Biomedical subjects

B W Day

Publications and source records attributed to B W Day.

58 records · Page 4Linked to original sources

PubMedDate not supplied

Enantiospecificity of covalent adduct formation by benzo[a]pyrene anti-diol epoxide with human serum albumin.

Human serum albumin was reacted with the (+)- and (-)-enantiomers of r-7,t-8-dihydroxy-t-9,t-10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene to determine if the chiral nature of the protein influences adduct formation. The alkylated proteins were analyzed directly by fluorescence line narrowing spectroscopy, and their spectra were compared to those of the model synthetic adducts N tau-(7,8,9-trihydroxy-r-7,t-8,t-9,c-10-tetrahydrobenzo[a]pyren-10- yl)histidine and 7,8,9-trihydroxy-r-7,t-8,t-9,c-10-tetrahydrobenzo[a]pyren- 10-yl N-t-BOC-alaninate ester. The results from these analyses indicated that different adducts were formed by the enantiomers of the diol epoxide. The adducted proteins were also enzymatically digested, and the 8,9-cis-dihydroxy-7,8,9,10-tetrahydrobenzo[a]pyrene-containing adducts and hydrolysis products were isolated by boronate affinity chromatography. Diode array UV, fast atom bombardment, and on-line atmospheric pressure ionization-mass spectral analysis of the HPLC purified products indicated that the more mutagenic and tumorigenic (+)-enantiomer forms carboxylic ester adducts with the protein at either Asp(187) or Glu(188), while the (-)-enantiomer forms N tau-histidine adducts at His(146). This previously unrealized enantiospecificity of the reaction of benzo[a]pyrene anti-diol epoxide with human serum albumin has important consequences for the application of the adducts as biomarkers of internal exposure.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗

PubMedDate not supplied

In vivo and in vitro reactions of toluene diisocyanate isomers with guinea pig hemoglobin.

Guinea pig hemoglobin (Hb) adducts of 2,4- and 2,6-toluene diisocyanate (2,4- and 2,6-TDI) were individually prepared, each at a Hb tetramer to diisocyanate ratio of 1:1, and compared with adducts of Hb from animals exposed to 1 ppm 2,4-TDI vapor. Each Hb sample was subjected to C4 HPLC chain/heme separation with UV detection. Survey of the LC fractions using an antiserum prepared to a heterologous TDI-protein indicated a difference in chain specificity and product types formed in vitro by the two isomers. Ionspray mass spectrometry (MS) revealed the chemistry of the adduction products. Carbamoylated products, formed from adduction by one TDI molecule (with one isocyanato group hydrolyzed to an amino group), were detected by MS with 2,4-TDI on each chain and with 2,6-TDI on the beta chain. Additionally, a quasi-molecular ion of a bis carbamoylation adduct was noted by MS in the in vitro 2,4-TDI-Hb adduct in the form of TDI-cross-linked alpha and beta chains. Ionspray MS analysis of the Hb isolated from guinea pigs exposed in vivo to 2,4-TDI indicated carbamoylation products with both the alpha and beta chains in which one of the two original isocyanato groups had been hydrolyzed to the amine. We also found evidence of an amine-nitroso adduct on the alpha chain in the in vivo sample. These results indicate that at least one of the isocyanato moieties (or a masked derivative) of 2,4-TDI survived passage through the lung, into the serum, and through the erythrocyte membrane to form adducts with Hb that were stable to dialysis, gel filtration, and reversed phase HPLC separation under acidic conditions. The presence of an apparent amine-nitroso adduct indicated in vivo formation of 2,4-diaminotoluene, a recognized animal carcinogen.

Administration, Inhalation↗

PubMedDate not supplied

Cytostatic and cytotoxic action of Z-1, 1-dichloro-2,3-diphenylcyclopropane in three human breast cancer cell lines.

Z-1,1-dichloro-2,3-diphenylcyclopropane (a.k.a. Analog II, AII) is known as a potential anti-breast cancer agent and has previously been studied as an antiestrogen (AE). We hypothesized that its activity is independent of estrogen receptor (ER) status. AII and its known and potential metabolites were synthesized and characterized by NMR and MS. ER positive/estrogen and AE sensitive MCF-7, ER positive/estrogen and AE resistant MCF-7/LY2, and ER negative/estrogen and AE resistant MDA-MB231 cells were used in metabolism, cytostasis, cytotoxicity, and serum binding/wash out assays. Bacterial mutation assays were performed with Salmonella typhimurim strain TM677. AII underwent slow solvolysis in culture medium to Z-2-chloro-1,3-diphenyl-2-propen-1-ol and its oxidized form Z-alpha-chlorochalcone (ZCC). ZCC was the major metabolite of AII in all three cell lines. Cytostasis and clonogenic assays showed AII to be cytostatic to each of the lines, and was more potent against MCF-7 and MCF-7/LY2 than MDA-MB231 cells. ZCC was cytotoxic, with IC50 values of 89, 0.5, and 170 nM in MCF-7, MCF-7/LY2, and MDA-MB231 cells, respectively. Cytotoxicity from ZCC was delayed compared to loss in cell viability, suggesting a non-necrotic mechanism. Serum protected against loss of cell viability caused by AII, but had no effect on the action of ZCC. The effects of ZCC could be partially reversed by washing the drug out of cells. The effects of AII persisted after wash out. AII was also shown to be nonmutagenic in forward Salmonella mutation assays both with and without metabolic activation. In conclusion, AII, a chemical with weak antiestrogenicity, anti-breast cancer activity, and low toxicity in whole animals, shows growth inhibitory properties against both ER positive and negative human breast cancer cells in culture. Its direct action appears to be cytostatic and longlived. AII is converted by the cells to a less-retained and -protein bound metabolite, ZCC, that is more cytotoxic. Neither AII nor ZCC appear to have mutagenic activity. Both AII and ZCC thus appear to have potential for use against estrogen-dependent and -independent human breast cancers.

Alkylation↗

PubMedDate not supplied

Cellular targets of the anti-breast cancer agent Z-1,1-dichloro-2,3-diphenylcyclopropane: type II estrogen binding sites and tubulin.

Z-1,1-dichloro-2,3-diphenylcyclopropane (a.k.a. Analog II, AII) is a known anti-breast cancer agent with apparent antiestrogenic effects and remarkably low toxicity in rodents. We have recently shown that AII and its major metabolite Z-alpha-chlorochalcone (ZCC) inhibit proliferation of both estrogen-responsive and -nonresponsive human breast cancer cells, suggesting its mechanism is not mediated by the type I estrogen receptor (ER). The present studies were performed to begin to define the molecular targets of AII and ZCC. Based on the compounds' structures and actions, we hypothesized that their effects could be due to interaction at type II estrogen binding sites (EBSII) and/or cellular microtubules. The affinities of AII ZCC and the positive control diethylstilbestrol (DES) for the ER (in MCF-7 and MCF-7/LY2 cells) and EBSII (in MCF-7, MCF 7/LY2, and MDA-MB231 cells) were determined with a whole cell assay for displacement of [3H]estradiol. The kinetics of their effects on cellular microtubules and cell cycle distribution of human breast cancer cells were measured by indirect immunofluorescence and flow cytometry. Their abilities to inhibit assembly of isolated tubulin in vitro were determined. AII, ZCC, and DES had similar affinities for the EBSII in the three cell lines. Neither AII nor ZCC displaced [3H]estradiol from the ER in MCF-7 cells, whereas DES did. The microtubule network of MDA-MB231 cells exposed to 100 microM AII or 10 microM ZCC began to disassemble within 1 hour of treatment and was completely diffuse after 6 hour of exposure to either drug. AII inhibited in vitro assembly of tubulin, with an IC50 of 6.7 +/- 0.9 microM, while ZCC was inactive below 40 microM. Both drugs caused accumulation of the cells in the G2/M phase of the cell cycle. The evidence suggests that the antitumor action of AII is mediated, at least in part, through the EBSII and/or perturbation of tubulin-microtubule dynamics. AII thus represents a new lead compound for design and discovery of novel antitumor agents directed against the EBSII and/or microtubules.

Antineoplastic Agents↗