ALBERT

Description: Background: Severe thrombocytopenia is an uncommon event in lower risk MDS patients, but it may significantly influence the prognosis. In fact, when it occurs, major bleeding may be a life-threatening complication. No licensed pharmacologic approach is nowadays available yet for these patients. Eltrombopag seems to be a very interesting product, but its efficacy and safeness are still to be better demonstrated. Romiplostim could be suitable too, but, at present, its safety is uncertain in MDS patients. Also danazol, an attenuated androgen, seems to have some ability to increase the platelet count in this context. Patients and methods: We retrospectively reviewed 17 thrombocytopenic patients affected by MDS, treated with danazol and observed for at least 6 months. Three patients of these had a therapy-related MDS. At the starting time of danazol therapy, the IPSS was “low” or “intermediate-1” in 16 cases; “intermediate-2” in 1 case. The IPSS-R was “very low”, “low” or “intermediate” in 16 cases; “very high” in 1 case. In 14 patients the platelet count was lower than 25x109/L, in the other 3 lower than 40x109/L, but with spontaneous bleeding. The initial dose was 600 mg/day for all the patients. The IWG criteria of response (Cheson 2006) were adopted. The outcomes were observed after 3 and 6 months from the beginning of therapy. Only descriptive statistical analysis was used. Results: At the beginning of therapy, the average platelet count of the 17 patients was 22.6 x109/L (S.D. 8.8, range 6-38). After 3 months, the therapy with danazol was ongoing in 16 patients (in 1 case the drug was discontinued due to renal failure). Platelet improvement, according to IWG criteria, was observed in 8 cases (47%). The average platelet count was 45.3x109/L (S.D. 32.9, range 4-133). The only one “high risk” patient did not show response. After 6 months danazol was still ongoing in 11 patients (in 5 cases the drug was stopped for inefficacy). The response according to IWG criteria was evident in 9 patients (52% of the initial 17 patients). The average platelet count was 66x109/L (S.D. 63.9, range 11-218). Adverse events recorded were as follows: increase in transaminases in 3 cases (in 2 of these the dose was reduced to 400 mg/day); severe but reversible renal failure in 1 case (the drug was stopped); moderate increasing of serum creatinine in 1 case (the drug was reduced to 400 mg/day); reversible cutaneous rush (the drug was reduced to 400mg/day); amenorrhea in 1 case (the only fertile woman in the series); weight loss and loss of appetite in 1 case, weight gain in 1 case. Conclusions This series confirms the efficacy of danazol to improve platelet count in approximately half of patients with severe thrombocytopenia due to “low-risk” MDS. In all patients with increased platelet count, the response was clinically significant. The response may not be immediate. Actually, there was an improvement of platelet count even after three months of therapy. The toxicity profile of this drug is low. The mechanism of action of danazol in MDS patients remains unclear. Waiting for more information on the efficacy and safety of eltrombopag from the clinical trials in progress, danazol may be a good therapeutic option for these patients. Disclosures Off Label Use: Danazol in MDS patients with severe trhombocytopenia.

Description: Introduction: Although blast-rich specimens immunophenotype studies in myelodysplastc syndromes (MDS) could associate bone marrow (BM) blast expression of CD7 and/or CD117 antigens with poor outcome (Ogata et al., Blood 2002), the prognostic role of markers of myeloid cell immaturity and committment in not enriched BM samples is largely unexplored. Patients and Methods: The expression of CD33, CD34 and CD117 antigens in not enriched BM samples of 50 newly diagnosed MDS was compared with both BM blast WHO category and IPSS score. Immunophenotyping was carried out by using the panel of quadruple monoclonal antibodies CD34/CD117/CD45/CD33, conjugated with the fluorochromes FITC, PE, PerCP, APC, respectively. Acquisition of information on 1x105 stained cells corresponding to the whole BM cellularity was assessed on a dual-laser FACSCalibur flow cytometer using the CellQUEST software (Becton Dickinson, San José CA USA). Multiple group comparisons were made using non parametric ANOVA for BM blasts; general linear model with Wald’s test and Kruskal-Wallis (KW) test to confirm significance was used for IPSS. Results: According to IPSS, 5 (10%) low risk, 27 (54%) intermediate risk-1, 14 (28%) intermediate risk −2 and 4 (8%) high risk pts were identified, respectively. The expression of CD33, CD34 and CD117 significantly correlated with both blast WHO category and IPSS, as shown in the Table 1. Interestingly, by analyzing the subset of 30 pts with BM blasts

Description: Abstract 3097 Poster Board III-34 Backgound Adolescent and young (

Description: Abstract 3951 Poster Board III-887 Background An increased incidence of second cancers has been reported in some lymphoproliferative disorders. Whether the predisposition to other malignancies is due to disease-related immune-suppression rather than to the carcinogenic role of therapy given to treat the hematologic disease is still poorly understood. Purpose The aims of this study were to assess the frequency, characteristics and factors predicting second cancers in patients with WM and to evaluate whether WM patients are at increased risk of developing other malignancies as compared to an age- and sex-matched control population. Patients and methods The clinical records of 220 consecutive WM patients seen in two Hematologic centres of Northern Italy from 1980 to 2008 were reviewed. All cancers, with exception of non-melanoma skin cancers, were considered for this analysis. Standardized incidence ratios (SIRs), defined as the ratio between observed and expected cases, were used to compare the incidence of second cancers in WM with that of the general population. The number of expected cancers was obtained from age, sex and calendar-specific incidence rates reported by AIRTUM (Associazione Italiana Registri Tumori) for Northern Italy. A multivariate Cox proportional hazards regression model was used to examine risk factors for the development of second cancers. Therapy was analyzed as a time-dependent covariate. Results The median age of patients was 61 years (range: 26-86), 131 (60%) were males and 89 females (40%). The median follow-up was 4.8 years (range: 0.5-25). Overall, 136 patients (62%) with symptomatic WM received therapy, whereas a watch-and-wait policy was adopted in 84 patients (38%) with asymptomatic WM. Among treated patients, first-line therapy consisted of chlorambucil in 93 cases (68%), cyclophosphamide-based regimens in 15 (11%) and nucleoside analogs-containing regimens in 17 (13%). Rituximab was associated to chemotherapy in 19 patients (14%). Details on therapy were not available in 11 cases (8%). Overall, 52 second cancers were observed in 49 patients (22%). Forty-six patients (94%) had one malignancy and 3 (6%) developed two cancers. The types of cancer were: gastrointestinal (n=9, 17%), lung (n=8, 15%), breast (n=7, 13%), urinary tract (n=6, 11%), prostate (n=5, 10%), diffuse large B cell lymphoma (n=5, 10%), myelodysplastic syndrome/acute leukemia (n=3, 6%), brain (n=3, 6%), thyroid (n=2, 4%), mouth (n=2, 4%), other cancers (n=2, 4%). The diagnosis of cancer preceded that of WM in 13 cases (27%), was concomitant (within 3 months) in 6 (12%) and subsequent in 30 (61%). The median time from diagnosis of WM to the occurrence of a subsequent cancer was 4.3 years (range: 0.2-12.9). The cumulative probability of developing a second cancer after WM was respectively 11% at 5 years, 21% at 10 years and 33% at 15 years. As compared to the control population, the risk of second cancer in WM was 1.66 times higher than expected (95% CI: 1.16-2.37, p=0.005), without significant difference between males and females (p=0.7). In multivariate analysis, the risk of second cancer was not influenced by age (p=0.91), sex (p=0.45), reduction of IgG (p=0.91) or IgA (p=0.58) levels. In a time-dependent analysis, therapy given for WM did not increase the risk of developing a second malignancy (hazard ratio: 1.12, p=0.76). Conclusions This study shows that WM are at higher risk of developing second cancers as compared to the general population. We did not find an association between the occurrence of second cancers and treatment of WM. Disease-related immune-suppression may predispose WM patients to develop other malignancies. Disclosures: No relevant conflicts of interest to declare.

Description: The role of an intensified program of autologous stem cell transplantation (ASCT) following a consolidation phase with alemtuzumab (A) in pts with chronic lymphocytic leukemia (B-CLL) who received a fludarabine-based regimen (Fbr) as debulking is still considered questionable. The reported evidence of a prolonged treatment free-survival and survival associated to the absence of minimal residual disease (MRD) in B-CLL pts treated with A doesn’t seem to justify an intensification with ASCT in case of MRD-. We have already reported our experience in B-CLL pts treated with Fbr who after a median period of discontinuation of 16 weeks, received A sc (10 mg x 3/w for six weeks) in order to obtain the maximum response of MRD negative remission. Pts obtaining a successful peripheral blood stem cell harvest (PBSC) were considered eligible for ASCT. After a longer follow-up period we analyze here the outcome of autografted pts. Furthermore results were compared with those of pts treated with the same regimen but excluded from transplant procedure. Overall 48 pts have been considered for the analysis. Twenty-nine pts underwent an ASCT. Mobilization regimen consisted in all but 1 pt of Ara-C (800 mg/ sqm/12h x 3 days) followed by granulocyte colony-stimulating factor (G-CSF) while the last patient received only G-CSF. Reason for exclusion from ASCT procedure in the 19 non transplanted pts was: 9 refusal, 4 progressive disease, 1 evolution to Richter syndrome, 2 priming failure, 3 physician decision. Initially, in the group of non-transplanted pts, 9 (47%) were in stage A, 9 (47%) B, 1 (6%) C; ZAP70 was positive in 4 (21%) cases. Response after consolidation with A was: 7 MRD- CR (37%), 7 MRD+ CR (37%), 1 PRn (5%), 4 PR (21%). As regards the transplanted pts: 9 (31%) were in stage Binet A, 16 (55%) B, 4 (14%) C; ZAP70 was positive in 10 (34%) cases. Disease status after A was as follows: 18 MRD- CR (62%), 6 MRD+ CR (21%), 5 PRn (17%). Median age at transplant was 55 years (range 44–64). In all pts a reassessment of response status was ruled out before transplant to exclude a disease progression. ASCT procedure was performed after a median of 12 mos from last A administration (range 6.5–16.8). One pt who reactivated a virus B hepatitis after consolidation was successfully transplanted after 16.8 mos interval from alemtuzumab. Conditioning regimen consisted of 12 Gy total body irradiation plus cyclophosphamide 120 mg/kg in 21 pts

Description: Polycythemia vera (PV) is a chronic myeloproliferative disorder with a propensity to develop myelofibrosis, a condition named post polycythemia vera myelofibrosis (post-PV MF). Survival and prognostic factors after transition to MF remain to be defined. We studied 68 patients with post-PV MF to define survival and prognostic factors for survival at diagnosis of post-PV MF. We also developed a dynamic prognostic model to predict survival at any time from diagnosis of post-PV MF. The median interval between the diagnosis of PV and that of post-PV MF was 13 years (range, 4–29.6 years). Patients with post-PV MF were observed for 181 person-years of follow-up. At diagnosis of post-PV MF, 43 (63%) of 68 patients had less than 65 years. During the follow-up, the incidence of thrombosis was 42 × 1000 person-years (95% CI: 19–93.5) and the incidence of leukemia was 50.3 × 1000 person-years (95% CI: 26–115). The median survival was 5.7 years. Multivariable Cox proportional hazard regression including age, hemoglobin value, platelet count, leukocyte count, and spleen size, showed that hemoglobin 〈 10 g/dL (P 〈 .001) and platelet count 〈 100 × 109/L (P= .026) were independent risk factors for survival. We stratified patients at diagnosis of post-PV MF, according to these factors, obtaining two risk groups with significantly different survival (P = .003): low risk (Hb 〉 10 g/dL and platelet count 〉 100 × 109/L) with a median survival of 7 years, and high risk (Hb 〈 10 g/dL or platelet count 〈 100 × 109/L) with a median survival of 2 years. The prognostic model retained significance after adjustment for age in a multivariable Cox proportional hazard regression (HR: 4.3, 95% CI: 1.6–11.4; P= .003). To assess whether this prognostic model may predict survival at any time from diagnosis of post-PV MF, we evaluated in a time-dependent analysis 64 patients who had longitudinal blood cell counts during follow-up. As first step, we evaluated univariate survival analysis with hemoglobin value 〈 10 g/dL and platelet count 〈 100 ×109/L as time-dependent covariates. Both time-dependent parameters affected survival (HR for hemoglobin 5.8, 95% CI: 2.2–15.2, P 〈 0.001; HR for platelets 4.5, 95% CI: 1.67-12, P=.002). As second step, we evaluated the prognostic model assessed at diagnosis as time-dependent covariate, to define whether the acquisition of one risk factor during follow-up may affect survival. The HR was 7.5 (95% CI: 2.4-23.4; P 〈 .001). The time-dependent prognostic model retained statistical significance after adjustment for age (P 〈 .001). In conclusion, in patients developing post-PV myelofibrosis, a prognostic model based on hemoglobin level 〈 10 g/dL and platelet count 〈 100 × 109/L may predict survival at diagnosis of post-PV MF and at any time thereafter.

Description: α and β-tryptase genes cluster on the short arm of human chromosome 16 and encode lineage-associated serine proteases that are abundantly expressed in mast-cells and, in trace amounts, in basophils. Under physiologic conditions no other myeloid cells express tryptases. However, in several myeloid leukemia cell lines and in AML blasts, the level of tryptase is elevated. In an attempt at correlating the levels of tryptase with cytogenetic features and the KIT and FLT3 mutational status, we analyzed serum samples collected at diagnosis from 150 AML and 57 ALL adult patients. The total serum concentration was determined by UniCAP 100 and UniCAP Tryptase Fluorenzyme Immunoassay Kit (Pharmacia-Upjohn, Uppsala, Sweden). The median value of tryptase level in the control group (50 healthy people; mean age 35 y, range 20–50; M/F= 26/24) amounted to less than 5 ng/ml, ranging from 1 to 15 ng/ml. We detected elevated tryptase levels (more than 15 ng/ml) in 66 out of 150 AML-patients (44%) and in 1 out of 57 ALL-patients (1.75%; median value 1.2 ng/ml) (p = 〈 0.0005, Fisher’s exact test ). In AMLs data showed that elevated tryptase values are significantly bound to patients with t(8;21) (n = 26, p =

Description: Background: Monoclonal antibodies directed towards specific cell surface antigens on neoplastic cells have been increasingly used in lymphoid malignancies in the last few years. Alemtuzumab is a humanized monoclonal antibody which binds the CD52 antigen highly expressed on B-CLL cells. Alemtuzumab significantly depletes both B and T lymphocytes increasing the risk of opportunistic infections. Cytomegalovirus (CMV) infection causes significant morbidity and mortality in the setting of immunodeficiency. CMV reactivation has been described in most of the reported series in which alemtuzumab has been administered. Aim: We investigated the incidence and management of CMV reactivation in a population of B-CLL patients treated with FAMP including regimens, followed by Alemtuzumab as consolidation. Methods: 35 B-CLL patients, 22 males and 13 females, median age 55 (range 39–64), responding to FAMP including regimens (32 FAMP alone, 3 FAMP+cyclophosphamide) received subcutaneous Campath-1H three times a week for six weeks in escalating doses up to 10 mg in order to treat residual disease. Monitoring of CMV reactivation by CMV pp65 antigenemia test was weekly performed, starting from the first week of alemtuzumab treatment until 4 weeks after discontinuation. Results: CMV pp65 antigenemia positive test was detected in 20 patients (57%). The median time for reactivation was 43 days (range 23–61) from the beginning of therapy. Among those twenty pts, 11 showed less than 10 positive cells. All the 9 pts (25.7%) showing positive test with more than 10 cells [median positive cells 44.5 (range 12–751) ] received pre-emptive treatment with oral Gancyclovir 1g thrice daily (oGCV). Only three of this group of 9 pts presented mild symptoms (fever and/or epigastric pain). Among the 11 pts presenting with positive antigenemia in less than 10 cells, seven received oGCV as pre-emptive therapy while the remaining four were not treated. Only one of the seven pts given pre-emptive therapy was symptomatic, presenting fever. The four pts not receiving treatment with oGCV showed a negativity at the subsequent test within a week from the first positive detection. Negativity of antigenemia test was obtained in all the pts receiving pre-emptive oGCV treatment. After short-time of discontinuation, alemtuzumab was restarted in all 20 cases tested positive for pp65 antigenemia. Organ involvement was never seen. Conclusion: Our survey shows that pp65 antigenemia is a sensitive test for monitoring CMV reactivation in pts treated with alemtuzumab. We suggest that the combination of FAMP and alemtuzumab is permissive to a higher risk of CMV reactivation in B-CLL pts. Pre-emptive treatment with oGCV is able to prevent the severe manifestations of CMV such as pneumonia and colitis and to permit the completion of the therapeutic program. To improve the cost-effectiveness of this strategy we propose: antigenemia monitoring not before the 4th week of treatment; closely repeating the test in case of positivity less than 10 cells; treating only in case of increase of the number of positive cells.

Description: Α and β tryptase genes cluster on the short arm of human chromosome 16 and encode lineage-associated serine proteases that are abundantly expressed in mast cells and, in trace amounts, in basophils. Under physiologic conditions no other myeloid cells express tryptases. However, in several myeloid leukemia cell lines and in AML blasts, the level of tryptase is elevated. In vitro, AML blasts are capable to produce and release the α-protryptase constitutively. Recently, elevated levels of serum tryptase have been detected in certain FAB subtypes of AML, particularly in AML-M4eo. In an attempt at correlating the levels of tryptase with FAB classification and cytogenetics, we have analyzed serum samples collected at diagnosis, from 103 AML and 57 ALL patients referred to our Institution. The total serum concentration was determined by UniCAP 100 and UniCAP Tryptase Fluorenzyme Immunoassay Kit (Pharmacia-Upjohn, Uppsala, Sweden). The median value of tryptase level in the control group (50 healthy people; mean age 35 y, range 20–50; M/F= 26/24) amounted to less than 5 ng/ml. We found elevated tryptase levels (more than 15 ng/ml) in 46 out of 103 AML-patients (44.6%) and in 1 out of 57 ALL-patients (1.75%) (p = 89.3 ng/ml) (p = 0.0001), in comparison with other cytogenetic groups (diploid, n= 43; t(15;17), n=10; other, n=29). Our data suggest that elevated serum tryptase levels at diagnosis may play a role as a marker for CBF AML subtypes.

Description: Fludarabine (F) in combination with cyclophosphamide (C) showed a relevant advantage over single-agent F in pts with relapsed CLL. Although minimal residual disease (MRD) remains detectable in many pts achieving CR, the combination of F and C seems to reduce MRD more efficiently. Still, pts in CR eventually relapse and require treatment, demonstrating the need for improved treatments able to further reduce or eliminate MRD and induce “better quality” and thus more durable responses. Alemtuzumab (CAM), anti-CD52 monoclonal antibody, acts synergistically with F in vitro and appears to have synergistic activity in vivo. Additionally, CAM is highly effective at clearing disease from bone marrow, the usual site of residual disease following purine analogue-based treatment. Therefore, we designed a phase II study to determine feasibility and efficacy, overall response rate (ORR)-duration of response-ability at clearing MRD, of a 4-weekly combination regimen consisting of F, C, and CAM (FCC). The study population is represented by pts with B-CLL with relapsed or refractory disease after at least one line of treatment. Subcutaneous route of administration of CAM has been adopted in this trial. MRD was measured by 4-color flow cytometry in the bone marrow. The FCC regimen consisted of F 40 mg/m2/d os (d 1–3), C 250 mg/m2/d os (d 1–3) and CAM 10 mg sc (d 1–3). This combination was repeated on d 29 for up to 6 cycles. The dose of CAM was increased after the first cohort of 10 treated pts from 10 mg to 20 mg sc. Currently, 25 pts have been enrolled in this trial. Median age was 57 years (range 42–79), 15/25 (60%) were male, 23/25 (92%) were in Binet stage B or C, median number of prior treatment regimens was 2 (range 1–4). In six (24%) pts 17p deletion was detected. IgVH unmutated was observed in 17 (68%) pts. At the moment of writing 19 pts are eligible for evaluation of toxicity and response. The ORR was 79%, with 7 (37%) pts achieving CR, 7 (37%) pts a PR, 1 (5%) pt a PRn. Three pts had SD, while 1 showed progression of the disease. MRD negativity was achieved in the bone marrow of 4/15 (27%) pts. Grade III-IV neutropenia episodes were observed in 43% of the administered courses while grade III-IV thrombocytopenia episodes were detected only in 8% of cycles. Four major infections were recorded: two sustained by Mycobacterium tuberculosis (1 cutis, 1 lung), one by Nocardia (lung) and one by E. coli (sepsis). The patient with pneumonia due to M. tuberculosis died because of respiratory failure. CMV reactivation occurred in 6 pts: no CMV disease was recorded. After a median follow up of 10 m (range 1–22) 73% of responding pts did not progressed. In conclusion, results from the interim analysis of this new, 4-weekly dosing FCC regimen suggest that combination therapy with F, C and CAM is feasible, safe, and effective in treating pts with relapsed and refractory CLL, even in those patients with inherent poor prognostic factors and who had received.