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Introduction: Myelodysplastic syndromes (MDS) are a heterogeneous group of hematopoietic disorders characterized by ineffective hematopoiesis and peripheral cytopenia, and their possible transformation into acute myeloid leukemia (AML). They typically affect the elderly but, when making treatment decisions, considering chronological age may be insufficient because it poorly correlates with patient frailty: the challenge is to select the optimal treatment in these patients by balancing efficacy and toxicity. Areas covered: This review discusses the rationale for and methods of personalizing the treatment of elderly MDS patients. Expert commentary: Decisions concerning treatment strategies for elderly MDS patients should be made after assessing their frailty on the basis of a geriatric assessment and an estimate of age-adjusted life expectancy. We suggest that all elderly MDS patients should undergo a timed up and go test (TUGT) as a preliminary means of identifying frail patients, and that all non-frail patients should then undergo a comprehensive geriatric assessment (CGA) in order to distinguish fit and pre-frail patients. Fit patients should receive standard dose treatment; pre-frail patients should receive individualized therapy; and frail patients should receive symptom-related therapy. A repeated CGA may be useful to evaluate the hematological, cognitive and socio-relational effects of MDS treatment.

Purpose: The biological and clinical heterogeneity of chronic myelomonocytic leukemia features renders its classification difficult. Moreover, because of the limited knowledge of the mechanisms involved in malignant evolution, chronic myelomonocytic leukemia remains a diagnostic and therapeutic challenge and a poor prognosis disease. We aimed to verify the biological and clinical significance of the discrimination, based on the leukocyte count, between myelodysplastic chronic myelomonocytic leukemia (MD-CMML) and myeloproliferative chronic myelomonocytic leukemia (MP-CMML)., Experimental Design: Peripheral blood samples from 22 patients classified as MD-CMML and 18 as MP-CMML were collected at different time points during disease course, and patients' clinical characteristics were examined. RAS mutational screening was done by sequencing and, for each substitution identified, a highly selective allele-specific PCR was set up to screen all specimens., Results: MP-CMML patients showed a significantly poorer survival (P = 0.003) and a higher frequency of RAS mutations (P = 0.033) by sequencing compared with MD-CMML. Overall, five MD-CMML patients progressed to myeloproliferative disease: in two, allele-specific PCR unveiled low levels of the RAS mutations predominating in the myeloproliferative phase at the time of myelodysplastic disease, documenting for the first time the expansion of a RAS mutated clone in concomitance with chronic myelomonocytic leukemia evolution. Moreover, one of the progressed patients harbored the FLT3-ITD and two MP-CMML patients presented with the JAK2 V617F substitution. All these lesions were mutually exclusive., Conclusions: Our results strongly suggest RAS mutations to function as a secondary event that contributes to development of the chronic myelomonocytic leukemia variant with the poorer prognosis (MP-CMML) and therefore advise their detection to be implemented in chronic myelomonocytic leukemia diagnostics and monitoring.

The main clinical problems of low-risk patients with myelodysplastic syndromes (MDS), as defined by the International Prognostic Scoring System, are infections and the need for frequent transfusions due to ineffective myelopoiesis and peripheral blood cytopenia. Promising results in treating MDS-related anemia have been obtained using high-dose recombinant human erythropoietin (rhEPO). To evaluate the molecular basis of the response to rhEPO, we used commercially available macro-arrays to investigate gene expression profiles in the glycophorin-expressing (Gly+) bone marrow (BM) erythroid cells of five responders (ERs) and five non-responders (ENRs) to rhEPO treatment. The cells were separated by means of positive selection using an immunomagnetic procedure, after which flow cytometry showed that their purity was more than 97% in all cases. The array data were validated by means of real time RT-PCR. The results showed that the genes responsible for proliferation/differentiation and DNA repair/stability were repressed in the BM Gly+ erythroid cells of the ENRs, but almost normally expressed in the ERs. Furthermore, the expression of genes involved in signal transduction suggested that the activity of the MAPK signaling pathway is inhibited in ERs. The different gene expression profiles of ERs and ENRs may provide a basis for early gene testing as a means of predicting the response to rhEPO of MDS patients with low endogenous EPO levels., ((c) 2008 Wiley-Liss, Inc.)

Hyperhomocysteinemia (HH) has been associated with cardiovascular and autoimmune diseases and oxidative cell damage. Myelodysplastic syndromes (MDS) are associated with autoimmunity (AI) and increased oxidative stress. We tested the association of HH and oxidative stress in 33 MDS patients, by measuring plasma homocysteine and malondialdehyde (MDA). HH was found in 42% of cases, (4/5) cases with associated cardiovascular events (CVE)(80%), and 9/15 cases with associated AI (60%). Thus in MDS, HH was significantly associated with AI/CVE (chi(2) : p=0.0011), and this association seems to be specific, as demonstrated by the comparison of MDS presenting AI/CVE with the ischemic cardiopathy/rheumatoid arthritis control group (13/20, 65% vs 19/69, 27%; chi(2) : p=0.0021). The levels of MDA indicated increased oxidative stress. Our data may suggest that in a subset of MDS, HH may simultaneously contribute to bone marrow myelodysplasia, CVE and AI pathogenesis, possibly through oxidative cell damage.

It is now well established that solid tumour growth depends on angiogenesis. However, less is known about the generation of new vessels in haematological malignancies and, in particular, in preleukaemic-myelodysplastic syndromes (MDS). In this study, bone marrow microvessel density (MVD) was assessed by immunohistochemistry and compared in trephine biopsies from 14 controls, five infectious disease (ID), 82 MDS, 15 acute myeloid leukaemia (AML) and 14 myeloproliferative disorder (MPD) patients. Statistical analysis (P < 0.001) demonstrated that MDS MVD was higher than in controls and ID (21 +/- 9 vs 6 +/- 2 and 10 +/- 8 respectively) but lower than AML (30 +/- 12) and MPD (40 +/- 12). Among MDS-FAB subtypes, MVD was significantly higher in RAEB-t, CMML and fibrosis subsets compared to RA, RARS and RAEB subsets (P= 0.008). To further investigate angiogenesis machinery, the expression of vascular endothelial growth factor (VEGF) was evaluated by means of immunohistochemistry in control, MDS, AML and MPD biopsies. Even though VEGF mRNA expression was reported in the past in AML cell cultures and cell lines, in our samples VEGF expression was found to be particularly strong in most of the megakaryocytes but significantly less prominent in other cell populations including blasts. Since our findings suggest a correlation between angiogenesis and progression to leukaemia, additional work is now warranted to determine what regulates the generation of new vessels in MDS and leukaemia.

Secondary myelodysplastic syndrome/acute myelogenous leukemia (MDS/AML) are today considered a primary complication of autologous hematopoietic stem cell transplantation. In our Center, 83 autografted patients underwent bone marrow (BM) biopsy and cytogenetic analysis at fixed intervals. Twelve patients developed non-clonal cytogenetic abnormalities and 10 patients clonal abnormalities, five of whom (three - 7, one - 5 and one t(9;11)) developed secondary MDS/AML. MDS was also diagnosed in two patients with a normal karyotype. In brief, seven patients (three males, four females; median age 36 years) developed MDS/AML 12-48 months (median 14) after autografting. The FAB diagnosis was AML-M2 in one, chronic myelomonocytic leukemia in two and refractory anemia with excess of blasts in transformation in four cases. Two patients presented a BM biopsy picture of MDS with fibrosis; none of them experienced leukemic transformation. Four MDS patients died, three of leukemic transformation and one of BM insufficiency; the two remaining patients are still living and untransformed. Our data underline the leukemogenic role of previous treatments, even if it is not possible to exclude that underlying disease and/or conditioning therapy may be involved.

Immunohistochemistry has been introduced as a means of increasing the diagnostic accuracy of bone marrow biopsy (BMB) in myelodysplastic syndromes (MDS); more recently the possibility of coupling immunostaining with other investigational techniques has broadened the spectrum of applications to the biology and physiopathology of MDS. Using panels of monoclonal antibodies (MoAbs), various histological classifications of MDS have been proposed as an alternative to the FAB criteria. The use of lineage-specific MoAbs has allowed a deeper insight into the dysplastic features of early hematopoietic precursors. The study of various gene products involved in the regulation of cell growth, proliferation and sensitivity to antineoplastic drugs, has revealed significant differences between MDS and morphologically-related disorders, particularly acute myelogenous leukemias (AML); these can be considered markers of a biological difference between the two groups of disorders and deserve consideration when designing therapeutic strategies for MDS. Both an increase in the percentage of cell positivity for the CD34 glycoprotein and a tendency of positive cells towards forming aggregates have been shown to be reliable predictors of leukemic transformation and survival, irrespective of the FAB subtype; furthermore, CD34 positivity has also proved to be a better prognostic factor than the presence of the abnormal localization of immature precursors (ALIP) on BMB. Finally, the simultaneous occurrence of "large" and CD34 positive aggregates can be proposed as a means of recognizing MDS patients with an exceedingly unfavourable prognosis, and who are therefore suitable for early aggressive therapy.

The effect of an ex vivo expansion culture system using multiple cytokine combinations was evaluated in 38 cases of myelodysplastic syndrome (MDS) with the aim of overcoming the defective in vitro growth of haemopoietic progenitor cells. A combination of four growth factors (GF) including SCF, IL-3, IL-6 and GM-CSF was identified as the optimal combination for expanding clonogenic progenitor cells in MDS bone marrow liquid cultures. The cultures of 50% of the patients (19/38) responded to GF stimulation (mean CFU-GM fold increase 15.65+/-48 at week 4) and showed morphological features of normal and/or dysplastic myeloid differentiation. In 12/38 cases (31%), complete unresponsiveness to multiple cytokine stimulation was observed; a small number of patients (7/38) showed progressive leukaemic growth along the cultures with the presence of 100% immature blasts at week 4. GM-CSF and c-kit receptors, analysed by immuno-histochemistry in 10 patients, were over-expressed in responding patients and either lacking or down-regulated in non-responders. Fluorescence in situ hybridization (FISH) analysis of cultured interphase cells of nine patients (trisomy 8 in eight patients) showed a clear-cut increase in the percentage of cells with three signals in the two responding patients, thus indicating the expansion of a MDS clone. Multiple cytokine liquid cultures seem to be able to override the refractoriness of MDS progenitor cells to GF stimulation in many cases, revealing a heterogeneity which may have prognostic implications and should be considered in ex-vivo and in vivo clinical trials with cytokine combinations.

The prognostic impact of bone marrow biopsy (BMB) histology and CD34 immunoreactivity was compared with that of the more conventional parameters (the FAB diagnosis, peripheral blood values, percentage of BM blasts and some common prognostic scores) in 100 MDS patients. Statistical correlations among the cytological, haematological, histological and immunohistochemical parameters and their relationship with clinical outcome were searched for. At univariate analysis, FAB classification (P < 0.001), pattern of blastic infiltration at BMB (P < 0.005), presence of CD34+ aggregates (P < 0.0005), percentage of blasts in BM aspirate (P < 0.0001) and percentage of CD34 positivity (P < 0.0001) proved to be linked to leukaemic transformation and, except for FAB classification, retained a high degree of prognostic significance in terms of survival. Leukaemic transformation occurred in 16/18 patients simultaneously presenting 'large' blastic infiltrates at BMB and CD34+ aggregates (P < 0.00001); 9/17 evaluable patients died within 12 months of diagnosis (P < 0.001)> Discriminant functions for leukaemic transformation and survival did not offer any advantage over univariate analysis in the prognostic work-up. The results indicate that the size of blastic aggregates and CD34 positivity allowed patients with a worse prognosis to be identified irrespective of their FAB subtype, but the prognostic impact is considerably greater when both parameters are simultaneously taken into account, as testified by the restricted and homogeneous subgroup of patients with both 'large' and CD34-positive aggregates.

Stem cell factor (SCF), the ligand of the c-kit receptor, is a potent enhancing cytokine for haematopoietic cells in the presence of IL-3, GM-CSF and erythropoietin (Epo). In the clonogenic assays of 63 MDS patients, the addition of rh-SCF + GM-CSF and/or IL-3 induced a significant increase (p < 0.001) in the number and size of CFU-GM. Never reaching the levels of controls, this increase was seen in all FAB subtypes, but particularly in RA. There was no significant increase in cluster formation, even in RAEB or RAEBt. Rh-SCF (10 ng/ml) led to mean increases of up to 26 times in the number of Epo-dependent BFU-E colonies, particularly in RA (p < 0.001) and RAEB (p < 0.05). Individual responses varied widely (especially in RA) from no response to supranormal levels. Added to the weekly refeed of 37 MDS LTBMC, SCF (10 ng/ml) induced only a 7% mean increase in both cell output and the number of clonogenic cells recovered in the supernatant. Immunohistochemical examination of the supernatant showed significant increases in differentiating myeloid cells in all examined cases, and in erythroid cells in 3 cases; blast cells increased in only 3 cases. These data suggest that rh-SCF is capable of at least partially reversing defective MDS myeloid haematopoiesis, and leads no overt risk of leukaemic transformation. Its potent effect on erythroid cells is encouraging for future clinical applications in patients, particularly if they are selected by means of in vitro tests.

In this study we describe the morphologic and immunohistochemical evaluation of bone marrow biopsies from 14 patients with therapy-related myelodysplastic syndromes (t-MDS). We employed CD34, anti-HLA-Dr, anti-elastase, CD68, anti-glycophorin, CD61 monoclonal antibodies immunostaining, and enzyme histochemistry for chloroacetate esterase. Moreover, we used PC10, a MAb raised against the proliferating cell nuclear antigen, to study the proliferative capacity of these marrows. Our data suggest that diagnosis of refractory anemia with excess of blasts (versus chronic myelomonocytic leukemia), the abnormal localization of immature precursors, marrow fibrosis, and augmented CD34 expression in the bone marrow biopsy are ominous prognostic factors at a statistically significant level (p < 0.0005). A combined morpho-immunohistochemical analysis of bone marrow biopsy correctly classifies t-MDS cases according to the biologic and clinical aggressiveness.

Bone marrow biopsy (BMB) has aroused growing interest as a possible aid in the diagnostic and prognostic evaluation of myelodysplastic syndromes (MDS). Previous reports have pointed out that MDS patients with blastic aggregates or severe bone marrow (BM) fibrosis are characterized by a worse clinical outcome. BMBs of 106 MDS patients were retrospectively reviewed, and relationships among the different histological parameters as well as clinicopathological correlations were looked for. Three patterns of BM blastic infiltration ("diffuse," "cluster," and "large") were recognized. Overt leukemic transformation and overall survival were selected as prognostic end points. BM infiltration was "diffuse" in 18, "cluster" in 48, and "large" in 40 cases. RAEB-t patients accounted for about half of the "large" cases, and none had a "diffuse" pattern (p < 0.01). Nineteen patients showed extensive BM fibrosis; most of them were characterized by "cluster" blastic infiltration and megakaryocyte hyperplasia. Leukemic transformation occurred in 67% of "large" cases (p< 0.001) and in none of the "cluster" cases with severe BM fibrosis (p < 0.01); however, survival was equally poor in these two groups because of early leukemic transformation (large cases) and BM failure (cluster cases). The FAB classification did not significantly correlate with prognosis. Patients with "cluster" BM infiltration and severe fibrosis can be regarded as a true separate MDS subset characterized by unique clinicopathological and prognostic features. Because of the subacute clinical behavior of most cases, and the poor performance status of many elderly patients, there is still controversy as to the best therapeutic approach in MDS. Histological analysis allowed two groups of MDS patients to be identified, both characterized by poor life expectancy, who could benefit from early aggressive chemotherapy.

Bone marrow biopsy (BMB) in myelodysplastic syndrome (MDS) frequently reveals a slight alteration in the reticulin stroma which does not have any clinical significance. However, in a minority of cases, full-blown bone marrow fibrosis (BMF) can be found. Primary MDS patients with BMF show distinct clinico-pathological features and an unfavourable prognosis mainly attributable to complications deriving from pancytopenia and continuous transfusions, while leukemic transformation occurs only rarely. Since BMF may characterize other hematological disorders, primary MDS with BMF should be included in the differential diagnosis particularly with malignant myelofibrosis (MM) and idiopathic myelofibrosis (IMF). Secondary MDS with BMF represent a variety of preleukemic conditions in subjects treated for previous neoplasias. Unlike the primary forms, they do not form a clearcut clinico-pathological entity.

Background: The myelodysplastic syndromes (MDS) are a group of haemopoietic stem cell disorders of unknown cause, occurring predominantly in elderly patients. They have a high mortality due to deaths from complications of cytopenias or transformation to leukaemia. No effective treatment is known. Preliminary studies have identified idarubicin (IDA), an anthracycline drug that can be administered orally, as an agent with possible activity in MDS., Methods: In a phase I-II study we administered oral IDA to 14 predominantly elderly patients (age range 40-79, median 74) with advanced forms of MDS, namely chronic myelomonocytic leukaemia (CMML) (one case), refractory anaemia with excess blasts (RAEB) (7 cases) or refractory anaemia with excess blasts in transformation (RAEB-t) (6 cases). IDA was given at a dose of 30 mg/m2 once each 2 weeks for 8 doses., Results: There was one complete remission (CR) and one partial response (PR), for an overall response rate of 14%. Mean survival from time of initiation of therapy was 10.0 months (+/- 8.0 months [SD]; median, 7.0 months). 5 patients developed severe side-effects, including 3 with life-threatening infections, and there were two deaths from cytopenias during the drug therapy., Conclusions: In this small uncontrolled trial, the overall response rate and extent of toxicity seem similar to those reported for other forms of experimental therapy for MDS, such as low-dose cytosine arabinoside. Further evaluation of the drug's potential in this difficult condition may be warranted.

Seventeen cases of myelodysplastic syndrome (10 primary and seven secondary to previous radio-chemotherapy), characterized by trilineage dysplasia, severe bone marrow fibrosis and a high number of megakaryocytes, are described. All of these patients had similar clinical and prognostic features consisting of pancytopenia, modest or absent visceral enlargement and poor survival. The use of CD61 antibodies, which recognize megakaryocytic cells at all stages of maturation, confirmed that these patients had a higher number of these cells than either normal subjects or patients affected by myelodysplastic syndrome (MDS) without fibrosis. Furthermore, primary and secondary MDS with fibrosis, although clinically and histopathologically similar, differed in terms of the number of megakaryoblasts which were significantly higher in primary forms (P less than 0.02). We conclude that MDS with fibrosis may represent a clinicopathological entity which needs to be distinguished from other MDS subtypes as well as from idiopathic myelofibrosis or malignant myelosclerosis.

Immunohistochemical studies were performed with monoclonal antibodies (MAbs) reactive on paraffin embedded bone marrow biopsies in 19 patients with myelodysplastic syndromes, 8 of them during r gamma-interferon treatment. CD15 MAbs stained mature myeloid cells predominantly located close to the bone marrow trabeculae. Anti-gpIIIa MAbs permitted precise identification of megakaryocytic cells including precursors and dysplastic megakaryocytes. Labelling with CD45 and CD68 MAbs, recognizing lymphocytes and macrophages respectively, was intense in patients in steady state, but progressively decreased during leukemic transformation. Increase in CD45+ and/or CD68+ cells was also observed in most bone marrow biopsies after 3 months of r gamma-interferon therapy.