Luís Cláudio Lima

DRUG METABOLISM

Prakash et al.  Nucl Receptor Res (2015) 2: 101178

P450 FAMILY

Prakash et al.  Nucl Receptor Res (2015) 2: 101178

Ahmed et al. Genomics Proteomics Bioinformatics. (2016) 14(5):298-313
P450 Family: CYP2C subfamily

∼390 kb gene cluster on

chromosome 10q23.3

Backman et al. Pharmacological Reviews (2016) 68:168–241

 P450 Family: CYP2C subfamily
 It is also expressed in extrahepatic tissues (the kidney,
lung, nasal mucosa, arteries, endothelial mucosa, and
heart)
CYP2C SUBSTRATES

-anticancer drug paclitaxel

-antidiabetics drugs such

as rosiglitazone and
pioglitazone

Backman et al. Pharmacological Reviews (2016) 68:168–241

 P450 Family: CYP2C8 subfamily

 CYP2C8 metabolizes more than 60 clinically used drugs

as well as endogenous substances including retinoic acid
and arachidonic acid.
CYP2C8∗2

SNPs CYP2C8∗3

CYP2C8∗4
 CYP2C8 appears to be less strongly affected by genetic
variation and consequently regulatory events may have a
more significant impact on variability.
 CYP2C8
polimorphisms:
Global distribution

Backman et al. Pharmacological Reviews (2016) 68:168–241

P450 Family: CYP2C8 subfamily

Daily and Aquilante.Pharmacogenomics(2009) 10 (9):1489–1510

(Ile269Phe in exon 5)

(Arg139Lys and Lys399Arg

in exons 3 and 8)
(Ile264Met in exon 5)
 CYP2C Genes and
Transcriptional Regulation
 Human CYP2C8 is Transcriptionally Regulated

Receptors CYP2C genes

HNF4 CAR / PXR GR 

-peroxisome proliferator-activated
receptor alpha (PPARα)
CYP3A4
PPARα Pathway

P.D. McMullen et al. Chemico-Biological Interactions 209 (2014) 14–24

WNT/β-Catenin Pathway

W. Kim, M. Kim and E. h. Jho. Biochemical Journal. (2013) 450: 9–21

 PPARα and WNT/β-Catenin Pathway

PXR-mediated induction of
CYP3A4 gene and protein
expression.
WNT/β-catenin pathway 
 PPARα-mediated induction of
CYP1A, CYP2C8, CYP3A4,
and CYP4A11 genes
 HYPOTHESIS and OBJECTIVE 

I. Peroxisome proliferator-activated receptor alpha

(PPARα) directly regulates the transcription of
CYP3A4.

II. CYP2C8 shares more common substrates with

CYP3A4.

III. Hepatic expression of CYP2C8 is rather strongly

correlated to CYP3A4
 HYPOTHESIS and OBJECTIVE 

 To assess the impact of PPARα polymorphisms,

(rs4253728 and rs4823613) on the expression and
activity of CYP2C8.

 To investigate the potential direct regulation of

CYP2C8 by PPARα in human hepatocytes.

 To elucidate the molecular basis for the modulation

of PPARα-mediated effects on CYP2C8 by the
WNT/β-catenin pathway.
 METHODS
150 patients of Caucasian ethnicity (71 males and 79 females)
Liver tissue and blood samples

mRNA expression (CYP2C8 – Western Blot (CYP2C8 – Liver

HepaRG Transfections with siRNAs
Liver tissue and HepaRG cells) tissue and HepaRG cells)
(PPARα and β-catenin)

Transfections and Luciferase

Chromatin Cytochrome P450 Reporter Analyses
Fluorescence-based
Immunoprecipitation Assay enzyme activities  pGL3-PPRE pos
Electromobility Shift Assays
pGL3-CYP2C8-Site C and A
 RESULTS AND DISCUSSION

CYP2C8 variation appeared to be

somewhat less pronounced;
Population Variability of Hepatic CYP2C8
 RESULTS AND DISCUSSION

Non-genetic Factors Influencing CYP2C8 Expression

 RESULTS AND DISCUSSION

Correlation between PPARα expression and

CYP2C8 Phenotypes
 RESULTS AND DISCUSSION

rs4253728 and rs4823613 SNPs significantly decreased levels

of CYP2C8 protein expression in homozygous carriers of the
minor allele.
 RESULTS AND DISCUSSION

Funcional impact of PPARα on CYP2C8: in vitro

study (treatment of HepaRG)

PPARα mediates both basal and inducible regulation

of CYP2C8 in human hepatocytes
 RESULTS AND DISCUSSION

Binding of PPARα to the CYP2C8 promoter in silico

 RESULTS AND DISCUSSION

Fluorescence-based
Electromobility Shift Assays

PPARα Directly Binds to the

Specific Motifs of CYP2C8
Promoter and Activates its
Expression.
 RESULTS AND DISCUSSION

In HepaRG cells β-catenin exerts a similar inhibitory

modulation on PPARα-mediated CYP2C8 induction
as previously shown for CYP3A4 regulation;
 CONCLUSION

 Their experiments suggested both constitutive and

inducible transactivation of CYP2C8 by PPARα.

 PPARα thus contributes to a transcriptional

network so far including CAR, PXR, GR, and
HNF4 as direct regulators of CYP2C8 expression,
while inducibility is also modulated via WNT/β-
catenin pathway.
OBRIGADO!