A phase II study of epirubicin in acute leukemia: a cooperative group study

A new doxorubicin analogue, epirubicin (EPI), was evaluated in 41 patients with acute leukemia at 11 Japanese institutions participating in a phase II study between January 1983 and July 1985; during this period 35 patients were considered evaluable. There were 25 males and 10 females with a median age of 43 years (range, 19-71 years) and the median PS of 2 (range, 0-4). EPI was given to 25 patients who had previously been treated with intensive combination chemotherapy, of whom 22 had already received anthracyclines. Ten patients had not been treated previously. Two dose schedules were explored. The higher dose schedule (18 cases) consisted of the administration of 24 to 60 mg/m2 for 3 to 5 consecutive days, and the lower dose schedule (17 cases) consisted of 11 to 20 mg/m2 for 5 to 7 days. Remissions were obtained in 7 patients (20%), 2 of whom showed complete remission and 5 partial remission. The remission duration was 2, 2, 3, 5, 16, 16 and 29+ weeks, respectively. The expected myelosuppression was universal. Stomatitis occurred in 15 patients, of which 7 cases were severe. This stomatitis occurred frequently in the higher dose schedule, and was thought to be a dose-limiting factor. In others, alopecia, G.I. symptoms, and diarrhoea (4 patients) were seen. These results from a cooperative group study indicated that EPI was an effective drug for the treatment of acute leukemia.     

Histologic typing of non-Hodgkin's lymphomas by in situ hybridization with DNA probes of oncogenes

Expression of six proto-oncogenes (fos, myc, myb, Ki-ras, Ha-ras, and N- ras) in 43 cases of non-Hodgkin's lymphoma was analyzed by means of in situ hybridization. Biotinylated DNA probes of the six oncogenes and those of immunoglobulin H (IgH) gene and T-cell receptor beta (TCR beta) chain gene were used. The results of in situ hybridization performed under blind conditions by IgH gene and TCR beta chain gene probes were compatible with those of typing by cell surface markers. The nuclear protein-related proto-oncogenes, fos, myc, and myb, were expressed in about 70% to 80% of all cases regardless of phenotype, histology, or histologic grade. On the contrary, genes of ras family were expressed in more limited numbers of cases except for the Ki-ras gene, which was more frequently expressed by cases of the T-cell immunophenotype with a high malignancy grade. The results of dot hybridization with RNA extracted from some cases were compatible with those of in situ hybridization, further demonstrating the specificity of in situ hybridization.     

Analyses of T-cell differentiation from hemopoietic stem cells in the G0 phase by an in vitro method.

Using differential radiation sensitivity of components of mouse embryonal thymus, an in vitro method for studying T-cell differentiation from hemopoietic stem cells (HSCs) in the G0 phase was established. Intrathymic T-cell precursors present in embryonal thymus were found to be quite radioresistant (up to 20 Gy), and consequently 25-Gy-irradiated embryonal thymic lobes were used. Thymic lobes (25-Gy irradiated) taken from mouse fetuses (gestation day 15) were placed in Millipore-HA culture plates supported on squares of gelatin foam sponge in 24-well culture plates in which neonatal thymus stromal cells were cultured. HSCs (10(5) cells per well) in the G0 phase were added to these thymic lobes and cocultured at 37 degrees C in a 5% CO2/95% air incubator. Half the culture medium was changed every week. After 3 weeks, a large number of colonies had formed. Immunohistochemical studies and fluorescence-activated cell sorter analyses revealed that the colonizing cells regularly develop and exhibit surface markers characteristic of T cells (Thy-1, IL-2R, L3T4, Lyt-2, etc.). In situ hybridization analyses revealed that mRNA expression for T-cell receptor (TCR) beta chains occurred within colonizing cells. Using a monoclonal antibody (F23.1), expression of TCR beta-chain variable domain (V beta 8) on the surface of these developing T cells was demonstrated. These cells responded to interleukin 2 and/or anti-CD3 monoclonal antibody, indicating functional T cells. This method will be useful in studying T-cell differentiation pathways from pluripotent HSCs and in clarifying the mechanisms involved in negative and positive selection of T cells within the thymus.     

(8S)-(glutathion-S-yl)dihydromorphinone, a novel metabolite of morphine from guinea pig bile

A novel glutathione-conjugated metabolite of morphine has been isolated from the bile of guinea pigs given morphine. The metabolite was separated by preparative HPLC on a reverse phase column (YMC-GEL C18) using methanol/water (1:4, v/v) as eluate and purified by HPLC on another reverse phase column (mu-Bondapak phenyl) using water/acetonitrile/trimethylamine/acetic acid (150:3:2:1, v/v) as a mobile phase. The unambiguous structure assignment of the metabolite was performed by fast atom bombardment mass spectrometry and 400 MHz fourier transform NMR spectrometric analysis, and it was identified as (8S)-glutathion-S-yl)dihydromorphinone, in comparison with the synthetic morphinone-glutathione adduct.     

Adriamycin-Lipiodol suspension for i. a. chemotherapy of hepatocellular carcinoma

Summary Physicochemical properties of two types of adriamycin preparation, suspensions and emulsions prepared for i.a. chemotherapy of hepatocellular cacinoma, were investigated. A suspension was prepared by dispersing adriamycin directly into the lipid contrast medium, Lipiodol, whereas an emulsion was obtained by emulsifying an aqueous solution of adriamycin into Lipiodol. The dispersibility of the drug in each preparation was examined microscopically. The chemical stability of and drug release from the preparation were determined by high-performance liquid chromatography and spectrophotometry, respectively. The suspension was then given to ten patients with primary hepatocellular carcinoma. The suspension maintained good dispersibility without coagulation of drug particles, whereas coalescence of aqueous droplets and the resultant phase separation occurred 4 h after preparation of the emulsion. Both preparations maintained the initial drug content for at least 1 week at room temperature. The release of adriamycin was more prolonged in the suspension than in the emulsion. After i.a. administration of the suspension, a selective accumulation of Lipiodol in the tumor and decrease in serum -fetoprotein (AFP) levels were found in most patients. A significant amount of adriamycin was still detected in hepatic speciments resected from two patients 1 and 2 months after treatment. These findings suggest that the adriamycin-Lipiodol suspension may be a useful preparation for targeting chemotherapy to hepatocellular carcinoma.     

Immature dendritic cells (CD11c+ CD3- B220- cells) present in mouse peripheral blood

It is well known that dendritic cells (DCs) are developed from the peripheral blood of mice when peripheral blood mononuclear cells (PBMCs) are cultured with GM-CSF. We have previously found that immature DCs are present in the blood even in humans. In the present study, we show that CD11c+ CD3- B220- cells in the mouse peripheral blood are immature DCs. The percentage of CD11c+ CD3- B220- cells in the (PBMCs) of normal mice ranges from 0.5 to 2.5%. The CD11c+ CD3- B220- cells in the PBMCs show dendrites, similar in shape to the CD11c+ CD3- B220- cells in the spleen, which are thought to be DCs definitely. However, they have practically no capacity to stimulate the proliferation of allogeneic T cells, and show a lower expression of MHC class II, B7-1 and B7-2 than CD11c+ CD3- B220- cells in the spleen. When the CD11c+ CD3- B220- cells in the PBMCs are cultured with GM-CSF, they show not only the potent ability to stimulate the proliferation of allogeneic T cells but also a higher expression of MHC class II, B7-1 and B7-2. Moreover, they migrate into the spleen when they are injected intravenously. These results suggest that CD11c+ CD3- B220- cells in the PBMCs are immature DCs, and that they migrate into the spleen, where they mature.     

Kimura's disease with unusual eosinophilic epithelioid granulomatous reaction: a finding possibly related to eosinophil apoptosis

We report and discuss a case of Kimura's disease with an unusual eosinophilic epithelioid granulomatous reaction. A 3-year-old Japanese boy with eosinophilia and a high concentration of IgE developed lymphadenopathy and multiple cervical masses. A lymph node biopsy demonstrated the infiltration of eosinophils in the stroma, which is consistent with the findings of Kimura's disease. Interestingly, a number of apoptotic eosinophils was detected in the infiltrating eosinophils. Multiple epithelioid granulomas with central eosinophilic abscesses and necrosis were also observed. Macrophages and giant cells had phagocytosed the apoptotic eosinophils at the edge of the granulomas. In situ terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling (TUNEL) assay showed that the TUNEL-positive eosinophils were both in the macrophages and in the central eosinophilic abscesses of the granulomas. These findings suggest that the eosinophils had undergone an accelerated apoptosis in this case of Kimura's disease, and that the epithelioid granulomas were produced by phagocytosis of the apoptotic eosinophils by macrophages.     

Three novel mutations of the fibrillin-1 gene and ten single nucleotide polymorphisms of the fibrillin-3 gene in Marfan syndrome patients

Marfan syndrome (MFS) is an autosomal dominant disorder of the extracellular matrix. Allelic variations in the gene for fibrillin-1 (FBN1) have been shown to cause MFS. To date, over 550 mutations have been identified in patients with MFS and related connective tissue diseases. However, about a half of MFS cases do not possess mutations in the FBN1 gene. These findings raise the possibility that variants located in other genes cause or modify MFS. To explore this possibility, firstly we analyzed FBN1 allelic variants in 12 Japanese patients with MFS, and secondly we analyzed fibrillin-3 gene (FBN3) in patients without FBN1 mutations using conformation sensitive gel electrophoresis (CSGE) and direct sequencing analysis. We identified three novel FBN1 mutations and ten FBN3 single nucleotide polymorphisms (SNPs). In this report, we could not detect a responsible mutation of the FBN3 gene for MFS. Although the number of the cases in this report is small, at least these results suggest that disease-causing mutations in exon regions of the FBN3 gene are very rare in MFS.Nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank databases under the accession numbers: AB177797, AB177798, AB177799, AB177800, AB177801, AB177802, AB177803     

Prevention of endotoxin shock by an antibody against leukocyte integrin {beta}2 through inhibiting production and action of TNF

Septic shock remains a serious disorder associated with highmortality. Accumulating evidence indicates that TNF is a majorand essential mediator of endotoxin shock. We report here thatadministration of an antibody against CD18 dramatically reducedendotoxin-induced shock inrabbits as revealed by preventionof severe hypotension, metabolic acidosis and a pathologicalchange suggestive of disseminated intravascular coagulationwith concomitant inhibition of elevation of plasma TNF activity.The anti-CD18 antibody also inhibited the hypotension inducedby administering recombinant TNF. Furthermore, an antibody againsta ligand for CD18 complexes, intercellular adhesion molecule-1,also prevented TNF-induced shock as well as endotoxin shockinrabbits. These observations suggest that adhesion of leukocytesto endothelium may be of primary importance in the action ofTNF as well as in the production of TNF in vivo and that theantibody against adhesion molecules could be of therapeuticbenefit in life-threatening septic shock in humans.