Genetic variants may influence the function of a gene:
- (i) directly, by introduction of loss of function (LOF) mutations, or missense mutation causing amino acid exchange
- (ii) indirectly, by modifying the abundance of the gene product (mRNA expression), thereby affecting protein expression levels.
The PK-phenotype can also be determined by multiple variations in different alleles, e.g. for medicinal products metabolized by multiple pathways. The detection of specific genetic variants in clinical routine helps to improve efficacy and safety of specific medicinal products in use, and for that, the genotype in question ultimately needs to be translated into a predicted phenotype.
Whereas the gene analysis based prediction for the phenotype is usually straightforward for homozygous variants that result in a loss of function of the respective gene product, it can be more complex for heterozygous LOF genetic variations or missense mutations.
Thus, phenotype assignment from genotypes where genetic variation(s) only modestly alter (increase or decrease) the respective enzyme activity is more difficult.
Additional factors that affect prediction of phenotype from genotype are environmental as well as endogenous in nature i.e.
- (i) patient compliance,
- (ii) diet,
- (iii) bioavailability,
- (iv) hepatic blood flow and function,
- (v) renal function,
- (vi) co-medication and
- (vii) dependency on the particular metabolic pathway.
Phenotyping for a specific metabolizing enzyme by e.g. therapeutic drug monitoring at different time intervals after administration is a valid method for measuring the impact of genetic variance on metabolism.
Compared to genotyping alone, this approach has the advantage of directly assessing functionality including contributions of several different gene variations or rare mutations of importance for the PK. The in vivo assessment of a phenotype relies on the use of specific probe drugs.
For simultaneous determination of activities of multiple CYP enzymes, it is recommended to use a phenotyping drug (probe) cocktail, in which different metabolites formed by different CYP enzymes are individually quantified in blood or urine.
Appropriate probe substances should be used, i.e. a compound must fulfill certain criteria such as i) selectivity for the enzyme in question, ii) having a specific, targeted, quantifiable metabolite, iii) should be relatively non-toxic and iv) not interacting with the parent substance or the metabolite from the enzymatic conversions.
The metabolic phenotype can vary substantially over time in the same individual due to physiological, pathological or dietary factors which must be taken into account when doing therapeutic drug monitoring. Genotyping gives a stable, lifelong insight into the functional status of the enzymes in question.
However due to the important contribution of rare genetic variants in each individual therapeutic drug monitoring takes into account all genetic variations of importance for the PK assessed, whereas Targeted genotyping can miss individual specific genetic variations, since these were not part of the testing SNP panel analyzed. For more discussion on this topic see the Guideline on the use of pharmacogenetic methodologies in the pharmacokinetic evaluation of medicinal products (EMA/CHMP/37646/2009).
Read also: Common and Rare Genetic Variants