Methylation of the p15(INK4b) gene in myelodysplastic syndromes is frequent and acquired during disease progression

p15(INK4b) gene is an inhibitor of cyclin-dependent kinase (CDK) 4 and CDK6 whose expression is induced by transforming growth factor (TGF)beta. Recent reports suggest frequent methylation of the p15(INK4b) gene promoter in leukemias, and it has been proposed that this methylation could be necessary for leukemic cells to escape TGF beta regulation. We investigated the methylation status of p15(INK4b) gene in 53 myelodysplastic syndromes (MDS) cases, including nine that had progressed to acute myeloid leukemia (AML), using a recently described sensitive method where polymerase chain reaction (PCR) is preceded by bisulfite modification of DNA (methylation specific PCR). p15(INK4b) methylation was observed in 20 of 53 (38%) of the cases. Twenty of the 24 patients with greater than 10% bone marrow blasts had p15(INK4b) methylation (including all nine patients who had progressed to AML) as compared with none of MDS patients with <10% bone marrow blasts. No correlation between karyotypic abnormalities and methylation status was found. Patients with p15(INK4b) methylation had a worse prognosis, but the prognostic significance of p15(INK4b) methylation was no more found by multivariate analysis, due to its strong correlation to the percentage of marrow blasts. In 10 MDS cases, sequential DNA samples were available. In five of them, methylation of the p15(INK4b) gene was detected at leukemic transformation, but not at diagnosis. Our results showed that methylation of the p15(INK4b) gene in MDS is correlated with blastic bone marrow involvement and increases with disease evolution toward AML. It suggests that proliferation of leukemic cells might require an escape of regulation of the G1 phase of the cell cycle, and possibly of TGF beta inhibitory effect.     

Mate guarding and risk assessment by male mountain baboons during inter-troop encounters

Humanized anti-CD33 monoclonal antibody HuM195 specifically targets myeloid leukemias in vivo and has been shown to produce molecular remissions in patients with acute promyelocytic leukemia who are in clinical remission. Previous human trials have used low intermittent dosing of HuM195 at 3 mg/m2/day, which is adequate to saturate all available CD33 sites in vivo. In the current trial, we investigated supersaturating doses of HuM195. Ten patients with relapsed or refractory myelogenous leukemia (nine acute myelogenous leukemias and one chronic myelogenous leukemia) were treated on days 1-4 and 15-18 with a 4-h daily infusion of HuM195 at three different dose levels: 12, 24, and 36 mg/m2/day. The total maximum dose of HuM195 was 576 mg. The most common toxicities were grade II fever and rigors, seen more frequently at the highest dose. Interestingly, a transient and reversible drop in hemoglobin of 1-3 g/dl was seen during the infusion in several patients. Flow cytometric analysis showed that antigen sites in the peripheral blood and bone marrow (BM) remained saturated with HuM195 during the entire 4-week trial period. At these high doses, the average plasma half-life of HuM195 was approximately 1 week, compared to 38 h, seen in previous studies. Human anti-HuM195 immune responses were not observed. One patient with acute myelogenous leukemia, whose disease was refractory to two rounds of chemotherapy, with < 10% blasts in his BM, achieved a complete remission, lasting > 32 months, at the first dose level. Another three patients showed a reduction in leukemic BM cells. These studies suggest that high doses of HuM195 achieve a long serum half-life, with tolerable toxicity and without immunogenicity. In addition, antileukemic activity was seen.     

Vitamin K2 selectively induces apoptosis of blastic cells in myelodysplastic syndrome: flow cytometric detection of apoptotic cells using APO2.7 monoclonal antibody

We have previously reported that vitamin K2 (VK2) but not VK1 has a potent apoptosis-inducing effect on freshly isolated leukemia cells from patients with various types of leukemia. By multi-color flow cytometric analysis using monoclonal antibody (mAb), APO2.7, which detects mitochondrial 7A6 antigen specifically expressed by cells undergoing apoptosis, we further investigated the apoptosis-inducing effect of VK2 on minor populations of leukemic blast cells in bone marrow from patients with myelodysplastic syndrome (MDS) and overt myeloid leukemia (post-MDS AML). Limiting dilution of CD95 (anti-Fas) mAb-treated apoptotic Jurkat cells with nonapoptotic CTB-1 cells revealed that APO2.7-positive Jurkat cells were consistently detectable by flow cytometry when present at levels of at least 5% in the CTB-1 suspension. In patient samples the gating area for leukemic clone was determined using cell surface antigen-specific mAbs conjugated with either fluorescein isothionate (FITC) or phycoerythrin (PE) and subsequently the cells stained with phycoerythrin cyanine (PE-Cy5)-conjugated APO2.7 mAb were assessed within the gating area of the leukemic clone for monitoring apoptosis. Treatment of the bone marrow mononuclear cells with 3-10 microM of VK2 (menaquinone-3, -4 and -5) in vitro potently induced apoptosis of the leukemic blast cells as compared with the untreated control cells in all 15 MDS patients tested. This effect was more prominent on blastic cells than that on mature myeloid cells such as CD34-/CD33+ gated cells. In addition, VK2 performed much less effectively on CD3-positive lymphoid cells. In contrast to VK2, VK1 did not show apoptosis-inducing activity. These data suggest that VK2 may be used for treatment of patients with MDS in blastic transformation.     

Indications, technique and risks in bone marrow transplantation in adulthood

The option of bone marrow transplantation (BMT) significantly improved prognosis of adult patients with hematologic malignancies aged less than 50 years. Allogeneic BMT using the marrow of an HLA-identical family member still provides the most effective method of BMT. Conventional indications for this form of BMT are chronic myeloid leukemia (CML), acute leukemias presenting with adverse risk factors, myelodysplastic syndromes and severe aplastic anemia. If performed early in the disease course (e.g. during the chronic phase of CML or first remission of acute leukemia and MDS) allogeneic BMT cures 50 to 60% of patients. About 20% die of therapy related complications, e.g. graft versus host disease (GvHD), fatal infections or venoocclusive disease of the liver (VOD) and about 20% of patients succumb to relapse of their hematologic disorder. 80% presenting with severe aplastic anemia can be cured, if allogeneic BMT is performed soon after diagnosis without previous immunosuppressive therapy and blood transfusions. BMT with the marrow of a matched unrelated donor or autologous BMT are increasingly used as alternative procedures. A rate of lethal complications as high as 50% hinders rapid extension of BMT with unrelated donors. Therefore, this form of BMT should be restricted to young patients with leukemias, who cannot achieve long-term remission with conventional chemotherapy (in case of acute leukemias) or alpha-interferon (in case of CML). Reconstitution of hematopoiesis is more rapid after peripheral blood stem cell transplantation (PBSCT) compared with autologous BMT. Therefore, PBSCT will replace autologous BMT in most cases. Most favourable results of PBSCT have been reported in patients with malignant lymphomas after relapse or inferior response to primary induction therapy. Due to the higher relapse rate autologous BMT is inferior to allogeneic BMT in leukemia patients. Trials are required to clarify the potential role of myeloablative therapy with stem cell support in the treatment of patients with solid tumors. Many of the preliminary results already published are unsatisfactory and data of larger trials are still lacking. Therefore, BMT or PBSCT cannot be recommended generally for the therapy of patients with solid tumors.     

Validation of the full and short forms of the CAMDEX interview for diagnosing dementia: evidence from a one-year follow-up study

BACKGROUND: Metaiodobenzylguanidine (MIBG) labeled with 131I has been used for targeted radiotherapy of neural crest tumors, with bone marrow suppression being the primary dose-limiting toxicity. The purpose of this study was to examine the engraftment and toxicity of higher myeloablative doses of 131I-MIBG with autologous bone marrow support. PROCEDURE: Twelve patients with refractory neuroblastoma were given infusions of their autologous, cryopreserved bone marrow following 1-4 doses of 131I-MIBG. The median cumulative administered activity per kilogram of 131I-MIBG was 18.0 mCi/kg (range 14.1-50.2 mCi/kg), the median total activity was 594 mCi (range 195-1,353 mCi), and the median cumulative whole body irradiation from 131I-MIBG was 426 cGy (range 256-800 cGy). A median of 2.5 x 10(8) viable cells/kg (range 0.9-4.7 x 10(8) cells/kg) was given in the bone marrow infusion. RESULTS: All 12 patients achieved an absolute neutrophil count > 500/microliter with a median of 19 days, but only 5/11 evaluable patients achieved red cell transfusion independence, in a median of 44 days; and 4/11 evaluable patients achieved platelet count > 20,000/microliter without transfusion, in a median of 27 days. CONCLUSIONS: Autologous bone marrow transplantation may allow complete hematopoietic reconstitution following ablative 131I-MIBG radiotherapy in patients with neuroblastoma. Risk factors for lack of red cell or platelet recovery include extensive prior chemotherapy, progressive disease at the time of transplant, especially in the bone marrow, and a history of prior myeloablative therapy with stem cell support.     

Radiological reasoning and its computer-based simulation. Reasons to use computer-based diagnostic systems developed on shells

p53 overexpression was studied immunohistochemically in paraffin-embedded bone marrow biopsies using a recently described technique for antigen retrieval based on microwave oven heating of paraffin sections. Using a monoclonal antibody (PAb1801) that reacts with human cellular p53, nuclear staining was detected in 7/11 (63%) therapy-related myelodysplastic syndromes and in 3/4 (75%) therapy-related acute myeloid leukemias. Conversely, staining for p53 was seen only in 9/40 (22%) cases of "primary" hematologic conditions (P < 0.007); these included myelodysplastic syndromes [#2], acute myeloid leukemia [#4], and chronic granulocytic leukemia in accelerated phase or blast crisis [#3]. Biopsies of normal controls and of chronic granulocytic leukemia in stable phase were consistently p53(-). Nine of the 10 karyotyped p53(+) acute myeloid leukemia/myelodysplastic syndrome cases showed complex cytogenetic findings with frequent involvement of chromosome 5 and/or 7. Only four of the 33 karyotyped p53(-) cases showed similar cytogenetic changes. Chromosome 17 involvement was present in four of 13 (31%) cytogenetically assessed p53+ cases, but in none of the p53(-). In univariate analysis, p53 expression in both MDS and AML was significantly associated with shorter survival. The frequent overexpression of p53 in therapy-related myelodysplastic syndromes, therapy-related acute myeloid leukemias and in accelerated phase/blast crisis, chronic granulocytic leukemia and its strong association with complex karyotypes suggests an important role of this gene in the pathogenesis of these leukemic conditions.     

Radioimmunotherapy of medullary thyroid cancer with iodine-131-labeled anti-CEA antibodies

This study evaluates the pharmacokinetics, dosimetry, toxicity and therapeutic potential of radiolabeled NP-4 and MN-14 anti-CEA antibodies in medullary thyroid cancer (MTC). METHODS: Eighteen patients with advanced MTC entered exploratory clinical studies with therapeutic doses of 131I-labeled NP-4 and MN-14 murine monoclonal antibodies (MAbs) reactive with carcinoembryonic antigen (CEA). Doses administered ranged from 46 mCi for 131I-MN-14 lgG to 195 mCi for 131I-MN-14 F(ab)2 in patients negative for human anti-mouse antibodies (HAMA). RESULTS: The radioconjugate blood half-life (T1/2) for the whole lgG was 42.5+/-5.0 hr compared to 18.8+/- 4.1 hr for the bivalent fragments. Tumor doses of 17.5+/-11.0 and 11.4+/-6.3 cGy/mCi were estimated for 131I-MN-14 lgG and F(ab)2, respectively. Tumor/red marrow dose ratios exceeded 3:1 for most lesions. Red marrow doses of up to 350 cGy generally could be delivered with < grade 4 toxicity. Seven of 14 evaluable patients showed evidence of anti-tumor effects lasting up to 26 months, based on physical exam, tumor markers or computed tomography. CONCLUSION: This study demonstrates that anti-CEA MAbs may be suitable for radioimmunotherapy of metastatic or recurrent MTC.     

The elbow joint: osseous and ligamentous structures

PURPOSE: The analogue 131I-metaiodobenzylguanidine (MIBG), which is specifically targeted to neuroblastoma cells, may provide more effective and less toxic treatment for neuroblastoma than conventional external-beam radiotherapy. We report a dose escalation study of 131I-MIBG to define dose-limiting toxicity without and with autologous bone marrow support. PATIENTS AND METHODS: Thirty patients with relapsed neuroblastoma were treated in groups of six with escalating doses of 3 to 18 mCi/kg of 131I-MIBG. After rapid escalation in the first three patients treated at 3 to 6 mCi/kg, treatment was escalated in 3-mCi/kg increments from 9 to 18 mCi/kg. Autologous tumor-free bone marrow was cryopreserved in all patients receiving 12 mCi/kg and more. Toxicity and response were assessed. RESULTS: Eighty percent of patients who received 12 mC/kg or more experienced grade 4 thrombocytopenia and/or neutropenia. Dose-limiting hematologic toxicity was reached at 15 mCi/kg, at which level two of five assessable patients required bone marrow reinfusion for absolute neutrophil count (ANC) of less than 200/microL for more than 2 weeks, and four of nine at the 18-mCi/kg level. Prolonged thrombocytopenia was common, with failure to become platelet-transfusion independent in nine patients. One patient with extensive prior treatment developed secondary leukemia and three became hypothyroid. Responses were seen in 37% of patients, with one complete response (CR), 10 partial response (PR), three mixed response, 10 stable disease, and six progressive disease. The minimum dose of 131I-MIBG for 10 of the 11 responders was 12 mCi/kg. CONCLUSION: Treatment with 131I-MIBG has mainly hematologic toxicity, which can be abrogated with bone marrow rescue. The high response rate in refractory disease suggests that this agent may be useful in combination with myeloablative chemotherapy and autologous stem-cell rescue to improve outcome in advanced neuroblastoma.     

Colony growth in cultures from bone marrow and peripheral blood after curative treatment for leukemia and severe aplastic anemia

The recovery of colony-forming cell numbers after curative treatment for leukemia and severe aplastic anemia (SAA) was studied. We examined 191 patients (85 acute myeloid leukemia [AML], 48 acute lymphocytic leukemia [ALL], 32 chronic myeloid leukemia [CML], 17 SAA, and nine myelodysplastic syndrome [MDS]) who were in hematologic remission 6 months to 13 years after either curative chemotherapy (n = 69) or allogeneic bone marrow transplantation (BMT) (n = 122) by culturing their precursor cells from bone marrow (BM) (n = 548) and peripheral blood (PB) (n = 529) in methylcellulose. Thirty-six BM donors and 25 PB donors served as controls. BM colony-forming cell numbers were abnormally low in all patients (p < 0.002) irrespective of underlying disorder and type of treatment (chemotherapy or irradiation). These numbers did not normalize with time--colony-forming cells were still strongly reduced up to 10 years after therapy, whether or not the patient had received an allogeneic bone marrow graft (p < 0.002). We also compared patients who remained in stable hematologic remission with those who later relapsed (6 months to 2 years after treatment). BM colony-forming cell numbers were significantly lower in patients who subsequently relapsed (p = 0.004). In contrast to BM cultures, we found normal colony-forming capacity by PB precursors in all patients. We conclude that (1) after chemotherapy or BMT, colony-forming cell numbers of BM in culture are permanently reduced; (2) this defect is probably due to a dysfunction of the BM environment rather than to a numerical reduction of the precursor cell pool; and (3) very low colony-forming capacity may be related to relapse.     

Allogeneic peripheral blood progenitor cells for treatment of relapse after bone marrow transplantation

Donor leukocyte infusions (DLI) are an effective therapy for patients who relapse with leukemia after bone marrow transplantation (BMT). Severe graft-versus-host disease and prolonged periods of pancytopenia compromise the success of this treatment in a substantial number of patients. We used filgrastim-mobilized peripheral blood progenitor cells (PBPCs), in some cases preceded by cytoreductive therapy, to circumvent some of the problems associated with DLI. Eleven patients (median age 41 years) received a total of 20 donor cell infusions. Their diagnosis was CML in hematological (two patients) or cytogenetic relapse (two patients), six patients suffered from acute myeloid leukemia (AM; n = 5) or Philadelphia chromosome-positive acute lymphoblastic leukemia (ALL Ph+). One patient had multiple myeloma (MM). All six patients with acute leukemias received cytoreductive therapy prior to PBPC infusions; three patients with CML were pretreated with IFN alpha. Four of four patients with CML responded to PBPC infusions and currently are in complete clinical and molecular remission for time periods between 1 and 12 months. Six of six patients with acute leukemias achieved a complete remission. All of them relapsed after a median remission duration of 24 weeks (range 11-49 weeks). Three patients relapsed at extramedullary sites (CNS, testes, skin). Four of six acute leukemia patients received further cytoreductive therapy. All patients responded again and are in complete remission for time periods between 14 and 615 days. Two patients with acute leukemias have died due to dissemination of the disease. The patient with MM did not respond and is alive with disease. Severe (grade III) acute GVHD developed in two of 11 patients, three patients developed grade II disease, six patients did not show any signs of GVHD. Extensive chronic GVHD has developed in two cases to date. Patients with chemotherapy prior to PBPC infusion developed neutropenia and thrombocytopenia with a maximum duration of 20 and 14 days, respectively; prolonged periods of neutropenia did not occur. Two patients developed long-lasting thrombocytopenia in spite of PBPC infusion, in one case followed by leukemic relapse. Repeated courses of chemotherapy and PBPC infusion were generally tolerated well; no early deaths due to treatment-related toxicity or GVHD were observed. We conclude that the use of allogeneic PBPC instead of DLI in patients with relapse after BMT is technically feasible and safe. The efficacy of PBPC infusions seems comparable to DLI in patients with CML. Patients with acute leukemias also achieved complete albeit transient remissions. Aggressive chemotherapy followed by PBPC infusions resulted in only limited duration of cytopenia. The usage of PBPC infusion instead of non G-CSF-mobilized donor cells for treatment of relapse after BMT may reduce pancytopenia-related complications and merits further investigation.     

Effects of adenosine, ATP, and UTP on chloride secretion by epithelia explanted from fetal rat lung

To determine the role of intensive chemotherapy and allogeneic bone marrow transplantation (BMT) in treatment of refractory anemia with excess of blasts (RAEB) or RAEB-t (in transformation), the outcome of 37 consecutive children, 12 with RAEB and 25 with RAEB-t, diagnosed between 1985 and 1995 was analyzed. Fourteen patients received intensive chemotherapy according to the AML-BFM protocols 83, 87, or 93 (group 1). Seven patients were treated less intensively with the 6-week consolidation phase as induction (group 2). Allogeneic BMT was performed in 10 children of group 1 and 2 after, and in eight (group 3) without prior chemotherapy. Eight children received minimal or no chemotherapy (group 4). Of 21 children (groups 1 and 2) 17 (81%) achieved complete or partial remission after chemotherapy, 12 of them (10 of group 1) remained in remission, eight after BMT. Five-year survival in 29 children treated intensively (groups 1-3) was 46%, SE 12%. Two of the other eight children (group 4) remained alive, one after spontaneous remission. Outcome after BMT was related to the blast count in the bone marrow prior to BMT. None of 10 children (including two with minimal or no chemotherapy) with < or = 12% blasts before BMT relapsed, in contrast to five of eight patients with a higher blast count (P log rank 0.02). We conclude that a substantial number of children with RAEB or RAEB-t can achieve remission with intensive AML-specific chemotherapy. In patients responding to intensive chemotherapy an increase in long-term survival after allogeneic BMT can be expected.     

Fas/APO-1 (CD95) expression in myelodysplastic syndromes

Increased apoptosis of myeloid precursors appears to contribute to the pathophysiology of cytopenias in myelodysplastic syndromes (MDS). Fas /APO-1(CD95) is a cell surface protein inducing an apoptotic signal after its binding to Fas ligand or to a functional anti-Fas antibody. Here we studied Fas expression by immunocytochemistry on marrow slides from 30 cases of MDS. Increased Fas expression in erythroblasts and/or immature granulocytes, compared to controls, was seen in 12 (40%) of the cases. In addition, in 16 of the 18 cases with > or = 5% marrow blasts, a variable proportion of blasts expressed Fas. Increased apoptosis was found by morphological analysis and/or TUNEL technique in marrow cells from 8 of the 26 cases analyzed (31%) The ability of Fas antigen to trigger apoptosis was studied after addition of a functional anti Fas antibody in 5 of the patients with Fas overexpression. Addition of this antibody, however, only lead to mild increase of apoptosis in immature granulocytes (but not other myeloid cells) in 2 of the 5 cases. Thus, increased Fas expression is seen in myeloid and/or blast cells in the majority of MDS cases. However, the relationship between this finding and increased apoptosis in MDS still remains to be established.     

Marrow ablative and immunosuppressive effects of 131I-anti-CD45 antibody in congenic and H2-mismatched murine transplant models

Targeted hematopoietic irradiation delivered by 131I-anti-CD45 antibody has been combined with conventional marrow transplant preparative regimens in an effort to decrease relapse. Before increasing the proportion of therapy delivered by radiolabeled antibody, the myeloablative and immunosuppressive effects of such low dose rate irradiation must be quantitated. We have examined the ability of 131I-anti-CD45 antibody to facilitate engraftment in Ly5-congenic and H2-mismatched murine marrow transplant models. Recipient B6-Ly5(a) mice were treated with 30F11 antibody labeled with 0.1 to 1.5 mCi 131I and/or total body irradiation (TBI), followed by T-cell-depleted marrow from Ly5(b)-congenic (C57BL/6) or H2-mismatched (BALB/c) donors. Engraftment was achieved readily in the Ly5-congenic setting, with greater than 80% donor granulocytes and T cells after 0.5 mCi 131I (estimated 17 Gy to marrow) or 8 Gy TBI. A higher TBI dose (14 Gy) was required to achieve engraftment of H2-mismatched marrow, and engraftment occurred in only 3 of 11 mice receiving 1.5 mCi 131I delivered by anti-CD45 antibody. Engraftment of H2-mismatched marrow was achieved in 22 of 23 animals receiving 0.75 mCi 131I delivered by anti-CD45 antibody combined with 8 Gy TBI. Thus, targeted radiation delivered via 131I-anti-CD45 antibody can enable engraftment of congenic marrow and can partially replace TBI when transplanting T-cell-depleted H2-mismatched marrow.     

Donor lymphocyte infusion for the treatment of relapse after allogeneic hematopoietic stem cell transplantation

The results of donor lymphocyte infusion (DLI) for treatment of relapse after bone marrow transplantation (BMT) are reviewed. Durable complete remission can be achieved at the molecular level for a majority (more than 70%) of patients with CML, when treated at early relapse. Results are less favourable for acute leukemias, although useful responses have been reported. Data are scarce though promising for myelodysplastic syndromes and multiple myeloma. Major treatment-associated toxicities are GVHD and bone marrow aplasia. The latter complication can be predicted by evaluating the level of residual donor-derived hematopoiesis. Modification of infused cells (CD8 negative selection or transduction with a suicide gene), addition of peripheral blood stem cells, and early implementation of escalating doses may counteract the complications and increase the response rate. Response rate is variably influenced by the presence of chronic GVHD after initial BMT, T-cell depleted BMT, underlying disease and stage at relapse, and the level of mixed chimerism. DLI is a direct demonstration of the graft-versus-leukemia effect (GVL). Because GVL after BMT is sometimes the predominant cause of cure, it may be advisable in such situations to redirect the conditioning regimens for BMT towards engraftment and less immediate cytotoxicity.     

Bulky lymphadenopathy in acute myeloid leukemia

Two cases of acute myeloid leukemia (AML) presenting with bulky adenopathy are reported. Both patients were febrile at admission and showed massive and diffuse lymph node involvement, hepatomegaly, and splenomegaly. Erythematopapular leukemic skin lesions were present in one case at the onset and developed in the other at the time of relapse. Anemia, thrombocytopenia, and moderate leukocytosis were present in both. The presence of immature cells in peripheral blood and bone marrow allowed a rapid diagnosis of AML, FAB M1, in one patient. In the other case, owing to the paucity of immature cells in peripheral blood and bone marrow, lymph node biopsy with histology, imprint cytology, and immunocytochemistry were essential for the diagnosis (AML, FAB M2, with trilineage dysplasia and basophilic involvement). Both patients achieved complete remission (CR), followed by an early relapse 3 months later. They underwent allogeneic bone marrow transplantation (BMT) from HLA identical siblings. One patient is actually alive and in CR at 6 months after BMT; the other patient showed a leukemic regrowth after transplantation and died 4 months later.     

Myeloid leukemia and myelodysplastic syndrome relapsing as granulocytic sarcoma (chloroma) after allogeneic bone marrow transplantation

Of 229 consecutive patients receiving allogeneic blood or bone marrow stem cell transplants for acute myeloid leukemia, chronic myeloid leukemia, or myelodysplastic syndrome between 1974 and 1996, 52 patients relapsed. The original tumor recurred as granulocytic sarcoma (chloroma) in three patients (1.3%). Chloroma was found in the ovary in two patients and in the central nervous system in one patient. None of these three patients had experienced > or = grade II acute or more than limited chronic graft-versus-host disease. The intervals between transplantation and recurrence with chloroma were 2, 6, and 13 years. Two patients received a second transplant, and all three died of treatment sequelae.     

Partial-pKD1 plasmids provide enhanced structural stability for heterologous protein production in Kluyveromyces lactis

We describe a patient with leukocytosis with all the stages of neutrophilic series, peripheral dominant myeloblast proliferation, marked dysplasia of myeloid and erythroid series, and extramedullary hematopoiesis of the lymph nodes. A cytogenetic study of the bone marrow cells showed normal karyotype, and molecular analysis of the leukemic cells showed negative for BCR-ABL by RT-PCR. After chemotherapy, the patient went into complete remission with a normal blood and bone marrow profile with no dysplasia. On relapse, the hematological findings showed a typical bone marrow dominant acute myeloid leukemia, with the leukemic cells having a chromosomal abnormality. The patient exhibited the combined features of myeloproliferative disorder, myelodysplastic syndrome, peripheral dominant myeloblast proliferation (so-called peripheral leukemia) and typical acute myeloid leukemia throughout the clinical course. This is thought to be a rare overlapping disease involving these distinct hematological conditions that do not usually occur in the same patient.     

Proteins and cells on PEG immobilized silicon surfaces

Functional adhesion of blood and marrow leukemic cells from 14 acute myeloid leukemia patients presenting with hyperleukocytosis was evaluated by performing cytoadhesion assays on purified (extracellular matrix proteins) and non-purified supports (MRC5 fibroblastic cell line). Results, in 30-min chromium release assay, show a mean +/- S.D. adhesion to fibronectin, collagen, and laminin respectively of 30 +/- 17%, 20 +/- 13%, 25 +/- 17% for blood leukemic cells and 18 +/- 11%, 11 +/- 10%, 11 +/- 8% for marrow leukemic cells. These differences between blood and marrow cells were statistically significant (respectively P = 0.005, P = 0.01 and P = 0.002), while no difference was noted regarding adhesion to non-purified supports. The higher adhesion of blood blast cells to purified supports was observed regardless of CD34 expression. No significant difference was observed in the expression of cell surface VLA-molecules (CD29, CD49b, CD49d, CD49e, CD49f) between blood and marrow blast cells. The addition of GM-CSF or G-CSF induced increased adhesion of marrow blasts and decreased adhesion of blood blasts leading to a loss of the difference between blood and marrow cells. In a 60-min chromium release assay, marrow blasts adhered even more than blood leukemic cells to fibronectin. In contrast, marrow blasts from 'aleukemic' acute myeloid leukemia patients did not show any modification regarding their adhesion to extracellular matrix proteins when co-cultured with growth factors.     

Myelodysplastic syndrome with bone marrow eosinophilia: clinical and cytogenetic features

We investigated the hematological and clinical status of 145 patients with de novo myelodysplastic syndrome (MDS), 14 of whom (10%) had eosinophilia in the bone marrow (MDS-Eo). Most of these 14 patients had severe anemia. Their bone marrow cells exhibited trilineage dysplasia and some morphological abnormalities in the eosinophils, including disproportion of eosinophilic granules, basophilic granules, a ring-shaped nucleus, and vacuolation in the cytoplasm. However, these abnormalities were less prominent than those of acute myelomonocytic leukemia with eosinophilia (FAB: M4Eo). Three of the 14 MDS-Eo patients had refractory anemia (RA), seven had RA with excess of blasts (RAEB), and four had RAEB in transformation. Cytogenetic analysis revealed chromosomal abnormalities in 12 of 13 MDS-Eo patients (92%), in particular, there were major karyotypic abnormalities (MAKA) in eight patients (62%). Cytotoxic agents were not effective in the treatment of four patients after leukemic transformation occurred. These four patients died of the leukemic transformation while seven died of bone marrow failure. The other three MDS-Eo patients are still alive; two of them have already transformed to a leukemic phase. The duration of survival of these patients was significantly shorter than that of the other MDS patients. These findings suggest that bone marrow eosinophilia in MDS may be a poor prognostic factor that is strongly related to the existence of MAKA.