Low IPSS Research Articles

Effect of Erythropoietin Treatment on Lipid Signalling Pathways in Low-Risk MDS Patients.

Abstract 2384Poster Board II-361 Introduction:Phosphoinositide-phospholipase (PI-PLC) C beta1, PI-PLCgamma1 and Akt are key enzymes in nuclear signal transduction pathways, affecting both cell cycle and differentiation in normal physiology and neoplastic transformation. Our group previously showed not only that the Akt/mTOR axis is activated in patients with high-risk MDS (Follo MY et al, Cancer Res 2007), but also that there is an inverse correlation between PI-PLCbeta1 expression and Akt activation (Follo MY et al, Leukemia 2008). Moreover, we recently demonstrated that patients belonging to all of the IPSS risk groups can display a PI-PLCbeta1 mono-allelic deletion, and that this cytogenetic alteration is associated with a higher risk of evolution into Acute Myeloid Leukemia (AML) (Follo MY et al, J Clin Oncol 2009). Erythropoietin (EPO) is an effective treatment of anemia in 40-60% of low-risk MDS, often inducing a prolonged response. Interestingly, the activation of the EPO receptor has been correlated to the PI3K/Akt axis, which in turn is linked to either PI-PLCbeta1 or PI-PLCgamma1 signalling, so that EPO could affect cell proliferation and apoptosis. The aim of this study was therefore to clarify the relationship between EPO treatment and lipid signalling pathways, to investigate their role as molecular targets or predictive factors during EPO therapy. In fact, in patients who are refractory or lose response to EPO there could be a specific activation or inhibition of pathways involved in both cell cycle and differentiation. Patients and Methods:In this study we examined the effect of EPO treatment on lipid signal transduction pathways in MDS patients. The study included 16 patients (IPSS risk: low or intermediate-1), with a favourable response to EPO in 8/16 (50%) of the cases. For each patient we had the opportunity to analyze the expression of PI-PLCbeta1, PI-PLCgamma1, p-Akt and PIP2, which is involved in both PI-PLCbeta1 and PI3K/Akt activation processes, before and during EPO treatment, in order to detect every change in both clinical and biological features. By FISH analysis, we firstly assessed the presence of PI-PLCbeta1 mono-allelic deletion. Then, we quantified PI-PLCbeta1 and PI-PLCgamma1 gene and protein expression, as well as PIP2 and the degree of Akt activation; mRNA levels were quantified by real-time PCR, whereas the protein amount was detected by both a immunocytochemical and a flow cytometric detection approach. Results:The PI-PLCbeta1 mono-allelic deletion was found in 5/16 (31%) low-risk MDS patients: 2 of them showed a rapid evolution into AML, whilst the remaining 3 cases did not respond to EPO treatment. The molecular analyses showed a specific increase in Akt/PI-PLCgamma1 pathway for responder patients, whereas most of the patients refactory to EPO displayed a slight decrease in p-Akt levels and an activation of PI-PLCbeta1 signalling during EPO administration, so that these patients seem to counteract the lack of one PI-PLCbeta1 allele by increasing PI-PLCbeta1 gene and protein expression. Conclusions:Our results, although obtained in a small number of cases, confirm the possible involvement of PI-PLCbeta1 pathways in the EPO signalling. Moreover, our data suggest that the presence of the PI-PLCbeta1 mono-allelic deletion is associated with a worse clinical outcome and with a lack of response to EPO treatment, even in low-risk MDS patients who apparently have a good response profile for EPO (recent diagnosis, absence of long-term transfusion dependence, low or intermediate-1 IPSS risk, serum EPO levels<500 U/L). In fact, in our series, patients with the PI-PLCbeta1 mono-allelic deletion showed an unfavourable outcome (either a rapid evolution into AML or refractoriness to EPO treatment). Moreover, our findings indicate that not only PI-PLCbeta1, but also Akt/PI-PLCgamma1 pathways are critical for cell survival and proliferation in MDS patients treated with EPO. Therefore, these signal transduction pathways could become in the future an important target for the development of innovative strategies for MDS. Disclosures:Finelli:Celgene: Consultancy.

Conventional and Molecular Cytogenetic Responses in Patients with Myelodysplastic Syndrome (MDS) with Deletion 5q Treated with Lenalidomide.

Abstract 1746Poster Board I-772 Background and objectivesInterstitial deletions of the long arm of chromosome 5 - del (5q) - are the most common chromosomal abnormalities in MDS. Del (5q) is detected in 14 % of the patients. Lenalidomide induces complete cytogenetic remissions and abolishes transfusion dependence in a significant proportion of these patients, apparently correlated with the malignant clone suppression. The objectives of this study were to assess clinical and cytogenetic responses to Lenalidomide and to establish the relationship between the number of 5q deleted cells detected by conventional cytogenetics (CG) and by fluorescence in situ hybridization (FISH) previously and after treatment. Due to its larger sensitivity, the FISH technique might be able to predict an eventual loss of response to treatment. Patients and methodsBone marrow samples were collected at diagnosis and every 6 months from the beginning of treatment with lenalidomide. CG and FISH (with LSI EGR1 (5q31)SO/D5S23, D5S721 (5p15.2) probe, from Vysis) studies were performed in all samples, even in the cases that a complete cytogenetic remission was confirmed by karyotype during the treatment follow-up. ResultsWe have treated 9 transfusion dependent MDS patients with lenalidomide, with a median age of 76 years (45-92). Four of the patients had one additional cytogenetic anomaly besides 5q deletion. Due to high clinical and cytological suspicion, one of the patients had a trisomy 9 by karyotype (2/20 metaphase) and was diagnosed of del (5q) by FISH, with a low clonal load (9,4% of deleted nuclei). Six patients were low IPSS risk, while 2 were Int-1 and one was Int-2 risk IPSS. All patients achieved complete erythroid response and transfusion independence (in a median of 8 weeks of treatment) and are still in response. Treatment was stopped in two patients after achieving complete hematologic response, maintaining normal hemoglobine levels for > 24 months. Five patients achieved cytogenetic remission, but 2 of them relapsed in a latter phase of the treatment (25 and 30 months). FISH analysis could not predict cytogenetic relapse in these two cases. ConclusionsOur results support the established high efficacy of lenalidomide in MDS with del (5q). In our experience a single additional cytogenetic anomaly did not compromise hematologic response. Moreover, in our study FISH analysis did not bring additional information to CG analysis in prediction of response and follow-up in the majority of the cases. DisclosuresNo relevant conflicts of interest to declare.

Alemtuzumab Treatment of Intermediate-1 (INT-1) Myelodysplasia Patients Is Associated with Sustained Improvement in Blood Counts and Cytogenetic Remissions.

Abstract 116▪▪This icon denotes an abstract that is clinically relevant.Myelodysplastic syndromes (MDS) are a diverse group of disorders characterized by cytopenias and ineffective hematopoiesis. Experimental evidence links bone marrow failure in a portion of MDS patients to a T cell-dominated autoimmune process. Immunosuppressive therapy (IST) can improve cytopenias in selected patients. Features associated with responsiveness to horse ATG in MDS are young age, low IPSS score, and the presence of HLA DR15 (Sloand E et al J Clin Oncol 2008 26(15):2505-11). The addition of cyclosporine to ATG improves response but continued use of this drug is associated with nephrotoxicity. Alemtuzumab, an anti-CD52 antibody, produces more profound lasting immunosuppression compared to ATG that is associated with transient lymphopenia. We conducted a non-randomized, off label, pilot, Phase II study of alemtuzumab (Campath®) in MDS patients who fit the criteria of those likely to respond to IST (Saunthararajah Y et al Blood. 2003 102(8):3025-7). Consecutive patients fulfilling the inclusion criteria received alemtuzumab 10mg IV for 10 days. Primary endpoints were changes in peripheral blood counts (platelets, absolute neutrophil count, reticulocyte count, hemoglobin). Secondary endpoints (in transfusion-dependent patients) included improvement in the transfusion requirements, duration of response, and late effects of treatment, relapse and survival. Median follow-up time was 11 months (range 1-46 months). Fifteen of 16 (93%) int-1 patients and 2 of 5 (40%) int-2 patients responded by 3 months following infusion. All responders were transfusion independent. Five of seven patients with abnormal cytogenetics at start of treatment had complete cytogenetic remission. One patient with monosomy 7 (constituting 65% of his bone marrow mononuclear cells by FISH) had a complete cytogenetic remission that has lasted 3.8 years thus far. Five of 9 (55%) patients evaluable at 9 months had completely normal blood counts. One patient developed immune thrombocytopenic purpura responsive to rituximab. Two int-2 patients developed leukemia and an additional int-2 patient with a history of smoking succumbed to small cell lung cancer. One death occurred in a non-responding int-1 patient due to disease progression. Two patients had declines in their counts (but not to pretreatment levels) and both responded to CsA. We monitored blood for EBV and CMV reactivation by polymerase chain reaction (PCR) weekly and 4 of the 22 patients became transiently positive for EBV but none developed disease and none of our patients had a significant infection while on drug. The response rate of int-1 patients at 3 months was superior to our previous study using ATG (53%; p=0.0071) but comparable to our results with ATG and CsA (93% p=1.0). Relapse was defined as requirement for additional treatment including cyclosporine; relapse-free survival of responders did not differ from those given ATG or ATG and CsA. The results from this pilot study indicate that Alemtuzumab is highly effective in producing durable responses in selected patients with int-1 MDS. [Display omitted] Disclosures:No relevant conflicts of interest to declare.

SNP-Array Based Karyotyping Complements Routine Cytogenetics in Diagnosis and Risk Stratification Schemes of MDS

While current pathomorphologic criteria distinguish MDS, MDS/MPD and MDSderived AML (sAML), these conditions share common unbalanced chromosomal abnormalities highly predictive of prognosis. Reflecting their clinical importance, IPSS is highly influenced by metaphase cytogenetics (MC), the standard for detection of chromosomal abnormalities. Due to its low resolution and need for cell divisions, MC can only detect abnormalities in 50% of patients with MDS, novel technologies have been developed that could improve the diagnostic yield. Among these methods, single nucleotide polymorphism arrays (SNP-A) have been adopted as a cytogenetic platform. SNP-A karyotyping can be performed on interphase cells and allows for detection of smaller lesions as well as for copy-neutral loss of heterozygosity (LOH). As a result, some reference laboratories offer this test for routine cytogenetic diagnostics. However, the clinical relevance of lesions detected by SNP-A remains unclear and, consequently, we designed this comprehensive study to clarify the clinical impact of cytogenetic results generated through a combined application of SNP-A and MC. A cohort of 352 patients (218 MDS, 59 MDS/MPD, 75 sAML) was studied using Affymetrix 250K (N=160), Affymetrix 6.0 (N=190) and both platforms (N=95). Controls included 362 and 118 samples analyzed by 250k and 6.0 arrays, respectively. CNAGv3 or GCv2.1 software was used for analysis. To avoid biological and technical artifacts, known copy number variants as well as areas of LOH<2.5Mba were excluded. All other lesions, if not present on MC, were confirmed in germ line DNA if possible. The median follow-up time was 29 months (95% CI; 21-38 mo) and for many patients serial samples were studied, showing sequential acquisition of chromosomal defects. Overall, the detection rate of chromosomal abnormalities was improved compared to MC (57% vs. 44%; p=.0096), MDS/MPD (66% vs. 39%, p=.0055) and sAML (63% vs. 45%, p=.048). Somatic uniparental disomy (UPD) accounted for a significant proportion of newly detected defects (26% of new lesions). Overall, UPD was detected in 100 patients (78 as the sole and 22 as an additional lesion). Chromosomal lesions were identified in 98/180 with normal MC. In 61% of patients with non-informative MC (N=18), SNP-A demonstrated an abnormal karyotype; those with newly detected lesions showed inferior outcomes in overall survival (OS) [16 vs. 36 mo, p=.03]. Analysis of OS showed that patients with new defects detected by SNP-A and previously normal MC have worse outcomes compared to those in whom normal chromosomes were confirmed (39 vs. 73 mo, p=.03). Moreover, additional lesions also conferred worse prognosis for patients with abnormal karyotype by MC (17 vs. 38 mo, p=.01). This trend was even more pronounced when specific subgroups were analyzed based on OS and event free survival (EFS). For example, detection of new lesions in MDS (OS p=.02), MDS/MPD (OS p=.009, EFS p=.01) or sAML (OS p=.02, EFS p=.008) results in significant worsening of OS and EFS as compared to patients with truly normal cytogenetics. Newly detected lesions allowed for more precise prognostication within established IPSS strata of MDS and MDS/MPD patients. The detection of SNP-A defects negatively impacted OS/EFS in patients with low IPSS (NR vs. 69 mo, p=<.0001; 42 vs. 112 mo, p=<.0001), decreased EFS in Int-1/2 (24 vs. 29 mo; p=<.0001) and decreased OS/ EFS in high IPSS categories (11 vs. 26 mo, p=<.0001; 7 vs. 14 mo, p=<.0001). In sAML, patients with SNP-A lesions, regardless of MC, have worse OS (6 vs. 21 mo, p=.009) and EFS (5 vs. 13 mo, p=.01). In summary, SNP-A karyotyping facilitates detection of lesions which can complement MC and contribute to a better molecular diagnosis. SNPdetected lesions have impact on prognostic parameters which will likely affect future risk stratification schemes and have prognostic significance regardless of MC in sAML. As a result, new molecular subtypes of MDS may be identified and better-defined recurrent lesions will contribute to identification of causative genes.

Aberrant Phenotype of blasts in Low and Intermediate-1 Risk Myelodysplastic syndromes Predicts Response of Growth Factor Treatment. a Prospective clinical Phase-II Study.

Erythropoietin (Epo) and G-CSF is standard treatment for patients with low and intermediate risk-I myelodysplastic syndromes (MDS) with low Epo levels and transfusion need. Recently, it was demonstrated that flow cytometry adds significantly in the distinction of clinically relevant subgroups in MDS with respect to transfusion dependency and progressive disease. (Van de Loosdrecht et al., Blood 2008, 111) We report a prospective clinical study of 48 patients with low and intermediate-I risk MDS treated with a standardized Epo/G-CSF regimen to address the question whether flow cytometric analysis was instrumental in predicting response. All patients started with Epo if symptomatic at a Hb of less than 6.2 mmol/l independent of endogenous erythropoietin level. Epo (NeoRecormon®) was started at a dose of 30.000U once weekly. In the absence of an increase in Hb of at least 0.6mmol/l within 6 weeks, Epo was escalated to 60.000U/weekly. If still no response was achieved, G-CSF was added (3 times weekly; dose dependent on weight). Hematological response was evaluated according to IWG2006 response criteria. (Cheson, et al. Blood,2006, 108) All patients were scheduled to Epo 60.000U/week and additional G-CSF was dosed in 46 patients. From these patients 17/48 (35%) responded to the standardized Epo/G-CSF regimen. Patients with a low IPSS (0, n=25) showed hematological improvement (HI) in 44% of the cases. Patients with intermediate-I risk MDS (0.5–1.0, n=22) showed a HI in 32% of the cases. When the WHO classification-based prognostic scoring system (WPSS) was used to classify patients into subgroups (Malcovati et al., J Clin Oncol 2007, 25), 43% of the patients with a very low WPSS score (0, n=7) responded to treatment, whereas 67% in the low WPSS (1, n=21) and none in the intermediate and high risk WPSS group (2 or 3, n=20). The levels of endogenous Epo significantly discriminated between responders and non responders (p=0.003). When a cut off of 100U/l was used, 14 of the 22 patients (64%) with an Epo level below 100U/l were responsive to Epo/G-CSF treatment with a median time to response of 11 months (SD 6 mo.); 23/26 patients with high Epo levels were non responsive/Epo levels correctly predicted response to treatment in 77% of the cases (37/48, p<0.001). Strikingly, aberrancies within the myeloid blasts compartment (defined by flow cytometry as CD45dimCD34+SSClow) were mainly seen in non-responding patients (22/31 non responders, 71%). Observed aberrancies were the expression of a lineage infidelity marker (CD5, CD7 or CD56) or a loss of the myeloid antigen CD33. Only 1 out of 17 patients that responded to treatment (6%) showed an aberrant phenotype (CD7); however, response duration in this particular patient was only 4 months as compared to a median time of 12 months in the other cases. The presence of flow cytometric aberrancies correctly predicted response to treatment in 79% of the cases (38/48 cases, p<0.001). None of the responders had both a high Epo level and flow cytometric aberrancies, whereas 3 responsive patients had a relatively high Epo level without flow cytometric aberrancies. Combining Epo levels and flow cytometric aberrancies increased correct prediction of response to therapy to 85%. In 5/8 (63%) of non responders with low Epo levels an aberrant phenotype was observed. Thus, when the presence of flow cytometric aberrancies is taken into account, it may prevent needless treatment with Epo in this group. To conclude, flow cytometry may add significantly to the well known validated predictive parameters for response to select patients who are likely to respond to Epo/G-CSF. The role of flow cytometry in prediction of response to new drugs such as lenalidomide in low/int-I risk MDS is currently under investigation.

Characteristics and Outcome of MDS with Isolated Del 20q: The GFM Experience in 64 Cases.

Background: Isolated del 20q is common in MDS, and considered of good prognosis in the IPSS, but no large series of MDS with del 20q have been reported.Methods: We studied characteristics and outcome of 64 MDS patients (pts) with isolated del 20q diagnosed between 1990 and 2008 in 5 centers of the GFM. They were also compared with MDS without del 20q included in the GFM registry (containing 995 karyotyped MDS diagnosed since 2003).Results: Median age of the 64 MDS with isolated del 20q was 72 (48–92), with 41 males and 23 females. They included 34 RA, 8 RARS, 16 RAEB, 4 CMML and 2 RAEB-T. Fifty two percent of them had less than 5% of marrow blasts, compared to 34% in the non del 20q pts included in the GFM registry (p=0.006). Despite a significantly higher proportion of low and int-1 IPSS risk (90% vs 78%, p=0.021) the isolated del 20q cohort had lower platelet count (mean 156 vs 212 G/l, p=0.014) and higher reticulocyte count (mean 84 vs 52 G/l, p=0.012) than non del 20 q pts included in the GFM. No significant difference was found for age, gender, FAB, ANC and Hb level between isolated del 20q and non del 20q pts. When the analysis was restricted to pts with less than 5% marrow blasts, the same differences were seen between isolated del 20q and non del 20q pts. On the other hand, 12 (19%) of the pts with isolated del 20q had Hb > 12g/dl and platelets < 100 G/L vs 80 of non del 20q patients (p=0.007). The 12 pts with isolated del 20q and plts <100G/L without anemia, were mostly men (9/12), with marrow blasts <5% in 83% (10/12), and all (100%) had ANC >1 G/l. None of those del 20q pts progressed to AML (compared to 17% of isolated del 20 q pts with Hb <12 g/dl and/or plts >100 G/l) and their median overall survival was 39 months vs 29 months, respectively (p=0.32). Median survival of MDS with isolated del 20q was 37 months and was significantly longer in pts with marrow blasts <5% (12 vs 60 months, p=0,0001), with low and int-1 IPSS score (39 vs 11 months, p=0.01) and with Hb >11 g/dl (53 vs 16 months, p=0.002). Survival could not be compared between isolated del 20q and non del 20q pts because of the short observation period of the GFM registry.Conclusion: MDS with isolated del 20q are characterized by usual absence of excess of blasts lower platelets and less anemia than other MDS. About 20% of them present with isolated thrombocytopenia. Prognosis is favorable only in the absence of excess of blasts.

Proposal of a New Prognostic Scoring System for Myelodysplastic Syndromes (MDS) Based on the Immunophenotypic Features of CD34+ Myeloid Precursors

Introduction. Myelodysplastic Syndromes (MDS) are hemopoietic clonal disorders characterized by a heterogeneous clinical course, which is currently predicted on the basis of clinical, morphologic and cytogenetic data. Only a few reports have studied the impact of flow cytometry immunophenotypic data on the prognosis of the disease.Objectives: To evaluate the immunophenotypic characteristics of CD34+ myeloid precursor cells in MDS and their association with the prognosis of the disease.Material & Methods: Overall, 39 patients with MDS - 8 with refractory anemia (RA), 9 RA with ringed sideroblasts (RARS), 12 refractory cytopenias with multilineage dysplasia (RCMD), 2 RA with excess of blasts (RAEB)-1, 5 RAEB-2, 2 with 5q- Syndrome and one unclassifiable MDS - were studied. Bone marrow cells were stained with an extensive panel of MoAbs.Results: the most frequent phenotypic abnormalities on CD34+ myeloid cells were: CD7+ (36%), CD25+ (33%) and CD56+ (15%) followed by overexpression of CD13 and/or CD33 (33%). Abnormal expression of CD7, CD25, CD13 and CD33 were evenly distributed within the WHO and IPSS subgroups of the disease, although CD56 expression was more frequently detected in low risk IPSS cases. From the prognostic point of view, patients that showed altered expression of CD25, CD56 and CD33 had a shorter median overall survival (OS): 30 vs 109 months (p = 0.0003), 19 vs 102 months (p=0.009), and 21 vs 102 months (p=0.009), for CD25, CD56 and CD33, respectively. Reactivity for CD7 and overexpression of CD13 did not influence OS. A phenotypic score was built where a score of 1 was given to abnormal CD25, CD56 or CD33 expression. Significantly different median OS rates (p<0.04) were found for cases with a score of 0 (101±11 months), 1 (47 ±6 months) and >1 (15 ± 5 months).Discussion: Our results show that flow cytometry immunophenotyping of CD34+ myeloid precursors provides clinically useful information for the prognostic stratification of MDS patients. Further studies including larger number of patients with a longer follow-up are necessary to confirm these results.

Improved Leukemia-Free and Overall Survival in Patients with Myelodysplastic Syndrome Receiving Iron Chelation Therapy: A Subgroup Analysis.

Myelodysplastic syndrome (MDS) is characterized by ineffective hematopoiesis and risk of progression to acute myeloid leukemia (AML). Many MDS patients (pts) require red blood cell (RBC) transfusions, risking iron overload (IOL)-related organ dysfunction. We previously showed in a multivariate analysis of 178 pts, improved survival in 18 pts with low and int-1 IPSS risk MDS and IOL receiving iron chelation therapy (ICT), and now examine the effect of ICT on AML-free survival (LFS). The effect of ICT on cytopenias and RBC transfusion requirements (TR); was also examined. To control for possible bias favoring ICT pts, a subgroup analysis was performed. Each of 18 ICT pts had a non-ICT control pt (CP) selected and these groups are matched for gender; neutrophil count (NC), platelet count (PLTC) and hemoglobin (Hb) at diagnosis (Dx); MDS subtype; no of cytopenias, karyotype; IPSS score; ECOG Performance Status; no of serious infections; initial ferritin level (FL); total RBC units (U) received; primary MDS treatment (Rx); and duration of follow-up (FU; all p=NS). Median age in ICT pts was 64 (range 32–70) years (y) vs. 78 (39–81)y for CP (p=0.04). Features of the 178 pts are previously reported. In a univariate analysis (n=178), factors significant for LFS were: MDS subtype; IPSS risk; increased FL; total RBCU transfused; ≥1 serious infection; and receipt of ICT (all p<0.05). Factors significant for LFS in a multivariate analysis (n=178) were: IPSS score (p=0.0001) and receipt of ICT (p=0.03). For low and int-1 IPSS, median LFS was not reached (NR) at 226 months (mo) for ICT pts and 40 mo for non-ICT pts (n=76) and 4y LFS was 64% and 42% respectively (p=0.002). In the subgroup analysis comparing ICT pts to 18 CP with matched features, at a median FU of 51.4 (7.1–225.8) mo and 44.8 (10.1–224) mo respectively, median LFS was NR at 226 mo vs. 40 mo in CP, and 4y LFS was 64% and 49% respectively (p=0.009). Median OS for ICT pts was NR at 226 mo vs. 40.5 mo for CP, and 4y OS was 64% and 49% respectively (p=0.01). There were 5 deaths (28%) in ICT pts (cardiac/IOL, n=2; infection, n=1; other MDS-related, n=1; AML, n=1) and 15 (83%) in CP (other MDS-related, n=6; AML, n=4; bleeding, n=2; infection, n=2; MDS-unrelated, n=1). Although non-ICT pts were older, only 1 death was age-related and all others were from MDS. One ICT pt developed AML at 15 mo from MDS Dx as did 4 non-ICT pts (p=0.06) at a median of 35 (19–71) mo; 2 pts received chemotherapy and both died of progressive AML. In ICT pts, mean ± standard error of the mean (sem) initial/pre-ICT FL was higher than in CP (4038±627 vs. 1759±1108 ug/L respectively, p=0.09), and FU levels decreased for ICT pts (3070±411, p=0.09) but not for CP (2185±996 ug/L, p=NS). There was no difference between ICT pts and CP in mean ± sem initial (2.0±0.3 and 2.0±0.5) and FU (3.4±0.9 and 7.4±4) x109/L NC; initial (236±32 and 104±25) and FU (164±24 and 82±20) x109/L PLTC; or initial (2.6±0.3 and 3.0±0.5) and FU (4.2±0.5 and 4.3±0.9) RBC-TR (no RBCU/4wk; all p=NS). In conclusion, LFS and OS in MDS pts with IOL receiving ICT were improved compared to non-ICT control pts matched for baseline features. These results support findings in the larger cohort of MDS pts and suggest there may be a beneficial effect of ICT on AML transformation and OS. Prospective studies of ICT in MDS pts are warranted.

Changes in Gene Expression Profiles during Treatment with Erythropoetic Growth Factors in Myelodysplastic Syndromes.

Background: Gene expression profiles have been associated with prognosis in myelodysplastic syndromes. WT1 is a tumor-suppressor gene coding for a transcription factor located on chromosome 11p13, which was originally identified for its involvement in the pathogenesis of the Wilms' tumor. In normal bone marrow, WT1 expression is low or undetectable, whereas it is aberrantly expressed in hematological malignancies. Evidence indicates that WT1 is important in the lineage-specific differentiation of hematopoietic cells and leukemogenesis. In myelodysplastic syndromes (MDS), WT1 expression has prognostic significance: it is directly correlated with the type of MDS, with IPSS score and with disease progression. Recent data demonstrate that WT1 is a potent activator of the EPO gene under normoxia and it is suggested that WT1 may regulate paracrine EPO synthesis in a tissue-specific manner. Bmi-1 is a transcriptional repressor gene which may be expressed restrictedly in stem cells and progenitors and is required to regulate the adult self-renewing hematopoietic and leukemic stem cells. It appears that it also plays an important role in providing cells the potential for proliferation. A number of reports on Bmi-1 provide perspectives on the close association of its expression with the progression of hematopoietic malignancies. Furthermore, flow cytometry has shown that Bmi-1 positivity in CD34+ cells is positively correlated with IPSS score.Introduction: We have designed a study to evaluate changes in gene expression profiles of bone marrow mononuclear cells of primary low and intermediate-1 IPSS risk MDS patients receiving erythropoetic growth factors (darbepoetin or high-dose rHuEpo alpha). Associations with response, changes in Hb, in percentage of CD34+ and apoptotic cells are evaluated. We present preliminary results in 6 patients.Methods: Bone marrow samples were obtained before and after 12 weeks of treatment. Mononucleated cells were cryopreserved and later thawed for total RNA extraction and cDNA synthesis. Gene expression profiling and the expressions of WT1 and Bmi-1 by RT-PCR were evaluated.Results: Baseline median Hb was 9,5 g/dL (interquartile range 8,7 10,1). Baseline mean Bmi-1 was 14011 (± SD 3927). All patients had a major erythroid response to treatment. Preliminary results demonstrate a significant decrease in WT1 from baseline median 1635 (interquartile range 1251–2150) to 1192 (interquartile range 1005–1365, P=0.046). [Display omitted] Discussion: Though few patients have yet been studied, it is suggested that WT1 decreases in patients responding to erythropoetic growth factors. Further evaluation with stratification for erythropoetic response in the extended study may furnish novel associations between gene expression, erythropoetic growth factors and prognosis in MDS patients.

Hematopoietic miRNAs Expression in Myelodysplastic Syndromes.

Myelodysplastic syndromes (MDS) are clonal hematopoietic stem-cell disorders that include a broad spectrum of entities characterized by ineffective hematopoiesis and risk of transformation into acute myeloid leukemia (AML). MicroRNAs (miRNA) act as negative expression regulators of important genes as those participating in cellular proliferation, apoptosis and carcinogenesis. In this study we analyzed the expression patterns of 26 miRNA hematopoietic-related genes in patients with MDS. Total RNA was extracted from 20 MDS patients (BM and PB) and 5 normal controls. Median age of patients was 70 (range, 55–82) years. Seven patients had refractory anemia with excess of blasts (RAEB-I, n=5; RAEB-II, n=2), 7 refractory cytopenia with multilineage dysplasia (RCMD), 3 RCMD with ringed sideroblasts (RCMD-RS), 1 chronic myelomonocytic leukemia (CMML), 1 refractory anemia with ring sideroblasts (RARS) and 1 MDS associated with isolated del(5q). 15 patients presented low risk IPSS (LR) (low or intermediate-1) and 5 patients had high risk IPSS (HR) (intermediate-2 and high risk). Twenty-six mature miRNAs were assessed by Stem-loop RT-PCR and Real time PCR in ABI PRISM 7500. MiRNA expression data was normalized to overall median and relative quantification was calculated with the 2−ΔΔCt method. The data were presented as log10 of relative quantity of target miRNA. Median of normal controls was used as calibrator for all samples. Data were analyzed by class comparison methods using BRB Array Tools version 3.4.0 and TIGR multiexperiment viewer. Hierarchical clustering analysis of BM categorized two clusters corresponding to BM-MDS and BM-controls. Then, miR-142-5p (p<0.001) and miR-18a (p<0.001) were differentially expressed between BM-MDS and BM-controls while miR-103 (p=0.003) was overexpressed and miR-18a (p=0.02) was underexpressed in PB-controls with respect to PB-MDS. The analysis of miR-10a and miR-103 in PB (p=0.035 and p=0.031) and miR-181a and miR-10a in BM (p=0.044 and p=0.02) allowed us to discriminate between LR and HR groups. We also found that miR-222 was overexpressed in AML regarding MDS samples (p<0.001). It has been previously described that high activity of miR-222 is necessary to maintain low p27KIP1 levels and continuous proliferation in certain cancer cell lines. In conclusion, this study confirms that miRNAs have a role in the pathogenesis of MDS and their transformation into AML.

Microarray Classification of Myelodysplastic Syndrome (MDS) Identifies Subgroups with Distinct Clinical Outcomes.

The MILE (Microarray Innovations in LEukemia) study has previously shown that gene expression signatures associated with initial leukaemia classifier (LCver7) give an overall cross-validation accuracy of >95% for distinct sub-classes of pediatric and adult leukemias. However, only 50% of the 174 MDS samples in the whole-genome microarray analysis (Stage 1) of the MILE study were correctly identified; the remainder showed AML-like or non-leukemia-like gene profiles. An external morphological review (DB & HL) according to FAB and WHO criteria, of the 174 slides was performed independently (blind) which resulted in 6 samples being reclassified as AML and 4 non-leukemia cases excluded from the study. A recently improved, hierarchical based algorithm correctly identified 100% of the confirmed MDS cases. In this study, using LCver7, the confirmed 164 samples had 50% MDS classifications (Class 17), 23.8% non-leukemia classifications (Class 18), and 22.6% AML classifications (Classes 13 or 14) with the remaining 3.7% having a classification tie between 2 or 3 Classes (due to low confidence). No 5q- syndrome patients had an AML call, whilst 68.3% of RAEB2 patients had an AML classification and none were Class 18. Similarly, 95.6% of Low IPSS patients were classified as Class 17 or 18, whilst all patients (n=5) with High IPSS had an AML call. The classification was independent of blast cells: 10.2% of Class 18 calls had >5% blasts; 28.2% of AML-like cases had <5% blasts. Outcome data (132 MDS patients) was correlated with Class: significant difference (p<0.028, (Kaplan-Meier)) was seen in overall survival; with p <0.004 if AML (Classes 13 & 14) was compared to “non AML” (Classes 17 & 18). Statistically significant differences were seen for time to transformation to AML between the classes (p<0.0001) and between AML and “non AML” (p<0.00007, Kaplan-Meier)) with a probability of transformation of 44% at 18 months for the AML group compared to <8% for the “non-AML” group. A further linear classifier has been used to discriminate patients who transform to AML within 18 months (poor prognosis) with patients with no transformation after >60 months (good prognostic group). Bioinformatic analysis of molecular mapped functions and canonical pathways showed that cell signalling processes were over-represented when comparing de novo AML (n=204) with MDS, from the MILE study, whilst signal transduction pathways were deregulated when comparing non-leukemia samples (n=71) with MDS. Similar pathways and functions were also deregulated when comparing the correctly classified MDS with Class 17 call against MDS with Class 18 call and MDS with AML Classes 13 or 14 calls. In conclusion, the use of microarrays within the initial study, solely intended for diagnostic purposes, has now evolved towards a position in which novel prognostic value may be gained from distinct gene expression signatures. This has also resulted in a better molecular understanding of the progression from non-leukemia, through MDS into full blown AML.

P116 Comparison of MDS patients with long survival to short survival patients. Analysis of cases with discrepancy between low IPSS and survival

P116 Comparison of MDS patients with long survival to short survival patients. Analysis of cases with discrepancy between low IPSS and survival

C025 Low IPSS score and bone marrow hypocellularity in MDS patients predict haematological responses to anti-thymocyte globulin

C025 Low IPSS score and bone marrow hypocellularity in MDS patients predict haematological responses to anti-thymocyte globulin

Long-Term Clinical Benefit of Lenalidomide (Revlimid) Treatment in Patients with Myelodysplastic Syndrome and Chromosome Deletion 5q.

Background: Myelodysplastic syndromes (MDS) are hematological disorders characterized by chronic anemia, and are often refractory to cytokine therapy. Two phase 2 studies evaluating lenalidomide in MDS (MDS-001 and MDS-003), demonstrated a high frequency of erythroid response in patients with a chromosome 5q31 deletion (List AF et al., N Eng J Med 2005; 352:549–557 and List AF et al., in press). Herein we report on the durability of response with long term follow-up approaching 4 years.Methods: All enrolled subjects were evaluated for frequency and duration of red blood cell transfusion independence (RBC-TI), change in hemoglobin (Hgb), pathologic and cytogenetic response, and safety.Results: Ten 5q31- patients achieved a cytogenetic and/or major erythroid response in MDS-001. Four of these patients have maintained a major response for at least 2 years (range 2.2 to 3.7 years). In MDS-003, 99 (67%) of 148 enrolled patients achieved RBC-TI. Of the 99 responders 83 were Low/Int-1, 3 were Int-2/High and 13 were not classified. Median increase in Hgb from baseline to maximum achieved during RBC-TI was 5.4 g/dL (range 1.1, 11.4). The estimated median duration of response was 115.9 weeks, and 52 (53%) of 99 responders remain on study with ongoing RBC-TI response as of 30 June 2006. Duration of response was at least 52 weeks for 63 (64%), 78 weeks for 52 (53%), and 104 weeks for 36 (36%) of 99 responding patients. Variables (or co-variates) associated with longer duration of TI in multivariate analysis included low baseline transfusion requirement (<4 units per 8 wks, p=0.002), 5q- syndrome (p=0.01) and low IPSS (p=0.02) classifications. Among 85 evaluable patients, major cytogenetic responses were achieved in 37 (44%) and minor responses in 24 (28%). Cytogenetic responses occurred in patients with complex karyotypes as well as those with an isolated 5q31 deletion. The frequencies of major and minor cytogenetic responses were similar among patients with and without 5q- syndrome. The most common drug-related adverse events (AE) were Grade 3/4 neutropenia and thrombocytopenia, reported in 87 (59%) and 74 (50%) of 148 patients, respectively although hematologic AEs led to study discontinuation in only 13 (9%) of patients. Pneumonia was the only serious adverse event that occurred in >10% of patients (15/148; 10%). Adverse events were manageable through supportive treatment and dose reduction.Conclusion: Lenalidomide is an effective treatment in MDS patients with an associated chromosome 5q31 deletion, with RBC-TI and major erythroid response maintained for median duration > 2 years.

Prognostic Factors and Response Duration in 419 MDS Treated with Erythropoietin±GCSF: The GFM Experience.

Background: EPO and its derivative darbepoietin alfa (DAR) are important treatments of anemia in lower risk MDS. Prognostic factors of response and of its duration have been recently updated (Blood , 2005, 106, 803–11) and we reanalyzed them in a large series of patients (pts) treated in France and Belgium.Patients: 419 MDS pts were treated with EPO (≥30000UI/wk for at least 12 wks) or DAR (300μg/wk)± GCSF in 25 GFM centers between 1998 and 2006 (160 prospectively analyzed in 3 consecutive trials, and 259 retrospectively analyzed). Median follow-up was 54 months, median age: 73.5 years. WHO classification: RA (14%), RCMD (16%), 5q- syndrome (4%), RARS (21%) RCMD-RS (13%), RAEB-1 (22%), RAEB-2 (6%), and also 4%CMML (FAB); karyotype: 64% FAV, 16% INT, and 4% UNFAV (16% failure or not done). IPSS: 34% LOW, 40% INT-1, 8% INT-2, 2% HIGH (16% unavailable). 185, 126, 80 and 28 pts received EPO alone (alfa or beta), DAR, EPO+G and DAR+G respectively. Median pre-treatment EPO level was 76 UI/l (only 7% pts>500 UI/l). All pts had Hb<10g/dl and 54% required RBC transfusions (including 36% with >2 RBC units/month).Results: 63% pts responded (IWG criteria: 43%HI-E major and 20% HI-E minor), including 57%, 63%, 57%, 66%, 63% with EPO alfa alone, beta alone, EPO+G, DAR alone, DAR+G response (p=ns). Median response duration was 20 mos (range 3–74 mos), 25 and 14 mos for major and minor responses (p= 0.001). Relapse was associated with treatment discontinuation (45%), progression to higher grade MDS (12%) or AML (13%), but without evident cause in 30% cases. In univariate analysis, significantly higher response rates were observed in pts with <10% blasts (p= 0.002), low and INT-1 IPSS score (p=0.001), transfusion <2 RBC units/month (p<0.0001), EPO level<200UI/l (p<0.0001) whereas no significant difference in response rate were seen between RA (69%,) RCMD (72%), RARS (59%), RCMD-RS (71%), RAEB-1 (60%) and 5q- syndrome (52%), and between cytogenetic groups. The response rate in RARS and RCMD-RS was similar with EPO or DAR alone (62.5%), and EPO or DAR+G (60%). In multivariate analysis, EPO <200UI/l (p=0.008), transfusion requirement (p<0.0001) and IPSS (p=0.047) remained predictive factors of response. Longer response duration was significantly associated with blasts<10% (20 mos vs 8 mos for blasts>10%, p=0.007), major response (vs minor), IPSS low-INT-1 (median 22 mos vs 8 mos for INT-2/HIGH, p=0.001) and in pts with absence of multilineage dysplasia (24 mos vs 16 mos, p=0.01). In multivariate analysis, blasts <10% and major response remained predictive factors of longer response.Conclusions:EPO level <200UI/l, RBC transfusions <2 units/month were confirmed as major prognostic factors of response to EPO±G.Good response rates were seen in RAEB-1, and with EPO alone, in RARS and RCMD-RS.Multilineage dysplasia was not associated with lower response rates, but with shorter response duration.Other prognostic factors of shorter response duration were minor response (vs major), and blasts >10%.A case control study with pts of the International MDS risk Analysis Workshop, who received RBC transfusion alone, is in progress to evaluate the impact of EPO treatment on leukemia-free survival and overall survival, and its results will be presented.

Improved Survival in Patients with Myelodysplastic Syndrome (MDS) Receiving Iron Chelation Therapy.

Patients (pts) with MDS and iron overload often receive iron chelation therapy (ICT), although there are no data demonstrating that this improves clinical outcome. Pts with thalassemia receiving ICT do have improved survival and a decrease in number of end-organ toxicities. We performed a retrospective review of 178 pts seen at St.Paul's Hospital in Vancouver, Canada, from January 1981 to April 2006, with a bone marrow diagnosis (Dx) of MDS. Clinical data were collected from the practice database, the Iron Chelation Program of British Columbia database, and by chart review. Pts receiving ICT were treated with desferroxamine 0.5–3g by subcutaneous infusion over 12 hours, 5 days per week. 105 were male and 73 female. MDS Dx were: RA, n=36; RARS, n=42; RAEB, n=28; RAEB-t or AML, n=16, CMMoL, n=25; other, n=31. Age at Dx was a median of 69 (18–94) years. Median absolute neutrophil count (ANC) was 1.6 (33–155) G/l, hemoglobin (Hgb) 96.5 (33–155) G/l, and platelet count 115 (7–644) G/l. Cytogenetic analysis was available in 128 pts; low risk (as defined by the IPSS), n=85; intermediate, n=22; high, n=21. Calculation of IPSS score was feasible in 133 pts; low risk, n=44; int-1, n=55; int-2, n=17; high, n=17. An elevated ferritin level, defined as a serum ferritin of ≥ 2000 ug/ml, was found in 28 pts. Clinical evidence of iron overload was present in 22 pts; CHF with no other contributing factor n=5; liver disease n=18; endocrine dysfunction, n=4; other, n=4; biopsy or imaging evidence was available in 6 pts. Of 18 pts receiving ICT, median duration of ICT was 15 (0–37) months (mo) and reasons for initiating ICT were: elevated ferritin, n=13; clinical and biochemical evidence of iron overload, n=3; number of transfusions received, n=2. In ICT pts, median ferritin level pre-ICT was 4215 (1500–8400) and post-ICT was 2659 (567–5228). In non-ICT pts with elevated ferritin, median ferritin after Dx was 1647 (265–5009) ug/L and at recent follow up was 3188 (763–12723) ug/L. There was a trend toward higher initial ferritin level in ICT pts (p<0.07) and significantly lower post-ICT ferritin in ICT pts compared to follow up ferritin in non-ICT pts (p<0.003). Documented causes of death in non-ICT pts were AML, n=22; MDS-related, n= 21; infection/sepsis, n=18 and non-MDS related, n=10. Documented causes of death in ICT pts were AML, n=1; MDS related, n=1; iron overload, n=1. Kaplan-Meier analysis showed that median overall survival (OS) for all pts was 36 (0.7–255.9) mo. Age showed a trend toward significance for OS (p<0.1); other factors that were significant included IPSS score, (p<0.0001); Dx, (p<0.0001); number of red blood cell units transfused, (p<0.0001); occurrence of ≥1 serious infectious episode, (p< 0.002); AML transformation, (p<0.0001); MDS-directed treatment, (p<0.04); elevated ferritin, (p<0.004); clinical evidence of iron overload, (p<0.001); and ICT, (p<0.001). In Cox regression analysis, the only factors significant for OS were IPSS score (p<0.008) and ICT (p<0.02). For pts with low or int-1 IPSS, median OS for pts receiving ICT was not reached at 160 mo vs. 40.1 (0.7–224) mo for non-ICT pts (p<0.03). In conclusion, although we were not able to demonstrate a decrease in organ dysfunction in pts receiving ICT for MDS, there was a significant improvement in OS. These are to our knowledge the first data documenting improvement in clinical outcome in pts with MDS receiving ICT.

Analysis of Apoptosis Regulatory Genes Expression in the Bone Marrow of Adult De Novo Myelodysplastic Syndromes (MDS).

Background: MDS cover a range of clonal stem cell disorders characterized by ineffective hematopoiesis associated by excessive intramedullary apoptosis of hematopoietic cells. The bcl2 family of proteins constitutes an essential component of the apoptotic machinery at the level of mitochondria and is involved in the acitvation of caspases a family of cytosolic proteases, the effector molecules of apoptosisAim: The aim of the present study was to examine caspases, granzyme B and bcl2 family mRNA expression and the degree of apoptosis in the bone marrow (BM) of adult de novo MDS and to correlate our findings with clinical parameters and prognosis.Methods. We studied 46 cases of MDS including 14 RA, 21 RAEB, 6 CMML and 5 RARS according to FAB criteria. The degree of apoptosis was determined by flow cytometry using the Annexin method and propidium iodine labelling (PI) on BM mononuclear cells. The expression of caspases 1, 2, 3,5,6,7,8 and 9 and Granzyme-B as well as of bclw, bclxL, bclxS, BFL1, BID, Bik, Bak, Bax, bcl2, Mcl1was determined using a multiprobe RNase Protection Assay System (Riboquant, BD Biosciences). A pool of RNA from normal bone marrow mononuclear cells was used as a normal control. The expression of each gene was compared to that of two housekeeping genes (GAPDH and L32) using the Image Master analysis Software. The level of each gene expression was compared to that obtained from the normal pool RNA. The expression of the genes and the degree of apoptosis were analyzed, taking into consideration haematological parameters, the FAB classification and the IPSS value.Results: The median value of apoptosis for all MDS cases was 4,07 (0.04–16.9) Apoptosis in the low risk group was higher but not significantly different from the high risk group (8.9 in the RA-RARS vs 4.3 in the RAEB, and CMML). A positive correlation was found between caspases 8, 5, 3, 2, 1 expression and the level ofapoptosis. Moreover a positive correlation was found between all caspases level of expression with the exception of caspase 7 that did not correlate with caspases 8 and 5. The same phenomenon was observed for the bcl2 family members with the exception of BFL1 that did not correlate with bclw, bclxS and bcl2. The level of expression as well as the percentage of positive cases for all caspases and granzyme B was not significantly different between different FAB subgroups and different IPSS risk categories. BFL1 and Mcl1 levels were significantly higher in patients with BM blasts >5%. Cases with ratio of BID gene expression >1 compared to normal pool were associated with IPSS values <=1. The epxression level of all other bcl2 family members and the percentage of positive cases were not significanlty different between different FAB and IPSS risk categories.Conclusion. The expression level of caspases 1,2,3,5,6,8,9 and granzyme B did not correlate with the FAB, calssification and the IPSS risk groups in patients with MDS. The antiapoptotic genes BFL1 and Mcl1 were significanlty higher in cases with BM blasts >5% while cases overexpressing the proapoptotic gene BID were mostly represented by the low risk IPSS subgroups. Larger number of cases need to be examined to draw definite conclusion about the role of these apoptosis regulatory genes in the pathogenesis and prognosis of MDS.

Expansion of Polyclonal CD4+CD25high Foxp3+ Regulatory T-Cells in High Risk Myelodysplastic Syndromes (MDS).

Foxp3+ regulatory T-cells(Tregs) play a central role in maintaining tolerance. A reduction in Tregs activity is a key feature of autoimmune diseases, whereas their expansion in malignant diseases leads to suppression of the host anti-tumour responses. Expansion of oligoclonal cytotoxic T cells, inhibition of MDS CD34+ progenitors proliferation and response to immunosuppressive therapy suggest that autoimmunity contributes to the pathophysiology of low risk MDS. However, this is not generally a feature of high risk MDS. We hypothesise that in high risk MDS patients an increased number of Tregs may suppress immune responses against the dysplastic clone. In early MDS, these cells may be reduced and or be associated with coexistent autoimmune diseases, an uncommon but recognised association of low risk MDS. We therefore studied CD4+ CD25high Foxp3+ and CD8+ CD25+Foxp3+ Tregs in peripheral blood of MDS patients and examined correlation with bone marrow blasts, cytogenetic Status, IPSS score and progression to AML. Clonality of CD4+CD25+ Tregs was assessed by TCR spectratype analysis of CDR3 size distribution and by CDR3 sequence analysis.52 patients with MDS (30 male, 22 female) with a mean age of 62 years (range 40 to 82 years) were studied. According to WHO classification, 5/52 (10%) had a diagnosis of 5q− syndrome, 9/52(17%) refractory anaemia (RA), 18/52 (35%) refractory cytopenia with multilineage dysplasia (RCMD), 16/52 (31%) refractory anaemia with excess blasts (RAEB) and 4/52 (7%) patients MDS/MPD (one with JAK2 V617F Mutation). Cytogenetic study shows normal pattern in 57%, 5q- in 22%, stable cytogenetic in 10% and complex findings in 11%. All samples were taken at diagnosis prior to any treatment. In addition 5 samples were analyzed pre and post 5-Azacytidine therapy. The absolute number of CD4+CD25highFoxp3+ regulatory T cells in 5q- syndrome was 0.5±0.28×107/l, RA0.6±0.56×107/l, RCMD1.42±0.97×107/l, RAEB2.8±2.2×107/l and MDS/MPD 2.9±2×107/l. In cases with <5% bone marrow blasts (RA, 5q− & RCMD)absolute number and percentage of Tregs was significantly lower than those with ≥5% BM blasts (p=0.001). The mean number was also significantly lower at 0.73±0.57×107/l in low risk cases(IPSS 0) compared with 2±1.5×107/l in intermediate and high risk groups (p=0.008). CD8+ Tregs were not significantly different between the subtypes of MDS and between low and high risk IPSS subgroups. Tregs number did not differ significantly between various cytogenetic subgroups. The spectratype of CD4+CD25+ TCR amplicons, showed a polyclonal pattern and the overall complexity of Vβ subfamilies was not different between low risk and high risk MDS, suggesting that the expanded Tregs in high risk MDS are not clonal and likely to arise by peripheral expansion rather than an antigen-driven response. CD4+ CD25high Foxp3+ Tregs in five patients were studied pre and post 5-Azacytidine. The numbers were significantly decreased after treatment in the one patient who responded to treatment (p=0.001), whereas Treg numbers were unchanged or increased in non responsive cases. In 5 RCMD cases with concomitant autoimmune diseases the percentage of Tregs was lower than other patients within the same subgroup of MDS, however, this did not achieve statistical significance.The findings indicate Tregs are altered in MDS and may be important in the pathophysiology of MDS. Monitoring of Tregs numbers can be a useful indicator for disease progression and response to immunosuppressive therapy.

Fatores de crescimento hemopoéticos nas síndromes mielodisplásicas

Fatores de crescimento hemopoéticos, como a eritropoetina e a associação da eritropoetina com o fator estimulante de colônias de granulócitos (G-CSF) são utilizados para manejo da anemia, particularmente nas síndromes mielodisplásicas de baixo risco (Anemia refratária e Anemia refratária com sideroblastos em anel e mielodisplasias com sistema de escore de prognóstico internacional de risco baixo ou intermediário I). O nível sérico de eritropoetina e a necessidade transfusional pré-tratamento podem identificar pacientes com melhor chance de resposta a essas terapêuticas de alto custo. O uso indiscriminado de G-CSF em portadores de neutropenia crônica não está indicado, devendo ser individualizado.

Karyotype analysis of 283 cases of myelodysplastic syndrome

To explore the implication of karyotype analysis in diagnosis and prognosis of myelodysplastic syndrome (MDS). The chromosomes were prepared with direct method, brief culture of cells and R-banding techniques, and then the karyotypic analysis was performed. Seventy-seven out of 283 patients (27.21%) had karyotypic abnormalities, including the numeral abnormalities of chromosomes and structural alterations. The most common chromosomal aberrations were +8, -20/20q-, -Y, translocation, -7/7q-, +9, -5/5q-. The rate of abnormal karyotype in refractory anemia with erythroblasts (RAEB) and refractory anemia erythroblasts-transformation (RAEB-t) was much higher than in refractory anemia (RA). Patients with abnormal karyotype or higher IPSS scores had a higher risk of transformation into acute leukemia than patients with normal karyotype or lower IPSS scores (P<0.05). MDS is a highly heterogenous disorder and karyotype analysis is helpful for its diagnosis and prognosis estimation.