St John's Wort: effect on CYP3A4 activity.

BACKGROUND: St John's Wort is a widely used herbal product. Information regarding its potential for drug interactions is required for responsible treatment of patients using St John's Wort. CYP3A4 is a metabolic enzyme implicated in most clinically significant drug-drug interactions. OBJECTIVE: To determine the in vivo effect of reagent-grade St John's Wort extract on CYP3A4 activity through evaluation of urinary 6-beta-hydroxycortisol/cortisol ratios. METHODS: Thirteen subjects ranging in age from 18 to 25 years participated in this unblinded, multiple-dose, single-treatment before-after trial conducted in a university-based pharmacokinetics and biopharmaceutics laboratory. Each subject ingested a 300-mg tablet of reagent-grade St John's Wort extract standardized to 0.3% hypericin three times a day for 14 days. Baseline and posttreatment CYP3A4 activity was assessed with the urinary 6-beta-hydroxycortisol/cortisol ratio after a 24-hour urine collection. RESULTS: The mean +/- SD urinary 6-beta-hydroxycortisol/cortisol ratio significantly increased (P = .003) from a baseline value of 7.1 +/- 4.5 to 13 +/- 4.9. The mean +/- SD percentage increase was 114% +/- 95%, with a range from -25% to 259%. All but one subject had an increase in the ratio. CONCLUSIONS: Treatment with St John's Wort for 14 days resulted in significant increases in the urinary 6-beta-hydroxycortisol/cortisol ratio. This finding suggests that St John's Wort is an inducer of CYP3A4.

Absorption of phenytoin from rectal suppositories formulated with a polyethylene glycol base.

STUDY OBJECTIVE: To compare phenytoin pharmacokinetics following administration of an oral suspension and a rectal suppository formulated with a polyethylene glycol base. DESIGN: Unblinded, single-dose, randomized, crossover trial. SETTING: University-affiliated pharmacokinetics and biopharmaceutics laboratory. SUBJECTS: Six healthy subjects. INTERVENTION: Subjects were given a single 200-mg dose of phenytoin as an oral suspension and a rectal suppository separated by a 1-week washout. MEASUREMENTS AND MAIN RESULTS: Blood for plasma phenytoin concentrations was obtained at baseline and 0.5, 1, 2, 4, 6, 8, 10, 12, and 24 hours after administration. Plasma was analyzed by high-performance liquid chromatography (coefficient of variation < 6%) for total phenytoin concentration. Phenytoin maximum concentration (Cmax), time to Cmax (Tmax), time to first measurable concentration (Tlag), and area under the curve from time zero to time of last measurable concentration (AUClast) were estimated for oral and rectal administration by WinNonlin (v 1.1) and compared using Wilcoxon's signed rank test (p<0.05 for statistical significance). Two subjects did not have detectable plasma phenytoin concentrations after rectal administration. For the other four subjects, median rectal Cmax was significantly lower than oral Cmax (0.4 vs 1.9 microg/ml, p=0.028), median rectal Tmax did not differ from oral Tmax (11.9 vs 8.0 hrs, p=0.465), and median rectal AUClast, although highly variable, was significantly lower than oral AUClast (5.4 vs 36.2 microg x hr/ml, p=0.046). No Tlag was seen after oral administration, but with rectal administration the median Tlag was 2 hours. The estimated relative bioavailability of rectal phenytoin suppositories based on AUC0-24 was 4.7%, with individual values ranging from 0-58.3%. CONCLUSION: It appears that absorption of phenytoin from polyethylene glycol rectal suppositories in healthy subjects is highly variable and unpredictable. Thus this formulation is not recommended.

Epitope mapping and topology of baculovirus-expressed HIV-1 gp160 determined with a panel of murine monoclonal antibodies.

To define protein folding patterns of HIV-1 Env subunit vaccines, we have isolated a set of 30 monoclonal antibodies (MAbs) from BALB/c mice immunized with a recombinant gp160 vaccine (rgp160) expressed in a baculovirus system. This article describes epitope mapping for the MAb panel and topology of the epitopes for rgp160 and a recombinant gp120 (rgp120) also expressed in a baculovirus system. The following results are reported: (1) rgp160 harbors a minimum of 4 antigenic domains, 3 mapping to the C1, C2, and C3/V4 regions of gp120 and 1 mapping to the cytoplasmic tail of gp41; (2) there are at least 3 adjacent or overlapping epitopes in each antigenic domain; (3) a minimum of 14 independent epitopes were mapped, all of which are continuous sites; (4) each of the epitopes is exposed on rgp160 without prior manipulation of the protein; and (5) by contrast, 6 of the 8 epitopes mapping to the C1, C2, and C3/V4 regions are not exposed on rgp120, but become exposed when the protein is denatured. Taken together, these results show that rgp160 and rgp120 are folded differently, illustrating the use of this MAb panel to compare epitope topographies of recombination HIV-1 Env proteins. This MAb panel may aid in the refinement of HIV-1 Env subunit vaccines.

Abrogation of tumorigenicity by MHC class II antigen expression requires the cytoplasmic domain of the class II molecule.

Transfection of syngeneic MHC class II genes into the lethal mouse SaI tumor abrogates the malignancy of the tumor in the autologous host, and protects the host against subsequent challenges with the wild type class II- tumor. We have hypothesized that the transfectants induce protective immunity by functioning as APC for tumor peptides, and stimulating tumor-specific Th cells. Recent in vitro studies suggest that Ag presentation by class II-restricted APC requires the cytoplasmic domain of the class II molecule, and may involve intracellular signaling via the cytoplasmic domain. To determine if the class II cytoplasmic domain is required for enhanced tumor-specific immunity, SaI mouse sarcoma cells were transfected with syngeneic Aak and Abk genes with truncated cytoplasmic domains. These transfectants are as malignant as wild type class II- SaI cells in autologous A/J mice. Stimulation of tumor-specific immunity by class II+ tumor cells is therefore dependent on the class II cytoplasmic region, and may involve intracellular signaling events.

Influence of major histocompatibility complex class I, class II and TLA genes on tumor rejection.

T lymphocytes recognize antigen associated with MHC class I and/or class II gene products. Recognition of malignant cells is therefore dependent on presentation of tumor associated antigen(s) by MHC molecules. We have studied immunity to tumors that have down-regulated class I expression. These studies demonstrate a requirement for class I antigens, but suggest that additional factors may also be required for tumor-specific immunity. The MHC also encodes TLA class I antigens, whose function is unknown. Our studies suggest that these molecules function is unknown. Our studies suggest that T lymphocytes, specifically in tumor cells that do not express H-2K or H-2D moieties. Other studies are aimed at improving tumor-specific Th cell generation by producing class II+ tumor cells. The success of these experiments indicates that this approach may be a potentially useful immunotherapy.