Significant antitumoral activity of cationic multilamellar liposomes containing human IFN-beta gene against human renal cell carcinoma.

PURPOSE: Immunotherapy is the most effective treatment against metastatic renal cell carcinoma (RCC). However, the response rate is approximately 15%. More effective therapy is, therefore, needed for patients with metastatic RCC. We then examined the antitumor effect of cationic multilamellar liposome containing human IFN-beta (huIFN-beta) gene (IAB-1) against RCC. EXPERIMENTAL DESIGN: Concentrations of huIFN-beta protein were measured by ELISA. The cytotoxicity of IAB-1 against human RCC (NC65, ACHN, and freshly isolated RCC cells), prostate and bladder cancer cell lines, and renal proximal tubule endothelial cells (RPTEC5899) was examined by the colorimetric method using tetrazolium salt. Apoptosis was assessed by the acridine-orange staining. For in vivo study, we used NC65 cells inoculated into severe combined immunodeficiency mouse. RESULTS: The RCC cells treated with IAB-1 secreted significant amounts of huIFN-beta protein continuously. Drastic in vitro cytotoxic effect of IAB-1 against RCC was observed. In contrast, treatment with 1000 IU/ml recombinant huIFN-beta protein resulted in weak cytotoxicity. The cytotoxic effect against prostate and bladder cancer cell lines was less than that against RCC. Furthermore, no significant cytotoxicity was observed in RPTEC5899 cells. Apoptosis was observed in the cells treated with IAB-1, but recombinant huIFN-beta failed to induce apoptosis. The size of NC65 tumors transfected with IAB-1 in mice was significantly smaller than that receiving injection of empty liposome or recombinant huIFN-beta protein. CONCLUSION: These findings indicate that IAB-1 may have an antitumor activity against human RCC by inducing apoptosis, suggesting its potential clinical application for gene therapy against RCC.     

Alteration of p53 Clonality Accompanying Colorectal Cancer Progression

The aim of this study was to clarify whether or not the status of gene alteration is heterogeneous in intramucosal carcinoma and homogeneous within invasive carcinoma. We selected 10 colorectal carcinoma cases (1 mucosal, 5 submucosal and 4 advanced carcinomas including 2 cases with lymph node metastasis) and analyzed the p53 gene sequence. Six colorectal cancers in this study showed heterogeneity in p53 mutations in cells from the intramucosal part. In the invasive part of a carcinoma, p53 mutation status was homogeneous intratumorally in all cases. These data indicate that, in regard to p53 gene alterations, colorectal cancers can be composed of various subclones when limited to the mucosa, but clonal selection occurs when one of these subclones commences invasion to the submucosa, generating a monoclonal invasive carcinoma.     

Carcinoma of the cervix: Value of dynamic magnetic resonance imaging in assessing early stromal invasion

Background The purpose of this study was to assess the accuracy of contrast-enhanced magnetic resonance imaging (dynamic MR imaging) in the evaluation of preinvasive and early invasive cancer of the cervix. Methods Twenty-nine women with untreated squamous cell carcinoma of the cervix with either no stromal invasion or early stromal invasion underwent pretreatment MR imaging and dynamic MR imaging within 4 weeks of surgical evaluation. The images were evaluated for tumor detection and compared with results of histologic examination of the surgical specimens. Results The lesions in 17 cases with histologically proven stromal invasion of 4 mm or greater were detected with dynamic MR imaging, whereas lesions in only 8 of these cases were detected with T2 imaging. In 9 cases with stromal invasion between 4.0 mm and 5.0 mm, lesions were represented as early phase focal enhancement on dynamic MR images, but not detected on T2-weighted images. In the 12 cases with less than 4 mm stromal invasion, no lesions were visualized on either T2-weighted images or dynamic MR images, except in 1 case of glandular involvement without stromal invasion that appeared as enhancement on early-phase dynamic MR imaging. Conclusion Dynamic MR imaging detected more lesions of early stromal invasion in pretreatment imaging for cervical cancer than nonenhanced MR imaging.     

Fetal alcohol spectrum disorders (FASD) among Japanese children of alcoholic mothers]

OBJECTIVE: This study was carried out to examine fetal alcohol spectrum disorders (FASD) among Japanese children of alcoholic mothers. This is the first report concerning FASD in Japan. METHODS: The subjects were 30 alcoholic women who were inpatients in the Kurihama Alcoholism Center and had given birth to children. They were subjected to a semi-structured interview by the author. Sixty healthy women who had not drunk during pregnancy were used as a control group, and they also underwent semi-structured interviews. The alcoholic women were divided into two groups, 13 who drunk during pregnancies and 17 who did not drink. Twenty children experienced of prenatal alcohol exposure and 40 children did not. The three groups; i.e., 13 alcoholic mothers who had drunk during pregnancy and their 20 children (ALD group), 17 alcoholic mothers who had not drunk during pregnancy and their 40 children (ALND group) and 60 non-alcoholic control mothers and their 80 children (Control group), were compared concerning the mothers' drinking problems and abnormal deliveries, children's birth weights, congenital abnormalities, abnormalities of the central nervous system and psychological problems. RESULTS: The mean age of onset of problem drinking of the mothers in the ALD group was significantly lower than that in the mothers of the ALND group, and some of the mothers in the ALD group showed alcohol dependence before their pregnancies. The mean birth weights of the children of the ALD group, ALND group and Control group were 2816 g, 3128 g and 3142 g, respectively and the differences were significant. The children of the ALD group had significantly more abnormal birth episodes, developmental retardation and psychiatric symptoms than those in the other two groups. Among 20 children in the ALD group, FASD was suspected in 6 children (10% of the children of alcoholic mothers). Six children had low birth weights, abnormal birth episodes, mental retardation and psychiatric symptoms. CONCLUSION: One third of the Japanese children of alcoholic mothers had experiences of prenatal alcohol exposure and 10% of them had suspected FASD abnormalities.     

Time-dependent inhibition and tetrahydrobiopterin depletion of endothelial nitric-oxide synthase caused by cigarettes.

Smoking causes a dysfunction in endothelial nitric-oxide synthase (eNOS), which is ameliorated, in part, by administration of tetrahydrobiopterin (BH(4)). The exact mechanism by which the nitric oxide deficit occurs is unknown. We have previously shown that aqueous extracts of chemicals in cigarettes (CE) cause the suicide inactivation of neuronal NO synthase (nNOS) by interacting at the substrate-binding site. In the current study, we have found that CE directly inactivates eNOS by a process that is not affected by the natural substrate l-arginine and is distinct from the mechanism of inactivation of nNOS. We discovered that CE causes a time-, concentration-, and NADPH-dependent inactivation of eNOS in an in vitro system containing the purified enzyme, indicating a metabolic component to the inactivation. The CE-treated eNOS but not nNOS was nearly fully reactivated upon incubation with excess BH(4), suggesting that BH(4) depletion is a potential mechanism of inactivation. Moreover, in the presence of CE, eNOS catalyzed the oxidation of BH(4) to dihydrobiopterin and biopterin by a process attenuated by high concentrations of superoxide dismutase but not catalase. We speculate that a redox active component in CE, perhaps a quinone compound, causes oxidative uncoupling of eNOS to form superoxide, which in turn oxidizes BH(4). The discovery of a direct inactivation of eNOS by a compound(s) present in tobacco provides a basis not only for further study of the mechanisms responsible for the biological effects of tobacco but also a search for a potentially novel inactivator of eNOS.     

Metabolism of the alpha,beta-unsaturated ketones, chalcone and trans-4-phenyl-3-buten-2-one, by rat liver microsomes and estrogenic activity of the metabolites.

When chalcone and trans-4-phenyl-3-buten-2-one (PBO) were incubated with liver microsomes of untreated rats in the presence of NADPH, 4-hydroxychalcone and trans-4-(4-hydroxyphenyl)-3-buten-2-one (4-OH-PBO), respectively, were formed as major metabolites. Two minor metabolites of chalcone, 4'-hydroxychalcone and 2-hydroxychalcone, were also observed. The oxidase activity affording 4-hydroxychalcone was inhibited by SKF 525-A, disulfiram, ketoconazole, and alpha-naphthoflavone. The oxidase activities leading to 4-hydroxychalcone and 4'-hydroxychalcone were enhanced in liver microsomes of 3-methylcholanthrene- and phenobarbital-treated rats, respectively. The activity generating 2-hydroxychalcone was enhanced in liver microsomes of 3-methylcholanthrene- and dexamethasone-treated rats. The oxidation of PBO to 4-OH-PBO was inhibited by SKF 525-A, ketoconazole, disulfiram, and sulfaphenazole. This activity was enhanced in liver microsomes of 3-methylcholanthrene-, acetone- and phenobarbital-treated rats. 4-Hydroxylation, 4'-hydroxylation, and 2-hydroxylation of chalcone were catalyzed by rat recombinant cytochrome P450 1A1, 1A2, and 2C6; by 1A1 and 2C6; and by 1A1 and 3A1, respectively. PBO was oxidized by cytochrome P450 1A1, 1A2, 2C6, and 2E1. Chalcone and PBO were negative in an estrogen reporter assay using estrogen-responsive human breast cancer cell line MCF-7. However, 4-hydroxychalcone, 2-hydroxychalcone, 4'-hydroxychalcone, and 4-OH-PBO exhibited estrogenic activity.     

Dendritic cells fused with allogeneic colorectal cancer cell line present multiple colorectal cancer-specific antigens and induce antitumor immunity against autologous tumor cells.

The aim of antitumor immunotherapy is to induce CTL responses against autologous tumors. Previous work has shown that fusion of human dendritic cells and autologous tumor cells induce CTL responses against autologous tumor cells in vitro. However, in the clinical setting of patients with colorectal carcinoma, a major difficulty is the preparation of sufficient amounts of autologous tumor cells. In the present study, autologous dendritic cells from patients with colorectal carcinoma were fused to allogeneic colorectal tumor cell line, COLM-6 (HLA-A2(-)/HLA-24(-)), carcinoembryonic antigen (CEA)(+), and MUC1(+) as an alternative strategy to deliver shared colorectal carcinoma antigens to dendritic cells. Stimulation of autologous T cells by the fusion cells generated with autologous dendritic cells (HLA-A2(+) and/or HLA-A24(+)) and allogeneic COLM-6 resulted in MHC class I- and MHC class II-restricted proliferation of CD4(+) and CD8(+) T cells, high levels of IFN-gamma production in both CD4(+) and CD8(+) T cells, and the simultaneous induction of CEA- and MUC1-specific CTL responses restricted by HLA-A2 and/or HLA-A24. Finally, CTL induced by dendritic cell/allogeneic COLM-6 fusion cells were able to kill autologous colorectal carcinoma by HLA-A2- and/or HLA-A24-restricted mechanisms. The demonstration of CTL activity against shared tumor-associated antigens using an allogeneic tumor cell line, COLM-6, provides that the presence of alloantigens does not prevent the development of CTL with activity against autologous colorectal carcinoma cells. The fusion of allogeneic colorectal carcinoma cell line and autologous dendritic cells could have potential applicability to the field of antitumor immunotherapy through the cross-priming against shared tumor antigens and provides a platform for adoptive immunotherapy.     

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Homepages are redesigned or modified very frequently; therefore,please note that comments in this section are based on the contentsof the homepage at the time of writing. View larger version (64K): [in this window] [in a new window]     ISCT: The Professional Organization for those Working in Cell-basedResearch and Therapy (http://www.celltherapy.org/) Established in 1992, International Society for Cellular Therapy(ISCT), formerly International Society for Hematotherapy andGraft Engineering (ISHAGE),