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Monoclonal gammopathy of uncertain significance (MGUS) may become symptomatic for autoimmune manifestations. We report on the prevalence and clinical course of immune thrombocytopenic purpura (ITP) observed in a consecutive series of 228 MGUS patients. At MGUS diagnosis, ITP was determined in 6/228 cases, accounting for a prevalence of 2630/100 000 [95% confidence interval (CI): 1210-5620]. One incidental ITP case occurred after 21 months of follow-up. After a follow-up of 681.3 patient-years, the crude incidence of ITP in MGUS was 146.8 per 100 000 patient-year (95% CI: 3.7-817.8). Overall, these observations point to an association between MGUS and ITP.

The aim of this study was to investigate the effects of a non-standard, intermittent imatinib treatment in elderly patients with Philadelphia-positive chronic myeloid leukaemia and to answer the question on which dose should be used once a stable optimal response has been achieved. Seventy-six patients aged ⩾65 years in optimal and stable response with ⩾2 years of standard imatinib treatment were enrolled in a study testing a regimen of intermittent imatinib (INTERIM; 1-month on and 1-month off). With a minimum follow-up of 6 years, 16/76 patients (21%) have lost complete cytogenetic response (CCyR) and major molecular response (MMR), and 16 patients (21%) have lost MMR only. All these patients were given imatinib again, the same dose, on the standard schedule and achieved again CCyR and MMR or an even deeper molecular response. The probability of remaining on INTERIM at 6 years was 48% (95% confidence interval 35–59%). Nine patients died in remission. No progressions were recorded. Side effects of continuous treatment were reduced by 50%. In optimal and stable responders, a policy of intermittent imatinib treatment is feasible, is successful in about 50% of patients and is safe, as all the patients who relapsed could be brought back to optimal response.

The purpose of this study was to develop a flow cytometric approach to the evaluation of marrow dysplasia in myelodysplastic syndromes (MDS). We first studied a cohort of 103 MDS patients as well as 46 pathological and healthy controls. Flow cytometry data were expressed as percentage of positive cells. Analysis of erythroid cells showed higher proportions of immature cells (P < 0.001) and decreased levels of CD71 expression on nucleated red cells (P = 0.02) in MDS. Analysis of myeloid cells showed lower proportions of CD10+ and higher proportions of CD56+ granulocytes (P < 0.001), and increased ratios of immature to mature cells (P = 0.007). Since no single immunophenotype could accurately differentiate MDS from other conditions, we used discriminant analysis for generating erythroid and myeloid classification functions using combinations of immunophenotypic parameters. These functions were prospectively validated in a testing cohort of 69 MDS patients and 46 pathological controls. A diagnosis of MDS was obtained in 60/69 cases (87%). No false-positive results were noticed among controls. Significant correlations between values of these functions and both degree of morphological dysplasia and the International Prognostic Scoring System were found. These findings indicate that flow cytometry evaluation of marrow dysplasia is feasible and may be useful in the work-up of individual MDS patients.

BACKGROUND Myeloma plasma cells interact with the bone marrow microenvironment which, in turn, supports their growth and protects them from apoptosis. In vitro studies have demonstrated the antitumor potential of zoledronic acid (ZOL) on myeloma cell lines, but few data are available on its effects on bone marrow stromal cells (BMSCs). The aim of the current study was to evaluate the antiproliferative and apoptotic effect of ZOL on BMSCs, as well as its effect on the expression of adhesion molecules. METHODS BMSCs, obtained from bone marrow mononucleated cells of 8 patients with multiple myeloma, were treated with increasing concentrations of ZOL for 3 days. Cytotoxic effect was analyzed by 3-(4-5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide; thiazolyl blue (MTT) assay whereas the induction of apoptosis was evaluated by flow cytometric detection of fluorescein isothiocyanate-labeled annexin V, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay, and nuclear changes. Moreover, expression of CD106, CD56, CD50, CD49d, CD44, and CD40 was analyzed by flow cytometry. Data were evaluated by the Friedman test. RESULTS After 3 days of exposure at concentrations of 10−4 to 10−5 M, ZOL induced a decrease in proliferation (P < 0.0001) and an increase in apoptosis (P < 0.002). Analysis of culture supernatants showed that myeloma BMSCs expressed interleukin (IL)-6, negligible levels of tumor necrosis factor-alpha, and no IL-1β. In vitro exposure to the lowest concentrations of ZOL decreased IL-6 production by BMSCs. Among the adhesion molecules, CD106, CD54, CD49d, and CD40, which were strongly expressed at baseline, showed a statistically significant reduction compared with controls after exposure to ZOL. CONCLUSIONS ZOL interfered with myeloma BMSCs by reducing proliferation, increasing apoptosis, and modifying the pattern of expression of adhesion molecules, especially those involved in plasma cell binding. These effects on BMSCs might explain the antitumor activity of ZOL. Cancer 2005;. © 2005 American Cancer Society.

BACKGROUND Acute leukemia (AL) may occur as rare and late event of polycythemia vera (PV). METHODS The current study included 23 patients who developed acute leukemia in a cohort of 414 consecutive PV patients with long-term observation (3208 person years of follow-up). Kaplan–Meier Product-Limit method was used to estimate the cumulative probability of survival; Gehan–Wilcoxon test was applied to compare survival in different groups of patients. RESULTS Median age was 68 years, and 18 patients (78%) were > 60 years of age. At diagnosis of AL, most patients had a white blood count > 10 × 109/L (n = 17; 74%), Hgb 50 × 109/L (n = 17; 74%). Of 14 patients in whom cytogenetic analysis was available at leukemic transformation, 12 showed high-risk abnormalities including complex karyotype (n = 10), del (7)(q22) sole (n = 1) and del (X)(q26) sole (n = 1), whereas 2 had a normal karyotype. In patients whose karyotype was available at diagnosis of PV, cytogenetic evolution was documented at progression to AL. Treatment consisted of supportive care and/or low-dose chemotherapy (n = 15), or induction chemotherapy (n = 8). This included idarubicin plus cytarabine (n = 3), high-dose cytarabine (n = 4), and fludarabine-based regimen (n = 1). Allogenic stem cell transplantation was offered to a single patient, who is alive at Day + 70. The outcome of patients was poor, with a median survival of 2.9 months (range, 0.6–20.1 mos), with no significant differences between palliation and intensive treatments. CONCLUSIONS AL following PV has distinct clinical and biologic features. Outcome of patients is poor irrespective of the treatment employed. Cancer 2005. © 2005 American Cancer Society.

The clinical outcome, response to treatment, and occurrence of acute complications were retrospectively investigated in 308 primary autoimmune hemolytic anemia (AIHA) cases and correlated with serological characteristics and severity of anemia at onset. Patients had been followed up for a median of 33 months (range 12-372); 60% were warm AIHA, 27% cold hemagglutinin disease, 8% mixed, and 5% atypical (mostly direct antiglobulin test negative). The latter 2 categories more frequently showed a severe onset (hemoglobin [Hb] levels ≤6 g/dL) along with reticulocytopenia. The majority of warm AIHA patients received first-line steroid therapy only, whereas patients with mixed and atypical forms were more frequently treated with 2 or more therapy lines, including splenectomy, immunosuppressants, and rituximab. The cumulative incidence of relapse was increased in more severe cases (hazard ratio 3.08; 95% confidence interval, 1.44-6.57 for Hb ≤6 g/dL; P < .001). Thrombotic events were associated with Hb levels ≤6 g/dL at onset, intravascular hemolysis, and previous splenectomy. Predictors of a fatal outcome were severe infections, particularly in splenectomized cases, acute renal failure, Evans syndrome, and multitreatment (4 or more lines). The identification of severe and potentially fatal AIHA in a largely heterogeneous disease requires particular experienced attention by clinicians.

Myelodysplastic syndromes and chronic myelomonocytic leukemia (CMML) are characterized by mutations in genes encoding epigenetic modifiers and aberrant DNA methylation. DNA methyltransferase inhibitors (DMTis) are used to treat these disorders, but response is highly variable, with few means to predict which patients will benefit. Here, we examined baseline differences in mutations, DNA methylation, and gene expression in 40 CMML patients who were responsive or resistant to decitabine (DAC) in order to develop a molecular means of predicting response at diagnosis. While somatic mutations did not differentiate responders from nonresponders, we identified 167 differentially methylated regions (DMRs) of DNA at baseline that distinguished responders from nonresponders using next-generation sequencing. These DMRs were primarily localized to nonpromoter regions and overlapped with distal regulatory enhancers. Using the methylation profiles, we developed an epigenetic classifier that accurately predicted DAC response at the time of diagnosis. Transcriptional analysis revealed differences in gene expression at diagnosis between responders and nonresponders. In responders, the upregulated genes included those that are associated with the cell cycle, potentially contributing to effective DAC incorporation. Treatment with CXCL4 and CXCL7, which were overexpressed in nonresponders, blocked DAC effects in isolated normal CD34+ and primary CMML cells, suggesting that their upregulation contributes to primary DAC resistance.

BACKGROUND: The INTERIM study (ClinicalTrials.gov NCT 00858806) showed that in elderly (> 65 years) Ph+ CML patients selected for a stable complete cytogenetic response (CCgR) lasting > 2 years, the policy of intermittent imatinib treatment (one month on/one month off) may affect the markers of residual disease (CCgR and major molecular response, MMR or MR3.0), but not the clinical outcomes (overall survival and progression-free survival) (Russo D et al, Blood 2013; 121(26):5138-44). AIMS: To update the results of the INTERIM Study, with a follow up ≥ 5 years. METHODS: After 4 years of follow up, patients continouing INTERIM treatment were monitored with peripheral blood RT-Q-PCR every 3 months according to the ELN-2013 guidelines. RESULTS: At 48thmonth, out of 76 patients enrolled in the INTERIM study, 13 (17%) had lost CCgR and MMR, 14 (18%) had lost MMR only and 50 patients (75%) continued INTERIM. The patients who had lost CCgR and/or MMR resumed imatinib continuously and all of them regained the CCgR and the MMR, within 3 to 12 months. No patient progressed to accelerated or blastic phase, or developed clonal chromosomal abnormalities in Ph+ cells, or BCR-ABL mutations. No patient complained of new or more severe side effects during the months “on”. After a follow up ≥ 5 years, 45/76 (59%) enrolled patients are on INTERIM, with a probability of maintaining intermittent administration of 59% (95% CI: 46-69). No patient lost the CCgR and only 9 additional patients lost the MMR while on intermittent treatment. All these patients resumed continuous imatinib treatment and regained the MMR. Thus, at ≥ 5 years, the probability of maintaing CCgR is 80% (95% CI 68-87) and the probability of maintaining the MMR is 61% (95% CI: 48-71). From start of INTERIM, 6 patients died but no deaths were related to CML progression (3 cases of other non haematological neoplasms, 1 stroke, 1 myocardial infarction, 1 chronic obstructive pulmonary disease).The PFS at ≥ 5 years is 94% (95% CI: 89-100) CONCLUSIONS: In summary, with a follow up ≥ 5 years, intermittent imatinib administration (INTERIM) confirmed to be safe, to produce a reversible increase of residual molecular disease in about one third of patients, but not to affect the long-term outcome. Aknowledgments: This work was supported in part by EuropeanLeukemiaNet (contract LSHC-CT-2004-503216) through the European Treatment and Outcome Study (EUTOS), supported by Novartis Oncology Europe, and COFIN 2009 Disclosures Russo: Celgene: Research Funding; Gilead: Research Funding; Novartis: Consultancy. Martinelli:Novartis: Speakers Bureau; Bristol-Meyers and Squibb: Speakers Bureau; Pfizer: Speakers Bureau. Soverini:Novartis: Consultancy, Honoraria; Bristol-Meyers Squibb: Consultancy, Honoraria; Ariad: Consultancy, Speakers Bureau. Turri:Novartis: Consultancy, Honoraria; Bristol-Meyers Squibb: Consultancy, Honoraria. Castagnetti:Novartis: Consultancy, Honoraria; Bristol-Meyers Squibb: Consultancy, Honoraria. Breccia:novartis: Consultancy; BMS: Consultancy; Celgene: Consultancy. Abruzzese:Novartis: Consultancy. Tiribelli:Novartis: Consultancy, Honoraria; Bristol-Meyers and Squibb: Consultancy, Honoraria. Rosti:Consultant: Consultancy, Speakers Bureau; Bristol-Meiers Squibb: Consultancy, Speakers Bureau; Ariad: Consultancy, Speakers Bureau.

Background. The therapeutic approach in thrombocythemic patients with Philadelphia negative chronic myeloproliferative neoplasms (Ph- MPN), in order to result cost-effective (primum non nocere), is commonly driven by the risk factors considered in the gradually updated guidelines. However, no studies were addressed to evaluate if and how the real-life therapeutic approach changed in the last decades. Objective. To evaluate, in a large series of thrombocythemic patients with Ph- MPN, the impact of clinical and biological characteristics at diagnosis on the therapeutic approach adopted before and after the publication of the Italian guidelines for essential thrombocythemia therapy [1]. Methods. The analysis considered, in the patients of the Registro Italiano Trombocitemie (RIT), the clinical and biological characteristics at diagnosis, and the treatment ongoing after 3, 6, 12, and >12 months from diagnosis (antiplatelet alone [AntiPLT]; cytoreductive alone [CYT]; CYT+AntiPLT). Results. The analyzed patients were 2418, 914 (38%) males and 1504 (62%) females, with a diagnosis (PVSG or WHO criteria) performed before and after 2005 in 51% and 49% of cases, respectively. The rate of ongoing treatment with AntiPLT, CYT, and CYT+AntiPLT increased as follows: at 3rd month 16%, 12%, and 31%; at 12th month 17%, 14%, and 39%; after 12 months 19%, 16%, and 55%, respectively. Patients treated with CYT or CYT+AntiPLT did not significantly differ in their characteristics at diagnosis. The analysis of data at the 3rd month (initial phase) showed that: 1) CYT±AntiPLT treatment, ongoing in 43% of patients, was significantly related, in univariate analysis, to male gender, older age, prior thrombosis, higher thrombocytosis, leukocytosis, higher HCT level, CVRFs, comorbidities, symptoms, splenomegaly, hepatomegaly, and bone marrow fibrosis grade >0 (no relationship with JAK2 V617F mutation, and prior hemorrhage); in multivariate analysis, it was significantly related to age >60 y, age 40-60 y, prior thrombosis, PLT >1000 x109/L, PLT 700-1000 x109/L, symptoms, and comorbidities. 2) patients with standard high risk (age >60 y, and/or prior thrombosis, and/or PLT >1500 x109/L ) were receiving CYT±AntiPLT (59%), AntiPLT (7%), and no treatment (34%). 3) patients with standard low risk were receiving CYT±AntiPLT (22%), AntiPLT (27%), and no treatment (51%). Low risk patients receiving CYT±AntiPLT had age 40-60 y (73%), CVRFs (59%), symptoms (53%), comorbidities (42%), PLT 1000-1500 x109/L (35%), PLT 700-1000 x109/L (42%), JAK2 V617F mutation (30%), WBC >10 x109/L (22%). 4) in patients receiving CYT±AntiPLT, the initial cytoreductive drug and the median age were: hydroxycarbamide (80%, 68 y), anagrelide (6%, 49 y), interferon alpha (9%, 42 y), pipobroman (2%, 72 y), busulfan (3%, 70 y). The AntiPLT drug mostly used was low dose ASA (86-90% of cases, at any age). 5) Patients diagnosed after 2005, compared with those diagnosed before, showed a higher rate of CYT±AntiPLT treatment when at standard high risk ( 64% vs 53 % p Conclusion. The initial (within the 3rd month) therapeutic approach in the thrombocythemic Ph- MPN patients of the RIT was after 2005 relatively compliant with the 2004 Italian guidelines. In fact, the rate of CYT±AntiPLT treatment in patients with standard high risk was higher than before (64% vs 53%, p [1] Barbui T et al. Haematologica 2004; [2] Harrison C et al. NEJM 2005; [3] Gisslinger H et al. NEJM 2013; [4] Passamonti F et al. Blood 2012; [5] Barbui T et al. Blood 2012 *The RIT is a project of the GIMEMA Foundation Disclosures Gugliotta: Shire : Honoraria.

In patients with myelodysplastic syndromes (MDS), the likelihood of having a sustained response to azacitidine is increased by maximizing treatment duration. This is important as prognosis postrelapse is poor. There is also the concern that early termination of treatment may result in rapid disease progression. We reviewed outcomes in 13 patients who discontinued azacitidine (decitabine in one patient) while still responding to the treatment. Most patients rapidly relapsed; median time to progression was 5.4 months. Reasons for treatment discontinuation included comorbidities, infections, and patient choice. These findings illustrate the risk of prematurely terminating azacitidine therapy in MDS.

Telomere shortening, a well-known marker of aging and cellular stress, occurs under several conditions in the hematopoietic compartment, including aplastic anemia and following iatrogenic noxae. We decided to verify whether pathological telomere erosion also arises in restored Philadelphia-negative (Ph-negative) hematopoiesis following successful treatment of chronic myeloid leukemia (CML). Eighty-one CML patients in complete cytogenetic remission were compared to 76 age-matched healthy subjects. Myeloid cells of CML patients had shorter telomeres than controls (6521 bp vs 7233 bp, p

Background Therapy-related myeloid neoplasms (t-MN), including myelodysplastic syndromes and acute myeloid leukemia (t-MDS and t-AML) are associated to clinical and biologic unfavorable prognostic features, including high levels of DNA methylation. Methods We retrospectively evaluated 50 t-MN patients (34 MDS and 16 AML) selected among all patients receiving azacitidine (AZA) at 10 Italian Hematology Centers. Patients had developed a t-MN at a median of 6.5 years (range 1.7- 29) after treatment of the primary tumor (hematological neoplasm, 27 patients; solid tumor, 23 patients). Results The overall response rate was 42% (complete remission: 10 patients, partial remission: 2 and hematological improvement: 8 patients) and was obtained after a median of 3 cycles (range 1–6). Median overall survival (OS) was 21 months (range 1–53.6+) from AZA start. OS was significantly better in patients with less than 20% blasts, in normal karyotype t-AML and when AZA was used as front-line treatment. This was confirmed by the multivariate analysis. Conclusions This study reports efficacy of AZA in the largest series of therapy-related MN patients treated with 5-AZA. Our data show that blasts and karyotype maintain their important prognostic role in t-MN also in the azacitidine era.

Abstract 1680 Background. Chronic myeloid leukemia (CML) is characterized by the presence of the BCR-ABL1 hybrid gene. Different types of BCR-ABL1 fusion transcripts can be found, but the most frequent are the e13a2 (b2a2) and the e14a2 (b3a2). In the tyrosine kinase inhibitors (TKIs) era, few data about the prognostic significance of the transcript type in early chronic phase (ECP) CML are available. Three larger studies suggested that the e13a2 transcript may have an adverse prognostic impact in ECP CML patients treated with imatinib (IM): Vega-Ruiz et al. (251 patients, ASH 2007) reported inferior molecular responses; Lucas et al. (71 patients, Haematologica 2009) reported lower cytogenetic response rates and lower event-free survival (EFS); the GIMEMA CML WP (493 patients, EHA 2011) reported a slower time to major molecular response (MMR) with inferior EFS and progression-free survival (PFS). To our knowledge this is the first evaluation of the prognostic influence of the BCR-ABL1 transcript type on the responses and the outcome of ECP CML treated frontline with nilotinib (NIL). Methods. The CML Italian Registry of Nilotinib includes 215 patients treated with NIL-based regimens. The patients were enrolled within 2 multicenter phase II studies conducted by the GIMEMA CML WP (ClinicalTrials.gov. NCT00481052 and NCT00769327) or treated at the “S. Orsola-Malpighi” University Hospital (Bologna, Italy), with NIL 300 mg BID or 400 mg BID as initial treatment. All the registered patients were analyzed. Patients expressing rare transcripts and patients with both b2a2 and b3a2 transcripts were excluded: 201 out of 215 patients were evaluable, 81 (40%) with e13a2 transcript and 120 (60%) with e14a2 transcript. Differences between groups were tested using χ2 test, Fisher exact test or t-test, as appropriate. Response monitoring: conventional cytogenetic examination (bone marrow) and QPCR (peripheral blood). Definitions: MMR: BCR-ABLIS ratio Results. The baseline characteristics of the 2 groups were comparable (no significant differences in age, Sokal/Hasford/EUTOS score distribution, clonal chromosomal abnormalities in Ph+ cells, NIL dose), except for the percentage of basophils in the peripheral blood, higher in patients with e14a2 transcript (3.4% vs 2.3%, p=0.01). The median observation was 29 months (range 18–47); 92% of the patients had at least 2 year observation. The CCgR and MMR rates at 12 months were comparable in the 2 groups. The time to MMR was longer for patients with e13a2 transcript (6 months vs 3 months, p=0.04), but the overall CCgR rates (93.8 vs 91.7, p=0.79) and the overall MMR rates (85.1 vs 90.0, p=0.38) were not significantly different in patients with e13a2 or e14a2 transcript, respectively. The probability of Overall Survival (OS), Progression-Free Survival (PFS) and Failure-Free Survival (FFS) were comparable: 91.4% vs 95.8% (p=0.61), 90.7% vs 95.0% (p=0.51), and 90.7% vs 88.7% (p=0.40) in patients with e13a2 and e14a2 transcript, respectively. Conclusions. In our experience, based on 201 early CP CML patients treated frontline with NIL with a minimum follow-up of 18 months, the BCR-ABL transcript type did not show any relevant prognostic impact. The time to MMR was longer in patients with e13a2 transcript, but no response and outcome differences have been observed so far. The number of observed events was low and a longer observation is required. Acknowledgments. European LeukemiaNet, COFIN, Bologna University, BolognAIL Disclosures: Castagnetti: Novartis Pharma: Consultancy, Honoraria, Speakers Bureau; Bristol Myers Squibb: Consultancy, Honoraria, Speakers Bureau. Gugliotta:Novartis: Consultancy, Honoraria; Bristol-Myers-Squibb: Consultancy, Honoraria. Breccia:Bristol Myers Squibb: Consultancy; Novartis: Consultancy. Cavazzini:Novartis Pharma: Honoraria; Bristol Myers Squibb: Honoraria. Turri:Novartis: Consultancy, Novartis Other; Bristol Myers Squibb: Bristol Myers Squibb, Bristol Myers Squibb Other, Consultancy. Soverini:Novartis: Consultancy; Bristol-Myers Squibb: Consultancy; ARIAD: Consultancy. Saglio:Novartis: Consultancy, Speakers Bureau; BMS: Consultancy, Speakers Bureau; Pfizer: Consultancy. Martinelli:Novartis: Consultancy, Honoraria, Speakers Bureau; Bristol-Myers-Squibb: Consultancy, Honoraria, Speakers Bureau. Baccarani:ARIAD, Novartis, Bristol Myers-Squibb, and Pfizer: Consultancy, Honoraria, Speakers Bureau. Rosti:Novartis Pharma: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Roche: Speakers Bureau; Pfizer: Speakers Bureau.

Abstract 4 Introduction Current treatment recommendations in polycythemia vera (PV) have emphasized to maintain the hematocrit (HCT) values Aim In a large scale randomized clinical trial (Cyto-PV) we prospectively determined the efficacy and safety of maintaining the recommended HCT target versus HCT levels in the range of 0.45–0.50 to prevent thrombotic events in PV patients. Methods Patients were eligible if they met WHO-2008 diagnostic criteria for PV. Both cases with newly diagnosed disease and previous treatment were centrally randomized to Arm A (HCT Results Arm A and Arm B included 182 and 183 patients respectively. At randomization, there were no significant differences between the two groups with respect to age, gender, years from diagnosis to recruitment, previous history of major thrombosis, bleeding, concomitant cardiovascular risk factors, and hematological presentation. Treatments were equally distributed with regard to phlebotomy, antiplatelet drugs, warfarin and hydroxyurea or their combination. After randomization, median HCT levels in arm A and Arm B during follow-up (median 31.0 months) were 0.44 and 0.48 respectively. A quarter of patients of arm A and Arm B failed to maintain the assigned HCT levels during the study period. Noticeable was that leukocyte levels remained higher in arm B than Arm A while no difference was revealed concerning the platelet count. Additionally, no difference in the safety profile was recognizable. As compared with arm B, the more intensive treatment aimed at maintaining the HCT Conclusion In this randomized clinical trial, the incidence of major cardiovascular events was 4 fold higher in patients who maintained HCT levels >0.45. Therefore, an HCT level Disclosures: No relevant conflicts of interest to declare.

BACKGROUND: The efficacy of azacitidine for the treatment of high-risk myelodysplastic syndromes has prompted the issue of its potential role even in the treatment of acute myeloid leukemia (AML). METHODS: The authors analyzed 82 patients with AML who were diagnosed according to World Health Organization criteria. The median patient age was 72 years (range, 29-87 years), and 27 patients (33%) had secondary AML. Of 62 patients with evaluable cytogenetics, 18 patients (29%) had a poor-risk karyotype, and 44 patients (71%) had an intermediate karyotype. Thirty-five patients (43%) received azacitidine as front-line treatment, and 47 patients (57%) had previously received 1 or more line of chemotherapy. RESULTS: The overall response rate was 32% (26 of 82 patients) and included 12 (15%) complete remissions (CRs), 4 (5%) CRs with incomplete blood count recovery (CRi), and 10 (12%) partial responses (PRs). Responses were observed more frequently among untreated patients compared with pretreated patients; in fact, 17 of 35 untreated patients (48%) responded, including 11 responses (31%) classified as CR/CRi. Conversely, only 9 of 47 pretreated patients (19%) responded, including 5 responses (11%) that were classified as CR/Cri. The response rate was significantly higher for untreated patients (P = .006) and those who had white blood cell counts

Abstract 2078 Background: Health-related quality of life profile (HRQOL) of patients diagnosed with high-risk myelodysplastic syndromes (MDS) can be compromised already at the time of diagnosis before receiving any kind of treatment. Clinical decision-making is challenging due to the poor prognosis and no data exist on the possible relationships between patient's HRQOL and the request of prognostic information on survival during consultation. Aim: The main objectives were to assess preferences for prognostic information of patients with high-risk MDS and the relationship between such preferences and patient characteristics including HRQOL. To date no such evidence exists in this population. Patients and Methods: Data were gathered through an ongoing international prospective observational study that recruits newly diagnosed patients with MDS. These patients typically have a limited life expectancy. At the time of diagnosis, and during one of the first clinical consultations in which treatment options were discussed, patients completed the European Organization for Research and treatment of Cancer, Quality of life Questionnaire (EORTC QLQ-C30). The EORTC QLQ-C30 is a psychometrically robust generic HRQOL cancer measure assessing both symptoms and functional aspects. Physicians also completed an extensive survey about their patient's preference for involvement in treatment decisions and whether the patient explicitly requested prognostic information for survival. Associations between request for prognostic information, HRQOL socio-demographic characteristics (i.e., living arrangements, age, gender, education) and clinical data including: performance status, comorbidity and disease severity (i.e. IPSS risk category intermediate 2 vs. high risk) were investigated using Fisher's exact test and Wilcoxon-Mann-Whitney test as appropriate. Results: Overall, 184 patients (36% female and 64% male) were analyzed. Mean age of patients was 70 years (range: 31–88). 65% explicitly requested information about expected survival at the time of diagnosis. The symptom profile of patients requesting prognostic information was better than those who did not in 7 out of the 8 symptoms evaluated. The largest clinically meaningful difference was found for fatigue with a mean score of 39 (SD:26) and 52 (SD:28) respectively for those requesting prognostic information versus those who did not. Request for prognostic information was significantly associated with younger age (P=.01) and fewer comorbidities (P=.04). In addition, better physical functioning (P=.009), better role functioning (P=.002) and a lower level of fatigue (P=.002) were also associated with a request for prognostic information during consultation. Additional supportive analysis revealed that patients with a higher overall mean symptom score did not request information about survival (P=.02). Conclusion: These data suggest that the majority of patients with high-risk MDS request prognostic information on survival from their physicians at the time of diagnosis. There is also an indication that patients who are more likely to request such information are those who are in better health condition reporting higher functional abilities and lower symptoms. Disclosures: No relevant conflicts of interest to declare.

Abstract 1973 Background. Hematological and extra-hematological adverse events associated to Hydroxyurea (HU) treatment in patients with chronic myeloproliferative neoplasms (MPN) are object of particular interest in the Ph-negative MPN since they can significantly limit the HU use. Objective. To evaluate the extra-hematological adverse events associated to HU treayment in a large cohort of Essential Thrombocythemia (ET) patients. Material and Methods. One thousand and seventy-five out of the 2005 ET patients registered in the RIT were treated with HU and are the object of this report. The patients, 641 females (59.6%) and 434 males (40.4%), diagnosed according to the PVSG or WHO criteria in 54 hematological centers of the RIT, started treatment with HU as first (92%) or second (8%) line, at median age of 65 years.The mean duration of HU treatment was 3.3 years. The HU treatment was withdrawn in 221 (20.5%) patients after a mean of 3.0 years.The administered dose of HU was 0.25–3.0 g/day (median 1), and a mean cumulative dose was 1113 g. The extra-hematological adverse events (EHAEs) observed during the HU treatment were distinguished in HU related AEs (HU-EHAEs), ET related AEs (ET-EHAEs) and HU or ET unrelated AEs (U-EHAEs).Results. During the HU treatment (3587 pt-y) 378 EHAEs were reported in 207 (19.3%) patients, being the HU-EHAEs 244 (6.8/100 pt-y) in 170 (15.8%) patients. In detail, the HU-EHAEs were: dermatological in 108 (48.3%) cases (38 hyperpigmentation, 26 leg ulcers, 22 maculo-papular rash, 10 lichenoid eruptions, 5 skin cancer, 4 alopecia); gastro-intestinal in 80 (32.8%) cases (37 nausea/vomiting, 30 diarrhea, 13 gastro-intestinal intolerance); systemic in 35 (14.3%) cases (28 fever, 7 fatigue); neurological in 19 (7.8%) cases (headache); miscellanea in 2 (0.8%) cases. Conclusion. This preliminary analysis in 1075 ET patients of the Registro Italiano Trombocitemia (RIT) treated with HU shows that the extra-hematological adverse events referred to the HU (HU-EHAEs) occurred with not negligible rate (6.8/100 pt-y) and need to be object of attention in the management of ET patients. Disclosures: No relevant conflicts of interest to declare.

Abstract 860 Background: Elderly CML patients treated with Imatinib (IM) in early chronic phase (CP) have similar cytogenetic response and survival compared with younger patients, but they show a lower compliance to standard IM therapy (400 mg/day). Aims: The aim of the study is to investigate if CCgR that has been achieved with standard (daily administration) IM therapy can be maintained with the same dose of IM given intermittently (INTERIM). Methods: The study population is represented by elderly patients (≥ 65 years old) with Ph+ CML and with stable CCgR after at least 2 years of standard IM therapy (daily administration). IM is given at the same dose that was given at the time of enrollment by the following intermittent schedule: 1 week on / 1 week off for the 1st month; 2 weeks on / 2 weeks off for the 2nd and 3rd month; 1 month on / 1 month off from the 4th month thereafter. In cases of loss of CCgR INTERIM was stopped and standard therapy (daily administration) was resumed. After 12 months, the patients who are in continuous CCgR are advised to continue the intermittent study schedule and to be followed indefinitely. The CgR status was evaluated at baseline (by conventional cytogenetics on bone marrow and FISH on peripheral-blood) and every 3 months during the study (only by FISH on peripheral-blood). If FISH (% of Ph+ cells) increased more than 1% in two consecutive examinations, evaluation of marrow cells metaphases was performed to confirm the loss of CCgR and to check for additional cytogenetic abnormalities. Quantitative molecular assessment of BCR-ABL transcript by RQ-PCR on peripheral blood was due at baseline and every 3 months during the study and mutational analysis of ABL was performed in case of loss of CCgR. Results: One-hundred and fourteen patients have been considered eligible, but 17 (15%) refused to enter into the protocol. Out of 97 enrolled patients, 87 started INTERIM, 5 patients (5%) went off the study for major protocol violation before the 3rd month and, at present, 82 patients are ongoing. Of these 82 patients, 52, 30 and 11 completed the 3rd, 6th and 9th month, respectively. The preliminary results of the first 6 months are here reported. The distribution of patients according to FISH results is shown in Fig. 1. Only 1/68 pts (at 6th month) showed an increased >1% in Ph+ cells by FISH but he maintained a CCgR when checked by conventional cytogenetic. As showed in Fig. 2, 96 to 87% of patients maintained a major molecular response MMR (≤0,1) according to International Scale (IS). Conclusions: This study is trying to test the minimum effective dose of Imatinib to maintain the CCgR in elderly CML patients with stable CCgR. The preliminary results at 6 months do not show negative trends both for cytogenetic and molecular response. Therefore, the study is ongoing and all patients are expected to complete the trial time (12 months). Disclosures: No relevant conflicts of interest to declare.

Abstract 943 Introduction: Arsenic trioxide (ATO) inhibits angiogenesis, induces apoptosis and differentiation in variety of malignant cells. Two multicenter phase II studies have documented that ATO has as a moderate activity both in low and high risk myelodysplasia with 19-21% haematological improvement. Ascorbic acid in “vitro” increases in tumor cells the activity of ATO by depleting GSH and inhibiting NF-kb. High EVI-1 levels have been reported to predicted the response to arsenic trioxide and thalidomide combination therapy. We report the results of a multicenter phase II study with combination of arsenic trioxide and acid ascorbic in myelodysplastic patients. Patient eligibility included:a) pts with 3q26 rearrangement or high EVI-1 levels; b) non-RAEB pts at low-intermediate/1 risk who failed a previous therapy; c) RAEB or high-intermediate-2 risk patients not elegible to chemotherapy and/or bone marrow transplant. Study design: Arsenic trioxide was administrated intravenously over 1 hour at the loading dose of 0.30 mg/kg/day for 5 consecutive days, followed by 0.25 mg/kg/day twice weekly for 15 weeks. Ascorbic acid 1000 mg was given IV within 30 minutes after each arsenic trioxide infusion. Response evaluation was scheduled after two and four months of treatment according to the IWG criteria (Cheson 2000) Patients: 44 pts were enrolled .Median age was 71 years (range 47–80). WHO categories were non-RAEB (15 pts), RAEB-1 (12 pts), RAEB-2 (17 pts). 23 patients had a low/intermediate-1 IPSS score; 18 had a high/intermediate-2 risk. Risk category, could not be assessed in 3 pts lacking of cytogenetic data. Before starting therapy EVI-1 was highly expressed in 13 out of 34 evaluated pts (38%); high WT-1 levels was documented in 25 out of 35 pts (71%). 31 pts (57%) were transfusion dependent at baseline. Results: Ten out of 44 evaluable patients obtained a response (23%). They included 1 complete remission, 2 major erythroid hematologic improvement (HI-E major), 3 minor HI-E, 2 major neutrophil improvemet (HI-N) and 2 major trilineage responses. Response rate was 35% in lower risk IPSS patients and 6% in higher risk pts (p 0.05).Only 3 pts with high EVI-1 expression achieved a response. In 8 out of ten responders, the response was evident within the first 8 weeks of treatment. Twenty three (52%) patients discontinued treatment because disease progression (11%), severe adverse events (32%), drug unrelated adverse events (5%), withdrawal of consent (5%). Severe neutropenia and thrombocytopenia were observed respectively in 45% and 23% pts. The other G3-4 adverse events observed were: ATRA-like syndrome (9%), cardiotoxicity (11%) infection (11%), hepatotoxicity (9%). No toxic death was observed. Summary and Conclusions: The combination of ATO and ascorbic acid is tolerable and it is active in about 25% of MDS patients. The addition of ascorbic acid to ATO does not increase neither the toxicity nor the response rate to ATO. The tolerability of this regimen is reduced in elderly and high risk patients. In our study high EVI-1 transcript levels were not confirmed to predict the response to ATO. Disclosures: Off Label Use: Arsenic trioxide is yet approved for the treatment of MDS, even if its efficacy has been shown in other phase II studies.

The aim of the study is to evaluate clinical features, treatments and outcome of patients with systemic mast cell disease (MCD) who arrived to the attention of hematologists. A retrospective study was conducted over 1995-2006 in patients admitted in 18 Italian hematological divisions. Twenty-four cases of advanced MCD were collected: 12 aggressive SM (50%), 8 mast cell leukemia (33%), 4 SM with associated clonal non-mast cell-lineage hematologic disease (17%). Spleen and liver were the principal extramedullary organ involved. The c-kit point mutation D816V was found in 13/18 patients in which molecular biology studies were performed (72%). Treatments were very heterogeneous: on the whole Imatinib was administered in 17 patients, alpha-Interferon in 8, 2-CdA in 3; 2 patients underwent allogeneic hematopoietic stem cell transplantation. The overall response rate to Imatinib, the most frequently employed drugs, was of 29%, registering one complete remission and four partial remission; all responsive patients did not present D816V c-kit mutation. Overall three patients (12%) died for progression of disease. We conclude that MCD is characterized by severe mediator-related symptoms but with a moderate mortality rate. D816V c-kit mutation is frequent and associated with resistance against Imatinib. Because of the rarity of these forms, an effective standard of care is lacking. More data are needed to find new and successful therapeutic strategies.

Summary Ph-negative chronic myeloproliferative disorders (CMPD) are characterized by constitutive Janus kinase-signal transducer and activator of transcription (JAK-STAT) activation. SOCS3, SOCS1 and PTPN6 (SHP1) are negative regulators of the JAK-STAT pathway. We investigated epigenetic and genetic inactivation of SOCS3, SOCS1 and PTPN6 in 112 CMPD and 20 acute myeloid leukaemia (AML) post-CMPD. SOCS3 methylation occurred at high frequency in both CMPD (46/112; 41·1%) and AML post-CMPD (10/17; 58·8%) and was associated with transcriptional silencing. In contrast, methylation of SOCS1 and PTPN6 was observed in only a fraction of CMPD (15/112, 13·4% for SOCS1; and 8/112, 7·1% for PTPN6) and AML post-CMPD (3/20, 15% for SOCS1; and 1/20, 5% for PTPN6). No somatic mutations of SOCS1 were found in CMPD. SOCS3, SOCS1 and PTPN6 methylation occurred in both JAK2V617F-positive (35·1% for SOCS3; 14·9% for SOCS1; 8·1% for PTPN6) and JAK2V617F-negative (57·1% for SOCS3; 14·3% for SOCS1; and 9·5% for PTPN6) CMPD. These data indicate that methylation of SOCS3 and, to a lesser extent, SOCS1 and PTPN6 is a frequent event in both JAK2V617F-positive and -negative CMPD and may act as an alternative or complementary mechanism to JAK2 mutations, enhancing cytokine signal transduction. The frequent inactivation of SOCS3 is a novel finding in CMPD with potential implications for the molecular pathology of these disorders.

5-azacytidine (AZA) significantly prolonged overall survival in higher-risk patients with myelodysplastic syndromes (MDS) in a large, international, randomized, phase III trial (AZA-001). However, data about efficacy and safety of AZA in lower risk MDS are less consistent and only few small studies have addressed this topic. Among a total of 246 MDS treated with AZA in 31 different Italian Institutions since 2005 within to a national patient named program, we evaluated 82 patients scored as low/int-1 IPSS risk MDS. Median age was 68 years (range 34–85), male/female ratio 50/32. According to WHO classification, there were 21 RA/RARS, 4 5q-syndromes, 20 RCMD, 24 RAEB-1, 5 RAEB-2, 4 CMMoL, and 4 MDS unclassified. Median time from diagnosis was 27 months (range 1–132). Sixty-eight patients (82.9%) were transfusion-dependent, sixty (74%) had received a prior treatment, mostly with erythropoiesis stimulating agents. AZA was administered as single drug in 61 patients (74.4%), while in the remaining subjects it was variously combined with growth factors, valproic acid or other agents. Forty-eight patients (58.5%) received a “standard” AZA dose of 75 mg/sqm/d s.c., thirty-four (41.5%) a fixed dose of 100 mg/d s.c. Single cycle treatment duration was 7 days in 45 patients (54.9%), < 7 days in 32 patients (39%), > 7 days in 3 patients (3.7%), unknown in 2 patients (2.4%). The median number of monthly cycles was 6 (range 1–21), and 63 patients (76.8%) completed at least 4 cycles. The most relevant toxicities observed (grade 3–4) were represented by myelosuppression (22%) and infections (6%). According to 2006-updated IWG criteria, overall response rate was 39% (47.5% in patients who had completed at least 4 cycles). In particular, complete response, partial response and hematological improvement occurred in 12.2%, 8.5% and 18.3% of patients (15.8%, 11.1% and 20.6% in those who were treated with at least 4 cycles), respectively. Stable or progressive disease was observed in 29.3%/25.6% and 30.2%/22.2% of patients receiving less than or at least 4 cycles, respectively. Response duration ranged from 1 to +21 months. There were no significant differences in response rate according to dose and schedule employed, although a slight trend in favour of 75 mg/sqm vs 100 mg fixed dose was seen (45.8% vs 29.4%, respectively). There was also no difference in the percentages of response according to age, previous treatment and transfusion dependence. Overall survival at 2 years was 62%. A survival benefit emerged for responding patients, compared to non responders (82% vs 57%) (p=0.015). A favourable trend was also observed for transfusion-independent patients, while age, pre-treatment and AZA dose did not influence survival. These data indicate that AZA may be safe and effective for a subset of patients with low/int-1 IPSS risk MDS, resistant or not suitable for alternative treatments. The efficacy may improve if at least 4 cycles are administered.

Lymphocytosis associated with viral infection is generally polyclonal or oligoclonal. In this article, we describe a case of transient monoclonal CD8+/CD57+ T-cell lymphocytosis with large granular lymphocyte (LGL) morphology occurring after primary CMV infection and review cases of virus-associated monoclonal CD8+ T-cell expansions reported in the literature. Several clinical features shared by virus-associated monoclonal CD8+ T-cell expansions suggest the reactive nature of the lymphocytosis. Based on this, our case report and those reported in the literature support the notion that T-cell receptor clonality per se is not necessarily indicative of malignancy. These observations further corroborate the need for a close follow-up before assigning the diagnosis of LGL leukemia to individuals developing monoclonal CD8+ T-cell expansions.

Hairy cell leukaemia (HCL) occasionally displays a monoclonal gammopathy, yet the association of HCL with paraproteinemic demyelinating neuropathy (PDN) has not been reported. We describe a HCL case complicated by PDN and high titers of monoclonal IgM against myelin associated glycoprotein (MAG). Heavy and light chains of the patient's anti-MAG monoclonal protein were consistent with those expressed by HCL cells. After treatment with cladribrine, remission of HCL strictly paralleled disappearance of the IgM monoclonal protein and of the serum anti-MAG activity, and led to PDN clinical and electrophysiological improvement. Purine analogs may represent a choice in IgM PDN associated with lymphoproliferative disorders.

Distinct forms of tyrosine kinase domain (TKD), juxtamembrane domain, exon 8, and internal tandem duplication (ITD) mutations of c-KIT, were observed in about 46% of core binding factor leukemia (CBFL) patients. To evaluate their prognostic significance, 67 adult patients with CBFL were analyzed to ascertain the c-KIT mutation status. In acute myeloid leukemia (AML) with t(8;21), the presence of c-KIT TKD mutation at codon 816 (TKD(816)) was associated with a high white blood cell count at diagnosis (median, 29.60 x 10(9)/L) and a higher incidence (33%) of extramedullary leukemia (EML) during the course of the disease. Data also showed that the TKD(816) mutated patients (n = 12) had a significantly higher incidence of relapse and a lower overall survival (OS) at 24 months, compared with the 17 c-KIT unmutated (c-KIT(-)) patients (90% vs 35.3%, P = .002; 25% vs 76.5%, P = .006, respectively). No difference in relapse incidence (P = .126) and OS (P = .474) was observed between the c-KIT mutated other than TKD(816) (n = 7) and the c-KIT(-) patients. These findings indicate that c-KIT TKD(816) mutation has a negative impact on the outcome of AML with t(8;21).

Myelodysplastic syndromes (MDS) and the related disorder chronic myelomonocytic leukemia (CMML) are characterized by abnormal DNA hypermethylation and the DNA methyltransferase inhibitors (DMTis) azacytidine and decitabine (DAC) are frequently used as frontline therapy in these patients. However, DMTis are ineffective for ~50% of the patients who must still undergo treatment for at least 6 months before they can be deemed resistant. Therefore, it is of critical importance to identify baseline molecular differences associated with DMTi sensitivity that can help (i) to improve patient risk-stratification at diagnosis and (ii) identify the underlying mechanisms of resistance to these agents. Previous efforts to identify baseline DNA methylation differences at promoter regions between DMTi responders and non-responders have not been successful, so we hypothesized that any potential differences would be located distally from promoter regions. For this purpose we studied 40 CMML patients at diagnosis, all of whom had been uniformly treated with DAC 20 mg/m2/day x 5 days as frontline therapy. After 6 cycles of therapy patients were classified as responders (n=19, hematological improvement or better), or non-responders (n=21, stable or progressive disease). Mutational analysis showed no significant differences in the frequency of mutations in TET2, ASXL1, DNMT3A, RUNX1, TP53, JAK2, KIT, KRAS, EZH2, IDH1/2 and spliceosome genes. Using Enhanced Reduced Representation Bisulfite Sequencing (ERRBS) we analyzed the baseline methylation status at ~3M CpG sites across the genome of 39/40 CMML patients. We identified 158 statistically significant differentially methylated regions (DMRs) (FDR Disclosures No relevant conflicts of interest to declare.

Background In adults, ITP displays variable clinical presentation and different response to immune suppressive agents. The pathophysiological differences underlying these different behaviours are mostly unknown and a better knowledge of this biological heterogeneity might help identifying more targeted and rationale treatments for this disease. Purpose To identify immunogenetic features distinguishing ITP patients from controls and different subsets of ITP patients based on clinical presentation and treatment responsiveness. Several biological analyses were based on the model of autoimmune lymphoprolipherative syndrome (ALPS), a pediatric disease due to genetic defects decreasing function of the Fas death receptor that displays polyclonal lymphoproliferation, expansion of TCRαβ+ T cells double negative for CD4 and CD8 (DN T cells) and autoimmune manifestations frequently including thrombocytopenia. ALPS is mostly due to deleterious mutations of the Fas gene, but the +1239A>C single nucleotide polymorphism (SNP) of the osteopontin gene (OPN) and several variations of the perforin gene (PRF1) can act as disease modifier. Finally, in the peripheral blood (PB) of ALPS, regulatory T cells (Treg), marginal zone B cells (MZB) and switched memory B cells (SNB) are decreased, whereas type 17 T helper cells (Th17) and production of IL-10 and IL-17 are increased. Patients and Methods Adult patients with ITP and matched controls were selected for the analyses and stratified in different clinical subsets according to the disease severity and responsiveness to therapy (asymptomatic, steroid or rituximab sensitive vs. steroid or rituximab refractory). Analyses included evaluation of Fas-mediated apoptosis, proportions of DN T cells in PB, mutation of OPN and PRF1, the percentage (%) of natural killer T cells (NKT), myeloid (mDC) and plasmacytoid dendritic cells (pDC), MZB, SMB, Treg, and Th17, intracellular production of IL-10 and IL12 in cultured mDC, intracellular and secreted IL-10 and IL-17 in cultured activated PBMC; finally, patients were evaluated for TCR monoclonality and, rituximab-untreated patients, also for BCR monoclonality. Results Analysis of Fas function and DN T cell expansion was assessed in 149 ITP patients and showed that they displayed higher frequency of defective Fas function than the controls (26/149, 17% vs. 5/100, 5%; PC SNP displayed a similar distribution in the patients and the controls. No statistical differences of all these immunogenetic parameters were found comparing patients refractory vs. sensitive to either rituximab or steroid treatments. TCR monoclonality was assessed in 74 patients and was detected in 3 of them (4%). BCR monoclonality was assessed in 14 patients in PB and bone marrow resulting always absent. Compared with controls, ITP patients showed a lower % of mDC, pDC and Treg, and increase % of MZB; on the contrary we couldn’t find difference in the % of NKT and SMB. Moreover, mDC showed higher median levels of IL-10 (2.98 vs 1.35 positive cells %, P = 0.030) and lower levels of IL-12 (1.54 vs 1.89 %, P = 0.0454), whereas activated PBMC showed lower levels of both IL-10 (0.215 vs 1.16 %, P < 0.0383) and IL-17 (0.545 vs 1.49 %, P < 0.0001). Conclusions These analyses detected some differences between ITP patients and controls suggesting that: a) some defects involved in ALPS development, and, in particular, defective Fas function and some rare PRF1 mutations may play a role also in adult ITP patients; b) no significant pattern of monoclonal T cell and B cell expansion were documented; c) the immunological analyses showed an altered distribution and activity of cell subsets involved in immune regulation, with substantial differences with those reported in patients with ALPS. Disclosures Zaja: Roche: Honoraria, Research Funding. Off Label Use: Rituximab in ITP. Gaidano:Roche: Membership on an entity's Board of Directors or advisory committees.

Autoimmune hemolytic anemia (AIHA) is a greatly heterogeneous condition both in terms of clinical presentation and response to treatment, usually classified as warm (WAIHA), cold (CAD), mixed, and atypical forms. The aim of this study was to identify predictors of outcome and response to therapy considering in particular the serological characteristics and the severity of anemia at onset. We evaluatedretrospectively 307 patients (112 M and 195 F, median age at diagnosis 63, range 1-97), diagnosed between 1978 and 2013 and followed-up for a median of 33 months (range 12-372); 60% of cases were WAIHA, 27% CAD, 8% mixed, and 5% atypical (14 DAT- and 1 DAT+ for IgA only). Hemoglobin values were lower in mixed (median 5.8, range 2-10.7 g/dL) atypical (6.2, 3-9), and in IgG+C3 DAT+ WAIHA (6.9, 2.9-11.5). Twenty-one subjects were diagnosed with Evans’ syndrome, the majority of them WAIHA, with a severe onset. Considering anemia at onset, 27% of cases had Hb levels 10 g/dL; the most severe cases were mainly mixed and atypical forms (P=0.0001). Regarding therapy, 47% of cases were treated with one therapy line only, 26% with two, 13% with three, and 4% with four or more lines. Sixty % of WAIHA received first line steroid therapy only, 20 CAD required no treatment, and patients with IgG+C DAT+ WAIHA, mixed, and atypical forms were more frequently treated with 2 or more therapy lines (P As regards complications, infections occurred in 26 cases (10 grade 3, 11 grade 4, and 5 grade 5), irrespective of serological AIHA type and severity at onset, and of the number of therapy lines; on the contrary, they were observed more frequently in splenectomized cases. Acute renal failure was recorded in 6 cases and was not associated with AIHA clinical or serological characteristics. A thrombotic event was recorded in 11% and was associated with severe onset, higher median LDH levels, and previous splenectomy. At the time of the analysis 63 cases (21%) have died, of whom 11 because of AIHA (3.6%); death was not associated with the severity of anemia at onset, nor with the serological type of AIHA; at variance, it was associated with infections (HR 11.47, 95% CI 3.43-38.4, p=0.0004), acute renal failure (HR 17.99, 95% CI 4.73-68.40, p=0.001), Evans’ syndrome (HR 6.8, 95% CI 1.99-23.63, P=0.0074), previous splenectomy (HR 3.21, 95% CI 0.92-11.25), and multi-treatment (4 or more lines of therapy; HR 9.1, 95% CI 2.41-34.36, p=0.0076). Death was not associated with thrombotic events, nor with the type of treatment, in particular immunosuppressants or rituximab. In conclusion, we showed that AIHA cases with a severe onset, mostly mixed and atypical forms, are frequently refractory to different therapies. Although obtained retrospectively, our results suggest to put forward rituximab among second line options, given its efficacy and safety. In addition, standard rituximab doses should be preferred in CAD, whereas lower doses may be equally effective in WAIHA and mixed forms. Finally, we suggest to defer splenectomy after rituximab, given the increased risk of thromboembolism, infections and fatal outcome in splenectomized patients. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

The nucleoside analog (na) 5-aza-2’-deoxycytidine (dac) has good, but heterogeneous, efficacy in the therapy of chronic myelomonocytic leukemia (cmml). Given that no standard therapy has been identified so far for cmml, there is urgent need to identify molecular markers that could support the choice of dac therapy and identify patients more prone to respond. Recently, we demonstrated that in mds patients the expression of uck1, involved in the activation of azacitidine, may influence the clinical response to this treatment (valencia et al, leukemia 2013). In the present study, we assessed the role of dac metabolizing enzymes as well as genetic alterations common to cmml patients in response to dac in a uniformly and prospectively treated cohort of cmml patients. Methods: Patients Forty cmml patients were treated with dac 20mg/m2/day for 5 days every 28 d. DNA and rna were extracted from bm mononuclear cells of 19 pts defined as responders to dac (r), and 21 as non-responders (nr). Hematological response was evaluated according to iwg 2006 criteria. Gene mutation. the fifteen most frequently mutated genes in cmml were sequenced at a mean depth of coverage of 520x (range 169–714x). Functional studies. the role of two main genes involved in dac metabolism: dck (dac activation) and dctd (dac deactivation) was evaluated by silencing both genes with sirnas. The experiments were performed in the mds-skm1 cell line and in primary cells of 6 cmml cases exposed in vitro to dac 10um for 48h. Gene expression. the expression level of genes hent1, hent2, dctd, hcnt3, cn-ii, dck and cda, involved in dac metabolism was evaluated by qrt-pcr in 38/40 cmml patients and by rnaseq in 14/40 patients. CDNA libraries were sequenced using the hiseq 2000. The counts of endogenous genes were normalized by ercc spike-in library controls, and differential expression analysis was performed using edger (v3.4.2) glm model. The differentially expressed genes were identified at the fdr cutoff of 0.05 and absolute fold change greater than 2. Results: in skm-1 cells and in cmml primary mononuclear cells, dctd silencing increased dac- induced apoptosis (annexin v-positive cells 20.2%±0.8% vs control 13.8%±0.5%; p=0.01). dck silencing led to a decrease in dac-induced apoptosis (annexin v-positive cells 8.8%±0.1% vs control 13.8%±0.5%; p=0.05). We could not detect by rnaseq a significant difference in the expression levels of na metabolizing enzymes between responders and non responders to dac. qrt-pcr confirmed these observations. The mutational frequencies (figure) in this cohort of cmml cases were: srsf2 50%, tet2 44.7%, asxl1 39.4%, nras 18.4%, runx1 and dnmt3a 10.5%, u2af1 and tp53 7.8%, kras, jak2 and kit 5.2%, ezh2, idh1, idh2 and sf3b1 2.6%. No single genetic alteration was significantly associated with shorter overall survival or resistance to dac. Conclusions: although we could clearly demonstrate that in vitro expression of dac metabolizing enzymes influenced response to dac, clinical resistance does not appear to be directly correlated with the expression of genes involved in dac metabolism nor to specific gene mutations and is likely determined by other clinical/molecular variables that remain to be identified. Figure 1 Figure 1. Disclosures Finelli: Janssen: Speaker, Speaker Other; Novartis: Speaker, Speaker Other; Celgene: Research Funding, Speaker Other.

We analyzed by immunocytochemistry the expression of survivin in bone marrow cells from 36 acute myeloid leukemia (AML) cases, from 98 patients with myelodysplastic syndrome (MDS), and from 41 non hemopathic subjects. Our aim was to evaluate whether abnormalites in survivin expression were associated with peculiar laboratory and clinical findings, altered apoptosis levels or altered proliferative rate. In normal samples survivin was never detectable. It was detected in almost all AML and MDS cases. In AML and in MDS with more than 5% bone marrow blasts survivin levels higher than in RA and RARS were observed (P=0.04). In MDS a tendential inverse correlation between survivin and TUNEL positivity was identified (P=0.08), whereas survivin expression was independent of the proliferative rate. Survivin levels did not predict disease progression in MDS; among AML patients treated with intensive polichemotherapy, survivin expression was significantly higher in resistant cases (P=0.01). Our findings confirm the high incidence of survivin expression in AML. Its abnormal expression also in MDS may play a role in promoting aberrantly increased cell viability and contribute to the altered homeostatic balance between cell growth and cell death.

Background and Objectives: In adult acute lymphoblastic leukaemia (ALL), unlike in childhood ALL, the percentage of long-term remitters and survivors has not improved significantly over the last decades. In the present analysis, we describe a series of adult ALL patients consecutively treated with the same regimen in order to analyse prognostic factors and treatment outcome as well as to define new risk-oriented strategies. Design and Methods: From 1990 to 1998, 102 newly diagnosed ALL patients were referred to our division, 83 of them were eligible for the present study. Median age was 31 years (range 13–76); 77.1% had B-lineage ALL and 22.9% T-lineage ALL; 36.1% showed associated myeloid markers. All patients received an induction phase treatment, consisting of a 4-week cycle with vincristine, daunorubicin, L-asparaginase and desametasone; the consolidation phase included cyclophosphamide, cytarabine, 6-mercaptopurine and central nervous system (CNS) prophylaxis, followed by three of months maintenance (methotrexate + 6-mercaptopurine), re-induction (4-week cycle with vincristine, adriamicin, desametasone), and 2-year maintenance with methotrexate + 6-mercaptopurine. Results: Complete remission (CR) was achieved in 66 patients (79.5%); 20.5% of patients were resistant. The relapse rate was 60.2%. There were 10 CNS relapses (accounting for 12% of all patients, 15% of all CRs and 20% of all relapses). One patient had an ovarian and 2 had a breast relapse. Eleven patients remained in first continuous CR after chemotherapy. Median overall survival (OS) and disease-free survival (DFS) were 1.8 and 1.0 years, respectively, with a median follow-up of 5.6 years (range 0.3–12.1). Initial white blood cell count ≤30 × 109/l, age Interpretation and Conclusions: In our cohort, we were able to define a small subgroup of adult ALL patients with a better outcome. However, the majority of patients is at a high risk of failure when treated with standard protocols. There is a need for new, more active regimens for these patients.

Secondary (or therapy-related) acute myelogenous leukemia (t-AML) and myelodysplastic syndrome (t-MDS) have been reported after chemotherapy with alkylating agents and/ or topoisomerase inhibitors for the treatment of Hodgkin disease, non-Hodgkin lymphoma, breast cancer, ovarian carcinoma or testicular carcinoma [1,2]. Th ese disorders usually have a very poor prognosis with conventional chemotherapy, with an estimated overall survival (OS) of less than 1 year, due to the association with several negative features, such as older age, poor risk karyotype and resistance to chemotherapy [1 – 3]. Recently, we reported a retrospective analysis of 50 patients with therapy-related myeloid neoplasms (t-MN) treated with azacitidine with an overall response rate of 42% and a median OS of 21 months. We found that OS was signifi cantly better in patients with fewer than 20% of blasts, normal karyotype and in patients treated frontline with azacitidine [4]. Quintas-Cardama and colleagues [5] recently proposed a new score for patients with t-MDS, which was created in a large series of 279 patients with a history of previous exposure to chemo- or radiotherapy for prior malignancies. Th e model was able to identify diff erent OS based on the negative prognostic weight of several features at baseline (each one with 1 point), such as age higher than 65 years, Eastern Cooperative Oncology Group (ECOG) performance score higher than 1, monosomy 7 or complex karyotype, refractory anemia with ringed sideroblasts (RARS) or refractory anemia with excess blasts types 1 and 2 (RAEB1 – 2) according to the World Health Organization (WHO) classifi cation, hemoglobin level less than 11 g/dL, platelet count less than 50 10 9 /L and transfusion requirement. Patients were divided into three prognostic groups and subdivided into good risk (0 – 2 points), with a median OS of 34 months, intermediate (3 – 4 points), with a median OS of 12 months, and poor risk (5 – 7 points), with a median OS of 5 months. Th e model was tested in 189 patients with t-MDS with a median OS of 26, 13 and 7 months, respectively. Not all patients received azacitidine, and the subset of patients treated with this drug were not specifi cally reported. We retrospectively applied this score to our series of patients uniformly treated with azacitidine with the aim of validating the new proposed t-MDS score in this setting. We applied the score in 34 patients with t-MDS (12 refractory cytopenia with multilineage dysplasia [RCMD], nine RAEB-1 and 13 RAEB-2), with an overall response rate to azacitidine of 50%. Six patients had a low score (17%), 15 patients had an intermediate score (43%) and 14 patients had a poor score (40%). Among the good risk group, one patient achieved a complete remission (CR), one patient a partial response (PR), two patients a hematological improvement (HI) and two patients stable disease. On the other hand, among the intermediate risk group of patients, six achieved a CR, three a HI and fi ve stable disease, and within poor risk patients, eight had stable disease, two patients achieved a HI and one patient a CR, whereas three patients experienced disease progression after a median of four cycles. Median OS estimated with the Kaplan – Meier method according to the score stratifi cation for low, intermediate and poor risk patients was 17, 11 and 8.6 months, respectively ( p 0.02). Similarly, leukemia-free survival (LFS) was also signifi cantly diff erent in patients with low, intermediate and poor risk score (83%, 73% and 57%, respectively, p 0.01). Two out of six low risk patients (33%) died during treatment, compared to four of 14 (28%) intermediate risk and nine of 14 (64%) poor risk patients ( p 0.02).

Several trials have suggested that intensive post-remission therapy may prolong the duration of complete remission (CR) in acute myeloid leukemia (AML). The purpose of this study was to evaluate the feasibility and the efficacy of high-dose cytarabine (HiDAC) consolidation chemotherapy followed by high-dose therapy and autologous infusion of peripheral blood progenitor cells (PBPC) mobilized by G-CSF in adult patients with AML in first CR. Fifteen consecutive AML patients underwent HiDAC consolidation chemotherapy, used as a method of in vivo purging, followed by G-CSF for the purpose of autologous PBPC collection. Eleven patients collected a median of 6.9x10(8)/kg peripheral blood mononuclear cells (MNC) (range 2.9-23) and a median of 6.67x10(6)/kg CD34+ cells (range 1.8-33.5) with a median of two aphereses (range 1-3). Two patients did not mobilize and two obtained an inadequate number of progenitor cells. The 11 patients with adequate collections received myeloablative chemotherapy followed by the infusion of PBPC. The median number of days to recover neutrophils and platelets was 12 and 13, respectively. After a median follow-up of 28.7 months (range 17.2-43.4), five out of 11 patients who underwent PBPC transplantation are still in CR, five have died in first relapse and one is alive in CR after relapse treated with salvage therapy and second PBPC infusion. These results demonstrate that HiDAC consolidation chemotherapy followed by autologous PBPC transplantation is a feasible procedure with minimal toxicity. Randomized studies should be performed to evaluate whether this form of consolidation may produce a significant improvement in leukemia-free survival.

Introduction Erythropoiesis stimulating agents (ESAs) have been demonstrated to reduce the transfusion requirements in International Prognostic Scoring System (IPSS) low/Int-1 risk MDS patients and response to ESAs is associated with an improved quality of life. Response to ESAs varies from 20 to 60% according to the different studies and an association between ESAs response and a better overall survival (OS), without any disease evolution to AML has been reported. Aim of our study is to analyze the real life use of ESAs in Italy and to assess which group of lower risk MDS patients can benefit more by this treatment. Methods From 1999 to 30 Jun 2013, 2487 patients affected by MDS were included in the Network of the Italian MDS Registries. Morphological MDS diagnosis was confirmed. Among them, 1411 IPSS low/int-1 patients with follow-up data available were selected. Furthermore we excluded the following cases: 87 patients who were treated at any time by azacytidine (n=61) or lenalidomide (n=26), 95 patients with WHO 2002 diagnosis of CMML and aCML, 96 patients with incomplete data, 84 patients unhomogeneously treated with ESAs other than EPO alpha. We obtained a final data base of 1049 cases. OS and leukemic evolution (LE) according to EPO treatment were analyzed using the Kaplan-Meier product-limit survival curve estimates and log-rank tests. Results Median age was 74 years, 59% of the patients were male and 41 % female. WHO 2002 classification was as follows: 296 (28 %) Refractory Anemia, 347 (33 %) refractory cytopenia with multilineage dysplasia, 94 (9%) refractory anemia with ringed sideroblasts, 46 (4 %) Del 5q, 188 (18 %) refractory anemia with excess of blasts and 78 (7 %) MDS unclassifiable. Median hemoglobin level at baseline was 10.0 g/dL. One hundred and twenty patients (11%) were previously transfused with a Hb baseline level less than 8.0 g/dL; 448 not yet transfused patients (43%) presented a mild anaemia with Hb ranging from 8 to 10 g/dL and 481 patients (46%) had a Hb level higher than 10 g/dL. Patients classification according to IPSS was as follows: low 55% (n=575); int-1 45% (n=474). 335 (32%) out of 1049 patients received EPO alpha treatment while the remaining 714 patients (68%) were supported without any ESAs treatment. The rate of erythroid response to EPO therapy was 61%. Median duration of response was 82 weeks. The response rate to EPO was significantly higher in non transfused than in transfused patients: 69% versus 14% respectively (p value < 0.001). Only 52 out of 1049 patients showed a LE and a higher incidence of LE in EPO treated patients was not observed. Considering the whole group of patients, the OS of EPO treated patients was not statistically different from untreated ones. However EPO responders patients showed a better OS in comparison to both the non responders and non-EPO treated patients (p value < 0.05). When we stratified patients according to the basal hemoglobin level (lower than 8 g/dL, from 8 to 10 g/dL and higher than 10 g/dL) in patients with Hb ranging from 8 to 10 g/dL we observed a significant improvement in OS following successful EPO treatment (median survival: EPO vs. non-EPO 64 vs. 43 months respectively; p 10 g/dL. Even if adverse events were not prospectively recorded in the Registry, no frequent thromboembolic events was reported in EPO group in comparison to non treated patients. Conclusions Our survey can suffer from selection bias due to the limits typical of a voluntary-based registry, but it can be considered a good real life picture of EPO alpha therapy in lower risk Italian MDS patients. Our data confirm that EPOa treatment is globally safe and that patients who can benefit more are the mild anemic untransfused subjects, with baseline Hb level ranging from 8 to 10 g/dL. Disclosures: Santini: Celgene: Honoraria; Novartis: Honoraria; GSK: Honoraria; Janssen: Honoraria.

Decitabine is a pyrimidine nucleoside analogue of cytidine and with azacytidine has been approved for treatment of MDS and non-proliferative CMML. Anyhow, few CMML cases were included in the registrative studies and CMML treatment still remains very arduous, being CMML an heterogenous disease by itself, with characteristics of both MDS and MPN and with a variable prognosis. Decitabine has been suggested to be particularly effective in CMML in a few studies. We evaluated efficacy and safety of decitabine 20mg/m2/day for 5 days every 28 days for a minimum of 6 cycles in a group of pretreated CMML patients. Evaluation of response was performed after 4 and 6 cycles of therapy, according to IWG 2006 criteria. Morphological diagnosis, evaluation of dysplastic signs and monocytic component was performed centrally after first on-site diagnosis. The mononuclear cell expression of decitabine metabolizing enzymes CDA, hENT1, DCK, and CN-II was determined. Between April 2010 and October 2011, 44 patients were enrolled from 15 Italian Hematology centers, 43 had a confirmed centralized diagnosis of CMML (M/F 30/13), and 42 were finally evaluable. According to WHO 27/42 pts were classified CMML-I with organomegaly and 15/42 CMML-II. Among these patients, 46% had received previous therapy. Proliferative CMML was diagnosed in 32/42 cases. Median age was 71 (42-84) yrs, median number of cycles 6 (1-28). Eleven patients received < 4 cycles, 16 from 4 to 6 cycles and 15 > 6 cycles. Eighty one percent of patients with CMML-I received > 4 cycles. All cycles were administered in an out-patient basis. Overall response rate (ORR) was 33%, mCR in 19%, CR in 14.3 %, PR 2.3% and SD in 34.9% and at 3 year-observation time 31% of patients are still alive (13/42). Median duration of response was 9 months. At present, 16.3% (7/42) of patients are still in treatment. Splenomegaly was significantly reduced in 28% of cases. No significative difference in response rate was present in CMML I vs CMML II, nor in proliferative CMML vs dysplastic ones. At analysis of karyotype, 57% of cases had no cytogenetic abnormality and there was no difference in RR between normal and abnormal cases, among which only 5% had complex karyotype. Median OS is 19 months (2-39). For CMML I cases OS is 26 months vs 15 months for CMML II (p Interruption of therapy with decitabine was mainly due to progressive disease (52% of drop-out cases), death (9%) and in 7% of the cases to toxicity of the drug. Grade 3/4 toxicity was in the majority of cases hematological, and present all over treatment courses. In 7/42 patients severe infections occurred, and 1/42 had grade 3/4 cardiac and gastrointestinal toxicities respectively. Conclusions CMML patients even when heavily pretreated, can be responsive to decitabine which can be administered for several cycles with good tolerability. Our data are confirming reported data on ORR and OS after decitabine therapy (Braun et al Blood 2011). OS of decitabine- responding patients is significantly prolonged. No clinical or cytogenetic characteristics seem to be predictive of response to decitabine. No significant difference in enzyme expression was demonstrated, although a trend in lower expression of hENT1 was noted for non responsive cases. Disclosures: Santini: Celgene: Honoraria; Janssen : Honoraria; Novartis: Honoraria; gsk: Honoraria.

tion. Of 22 evaluable pts, 5 had early death (

Background: Erythroid stimulating agents (ESAs) are a common treatment for lower risk anaemic MDS patients (pts). The response rate varies from 20 to 60% according to the different published studies and ESAs are recommended in International Guidelines. Many clinical and biological parameters have been proposed as predictors of response. Little is known about ESAs therapy in real life. Purpose:We performed an analysis based on the network of regional Italian MDS Registries in order to assess which group of patients could benefit more of ESAs treatment. Materials and Methods: From 2487 pts enrolled in the Registry from 1999 to November 2012, we selected a group of 1411 low or intermediate-1 cases with follow-up data available. No differences in leukemic evolution rate have been observed by ESAs treatment (p=0,279). Subsequently, in order to study the effect of ESAs on survival, we excluded the 92 pts who experienced a leukemic evolution and the cases whose evolution was influenced by lenalidomide and/or azacytidine treatments. Cases treated with ESAs other than erythropoietin alpha (EPOa) were excluded because of the low number of observations and/or incomplete information and/or inhomogeneous dosages. Finally, we obtained a data base of 1110 pts treated by EPOa or observation only. We evaluated Overall Survival (OS) according to EPOa treatment considering the degree of anaemia at baseline. Results: The 1110 pts were subdivided according to Hb level as follows: 123 (11%) with Hb 10 g/dL. Three hundred and fifty six pts were treated by EPOa. The difference in OS by EPOa treatment in the whole cohort was not statistically significant. However, when considering patients not yet transfusion dependent with Hb levels ranging from 8 g/dL to 10 g/dL, there was a clear statistically significant difference in OS: median time 216 months in EPOa treated group vs 99 months in non treated pts (p=0,002). The influence of EPOa on OS in pts with Hb 10 g/dL did not reached a significant level. Additionally, no statistically significant difference was observed in deaths for thrombotic events. Conclusions: Our real life experience confirms that EPOa treatment is safe inMDS patients and it is particularly useful in prolonging OS of the selected group of lower risk pts with mild anaemia not yet transfusion dependent.

Abstract 4446 Life expectancy of CML patients has greatly improved in tyrosine-kynase inhibitor (TKI) era, but still some questions remain about the management of suboptimal responders (SR) to imatinib standard dose. This group of patients appears to be heterogeneous, with significant differences in terms of event-free survival between cytogenetic SR and molecular SR (Alvarado et al, Cancer 2009) European Leukemia Net (ELN) recommendations did not clarify those differences and there isn't a clear agreement on SR: maintaining imatinib at standard or higher dose or switching to another TKI are all considered acceptable options (Baccarani et al, JCO 2009). We retrospectively analyzed 63 CML patients, diagnosed in chronic phase between 1988 and 2011, SR to imatinib 400 mg/d, treated according to the 3 different ELN options. Fifty-two patients received imatinib front line and 11 had been previously treated with an interferon based therapy. Sokal score, evaluable in 44 patients, was high in 7, intermediate in 24 and low in 12, respectively. Twenty-five patients were cytogenetic SR and 38 molecular SR. The median follow-up from diagnosis was 76 months (range 10–292). At suboptimal response detection 47 patients (74%) continued imatinib 400 mg/d (30 of them afterword switched to high dose imatinib or new TKI), 12 patients (19%) increased imatinib dose to 600 mg/d (7) or 800 mg/d (5) (8 of them later changed TKI) and only 4 patients switched immediately to new TKI. Twenty-three percent of the 47 patients who continued imatinib 400 mg/d obtained a stable complete cytogenetic response (CCyR) and major molecolar response (MMR) while 27% underwent to a failure. Globally thirty-tree SR patients increased imatinib dose, 36% at suboptimal response detection and 64% after a median of further 12 months of standard dose treatment (range 3–50): 48% of them obtained a durable CCyR and MMR. Twenty-six evaluable patients switched to new TKI (9 to nilotinib and 17 to dasatinib), 13 after high dose imatinib: 62% of the patients achieved a stable CCgR and MMR. Both high dose imatinib and new TKI were significantly more effective in achieving CCyR and MMR than maintaining imatinib 400 mg/d (p Considering separately the subgroup of cytogenetic SR patients, a stable CcyR has been reached by 35% of patients continuing imatinib standard dose, by 50% of the patients who increased imatinib dose and by the totality of the patients treated with new TKI (option significantly superior to the other two, p Among molecular SR, 26% of the patients obtained a stable MMR with imatinib 400 mg/d, 52% with imatinib 600 or 800 mg/d and 63% switching to new TKI. The difference was statistically significant between new TKI and imatinib 400 standard dose only (p Cytogenetic SR maintained at imatinib 400 mg/d had a higher risk of event (defined as loss of hematologic or cytogenetic response or death) compared to molecular SR (40% vs 5%, p Two patients progressed to accelerated phase (clonal evolution) but then obtained an optimal response after switching to new TKI; 2 patients died of unrelated disease. Among the 61 living patients, 71% was in MMR, 26% in CCyR only and 3% didn't reach CCyR (for these patients the efficacy of the therapeutic change is not evaluable yet). In our casistics cytogenetic SR had a higher risk of negative events than molecular SR, as reported in literature, although they obtained similar responses after changing therapeutic strategy. No clear advantage in maintaining imatinib 400 mg/d after suboptimal response was observed, since it led to a few optimal responses and was associated with a significant risk of treatment failure. Imatinib dose increment might represent a more reasonable option, at least for molecular SR. Considering the global casistic no significant difference in response rates were found between new TKI and high dose imatinib even if dasatinib and nilotinib showed a trend towards a superior efficacy in patients mostly unresponsive to the last option, suggesting that an earlier switch to new TKI might further increase the proportion of optimal responders. For cytogenetic SR the switch to new TKI brought better results than those obtained with high dose imatinib: therefore it seems to be the best choice in this subgroup of patients. Disclosures: No relevant conflicts of interest to declare.

Abstract 2192 Background. In adults, ITP displays variable clinical presentation and different response to steroid, Rituximab, and other immune suppressive agents. The pathophysiological differences underlying these different behaviours are mostly unknown and a better knowledge of this biological heterogeneity might help identifying more targeted and rationale treatments for this disease. Purpose. To identify immunogenetic features distinguishing ITP patients from control and different subsets of ITP patients based on clinical presentation and responsiveness to steroid or Rituximab. Several biological analyses were based on the model of autoimmune lymphoprolipherative syndrome (ALPS), a pediatric disease due to genetic defects decreasing function of the Fas death receptor involved in shutting off the immune response and cytotoxic cell function. ALPS displays polyclonal lymphoproliferation, peripheral blood (PB) expansion of TCRαβ+ T-cells double negative for CD4 and CD8 (DN T-cells) and autoimmune manifestations frequently including thrombocytopenia. ALPS is mostly due to deleterious mutations of the Fas gene, but the +1239A>C single nucleotide polymorphism (SNP) of the osteopontin gene (OPN) and several variations of the perforin gene (PRF1) can act as disease modifier. Moreover, defective Fas function and these OPN and PRF1 variations are also displayed by subsets of patients with multiple sclerosis, type-I diabetes mellitus, systemic lupus erythematosus, progressive sclerosis, and chronic inflammatory demyelinating polyneuropathy. Patients and Methods. Adult patients with ITP and matched controls were selected for the analyses and stratified in different clinical subsets according to the disease severity and responsiveness to therapy (asymptomatic, steroid or Rituximab sensitive vs. steroid or Rituximab refractory). Analyses included evaluation of Fas-mediated apoptosis in T cell cultures; proportions of DN-T cells in PB; typing of the +1239A>C SNP of OPN and sequencing of PRF1. All patients were also investigated for TCR monoclonality, whereas BCR monoclonality was analysed in Rituximab-untreated patients only. Results. Analysis of Fas function and DN T-cell expansion was assessed in 100 ITP patients and showed that they displayed higher frequency of defective Fas function than the controls (17/100 vs. 5/100; PC SNP displayed a similar distribution in the patients and the controls. TCR monoclonality was assessed in 76 patients and was detected in 4 of them (5%). BCR monoclonality was assessed in 17 patients in PB and bone marrow and it was always absent. No statistical differences of these parameters were detected comparing patients refractory vs. sensitive to either Rituximab or steroid treatments. However, a trend was found for DN T-cell expansion that tended to be more frequent in Rituximab resistant vs. sensitive patients (0/13 vs. 4/20, P=0.13). Conclusions. These preliminary analyses detected some differences between ITP patients and controls suggesting that defects involved in ALPS development may play a role in adult ITP too. Increasing the patient number is needed to confirm these data and, possibly, to detect differences between clinical subgroups. Disclosures: Off Label Use: Rituximab in ITP.