Enantioselective, mechanism-based inactivation of guinea pig hepatic cytochrome P450 by N-(alpha-methylbenzyl)-1-aminobenzotriazole

N-Aralkylated derivatives of 1-aminobenzotriazole are well-established, mechanism-based inhibitors of cytochrome P450 (CYP or P450). In this study, the kinetics of inactivation of CYP2B-dependent 7-pentoxyresorufin O-depentylation (PROD) and CYP1A-dependent 7-ethoxyresorufin O-deethylation (EROD) activities by enantiomers of N-(alpha-methylbenzyl)-1-aminobenzotriazole (alphaMB) were compared. The racemic mixture (+/-)-alphaMB, as well as the enantiomers (-)-alphaMB and (+)-alphaMB, produced a time-, concentration-, and NADPH-dependent loss of PROD and EROD activity in hepatic microsomes from phenobarbital-treated guinea pigs. The rates of PROD inactivation by (-)-alphaMB were significantly faster than for (+)-alphaMB. Consistent with this, the derived maximal kinact was also significantly greater for (-)-alphaMB than for (+)-alphaMB (0.49 vs. 0.35 min-1). In contrast, the concentrations required for the half-maximal rate of inactivation (Ki) were equivalent for (-)-alphaMB and (+)-alphaMB, whereas the degree of competitive inhibition of PROD activity was greater for (+)-alphaMB. No significant differences were found among (-)-alphaMB, (+)-alphaMB, and (+/-)-alphaMB with respect to mechanism-based inactivation (kinact = 0.18, 0.16, and 0.17 min-1, respectively) or competitive inhibition of EROD activity. No differences were found for the maximal extent of PROD or EROD inhibition or the loss of spectral P450 after an extended 30-min incubation with the inhibitors. We conclude that mechanism-based inactivation of guinea pig CYP2B, but not CYP1A, isozymes by alphaMB occurs in a stereoselective manner, most likely as a result of a difference in the balance between metabolic activation and deactivation for the alphaMB enantiomers.     

Biodegradation pathway of 2-chlorodibenzo-p-dioxin and 2-chlorodibenzofuran in the biphenyl-utilising strain JB1

The biphenyl-utilising Burkholderia (previously Alcaligenes) strain JB1 is also able to degrade a number of chlorinated dibenzo-p-dioxins and dibenzofurans. In this study, 4-chlorocatechol and a chlorotrihydroxydiphenyl ether were identified as metabolites of 2-chlorodibenzo-p-dioxin. 5-Chlorosalicylic acid and a chlorotrihydroxybiphenyl were metabolites of 2-chlorodibenzofuran. These results show that degradation of these compounds follows pathways in which the initial reaction is angular dioxygenation, followed by cleavage of an ether bridge. This pathway is similar to that used by dibenzofuran-degrading strains such as Sphingomonas sp. strain RW1.     

Efficacy of low-dose dopamine in preventing amphotericin B nephrotoxicity in bone marrow transplant patients and leukemia patients

This study evaluated the efficacy of low-dose dopamine for prevention of amphotericin B-induced nephrotoxicity in autologous bone marrow transplant and leukemia patients. Seventy-one patients undergoing cytoreductive therapy who required amphotericin B were randomly assigned in an unblinded fashion to a group receiving continuous-infusion low-dose dopamine (3 microgram/kg/min) or a group receiving no dopamine. Amphotericin B was dosed at 0.5 or 1.0 mg/kg/day based on computerized tomography scan results or presence of positive blood cultures. No patient received saline boluses. The rate of nephrotoxicity, severity as graded by Southwest Oncology Group toxicity criteria, and time to each grade of nephrotoxicity were compared between the two groups. Eighty percent of the no-dopamine group and 66.7% of the dopamine group developed nephrotoxicity, defined as a 1.5-fold or greater increase in baseline serum creatinine level (P = 0.20). No statistical difference was noted at any grade of nephrotoxicity between the two groups. Thirty-four percent of patients in the no-dopamine group versus 17.6% in the dopamine group had a 2.5-fold or greater increase in serum creatinine level, which was not statistically significant (P = 0.0888). Ten patients developed grade IV nephrotoxicity and were withdrawn from the study, 7 in the no-dopamine group and 3 in the dopamine group (P = 0.19). The time to each grade of nephrotoxicity was also not significantly different for the two groups. Eleven adverse drug reactions were reported in the dopamine group in comparison to one in the no-dopamine group. Thus, dopamine offers little in the way of prevention of nephrotoxicity associated with amphotericin B therapy. Although the significance of drug reactions in the dopamine group is not clearly established due to lack of cardiac monitoring in the no-dopamine group, dopamine therapy is not without complications.     

Inactivation of the mitogen-activated protein kinase Mps1 from the rice blast fungus prevents penetration of host cells but allows activation of plant defense responses

The rice blast fungus, Magnaporthe grisea, generates enormous turgor pressure within a specialized cell called the appressorium to breach the surface of host plant cells. Here, we show that a mitogen-activated protein kinase, Mps1, is essential for appressorium penetration. Mps1 is 85% similar to yeast Slt2 mitogen-activated protein kinase and can rescue the thermosensitive growth of slt2 null mutants. The mps1-1Delta mutants of M. grisea have some phenotypes in common with slt2 mutants of yeast, including sensitivity to cell-wall-digesting enzymes, but display additional phenotypes, including reduced sporulation and fertility. Interestingly, mps1-1Delta mutants are completely nonpathogenic because of the inability of appressoria to penetrate plant cell surfaces, suggesting that penetration requires remodeling of the appressorium wall through an Mps1-dependent signaling pathway. Although mps1-1Delta mutants are unable to cause disease, they are able to trigger early plant-cell defense responses, including the accumulation of autofluorescent compounds and the rearrangement of the actin cytoskeleton. We conclude that MPS1 is essential for pathogen penetration; however, penetration is not required for induction of some plant defense responses.