Pharmacokinetics, microbial response, and pulmonary outcomes of multidose intravenous azithromycin in preterm infants at risk for Ureaplasma respiratory colonization.

The study objectives were to refine the population pharmacokinetics (PK) model, determine microbial clearance, and assess short-term pulmonary outcomes of multiple-dose azithromycin treatment in preterm infants at risk for Ureaplasma respiratory colonization. Fifteen subjects (7 of whom were Ureaplasma positive) received intravenous azithromycin at 20 mg/kg of body weight every 24 h for 3 doses. Azithromycin concentrations were determined in plasma samples obtained up to 168 h post-first dose by using a validated liquid chromatography-tandem mass spectrometry method. Respiratory samples were obtained predose and at three time points post-last dose for Ureaplasma culture, PCR, antibiotic susceptibility testing, and cytokine concentration determinations. Pharmacokinetic data from these 15 subjects as well as 25 additional subjects (who received either a single 10-mg/kg dose [n = 12] or a single 20-mg/kg dose [n = 13]) were analyzed by using a nonlinear mixed-effect population modeling (NONMEM) approach. Pulmonary outcomes were assessed at 36 weeks post-menstrual age and 6 months adjusted age. A 2-compartment model with all PK parameters allometrically scaled on body weight best described the azithromycin pharmacokinetics in preterm neonates. The population pharmacokinetics parameter estimates for clearance, central volume of distribution, intercompartmental clearance, and peripheral volume of distribution were 0.15 liters/h · kg(0.75), 1.88 liters · kg, 1.79 liters/h · kg(0.75), and 13 liters · kg, respectively. The estimated area under the concentration-time curve over 24 h (AUC24)/MIC90 value was ∼ 4 h. All posttreatment cultures were negative, and there were no drug-related adverse events. One Ureaplasma-positive infant died at 4 months of age, but no survivors were hospitalized for respiratory etiologies during the first 6 months (adjusted age). Thus, a 3-day course of 20 mg/kg/day intravenous azithromycin shows preliminary efficacy in eradicating Ureaplasma spp. from the preterm respiratory tract.

Opioids and efflux transporters. Part 4: influence of N-substitution on P-glycoprotein substrate activity of noroxymorphone analogues.

The efflux transporter protein P-glycoprotein (P-gp) is capable of affecting the central distribution of diverse neurotherapeutics, including opioid analgesics, through their active removal from the brain. P-gp located at the blood brain barrier has been implicated in the development of tolerance to opioids and demonstrated to be up-regulated in rats tolerant to morphine and oxycodone. We have previously examined the influence of hydrogen-bonding oxo-substitutents on the P-gp-mediated efflux of 4,5-epoxymorphinan analgesics, as well as that of N-substituted analogues of meperidine. Structure-activity relationships (SAR) governing N-substituent effects on opioid efficacy is well-established, however the influence of such structural modifications on P-gp-mediated efflux is unknown. Here, we present SAR describing P-gp recognition of a short series of N-modified 4,5-epoxymorphinans. Oxymorphone, naloxone, naltrexone, and nalmexone all failed to demonstrate P-gp substrate activity, indicating these opioid scaffolds contain structural features that preclude recognition by the transporter. These results are examined using mathematical molecular modeling and discussed in comparison to other opioid scaffolds bearing similar N-substituents.

Azithromycin to prevent bronchopulmonary dysplasia in ureaplasma-infected preterm infants: pharmacokinetics, safety, microbial response, and clinical outcomes with a 20-milligram-per-kilogram single intravenous dose.

Ureaplasma respiratory tract colonization is associated with bronchopulmonary dysplasia (BPD) in preterm infants. Previously, we demonstrated that a single intravenous (i.v.) dose of azithromycin (10 mg/kg of body weight) is safe but inadequate to eradicate Ureaplasma spp. in preterm infants. We performed a nonrandomized, single-arm open-label study of the pharmacokinetics (PK) and safety of intravenous 20-mg/kg single-dose azithromycin in 13 mechanically ventilated neonates with a gestational age between 24 weeks 0 days and 28 weeks 6 days. Pharmacokinetic data from 25 neonates (12 dosed with 10 mg/kg i.v. and 13 dosed with 20 mg/kg i.v.) were analyzed using a population modeling approach. Using a two-compartment model with allometric scaling of parameters on body weight (WT), the population PK parameter estimates were as follows: clearance, 0.21 liter/h × WT(kg)(0.75) [WT(kg)(0.75) indicates that clearance was allometrically scaled on body weight (in kilograms) with a fixed exponent of 0.75]; intercompartmental clearance, 2.1 liters/h × WT(kg)(0.75); central volume of distribution (V), 1.97 liters × WT (kg); and peripheral V, 17.9 liters × WT (kg). There was no evidence of departure from dose proportionality in azithromycin exposure over the tested dose range. The calculated area under the concentration-time curve over 24 h in the steady state divided by the MIC90 (AUC24/MIC90) for the single dose of azithromycin (20 mg/kg) was 7.5 h. Simulations suggest that 20 mg/kg for 3 days will maintain azithromycin concentrations of >MIC50 of 1 µg/ml for this group of Ureaplasma isolates for ≥ 96 h after the first dose. Azithromycin was well tolerated with no drug-related adverse events. One of seven (14%) Ureaplasma-positive subjects and three of six (50%) Ureaplasma-negative subjects developed physiologic BPD. Ureaplasma was eradicated in all treated Ureaplasma-positive subjects. Simulations suggest that a multiple-dose regimen may be efficacious for microbial clearance, but the effect on BPD remains to be determined.

Induction of xenobiotic receptors, transporters, and drug metabolizing enzymes by oxycodone.

Perturbations of the expression of transporters and drug-metabolizing enzymes (DMEs) by opioids can be the locus of deleterious drug-drug interactions (DDIs). Many transporters and DMEs are regulated by xenobiotic receptors [XRs; e.g., pregnane X receptor (PXR), constitutive androstane receptor (CAR), and Aryl hydrocarbon receptor (AhR)]; however, there is a paucity of information regarding the influence of opioids on XRs. The objective of this study was to determine the influence of oxycodone administration (15 mg/kg intraperitoneally twice daily for 8 days) on liver expression of XRs, transporters, and DMEs in rats. Microarray, quantitative real-time polymerase chain reaction and immunoblotting analyses were used to identify significantly regulated genes. Three XRs (e.g., PXR, CAR, and AhR), 27 transporters (e.g., ABCB1 and SLC22A8), and 19 DMEs (e.g., CYP2B2 and CYP3A1) were regulated (P < 0.05) with fold changes ranging from -46.3 to 17.1. Using MetaCore (computational platform), we identified a unique gene-network of transporters and DMEs assembled around PXR, CAR, and AhR. Therefore, a series of transactivation/translocation assays were conducted to determine whether the observed changes of transporters/DMEs are mediated by direct activation of PXR, CAR, or AhR by oxycodone or its major metabolites (noroxycodone and oxymorphone). Neither oxycodone nor its metabolites activated PXR, CAR, or AhR. Taken together, these findings identify a signature hepatic gene-network associated with repeated oxycodone administration in rats and demonstrate that oxycodone alters the expression of many transporters and DMEs (without direct activation of PXR, CAR, and AhR), which could lead to undesirable DDIs after coadministration of substrates of these transporters/DMEs with oxycodone.

Increased oxidative-modifications of cytosolic proteins in 3,4-methylenedioxymethamphetamine (MDMA, ecstasy)-exposed rat liver.

It is well established that 3,4-methylenedioxymethamphetamine (MDMA, ecstasy) causes acute liver damage in animals and humans. The aim of this study was to identify and characterize oxidative modification and inactivation of cytosolic proteins in MDMA-exposed rats. Markedly increased levels of oxidized and nitrated cytosolic proteins were detected 12 h after the second administration of two consecutive MDMA doses (10 mg/kg each). Comparative 2-DE analysis showed markedly increased levels of biotin-N-methylimide-labeled oxidized cytosolic proteins in MDMA-exposed rats compared to vehicle-treated rats. Proteins in the 22 gel spots of strong intensities were identified using MS/MS. The oxidatively modified proteins identified include anti-oxidant defensive enzymes, a calcium-binding protein, and proteins involved in metabolism of lipids, nitrogen, and carbohydrates (glycolysis). Cytosolic superoxide dismutase was oxidized and its activity significantly inhibited following MDMA exposure. Consistent with the oxidative inactivation of peroxiredoxin, MDMA activated c-Jun N-terminal protein kinase and p38 kinase. Since these protein kinases phosphorylate anti-apoptotic Bcl-2 protein, their activation may promote apoptosis in MDMA-exposed tissues. Our results show for the first time that MDMA induces oxidative-modification of many cytosolic proteins accompanied with increased oxidative stress and apoptosis, contributing to hepatic damage.

The ethanol metabolite acetaldehyde increases paracellular drug permeability in vitro and oral bioavailability in vivo.

Alcohol consumption leads to the production of the highly reactive ethanol metabolite, acetaldehyde, which may affect intestinal tight junctions and increase paracellular permeability. We examined the effects of elevated acetaldehyde within the gastrointestinal tract on the permeability and bioavailability of hydrophilic markers and drug molecules of variable molecular weight and geometry. In vitro permeability was measured unidirectionally in Caco-2 and MDCKII cell models in the presence of acetaldehyde, ethanol, or disulfiram, an aldehyde dehydrogenase inhibitor, which causes acetaldehyde formation when coadministered with ethanol in vivo. Acetaldehyde significantly lowered transepithelial resistance in cell monolayers and increased permeability of the low-molecular-weight markers, mannitol and sucrose; however, permeability of high-molecular-weight markers, polyethylene glycol and inulin, was not affected. In vivo permeability was assessed in male Sprague-Dawley rats treated for 6 days with ethanol, disulfiram, or saline alone or in combination. Bioavailability of naproxen was not affected by any treatment, whereas that of paclitaxel was increased upon acetaldehyde exposure. Although disulfiram has been shown to inhibit multidrug resistance-1 P-glycoprotein (P-gp) in vitro, our data demonstrate that the known P-gp substrate paclitaxel is not affected by coadministration of disulfiram. In conclusion, we demonstrate that acetaldehyde significantly modulates tight junctions and paracellular permeability in vitro as well as the oral bioavailability of low-molecular-weight hydrophilic probes and therapeutic molecules in vivo even when these molecules are substrates for efflux transporters. These studies emphasize the significance of ethanol metabolism and drug interactions outside of the liver.

Differential activation of pregnane X receptor and constitutive androstane receptor by buprenorphine in primary human hepatocytes and HepG2 cells.

Buprenorphine is a partial µ-opioid receptor agonist used for the treatment of opioid dependence that has several advantages over methadone. The principal route of buprenorphine disposition has been well established; however, little is known regarding the potential for buprenorphine to influence the metabolism and clearance of other drugs by affecting the expression of drug-metabolizing enzymes (DMEs). Here, we investigate the effects of buprenorphine on the activation of pregnane X receptor (PXR) and constitutive androstane receptor (CAR), as well as the induction of DMEs, in both HepG2 cells and human primary hepatocytes (HPHs). In HepG2 cells, buprenorphine significantly increased human PXR-mediated CYP2B6 and CYP3A4 reporter activities. CYP2B6 reporter activity was also enhanced by buprenorphine in HepG2 cells cotransfected with a chemical-responsive human CAR variant. Real-time reverse transcription-polymerase chain reaction analysis revealed that buprenorphine strongly induced CYP3A4 expression in both PXR- and CAR-transfected HepG2 cells. However, treatment with the same concentrations of buprenorphine in HPHs resulted in literally no induction of CYP3A4 or CYP2B6 expression. Further studies indicated that buprenorphine could neither translocate human CAR to the nucleus nor activate CYP2B6/CYP3A4 reporter activities in transfected HPHs. Subsequent experiments to determine whether the differential response was due to buprenorphine's metabolic stability revealed a dramatically differential rate of elimination for buprenorphine between HPHs and HepG2 cells. Taken together, these studies indicate that metabolic stability of buprenorphine defines the differential induction of DMEs observed in HepG2 and HPHs, and the results obtained from PXR and CAR reporter assays in immortalized cell line require cautious interpretation.

Characterization of the transport, metabolism, and pharmacokinetics of the dopamine D3 receptor-selective fluorenyl- and 2-pyridylphenyl amides developed for treatment of psychostimulant abuse.

The recent discovery of novel high-affinity and selective dopamine D3 receptor (DA D3R) antagonists and partial agonists has provided tools with which to further elucidate the role DA D3R plays in substance abuse. The present study was conducted to evaluate the transport, metabolism, pharmacokinetics, and brain uptake of the DA D3R-selective fluorenyl amides, NGB 2904 [N-(4-(4-(2,3-dichlorophenyl)piperazin-1-yl)butyl)-9H-fluorene-2-carboxamide] fumarate) and JJC 4-077 [N-(4-(4-(2,3-dichlorophenyl)piperazin-1-yl)-3-hydroxybutyl)-9H-fluorene-2-carboxamide hydrochloride], and the 2-pyridylphenyl amides, CJB 090 [N-(4-(4-(2,3-dichlorophenyl)piperazin-1-yl)butyl)-4-(pyridine-2-yl)benzamide hydrochloride] and PG 01037 [N-(4-(4-(2,3-dichlorophenyl)piperazin-1-yl)-trans-but-2-enyl)-4-(pyridine-2-yl)benzamide hydrochloride], all of which have been studied in animal models of psychostimulant abuse. Additional screening with a panel of human and rat Supersomes was performed for NGB 2904 and PG 01037. Drug-stimulated ATPase activation assays and bidirectional transport and efflux assays were used to test for substrate specificity of NGB 2904 and PG 01037 for human and rat efflux transporters. All compounds exhibited moderate elimination half-lives, ranging from 1.49 to 3.27 h, and large volumes of distribution (5.95-14.19 l/kg). The brain-to-plasma ratios ranged from 2.93 to 11.81 and were higher than those previously reported for cocaine. Brain exposure levels of NGB 2904 and PG 01037 were significantly reduced after intraperitoneal administration compared with intravenous administration. The metabolism of these compounds was mediated primarily by CYP3A subfamilies. PG 01037 was a P-glycoprotein-transported substrate. Higher doses of these compounds are often required for in vivo action, suggesting decreased bioavailability via extravascular administration that may be attributed to high drug efflux and hepatic metabolism. These studies provide important preclinical information for optimization of next-generation D3R selective agents for the treatment of drug addiction.

Regulation of gene expression in brain tissues of rats repeatedly treated by the highly abused opioid agonist, oxycodone: microarray profiling and gene mapping analysis.

Although oxycodone is the most often used opioid agonist, it remains one of the most understudied drugs. We used microarray analysis to better understand the global changes in gene expression in brain tissues of rats repeatedly treated with oxycodone. Many genes were significantly regulated by oxycodone (e.g., Fkbp5, Per2, Rt1.Dalpha, Slc16a1, and Abcg2). Validation of the microarray data by quantitative real-time-polymerase chain reaction (Q-PCR) indicated that there was a strong significant correlation (r = 0.979, p < 0.0000001) between the Q-PCR and the microarray data. Using MetaCore (a computational platform), many biological processes were identified [e.g., organic anion transport (p = 7.251 x 10(-4)) and regulation of immune response (p = 5.090 x 10(-4))]. Among the regulated genes, Abcg2 mRNA was up-regulated by 2.1-fold, which was further confirmed by immunoblotting (1.8-fold up-regulation). Testing the Abcg2 affinity status of oxycodone using an Abcg2 ATPase assay suggests that oxycodone behaves as an Abcg2 substrate only at higher concentrations (> or = 500 microM). Furthermore, brain uptake studies demonstrated that oxycodone-induced Abcg2 up-regulation resulted in a significant (p < 0.05) decrease (approximately 2-fold) in brain/plasma ratios of mitoxantrone. These results highlight markers/mediators of neuronal responses and identify regulatory pathways involved in the pharmacological action of oxycodone. These results also identify genes that potentially modulate tolerance, dependence, immune response, and drug-drug interactions. Finally, our findings suggest that oxycodone-induced up-regulation of Abcg2 enhanced the efflux of the Abcg2 substrate, mitoxantrone, limiting its brain accumulation and resulting in an undesirable drug-drug interaction. Extrapolating these results to other Abcg2 substrates (e.g., daunorubicin and doxorubicin) indicates that the brain uptake of these agents may be affected if they are administered concomitantly with oxycodone.

Trends in Research and Graduate Affairs in Schools and Colleges of Pharmacy, Part 3: Underrepresented Minorities.

Objective. To examine the landscape of research and graduate affairs nationally and within schools and colleges of pharmacy. This report, part 3 of a three-part series, focuses on underrepresented minority (URM) faculty members and students, with a focus on recruitment and retention. Findings. There has been a substantial increase in recruitment of Asian faculty members by schools of pharmacy over the last 10 years, but there has been only minimal changes in the numbers of Black and Hispanic faculty numbers, which reflects the challenges in recruitment and retention of URM faculty members. Consistently low enrollment of Black and Hispanic graduate students over a 10-year period demonstrates that pharmacy schools could improve their stated diversity initiatives and goals. Despite an overall increase in PhDs conferred over the last 10 years, international students continue to receive the majority of degrees conferred. Graduation rates of Black and Hispanic students have remained low, suggesting that continued and sustained efforts are needed to recruit, support, and graduate URM students. Summary. Pharmacy schools must make a focused investment and effort toward increasing the diversity of their graduate enrollees by modeling their recruitment, enrollment, and retention strategies after national programs and best practices. Because there is a direct link between the number of faculty role models and the recruitment of students, pharmacy schools must enhance the recruitment, retention, and success of URM faculty members. Further, pharmacy schools should provide inclusion training to encourage better communication with URM advisees.

Trends in Research and Graduate Affairs in Schools and Colleges of Pharmacy, Part 2: Students.

Objective. To examine the landscape of research and graduate affairs nationally and within schools and colleges of pharmacy. This report, part 2 of a three-part series, focuses on characteristics of full-time PhD enrollees and graduates in schools and colleges of pharmacy, and career planning and preparation in graduate programs. Findings. Despite a 41% increase in funding awarded by the National Institutes of Health (NIH) to schools and colleges of pharmacy over the last 10 years, NIH funding per principal investigator only increased 14% and graduate student enrollment increased just 6% during the period. However, there was a 15% increase in PhD degrees conferred in the 10-year period, which is evidence that degree completion time decreased. The number of female graduates from pharmacy schools consistently increased, and outpaced growth in the number of male graduates by more than 10%. Most graduate programs do not include training for industry-specific skills, abilities, and experiences to better prepare graduates for nonacademic careers, although national programs have been recognized as vital to graduate student career preparation. Summary. Graduate biomedical science programs and faculty members must recognize that academia is an "alternative" career choice for their trainees, and provide job skills training to support the majority of nonacademic career choices, without compromising the rigorous training in basic biomedical disciplines.

Trends in Research and Graduate Programs in Schools and Colleges of Pharmacy, Part 1: Programs.

Objective. To examine the landscape of research and graduate education nationally and within schools and colleges of pharmacy. This report is part 1 of a three-part series and focuses on graduate programs' research funding and science faculty composition and diversity. Findings. Between FY2008 and FY2017, the number of full-time faculty members in schools and colleges of pharmacy increased 36%. The number of pharmacy schools with National Institutes of Health (NIH) awards increased by 15%, while NIH grants per faculty principal investigator (PI) increased by 31%. However, unadjusted for inflation, the mean NIH dollar amount per-faculty member PI increased just 14% and the mean NIH dollar amount per-school declined 7%, indicating that number of funded faculty outpaced dollars available. Proportionately, the percentage of science faculty members at pharmacy schools decreased from 47% to 43%. Only 15 public, research-intensive schools and colleges of pharmacy received more than half of the combined FY2017 NIH funding and total funding, while all other public and private schools and colleges of pharmacy shared the remaining funds. Interdisciplinary programs are developing slowly, and may help to diversify and increase future funding. Proportions of tenured and tenure-track positions are declining, but biological sciences and social and administrative sciences disciplines are growing and women faculty are making significant gains in these fields and at the assistant professor rank. Summary. Research-intensive schools and colleges of pharmacy are best-positioned to lead the academy to reframe graduate education to build interdisciplinary team skills and attract more diverse funding and science faculty members.

Randomised trial of azithromycin to eradicate Ureaplasma in preterm infants.

OBJECTIVE: To test whether azithromycin eradicates Ureaplasma from the respiratory tract in preterm infants. DESIGN: Prospective, phase IIb randomised, double-blind, placebo-controlled trial. SETTING: Seven level III-IV US, academic, neonatal intensive care units (NICUs). PATIENTS: Infants 240-286 weeks' gestation (stratified 240-266; 270-286 weeks) randomly assigned within 4 days following birth from July 2013 to August 2016. INTERVENTIONS: Intravenous azithromycin 20 mg/kg or an equal volume of D5W (placebo) every 24 hours for 3 days. MAIN OUTCOME MEASURES: The primary efficacy outcome was Ureaplasma-free survival. Secondary outcomes were all-cause mortality, Ureaplasma clearance, physiological bronchopulmonary dysplasia (BPD) at 36 weeks' postmenstrual age, comorbidities of prematurity and duration of respiratory support. RESULTS: One hundred and twenty-one randomised participants (azithromycin: n=60; placebo: n=61) were included in the intent-to-treat analysis (mean gestational age 26.2±1.4 weeks). Forty-four of 121 participants (36%) were Ureaplasma positive (azithromycin: n=19; placebo: n=25). Ureaplasma-free survival was 55/60 (92% (95% CI 82% to 97%)) for azithromycin compared with 37/61 (61% (95% CI 48% to 73%)) for placebo. Mortality was similar comparing the two treatment groups (5/60 (8%) vs 6/61 (10%)). Azithromycin effectively eradicated Ureaplasma in all azithromycin-assigned colonised infants, but 21/25 (84%) Ureaplasma-colonised participants receiving placebo were culture positive at one or more follow-up timepoints. Most of the neonatal mortality and morbidity was concentrated in 21 infants with lower respiratory tract Ureaplasma colonisation. In a subgroup analysis, physiological BPD-free survival was 5/10 (50%) (95% CI 19% to 81%) among azithromycin-assigned infants with lower respiratory tract Ureaplasma colonisation versus 2/11 (18%) (95% CI 2% to 52%) in placebo-treated infants. CONCLUSION: A 3-day azithromycin regimen effectively eradicated respiratory tract Ureaplasma colonisation in this study. TRIAL REGISTRATION NUMBER: NCT01778634.

Methadone induces the expression of hepatic drug-metabolizing enzymes through the activation of pregnane X receptor and constitutive androstane receptor.

Methadone (MD) is the most established substance abuse pharmacotherapy of choice for the management of heroin dependence. To date, drug-drug interactions involving MD have been characterized asymmetrically among existing reports, which describe how other drugs affect the metabolic or pharmacokinetic profiles of MD; however, limited information is available regarding the potential for MD to influence similar fates of coadministered drugs. Moreover, little to no mechanistic evidence has been explored. Here, we show that MD induces hepatic drug-metabolizing enzymes (DMEs) through the activation of pregnane X receptor (PXR) and constitutive androstane receptor (CAR). Real-time polymerase chain reaction analysis of human hepatocyte cultures revealed that MD induces the mRNA expression of CYP2B6, CYP3A4, UGT1A1, and multidrug resistance 1 in a concentration-related manner, with the maximal induction of CYP2B6 challenging that of the induction by rifampicin. Furthermore, MD-mediated induction of CYP2B6 and CYP3A4 proteins was observed in Western blot analysis. In cell-based reporter assays, MD significantly increased human (h) PXR-mediated CYP2B6 reporter activities but exhibited minimal effect on hCAR activation as a result of the constitutive activity of hCAR in HepG2 cells. Further studies revealed that treatment with MD resulted in significant nuclear accumulation of adenovirus/enhanced yellow fluorescent protein tagged-hCAR in human hepatocytes, which has been regarded as the initial step of CAR activation. Additional analysis of the two enantiomers of MD, R-(-)-MD (active) and S-(+)-MD (inactive), indicates the lack of stereoselectivity pertaining to MD-mediated DME induction. Overall, our results show that MD induces the hepatic expression of multiple DMEs by activating PXR- and CAR-mediated pathways.

Estradiol and progesterone-mediated regulation of P-gp in P-gp overexpressing cells (NCI-ADR-RES) and placental cells (JAR).

The effect of progesterone and estrogen treatment on the expression and function of P-glycoprotein (P-gp) was evaluated in JAR cells and a P-gp overexpressing cell line, NCI-ADR-RES. Western blot analysis and real-time Q-PCR were used to evaluate P-gp protein and MDR1 mRNA expression respectively in the cells following incubation with progesterone (P4) and/or beta-estradiol (E2). Cellular uptake studies of the P-gp substrates, saquinavir and paclitaxel, were performed to evaluate function. Treatment with either E2 or P4 resulted in a significant increase in P-gp protein levels in the NCI-ADR-RES cells at concentrations of or greater than 100 nM or 10 nM, respectively. JAR cells also had increased levels of P-gp with 100 nM of P4 but were much more sensitive to E2 showing increased P-gp at a concentration of 1 nM. Furthermore, E2 or P4 treatment resulted in a significant decrease in cellular uptake of the P-gp substrates tested in these cells lines. Based on mRNA quantitation, a transient increase (2-fold) in MDR1 levels was observed at 8 h postincubation with either E2 or P4, while MDR1 levels remained high in the JAR cells treated with E2 for 72 h postincubation. The addition of actinomycin D, a transcription inhibitor negated the increase in P-gp by P4 and E2. P4 and E2 increase P-gp expression and function in NCI-ADR-RES and JAR cells with the ERalpha-expressing cells (JAR) much more sensitive to E2. Furthermore, transcriptional regulation by E2 and P4 likely contributes to the modulation of P-gp levels.

Trends in the Pharmacist Workforce and Pharmacy Education.

This commentary is an observation of longitudinal trends in national data on the pharmacist workforce and pharmacy education. Data indicate seismic shifts in supply and demand, from critical shortage to imminent oversupply. The change in the profession to employing more patient-care focused jobs has been observed as slow and minimal, although academia has focused on the clinical training and rapidly increased enrollments. Pharmacy is on the brink of transforming the profession, but several important changes are still required to alter the current trajectories of supply and demand. Pharmacy schools, associations, and employers must devote all energies to immediate and significant actions that tip the balance in favor of pharmacists of the future.

A Systematic Review of Entrepreneurship in Pharmacy Practice and Education.

Objective. To review literature pertaining to entrepreneurship in pharmacy practice, education, and the knowledge, skills, and attitudes (KSAs) identified for pharmacist entrepreneurs. Findings. In terms of pharmacy practice, entrepreneurship was most frequently identified with innovation and creativity to develop new opportunities for pharmacists. The most frequent role for entrepreneurship in pharmacy education was related to schools putting a greater emphasis on innovation, creativity, or divergent thinking. Risk-taking and creativity/innovation were the most frequently identified KSAs, with 17 (63.0%) manuscripts mentioning these as important for a pharmacist entrepreneur. Other KSAs pertaining to pharmacy entrepreneurship that were mentioned in the articles included self-starter, management, proactivity, communication, strategic planning, positivity, decision-making, teamwork, versatility, marketing, critical thinking, competitiveness, proposal development, numeracy, technology, self-reflection, persistence, social responsibility, and cultural competence. Summary. No consensus for entrepreneurship in pharmacy practice or education currently exists. In order to improve instructional design and assessment for pharmacy entrepreneurship education, a core set of KSAs for a pharmacist entrepreneur construct must be identified. The most commonly cited KSAs in related literature that are not already part of the Accreditation Council for Pharmacy Education standards include risk-taking, strategic planning, marketing, competitiveness, and social responsibility. These may serve as a starting point for enhancing pharmacy curricula to embrace pharmacist entrepreneurship.

Pharmapreneur - Defining a Framework for Entrepreneurship in Pharmacy Education.

Objective. To determine the role of entrepreneurism within the broader missions of schools of pharmacy and develop an educational framework to produce pharmacist entrepreneurs. Methods. Following a systematic review and six semi-structured interviews, a three-round Delphi process was conducted with an expert panel comprised of successful entrepreneurs, pharmacy faculty members and administrators, students, and community members. Participants were asked about the role of entrepreneurship in a pharmacy school's mission, how they would define a pharmacist entrepreneur, and to identify the knowledge, skills, and attitudes (KSAs) expected to be successful as a pharmacist entrepreneur. A model for entrepreneur education was also developed in accordance with Bloom's taxonomy. Participant agreement and rankings were reported. Results. Based on the semi-structured interviews and the results from the Delphi process, the following framework for a pharmacist entrepreneur was proposed along with a list of KSAs: identifies, creates, and pursues new opportunities; successfully implements new ideas into practice; is willing to take risks; fills unmet needs; creates new value through innovation; is responsive to change; makes sacrifices; includes social and intrapreneurship; leverages existing knowledge, skills, and resources; goes beyond traditional roles for pharmacists; and improves patient care. Recommendations for entrepreneurship instruction, guided by Bloom's taxonomy of cognitive processes, were created. Conclusion. According to our expert panel, a pharmacist entrepreneur combines several characteristics identified with a more traditional entrepreneur construct with the characteristics of an individual devoted to achieving outcomes beyond one's personal gain. Additional research to inform implementation and assessment of entrepreneurship within pharmacy curricula would provide more specific guidance for instructional design and accreditation evaluations.

Mechanism of 3,4-methylenedioxymethamphetamine (MDMA, ecstasy)-mediated mitochondrial dysfunction in rat liver.

Despite numerous reports citing the acute hepatotoxicity caused by 3,4-methylenedioxymethamphetamine (MDMA) (ecstasy), the underlying mechanism of organ damage is poorly understood. We hypothesized that key mitochondrial proteins are oxidatively modified and inactivated in MDMA-exposed tissues. The aim of this study was to identify and investigate the mechanism of inactivation of oxidatively modified mitochondrial proteins, prior to the extensive mitochondrial dysfunction and liver damage following MDMA exposure. MDMA-treated rats showed abnormal liver histology with significant elevation in plasma transaminases, nitric oxide synthase, and the level of hydrogen peroxide. Oxidatively modified mitochondrial proteins in control and MDMA-exposed rats were labeled with biotin-N-maleimide (biotin-NM) as a sensitive probe for oxidized proteins, purified with streptavidin-agarose, and resolved using 2-DE. Comparative 2-DE analysis of biotin-NM-labeled proteins revealed markedly increased levels of oxidatively modified proteins following MDMA exposure. Mass spectrometric analysis identified oxidatively modified mitochondrial proteins involved in energy supply, fat metabolism, antioxidant defense, and chaperone activities. Among these, the activities of mitochondrial aldehyde dehydrogenase, 3-ketoacyl-CoA thiolases, and ATP synthase were significantly inhibited following MDMA exposure. Our data show for the first time that MDMA causes the oxidative inactivation of key mitochondrial enzymes which most likely contributes to mitochondrial dysfunction and subsequent liver damage in MDMA-exposed animals.

Opioids and efflux transporters. Part 2: P-glycoprotein substrate activity of 3- and 6-substituted morphine analogs.

Continuing our studies investigating opioids with reduced P-glycoprotein (P-gp) substrate activity, a series of known 3- and 6-hydroxy, -methoxy, and -desoxymorphine analogs was synthesized and analyzed for P-gp substrate activity and opioid binding affinity. 6-Desoxymorphine ( 7) showed high affinity for opioid receptors and did not induce P-gp-mediated ATP hydrolysis. Additionally, 7 demonstrated morphine-like antinociceptive potency in mice, indicating this compound as an ideal lead to further evaluate the role of P-gp in opioid analgesic tolerance development.