) (Figure). The estimated 5-year risk of leukemic transformation was 17%, 12% and 13%, respectively. Conclusions With long-term follow up, myelofibrosis patients receiving JAKi therapy exhibited similar overall and leukemia-free survival as compared to two control populations (DIPSS-plus matched standard therapy and pomalidomide-treated patients). Baseline DIPSS-plus status was a powerful predictor of overall survival in JAKi treated patients. While underscoring the limitation of a non-head-to-head retrospective comparison, there was no significant difference in overall or leukemia-free survival when comparing individual JAKi drugs. Disclosures: Pardanani: Bristol Myers Squibb: Clinical trial support Other; Sanofi: Clinical trial support, Publication support services, Clinical trial support, Publication support services Other; PharmaMar: Clinical trial support, Clinical trial support Other; JW Pharmaceutical Corp.: Clinical trial support, Clinical trial support Other. Off Label Use: Use of Ruxolitinib, Momelotinib, Fedratinib and Pomalidomide for treatment of Myelofibrosis in Clinical Trial setting.

Abstract Background JAK inhibitors (JAKi) currently represent a frontline treatment modality in patients with symptomatic intermediate- or high-risk myelofibrosis (MF). While the efficacy of JAKi in decreasing spleen size and improving disease-related symptoms is well documented, the effect of this drug class on overall survival and risk of leukemic transformation in MF patients is less well understood, primarily due to short follow up in individual patient cohorts. Furthermore, there is scant information on baseline predictors of overall and leukemia-free survival, and also spleen and/or anemia responses, at the onset of JAKi therapy. We conducted a sponsor-independent single-center retrospective review of patients treated with JAKi with long follow up to address the aforementioned issues. Here, we report our findings regarding baseline predictors of clinical outcome and treatment response in MF patients treated with JAKi. Methods Clinical, cytogenetic and molecular data were retrospectively collected on patients treated with one or more JAKi on a clinical trial at the Mayo Clinic, Rochester, MN. Baseline clinical data pertained to the onset of treatment with the first JAKi. Karyotype information was available for all patients. Screening for JAK2V617F, MPLW515, ASXL1, and SRSF2 mutations was performed as previously described (Leukemia. 2013 Apr 26. doi: 10.1038/leu.2013.119). Spleen size was measured by palpation by clinical investigators. Anemia and spleen responses were adjudicated as per IWG criteria (Tefferi, Blood, 2006). Information on survival and leukemic transformation was updated in July 2013. Results A total of 157 MF patients were studied (60% male; median age 65 years, range 34-89 years). Ninety seven patients (62%) had primary MF, 42 (27%) post-polycythemia vera MF, and 18 (11%) post-essential thrombocythemia MF. The DIPSS-plus distribution was: Low/Int-1 20 (13%), Int-2 66 (42%), and High 71 (45%). Other characteristics were: JAK2V617F positive 78%, unfavorable karyotype 17%, hemoglobin <10 g/dL 58%, leukocyte count >25 x 109/L 31%, platelet count <100 x 109/L 15%, peripheral blasts ≥1% 69%, red blood cell transfusion-dependent 37%, and constitutional symptoms 56%. Forty one patients (38%) were ASXL1 mutated (n=108) and 20 (18%) SRSF2 mutated (n=110). One hundred and thirty nine patients (89%) were evaluable for spleen response (median spleen size by palpation 19 cm, range 6-32 cm) and 91 (58%) for anemia response by IWG criteria. The first JAKi was momelotinib (CYT387) in 79 patients (50%), ruxolitinib in 51 (33%) and fedratinib (SAR302503) in 27 (17%). Median follow up from first JAKi treatment was 30 months (range 1-67 months); during this period, 72 deaths (46%) and 14 (9%) leukemic transformations have been documented. The median overall survival of the entire cohort was 41 months. Multivariable analysis identified the following DIPSS-plus independent predictors for survival: palpable spleen size >20 cm (HR 2.5), ASXL1 mutated status (HR 3.1) and SRSF2 mutated status (HR 3.8) (all p<0.01) (Figure). In contrast, there was no difference in survival with regards to the first JAKi used for MF treatment (p=0.3). Multivariable analysis identified the following predictors for leukemia-free survival: SRSF2 mutated (HR 9.7), unfavorable karyotype (HR 8.7), and palpable spleen size >20 cm (HR 9.1) (all p<0.01). Predictors of spleen response included baseline spleen size (≤20 cm 56%, >20 cm 34%; p=0.01), with borderline significance for ASXL1 mutation status (unmutated 51%, mutated 31%; p=0.06). Predictors of anemia response included RBC transfusion status at baseline (transfusion dependent 38%, transfusion independent 12%; p<0.01) and JAK2V617F status (unmutated 46%, mutated 22%; p=0.02). Conclusions Larger baseline spleen size and ASXL1 and SRSF2 mutated status predicted for inferior overall survival, independent of DIPSS-plus, at the onset of first JAKi treatment in MF patients. Similarly, multivariable analysis identified larger baseline spleen size and presence of unfavorable karyotype and SRSF2 mutations as being predictive for inferior leukemia-free survival. Spleen response rates were significantly superior in patients with smaller baseline spleen size and possibly ASXL1 unmutated status. Anemia response rates were significantly higher in patients who were transfusion-dependent at baseline and had JAK2V617F unmutated status. Disclosures: Pardanani: Bristol Myers Squibb: Clinical trial support Other; Sanofi: Clinical trial support, Publication support services, Clinical trial support, Publication support services Other; PharmaMar: Clinical trial support, Clinical trial support Other; JW Pharmaceutical Corp.: Clinical trial support, Clinical trial support Other. Off Label Use: Use of Ruxolitinib, Momelotinib, Fedratinib and Pomalidomide for treatment of Myelofibrosis in Clinical Trial setting.

Abstract Background: The combination of MPD and amyloidosis is rare, and this patient population's clinical outcome is not well studied. Methods: Pts with a MPD and amyloidosis were identified via an clinical note search engine that searches through the electronic medical records of patients seen at Mayo Clinic Rochester, Jacksonville, and Scottsdale between 1990 and 2016. Terms used included amyloid or amyloidosis, chronic myelogenous leukemia or CML, essential thrombocytopenia or ET, and polycythemia vera or PV. Demographic and clinical data were abstracted from the medical record. Pts with both disorders were analyzed and their mortality rates along with median time to death were calculated. Prevalences at the Mayo Clinic were calculated for the years 2014 and 2015. Results: Twenty-three pts diagnosed with both a MPD and amyloidosis were identified. Thirteen (56.5%) were male, 10 (43%) were female. Eleven (47.8%) were initially diagnosed at the Mayo Clinic. Types of amyloidosis were as follows: Eleven (47%) had immunoglobulin light-chain (AL), four had localized (17%), two (8.7%) had wild-type transthyretin (ATTR), one (4.3%) had mutant ATTR, and five (21.8%) were unknown. Types of MPD were as follows: Seven (30%) had polycythemia vera (PV), seven (30%) had chronic myelogenous leukemia (CML), five (22%) had myelofibrosis, and four (17%) had essential thrombocytosis (ET). Fifteen (65%) were initially diagnosed with a MPD. Median time to last follow-up from second diagnosis was 1.7 years, and median time to death following second diagnosis was 1.4 years. The mortality rate was 87% in the total population. The median time to death for AL and PV was 2.7 years, AL and CML 1.0 years, and AL and ET 1.17 years. Myelofibrosis did not occur with AL. The most common combination was AL and PV, which accounted for five (22%) of the cases. Treatment regimens for the patients with AL were varied. Multiple drugs were combined with dexamethasone including melphalan, velcade, pomalidomide, lenalidomide, doxorubicin, and revlimid. Cyclophosphamide, bortezomib, and dexamethasone (CyBorD) were used in two cases. The mean time to diagnosis of amyloidosis from symptom onset was 10.3 months. Prevalences of AL and a MPD in 2014 and 2015 at the Mayo Clinic were 2.8% and 1.5%, respectively. Conclusions: The mortality rate for the combined diagnoses is high. The most common combination of diagnoses was that of AL and PV, which was associated with a mean time to death of 2.7 years. Disclosures Al-Kali: Celgene: Research Funding; Onconova Therapeutics, Inc.: Research Funding.

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 Spliceosome pathway gene mutations are recurrent in myeloid malignancies with the highest frequencies reported for myelodysplastic syndromes associated with ring sideroblasts (MDS-RS; 85%), MDS without RS (44%) and chronic myelomonocytic leukemia (CMML; 55%) (Nature 2011;478:64). SF3B1 mutations were the most frequent (75%) in MDS-RS and SRSF2 mutations in CMML (28%); U2AF1 mutational frequencies were 12% in MDS without RS and 8% in CMML. We have previously described SRSF2 (Blood. 2012;120:4168) and SF3B1 (Leukemia. 2012;26:1135) mutations in 17% and 6.5% of patients with primary myelofibrosis (PMF); prognostic relevance was shown for the former but not the latter. Objectives The objectives of the current study were to i) describe the incidence of U2AF1 mutations in PMF and their correlation with clinical features, karyotype and other mutations and ii) examine the prognostic significance of U2AF1 mutations in PMF, in the context of both conventional prognostic models and other prognostically-relevant mutations. Methods Information on clinical and laboratory parameters and karyotype was available in all study patients, at time of referral, which coincided with time of sample collection for mutation screening. Risk stratification was according to the Dynamic International Prognostic Scoring System (DIPSS)-plus system. U2AF1 and other mutations were analyzed using standard PCR techniques and bidirectional sequencing; for U2AF1, two hot spots that included residues S34 and Q157 were amplified. Results A total of 251 PMF patients (median age 63 years; 160 males) were studied. DIPSS-plus risk distribution was high in 32%, intermediate-2 in 38%, intermediate-1 in 17% and low in 13% of the patients. The frequency of each DIPSS-plus adverse feature was as follows: age >65 years (42%), transfusion need (32%), hemoglobin <10 g/dL (47%), leukocyte count >25 x 10(9)/L, (16%), platelet count <100 x 10(9)/L (22%), ≥1% blasts (57%), constitutional symptoms (35%), and unfavorable karyotype (9%). Karyotype was normal in 156 (63%) patients. At a median follow-up of 48 months, 158 (63%) deaths and 27 (11%) leukemic transformations were recorded. Mutational frequencies Forty-one (16.3%) patients harbored U2AF1 mutations: 16 (39%) Q157P, 10 (24%) Q157R, 8 (20%) S34F, 4 (10%) S34Y and one each for Q157P/E159A, Q157R/S34Y and Q157-Y158insYE. Frequencies for other mutations were 11% for SRSF2, 7.3% for SF3B1, 31% for ASXL1, 5.5% for EZH2, 5% for IDH1/2 and 58% for JAK2V617F. U2AF1 mutations were usually, but not always, exclusive of other spliceosomal mutations: one patient expressed all three spliceosomal mutations. The frequency of any one of the three spliceosomal mutations was 34%. Clinical correlates U2AF1 mutations were significantly associated with older age (p=0.02), JAK2V617F (p=0.002), mutant ASXL1 (p=0.04), transfusion need (p<0.0001), hemoglobin <10 g/dL (p<0.0001), platelets <100 x 10(9)/L (p<0.0001) and normal karyotype (p=0.006). The associations with anemia, thrombocytopenia, JAK2V617F and normal karyotype were inter-independent; the frequency of U2AF1 mutations in the presence of anemia was 29%, transfusion need 36%, thrombocytopenia 35%, JAK2V617F 23% and normal karyotype 21%. Prognostic interactions U2AF1 mutations were associated with inferior overall (p=0.004) but not leukemia-free (p=0.6) survival. However, the survival association was fully accounted for by the above-mentioned clustering of U2AF1 mutations with anemia and thrombocytopenia. Similarly, although multivariable analysis of U2AF1, SRSF2, ASXL1, EZH2 and IDH mutations identified the first three as being independently predictive of poor survival, only ASXL1 and SRSF2 remained significant when either anemia or thrombocytopenia was included as a co-variate. Conclusions U2AF1 mutations are the most frequent spliceosome pathway mutations in PMF, cluster with normal karyotype and JAK2V617F, and are strongly associated with anemia and thrombocytopenia; the latter suggests a pathogenetic contribution to ineffective hematopoiesis in PMF. The current study also suggests that more than one third of patients with PMF carry a spliceosome mutation. Disclosures: No relevant conflicts of interest to declare.