Pharmacogenetics

Julien De Greef, Leïla Blekhir, Laure Elens (LDRI) and Vincent Haufroid

Pharmacogenetics (PGx), and by extension pharmacogenomics, is the study of how our genes affect clinical response to drugs. In this definition, clinical response should be considered in a broad sense and include both drug efficacy and adverse drug reactions (ADRs).This relatively new field combines pharmacology (the science of drugs) and genomics (the study of genes and their functions) to propose drug and/or dose recommendations that will be tailored to a person’s genetic makeup

Our research in PGx at the Louvain centre for Toxicology and Applied Pharmacology (LTAP) is translational in essence. We combine analytical developments (for Therapeutic Drug Monitoring purposes, TDM), pharmacokinetics (PK) modeling, and we use clinical samples and in vitro cellular models.

We are particularly interested in a few pharmacotherapeutic areas including immunosuppressive drugs used in transplantation, antiretrovirals used in HIV therapy, and anticancer drugs.

As an illustration, our research work carried out in the field of HIV therapy is briefly summarised below:

In previous works, carried out in the framework of two doctoral theses (L. Elens and L. Belkhir), we measured plasma and/or intracellular concentrations of several human immunodeficiency virus (HIV) antiretrovirals (ARVs). We used these PK data for PGx studies to predict the clinical response to treatment, both in terms of tolerance (reduction of side effects) but also in terms of therapeutic efficacy (better drug penetration at the target level, viral load reduction, restoring or improving immunological function).

As a first step, we have developed and validated analytical quantification tools to measure the concentration of several ARVs in the plasma of HIV-positive patients but also directly at their site of pharmacological action, i.e. in the mononuclear fraction of blood cells. An original method for the quantification of 10 ARVs in plasma by liquid chromatography with ultra-violet diode array detection (LC-DAD) has been published in the best journal in the field of clinical chemistry [1]. An even more original method for the quantification of ARVs in peripheral blood mononuclear cells (PBMCs) by liquid chromatography with tandem mass spectrometry (LC-MS/MS) has also been published [2],[3]. All patients treated at CUSL or in other institutions can now benefit from this therapeutic drug monitoring (TDM) allowing a posteriori dose-adjustments in order to avoid sub-therapeutic concentrations (risk of emergence of resistance) or concentrations considered as toxic or at higher risk of side effects.

These tools subsequently made it possible to carry out PGx clinical studies designed to identify genetic factors influencing plasma concentrations and/or intracellular accumulation (and hence the therapeutic activity) of ARVs.

As part of Laure Elens' thesis, we have identified a genetic factor highly predictive for efavirenz (EFV) toxicity, i.e. a polymorphism in CYP2B6 responsible for the biotransformation of EFV into its inactive metabolite. This study was published in one of the reference journals in the field of pharmacogenetics [4]. Screening for this genetic factor of toxicity (associated with poor compliance) can now be implemented, either prospectively with the therapeutic option of drug substitution or dose reduction in patients carrying the allele associated with a decrease in CYP2B6 activity, or retrospectively when neurological symptoms appear. We have also been able to demonstrate that the intracellular accumulation of lopinavir (LPV), a protease inhibitor (PI), was influenced by a polymorphism in the gene coding for an efflux protein (MRP2 or ABCC2) expressed on the surface of mononuclear cells, and lymphocytes in particular. Interestingly, these effects, observed during a clinical study performed in HIV-positive patients treated at CUSL and published in another reference journal in the field of pharmacogenetics [5], have been validated subsequently in an experimental study using cell lines stably expressing the relevant efflux protein presenting with (or not, wild-type protein) the identified genetic polymorphism [6]. This work was a very nice example of translational research from bed to bench.

A few years later, as part of Leila Belkhir's thesis, we were able to demonstrate, for the first time ever, that the drug interaction between etravirine (ETV) and darunavir (DRV) is mainly mediated by genetic factors involving a variant in the gene coding for a biotransformation enzyme, CYP3A5. Indeed, a CYP-enzyme inducer such as ETV exerts a different action depending on whether the patient is genetically expressing CYP3A5 or not [7]. The mechanism that we have elucidated allows understanding why such an interaction had never been demonstrated in patients of Caucasian origin (mainly CYP3A5 non-expressers), whereas this effect can become very significant in patients expressing the CYP3A5 enzyme (the majority of patients of African origin). CYP3A5 expressers have an increased risk of infra-therapeutic concentrations of DRV and, therefore, an increased risk of virologic failure, and emergence of drug-resistant viruses.

We have also demonstrated that genetic variants associated with a decrease in glucuronoconjugation activity (UGT1A1) have an impact on the plasma concentrations of raltegravir (RAL) and its RAL-Glu metabolite, making UGT1A1-deficient patients more sensitive to the side effects of these drugs. This work was published in Scientific Reports [8]. These results have also opened doors for the understanding of the mechanisms explaining the appearance of important side effects in some patients treated with the most recent molecules of this pharmacological class, such as dolutegravir (DTG). This drug, as well as the latest molecules in the same pharmacological class (bictegravir and cabotegravir) are the focus of an ongoing research project (Julien DeGreef’s thesis).

Very recently, using population PK modeling and simulations (collaboration with the group of Laure Elens at LDRI), we explored the feasibility of alternative dosing regimens (e.g. short cycle therapy) for DRV/cobicistat in HIV-positive patients. This work has been published in the reference journal in the field of clinical PK [9]. Such popPK approach could also easily be implemented for the most recent ARVs.

We have also reviewed the main PGx indications in the field of ARV treatments, with particular attention to the cytochrome P450 system [10].