Synthesis, conformation and antiproliferative activity of isothiazoloisoxazole 1,1-dioxides.

Sixteen new isothiazoloisoxazole 1,1-dioxides, one new isothiazolotriazole and one new isothiazolopyrazole have been synthesised by using 1,3-dipolar cycloadditions to isothiazole 1,1-dioxides. One sub-set of these isothiazoloisoxazoles showed low µM activity against a human breast carcinoma cell line, whilst a second sub-set plus the isothiazolotriazole demonstrated an interesting restricted rotation of sterically hindered bridgehead substituents. A thiazete 1,1-dioxide produced from one of the isothiazole 1,1-dioxides underwent conversion into an unknown 1,2,3-oxathiazolin-2-oxide upon treatment with Lewis acids, but was inert towards 1,3-dipoles and cyclopropenones. Six supporting crystal structures are included.

A clinical trial of retroviral-mediated transfer of a rev-responsive element decoy gene into CD34(+) cells from the bone marrow of human immunodeficiency virus-1-infected children.

Genetic modification of hematopoietic stem cells with genes that inhibit replication of human immunodeficiency virus-1 (HIV-1) could lead to development of T lymphocytes and monocytic cells resistant to HIV-1 infection after transplantation. We performed a clinical trial to evaluate the safety and feasibility of this procedure, using bone marrow from four HIV-1-infected pediatric subjects (ages 8 to 17 years). We obtained bone marrow, isolated CD34(+) cells, performed in vitro transduction with a retroviral vector carrying a rev-responsive element (RRE) decoy gene, and reinfused the cells into these subjects with no evidence of adverse effects. The levels of gene-containing leukocytes in peripheral blood samples in the 1 year after gene transfer/cell infusion have been extremely low. These observations support the potential of performing gene therapy for HIV-1 using hematopoietic cells, but emphasize the need for improved gene transfer techniques.

Increased gene transfer into human CD34+ progenitor cells using retroviral vectors produced by a canine packaging cell line.

Using retroviral supernatants derived from the amphotropic murine packaging cell line PA317 and the amphotropic canine packaging cell line (DA), cord blood and mobilized peripheral blood CD34+ cells were transduced with the vector LN (neomycin resistance) and the vector L-TR/TAT neo (neomycin resistance in conjunction with a double-hammerhead ribozyme conferring anti-HIV activity). Different multiplicities of infection (MOI) were applied in the setup according to vector titrations on NIH-3T3 cells. PA317-based supernatants were tested at MOI of 10 and 30. Purified concentrated DA-derived vector preparations were tested at MOI of 10, 30, 100, and 300. Immediately after transduction, CD34+ cells were plated into colony assays in the presence and absence of G418 to evaluate the amount of gene transfer and potential toxic effects of the vectors on colony growth. The remaining cells were subjected to G418 selection in liquid culture for 12 days and subsequently challenged with HIV-1JR-FL to test for efficacy of the anti-HIV gene in macrophages derived from transduced CD34+ cells. Transduction by the PA317-packaged vectors was maximal at the lowest MOI used and did not increase with increasing MOI. In contrast, transduction by the DA-packaged vectors could be progressively increased using increased MOI. The net transduction efficiency per unit of reverse transcriptase activity in the DA vector preparations was 8.7-fold higher than in the PA317 vector supernatants. HIV-1 challenge of the cells transduced by the ribozyme vector derived from the PA317 packaging cells resulted in a 1.5 log inhibition of p24 output compared with the control cells containing neomycin resistance only. A 2.5 log inhibition of p24 output could be observed in the cell population transduced with DA-packaged vector supernatants. Compared with retroviral supernatants from PA317 packaging cell lines, DA packaging line-derived vector preparations demonstrated higher transduction efficiency into CD34+ cells, particularly at higher MOI, and increased efficacy of the transferred anti-HIV gene when challenged with HIV-1JR-FL. The increase in transduction efficiency may be due to a higher ratio of intact vs. defective vector particles in the DA-derived vector preparations.

Efficacy and compliance with cholestyramine bar versus powder in the treatment of hyperlipidemia.

PURPOSE: The purpose of the study was to compare the powder and the bar forms of cholestyramine to determine efficacy and patient compliance. SUBJECTS AND METHODS: A prospective, randomized trial was conducted that included 83 healthy men and women with hyperlipidemia greater than the 90th percentile for low-density lipoprotein (LDL) or total cholesterol. Patients were randomly assigned to receive either cholestyramine powder, two packets (8 g), twice daily, or cholestyramine confectionery bar, in maple or mint flavors, two bars (8 g), twice daily. Fasting serum total cholesterol, LDL cholesterol, high-density lipoprotein (HDL) cholesterol, and triglycerides were measured at baseline, after 6 to 8 weeks of following the American Heart Association Step I diet alone, and after 8 weeks of taking either the cholestyramine bar or powder. RESULTS: Total cholesterol decreased significantly (p less than 0.01) by 16% in the bar group and 17% in the powder group. LDL cholesterol decreased by 28% and 29% in the bar and powder groups, respectively (p less than 0.01). There was no significant change in HDL cholesterol. Triglycerides increased in both groups, by 29% in the bar group and by 25% in the powder group. There was no difference between bar and powder in the effect on blood lipids. The majority of the lipid-lowering effect was seen within 14 days. Mean patient endpoint compliance with the therapy was 91.8 +/- 3.6% in the bar group and 94.8 +/- 2.1% in the powder group. There was no difference between groups. CONCLUSION: The cholestyramine confectionery bar is as effective as cholestyramine powder in the treatment of hyperlipidemia. The majority of the lipid-lowering effect is seen within 14 days of therapy. Although patient compliance is comparable between the two forms, gastrointestinal side effects were slightly greater with the bar form. Therefore, although the bar offers an alternative form of therapy, there appears to be no advantage with regard to patient compliance or palatability.