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Dietary Supplements: A Framework for Evaluating Safety
likely to cause interactions. These in vitro studies and other approaches have focused on determining which drugs affect metabolizing enzymes and transporters and could similarly be used to determine which dietary supplements may lead to interactions. Whether an interaction predicted on the basis of in vitro studies actually occurs clinically will depend on whether the dietary supplement compound attains a concentration in vivo adequate to reproduce the effect observed in vitro, as discussed in more detail below.
In Vitro Prediction of Pharmacokinetic Effects
In vitro studies for determining which xenobiotics affect transporters and metabolic enzymes ideally employ human transporter proteins or human metabolic enzymes. For example, subcellular fractions of human liver tissue are commonly used, as are whole-cell models such as isolated human hepatocytes (Li, 1997; Sinz, 1999), liver slices (Ferrero and Brendel, 1997), and cell lines derived from human cancer cells (Yee and Day, 1999). Human transporters and enzymes can also effectively be studied by expressing them in other cell types (Crespi and Penman, 1997; Rodrigues, 1999). Changes in either the activity or amount of enzyme or transporter are detected with activity assays, pharmacological assays, and immunochemical or mRNA assays that detect changes in protein or transcription (Li, 1997).
In vitro assays for predicting possible interactions are a well-accepted staple of the drug development process. The limitation to using these assays to predict clinical interactions lies, like most in vitro assays, in relating the dose at which enzyme or transporter effects are observed with the amount of unbound xenobiotic present at the active site in vivo. If information about the concentration of xenobiotic reached in vivo is available, a comparison of a dietary supplement ingredient’s inhibitory binding constants (Ki) for the CYP enzymes and the in vivo concentration (Cmax) may place the in vitro information in the appropriate perspective.
Animal and Human In Vivo Data in Predicting Pharmacokinetic Effects
Given the inter- and intraspecies differences in xenobiotic metabolizing enzymes, it is ideal to study xenobiotic metabolism using human cells, subcellular fractions of human tissue, or heterologously expressed human proteins (see Health Canada, 2000), although information about effects on animal proteins may serve as a preliminary indicator of concern. The study of human proteins in transgenic animals may improve ability to relate effects observed in animals or animal cells to humans.
Humans themselves may also be studied to determine if a given xenobiotic may cause an observable interaction. Such tests are usually designed to compare the levels of a test substrate with and without the