The Competitive Counterflow Assay for Identifying Drugs Transported by Solute Carriers: Principle, Applications, Challenges/Limits, and Perspectives

The identification of substrates for solute carriers (SLCs) handling drugs is an important challenge, owing to the major implication of these plasma membrane transporters in pharmacokinetics and drug-drug interactions. In this context, the competitive counterflow (CCF) assay has been proposed as a practical and less expensive approach than the reference functional uptake assays for discriminating SLC substrates and non-substrates. The present article was designed to summarize and discuss key-findings about the CCF assay, including its principle, applications, challenges and limits, and perspectives. The CCF assay is based on the decrease of the steady-state accumulation of a tracer substrate in SLC-positive cells, caused by candidate substrates. Reviewed data highlight the fact that the CCF assay has been used to identify substrates and non-substrates for organic cation transporters (OCTs), organic anion transporters (OATs), and organic anion transporting polypeptides (OATPs). The performance values of the CCF assay, calculated from available CCF study data compared with reference functional uptake assay data, are, however, rather mitigated, indicating that the predictability of the CCF method for assessing SLC-mediated transportability of drugs is currently not optimal. Further studies, notably aimed at standardizing the CCF assay and developing CCF-based high-throughput approaches, are therefore required in order to fully precise the interest and relevance of the CCF assay for identifying substrates and non-substrates of SLCs.