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The effects of teprenone (6,10,14,18-tetramethyl-5,9,13, 17-nonadecatetraen-2-one) on changes in gastric mucosal blood flow, adenosine triphosphate (ATP) content, and incidence of histologic lesions were evaluated in rat gastric mucosa after hemorrhage and retransfusion.Teprenone (100 mg/kg) was administered orally once a day for 3 consecutive days. On the 3rd day hemorrhage was induced, withdrawn blood (retransfusion) was returned, and the above variables were determined.Teprenone significantly inhibited the decreases in blood flow and index of mucosal oxygen saturation (ISO2) during hemorrhage in the corpus and antral mucosa. However, no effect of teprenone was observed on systemic blood pressure and ATP levels after hemorrhage and retransfusion. Teprenone significantly (p0.05) decreased both the incidence of ischemic lesions and the increase in the severity of lesions after retransfusion in both mucosal regions.From these results, it is concluded that the protective effect of teprenone on blood flow was partly responsible for its inhibitory effect on the incidence of lesions in the rat stomach in this hypovolemic shock model, although the former effect might be not a direct effect on systemic vascular tone.

The relationship of gastric mucosal hemodynamics, energy metabolism, and acid to antral histological lesion incidence following hemorrhage and retransfusion was investigated in rats. Decrease of mucosal blood flow and of the index of mucosal oxygen saturation (ISO2) during hemorrhage were slightly greater in the corpus than in the antrum. ATP content and redox ratio in the corpus were decreased by shock; however, these hardly changed in the antrum. The index of tissue hemoglobin concentration (IHb) recovered to above the prehemorrhage value only in the antrum after retransfusion. Histological lesion incidence in the antral mucosa was more markedly increased by acid instillation and/or retransfusion than in the corpus. From these results, it was concluded that the decrease of blood flow is related as an underlying factor to induction of ischemic damage in the antral mucosa and that intraluminal acid and/or postischemic hyperemia play a causative role in the development of histologic lesions in the antral mucosa during shock and retransfusion in the rat stomach.

Overexpression of NF-kappa B reportedly plays anti-apoptotic roles in the growth of AML cells. Control of AML cell growth was attempted using a replication-defective herpes simplex virus-1 vector, T0I kappa B alpha, overexpressing mutant I kappa B alpha to inhibit NF-kappa B in vitro. T0I kappa B alpha displays defective ICP4/ICP22/ICP27, isogenic thymidine kinase, and mutant I kappa B alpha. T0Z.1 expressing lacZ instead of I kappa B was used for controls. Infection of T0I kappa B alpha at 15 multiplicity of infection (MOI) with cells of AML lines, HL60, K562, and NB4 displaying90% infection efficiency and tumor killing in vitro. Use of 10 microM of Ara-C alone was clinically equivalent to high-dose Ara-C, displaying 11% tumor killing. Neither ganciclovir (GCV) nor Ara-C enhanced T0I kappa B- alpha mediated tumor killing. Attenuation of NF-kappa B by T0I kappa B alpha was confirmed by EMSA. T0I kappa B alpha induced caspase-3 activity, with subsequent apoptosis confirmed by colorimetric and TUNEL assays. Fresh AML cells from 8 patients were infected with T0I kappa B alpha at 3 MOI, with or without GCV or 10 microM of Ara-C in vitro. Infection efficiency was 10%. T0I kappa B alpha displayed 8-15% tumor killing, superior to Ara-C in 6 of the 8 patients. Administration of Ara-C enhanced tumor killing in 5 of these 6 cases. Our results suggest that T0I kappa B alpha-mediated gene therapy induces apoptosis of AML cells in vitro.

A 41-year-old man visited his doctor in May 2000 because of a sore throat and high fever. His symptoms did not improve, despite administration of antibiotics and nonsteroidal anti-inflammatory drugs. Since a chest X-ray examination revealed an anterior mediastinal bulky tumor, he was referred and admitted to our hospital on June 21, 2000. The peripheral white blood cell count was 44,540/microliter with 74% myeloblasts. Bone marrow aspiration revealed a hypercellular marrow with 82% myeloblasts, which were negative for peroxidase and alpha-naphthyl butylate esterase staining. Blast cells were positive for CD7, CD13, CD33, CD34, and HLA-DR, and negative for CD56. A needle biopsy specimen of the mediastinal tumor consisted of myeloblasts. We diagnosed the patient as having CD7 (+) acute myeloid leukemia (AML) (M0) with a bulky mediastinal mass based on the surface marker analysis, although the clinical features resembled myeloid/NK precursor acute leukemia. The patient achieved a complete remission after two courses of induction therapy. We are planning an allogeneic stem cell transplantation during his first remission because of the high risk of relapse.

Cefozopran (CZOP) and amikacin (AMK) were used concomitantly to treat infections complicated by hematological diseases. A total of 103 subjects were evaluated, and the all over efficacy rate was 69.9%. Acute leukemia was found in the largest number of patient, 57, followed by 29 cases of malignant lymphoma and 7 cases of myelodysplastic syndrome. By type of infection, patients having unknown origin were the largest in number, being 66, and the efficacy rate was 71.2%. The efficacy rates for sepsis, pneumonia and upper respiratory infection were 42.9% (7 cases), 71.4% (14 cases) and 90% (10 cases) respectively. The efficacy rates by neutrophil counts before administration of CZOP and AMK and at 1 week after administration were both 53.3% in the group of less than 100/microliter, both 60% in the group of less than 500/microliter. The efficacy rate by neutrophil counts at 1 week after administration was 58.6% in the group of less than 100/microliter. The efficacy rate was 75.4% in the group of granulocyte colony stimulating factor (G-CSF) concomitant usage, and 61.9% in the group of non-concomitant usage group. The efficacy rates by serum albumin levels before administration of CZOP and AMK and at 1 week after administration were both 92.9% in the group of over than 4 g/dl, both 50% in the group of less than 3 g/dl. Concomitant treatment with CZOP and AMK exhibited a high level of safety and efficacy rates in infections complicated by hematological diseases.

We evaluated efficacy and safety of monotherapy with CZOP (1-2 g x 2/day) and combination therapy with CZOP (1-2 g x 2/day) and AMK (200 mg x 2/day) for infections in patients with hematological diseases. Efficacy was evaluated in 71 patients of monotherapy group and 70 patients of combination therapy group. Underlying diseases were mostly leukemia and lymphoma. Infections included sepsis, suspected sepsis, pneumonia and so on. Efficacy in CZOP monotherapy was excellent in 21 patients (31.3%), good in 23 patients (34.3%), fair in 5 patients (7.5%) and the efficacy rate was 65.7%. On the other hand, in combination therapy, each was 14 patients (21.2%), 23 patients (34.8%), 12 patients (18.2%) and the efficacy rate was 56.1%. Side effects such as eruption were noted in 2 patients. Abnormal laboratory findings were noted in 9 patients. All side effects as well as abnormal laboratory findings were minimal. It was concluded that CZOP monotherapy was effective in the treatment of various infections accompanying hematological diseases.

We identified a novel human long fatty acyl CoA synthetase 2 gene, ACS2, as a new ETV6 fusion partner gene in a recurrent t(5;12)(q31;p13) translocation in a patient with refractory anemia with excess blasts (RAEB) with basophilia, a patient with acute myelogenous leukemia (AML) with eosinophilia, and a patient with acute eosinophilic leukemia (AEL). ACS2 is expressed in the brain and bone marrow and is highly conserved in man and rats. The resulting ETV6/ACS2 fusion transcripts showed an out-frame fusion of exon 1 of ETV6 to exon 1 of ACS2 in the AEL case, an out-frame fusion of exon 1 of ETV6 to exon 11 of ACS2 in the AML case, and a short in-frame fusion of ETV6 exon 1 to the 3' untranslated region of ACS2 in the RAEB case. Reciprocal ACS2/ETV6 transcripts were identified in two of the cases. Fluorescence in situ hybridization (FISH) analysis with ETV6 cosmids on 12p13, and BACs and P1s on 5q31, demonstrated that the 5q31 breakpoints of the AML and AEL cases involved the 5' portion of the ACS2 gene, and that the 5q31, breakpoint of the RAEB case involved the 3' portion of the ACS2 gene. None of the resulting chimeric transcripts except for the ACS2/ETV6 transcript in the RAEB case led to a fusion protein. Disruption of the second ETV6 allele by t(12;19) was detected in the AML case by FISH analysis. These observations suggest that the disruption of ETV6 and/or ACS2 may lead to the pathogenesis of hematologic malignancies with t(5;12)(q31;p13).

The Wilms tumor gene (WT1) has been reported to be a prognostic factor and a marker for the detection of minimal residual disease (MRD) in acute leukemia. Using competitive polymerase chain reaction procedures, we examined the expression of the WT1 gene in acute leukemia patients with several tumor-specific DNA markers, including bcr/abl, PML/RAR alpha, and AML1/MTG8. A strong correlation was observed between the levels of WT1 and PML/RAR alpha expression. However, AML1/MTG8 transcripts were detected at all stages of the disease even when the expression level of WT1 gene was low. From these findings, we concluded that monitoring the WT1 expression level is a useful means of determining the effectiveness of chemotherapy, and that WT1 is an effective marker for the detection of MRD, especially in acute myeloid leukemia patients with AML1/MTG8.

A 78-year-old man was diagnosed as leukocytosis in February 1994. Physical examination revealed marked hepatosplenomegaly. A peripheral blood examination disclosed 95,090/microliter leukocytes without hiatus leukemicus, 6.5 g/dl Hb, and 15.0 x 10(4)/microliter platelets. The neutrophil alkaline phosphatase score was 27, and serum VB12 was above 1,600pg/ml. IgG was identified as monoclonal immunoglobulin of type lambda. Bone marrow specimens demonstrated marked granulocytic hyperplasia. Neither the Philadelphia chromosome (Ph1) nor BCR gene rearrangement was detected; hence, the diagnosis of Ph1 (-) chronic myeloid leukemia (CML) was made. The patient was treated with hydroxyurea and low-dose VP-16 with no improvement, and died of pneumonia and sepsis in June 1995. This case was considered to be consistent with atypical CML (aCML) according to the FAB classification because monocytosis was not observed. It seems likely and interesting that the coexistent monoclonal gammopathy and aCML might have arisen from common abnormal hematopoietic stem cells.

A 19-year-old male patient with virus associated hemophagocytic syndrome (VAHS) began receiving chemotherapy including etoposide (cumulative dose of 900 mg/m2 intravenously) and Ara-C (cumulative dose of 360 mg/m2 intravenously) in July 1994. He achieved complete remission, but developed acute myelomonocytic leukemia (AML, FAB M4) with t(9;11)(p22;q23) in March 1997 and a rearrangement of the MLL gene was also recognized. The MLL gene rearrangement is closely associated with secondary leukemia with an 11q23 translocation. It is highly likely that this case of AML was caused by the cytostatic treatment the patient received, including etoposide for VAHS.

Cefpirome (CPR) and amikacin (AMK) were used concomitantly to treat infections complicated by hematological diseases. A total of 100 subjects were evaluated, and the allover efficacy rate was 72.0%. Acute leukemia was found in the largest number of patient, 55, followed by 12 cases of malignant lymphoma and 6 cases of chronic myelogenous leukemia. By type of infection, patients having suspected sepsis were the largest in number, being 50, and the efficacy rate was 68.0%. The efficacy rates for sepsis and pneumonia were 57.1% (7 cases) and 61.1% (18 cases), respectively. The efficacy rates by neutrophil counts before administration of CPR and AMK and at 7 days after administration were both 71.9% in the group of less than 500/microliter, both 60.0% in the group of less than 100/microliter. The efficacy rate was 75.0% in the group of granulocyte colony stimulating factor (G-CSF) concomitant usage, and 70.0% in the non-concomitant usage group. Concomitant treatment with CPR and AMK exhibited a high level of safety and efficacy rates in infections complicated by hematological diseases and high.

A 56-year-old man was admitted to our hospital in September, 1996. Chromosomal translocation (15; 17) and a PT-PCR study for PML-RAR alpha mRNA were positive in bone marrow aspirates, and acute promyelocytic leukemia was diagnosed. After CR was obtained with all-trans retinoic acid (ATRA) followed up with chemotherapy, the RT-PCR became negative. When he was readmitted in April, 1997, skin eruption on his chest and extremities were observed. Specimens taken for biopsy revealed leukemia cutis, and RT-PCR became positive in the same specimen. Bone marrow PT-PCR was also positive without abnormal promyelocytes. Although he was treated with oral ATRA 80 mg/day again, no significant improvement in leukemia cutis was noted. After combined therapy with Ara-C and acularubicin, skin eruption disappeared and bone marrow RT-PCR became negative. A second CR was then obtained. Although it is unknown whether the administration of ATRA is related to extramedullary relapse or not, we recommend combined chemotherapy for such cases.

A 51-year-old man was admitted to our hospital in December 1993, because of fatigue. Peripheral blood tests showed a WBC of 49,400/microliter with 36% plasma cells and 35% monocytes, Hb 14.5 g/dl, and Plt 137,000/microliter. Bone marrow aspirate revealed hypercellularity with 48.7% plasma cells and 22.4% monocytes. Plasma cells in blood were positive for CD38 and PCA-1. Serum calcium, IgA and M-CSF levels were elevated to 14.1 mg/dl, 2,337 mg/dl and 2.7 ng/ml, respectively. Immunoelectrophoresis of serum and urine revealed IgA lambda type M protein and lambda type Bence Jones protein, respectively. Rearrangements of immunoglobulin heavy chain and light chain were demonstrated by Southern blotting analysis. Plasma cell leukemia (IgA lambda type) was diagnosed. He was treated with combination chemotherapy and IFN-alpha and achieved complete remission. However, he suffered a meningeal relapse in February 1995, and died in April 1996. It seems likely that the enhanced production of M-CSF by myeloma cells and/ or activated B cells stimulated monocyte production.

A multicenter randomized controlled study was undertaken in order to determine whether epoetin beta (EPO) ameliorates the anemia in aplastic anemia (AA) patients treated with granulocyte colony-stimulating factor (G-CSF). Enrolled patients were randomized into 3 groups: group C receiving G-CSF alone as the control; group L receiving G-CSF and 200 IU/kg of EPO; group H receiving G-CSF and 400 IU/kg of EPO. Throughout the study, the dose and the administration interval of G-CSF were adjusted to maintain neutrophil counts between 1000 and 5000 microliters EPO was administered subcutaneously for 12 wk as the first step in treatment and when favorable effects were observed over this period, treatment was continued for another 12 wk as the second step in treatment. Significant erythroid responses were defined as increases in untransfused hemoglobin values1.0 g/dl or50% decreases in RBC transfusion requirements over the treatment period. Of 131 patients enrolled, 88 patients allocated to groups L and H were evaluated for toxicity to EPO and 110 were evaluated for erythroid responses. Four of the 31 patients (12.9%) in group C, 6 of the 41 patients (14.6%) of group L, and 14 of the 38 patients (36.8%) of group H showed erythroid responses in the first step in treatment. The erythroid responses of group H were significantly higher than those of the other 2 groups (p0.05). The significant effects of EPO were due to erythroid responses in non-severe AA. Responding patients were significantly different from non-responders with regard to disease severity, hemoglobin concentration, reticulocyte count, serum endogenous erythropoietin levels and serum transferrin receptors; non-severe AA patients were more likely to respond to EPO, and responding patients had lower serum EPO and higher hemoglobin concentration, reticulocyte count and serum transferrin receptors than non-responders. The response rate increased in the second step in treatment, suggesting that long-term treatment improved the efficacy of EPO. No serious side-effects were observed. From these results, we conclude that EPO given in combination with G-CSF is a safe and effective alternative for the treatment of anemia of a subset of AA patients.

A 33-year-old man was admitted to our hospital because of thrombocytopenia found on a periodic physical examination. Splenomegaly was recognized Peripheral blood showed WBC 4,510/microliters, Hb 12.5 g/dl, and Plt 40,000/microliters. Increased serum levels of acid phosphatase and angiotensin converting enzyme were observed on laboratory tests. Bone marrow aspirate revealed Gaucher cells. Decreased beta-glucosidase activity was demonstrated in blood leukocytes, cultured blood lymphocytes, and cultured bone marrow fubroblasts from the patient. His glucocerebrosidase genotype was T1448C/C1504T (L444P/R463C). Since neurological examination and skeletal X ray results were normal, Gaucher disease type 1 was diagnosed.

We studied the antileukemic effect of recombinant human granulocyte colony-stimulating factor (rhG-CSF) by using a radiation-induced murine myeloid leukemia cell line C2M-A5. Intravenous inoculation of C2M-A5 cells into C3H/He mice resulted in the development of myeloid leukemia. However, the leukemic death of the mice was completely suppressed by the subcutaneous injections of rhG-CSF. In order to clarify the mechanism of the suppression, effects of rhG-CSF on C2M-A5 cells were studied in vitro. While C2M-A5 cells grew exponentially in the absence of rhG-CSF, the viability, the growth, and the self-renewal capacity of C2M-A5 cells were all suppressed in cultures in the presence of rhG-CSF. Preincubation with rhG-CSF for 48 h deprived C2M-A5 cells of the ability to induce leukemia in syngeneic mice. Morphological examination revealed the appearance of apoptotic changes of C2M-A5 cells in cultures containing rhG-CSF over the 2-day incubation period. In gel electrophoresis, the DNA from C2M-A5 cells incubated with rhG-CSF for 48 h showed a ladder of degradated DNA bands compatible with apoptosis. From these results, we concluded that the apoptosis of C2M-A5 cells played a key role in the antileukemic effect of rhG-CSF.

A 45-year-old male was hospitalized on September 2, 1989 with chief complaints of general fatigue and fever. Physical examination revealed hepatomegaly and massive splenomegaly. Laboratory tests on admission showed Hb of 7.5g/dl, PLT 4.8 x 10(4)/microliters and WBC 9,610/microliters with 81% hairy cells. Bone marrow aspirate demonstrated 55.1% hairy cells and moderate myelofibrosis. Cytochemically, hairy cells were positive for tartrate-resistant acid phosphatase (TRAP). Surface markers were SmIg G+ A+ kappa +, CD11b+, CD11c+, CD19+, CD20+, CD21-, CD25+, HC2+, HLA-DR+. From these findings, a diagnosis of hairy cell leukemia (HCL) was made. After administration of deoxycoformycin (DCF) at a dose of 5.0mg/m2 1-2 times monthly, splenomegaly disappeared, as did hairy cells from the peripheral blood. Hematological level returned to within normal range except for the presence of 1.2% hairy cells and mild myelofibrosis in bone marrow aspirates. DCF has so far been effective for this patient. While DCF has been reported to be effective in the treatment of HCL in the West, it has not been determined in Japanese patients with HCL, who have different hematologic features from those of HCL patients in the West.

The therapeutic and hematological effects of recombinant human granulocyte colony-stimulating factor (rhG-CSF) in combination with cyclophosphamide (CY) were investigated in a murine myeloid leukemia model. Ten daily administrations of rhG-CSF following CY prolonged the survival time of leukemic mice more than either agent alone. Hematological examination indicated that this effect was attributable to suppression with rhG-CSF of the leukemic repopulation after CY injection. In addition, rhG-CSF accelerated recovery from CY-induced neutropenia. Based on these hematological changes, a treatment regimen was established consisting of a single injection of CY on day 1 and daily injections of rhG-CSF on days 2-6; this combination treatment was given to the leukemic mice for up to four cycles, with a pause of one day between each cycle. The leukemic mice completed each cycle of treatment with few failures, and it resulted in a long survival time for the leukemic mice. The mean survival time of the mice receiving four cycles of treatment was 47 days, 30 days longer than that of the untreated mice. Hematological examination performed at the end of each cycle showed that the leukemic cell population was controlled at a level equal to or below the pre-treatment level, and peripheral blood neutrophils were maintained at a level equal to or above the normal level. These results indicate the possible effectiveness of combining rhG-CSF with chemotherapeutic drugs in controlling leukemic cell growth, and the effectiveness of rhG-CSF in enhancing neutrophil recovery after chemotherapy. However, it was found that the leukemic cells became resistant to treatment with rhG-CSF after four cycles of combination treatment, suggesting that great care should be taken in the clinical application of rhG-CSF, even when the growth of acute myelogenous leukemia cells is not apparently stimulated by it.

Twenty-three patients with bicytopenia or pancytopenia were retrospectively studied. The patients with underlying disorders, blast count of more than 5% on bone marrow (BM) aspirate, blast count of more than 1% on peripheral blood or ringed sideroblast count of more than 15% on BM aspirate were excluded. According to Yoshida's criteria, 23 patients were classified into 6 subtypes [AA (aplastic anemia)1: typical AA, AA2: atypical AA, MDS (myelodysplastic syndrome)3: typical RA (refractory anemia, MDS4-6: atypical RA], and AA1 7 cases; AA2 2 cases; MDS3 5 cases; MDS4 1 case; MDS5 2 cases; MDS6 6 cases. To clarify the clinical features of atypical RA group (MDS4-6), we investigated ferrokinetics, RBC life span, karyotype, serum Epo (erythropoietin) concentration, response to therapy and prognosis. Results were as follows: 1) all three RA patients who were younger than 30 years old were included in atypical RA group, 2) in ferrokinetics study PID (plasma iron disappearance time) values of MDS4 and MDS6 patients ranged between those of AA1 and those of MDS3 patients (5 of 7 patients), 3) two cases who developed leukemia belonged to typical RA group, 4) patients with atypical RA showed response to therapy and their prognosis were better than those with typical RA. These observations suggest that atypical RA have different clinical features from typical RA.

The clonal origin of radiation-induced thymic lymphoma was studied in mice with cellular mosaicism for phosphoglycerate kinase (PGK). Repeated whole-body X-irradiations (4 doses, 1.7 Gy each) with intervals of 7 days resulted in development of thymic lymphomas in the mosaic mice. PGK from all lymphomas gave only a single spot on electrophoresis. The results demonstrate the single cell origin of the thymic lymphoma.

Dialyzed urine of mice bearing leukocytosis-inducing fibrosarcoma stimulated granulocyte colony formation in semisolid agar culture of human bone marrow cells. Removal of phagocytic cells prior to stimulation did not interfere with the formation of these colonies in the culture. On the other hand, macrophage colonies were predominantly produced when murine bone marrow cells were stimulated by the dialyzed mouse urine. The activity of colony-stimulating factor (CSF) in the urine of normal mice was less than 1/100 of that in the urine of tumor-bearing mice. DEAE-cellulose column chromatography separated the activity stimulating human granulocyte colony formation from that stimulating murine macrophage colony formation. Further purification showed that a sialoglycoprotein with an apparent molecular weight of 80,000 corresponded to the macrophage CSF, which was devoid of activity toward human cells. The molecular properties of the human-active granulocyte CSF could not be studied further, because it was quite unstable.