Jun Sheng Wang

Two CES1 Gene Mutations Lead to Dysfunctional Carboxylesterase 1 Activity in Man: Clinical Significance and Molecular Basis

The human carboxylesterase 1 (CES1) gene encodes for the enzyme carboxylesterase 1, a serine esterase governing both metabolic deactivation and activation of numerous therapeutic agents. During the course of a study of the pharmacokinetics of the methyl ester racemic psychostimulant methylphenidate, profoundly elevated methylphenidate plasma concentrations, unprecedented distortions in isomer disposition, and increases in hemodynamic measures were observed in a subject of European descent. These observations led to a focused study of the subjects CES1 gene. DNA sequencing detected two coding region single-nucleotide mutations located in exons 4 and 6. The mutation in exon 4 is located in codon 143 and leads to a nonconservative substitution, p.Gly143Glu. A deletion in exon 6 at codon 260 results in a frameshift mutation, p.Asp260fs, altering residues 260-299 before truncating at a premature stop codon. The minor allele frequency of p.Gly143Glu was determined to be 3.7%, 4.3%, 2.0%, and 0% in white, black, Hispanic, and Asian populations, respectively. Of 925 individual DNA samples examined, none carried the p.Asp260fs, indicating it is an extremely rare mutation. In vitro functional studies demonstrated the catalytic functions of both p.Gly143Glu and p.Asp260fs are substantially impaired, resulting in a complete loss of hydrolytic activity toward methylphenidate. When a more sensitive esterase substrate, p-nitrophenyl acetate was utilized, only 21.4% and 0.6% catalytic efficiency (V(max)/K(m)) were determined in p.Gly143Glu and p.Asp260fs, respectively, compared to the wild-type enzyme. These findings indicate that specific CES1 gene variants can lead to clinically significant alterations in pharmacokinetics and drug response of carboxylesterase 1 substrates.

Risperidone and paliperidone inhibit p-glycoprotein activity in vitro.

Risperidone (RSP) and its major active metabolite, 9-hydroxy-risperidone (paliperidone, PALI), are substrates of the drug transporter P-glycoprotein (P-gp). The goal of this study was to examine the in vitro effects of RSP and PALI on P-gp-mediated transport. The intracellular accumulation of rhodamine123 (Rh123) and doxorubicin (DOX) were examined in LLC-PK1/MDR1 cells to evaluate P-gp inhibition by RSP and PALI. Both compounds significantly increased the intracellular accumulation of Rh123 and DOX in a concentration-dependent manner. The IC50 values of RSP for inhibiting P-gp-mediated transport of Rh123 and DOX were 63.26 and 15.78 μM, respectively, whereas the IC50 values of PALI were >100 μM, indicating that PALI is a less potent P-gp inhibitor. Caco-2 and primary cultured rat brain microvessel endothelial cells (RBMECs) were utilized to investigate the possible influence of RSP on intestinal absorption and blood–brain barrier (BBB) transport of coadministered drugs that are P-gp substrates. RSP, 1–50 μM, significantly enhanced the intracellular accumulation of Rh123 in Caco-2 cells by inhibiting P-gp activity with an IC50 value of 5.87 μM. Following exposure to 10 μM RSP, the apparent permeability coefficient of Rh123 across Caco-2 and RBMECs monolayers was increased to 2.02 and 2.63-fold in the apical to basolateral direction, but decreased to 0.37 and 0.21-fold in the basolateral to apical direction, respectively. These data suggest that RSP and PALI, to a lesser extent, have a potential to influence the pharmacokinetics and hence the pharmacodynamics of coadministered drugs via inhibition of P-gp-mediated transport. However, no human data exist that address this issue. In particular, RSP may interact with its own active metabolite PALI by promoting its brain concentration through inhibiting P-gp-mediated efflux of PALI across endothelial cells of the BBB.

Characterization of P-glycoprotein inhibition by major cannabinoids from marijuana

The ATP-dependent drug efflux transporter P-glycoprotein (P-gp) plays a significant role in the absorption and disposition of many compounds. The purpose of this study was to investigate the possible interaction of P-gp with each of four major mari-juana constituents: Δ9-tetrahydrocannabinol (THC), 11-nor-Δ9-tetrahydrocannabinol-carboxylic acid (THC-COOH), cannabinol (CBN), and cannabidiol (CBD). The results of a P-gp ATPase activity screen showed that THC-COOH, CBN, THC, and CBD all stimulated P-gp ATPase activity with a Michaelis-Menten parameter (Vmax/Km) value of 1.3, 0.7, 0.1, and 0.05, respectively. Furthermore, CBD showed a concentration-dependent inhibitory effect on verapamil-stimulated ATPase activity with an IC50 value of 39.6 μM, whereas all other tested cannabinoids did not display appreciable inhibitory effects. Thus, the inhibitory effects of CBD on P-gp transport were further studied. At concentrations ranging from 5 to 100 μM, CBD robustly enhanced the intracellular accumulation of known P-gp substrates rhodamine 123 and doxorubicin in a concentration-dependent manner in Caco-2 and LLC-PK1/MDR1 cells. An IC50 value of 8.44 μM was obtained for inhibition of P-gp function in LLC-PK1/MDR1 cells as determined by flow cytometry using rhodamine 123 as a fluorescence probe. Following exposure to 30 μM CBD, the apparent permeability coefficient of rhodamine 123 across Caco-2 and rat brain microvessel endothelial cell monolayers was increased to 2.2- and 2.6-fold in the apical-to-basolateral direction but decreased to 0.69- and 0.47-fold in the basolateral-to-apical direction, respectively. These findings indicate that CBD significantly inhibits P-gp-mediated drug transport, suggesting CBD could potentially influence the absorption and disposition of other coadministered compounds that are P-gp substrates.

Therapeutic drug monitoring of psychoactive drugs during pregnancy in the genomic era: challenges and opportunities

Various symptoms of mental illness occur commonly during pregnancy. It is estimated that serious mental disorders, including major depression, bipolar disorder, schizophrenia, panic and other anxiety disorders, occur with a frequency of 10 to 25% in community samples of US women in their child-bearing years. As a result, approximately a third of all women take at least one psychoactive drug during pregnancy. Fetal drug exposure has been documented for all psychoactive drugs studied to date. However, the rate and extent of placental transfer within and between psychoactive drug classes remains ill defined. The contribution of various genetic factors such as the role of polymorphic drug metabolizing enzymes and drug transporters in controlling the variability of fetal drug exposure is also unclear. Therapeutic drug monitoring (TDM) has traditionally played an important role in psychiatric pharmacotherapy during pregnancy to ensure an adequate drug dose to achieve desired benefits while avoiding excessive fetal accumulation for drugs. In the genomic era, individualized treatment with specific drugs tailored to the mothers and fetuss genotype should eventually become the standard of care. Several methodological problems need to be overcome for this prediction to become reality. One approach to this goal taken by the Specialized Center of Research on Sex and Gender Factors Affecting Womens Health at the Emory University Womens Mental Health Program is described. This research is grounded on TDM of pregnant women receiving antidepressants, antipsychotics, anti-epileptic drugs and mood stabilizers. The use of pharmacokinetic and pharmacogenetic models to predict maternal plasma drug concentrations, fetal drug exposure, and maternal and neonatal outcomes, is expected to improve our understanding of dose-response relationships of psychoactive drugs in pregnancy.

Pharmacokinetics of Olanzapine After Single‐Dose Oral Administration of Standard Tablet Versus Normal and Sublingual Administration of an Orally Disintegrating Tablet in Normal Volunteers

Olanzapine (OLZ) is a second‐generation antipsychotic agent available in 2 solid oral dosage forms, a standard oral tablet (SOT) and an orally disintegrating tablet (ODT). This study assessed the absorption of each by different routes of administration. Secondarily, the influence of P‐glycoprotein (P‐gp) genotype was assessed. It was hypothesized that more rapid absorption of the OLZ ODT would occur when administered sublingually versus standard oral administration. A randomized, 3‐way crossover study assessed the 5‐mg OLZ formulations in healthy volunteers (n = 10). Blood was collected (0–8 hours) to assess OLZ pharmacokinetics using liquid chromatography/mass spectrometry. Both routes of ODT administration resulted in more measurable early concentraions relative to SOT. However, there were no statistically significant differences observed between any of the OLZ exposures for observed pharmacokinetic parameters (Cmax, Tmax, AUC0–8h). The homozygous TT genotype for P‐gp resulted in an increased AUC of OLZ for SOT administration but not for either condition where sublingual absorption could occur.

Antipsychotic Drugs Inhibit the Function of Breast Cancer Resistance Protein

The ABCG2 transporter breast cancer resistance protein (BCRP) has been identified in several physiological sites. It has been suggested to play an important role in disposition of many drugs and environmental toxins. We investigated the effects of several antipsychotic drugs, including risperidone, 9-hydroxy-risperidone (paliperidone), olanzapine, quetiapine, clozapine, haloperidol and chlorpromazine, and a positive control inhibitor Ko143 on functions of BCRP in MCF7 and BCRP over-expressing MCF7/MX100 cell lines using a BCRP prototypical substrate mitoxantrone. Our findings indicated that the tested antipsychotics rank order of potency of inhibition of BCRP according to concentrations required to reach 50% of maximum inhibition (IC(50)) was as follows: Ko143 (0.07 microM) > risperidone (38.1 microM) > clozapine (42.0 microM) > paliperidone (51 microM) > chlorpromazine (52.2 microM) > quetiapine (66.1 microM) > olanzapine = haloperidol (>100.0 microM). We further tested the effects of various concentrations of risperidone on the BCRP-mediated transport of oestrone-3-sulfate in a colon carcinoma cell line, Caco-2, a widely used model to study drug absorption. Our findings show that risperidone at concentrations ranging from 1 to 100 microM significantly inhibited intracellular accumulation of oestrone-3-sulfate in Caco-2 cell monolayers. The present results suggest that a potential source of pharmacokinetic interactions exists between BCRP substrates and several antipsychotics.

Aripiprazole brain concentration is altered in P-glycoprotein deficient mice

P-glycoprotein (P-gp) is a transporter that mediates the tissue disposition of numerous drugs. To evaluate the role of P-glycoprotein (P-gp) in aripiprazole tissue distribution and penetration across the blood-brain barrier, mice deficient in the P-gp gene (Abcb1a/b-/-) were dosed intraperitoneally with 2 microg/g mouse of the antipsychotic drug aripiprazole. Wildtype FVB mice were administered the same dose as transgenic animals. At one, two, and three hours after dosing, blood and tissue samples were collected and assayed for aripiprazole concentration by HPLC. Deficiency of P-gp did not result in significantly altered plasma drug concentrations but had dramatic effects on drug concentrations in brain tissue. At 1, 2, and 3 h after dosing, aripiprazole brain concentrations in the Abcb1a/b-/- mice were 4.6-, 4.1- and 3.0-fold higher, respectively (P<0.01), compared with the wildtype mice. Increases in drug concentration were also observed in testes and muscle in Abcb1a/b -/- mice. All other tissues including gut, lung, heart, kidney, liver, and spleen did not show significant differences between the two groups. These data provide evidence that aripiprazole is a transportable substrate of P-gp. Thus, factors influencing P-gp activity within the blood brain barrier in humans may have implications for the therapeutic effects and tolerability of aripiprazole.

The emerging importance of transporter proteins in the psychopharmacological treatment of the pregnant patient

P-glycoprotein, breast cancer resistance protein, and multidrug resistance proteins have physiological functions in placental tissue. Several antidepressants, antipsychotics, and anti-epileptic drugs have been found to be substrates of P-glycoprotein and other transporters. The extent that drugs pass through the placental barrier is likely influenced by drug transporters. The rational choice of psychoactive drugs to treat mental illness in women of child-bearing age should incorporate knowledge of both drug disposition as well as expected pharmacologic effects. This review summarizes the current data on drug transporters in the placental passage of medications, with a focus on medications used in clinical psychopharmacology.

The role of the polymorphic efflux transporter P-glycoprotein on the brain accumulation of d-methylphenidate and d-amphetamine

The psychostimulant medications methylphenidate (MPH) and amphetamine (AMP), available in various ratios or enantiopure formulations of their respective active dextrorotary isomers, constitute the majority of agents used in the treatment of attention-deficit/hyperactivity disorder (ADHD). Substantial interindividual variability occurs in their pharmacokinetics and tolerability. Little is known regarding the potential role of drug transporters such as P-glycoprotein (P-gp) in psychostimulant pharmacokinetics and response. Therefore, experiments were carried out in P-gp knockout (KO) mice versus wild-type (WT) mice after intraperitoneal dosing (2.5 mg/kg) of d-MPH or (3.0 mg/kg) of d-AMP. After the administration of each psychostimulant, locomotor activity was assessed at 30-min intervals for 2 h. Total brain-to-plasma drug concentration ratios were determined at 10-, 30-, and 80-min postdosing time-points. The results showed no statistically supported genotypic difference in d-AMP-induced locomotor activity stimulation or in brain-to-plasma ratio of d-AMP. As for d-MPH, the P-gp KO mice had 33% higher brain concentrations (p < 0.05) and 67.5% higher brain-to-plasma ratios (p < 0.01) than WT controls at the 10-min postdosing timepoint. However, in spite of elevated brain concentrations, d-MPH-induced locomotor activity increase was attenuated for P-gp compared with that for WT mice. These data indicate that P-gp has no apparent effect on the pharmacokinetics and pharmacodynamics of d-AMP. In addition, d-MPH is a relatively weak P-gp substrate, and its entry into the brain may be limited by P-gp. Furthermore, the mechanism by which d-MPH-induced locomotor activity was attenuated in P-gp KO mice remains to be elucidated.