Modification of the ultrafiltration technique to overcome solubility and non-specific binding challenges associated with the measurement of plasma protein binding of corticosteroids

Taylor, S.; Harker, A.J.

doi:10.1016/j.jpba.2005.10.031

Cited by 80 publications

(50 citation statements)

References 14 publications

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“…Binding of SCY-635 to human plasma proteins was determined by ultrafiltration by methods adapted to reduce nonspecific binding (27). CsA, SCY-635, or a control compound (warfarin, imipramine, or carbamezapine) was added to pooled fresh human plasma (Bioreclamation, Liverpool, NY), and the mixture was incubated at 37°C for 30 min before centrifugation.…”

Section: Methodsmentioning

confidence: 99%

SCY-635, a Novel Nonimmunosuppressive Analog of Cyclosporine That Exhibits Potent Inhibition of Hepatitis C Virus RNA Replication In Vitro

Hopkins

Scorneaux

Huang

et al. 2010

Antimicrob Agents Chemother

136

125

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SCY-635 is a novel nonimmunosuppressive cyclosporine-based analog that exhibits potent suppression of hepatitis C virus (HCV) replication in vitro. SCY-635 inhibited the peptidyl prolyl isomerase activity of cyclophilin A at nanomolar concentrations but showed no detectable inhibition of calcineurin phosphatase activity at concentrations up to 2 M. Metabolic studies indicated that SCY-635 did not induce the major cytochrome P450 enzymes 1A2, 2B6, and 3A4. SCY-635 was a weak inhibitor and a poor substrate for P-glycoprotein. Functional assays with stimulated Jurkat cells and stimulated human peripheral blood mononuclear cells indicated that SCY-635 is a weaker inhibitor of interleukin-2 secretion than cyclosporine. A series of two-drug combination studies was performed in vitro. SCY-635 exhibited synergistic antiviral activity with alpha interferon 2b and additive antiviral activity with ribavirin. SCY-635 was shown to be orally bioavailable in multiple animal species and produced blood and liver concentrations of parent drug that exceeded the 50% effective dose determined in the bicistronic con1b-derived replicon assay. These results suggest that SCY-635 warrants further investigation as a novel therapeutic agent for the treatment of individuals who are chronically infected with HCV.Hepatitis C virus (HCV) is a member of the Flaviviridae family, which comprises three distinct genera, including the flaviviruses (such as yellow fever virus, dengue virus, West Nile virus, and Japanese encephalitis virus), the pestiviruses (bovine viral diarrhea virus and classical swine fever virus), and the hepaciviruses (of which HCV is the only member) (16). HCV is highly polymorphic, and current taxonomic schemes recognize six major genotypes and several subtypes. Although no strict relationship exists between the genotype and the severity of HCV disease or the clinical outcome, numerous clinical studies indicate that patients who are infected with genotype 1 viruses are less responsive to antiviral therapy than individuals who are infected with genotypes 2 through 6 (10, 11). Chronic infection with HCV now represents a major global health problem, with approximately 170 million people worldwide being infected (26). The current standard of care for chronic hepatitis C virus infection involves treatment for up to 1 year with combination chemotherapy of pegylated alpha interferon coadministered with ribavirin. At this time, there are no approved drugs specifically indicated for the treatment of patients who do not respond to first-line therapy. Complete clearance of the virus is achieved in approximately 50% of all HCV-infected patients who initiate therapy (10, 11), and the response rates are related to viral factors (the genotype and the viral load), as well as multiple host factors (the presence of liver fibrosis, cirrhosis, ethnicity, coinfection with HIV type 1 [HIV-1], alcohol consumption, and metabolic disorders).At this time, the combined action of interferon and ribavirin against HCV infection is poorly understood. The e...

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Section: Methodsmentioning

confidence: 99%

SCY-635, a Novel Nonimmunosuppressive Analog of Cyclosporine That Exhibits Potent Inhibition of Hepatitis C Virus RNA Replication In Vitro

Hopkins

Scorneaux

Huang

et al. 2010

Antimicrob Agents Chemother

136

125

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“…Preliminary studies showed that a significant fraction of [ 3 H]AA-I binds to the collection tubes in the absence of protein, resulting in overestimation of the fraction bound. Therefore, we used a method designed to reduce nonspecific binding of lipophilic compounds, as reported by others (Taylor and Harker, 2006).…”

Section: Methodsmentioning

confidence: 99%

Physiological and Molecular Characterization of Aristolochic Acid Transport by the Kidney

Dickman

Sweet²,

Bonala³

et al. 2011

J Pharmacol Exp Ther

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Consumption of herbal medicines derived fromAristolochia plants is associated with a progressive tubulointerstitial disease known as aristolochic acid (AA) nephropathy. The nephrotoxin produced naturally by these plants is AA-I, a nitrophenanthrene carboxylic acid that selectively targets the proximal tubule. This nephron segment is prone to toxic injury because of its role in secretory elimination of drugs and other xenobiotics. Here, we characterize the handling of AA-I by membrane transporters involved in renal organic anion transport. Uptake assays in heterologous expression systems identified murine organic anion transporters (mOat1, mOat2, and mOat3) as capable of mediating transport of AA-I. Kinetic analyses showed that all three transporters have an affinity for AA-I in the submicromolar range and thus are likely to operate at toxicologically relevant concentrations in vivo. Structure-activity relationships revealed that the carboxyl group is critical for high-affinity interaction of AA-I with mOat1, mOat2, and mOat3, whereas the nitro group is required only by mOat1. Furthermore, the 8-methoxy group, although essential for toxicity, was not requisite for transport. Mouse renal cortical slices avidly accumulated AA-I, achieving slice-to-medium concentration ratios Ͼ10. Uptake by slices was sensitive to known mOat1 and mOat3 substrates and the organic anion transport inhibitor probenecid, which also blocked the production of DNA adducts formed with reactive intracellular metabolites of AA-I. Taken together, these findings indicate that OAT family members mediate high-affinity transport of AA-I and may be involved in the site-selective toxicity and renal elimination of this nephrotoxin.

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“…We measured drug binding to BSA and two enzyme sources, insect cell membranes and HLM, by a newly developed ultrafiltration assay. Because ultrafiltration systems often "suffer" from high NSB f values (Lee et al, 2003;Taylor and Harker, 2006), we took care to minimize, determine, and take into account the nonspecific binding to the filter device, regardless of whether the binding was to the filter itself or to the walls of the tube. Our results concerning AZT binding to BSA, negligible binding, are the same as in a previous study that used a dialysis system (Rowland et al, 2007).…”

Section: Albumin Effect In Ugt2b7 and Ugt1a9mentioning

confidence: 99%

Bovine Serum Albumin Decreases K_m Values of Human UDP-Glucuronosyltransferases 1A9 and 2B7 and Increases V_max Values of UGT1A9

Manevski¹,

Moreolo²,

Yli‐Kauhaluoma³

et al. 2011

Drug Metab Dispos

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ABSTRACT:The human UDP-glucuronosyltransferase (UGT) enzymes UGT1A9 and UGT2B7 play important roles in the hepatic glucuronidation of many drugs. The presence of bovine serum albumin (BSA) during in vitro assays was recently reported to lower the K m values of both these UGTs for their aglycone substrates without affecting the corresponding V max values. Nonetheless, using the specific substrates entacapone and zidovudine (AZT) for UGT1A9 and UGT2B7, respectively, and using an improved ultrafiltration method for measuring drug binding to BSA and to biological membranes, we found that the presence of BSA during the glucuronidation reaction leads to a large increase in the V max value of UGT1A9, in addition to lowering its K m value. On the other hand, in the case of UGT2B7, our results agree with the previously described effect of BSA, namely lowering the K m value without a large effect on the enzyme's V max value. The unexpected BSA effect on UGT1A9 was independent of the expression system because it was found in a recombinant enzyme that was expressed in baculovirus-infected insect cells as well as in the native enzyme in human liver microsomes. Moreover, the effect of BSA on the kinetics of 4-methylumbelliferone glucuronidation by recombinant UGT1A9 was similar to its effect on entacapone glucuronidation. Contrary to the aglycone substrates, the effect of BSA on the apparent K m of UGT1A9 for the cosubstrate UDP-␣-D-glucuronic acid was nonsignificant. Our findings call for further investigations of the BSA effects on different UGTs and the inhibitors that it may remove.

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Modification of the ultrafiltration technique to overcome solubility and non-specific binding challenges associated with the measurement of plasma protein binding of corticosteroids

Cited by 80 publications

References 14 publications

SCY-635, a Novel Nonimmunosuppressive Analog of Cyclosporine That Exhibits Potent Inhibition of Hepatitis C Virus RNA Replication In Vitro

SCY-635, a Novel Nonimmunosuppressive Analog of Cyclosporine That Exhibits Potent Inhibition of Hepatitis C Virus RNA Replication In Vitro

Physiological and Molecular Characterization of Aristolochic Acid Transport by the Kidney

Bovine Serum Albumin Decreases K_m Values of Human UDP-Glucuronosyltransferases 1A9 and 2B7 and Increases V_max Values of UGT1A9

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Modification of the ultrafiltration technique to overcome solubility and non-specific binding challenges associated with the measurement of plasma protein binding of corticosteroids

Cited by 80 publications

References 14 publications

SCY-635, a Novel Nonimmunosuppressive Analog of Cyclosporine That Exhibits Potent Inhibition of Hepatitis C Virus RNA Replication In Vitro

SCY-635, a Novel Nonimmunosuppressive Analog of Cyclosporine That Exhibits Potent Inhibition of Hepatitis C Virus RNA Replication In Vitro

Physiological and Molecular Characterization of Aristolochic Acid Transport by the Kidney

Bovine Serum Albumin Decreases Km Values of Human UDP-Glucuronosyltransferases 1A9 and 2B7 and Increases Vmax Values of UGT1A9

Contact Info

Product

Resources

About

Bovine Serum Albumin Decreases K_m Values of Human UDP-Glucuronosyltransferases 1A9 and 2B7 and Increases V_max Values of UGT1A9