Cytotoxicity of novel sulfanilamides towards sensitive and multidrug-resistant leukemia cells.

Novel sulfa Schiff bases were synthesized and characterized by a reaction between aromatic sulfonamides and aromatic aldehydes or heterocyclic ketones in equimolar ratios. Their cytotoxicity was evaluated by the resazurin assay towards human sensitive CCRF-CEM and multidrug-resistant CEM/ADR5000 leukemia cells. Three of the tested compounds viz., 4-(anthracen-9-ylmethyleneamino)-N-(pyrimidin-2-yl)benzenesulfonamide (4), 4-(anthracen-9- ylmethyleneamino)benzenesulfonamide, (5) and 4-((3-phenylallylidene)amino)benzene-sulfonamide, (6) were cytotoxic (IC50 values: 5.38-19.96 µM). CEM/ADR5000 cells were not cross-resistant to these compounds, indicating activity against otherwise drug-resistant tumors. Compound 6 inhibited P-glycoprotein by increasing doxorubicin accumulation and reducing expression of P-glycoprotein in CEM/ADR5000 cells. A human P-glycoprotein homology model was used for molecular docking studies. Compound 6 and verapamil (a well-known P-glycoprotein inhibitor) docked with similar binding energies to the same binding pocket.

Direct recovery of herpes simplex virus (HSV)-specific T lymphocyte clones from recurrent genital HSV-2 lesions.

T lymphocytes were recovered from recurrent vesicular and ulcerative herpes simplex virus type 2 (HSV-2) lesions from 4 patients and cloned without exogenous secondary antigenic stimulation. Resultant cultures were screened for antigen-specific proliferative and cytotoxic responses. Of the 39 HSV-specific clones recovered, all were CD3-positive; 38 were CD4-positive and 1 was CD8-positive. Of the T cell clones recovered directly from lesions, 6%-10% had HSV-specific proliferative responses, in contrast to a peripheral blood precursor frequency of HSV-specific CD4 cells of approximately 1/10(3) to 1/10(4) as measured by limiting dilution assays. CD4+ clones studied restricted by HLA-DR, -DP, and -DQ were detected and both type-common and type-specific CD4+ T cell clones were recovered. HSV-specific T cells localize to recurrent human genital HSV-2 lesions and can be obtained directly from early lesions. The ability to isolate HSV-specific T cell clones from lesions allows the study of antigenic specificities and functional properties of tissue-resident antigen-specific T lymphocytes.

Herpes simplex virus infection of human fibroblasts and keratinocytes inhibits recognition by cloned CD8+ cytotoxic T lymphocytes.

CD8+ cytotoxic T lymphocytes (CTL) clones with specificity for herpes simplex virus (HSV) were derived from two donors with genital HSV-2 infection. These CTL clones specifically lysed HSV-infected autologous B lymphoblastoid cells, but not HSV-infected fibroblasts. Exogenous peptide loading sensitized both cell types to lysis by an HSV-specific CTL clone of known specificity. HSV infection rendered fibroblasts refractory to peptide sensitization. HSV infection also rendered fibroblasts and keratinocytes insensitive to lysis by allospecific CD8+ CTL clones. Lysis of B lymphoblastoid cells in this system was only slightly reduced by HSV infection. Reduction of fibroblast allospecific lysis was dose and time dependent and was blocked by acyclovir, indicating the involvement of a late HSV gene product. HSV caused a reduction of fibroblast cell surface HLA class I antigen, at least in part due to reduction of synthesis of heavy chain-beta 2 microglobulin heterodimers. These results suggest that HSV-induced blockade of antigen presentation by cutaneous cells to CD8+ CTL may be a mechanism by which HSV limits or evades the immune response of the host.

Comparison of monoclonal antibodies for rapid detection of cytomegalovirus in spin-amplified plate cultures.

Cytomegalovirus (CMV) was detected in 56 of 275 specimens (20%); 50 of 56 (89%) were detected by conventional culture, and 37 (66%) were detected by rapid assay at 72 h with a commercial monoclonal antibody and a pooled monoclonal antibody. Although the two antibodies were equally sensitive at 72 h, the pooled antibody gave a brighter, more easily detected signal. Other viruses were isolated from 9 specimens (3.3%) by conventional culture. Use of rapid assays alone fails to detect slow-growing CMV and non-CMV viral pathogens.

Chronic-dose acyclovir to suppress frequently recurring genital herpes simplex virus infection: effect on antibody response to herpes simplex virus type 2 proteins.

To determine the effect of prolonged suppressive acyclovir therapy on the antibody response to herpes simplex virus type 2 (HSV-2) proteins, we studied sequential sera from 33 patients with frequently recurring (six or more recurrences per year) genital herpes. Twenty-two patients received 400 mg of oral acyclovir and 11 received placebo, twice daily for one year. Sera collected at enrollment, after six months and 12 months of therapy, and during the first recurrence after cessation of therapy were evaluated by western blot for levels of antibodies to HSV-2, gB, gG, gC/gE, VP16, and gD. Mean levels declined by 27%-39% after one year of acyclovir. The magnitude of the decrease in antibody levels was not correlated with disease severity either during or after therapy. Patients with high relative antibody levels to gB after therapy had more-severe first recurrences after therapy than did patients with antibody levels to gB less than or equal to the median. Antibody levels were not restored after the first untreated recurrence.

Analysis of a major target of the human immune response to cytomegalovirus using monoclonal antibodies.

Two murine monoclonal antibodies (C6D1 and D2B1) were found to react with a set of cytomegalovirus (CMV)-infected cell polypeptides, which comprise a major target of the human immune response to CMV. C6D1 reacted with proteins of 50 kilodaltons (KD) and 40KD molecular weight; D2B1 reacted with these two proteins plus a third of 35KD. Western blot analysis demonstrated that these protein targets also react with serum antibody from patients with acute or latent CMV infection. Immunofluorescence staining of CMV-infected diploid fibroblast cells by C6D1 and D2B1 showed that the protein targets are found in the nucleus throughout the course of viral infection. The proteins were shown to be late proteins dependent on viral DNA synthesis for their expression. Not all wild-type CMV strains tested expressed proteins that react with C6D1 and D2B1. Using an immunofluorescence stain of diploid fibroblasts infected with CMV strains from infected patients, we found that 70 of 76 (92%) wild-type strains reacted with C6D1 and 23 of 24 (96%) with D2B1. One strain was not reactive with either C6D1 or D2B1. Western blot analysis of 11 wild-type strains revealed that two isolates either lack the C6D1 and D2B1 protein targets or have forms of these proteins that migrate at different molecular weights.

Antigenic and biochemical analysis of field isolates of influenza B virus: evidence for intra- and inter-epidemic variation.

Detailed antigenic analysis using a panel of monoclonal antibodies was carried out on the haemagglutinin antigen of 53 influenza B viruses isolated from an epidemic in a single school. Thirteen distinguishable antigenic groupings of influenza B viruses could be detected but 26 of the viruses were in two groups (III and IV) which co-existed during the entire epidemic. Antigenically distinguishable influenza B viruses were isolated from an epidemic in a second nearby school. Influenza B viruses isolated from the two schools could be further distinguished by different electrophoretic mobilities of NS1 polypeptides and of genes 1, 2, 3 and 6, whereas viruses from a single school epidemic were very closely related as regards these biochemical characteristics. The findings are consistent with the hypothesis that the outbreak was initiated by a single individual who excreted antigenic mutants of which predominantly two spread and co-existed during the epidemic, although the additional occurrence of random mutations during the evolution of the epidemic cannot be excluded.