Impact of Rifabutin or Rifampin on Bedaquiline Safety, Tolerability, and Pharmacokinetics Assessed in a Randomized Clinical Trial with Healthy Adult Volunteers.

Bedaquiline is a diarylquinoline that specifically inhibits mycobacterial ATP synthase. Bedaquiline has been used to effectively treat tuberculosis (TB) caused by drug-susceptible and drug-resistant Mycobacterium tuberculosis Rifamycins are a cornerstone of combination drug regimens for the treatment of TB. This phase 1, open-label, randomized, controlled trial evaluated the effect of steady-state dosing of rifabutin or rifampin on the safety, tolerability, and pharmacokinetics of bedaquiline given as a single dose. Thirty-three healthy subjects were enrolled to receive a 400-mg single oral dose of bedaquiline at two time points, on study days 1 and 29. Subjects were randomly assigned to once daily oral doses of rifabutin (300 mg/day, n = 17) or rifampin (600 mg/day, n = 16) during period 2 from days 20 to 41. Serial blood sampling for bedaquiline measurement occurred on days 1 and 29 through 336 h after bedaquiline administration. The day 29 bedaquiline pharmacokinetic parameter estimates were compared to the corresponding day 1 estimates for each rifamycin group. Steady-state rifampin reduced bedaquiline AUC0-336 approximately 45%, from 47.69 h·µg/ml in period 1 to 26.33 h·µg/ml in period 2. Bedaquiline apparent clearance accelerated 24% in rifampin-treated subjects from 6.59 liters/h in period 1 to 8.19 liters/h in period 2. Steady-state rifabutin resulted in little quantitative impact on bedaquiline exposure but was associated with grade 3 and 4 adverse events before and after the day 29 bedaquiline dose. Dosage adjustments may therefore be necessary to ensure that bedaquiline plasma concentrations reach therapeutic levels safely when combining bedaquiline and rifamycins in TB treatment regimens. (This single-site, randomized, open-label, prospective study in healthy adult volunteers was registered at Clinicaltrials.gov under registration no. NCT01341184.).

A Phase 1 Pharmacokinetic and Safety Study of Extended-Duration, High-dose Cefixime for Cephalosporin-resistant Neisseria gonorrhoeae in the Pharynx.

BACKGROUND: There are no fully oral recommended treatment regimens for gonorrhea. Inadequately treated pharyngeal gonococcal infections are a likely reservoir for transmission and development of antimicrobial resistance. We sought to determine an oral cefixime dosing regimen that would theoretically treat pharyngeal infections by gonococci with minimum inhibitory concentrations 0.5 µg/mL. METHODS: We conducted an open-label, nonrandomized, phase I pharmacokinetic and safety study of cefixime in 25 healthy male and female volunteers divided into 4 dosing cohorts (cohort A, 400 mg; cohort B, 800 mg; cohort C, 1200 mg; and cohort D, 800 mg every 8 hours × 3 doses [total dose 2400 mg]) with a target serum concentration of at least 2.0 µg/mL for more than 20 hours. Cefixime concentrations from serum and pharyngeal fluid were determined with use of a validated liquid chromatography-tandem mass spectrometry assay. Safety measures included laboratories, physical examinations, and symptom diaries. RESULTS: None of the single-dose regimens attained the target concentration; however, 50% of subjects in cohort D attained the target concentration. Variation in absorption and protein binding contributed to differences in concentrations. Pharyngeal fluid concentrations were negligible. The single-dose regimens were well tolerated; the multidose regimen resulted in mild to moderate gastrointestinal symptoms in 43% of subjects. CONCLUSIONS: None of the dosing regimens achieved the target concentration. However, the proposed theoretical target was extrapolated from penicillin data; there are no empirically derived pharmacokinetic/pharmacodynamic criteria for pharyngeal gonorrhea. Under alternative cephalosporin-specific therapeutic goals, the multidose regimen may be effective, although the absence of cefixime in pharyngeal fluid is concerning. A clinical trial evaluating efficacy and defining pharmacokinetic/pharmacodynamic outcomes may be warranted.

Safety, Tolerability, Systemic Exposure, and Metabolism of CRS3123, a Methionyl-tRNA Synthetase Inhibitor Developed for Treatment of Clostridium difficile, in a Phase 1 Study.

Clostridium difficile causes antibiotic-associated diarrhea and is a major public health concern. Current therapies disrupt the protective intestinal flora, do not reliably prevent recurrent infections, and will be decreasingly effective should less susceptible strains emerge. CRS3123 is an oral agent that inhibits bacterial methionyl-tRNA synthetase and has potent activity against C. difficile and aerobic Gram-positive bacteria but little activity against Gram-negative bacteria, including anaerobes. This first-in-human, double-blind, placebo-controlled, dose escalation study evaluated the safety and systemic exposure of CRS3123 after a single oral dose in healthy adults. Five cohorts of eight subjects each received CRS3123 or placebo in a 3:1 ratio. Doses for the respective active arms were 100 mg, 200 mg, 400 mg, 800 mg, and 1,200 mg. Blood and urine were collected for pharmacokinetic analysis. CRS3123 concentrations were measured with validated LC-MS/MS techniques. There were no serious adverse events or immediate allergic reactions during administration of CRS3123. In the CRS3123-treated groups, the most frequent adverse events were decreased hemoglobin, headache, and abnormal urine analysis; all adverse events in the active-treatment groups were mild to moderate, and their frequency did not increase with dose. Although CRS3123 systemic exposure increased at higher doses, the increase was less than dose proportional. The absorbed drug was glucuronidated at reactive amino groups on the molecule, which precluded accurate pharmacokinetic analysis of the parent drug. Overall, CRS3123 was well tolerated over this wide range of doses. This safety profile supports further investigation of CRS3123 as a treatment for C. difficile infections. (This study has been registered at ClinicalTrials.gov under identifier NCT01551004.).

Dosing Colistimethate Every 8 h Results in Higher Plasma Concentrations of Active Colistin Than Every 12-Hourly Dosing without Increase in Nephrotoxicity: A Phase 1 Pharmacokinetics Trial in Healthy Adult Volunteers.

Despite its use for decades, pharmacokinetic (PK) and safety studies on colistin are limited. We conducted a phase l, open-label trial to evaluate the safety and PK of multiple doses of intravenous (IV) and aerosolized colistimethate sodium (CMS) administered separately and in combination. In total, 31 healthy adults were enrolled into three cohorts of 9, 10, and 12 participants, respectively. Each cohort received increasing doses of CMS over three dosing periods as follows: Period 1 (IV only), 2.5 mg/kg every 12 h (q12h) to 3.3 mg/kg every 8 h (q8h); Period 2 (aerosolized only), 75 mg 2-4 doses, and Period 3 (combined IV aerosolized), in which was Periods 1 and 2 combined. Safety assessments, serum and lung concentrations of colistin analytes (colistin A, colistin B, CMS A, and CMS B), and kidney biomarkers were measured at specified time points. Increasing the CMS dose from 2.5 mg/kg q12h to q8h resulted in a 33% increase in serum colistin A concentrations from 3.9 µg/mL to 5.3 µg/mL-well above the accepted target of 2 µg/mL for 6 h after dosing, without evidence of nephrotoxicity. However, there was an increase in neurotoxicity, primarily perioral and lingual paresthesias, and self-limited ataxia. IV administration did not increase the lung concentrations of colistin.

A two-part phase 1 study to establish and compare the safety and local tolerability of two nasal formulations of XF-73 for decolonisation of Staphylococcus aureus: A previously investigated 0.5mg/g viscosified gel formulation versus a modified formulation.

OBJECTIVES: Successful decolonisation of nasal Staphylococcus aureus (SA) carriage by mupirocin is limited by increasing drug resistance. This randomised, open-label, phase 1 study compared the safety and local tolerability of two nasal formulations of XF-73, a novel porphyrinic antibacterial with rapid intrinsic activity against SA. METHODS: The study was performed in 60 healthy adults. In Part 1, eight non-SA carriers were randomised to groups of four subjects each and were treated with XF-73 concentrations of 0.5mg/g 2% gel or 2.0mg/g 2% gel. In Part 2, 52 persistent SA carriers were randomised to groups of 13 subjects each and were treated with XF-73 concentrations of 0.5mg/g 2% gel, 2.0mg/g 2% gel, 0.5mg/g 4% gel or 4% viscosified placebo gel. Plasma pharmacokinetic and pharmacodynamic studies were performed. Antistaphylococcal activity was assessed as the presence/absence of SA and by quantification of colonisation using a semiquantitative scale (SA score). RESULTS: 56 subjects (8/8 from Part 1 and 48/52 from Part 2) completed the study, with 47/60 comprising the pharmacokinetic population and 48/60 the pharmacodynamic population. There was no measurable systemic absorption of XF-73. XF-73 treatment was associated with rapid reduction in SA score in all subjects. The most common treatment-emergent adverse events (TEAEs) were rhinorrhoea and nasal dryness (15.5% each in Parts 1 and 2). TEAEs were mild and resolved spontaneously. CONCLUSION: XF-73 was well tolerated with minimal side effects at doses of 0.5mg/g 2% gel and 2.0mg/g 2% gel. These findings support further development of XF-73.

Phytoestrogens regulate mRNA and protein levels of guanine nucleotide-binding protein, beta-1 subunit (GNB1) in MCF-7 cells.

Phytoestrogens (PEs) are non-steroidal ligands, which regulate the expression of number of estrogen receptor-dependent genes responsible for a variety of biological processes. Deciphering the molecular mechanism of action of these compounds is of great importance because it would increase our understanding of the role(s) these bioactive chemicals play in prevention and treatment of estrogen-based diseases. In this study, we applied suppression subtractive hybridization (SSH) to identify genes that are regulated by PEs through either the classic nuclear-based estrogen receptor or membrane-based estrogen receptor pathways. SSH, using mRNA from genistein (GE) treated MCF-7 cells as testers, resulted in a significant increase in GNB1 mRNA expression levels as compared with 10 nM 17beta estradiol or the no treatment control. GNB1 mRNA expression was up regulated two- to fivefold following exposure to 100.0 nM GE. Similarly, GNB1 protein expression was up regulated 12- to 14-fold. GE regulation of GNB1 was estrogen receptor-dependent, in the presence of the anti-estrogen ICI-182,780, both GNB1 mRNA and protein expression were inhibited. Analysis of the GNB1 promoter using ChIP assay showed a PE-dependent association of estrogen receptor alpha (ERalpha) and beta (ERbeta) to the GNB1 promoter. This association was specific for ERalpha since association was not observed when the cells were co-incubated with GE and the ERalpha antagonist, ICI. Our data demonstrate that the levels of G-protein, beta-1 subunit are regulated by PEs through an estrogen receptor pathway and further suggest that PEs may control the ratio of alpha-subunit to beta/gamma-subunits of the G-protein complex in cells. J. Cell. Physiol. 219: 584-594, 2009. (c) 2009 Wiley-Liss, Inc.

Development and validation of an LC-MS/MS method for the simultaneous determination of bedaquiline and rifabutin in human plasma.

This article describes the simultaneous determination of bedaquiline fumarate (TMC-207) and rifabutin in human plasma by stable isotope dilution tandem mass spectrometry. The methodology was developed for an investigation of potential drug-drug interactions of the two anti-tuberculosis drugs when given together to healthy human volunteers. Use of the two drugs in combination to treat disease caused by Mycobacterium tuberculosis is contemplated as the bacterium becomes resistant to many currently available drugs.

Effects of Rifamycin Coadministration on Bedaquiline Desmethylation in Healthy Adult Volunteers.

There is an urgent need to identify safe and effective combination treatments for multidrug-resistant (MDR) Mycobacterium tuberculosis infection (TB). Bedaquiline, a new diarylquinoline, is approved for the treatment of MDR pulmonary TB in combination with other drugs, which could include rifabutin, which is also used to treat drug-resistant TB. Both rifabutin and bedaquiline are metabolized via cytochrome P450 3A4, and rifabutin is an inducer of this enzyme. Bedaquiline is metabolized into its primary N-monodesmethyl metabolite, M2, and further desmethylated into an N-didesmethyl metabolite, M3. Both metabolites are cytotoxic and induce phospholipidosis. The effect of rifabutin on the generation and disposition of the 2 metabolites was investigated in healthy adult volunteers coadministered bedaquiline and either rifabutin or rifampin. Subjects received single oral doses (400 mg) of bedaquiline on days 1 and 29. Oral rifabutin (300 mg) or rifampin (600 mg) were given daily on days 20-41. In the rifabutin group maximum M2 concentrations (Cmax ) increased significantly (P < .001) from 47.59 to 79.53 ng/mL, and clearance slowed slightly (P = .01). This resulted in significantly (P < .001) increased overall exposure (area under the concentration-time curve [AUC0-τ ]). Peak concentrations of M3 increased approximately 3-fold with little decline thereafter. In rifampin recipients M2 Cmax doubled (48.44 to 101.52 ng/mL), but M2 clearance and time to Cmax significantly (P < .001) increased, and AUC0-∞ and mean residence time significantly decreased (P < .001). Peak M3 concentrations increased 4-fold and rapidly declined. Although both rifamycins accelerate desmethylation of bedaquiline and M2, differences in clearance resulted in sustained elevations of both metabolites during rifabutin, but not rifampin, treatment.

Developmental neurotoxicity and autism: A potential link between indoor neuroactive pollutants and the curious birth order risk factor.

Epidemiological and demographic studies find an increased risk of autism among first-borns. Toxicological studies show that some semi-volatile substances found in infant products produce adverse effects in neural and endocrine systems of animals, including behavioral and developmental effects. Several factors elevate the exposure of human infants to these chemicals. The highest exposures found in infants are comparable to the exposures that induce neural toxicity in animals. A review of these literatures suggests a linking hypothesis that could bridge the epidemiological and toxicological lines of evidence: an infant's exposure to neuroactive compounds emitted by infant products is increased by product newness and abundance; exposure is likely maximized for first-born children in families that can afford new products. Exposure is reduced for subsequently-born children who reuse these now neuroactive-depleted products. The presence of neuroactive chemical emissions from infant products has implications for birth-order effects and for other curious risk factors in autism, including gender, socioeconomic status, and season-of-birth risk factors.

Does F1-ATPase have a catalytic site that preferentially binds MgADP?

During ATP synthesis, ATP synthase has to bind MgADP in the presence of an excess of MgATP. Thus, for efficient ATP synthesis it would be desirable if incoming substrate could be bound to a catalytic site with a preference for MgADP over MgATP. We tested three hypotheses predicting the existence of such a site. However, our results showed that, at least in absence of an electrochemical proton gradient, none of the three catalytic sites has a higher affinity for MgADP than for MgATP.

Molecular and biochemical effects of a kola nut extract on androgen receptor-mediated pathways.

The low incidence of prostate cancer in Asians has been attributed to chemopreventative properties of certain chemicals found in their diet. This study characterized the androgenic and chemopreventative properties of the Jamaican bush tea "Bizzy," using androgen receptor positive and negative cell lines. Exposure of prostate cells to Biz-2 resulted in a growth inhibition (GI(50)) of 15 ppm in LNCaP cells and 3.6 ppm in DU145 cells. Biz-2 elicited a 2-fold increase in the mRNA of the anti-apoptotic gene Bcl2, with a 10-fold increase in that of the proapoptotic gene Bax. We observed a 2.4- to 7.5-fold change in apoptotic cells in both cell lines. Biz-2 at 10 ppm elicited a time- and dose-dependent stimulation of both the protein and mRNA levels of several androgen-regulated genes. Biz-2 caused a 36% decrease in PSA secretion and a significant increase in PSA mRNA. The relative binding affinity (IC(50)) of Biz-2 for AR was 2- to 5-fold lower than that of the synthetic androgen R1881. Biz-2 was found to be a specific ligand for the AR in that the natural ligand, DHT, and the anti-androgen, flutamide, displaced Biz-2 bound to AR and inhibited Biz-2-induced transcription and PSA secretion. This study provided evidence that Biz-2 extract possesses the ability to modulate prostate cancer cell biology in an AR-dependent manner.

Characterization of Bizzy Nut extracts in estrogen-responsive MCF-7 breast cancer cells.

Kola acuminate, also known as Bizzy Nut or Kola Nut, is a natural product that contains bioactive chemicals that possess hormonal properties. The purpose of this study was to characterize the putative phytoestrogenic compounds present in Bizzy Nut for estrogenic-like activity. As an initial step, five extracts (E1 - hexane, E2 - ether, E3 - acetone, E4 - methanol and E5 - water) were sequentially generated using solid-liquid phase extraction and their bioactivity was examined in MCF-7, MDA-MB-468 and LNCaP cancer cell models. MTT cell viability, dye exclusion, caspase activity and microscopic assessment of apoptotic cells demonstrated that extracts of Bizzy were cytotoxic to MCF-7, MDA-MB 468 and LNCaP cells. In MCF-7 cells, the acetone extract (E3) at 100 ppm elicited a potent cytotoxic response with a growth-inhibitory concentration (GI(50)) of 67 ppm. In contrast, E3 stimulated growth in LNCaP cells. The ether extract (E2) showed a dose-dependent cytotoxic response with a GI(50) of 13 ppm in the LNCaP cell line. Examination of the apoptotic response induced by E2 and E3 paralleled the level of cell cytotoxicity observed in both cell lines. The methanol extract (E4) was the only extract that showed a time-, dose-, and estrogen-receptor-dependent stimulation of pS2 gene expression. On the other hand, the acetone extract (E3), which showed the highest degree of cytotoxicity, showed no transcription stimulation of pS2 in MCF-7 cells. Altogether, these data indicate that Bizzy contains unique active hormonal compounds that have specific biological properties that are cell line-dependent.

Identification and characterization of a phytoestrogen-specific gene from the MCF-7 human breast cancer cell.

Phytoestrogens are a group of compounds present in human diet that display estrogenic-like properties. Several studies have demonstrated that populations who consume large quantities of phytoestrogens have a reduced risk of estrogen-dependent cancers. Although it has been shown that certain phytoestrogens modulate estrogen action, their biological role in cancer reduction remains unclear. Through the use of differential display reverse transcriptase-polymerase chain reaction and representational difference analysis of cDNA, we have identified several phytoestrogen-responsive genes from the human breast cancer cell MCF-7. Two of these genes, PE-13.1 and pRDA-D, have been characterized in greater detail in this study. These genes were not previously known to be regulated by phytoestrogen or estradiol. PE-13.1 is a novel gene that specifies the coding of a 1.10-kb mRNA transcript. Northern blot analysis confirmed that the PE-13.1 transcript is up-regulated by phytoestrogens (Genistein, sevenfold; Zearalenone, twofold) and is nonresponsive to estradiol. Conversely, the pRDA-D transcript was down-regulated by both phytoestrogens and estradiol. The antiestrogen ICI-182,780 inhibits the expression of PE-13.1 and reverses the inhibition of pRDA-D expression induced by phytoestrogens and estradiol. Analysis of the tissue distribution of PE-13.1 transcript by RNA blot reveals that this transcript is expressed in both normal and tumor tissues. This report demonstrates for the first time the presence of two phytoestrogen-responsive genes that may be used as molecular markers in understanding the role dietary estrogen plays in cancer prevention.