In Vitro Drug Interaction Studies
 

Cytochrome P450 Enzyme- and Transporter-Mediated Drug Interactions 
 

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This final guidance provides recommendations for ǯǻǺǰȁǯȀǵǺdz and evaluating studies to determine 

the drug-drug interaction (DDI) potential of an investigational drug product.˃ This guidance focuses

on in vitro approaches to evaluate the interaction potential between investigational drugs with 

cytochrome P450 enzymes (CYPs) and transporters as well as how in vitro results can inform future 

clinical DDI studies.2 

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Why is this guidance important? 

Patients frequently use more than one medication at a time. Unanticipated, unrecognized, or mismanaged 

DDIs are an important cause of morbidity and mortality associated with prescription drug use and have 

occasionally caused the withdrawal of approved drugs from the market. This guidance outlines a general 

framework for conducting in vitro experiments and interpreting in vitro study results to determine the 

potential for clinical DDIs. 

Why does the guidance focus on cytochrome P450 enzyme 

and transporter-mediated drug interactions? 

CYP enzyme and transporter-mediated drug interactions are some of the most common mechanisms for affecting drug 

absorption, distribution, metabolism, and excretion (ADME) that can cause a DDI. Drug metabolism primarily occurs in 

the liver and intestinethrough the CYP family of enzymes. Membrane transporters can have clinically relevant effects 

on the pharmacokinetics and pharmacodynamics of a drug in various organs and tissues by ǭDzDzDZǯȀǵǺdz its absorption, 

distribution, and elimination. 

In vitro evaluation 

of the DDI potential of 

an investigational new 

drug should involve 

1 ǘǻǾǭǰǰǵȀǵǻǺǭǸǵǺDzǻǾǹǭȀǵǻǺ ̇ǿȁǯǴǭǿDZȄǭǹǼǸDZǿǻDzǿȁǮǿȀǾǭȀDZǿ ̇ǵǺǴǵǮǵȀǻǾǿ ̇ǭǺǰǵǺǰȁǯDZǾǿǻDzǯȅȀǻǯǴǾǻǹDZǢˆˇ˂DZǺȆȅǹDZǿǭǺǰȀǾǭǺǿǼǻǾȀDZǾǿ ̇ǼǸDZǭǿDZǿDZDZȀǴDZDzǻǸǸǻȃǵǺdzȃDZǮǿǵȀDZ: https://www.fda.gov/drugs/drug-interactions-labeling/drug-development-and-drug-interactions. 2 ǛDzǭǺǵǺȂǵȀǾǻǭǿǿDZǿǿǹDZǺȀǿȁdzdzDZǿȀǿǭǯǸǵǺǵǯǭǸǖǖǛǿȀȁǰȅǿǴǻȁǸǰǮDZǯǻǺǰȁǯȀDZǰ ̇ǿǼǻǺǿǻǾǿǿǴǻȁǸǰǾDZDzDZǾȀǻȀǴDZ˄˂˄˂DzǵǺǭǸǘǖǓdzȁǵǰǭǺǯDZDzǻǾǵǺǰȁǿȀǾȅDZǺȀǵȀǸDZǰǕǸǵǺǵǯǭǸǖǾȁdzǛǺȀDZǾǭǯȀǵǻǺǥȀȁǰǵDZǿ˱ǕȅȀǻǯǴǾǻǹDZǢˆˇ˂ǗǺȆȅǹDZ˭ǭǺǰǦǾǭǺǿǼǻǾȀDZǾ˭ǟDZǰǵǭȀDZǰǖǾȁdzǛǺȀDZǾǭǯȀǵǻǺǿ ̆

To learn more about in vitro cytochrome P450 enzyme and transporter-mediated drug interaction studies, read the guidance: 

https://www.fda.gov/media/134582/download 

the following: 

Ǜnforming the likelihood of

clinical DDI as well as the need 

for clinical DDI studies 

Ǘstimating the contribution

of enzymes and transporters 

to the drug’s disposition 

Ǖharacterizing the effect

of the drug on enzymes

and transporters 

1 2 3 
 

Taking more than one drug at the same time can alter the processes 

by which one or more of drugs is absorbed, distributed, metabolized, 

or excreted. This alteration is referred to as a drug interaction and 

can modify the drug’s effectiveness or cause unexpected side effects. 

During drug development, it is not feasible to evaluate interaction 

with every potential coadministered drug in clinical trials. Therefore, 

evaluation often starts with in vitro experiments to identify mechanisms underlying potential DDIs. This guidance outlines a general 

framework for conducting in vitro experiments and interpreting in 

vitro study results to determine the potential for clinical DDIs. 

Guidance Snapshots are a communication tool and are not a substitute for the guidance document. To 

learn more about in vitro drug interaction studies, read the guidance:

https://www.fda.gov/media/134582/download 

Background About the Guidance 

Guidance Recap Podcast – Hear 

Highlights Straight From FDA Staff 

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Sponsors should conduct JOWitro experiments to identify potential factors influencing drug 

disposition to elucidate potential DDI mechanisms and to yield kinetic parameters for use in 

further analyses. Results of in vitro experiments, along with clinical PK datBBOE mechanistic 

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can inform the need for and proper design of potential future clinical studies. 

Drug Development Timeline – When to Apply 

the Guidance Recommendations? 

Basic 

Research 

Prototype 

Design or 

Discovery 

Preclinical 

Development 

Clinical Development 

Phase I Phase 2 Phase 3 

FDA Filing/ 

Approval 

Post-Market/ 

Post-Approval 

Studies 

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