Permeability & Transporter Services

Drug-transporter interactions are of significant importance in drug discovery and development. These interactions affect all aspects of absorption, distribution, metabolism, excretion and toxicity and can also be the cause of clinically relevant drug-drug interactions.

The USA Food and Drug Administration (FDA) guidance (In Vitro Metabolism and Transporter-Mediated Drug-Drug Interaction Studies, Guidance for Industry, draft October 2017)[1] and European Medicines Agency (EMA) guidance (Guideline on the Investigation of Drug Interactions, adopted 2012)[2] recommend the assessment of drug-transporter interactions in vitro and their translation into the planning of clinical drug-drug interaction studies.

Identifying potential interactions early in drug discovery can help clients make the right compound selection decisions and mitigate any potential issues prior to candidate selection or clinical trials. XenoGesis has extensive experience in this field and offers a range of assays to assess drug-transporter interactions in vitro including:

  • cell-free vesicle-based assays
  • cell lines expressing individual transporters in monolayer or suspension cultures
  • polarised cells expressing combinations of uptake and efflux transporters (Caco 2, MDCK-II)
  • hepatic uptake using primary hepatocytes

Investigational drug as a transporter substrate

The FDA guidelines state that “Sponsors should evaluate most investigational drugs in vitro to determine whether they are substrates of P-gp and BCRP.”

The FDA and EMA state that a drug should be investigated as a substrate of OATP1B1 and/or OATP1B3 when hepatic metabolism and/or biliary excretion constitute ≥ 25% of total drug clearance.

The FDA recommends that drugs that are ≥ 25% eliminated via active renal secretion should be investigated as potential substrates of OAT1, OAT3, OCT2, MATE1 and MATE2-K. The EMA guidelines recommend that if an investigational drug is or may be ≥ 25% eliminated through renal, biliary or intestinal secretion, the candidate main transporter(s) involved should be identified through in vitro studies.

Other transporters should be investigated as and when their involvement appears likely based on experience from related compounds or the scientific literature. XenoGesis works with clients to help identify the transporters relevant for each project.

Investigational drug as a transporter inhibitor

As with the FDA guidelines, the EMA recommends evaluating whether an investigational drug is an inhibitor of P-gp, BCRP, OATP1B1, OATP1B3, OAT1, OAT3 and OCT2. In addition, inhibition studies with OCT1, MATE1, MATE2 and BSEP should be considered.

Transporter Assays Offered

SLC Transporters

Most solute carrier (SLC) transporters are secondary active transporters and use the electrochemical potential stored in ion gradients such as sodium, protons or bicarbonate.

TransporterRecommended by
OATP1B1EMA | FDA
OATP1B3EMA | FDA
OAT1EMA | FDA
OAT2
OAT3EMA | FDA
OCT1EMA
OCT2EMA | FDA
OCTN2
MATE1EMA
MATE2-kEMA
OATP1A2
OATP2B1
PepT1
PepT2
NTCP

ABC transporters

ATP-binding cassette (ABC) transporters are primary active transporters that use ATP as an energy source in order to translocate their substrates across biological membranes and can generate huge concentration gradients.

TransporterRecommended by
MDR1 (P-gp)EMA | FDA
BCRPEMA | FDA
BSEPEMA
MRP1
MRP2
MRP3

[1] https://www.fda.gov/regulatory-information/search-fda-guidance-documents/vitro-metabolism-and-transporter-mediated-drug-drug-interaction-studies-guidance-industry

[2] https://www.ema.europa.eu/en/documents/scientific-guideline/guideline-investigation-drug-interactions_en.pdf

Drug transporters: An overview
Drug transporters belong to three categories of transport proteins: ATP-binding cassette (ABC) transporters, solute carriers (SLC) and P-type ATPases.


ABC transporters
Most eukaryotic ABC transporters are efflux pumps. Exceptions are the cystic fibrosis transmembrane conductance regulator (CFTR) and the sulfonylurea receptors (SUR), which are ion channels. ABC transporters are primary active transporters, i.e. they use ATP as an energy source in order to translocate their substrates across biological membranes and can generate huge concentration gradients. The ATP binding motifs have been highly preserved during evolution and can, therefore, be used to identify ABC superfamily members. ABC transporters of particular pharmacological interest include P-glycoprotein (P-gp), also known as multidrug resistance protein 1 (MDR1), which is the first member of the ABCB family (gene symbol ABCB1). Other ABC transporters, for which an involvement in drug ADMET has been shown, are the Multidrug Resistance-Associated Proteins 1 (MRP1, ABCC1) and MRP2 (ABCC2), also known as canalicular Multi-specific Organic Anion Transporter (cMOAT), the Breast Cancer Resistance Protein (BCRP, ABCG2) and the Bile Salt Export Pump (BSEP, ABCB11).


SLC transporters
Most SLC transporters are secondary active transporters, i.e. they use the electrochemical potential stored in ion gradients such as sodium, protons or bicarbonate. Like primary active transporters, secondary active transporters can pump their substrates against a concentration gradient. Exceptions are the equilibrative nucleoside transporters (ENTs, SLC29).

SLC transporters that are of particular interest in drug R&D include the organic anion transporting polypeptides (OATPs), organic anion transporters (OATs), organic cation transporters (OCTs), the bile salt uptake transporters NTCP and ASBT as well as the multidrug and toxin extrusion transporters (MATEs).


P-type ATPases
P-type ATPases are an evolutionary very old class of proteins that translocate ions and lipids across biological membranes. Like ABC transporters, the P-type ATPases are primary active transporters.

Two members, namely ATP7A and ATP7B are involved in the detoxification of copper. Both these proteins have been shown to transport the platinum-containing cancer drugs cisplatin, carboplatin and oxaliplatin and might, therefore, play a role in drug resistance.

FDA guidelines

“Development of a drug should include identification of the principal routes of elimination, quantitation of the contribution by enzymes and transporters to drug disposition, and characterization of the mechanism of drug-drug interactions.”

FDA Guidance for Industry, Drug Interaction Studies —Study Design, Data Analysis, Implications for Dosing, and Labeling Recommendations.

Investigational drug as a transporter substrate

The current American Food and Drug Administration (FDA) draft guidelines suggest that “all investigational drugs should be evaluated in vitro to determine whether they are a potential substrate of P-gp and BCRP”.

When hepatic metabolism and/or biliary excretion constitute ≥25% of total drug clearance, then it should be investigated if this drug is a substrate of OATP1B1 and/or OATP1B3.”

Similarly, drugs that are ≥25% eliminated via active renal secretion should be investigated as potential substrates of OAT1, OAT3 and OCT2.

Other transporters should be investigated as and when their involvement appears likely based on experience from related compounds or the scientific literature.

Investigational drug as a transporter inhibitor

The current FDA draft guidelines also recommend clarifying whether an investigational drug is an inhibitor of all the above transporters.

http://www.fda.gov/Drugs/DevelopmentApprovalProcess/DevelopmentResources/DrugInteractionsLabeling/ucm093664.htm

EMA guidelines

In vitro inhibition studies are recommended to investigate whether the investigational drug inhibits any of the transporters known to be involved in clinically relevant in vivo drug interaction studies.”

European Medicines Agency: Guideline on the investigation of drug interactions (2012)

Investigational drug as a transporter substrate

In agreement with the FDA guidance, the EMA guidance recommends investigating whether an investigational drug is a substrate of OATP1B1 and OATP1B3, if it is ≥25% eliminated hepatically.

If an investigational drug is or may be ≥25% eliminated through renal, biliary or intestinal secretion, then the candidate main transporter(s) involved should be identified through in vitro studies.

Investigational drug as a transporter inhibitor

Like the FDA guideline, the EMA guideline suggests exploring whether an investigational drug is an inhibitor of P-gp, BCRP, OATP1B1, OATP1B3, OAT1, OAT3 and OCT2. In addition, inhibition studies with OCT1, MATE1 and MATE2 and BSEP should be considered.