Xenophon Yataganas

Prolonged administration of erythropoietin increases erythroid response rate in myelodysplastic syndromes: a phase II trial in 281 patients

Summary. Treatment with recombinant human erythropoietin (rHuEpo) improves anaemia in approximately 20% of patients with myelodysplastic syndromes (MDS). We investigated the potential advantage of a prolonged administration of rHuEpo to achieve higher erythroid response rates (RR) in 281 MDS patients: 118 with refractory anaemia (RA), 77 with refractory anaemia and ringed sideroblasts (RARS), 59 with refractory anaemia with excess of blasts and blast count < 10% (RAEB‐I), and 27 with RAEB and blast count between 11–20% (RAEB‐II). rHuEpo was given subcutaneously at a dose of 150 U/kg thrice weekly, for a minimum of 26 weeks. Response to treatment was evaluated after 12 and 26 weeks of therapy. The overall RR was 45·1%; the RR for RA, RARS, RAEB‐I and RAEB‐II were 48·3%, 58·4%, 33·8% and 13% respectively. A significant increase in RR was observed at week 26 in RA, RARS and RAEB‐I patients, as the response probability increased with treatment duration. The RR was higher in the good cytogenetic prognostic group and serum Epo level of > 150 U/l at baseline predicted for non‐response. The median duration of response was 68 weeks and the overall risk of leukaemic transformation was 21·7%. These results suggest that prolonged administration of rHuEpo produces high and long‐lasting erythroid RR in MDS patients with low blast counts, particularly in those with pretreatment serum Epo levels of < 150 U/l and good cytogenetic prognosis.

Tartrate-resistant acid phosphatase isoform 5b: a novel serum marker for monitoring bone disease in multiple myeloma.

Tartrate‐resistant acid phosphatase isoform‐5b (TRACP‐5b), a new marker reflecting osteoclast activity, and osteoprotegerin (OPG) were measured in 121 patients with multiple myeloma (MM) at diagnosis, and in 63 of them during pamidronate administration, to define their correlation with the extent of bone disease and disease activity in MM. Radiographic evaluation of the skeleton, measurement of other markers of bone remodelling, including N‐terminal cross‐linking telopeptide of type‐I collagen (NTX), bone alkaline phosphatase and osteocalcin and of markers of disease activity (beta2‐microglobulin, paraprotein, interleukin‐6 (IL‐6), were also performed. Levels of TRACP‐5b were increased (p < .0001), while OPG was decreased in MM patients compared to controls (p < .01). TRACP‐5b levels were associated with the radiographically assessed severity of bone disease (p < .0001) as well as with levels of NTX, IL‐6 and beta2‐microglobulin (p < .001, for each biochemical parameter, respectively). The combination of pamidronate with VAD‐chemotherapy produced a reduction in TRACP‐5b, NTX, IL‐6, paraprotein and beta2‐microglobulin levels from the 2nd month of treatment, with no effect on bone formation and OPG. A strong correlation was observed between changes in TRACP‐5b and changes in NTX, IL‐6 and beta2‐microglobulin, while TRACP‐5b predicted the disease progression in 5 patients. These findings suggest that TRACP‐5b is increased in MM, reflects the extent of myeloma bone disease and may have a predictive value. TRACP‐5b has also proved to be very useful for monitoring antimyeloma treatment, which had no effect on OPG levels.

Effect of pamidronate administration on markers of bone turnover and disease activity in multiple myeloma.

Abstract: Aim: Bisphosphonates are potent inhibitors of osteoclastic activity and are used in the treatment of multiple myeloma (MM) in combination with chemotherapy. The effect of pamidronate on markers of bone resorption [cross‐linked N‐telopeptides of type I collagen (NTx)], markers of bone formation [serum alkaline phosphatase (BAP) and osteocalcin (OSC)], interleukin‐6 (IL‐6), β2‐microglobulin, CRP, paraprotein and disease‐related pain and skeletal events has been evaluated in 62 newly diagnosed patients with MM. Patients and methods: The patients were randomly assigned to two groups: the first included 32 patients under chemotherapy and pamidronate (group I) and the second 30 patients on chemotherapy only (group II). Pamidronate was administered at a monthly dose of 90 mg iv, and the above parameters were evaluated at the beginning of this study and after 1, 3, 6, 9, 12 and 14 months of treatment. Results: The addition of pamidronate to chemotherapy resulted in a significant reduction of NTx, IL‐6 and paraprotein from the 3rd month and of β2‐microglobulin, CRP and pain from the 6th month of treatment. No changes of NTx, IL‐6, β2‐microglobulin, CRP or skeletal events were observed in patients of group II, while paraprotein was significantly reduced after 6 months of treatment. The differences in NTx, IL‐6, paraprotein and β2‐microglobulin were statistically significant between the two groups. Multivariate analysis revealed a significant correlation between changes of NTx, changes of IL‐6 in both groups and reduction of pain and paraprotein in group I. Conclusions: These results suggest that pamidronate may have a synergistic action with chemotherapy in decreasing osteoclastic activity, in reducing markers of myeloma activity and myeloma related pain and in improving the quality of life in patients with MM.

Pamidronate is superior to ibandronate in decreasing bone resorption, interleukin‐6 and β2‐microglobulin in multiple myeloma

Abstract: Objectives: Bisphosphonates have been found to reduce skeletal events in patients with multiple myeloma (MM). This is the first randomised trial to compare the efficacy of pamidronate and ibandronate, a third‐generation aminobisphosphonate, in bone turnover and disease activity in MM patients. Methods: Patients with MM, stage II or III, were randomly assigned to receive either pamidronate 90 mg (group I: 23 patients) or ibandronate 4 mg (group II: 21 patients) as a monthly intravenous infusion in addition to conventional chemotherapy. Skeletal events, such as pathologic fractures, hypercalcaemia, and bone radiotherapy were analysed. Bone resorption markers [N‐terminal cross‐linking telopeptide of type‐I collagen (NTX) and tartrate‐resistant acid phosphatase type 5b (TRACP‐5b)], bone formation markers (bone alkaline phosphatase and osteocalcin), markers of disease activity (paraprotein, CRP, β2‐microglobulin), and interleukin‐6 (IL‐6) were also studied. Results: In both groups, the combination of chemotherapy with either pamidronate or ibandronate produced a reduction in bone resorption and tumour burden as measured by NTX, IL‐6, paraprotein, CRP, and β2‐microglobulin from the second month of treatment, having no effect on bone formation. TRACP‐5b also had a significant reduction in the pamidronate group from the second month of treatment and in the ibandronate group from the sixth month. However, there was a greater reduction of NTX, IL‐6, and β2‐microglobulin in group I than in group II, starting at the second month of treatment (P = 0.002, 0.001, and 0.004, respectively) and of TRACP‐5b, starting at the fourth month (P = 0.014), that being continued throughout the 10‐month follow‐up of this study. There was no difference in skeletal events during this period. A significant correlation was observed between changes of NTX and changes of TRACP‐5b, IL‐6, and β2‐microglobulin from the second month for patients of both groups. Conclusions: These results suggest that a monthly dose of 90 mg of pamidronate is more effective than 4 mg of ibandronate in reducing osteoclast activity, bone resorption, IL‐6, and possibly tumour burden in MM. TRACP‐5b has also proved to be a useful new marker for monitoring bisphosphonates treatment in MM.

Somatic hypermutation of immunoglobulin variable region genes: focus on follicular lymphoma and multiple myeloma

Summary: Analysis of the rearranged immunoglobulin variable region gene hypermutation has provided important information concerning the clonal history and ontogenetic origin of various B‐cell lymphoproliferative disorders. Under the selective pressure of antigen, mutational events in immunoglobulin genes will fine tune survival of B‐cell clones bearing immunoglobulin with high affinity for antigen. Our studies aimed at analyzing neoplastic disorders originating from germinal and post‐germinal center B‐cells: follicular lymphoma and multiple myeloma. respectively. Despite the already acknowledged evidence for a selectable distribution of mutations within the clonal immunoglobulin variable heavy chain genes, very little is known about the contribution of light chains in the process of antigen selection. In follicular lymphoma. a more limited pattern of somatic mutation with less evidence of antigen selection was observed in variable K light chain genes (40%) than in their partner heavy chain genes (80%). In myeloma, hypermutation of variable light chain genes, with a distribution suggestive of antigen selection, was frequently observed. Based on these data and recent reports it appears that the light chain expressed by the clonogenic myeloma B‐cells plays a pivotal role in the antigen selection process. Additionally, abortive K light chain variable region genes in X‐expressing myeloma carried a significant number of somatic mutations indicating that the cell of origin is open to the hypermutation machinery at that particular developmental stage irrespective of antigen selection.

Evaluation of the clinical relevance of the expression and function of P-glycoprotein, multidrug resistance protein and lung resistance protein in patients with primary acute myelogenous leukemia.

The multidrug resistance (MDR) transporter-proteins P-glycoprotein (Pgp), multidrug resistance protein (MRP) and lung resistance protein (LRP) have been associated with treatment failure. The aim of this study was to investigate prospectively the clinical significance of expression and function of the MDR proteins, considering other prognostic factors, such as age, immunophenotype, and cytogenetics. Mononuclear cells of peripheral blood or bone marrow from 61 patients with de novo acute myelogenous leukemia (AML) were analyzed. The monoclonal antibodies JSB1, MRPm6 and LRP56 were used for expression studies. Accumulation and retention studies were performed using the substrates Daunorubicin, Calcein-AM, Rhodamine-123 and DiOC(2) in the presence or absence of the modifiers Verapamil, Genistein, Probenecid, BIBW22S and PSC833. Induction treatment consisted of a 3+7 combination of Ida/Ara-C for patients < or = 60 years of age and a 3+5 Ida/VP-16 combination per OS for patients >60. MDR function was expressed as the ratio of mean fluorescence intensity substrate in the presence of modifier over the substrate alone (resistance index, RI). Patients with advanced age, low CD15 expression and high RI for accumulation of DiOC(2) in the presence of BIBW22S had significantly lower complete remission (CR) rates. No factor was prognostic for event-free survival analysis, which was limited to remitters only. Overall survival was shorter in patients with advanced age, poor prognosis cytogenetics, high CD7 expression, and high RI for Daunorubicin efflux modulated by Verapamil. These results suggest that MDR transporter-proteins have a limited role in the treatment failure of patients treated with Idarubicin-based regimens.

Angio-immunoblastic lymphadenopathy terminating as Hodgkin's disease.

The clinical course of a 33‐year‐old man with generalized lymphadenopathy bearing all physical, laboratory and histologic characteristics of “angio‐immunoblastic lymphadenopathy with dysproteinemia” (AILD) is described. Therapy was without significant benefit and the patient died 22 months after initial diagnosis. At autopsy in addition to the characteristic cellular polymorphism of AILD, numerous Hodgkins cells and Sternberg‐Reed cells were identified in the lymph nodes and spleen. Pleomorphic cellular infiltrates containing an increased number of immunoblasts and some giant cells were found also in the portal spaces of the liver. The evolution of Hodgkins disease (H.D.) from AILD suggests that the latter may have represented a reaction to the agent which causes H.D.

Analysis of the κ light chain variable region in multiple myeloma

The study of immunoglobulin heavy chain gene rearrangements in multiple myeloma has revealed extensive divergence from the germline sequences, but no intraclonal diversity with disease evolution. Our study investigated the state of the rearranged κ light chain variable region (Vκ) gene segments, as well as abortive Vκ family gene usage in cases of multiple myeloma expressing λ light chain. We studied 11 cases of κ and five cases of λ light chain‐expressing multiple myeloma. Total cellular RNA was extracted from the bone marrow of patients with overt disease and subjected to reverse transcription‐polymerase chain reaction (RT‐PCR) analysis to amplify clonally rearranged variable region sequences. Direct nucleotide sequencing by the dideoxy‐chain termination method was performed on the RT‐PCR products. We did not observe preferential usage of certain Vκ gene families. Mutation frequencies of the Vκ segments varied in number. In the majority of cases, extensive somatic mutations occurred within the complementarity determining regions (CDRs) of Vκ, whereas only a limited degree of divergence from the germline was observed in others. In all cases studied, replacement mutations tended to cluster in the CDRs, a finding compatible with an antigen‐driven somatic hypermutation process. In 3/5 cases of λ light‐chain expressing multiple myeloma, abortively rearranged Vκ gene segments were amplified from genomic DNA; in two cases a non‐templated nucleotide insertion rendering the Vκ sequences out‐of‐frame was observed, and in the third a stop codon was identified in the open reading frame of the Vκ sequence. Somatic mutations were observed in all cases of abortive Vκ genes studied; however, their distribution does not suggest selection by antigen.

Detection of CD55 and/or CD59 deficient red cell populations in patients with aplastic anaemia, myelodysplastic syndromes and myeloproliferative disorders.

Paroxysmal nocturnal haemoglobinuria (PNH) is an acquired clonal stem cell disorder characterized by intravascular haemolysis, venous thrombosis, marrow hypoplasia, frequent episodes of infection, and rarely leukaemic conversion. At the cellular level, PNH is characterized by the decrease or absence of glycosylphosphatidylinositol (GPI)-anchored molecules, such as CD55 and CD59, from the cell surface. PNH-like clones have been described in various haematological disorders. The link between PNH and aplastic anaemia (AA) has been established but the relationship of PNH with myelodysplastic syndromes (MDS) or myeloproliferative disorders (MPD) remains unclear. In this study, the presence of CD55 and/or CD59 defective (PNH-like) red cell populations was evaluated in 21 patients with AA, 133 with MDS, 197 with MPD, 7 with PNH and in 121 healthy blood donors using the Sephacryl Gel Test microtyping system. Red cell populations deficient in both molecules CD55 and CD59 were detected in 33.3% of AA patients, in 16.5% of MDS patients (50% with hypoplastic bone marrow), in 14.2% of MPD patients (more often in essential thrombocythemia, 21.2%) and in all PNH patients. CD55 deficient red cell populations were found in 14.2% of patients with AA, 12.7% of patients with MDS and 21.3% of patients with MPD. CD59 deficient populations were found in 9.5% of AA patients, 2.2% of MDS patients and 2% of MPD patients. These results indicate an association between PNH, AA and MDS or even between PNH and MPD. Further investigation is necessary to work out the mechanisms of this association, and to define classification criteria for borderline cases, where diagnosis is difficult.

An electron microscopic study of the nuclear abnormalities in erythroblasts in beta-thalassaemia major.

Summary. Erythroblasts in the peripheral blood of 20 splenectomized patients with β‐thalassaemia were examined by electron microscopy. A number of alterations in the structure and integrity of the nucleus and nuclear membrane was noted and these included apparent widening of the nuclear pores, partial absence and areas of reduplication of the nuclear membrane. Intranuclear inclusions were found, some identical to the electron‐dense inclusions (Heinz bodies) present in the cytoplasm, while others contained aggregates of finer particles, probably iron micelles. Some erythroblast nuclei appeared to contain portions of cytoplasm with similar inclusions. Nuclear alterations were most prominent when inclusion bodies were abundant in the cytoplasm and when they were in close approximation to or in contact with nuclear material. These nuclear abnormalities may be related to the recognized disturbance in proliferation of thalassaemic erythroblasts and the ineffective erythro‐poiesis encountered in this disease.