In vitro transporter assays
Understanding whether your compound interacts with drug transporters is an important stage in the drug development process. In vitro transporter interaction assays are used to identify if clinical drug-drug interaction studies are required.
Contact us now to discuss how Cyprotex can help you assess your compound’s potential for drug transporter interactions.
The white paper published in Nature Reviews in Drug Discovery (March 2010)1 by the International Transporter Consortium and the draft FDA guidance for industry on drug interaction studies (Feb 2012)2 provides recommendations on the seven most relevant drug transporters for evaluation in the drug discovery and development process:
The European Medicines Agency (EMA) Draft Guideline on the Investigation of Drug Interactions (April 2010)3 also recommends evaluating an additional two transporters:
In addition to these broadly recommended transporter studies, there are a number of other potentially clinically relevant transporters which may be important for particular drug-discovery programmes. Cyprotex can also help you with these. Please contact us.
Cyprotex provides an extensive portfolio of drug transporter services through a combination of transporter assays performed at our own laboratories and those performed at the laboratories of our partner, Solvo Biotechnology. Regardless of the location in which your assays are performed, all of your assays are managed seamlessly for you by your Cyprotex Study Manager.
Contact us now to discuss how Cyprotex can help you assess your compound’s potential for drug transporter interactions.
P-gp Interactions
P-gp (P-glycoprotein) is one of the most well-recognised efflux transporters. It is expressed in many tissues, including the intestine, brain, and kidney. P-gp inhibition has been shown to be responsible for several clinical drug-drug interactions. For example, clarithromycin can inhibit the transport of the P-gp substrate digoxin, resulting in an elevation of plasma levels and a decrease in renal clearance4.
The FDA has outlined recommendations for studying P-gp inhibition and substrate identification in its Draft Guidance for Industry, Drug Interaction Studies – Study Design, Data Analysis, Implications for Dosing, and Labeling Recommendations, 20122. This guidance follows the recommendations issued in the March 2010 white paper on drug transporters published in Nature Reviews in Drug Discovery by the International Transporter Consortium1. Cyprotex follow the recommended decision tree for identifying if an in vivo drug-drug interaction study is required using FDA-recommended in vitro protocols.
Cyprotex provide a P-gp substrate identification assay using bidirectional Caco-2 in the presence of the P-gp inhibitor, verapamil. We also offer bidirectional drug transport services in MDR1-MDCK cells to identify P-gp substrates and to screen for P-gp inhibitors.
BCRP Interactions
BCRP (Breast Cancer Resistance Protein) is an efflux transporter expressed in several tissues such as the gastrointestinal tract, liver, brain endothelium, mammary tissue, testis, and placenta. BCRP is known to play a role in clinical drug-drug interactions. In addition, clinically relevant genetic polymorphisms have been shown to impact on the PK (e.g., irinotecan5, rosuvastatin6, sulfasalasine7 and topotecan8) and toxicity (e.g., gefitinib-induced diarrhoea9) of marketed drugs. The International Transporter Consortium1 and the draft FDA guidance2 recommend investigating BCRP due to BCRP’s clinical importance in the absorption and disposition of drugs.
The International Transporter Consortium1 and the draft FDA guidance2 recommend using Caco-2 to study BCRP interactions. Cyprotex identify BCRP substrates by investigating bidirectional Caco-2 permeability in the presence of a selective BCRP inhibitor, fumitremorgin C.
OATP1B1 Interactions
Organic Anion Transporting Polypeptide 1B1 (OATP1B1) is expressed on the sinusoidal membrane of hepatocytes where it is responsible for the uptake of several marketed drugs including some statins10. There is evidence for clinical drug-drug interactions involving cyclosporine which appear to be mediated, at least in part, by inhibition of OATP1B111, 12.
The International Transporter Consortium1 and the draft FDA guidance2 recommend investigating for potential OATP1B1 substrates and inhibitors due to the role of OATP1B1 in drug-drug interactions and the impact of genetic polymorphism of this transporter on therapy outcome and toxicity. It is only necessary to evaluate potential OATP1B1 substrates when hepatic clearance of the investigational drug is significant (e.g., clearance through hepatic or biliary clearance is more than or equal to 25% of the total clearance).
OATP1B3 Interactions
Organic Anion Transporting Polypeptide 1B3 (OATP1B3) is an uptake transporter expressed on the sinusoidal membrane of hepatocytes. It has a substrate specificity that overlaps somewhat with OATP1B113. Because of the prominent expression of these transporters on the basolateral membrane of hepatocytes, they represent a critical mechanism for chemical uptake into the liver.
The International Transporter Consortium1 and the draft FDA guidance2 recommend the evaluation of new chemical entities for their potential to act as substrates or inhibitors of OATP1B3 in vitro. It is only necessary to evaluate potential OATP1B3 substrates when hepatic clearance of the investigational drug is significant (e.g., clearance through hepatic or biliary clearance is more than or equal to 25% of the total clearance).
OCT2 Interactions
Organic Cation Transporter 2 (OCT2) is a member of the SLC family of transporters (SLC22A2). This transporter is expressed on the cells of the kidney proximal tubules where it is involved in the renal clearance of drug substrates14. Drug-drug interactions involving OCT2 may result in decreased renal clearance of the victim drug and a corresponding increase in exposure (e.g., cimetidine interaction with metformin15).
The International Transporter Consortium1 and the draft FDA guidance2 recommend the use of cell lines expressing recombinant OCT2 to study OCT2 interactions in an in vitro setting. It is only necessary to evaluate potential OCT2 substrates when renal active secretion of the investigational drug is significant (e.g., active secretion by the kidney is more than or equal to 25% of total clearance).
OAT1 Interactions
Organic Anion Transporter 1 (OAT1) is part of the SLC superfamily (SLC22A6). It is a transmembrane protein expressed predominantly in the basolateral membrane of proximal tubular cells of the kidneys. It plays a central role in renal organic anion transport16. OAT1 is involved in the uptake of a wide range of relatively small and hydrophilic organic anions from plasma into the cytoplasm of the proximal tubular cells of the kidneys for subsequent exit across the apical membrane for excretion via the urine17. It has an essential role in the disposition of NSAIDs, antiviral drugs, diuretics, antitumour drugs and ß-lactam antibiotics16.
The International Transporter Consortium1 and the draft FDA guidance2 recommend the use of transfected cell lines to investigate if new chemical entities are substrate or inhibitors of OAT1. It is only necessary to evaluate potential OAT1 substrates when renal active secretion of the investigational drug is significant (e.g., active secretion by the kidney is more than or equal to 25% of total clearance).
OAT3 Interactions
Organic Anion Transporter 3 (OAT3) is part of the SLC superfamily (SLC22A8). Like OAT1, it is primarily expressed in the basolateral membrane of proximal tubular cells of the kidneys, facilitating its role is in the renal transport of organic anions16.
OAT3 exhibits a broader substrate specificity than OAT1, and accepts amphipathic and hydrophilic organic anions and some organic cations16. Drugs which are renally cleared and are actively secreted by OATs may be susceptible to increases in AUC as a result of reduced clearance. Examples of these clinically relevant interactions include the interaction of probenecid with acyclovir, resulting in a 32% decline in renal clearance, a 40% increase in AUC, and an 18% increase in the terminal plasma half-life18.
The International Transporter Consortium1 and the draft FDA guidance2 recommend screening for drug interaction potential using cell lines expressing OAT3. It is only necessary to evaluate potential OAT3 substrates when renal active secretion of the investigational drug is significant (e.g., active secretion by the kidney is more than or equal to 25% of total clearance).
BSEP Interactions
The Bile Salt Export Pump (BSEP) is a member of the ATP binding cassette family of transporters and is located on the canalicular membrane of hepatocytes. It is involved in transport of taurocholate and other cholate conjugates from hepatocytes to the bile and plays an important function in bile formation and bile flow19.
Evaluating the inhibitory potential of new chemical entities on the transporter BSEP is recommended by the European Medicines Agency3 for detecting pharmacodynamic interactions as well as for adequate safety monitoring during drug development.
OCT1 Interactions
Organic Cation Transporter 1 (OCT1) is a member of the SLC superfamily (SLC22A1) and is located on the basolateral membrane of hepatocytes, enterocytes, and renal proximal tubular cells20. It mediates facilitated transport of small (hydrophilic) organic cations21. The OCTs have been implicated in several clinically relevant drug interactions. For example co-administration of cimetidine with metformin increased the AUC of metformin by 50% and reduced the renal clearance by 27%15.
The European Medicines Agency3 recommends evaluating the potential of new chemical entities for their ability to inhibit OCT1 in vitro.
Contact us now to discuss how Cyprotex can help you assess your compound’s potential for drug transporter interactions.
References
1 The International Transporter Consortium (2010) Nat Rev Drug Disc 9; 215–236
2 Draft FDA Guidance for Industry – Drug Interaction Studies – Study Design, Data Analysis, Implications for Dosing, and Labeling Recommendations, February 2012
3 The European Medicines Agency (EMA) Draft Guideline on the Investigation of Drug Interactions (2010)
4 Wakasugi H et al. (1998) Clin Pharmacol Ther 64; 123–128
5 Zhou Q et al. (2005) Br J Clin Pharmacol 59; 415–424
6 Zhang W et al. (2006) Clin Chim Acta 373; 99–103
7 Yamasaki Y et al. (2008) Clin Pharmacol Ther 84(1); 95–103.
8 Sparreboom A et al. (2005) Cancer Biol Ther 4; 650–658
9 Cusatis G et al. (2006) J Natl Cancer Inst 98; 1739–1742
10 Chen C et al. (2005) Drug Metab Dispos 33; 537–546
11 Neuvonen PJ et al. (2006) Clin Pharmacol Ther 80; 565–581
12 Shitara Y et al, (2003) J Pharmacol Exp Ther 304; 610-616
13 Klaassen CD & Aleksunes LM (2010) Pharmacol Rev 62(1); 1–96
14 Aoki M et al., (2008) Am J Physiol Renal Physiol 295; F165-F170
15 Somogyi A et al. (1987) Br J Clin Pharmacol 23; 545-551
16 Nozaki Y et al., (2007) J Pharmacol Exp Ther 321; 362–369
17 El-Sheikh AAK et al., (2008) Eur J Pharmacol 585; 245-255
18 Laskin OL et al., (1982) Antimicrob Agents Chemother 21(5); 804-7
19 Stieger B et al., (2007) Pflügers Archiv Eur J Physiol 453; 611-620
20 Jonker JW and Schinkel AH (2004) J Pharmacol Exp Ther 308(1); 2-9
21 Jonker JW et al, (2001) Mol Cell Biol 21(16); 5471–5477
By Dr. Katya Tsaioun, Chief Scientific Officer, Cyprotex
Wiley & Sons, March 2011
