32 results on '"Vuppugalla R"'
Search Results
2. Characterization of BMS-911543, a functionally selective small-molecule inhibitor of JAK2
- Author
-
Purandare, A V, primary, McDevitt, T M, additional, Wan, H, additional, You, D, additional, Penhallow, B, additional, Han, X, additional, Vuppugalla, R, additional, Zhang, Y, additional, Ruepp, S U, additional, Trainor, G L, additional, Lombardo, L, additional, Pedicord, D, additional, Gottardis, M M, additional, Ross-Macdonald, P, additional, de Silva, H, additional, Hosbach, J, additional, Emanuel, S L, additional, Blat, Y, additional, Fitzpatrick, E, additional, Taylor, T L, additional, McIntyre, K W, additional, Michaud, E, additional, Mulligan, C, additional, Lee, F Y, additional, Woolfson, A, additional, Lasho, T L, additional, Pardanani, A, additional, Tefferi, A, additional, and Lorenzi, M V, additional
- Published
- 2011
- Full Text
- View/download PDF
3. A-195 Preliminary Results from an Ongoing Phase 2, Open-Label, Multicenter, Single-Arm Study Assessing an Every-4-Week Dosing Schedule of Mogamulizumab in Patients with Cutaneous T-Cell Lymphoma.
- Author
-
Scarisbrick, J., Querfeld, C., Akilov, O., Bagot, M., Córdoba, R., Cowan, R., García -Sancho, A.M., Geskin, L.J., Huen, A.O., Jadwani, J., Liu, Y., Zhao, H., Morris, S., Ortiz-Romero, P.L., Patel, A., Pinter-Brown, L.C., Pujol, R.M., Quaglino, P., Saba, N.S., and Vuppugalla, R.
- Subjects
- *
THERAPEUTIC use of antineoplastic agents , *THERAPEUTIC use of monoclonal antibodies , *ANTINEOPLASTIC agents , *DRUG administration , *CONFERENCES & conventions , *CUTANEOUS T-cell lymphoma , *MONOCLONAL antibodies - Published
- 2024
- Full Text
- View/download PDF
4. Prospective prediction of plasma pharmacokinetics of a novel immune-modulating agent in cancer patients after intra-tumoral administration: translation from non-clinical species to humans.
- Author
-
Vuppugalla R, Sane R, Wichroski M, Gavai AK, Boyanapalli S, and Yang Z
- Subjects
- Biological Availability, Humans, Pharmacokinetics, Prospective Studies, Antineoplastic Agents, Neoplasms drug therapy, Pharmaceutical Preparations
- Abstract
Intra-tumoral (I-TUMOUR) delivery is being widely explored for novel anti-cancer agents. This route is anticipated to result in high tumour concentrations leading to better efficacy and safety. Prediction of human systemic pharmacokinetics (PK) from non-clinical species facilitates understanding of pharmacokinetic-pharmacodynamic relationships, efficient dose selection, and risk assessment of novel drugs. However, there is limited knowledge on the predictability of human pharmacokinetics following I-TUMOUR delivery.In this publication, we present a case study wherein human systemic PK of a novel agent administered intra-tumourally was prospectively predicted and compared with observed human PK.Simple allometry was used to project the human clearance (10.5 mL/min/kg) and steady-state volume of distribution (1.4 L/kg) after intravenous (IV) dosing. Using these IV PK parameters and assuming rapid absorption and complete I-TUMOUR bioavailability, human plasma PK profile was simulated. The projected 30 min concentrations and AUC
(0-6h) were within 1.9 to 2.5-fold and 1 to 1.4-fold of the observed PK indicating a reasonable concordance between predicted and observed PK.To our knowledge, this is the first article that prospectively projected human pharmacokinetics after I-TUMOUR dosing. The results from this study indicate that similar approaches can be used to project the human PK of other I-TUMOUR agents.- Published
- 2021
- Full Text
- View/download PDF
5. BMS-813160: A Potent CCR2 and CCR5 Dual Antagonist Selected as a Clinical Candidate.
- Author
-
Cherney RJ, Anjanappa P, Selvakumar K, Batt DG, Brown GD, Rose AV, Vuppugalla R, Chen J, Pang J, Xu S, Yarde M, Tebben AJ, Paidi VR, Cvijic ME, Mathur A, Barrish JC, Mandlekar S, Zhao Q, and Carter PH
- Abstract
BMS-813160 (compound 3 ) was identified as a potent and selective CCR2/5 dual antagonist. Compound 3 displayed good permeability at pH = 7.4 in PAMPA experiments and demonstrated excellent human liver microsome stability. Pharmacokinetic studies established that 3 had excellent oral bioavailability and exhibited low clearance in dog and cyno. Compound 3 was also studied in the mouse thioglycollate-induced peritonitis model, which confirmed its ability to inhibit the migration of inflammatory monocytes and macrophages. As a result of this profile, compound 3 was selected as a clinical candidate., Competing Interests: The authors declare no competing financial interest., (© 2021 American Chemical Society.)
- Published
- 2021
- Full Text
- View/download PDF
6. Discovery of BMS-753426: A Potent Orally Bioavailable Antagonist of CC Chemokine Receptor 2.
- Author
-
Yang MG, Xiao Z, Zhao R, Tebben AJ, Wang B, Cherney RJ, Batt DG, Brown GD, Cvijic ME, Duncia JV, Gallela MA, Gardner DS, Khandelwal P, Malley MF, Pang J, Rose AV, Santella JB 3rd, Sarjeant AA, Xu S, Mathur A, Mandlekar S, Vuppugalla R, Zhao Q, and Carter PH
- Abstract
To improve the metabolic stability profile of BMS-741672 ( 1a ), we undertook a structure-activity relationship study in our trisubstituted cyclohexylamine series. This ultimately led to the identification of 2d (BMS-753426) as a potent and orally bioavailable antagonist of CCR2. Compared to previous clinical candidate 1a , the tert -butyl amine 2d showed significant improvements in pharmacokinetic properties, with lower clearance and higher oral bioavailability. Furthermore, compound 2d exhibited improved affinity for CCR5 and good activity in models of both monocyte migration and multiple sclerosis in the hCCR2 knock-in mouse. The synthesis of 2d was facilitated by the development of a simplified approach to key intermediate (4 R )- 9b that deployed a stereoselective reductive amination which may prove to be of general interest., Competing Interests: The authors declare no competing financial interest., (© 2021 American Chemical Society.)
- Published
- 2021
- Full Text
- View/download PDF
7. Quantification of in vivo site-specific Asp isomerization and Asn deamidation of mAbs in animal serum using IP-LC-MS.
- Author
-
Mehl JT, Sleczka BG, Ciccimaro EF, Kozhich AT, Gilbertson DG, Vuppugalla R, Huang CS, Stevens B, Mo J, Deyanova EG, Wang Y, Huang RY, Chen G, and Olah TV
- Subjects
- Amino Acid Sequence, Animals, Antibodies, Monoclonal blood, Chromatography, Liquid methods, Humans, Immunoprecipitation methods, Isomerism, Macaca fascicularis, Male, Tandem Mass Spectrometry methods, Antibodies, Monoclonal chemistry, Asparagine analysis, Aspartic Acid analysis
- Abstract
Background: Isomerization of aspartic acid and deamidation of asparagine are two common amino acid modifications that are of particular concern if located within the complementarity-determining region of therapeutic antibodies. Questions arise as to the extent of modification occurring in circulation due to potential exposure of the therapeutic antibody to different pH regimes., Results: To enable evaluation of site-specific isomerization and deamidation of human mAbs in vivo, immunoprecipitation (IP) has been combined with LC-MS providing selective enrichment, separation and detection of naive and modified forms of tryptic peptides comprising complementarity-determining region sequences., Conclusion: IP-LC-MS can be applied to simultaneously quantify in vivo drug concentrations and measure the extent of isomerization or deamidation in PK studies conducted during the drug discovery stage.
- Published
- 2016
- Full Text
- View/download PDF
8. Ultrasensitive quantitative LC-MS/MS of an inhibitor of apoptosis protein's antagonist in plasma using protein target affinity extraction.
- Author
-
Discenza LN, Cornelius G, Gan J, Szapiel N, Talbott RL, Chaudhry C, Roy A, Borzilleri RM, Vuppugalla R, Stefanski K, Moore R, D'Arienzo CJ, Olah TV, and Mehl JT
- Subjects
- Animals, Dogs, Female, Humans, Immobilized Proteins antagonists & inhibitors, Immobilized Proteins chemistry, Inhibitor of Apoptosis Proteins chemistry, Isoquinolines chemistry, Isoquinolines pharmacology, Male, Oligopeptides chemistry, Oligopeptides pharmacology, Small Molecule Libraries chemistry, Small Molecule Libraries pharmacology, Blood Chemical Analysis methods, Chemical Fractionation methods, Chromatography, Liquid methods, Inhibitor of Apoptosis Proteins antagonists & inhibitors, Isoquinolines analysis, Limit of Detection, Oligopeptides analysis, Small Molecule Libraries analysis, Tandem Mass Spectrometry methods
- Abstract
Background: A target protein-based affinity extraction LC-MS/MS method was developed to enable plasma level determination following ultralow dosing (0.1-3 µg/kg) of an inhibitor of apoptosis proteins molecule. Methodology & results: Affinity extraction (AE) utilizing immobilized target protein BIR2/BIR3 was used to selectively capture the inhibitor of apoptosis proteins molecule from dog plasma and enable removal of background matrix components. Pretreatment of plasma samples using protein precipitation was found to provide an additional sensitivity gain. A LLOQ of 7.8 pM was achieved by combining protein precipitation with AE. The method was used to support an ultralow dose dog toxicity study., Conclusion: AE-LC-MS/MS, utilizing target protein, is a highly sensitive methodology for small molecule quantification with potential for broader applicability.
- Published
- 2016
- Full Text
- View/download PDF
9. Pharmacology of smac mimetics; chemotype differentiation based on physical association with caspase regulators and cellular transport.
- Author
-
Talbott RL, Borzilleri RM, Chaudhry C, Fargnoli J, Shen H, Fairchild C, Barnhart B, Ortega M, McDonagh TE, Vuppugalla R, Vite GD, Hunt JT, Gottardis M, and Naglich JG
- Subjects
- ATP Binding Cassette Transporter, Subfamily B metabolism, Animals, Apoptosis drug effects, Apoptosis Regulatory Proteins, Biomimetics methods, Cell Differentiation drug effects, Cell Line, Tumor, Cell Proliferation drug effects, Female, HCT116 Cells, Humans, Inhibitor of Apoptosis Proteins antagonists & inhibitors, Melanoma drug therapy, Melanoma metabolism, Mice, Inbred BALB C, Mice, Nude, Protein Binding drug effects, Protein Structure, Tertiary drug effects, Antineoplastic Agents pharmacology, Biological Transport drug effects, Caspase 3 metabolism, Intracellular Signaling Peptides and Proteins antagonists & inhibitors, Mitochondrial Proteins antagonists & inhibitors
- Abstract
Cellular levels of inhibitor of apoptosis (IAP) proteins are elevated in multiple human cancers and their activities often play a part in promoting cancer cell survival by blocking apoptotic pathways, controlling signal transduction pathways and contributing to resistance. These proteins function through interactions of their BIR (baculoviral IAP repeat) protein domains with pathway components and these interactions are endogenously antagonized by Smac/Diablo (second mitochondrial activator of caspases/direct IAP binding protein with low isoelectric point). This report describes development of synthetic smac mimetics (SM) and compares their binding, antiproliferative and anti-tumor activities. All dimeric antagonists inhibit in vitro smac tetrapeptide binding to recombinant IAP proteins, rescue IAP-bound caspase-3 activity and show anti-proliferative activity against human A875 melanoma cells. One heterodimeric SM, SM3, binds tightly to IAP proteins in vitro and slowly dissociates (greater than two hours) from these protein complexes compared to the other antagonists. In addition, in vitro SM anti-proliferation potency is influenced by ABCB1 transporter (ATP-binding cassette, sub-family B; MDR1, P-gp) activities and one antagonist, SM5, does not appear to be an ABCB1 efflux pump substrate. All dimeric smac mimetics inhibit the growth of human melanoma A875 tumors implanted in athymic mice at well-tolerated doses. One antagonist, SM4, shows broad spectrum in vivo anti-tumor activity and modulates known pharmacodynamic markers of IAP antagonism. These data taken together demonstrate the range of diverse dimeric IAP antagonist activities and supports their potential as anticancer agents., (Copyright © 2015 Elsevier Inc. All rights reserved.)
- Published
- 2015
- Full Text
- View/download PDF
10. Comparison of physiologic and pharmacologic parameters in Asian and mauritius cynomolgus macaques.
- Author
-
Kozlosky JC, Mysore J, Clark SP, Burr HN, Li J, Aranibar N, Vuppugalla R, West RC, Mangipudy RS, and Graziano MJ
- Subjects
- Animals, Asian People, Female, Humans, Male, Mauritius, Macaca fascicularis physiology, Organ Size physiology
- Abstract
This comparative study was conducted to assess background physiologic and pharmacologic parameters of cynomolgus macaques (Macaca fascicularis) from Cambodia, from a mixed Asian source (Cambodia, Vietnam and Indonesia), and from Mauritius. This evaluation provides a comprehensive assessment of several of these parameters in a single study. Ten male and 10 female captive-bred, age-matched macaques from each source were evaluated. Criteria for evaluation included weight gain, assessment of drug metabolizing enzyme activity, metabolomic analysis, immunologic assessments (lymphocyte subsets, TDAR, and serum Ig isotyping), clinical pathology evaluations, physical (respiratory, neurologic, cardiovascular, and ophthalmologic) examinations, pathogen screening, organ weights, and gross and microscopic pathology analyses. The results of this evaluation indicate that, compared to macaques of Asian origin, macaques from Mauritius had the lowest incidence and/or severity of spontaneous pathologic findings in several organs and tissues (lymphoid organs, stomach, kidney, urothelium, heart, arteries and lung) and better testicular maturity at a given age with minimal variability in organ weights. Although slight differences were observed in other parameters, none were considered detrimental to the use of macaques of Asian or Mauritius origin in pharmaceutical candidate safety studies with the use of a consistent source, concomitant controls, and appropriate background knowledge and screening., (Copyright © 2015 Elsevier Inc. All rights reserved.)
- Published
- 2015
- Full Text
- View/download PDF
11. Discovery of a Highly Selective JAK2 Inhibitor, BMS-911543, for the Treatment of Myeloproliferative Neoplasms.
- Author
-
Wan H, Schroeder GM, Hart AC, Inghrim J, Grebinski J, Tokarski JS, Lorenzi MV, You D, Mcdevitt T, Penhallow B, Vuppugalla R, Zhang Y, Gu X, Iyer R, Lombardo LJ, Trainor GL, Ruepp S, Lippy J, Blat Y, Sack JS, Khan JA, Stefanski K, Sleczka B, Mathur A, Sun JH, Wong MK, Wu DR, Li P, Gupta A, Arunachalam PN, Pragalathan B, Narayanan S, K C N, Kuppusamy P, and Purandare AV
- Abstract
JAK2 kinase inhibitors are a promising new class of agents for the treatment of myeloproliferative neoplasms and have potential for the treatment of other diseases possessing a deregulated JAK2-STAT pathway. X-ray structure and ADME guided refinement of C-4 heterocycles to address metabolic liability present in dialkylthiazole 1 led to the discovery of a clinical candidate, BMS-911543 (11), with excellent kinome selectivity, in vivo PD activity, and safety profile.
- Published
- 2015
- Full Text
- View/download PDF
12. Structure-Based Design of Selective Janus Kinase 2 Imidazo[4,5-d]pyrrolo[2,3-b]pyridine Inhibitors.
- Author
-
Hart AC, Schroeder GM, Wan H, Grebinski J, Inghrim J, Kempson J, Guo J, Pitts WJ, Tokarski JS, Sack JS, Khan JA, Lippy J, Lorenzi MV, You D, McDevitt T, Vuppugalla R, Zhang Y, Lombardo LJ, Trainor GL, and Purandare AV
- Abstract
Early hit to lead work on a pyrrolopyridine chemotype provided access to compounds with biochemical and cellular potency against Janus kinase 2 (JAK2). Structure-based drug design along the extended hinge region of JAK2 led to the identification of an important H-bond interaction with the side chain of Tyr 931, which improved JAK family selectivity. The 4,5-dimethyl thiazole analogue 18 demonstrated high levels of JAK family selectivity and was identified as a promising lead for the program.
- Published
- 2015
- Full Text
- View/download PDF
13. Dimeric Macrocyclic Antagonists of Inhibitor of Apoptosis Proteins for the Treatment of Cancer.
- Author
-
Zhang Y, Seigal BA, Terrett NK, Talbott RL, Fargnoli J, Naglich JG, Chaudhry C, Posy SL, Vuppugalla R, Cornelius G, Lei M, Wang C, Zhang Y, Schmidt RJ, Wei DD, Miller MM, Allen MP, Li L, Carter PH, Vite GD, and Borzilleri RM
- Abstract
A series of dimeric macrocyclic compounds were prepared and evaluated as antagonists for inhibitor of apoptosis proteins. The most potent analogue 11, which binds to XIAP and c-IAP proteins with high affinity and induces caspase-3 activation and ultimately cell apoptosis, inhibits growth of human melanoma and colorectal cell lines at low nanomolar concentrations. Furthermore, compound 11 demonstrated significant antitumor activity in the A875 human melanoma xenograft model at doses as low as 2 mg/kg on a q3d schedule.
- Published
- 2015
- Full Text
- View/download PDF
14. Discovery of potent heterodimeric antagonists of inhibitor of apoptosis proteins (IAPs) with sustained antitumor activity.
- Author
-
Perez HL, Chaudhry C, Emanuel SL, Fanslau C, Fargnoli J, Gan J, Kim KS, Lei M, Naglich JG, Traeger SC, Vuppugalla R, Wei DD, Vite GD, Talbott RL, and Borzilleri RM
- Subjects
- Animals, Antineoplastic Agents chemical synthesis, Antineoplastic Agents chemistry, Cell Line, Tumor, Cell Proliferation drug effects, Dose-Response Relationship, Drug, Drug Screening Assays, Antitumor, Humans, Mice, Molecular Structure, Neoplasms, Experimental pathology, Proline chemical synthesis, Proline chemistry, Structure-Activity Relationship, Antineoplastic Agents pharmacology, Drug Discovery, Inhibitor of Apoptosis Proteins antagonists & inhibitors, Neoplasms, Experimental drug therapy, Proline pharmacology
- Abstract
The prominent role of IAPs in controlling cell death and their overexpression in a variety of cancers has prompted the development of IAP antagonists as potential antitumor therapies. We describe the identification of a series of heterodimeric antagonists with highly potent antiproliferative activities in cIAP- and XIAP-dependent cell lines. Compounds 15 and 17 further demonstrate curative efficacy in human melanoma and lung cancer xenograft models and are promising candidates for advanced studies.
- Published
- 2015
- Full Text
- View/download PDF
15. Quantification of human mAbs in mouse tissues using generic affinity enrichment procedures and LC-MS detection.
- Author
-
Sleczka BG, Mehl JT, Shuster DJ, Lewis KE, Moore R, Vuppugalla R, Rajendran S, D'Arienzo CJ, and Olah TV
- Subjects
- Amino Acid Sequence, Animals, Antibodies, Monoclonal blood, Antibodies, Monoclonal metabolism, Antibodies, Monoclonal, Humanized analysis, Antibodies, Monoclonal, Humanized blood, Antibodies, Monoclonal, Humanized metabolism, Chromatography, Affinity, Female, Humans, Liver metabolism, Mice, Mice, Inbred C57BL, Molecular Sequence Data, Peptides analysis, Regression Analysis, Skin metabolism, Trypsin metabolism, Ustekinumab, Antibodies, Monoclonal analysis, Chromatography, High Pressure Liquid, Tandem Mass Spectrometry
- Abstract
Background: The disease state can modulate the penetration of large antibody-sized therapeutic molecules into affected tissues. Suitable bioanalytical methods are required for the quantitative analysis of drug tissue levels to enable a better understanding of the parameters influencing drug penetration and target engagement., Results: Described is a sensitive and selective LC-MS/MS assay for the quantification of human mAb molecules in mouse tissues. By homogenizing tissues directly into serum, a common serum calibration curve can be used for multiple tissues. A generic procedure was used for affinity enrichment. An analytical range of 20 - 20,000 ng/ml was achieved in serum., Conclusion: The method described here can be applied for the quantitative analysis of mAb and Fc-fusion therapeutic molecules in a variety of animal tissue matrices.
- Published
- 2014
- Full Text
- View/download PDF
16. Impact of nonlinear midazolam pharmacokinetics on the magnitude of the midazolam-ketoconazole interaction in rats.
- Author
-
Vuppugalla R, Zhang Y, Chang S, Rodrigues AD, and Marathe PH
- Subjects
- Administration, Intravenous, Administration, Oral, Animals, Blood Proteins metabolism, Chromatography, Liquid, Computer Simulation, Drug Interactions, Humans, Hypnotics and Sedatives administration & dosage, Male, Mass Spectrometry, Microsomes, Liver metabolism, Midazolam administration & dosage, Models, Chemical, Rats, Rats, Sprague-Dawley, Antifungal Agents pharmacology, Hypnotics and Sedatives pharmacokinetics, Ketoconazole pharmacology, Midazolam pharmacokinetics
- Abstract
Numerous groups have described the rat as an in vivo model for the assessment and prediction of drug-drug interactions (DDIs) in humans involving the inhibition of cytochrome P450 3A forms. Even for a well-established substrate-inhibitor pair like midazolam-ketoconazole, however, the magnitude of the DDI in rats (e.g. 1.5- to 5-fold) does not relate to what is observed clinically (e.g. 5- to 16-fold). Because nonlinear substrate pharmacokinetics (PK) may result in a weaker interaction, it was hypothesized that the lower magnitude of interaction observed in rats was due to the saturation of metabolic pathway(s) of midazolam at the doses used (10-20 mg/kg). Therefore, the inhibitory effects of ketoconazole were reevaluated at lower oral (1 and 5 mg/kg) and intravenous (IV) (1 mg/kg) doses of midazolam. In support of the hypothesis, oral exposure at 5 mg/kg dose of midazolam was 18-fold higher compared to that at 1 mg/kg. Furthermore, when the interaction was investigated at the lower midazolam dose (1 mg/kg), ketoconazole increased the IV and oral exposure of midazolam by 7-fold and 11-fold, respectively. A weaker DDI (1.5- to 1.8-fold) was observed at the higher oral midazolam dose. Collectively, these results suggest that the lower reported interaction in rats is likely due to saturation of midazolam clearance at the doses used. Therefore, when the rat is used as a DDI model to screen and differentiate compounds, or predict CYP3A inhibition in humans, it is important to use low doses of midazolam and ensure linear PK.
- Published
- 2012
- Full Text
- View/download PDF
17. Evaluation of six proton pump inhibitors as inhibitors of various human cytochromes P450: focus on cytochrome P450 2C19.
- Author
-
Zvyaga T, Chang SY, Chen C, Yang Z, Vuppugalla R, Hurley J, Thorndike D, Wagner A, Chimalakonda A, and Rodrigues AD
- Subjects
- 2-Pyridinylmethylsulfinylbenzimidazoles pharmacology, Aryl Hydrocarbon Hydroxylases metabolism, Biotransformation, Cells, Cultured, Computer Simulation, Cytochrome P-450 CYP2C19, Dealkylation, Dexlansoprazole, Diazepam metabolism, Dose-Response Relationship, Drug, Drug Interactions, Esomeprazole pharmacokinetics, Hepatocytes drug effects, Hepatocytes enzymology, Humans, Kinetics, Lansoprazole, Liver enzymology, Microsomes, Liver enzymology, Models, Biological, NADP metabolism, Omeprazole pharmacokinetics, Pantoprazole, Proton Pump Inhibitors pharmacokinetics, Rabeprazole, Recombinant Proteins antagonists & inhibitors, Recombinant Proteins metabolism, Substrate Specificity, Aryl Hydrocarbon Hydroxylases antagonists & inhibitors, Enzyme Inhibitors pharmacology, Esomeprazole pharmacology, Liver drug effects, Omeprazole pharmacology, Proton Pump Inhibitors pharmacology
- Abstract
Six proton pump inhibitors (PPIs), omeprazole, lansoprazole, esomeprazole, dexlansoprazole, pantoprazole, and rabeprazole, were shown to be weak inhibitors of cytochromes P450 (CYP3A4, -2B6, -2D6, -2C9, -2C8, and -1A2) in human liver microsomes. In most cases, IC₅₀ values were greater than 40 μM, except for dexlansoprazole and lansoprazole with CYP1A2 (IC₅₀ = ∼8 μM) and esomeprazole with CYP2C8 (IC₅₀ = 31 μM). With the exception of CYP2C19 inhibition by omeprazole and esomeprazole (IC₅₀ ratio, 2.5 to 5.9), there was no evidence for a marked time-dependent shift in IC₅₀ (IC₅₀ ratio, ≤ 2) after a 30-min preincubation with NADPH. In the absence of preincubation, lansoprazole (IC₅₀ = 0.73 μM) and esomeprazole (IC₅₀ = 3.7 μM) were the most potent CYP2C19 inhibitors, followed by dexlansoprazole and omeprazole (IC₅₀ = ∼7.0 μM). Rabeprazole and pantoprazole (IC₅₀ = ≥ 25 μM) were the weakest. A similar ranking was obtained with recombinant CYP2C19. Despite the IC₅₀ ranking, after consideration of plasma levels (static and dynamic), protein binding, and metabolism-dependent inhibition, it is concluded that omeprazole and esomeprazole are the most potent CYP2C19 inhibitors. This was confirmed after the incubation of the individual PPIs with human primary hepatocytes (in the presence of human serum) and by monitoring their impact on diazepam N-demethylase activity at a low concentration of diazepam (2 μM). Data described herein are consistent with reports that PPIs are mostly weak inhibitors of cytochromes P450 in vivo. However, two members of the PPI class (esomeprazole and omeprazole) are more likely to serve as clinically relevant inhibitors of CYP2C19.
- Published
- 2012
- Full Text
- View/download PDF
18. PhRMA CPCDC initiative on predictive models of human pharmacokinetics, part 2: comparative assessment of prediction methods of human volume of distribution.
- Author
-
Jones RD, Jones HM, Rowland M, Gibson CR, Yates JW, Chien JY, Ring BJ, Adkison KK, Ku MS, He H, Vuppugalla R, Marathe P, Fischer V, Dutta S, Sinha VK, Björnsson T, Lavé T, and Poulin P
- Subjects
- Access to Information, Administration, Intravenous, Animals, Computer Simulation, Cooperative Behavior, Dogs, Drug Evaluation, Preclinical, Humans, Interdisciplinary Communication, Models, Statistical, Pharmaceutical Preparations administration & dosage, Pharmaceutical Preparations blood, Program Development, Program Evaluation, Protein Binding, Rats, Reproducibility of Results, Species Specificity, Databases, Pharmaceutical, Drug Discovery methods, Models, Biological, Pharmaceutical Preparations metabolism, Pharmacokinetics
- Abstract
The objective of this study was to evaluate the performance of various empirical, semimechanistic and mechanistic methodologies with and without protein binding corrections for the prediction of human volume of distribution at steady state (Vss ). PhRMA member companies contributed a set of blinded data from preclinical and clinical studies, and 18 drugs with intravenous clinical pharmacokinetics (PK) data were available for the analysis. In vivo and in vitro preclinical data were used to predict Vss by 24 different methods. Various statistical and outlier techniques were employed to assess the predictability of each method. There was not simply one method that predicts Vss accurately for all compounds. Across methods, the maximum success rate in predicting human Vss was 100%, 94%, and 78% of the compounds with predictions falling within tenfold, threefold, and twofold error, respectively, of the observed Vss . Generally, the methods that made use of in vivo preclinical data were more predictive than those methods that relied solely on in vitro data. However, for many compounds, in vivo data from only two species (generally rat and dog) were available and/or the required in vitro data were missing, which meant some methods could not be properly evaluated. It is recommended to initially use the in vitro tissue composition-based equations to predict Vss in preclinical species and humans, putting the assumptions and compound properties into context. As in vivo data become available, these predictions should be reassessed and rationalized to indicate the level of confidence (uncertainty) in the human Vss prediction. The top three methods that perform strongly at integrating in vivo data in this way were the Øie-Tozer, the rat -dog-human proportionality equation, and the lumped-PBPK approach. Overall, the scientific benefit of this study was to obtain greater characterization of predictions of human Vss from several methods available in the literature., (Copyright © 2011 Wiley-Liss, Inc.)
- Published
- 2011
- Full Text
- View/download PDF
19. PhRMA CPCDC initiative on predictive models of human pharmacokinetics, part 1: goals, properties of the PhRMA dataset, and comparison with literature datasets.
- Author
-
Poulin P, Jones HM, Jones RD, Yates JW, Gibson CR, Chien JY, Ring BJ, Adkison KK, He H, Vuppugalla R, Marathe P, Fischer V, Dutta S, Sinha VK, Björnsson T, Lavé T, and Ku MS
- Subjects
- Access to Information, Administration, Intravenous, Administration, Oral, Animals, Computer Simulation, Cooperative Behavior, Drug Evaluation, Preclinical, Humans, Interdisciplinary Communication, Models, Statistical, Pharmaceutical Preparations administration & dosage, Pharmaceutical Preparations blood, Pharmaceutical Preparations chemistry, Program Development, Program Evaluation, Reproducibility of Results, Risk Assessment, Risk Factors, Species Specificity, Databases, Pharmaceutical, Drug Discovery methods, Models, Biological, Pharmaceutical Preparations metabolism, Pharmacokinetics
- Abstract
This study is part of the Pharmaceutical Research and Manufacturers of America (PhRMA) initiative on predictive models of efficacy, safety, and compound properties. The overall goal of this part was to assess the predictability of human pharmacokinetics (PK) from preclinical data and to provide comparisons of available prediction methods from the literature, as appropriate, using a representative blinded dataset of drug candidates. The key objectives were to (i) appropriately assemble and blind a diverse dataset of in vitro, preclinical in vivo, and clinical data for multiple drug candidates, (ii) evaluate the dataset with empirical and physiological methodologies from the literature used to predict human PK properties and plasma concentration-time profiles, (iii) compare the predicted properties with the observed clinical data to assess the prediction accuracy using routine statistical techniques and to evaluate prediction method(s) based on the degree of accuracy of each prediction method, and (iv) compile and summarize results for publication. Another objective was to provide a mechanistic understanding as to why one methodology provided better predictions than another, after analyzing the poor predictions. A total of 108 clinical lead compounds were collected from 12 PhRMA member companies. This dataset contains intravenous (n = 19) and oral pharmacokinetic data (n = 107) in humans as well as the corresponding preclinical in vitro, in vivo, and physicochemical data. All data were blinded to protect the anonymity of both the data and the company submitting the data. This manuscript, which is the first of a series of manuscripts, summarizes the PhRMA initiative and the 108 compound dataset. More details on the predictability of each method are reported in companion manuscripts., (Copyright © 2011 Wiley-Liss, Inc.)
- Published
- 2011
- Full Text
- View/download PDF
20. PHRMA CPCDC initiative on predictive models of human pharmacokinetics, part 5: prediction of plasma concentration-time profiles in human by using the physiologically-based pharmacokinetic modeling approach.
- Author
-
Poulin P, Jones RD, Jones HM, Gibson CR, Rowland M, Chien JY, Ring BJ, Adkison KK, Ku MS, He H, Vuppugalla R, Marathe P, Fischer V, Dutta S, Sinha VK, Björnsson T, Lavé T, and Yates JW
- Subjects
- Access to Information, Administration, Intravenous, Administration, Oral, Animals, Computer Simulation, Cooperative Behavior, Drug Evaluation, Preclinical, Gastrointestinal Absorption, Humans, Interdisciplinary Communication, Metabolic Clearance Rate, Models, Statistical, Pharmaceutical Preparations administration & dosage, Pharmaceutical Preparations blood, Program Development, Program Evaluation, Reproducibility of Results, Species Specificity, Databases, Pharmaceutical, Drug Discovery methods, Models, Biological, Pharmaceutical Preparations metabolism, Pharmacokinetics
- Abstract
The objective of this study is to assess the effectiveness of physiologically based pharmacokinetic (PBPK) models for simulating human plasma concentration-time profiles for the unique drug dataset of blinded data that has been assembled as part of a Pharmaceutical Research and Manufacturers of America initiative. Combinations of absorption, distribution, and clearance models were tested with a PBPK approach that has been developed from published equations. An assessment of the quality of the model predictions was made on the basis of the shape of the plasma time courses and related parameters. Up to 69% of the simulations of plasma time courses made in human demonstrated a medium to high degree of accuracy for intravenous pharmacokinetics, whereas this number decreased to 23% after oral administration based on the selected criteria. The simulations resulted in a general underestimation of drug exposure (Cmax and AUC0- t ). The explanations for this underestimation are diverse. Therefore, in general it may be due to underprediction of absorption parameters and/or overprediction of distribution or oral first-pass. The implications of compound properties are demonstrated. The PBPK approach based on in vitro-input data was as accurate as the approach based on in vivo data. Overall, the scientific benefit of this modeling study was to obtain more extensive characterization of predictions of human PK from PBPK methods., (Copyright © 2011 Wiley-Liss, Inc.)
- Published
- 2011
- Full Text
- View/download PDF
21. PhRMA CPCDC initiative on predictive models of human pharmacokinetics, part 3: comparative assessement of prediction methods of human clearance.
- Author
-
Ring BJ, Chien JY, Adkison KK, Jones HM, Rowland M, Jones RD, Yates JW, Ku MS, Gibson CR, He H, Vuppugalla R, Marathe P, Fischer V, Dutta S, Sinha VK, Björnsson T, Lavé T, and Poulin P
- Subjects
- Access to Information, Administration, Intravenous, Animals, Area Under Curve, Computer Simulation, Cooperative Behavior, Dogs, Drug Evaluation, Preclinical, Humans, Interdisciplinary Communication, Metabolic Clearance Rate, Models, Statistical, Pharmaceutical Preparations administration & dosage, Pharmaceutical Preparations blood, Program Development, Program Evaluation, Protein Binding, Rats, Reproducibility of Results, Species Specificity, Databases, Pharmaceutical, Drug Discovery methods, Models, Biological, Pharmaceutical Preparations metabolism, Pharmacokinetics
- Abstract
The objective of this study was to evaluate the performance of various allometric and in vitro-in vivo extrapolation (IVIVE) methodologies with and without plasma protein binding corrections for the prediction of human intravenous (i.v.) clearance (CL). The objective was also to evaluate the IVIVE prediction methods with animal data. Methodologies were selected from the literature. Pharmaceutical Research and Manufacturers of America member companies contributed blinded datasets from preclinical and clinical studies for 108 compounds, among which 19 drugs had i.v. clinical pharmacokinetics data and were used in the analysis. In vivo and in vitro preclinical data were used to predict CL by 29 different methods. For many compounds, in vivo data from only two species (generally rat and dog) were available and/or the required in vitro data were missing, which meant some methods could not be properly evaluated. In addition, 66 methods of predicting oral (p.o.) area under the curve (AUCp.o. ) were evaluated for 107 compounds using rational combinations of i.v. CL and bioavailability (F), and direct scaling of observed p.o. CL from preclinical species. Various statistical and outlier techniques were employed to assess the predictability of each method. Across methods, the maximum success rate in predicting human CL for the 19 drugs was 100%, 94%, and 78% of the compounds with predictions falling within 10-fold, threefold, and twofold error, respectively, of the observed CL. In general, in vivo methods performed slightly better than IVIVE methods (at least in terms of measures of correlation and global concordance), with the fu intercept method and two-species-based allometry (rat-dog) being the best performing methods. IVIVE methods using microsomes (incorporating both plasma and microsomal binding) and hepatocytes (not incorporating binding) resulted in 75% and 78%, respectively, of the predictions falling within twofold error. IVIVE methods using other combinations of binding assumptions were much less accurate. The results for prediction of AUCp.o. were consistent with i.v. CL. However, the greatest challenge to successful prediction of human p.o. CL is the estimate of F in human. Overall, the results of this initiative confirmed predictive performance of common methodologies used to predict human CL., (Copyright © 2011 Wiley-Liss, Inc.)
- Published
- 2011
- Full Text
- View/download PDF
22. PhRMA CPCDC initiative on predictive models of human pharmacokinetics, part 4: prediction of plasma concentration-time profiles in human from in vivo preclinical data by using the Wajima approach.
- Author
-
Vuppugalla R, Marathe P, He H, Jones RD, Yates JW, Jones HM, Gibson CR, Chien JY, Ring BJ, Adkison KK, Ku MS, Fischer V, Dutta S, Sinha VK, Björnsson T, Lavé T, and Poulin P
- Subjects
- Access to Information, Administration, Intravenous, Administration, Oral, Animals, Biological Availability, Computer Simulation, Cooperative Behavior, Dogs, Drug Evaluation, Preclinical, Gastrointestinal Absorption, Humans, Interdisciplinary Communication, Metabolic Clearance Rate, Models, Statistical, Pharmaceutical Preparations administration & dosage, Pharmaceutical Preparations blood, Program Development, Program Evaluation, Rats, Reproducibility of Results, Species Specificity, Databases, Pharmaceutical, Drug Discovery methods, Models, Biological, Pharmaceutical Preparations metabolism, Pharmacokinetics
- Abstract
The objective of this study was to evaluate the performance of the Wajima allometry (Css -MRT) approach published in the literature, which is used to predict the human plasma concentration-time profiles from a scaling of preclinical species data. A diverse and blinded dataset of 108 compounds from PhRMA member companies was used in this evaluation. The human intravenous (i.v.) and oral (p.o.) pharmacokinetics (PK) data were available for 18 and 107 drugs, respectively. Three different scenarios were adopted for prediction of human PK profiles. In the first scenario, human clearance (CL) and steady-state volume of distribution (Vss ) were predicted by unbound fraction corrected intercept method (FCIM) and Øie-Tozer (OT) approaches, respectively. Quantitative structure activity relationship (QSAR)-based approaches (TSrat-dog ) based on compound descriptors together with rat and dog data were utilized in the second scenario. Finally, in the third scenario, CL and Vss were predicted using the FCIM and Jansson approaches, respectively. For the prediction of oral pharmacokinetics, the human bioavailability and absorption rate constant were assumed as the average of preclinical species. Various statistical techniques were used for assessing the accuracy of the simulation scenarios. The human CL and Vss were predicted within a threefold error range for about 75% of the i.v. drugs. However, the accuracy in predicting key p.o. PK parameters appeared to be lower with only 58% of simulations falling within threefold of observed parameters. The overall ability of the Css -MRT approach to predict the curve shape of the profile was in general poor and ranged between low to medium level of confidence for most of the predictions based on the selected criteria., (Copyright © 2011 Wiley-Liss, Inc.)
- Published
- 2011
- Full Text
- View/download PDF
23. Evaluation of cynomolgus monkey pregnane X receptor, primary hepatocyte, and in vivo pharmacokinetic changes in predicting human CYP3A4 induction.
- Author
-
Kim S, Dinchuk JE, Anthony MN, Orcutt T, Zoeckler ME, Sauer MB, Mosure KW, Vuppugalla R, Grace JE Jr, Simmermacher J, Dulac HA, Pizzano J, and Sinz M
- Subjects
- Adult, Amino Acid Sequence, Animals, Bridged Bicyclo Compounds blood, Bridged Bicyclo Compounds pharmacokinetics, Bridged Bicyclo Compounds pharmacology, Cell Line, Cell Line, Tumor, Cell Survival drug effects, Cloning, Molecular, Cytochrome P-450 CYP3A genetics, Cytochrome P-450 CYP3A metabolism, Drug Interactions genetics, Enzyme Induction drug effects, Enzyme Induction genetics, Female, Gene Expression drug effects, Gene Expression genetics, Hepatocytes drug effects, Hepatocytes enzymology, Humans, Hypericum chemistry, Macaca mulatta, Male, Midazolam blood, Midazolam metabolism, Midazolam pharmacokinetics, Middle Aged, Models, Animal, Molecular Sequence Data, Phloroglucinol analogs & derivatives, Phloroglucinol blood, Phloroglucinol pharmacokinetics, Phloroglucinol pharmacology, Plant Extracts blood, Plant Extracts pharmacokinetics, Pregnane X Receptor, Receptors, Steroid genetics, Rifampin blood, Rifampin pharmacokinetics, Rifampin pharmacology, Sequence Homology, Amino Acid, Terpenes blood, Terpenes pharmacokinetics, Terpenes pharmacology, Transcriptional Activation drug effects, Transcriptional Activation genetics, Transfection, Cytochrome P-450 CYP3A biosynthesis, Hepatocytes metabolism, Macaca fascicularis, Receptors, Steroid metabolism, Xenobiotics pharmacokinetics
- Abstract
Monkeys have been proposed as an animal model to predict the magnitude of human clinical drug-drug interactions caused by CYP3A4 enzyme induction. To evaluate whether the cynomolgus monkey can be an effective in vivo model, human CYP3A4 inducers were evaluated both in vitro and in vivo. First, a full-length pregnane X receptor (PXR) was cloned from the cynomolgus monkey, and the sequence was compared with those of rhesus monkey and human PXR. Cynomolgus and rhesus monkey PXR differed by only one amino acid (A68V), and both were highly homologous to human PXR (approximately 96%). When the transactivation profiles of 30 compounds, including known inducers of CYP3A4, were compared between cynomolgus and human PXR, a high degree of correlation with EC(50) values was observed. These results suggest that cynomolgus and human PXR respond in a similar fashion to these ligands. Second, two known human CYP3A4 inducers, rifampicin and hyperforin, were tested in monkey and human primary hepatocytes for induction of CYP3A enzymes. Both monkey and human hepatocytes responded similarly to the inducers and resulted in increased RNA and enzyme activity changes of CYP3A8 and CYP3A4, respectively. Lastly, in vivo induction of CYP3A8 by rifampicin and hyperforin was shown by significant reductions of midazolam exposure that were comparable with those in humans. These results show that the cynomolgus monkey can be a predictive in vivo animal model of PXR-mediated induction of human CYP3A4 and can provide a useful assessment of the resulting pharmacokinetic changes of affected drugs.
- Published
- 2010
- Full Text
- View/download PDF
24. Effect of commonly used organic solvents on the kinetics of cytochrome P450 2B6- and 2C8-dependent activity in human liver microsomes.
- Author
-
Vuppugalla R, Chang SY, Zhang H, Marathe PH, and Rodrigues DA
- Subjects
- Acetonitriles pharmacology, Bupropion analogs & derivatives, Bupropion metabolism, Cytochrome P-450 CYP2B6, Cytochrome P-450 CYP2C8, Dimethyl Sulfoxide pharmacology, Ethanol pharmacology, Humans, Hydroxylation drug effects, Kinetics, Methanol pharmacology, Microsomes, Liver enzymology, Microsomes, Liver metabolism, Paclitaxel metabolism, Taxoids metabolism, Aryl Hydrocarbon Hydroxylases metabolism, Cytochrome P-450 Enzyme System metabolism, Microsomes, Liver drug effects, Solvents pharmacology
- Abstract
The effect of common organic solvents on the activities of various human cytochromes P450 has been reported. However, very little is known about their influence on CYP2B6 and CYP2C8 enzymes. The purpose of this study was to investigate the effect of solvents on the kinetics of representative CYP2B6 (bupropion hydroxylase) and CYP2C8 (paclitaxel hydroxylase) reactions in human liver microsomes. Methanol, ethanol, dimethyl sulfoxide (DMSO), and acetonitrile were studied at increasing volumes (v/v). Acetonitrile, DMSO, and ethanol were shown to increase the Km and decrease the intrinsic clearance (CLint) of CYP2B6-mediated bupropion hydroxylation in a concentration-dependent manner. These solvents did not noticeably alter the Vmax at concentrations of < or =1% (v/v). Unlike the other solvents studied, the effect of methanol (< or =0.5%, v/v) on CYP2B6 kinetics was negligible. Both DMSO and ethanol increased the Km and decreased the CL(int) of CYP2C8-mediated paclitaxel hydroxylation in a concentration-dependent manner. Acetonitrile had minimal influence on CYP2C8 enzyme kinetics at concentrations of < or =1% (v/v). Methanol decreased the Km of paclitaxel at low concentrations followed by an increase at concentrations of > or =2% (v/v). This differential influence on Km resulted in an increased CLint at low concentrations followed by a decrease at high concentrations. The studied solvents had minimal influence on Vmax of paclitaxel. Collectively, DMSO and ethanol were not suitable for characterizing CYP2B6- and CYP2C8-mediated reactions because they showed concentration-dependent inhibition. Methanol and acetonitrile at concentrations of < or =0.5% and < or =1% (v/v) appeared to be suitable for the measurement of CYP2B6- and CYP2C8-mediated activities, respectively.
- Published
- 2007
- Full Text
- View/download PDF
25. Route-dependent stereoselective pharmacokinetics of tramadol and its active O-demethylated metabolite in rats.
- Author
-
Parasrampuria R, Vuppugalla R, Elliott K, and Mehvar R
- Subjects
- Analgesics, Opioid administration & dosage, Animals, Gastrointestinal Tract metabolism, Liver metabolism, Male, Rats, Rats, Sprague-Dawley, Stereoisomerism, Tramadol administration & dosage, Analgesics, Opioid pharmacokinetics, Tramadol pharmacokinetics
- Abstract
The effects of route of administration on the stereoselective pharmacokinetics of tramadol (T) and its active metabolite (M1) were studied in rats. A single 20 mg/kg dose of racemic T was administered through intravenous, intraperitoneal, or oral route to different groups of rats, and blood and urine samples were collected. Samples were analyzed using chiral chromatography, and pharmacokinetic parameters (mean +/- SD) were estimated by noncompartmental methods. Following intravenous injection, there was no stereoselectivity in the pharmacokinetics of T. Both enantiomers showed clearance values (62.5 +/- 27.2 and 64.4 +/- 39.0 ml/min/kg for (+)- and (-)-T, respectively) that were equal or higher than the reported liver blood flow in rats. Similar to T, the area under the plasma concentration-time curves (AUCs) of M1 did not exhibit stereoselectivity after intravenous administration of the parent drug. However, the systemic availability of (+)-T was significantly (P < 0.05) higher than that of its antipode following intraperitoneal (0.527 +/- 0.240 vs. 0.373 +/- 0.189) and oral (0.307 +/- 0.136 vs. 0.159 +/- 0.115) administrations. The AUC of the M1 enantiomers, on the other hand, remained mostly nonstereoselective regardless of the route of administration. Pharmacokinetic analysis indicated that the stereoselectivity in the pharmacokinetics of oral T is due to stereoselective first pass metabolism in the liver and, possibly, in the gastrointestinal tract. The direction and extent of stereoselectivity in the pharmacokinetics of T and M1 in rats were in agreement with those previously reported in humans, suggesting that the rat may be a suitable model for enantioselective studies of T pharmacokinetics.
- Published
- 2007
- Full Text
- View/download PDF
26. Hepatic disposition of the cytochrome P450 2E1 marker chlorzoxazone and its hydroxylated metabolite in isolated perfused rat livers.
- Author
-
Mehvar R and Vuppugalla R
- Subjects
- Animals, Bile chemistry, Biomarkers, Cytochrome P-450 CYP2E1, In Vitro Techniques, Male, Muscle Relaxants, Central metabolism, Perfusion, Rats, Rats, Sprague-Dawley, Chlorzoxazone analogs & derivatives, Chlorzoxazone metabolism, Liver metabolism
- Abstract
The steady-state disposition of chlorzoxazone (CZX) and its hydroxylated metabolite 6-hydroxychlorzoxazone (HCZX) was determined in a single-pass isolated perfused rat liver (IPRL) model using constant CZX concentrations of 10-200 microM. The concentrations of CZX, HCZX, and/or HCZX glucuronide in the perfusate, bile, and liver tissues were measured and kinetic parameters calculated. Upon an increase in CZX inlet concentrations from 10 to 200 microM, its extraction ratio sharply declined from 0.681 to 0.087. This was associated with a saturable formation of HCZX, which was rapidly and completely metabolized to its glucuronide conjugate. Whereas the biliary excretion of CZX was negligible, that of HCZX was substantial (up to 40% of the generated metabolite). Overall, 79-93% of the CZX dose (10-200 microM) was recovered in our model as CZX and HCZX. Additionally, HCZX accounted for 56% (200 microM) to 71% (10 microM) of the extracted CZX dose. Further, a preliminary study using the preformed HCZX showed a complete (100%) recovery of the metabolite as its conjugate. Therefore, the unrecovered portion of CZX dose in our study (7-21% of the administered dose or 29-44% of the extracted dose at inlet CZX concentrations of 10-200 microM) is most likely due to parallel metabolism of CZX to other metabolites.
- Published
- 2006
- Full Text
- View/download PDF
27. Selective effects of nitric oxide on the disposition of chlorzoxazone and dextromethorphan in isolated perfused rat livers.
- Author
-
Vuppugalla R and Mehvar R
- Subjects
- Alcohol Oxidoreductases metabolism, Animals, Chlorzoxazone metabolism, Cytochrome P-450 CYP2E1 metabolism, Cytochrome P450 Family 2, Dextrorphan metabolism, Enzyme Inhibitors pharmacology, Glucuronosyltransferase metabolism, In Vitro Techniques, Isosorbide Dinitrate pharmacology, Kinetics, Liver enzymology, Male, Nitroprusside pharmacology, Perfusion, Rats, Rats, Sprague-Dawley, Umbelliferones metabolism, Chlorzoxazone analogs & derivatives, Cytochrome P-450 CYP2E1 Inhibitors, Dextromethorphan metabolism, Glucuronosyltransferase antagonists & inhibitors, Liver drug effects, Nitric Oxide Donors pharmacology
- Abstract
The rapid and direct effects of nitric oxide (NO) donors sodium nitroprusside (SNP) and isosorbide dinitrate (ISDN) on the hepatic and biliary disposition of chlorzoxazone (CZX), a marker of CYP2E1, and dextromethorphan (DEM), a marker of CYP2D1, were studied in a single-pass isolated perfused rat liver model. Livers (n = 30) were perfused with constant concentrations of NO donors (0-120 min) in addition to infusion of CZX or DEM (60-120 min), and periodical outlet and bile samples were collected. Both ISDN and SNP significantly reduced (30 and 60%, respectively) the hepatic extraction ratio of CZX and decreased (50 and 70%, respectively) the recovery of the CYP2E1-mediated metabolite, 6-hydroxychlorzoxazone, in the outlet perfusate and bile. As for DEM, both NO donors increased (up to 3.5-fold) the recovery of the CYP2D1-mediated metabolite dextrorphan (DOR) in the outlet perfusate. However, this was associated with a simultaneous decrease (50-75%) in the excretion of the metabolite into the bile, thus resulting in no change in the overall recovery of DOR as a result of NO donor treatment. The decrease in the biliary excretion of DOR was caused by NO-induced simultaneous reductions in both the conjugation of DOR and biliary clearance of DOR conjugate. Additionally, both SNP and ISDN significantly reduced the metabolism of DEM to 3-hydroxymorphinan, which is mostly regulated by CYP3A2. These studies in an intact liver model confirm the selectivity of the inhibitory effects of NO donors on cytochrome P450 enzymes, which was recently reported in microsomal studies, and expand these inhibitory effects to conjugation pathways.
- Published
- 2006
- Full Text
- View/download PDF
28. Enzyme-selective effects of nitric oxide on affinity and maximum velocity of various rat cytochromes P450.
- Author
-
Vuppugalla R and Mehvar R
- Subjects
- Animals, Cytochrome P-450 Enzyme System pharmacokinetics, Isoenzymes metabolism, Isoenzymes pharmacokinetics, Liver drug effects, Male, Nitric Oxide pharmacokinetics, Nitric Oxide Donors pharmacology, Rats, Rats, Sprague-Dawley, Cytochrome P-450 Enzyme System metabolism, Liver enzymology, Nitric Oxide metabolism
- Abstract
Nitric oxide (NO) has recently been shown to decrease cytochrome P450 (P450) enzyme activity rapidly (< or =30 min), concentration dependently, and enzyme-selectively in the rat liver. Interestingly, among all the studied P450 enzymes, only CYP2D1 was not affected by NO donors. However, these studies were conducted using only a single concentration of the substrates, thus lacking information about the possible simultaneous changes in both maximum velocity (Vmax) and affinity (Km) of the enzymes. In the present study, we systematically evaluated the effects of NO on the enzyme kinetic parameters of marker substrates for a range of P450 enzymes, including 2D1. Livers were perfused (1 h) in the absence (control) or presence of two NO donors with different mechanisms of NO release. At the end of the perfusion, microsomes were prepared and used for kinetic analysis. Except for 2D1, NO reduced the Vmax of all the model reactions studied, although to a varying degree. However, the effects of NO donors on Km were more diverse. Whereas the Km values for testosterone 6beta-hydroxylation (3A2) and 16alpha-hydroxylation (2C11) significantly decreased, the values for chlorzoxazone 6-hydroxylation (2E1), dextromethorphan N-demethylation (3A2), and high affinity ethoxyresorufin O-dealkylation (1A1/2) significantly increased in the presence of NO donors. Furthermore, the Km values for the high-affinity component of dextromethorphan O-demethylation and benzyloxyresorufin O-dealkylation remained unchanged. These results indicate that NO can potentially change both the Vmax and Km of various substrates selectively and confirm our previous findings that the activity of CYP2D1 is not affected by NO donors.
- Published
- 2005
- Full Text
- View/download PDF
29. Short-term inhibitory effects of nitric oxide on cytochrome P450-mediated drug metabolism: time dependency and reversibility profiles in isolated perfused rat livers.
- Author
-
Vuppugalla R and Mehvar R
- Subjects
- Animals, Bile physiology, Cytochromes b5 metabolism, Enzyme Inhibitors pharmacology, Heme metabolism, In Vitro Techniques, Male, Microsomes, Liver drug effects, Microsomes, Liver enzymology, Nitric Oxide Donors pharmacology, Nitroprusside pharmacology, Proteins metabolism, Rats, Rats, Sprague-Dawley, Sulfhydryl Compounds metabolism, Time Factors, Tyrosine metabolism, Cytochrome P-450 Enzyme Inhibitors, Cytochrome P-450 Enzyme System metabolism, Liver drug effects, Liver enzymology, Nitric Oxide pharmacology, Pharmaceutical Preparations metabolism, Tyrosine analogs & derivatives
- Abstract
Nitric oxide (NO) is implicated as a mediator in the decreased catalytic activities of cytochrome P450 (P450) enzymes during inflammation or infection. Here, we examined the time course and the reversibility of the NO effect on P450s using isolated perfused rat livers. Livers were perfused at a constant rate with the NO donor sodium nitroprusside (SNP) for 0.5 or 1 h, followed by washout periods of 0 to 2.5 h. At the end of perfusion, microsomes were prepared and analyzed for P450 activities and other metabolic markers. Whereas 0.5 h of NO exposure caused an irreversible decline (approximately 30%) in total P450 content, a greater decline after 1 h of NO (approximately 55%) was mostly (approximately 30%) reversible, a pattern identical to that observed for the microsomal heme content. NO exposure also caused an enzyme-selective and time-dependent decline in P450 activities. Whereas the pattern of decline and reversibility of activities were qualitatively similar for CYP3A2, 2C11, 2E1, and 1A1/2, they differed for 2B1/2 and 2D1 in that the decline in the activity was delayed (1 h) for 2B1/2 and not observed for 2D1. This may be attributed to the accessibility of heme or cysteine thiolate and/or the presence/reactivity of critical cysteinyl amino acid residues in various P450 enzymes. Additionally, for most enzymes, the activity showed a biphasic decline, one within 1 h of SNP perfusion and another after 2 h of washout. This was associated with an identical biphasic decline in the microsomal free thiols, presumably due to the rapid and slow reaction of NO and peroxynitrite, respectively, with critical P450 thiols. The short-term effects of NO on P450 are time-dependent and enzyme-selective, with both reversible and irreversible mechanisms.
- Published
- 2004
- Full Text
- View/download PDF
30. Hepatic disposition and effects of nitric oxide donors: rapid and concentration-dependent reduction in the cytochrome P450-mediated drug metabolism in isolated perfused rat livers.
- Author
-
Vuppugalla R and Mehvar R
- Subjects
- Animals, Dose-Response Relationship, Drug, In Vitro Techniques, Male, Nitric Oxide metabolism, Perfusion, Rats, Rats, Sprague-Dawley, Cytochrome P-450 Enzyme System metabolism, Liver drug effects, Liver metabolism, Nitric Oxide Donors pharmacology
- Abstract
Various mechanisms, including high levels of cytokines and nitric oxide (NO), have been proposed as mediators for inflammation-induced cytochrome 450 down-regulation. However, the contribution of each of these mediators to the observed effects is controversial. We used an isolated perfused rat liver (IPRL) model to test the direct effects of NO donors on CYP450 down-regulation in the absence of cytokines or other confounding in vivo factors. Our hypothesis was that NO rapidly and concentration-dependently decreases CYP450 activities in IPRL. Livers were perfused (60 min) with 50 to 500 microM sodium nitroprusside (SNP) or 100 to 500 microM isosorbide dinitrate (ISDN) as NO donors, and the perfusate and biliary disposition of SNP, ISDN, and generated nitrate/nitrite (NO(x)) were determined. Additionally, at the end of perfusion, catalytic activities and protein levels of various cytochrome isoenzymes were measured. Both SNP and ISDN exhibited linear hepatic disposition with extraction ratios of approximately 0.30 and 0.50, respectively. Furthermore, although in small amounts, both NO donors and NO(x) were found in the bile. Except for CYP2D1, the catalytic activities of all the studied isoenzymes were substantially (up to 85%) decreased by both NO donors. However, the apoprotein levels of isoenzymes remained largely unchanged. Additionally, the inhibitory effects of NO donors were concentration-dependent, with the concentrations of SNP producing one-half of maximum inhibition being in the order of 2C11 > 2B1/2 > 2E1 = 3A2 > 1A1/2. These studies indicate that the effects of NO on the down-regulation of cytochrome 450 catalytic activity are rapid, concentration-dependent, and isoenzyme-selective.
- Published
- 2004
- Full Text
- View/download PDF
31. A simple HPLC method for the simultaneous analysis of insulin and ovomucoid.
- Author
-
Vuppugalla R, Agarwal V, and Khan MA
- Subjects
- Animals, Calibration, Chickens, Chromatography, High Pressure Liquid, Humans, Recombinant Proteins analysis, Reference Standards, Reproducibility of Results, Solutions, Spectrophotometry, Ultraviolet, Verapamil chemistry, Insulin analysis, Ovomucin analysis
- Abstract
An analytical HPLC method is reported for the simultaneous determination of insulin and its enzyme inhibitor, chicken ovomucoid. Verapamil was used as an internal standard. The elution was achieved using a gradient technique (10-15% B for 4 min, 15-35% B from 5th to 11th min and 35-10% B from 12th to 22nd min). The mobile phase used was 0.05% v/v trifluoroacetic acid (TFA) in water and 0.05% v/v TFA in acetonitrile with a flow rate of 1.2 ml/min. The analytes were detected at 210 nm after resolution using a reversed phase C-18 column. Insulin, ovomucoid and verapamil (IS) were eluted at 11.9, 14.2, and 18 min, respectively, free from any interfering endogenous peaks during a run time of 22 min. Linear relationships were observed between the detector response and the concentrations of the analytes (0.05-1 I.U/ml for insulin (r2 = 0.9975) and 5-100 microg/ml for the chicken ovomucoid (r2 = 0.9993)). The assay was found to be highly selective and sensitive due to the absence of any interfering peaks. The lower C.V and % error values of the assay indicates that the assay could accurately and precisely quantitate both insulin and ovomucoid in the examined concentration range. This method can be used for the simultaneous quantitation of insulin and chicken ovomucoid.
- Published
- 2003
32. Microsomal cytochrome P450 levels and activities of isolated rat livers perfused with albumin.
- Author
-
Vuppugalla R, Shah RB, Chimalakonda AP, Fisher CW, and Mehvar R
- Subjects
- Alanine Transaminase metabolism, Animals, Aspartate Aminotransferases metabolism, Bile drug effects, Bile metabolism, Cytokines metabolism, In Vitro Techniques, Isoenzymes metabolism, Liver drug effects, Liver enzymology, Liver metabolism, Male, Microsomes, Liver drug effects, NADH Dehydrogenase metabolism, Nitrates metabolism, Nitrites metabolism, Organ Size drug effects, Perfusion, Rats, Rats, Sprague-Dawley, Serum Albumin, Bovine pharmacokinetics, Cytochrome P-450 Enzyme System metabolism, Microsomes, Liver enzymology, Serum Albumin, Bovine pharmacology
- Abstract
Purpose: We recently showed that the perfusion of isolated rat livers with perfusates containing bovine serum albumin (BSA) would significantly stimulate the release of tumor necrosis factor (TNF)-alpha. Here, we hypothesize that BSA-induced increase in the release of TNF-alpha, and possibly other cytokines, would affect cytochrome P450 (CYP)-mediated drug metabolism., Methods: Rat livers were perfused ex vivo for 1, 2, or 3 h with a physiologic buffer containing or lacking 1% BSA (n = 4-5/group). At the end of perfusion, liver microsomes were prepared and analyzed for their total CYP, CYP2E1, CYP3A2, and CYP2C11 protein contents and the activities of cytochrome c reductase, CYP2E1, CYP3A2, CYP2C11, CYP2E1, CYP2D1, CYP1A1, and CYP2B1/2. In addition, the concentrations of various cytokines and nitric oxide were quantified in the outlet perfusate., Results: In the absence of BSA, the perfusate levels of all measured cytokines and nitric oxide were low. However, when the perfusate contained BSA, the levels of TNF-alpha, interleukin-6, and nitric oxide increased significantly (p < 0.005). Perfusion of the livers for 3 h with the BSA-containing perfusate resulted in significant (p < 0.05) decreases in the total CYP (41%), CYP2E1 (59%), CYP3A2 (68%), and CYP2C11 (50%) protein contents and activities of cytochrome c reductase (31%), CYP2E1 (66%), CYP3A2 (54%), and CYP2G11 (51%). In contrast, perfusion of livers for 1 or 2 h with the BSA perfusate did not have any significant effect on CYP-mediated metabolism. The CYP1A2, CYP2D1, and CYP2B1/2 activities were not affected by BSA, regardless of perfusion time., Conclusion: Addition of BSA to perfusates, which is a routine practice in isolated rat liver studies, can reduce CYP-mediated drug metabolism by a mechanism independent of protein-binding effect.
- Published
- 2003
- Full Text
- View/download PDF
Catalog
Discovery Service for Jio Institute Digital Library
For full access to our library's resources, please sign in.