Caroline Fradette

Pomelo juice, but not cranberry juice, affects the pharmacokinetics of cyclosporine in humans

Cyclosporine (INN, ciclosporin) is a cytochrome P450 (CYP) 3A and P‐glycoprotein (P‐gp) substrate whose bioavailability increases when administered with grapefruit juice. It is unknown whether pomelo, a closely related citrus fruit, interacts with cyclosporine in humans. In addition, a case study reports that cranberry juice interacts with warfarin, a drug with a narrow therapeutic range. Cranberries have a high content of flavonoids, compounds with various metabolic effects, including interaction with P‐gp in vitro. Although the effect of flavonoids is less evident in vivo, cranberry juice has become a very popular beverage, and it was deemed important to investigate whether it has an effect on the disposition of cyclosporine, another drug with a narrow therapeutic range.

Animal models of acute moderate hypoxia are associated with a down- regulation of CYP1A1, 1A2, 2B4, 2C5 and 2C16 and up-regulation of CYP3A6 and P-glycoprotein in liver

In humans, indirect evidence suggests that hypoxia reduces the rate of biotransformation of drugs cleared by cytochrome P450 (P450) subfamilies CYP1A, 2B, and 2C. The aim of this study was to assess whether acute moderate hypoxia modulates the expression of CYP2B4, 2C5, and 2C16 in vivo, and to determine whether the changes in hepatic P450 are conveyed by serum mediators. Moreover, because hypoxia increases the expression of P-glycoprotein in vitro, we examined whether in vivo acute moderate hypoxia modulates the expression of several membrane transporters in the liver. Rabbits and rats were exposed to a fractional concentration of oxygen of 8% for 48 h to generate a stable arterial partial pressure of O2 of 34 ± 1 mm Hg. Compared with rabbits breathing room air, hypoxia in rabbits reduced the amount of CYP1A1, 1A2, 2B4, 2C5, and 2C16 proteins and increased the expression of CYP3A6. Sera of rabbits with hypoxia were fractionated by size exclusion chromatography, the fractions were tested for their ability to modify the expression of P450 isoforms, and serum mediators were identified through neutralization experiments. The serum mediators responsible for the down-regulation of P450 isoforms were interferon-γ, interleukin-1β (IL-1β), and IL-2. In vivo, in rats, hypoxia increased the mRNA and protein expression of P-glycoprotein but did not affect the mRNA of breast cancer resistance protein and organic anion-transporting polypeptide 2. It is concluded that in vivo, hypoxia down-regulates rabbit hepatic CYP1A1, 1A2, 2B4, 2C5, and 2C16 and up-regulates CYP3A6. CYP3A11 and P-glycoprotein were up-regulated in the livers of hypoxic rats.

Hypoxia-induced down-regulation of CYP1A1/1A2 and up-regulation of CYP3A6 involves serum mediators

Acute moderate hypoxia modifies the catalytic activity and expression of certain isoenzymes of hepatic cytochrome P450 (P450). The aim of this study was to document whether hypoxia affects hepatic P450 directly or through the release of serum mediators. Rabbits were subjected to a FiO2 of 8% for 48 h, sacrificed, and serum and hepatocytes were isolated; hepatocytes from control and rabbits with hypoxia were incubated with serum from control and hypoxic rabbits for 4 and 24 h, and total P450 content, CYP1A1, 1A2 and 3A6 activities and expressions were assessed. Sera were fractionated by size exclusion chromatography and fractions tested for their ability to modify activity and amount of P450, and serum mediators were identified through neutralization experiments. Total serum and fractions with proteins of 15–23 and 65–94 kDa of Mr reduced P450 content and expression of CYP1A1, 1A2 and 3A6, as well as CYP1A1, 1A2 and 3A6 mRNA. Total serum and the fraction with 32–44 kDa proteins increased CYP3A6 activity and protein and mRNA. The serum mediators implicated in the decrease in activity and expression of CYP1A1, 1A2 and 3A6 were interferon‐γ (IFN‐γ), interleukin‐1β (IL‐1β) and IL‐2. Erythropoietin (Epo) was partly responsible for the increase in P450 content and CYP3A6 expression. In conclusion, acute moderate hypoxia diminishes the activity and expression of CYP1A1, 1A2 and CYP1A1, 1A2 mRNA, and increases CYP3A6 protein, activity and CYP3A6 mRNA. Several mechanisms contribute to these changes in P450, among them the release of cytokines acting as serum mediators.

Hypoxia-inducible factor-1 and activator protein-1 modulate the upregulation of CYP3A6 induced by hypoxia

Moderate hypoxia in vivo and serum from rabbits subjected to moderate hypoxia (SHYPO) in vitro reduce CYP1A1 and 1A2 P450 isoforms and upregulate CYP3A6. The aim of this project was to investigate the signal transduction pathways implicated in the upregulation of CYP3A6 expression by hypoxia. Hypoxia in vivo and SHYPO in vitro increased the expression of hypoxia‐inducible factor‐1α (HIF‐1α) and c‐jun, as well as CYP3A6. By electrophoresis mobility shift assay, it was shown that HIF‐1 and activator protein‐1 (AP‐1) bind to CYP3A6 oligonucleotide probe after exposure to hypoxia in vivo and SHYPO in vitro. The effects of hypoxia in vivo or SHYPO in vitro were reproduced by CoCl2 and lead acetate, activators of HIF‐1 and AP‐1, respectively. PD98059, a p42/44 MAPK inhibitor, prevented the increase of CYP3A6 and c‐jun, but did not impede the increase of HIF‐1α and binding to CYP3A6 oligonucleotide probe. Genistein, an inhibitor of protein tyrosine kinases (PTKs), prevented the increase in HIF‐1α, c‐jun and CYP3A6, as well as HIF‐1 and AP‐1 binding to CYP3A6 oligonucleotide probe. Moreover, hypoxia in vivo induced constitutive androstane receptor (CAR) as well as CAR binding to the CYP3A6 oligonucleotide probe, but not the pregnane X receptor. In conclusion, hypoxia in vivo and SHYPO induce the expression of CYP3A6. The in vitro induction of CYP3A6 by SHYPO is PTK‐ and p42/44 MAPK‐dependent. The present data support the hypothesis that HIF‐1 and AP‐1 are part of the signalling pathway leading to CYP3A6 induction by hypoxia.

5-Hydroxytryptamine is biotransformed by CYP2C9, 2C19 and 2B6 to hydroxylamine, which is converted into nitric oxide.

There is circumstantial evidence suggesting that 5‐hydroxytryptamine (5‐HT) could be biotransformed by enzymatic systems other than monoamino oxidase A, and that the isoforms of cytochrome P450 may be a source of nitric oxide. This study aimed to assess whether cytochrome P450 contributes to 5‐HT biotransformation, and to provide evidence that 5‐HT metabolism generates nitric oxide. Addition of 5‐HT to cultured hepatocytes yielded 5‐hydroxyindol acetic acid, a formation modulated by cytochrome P450 enzyme inducers and inhibitors. Recombinant human CYP2B6, 2C9 and 2C19 biotransformed 5‐HT in 5‐hydroxyindol acetic acid, but not CYP1A2, 2D6 or 3A4. Cultured hepatocytes with 5‐HT generated nitric oxide, the amount of which was altered by cytochrome P450 enzyme inducers and inhibitors. In the presence of CYP2B6, 2C9 and 2C19, 5‐HT relaxed precontracted isolated aortic rings, with or without endothelium, an effect prevented by the addition of methylene blue and an inhibitor of catalase, but not by myoglobin. In the absence of catalase, hydroxylamine was always assayed as a byproduct of 5‐HT metabolism. In conclusion, CYP2B6, 2C9 and 2C19 biotransform 5‐HT, yielding hydroxylamine, which is converted to nitric oxide in the presence of catalase.

The utility of the population approach applied to bioequivalence in patients: comparison of 2 formulations of cyclosporine.

Mixed-effect modeling was used to compare the population pharmacokinetics of 2 formulations of cyclosporine in patients. An open-label, multicenter, conversion study in stable, 6-month post-renal allograft recipients was conducted to compare the safety and pharmacokinetics of oral Pliva Cyclosporine Soft Gelatin Capsules (USP Modified) with Neoral® (cyclosporine soft gelatin capsules, USP Modified) in stable post-renal transplant patients. Blood samples were collected predose and for 12 hours postdose on days 1, 14, 15, 28, and 29. Whole-blood samples were analyzed for cyclosporine using high-performance liquid chromatography and mass spectroscopy. Estimates of pharmacokinetic parameters were generated using noncompartmental and population compartmental pharmacokinetic analysis. Moreover, the effects of demographic factors on the pharmacokinetics of cyclosporine were evaluated using the nonlinear mixed-effects modeling program NONMEM®. The rate and extent of bioavailability of cyclosporine did not differ between Pliva Cyclosporine Soft Gelatin Capsules and Neoral®. In the final model, gender and actual body weight significantly affected the central and peripheral volumes of distribution. In addition, the pharmacokinetics of cyclosporine was defined robustly in this patient population using population pharmacokinetic approaches. Results indicate that the Pliva Cyclosporine Soft Gelatin Capsules and Neoral® are bioequivalent when administered to renal transplant patients. Pliva Cyclosporine Soft Gelatin Capsules can then be substituted for Neoral® in stabilized patients without anticipating dose adjustments.

The effect and clinical consequences of hypoxia on cytochrome P450, membrane carrier proteins activity and expression

Introduction: Chronic pulmonary disease and heart failure reduce drug clearance and consequently enhance adverse drug reactions. The mechanisms of action underlying the regulation of cytochrome P450 (CYP) isoforms and membrane carrier proteins by hypoxia, and the clinical consequences of the regulation of CYP by hypoxia, alone or combined with other conditions have been elucidated in the last decades. Overall, a reduced drug clearance appears to be associated with hypoxemia. Areas covered: In this review, the mechanisms of action underlying hypoxia-induced regulation of CYP enzymes are discussed. The authors also revise the effects of hypoxia on serum mediators, signal transduction pathways, orphan nuclear receptors, transcription factors and post-transcriptional mechanisms regulating CYP and membrane carrier proteins expression. Additionally, the paper also discusses the clinical repercussions of hypoxia-induced changes in CYP and membrane carrier proteins activity. Expert opinion: Acute systemic hypoxia down-regulates selected CYP isoforms and up-regulates CYP3A4 and P-glycoprotein, changing the metabolic clearance of drugs and endogenous compounds biotransformed by these isoforms as well as the kinetics. In patients with acute hypoxia, the dosage of drugs, biotransformed by CYP isoforms, may need to be adjusted. Tissue hypoxia enhances the expression of efflux membrane carrier proteins, increasing the probability of drug resistance.

Modulation of CYP1A2 and CYP3A6 catalytic activities by serum from rabbits with a turpentine-induced inflammatory reaction and interleukin 6.

Inflammatory reactions reduce the activity of cytochrome P450 isoforms. The aim of the study was to determine the mechanisms underlying the decrease in CYP1A2 and CYP3A6 catalytic activities produced by serum from rabbits with a turpentine-induced inflammatory reaction (STIIR) and interleukin 6 (IL-6). STIIR and IL-6 were incubated with cultured primary hepatocytes from control rabbits (HCONT), and from rabbits with a turpentine-induced inflammatory reaction (HTIIR) in the absence or presence of pyrrolidine dithiocarbamate (PDTC), an antioxidant and inhibitor of nuclear factor κB transcription; 2′-amino-3′-methoxyflavone (PD98059), an inhibitor of extracellular signal-related kinase (Erk1/2); 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imidazole (SB203580), an inhibitor of p38MAPK; Nω-nitro-l-arginine methyl ester, an inhibitor of nitric-oxide synthase 2 (NOS2); the combination of PDTC, PD98059, and SB203580; and genistein, an inhibitor of Janus-associated protein tyrosine kinase (JAK). After 4 and 24 h of incubation of HCONT with STIIR and IL-6, CYP1A2 activity was reduced without changes in expression; the reduction in activity was partially prevented by the inhibition of JAK, Erk1/2, and NOS2. In HCONT, STIIR and IL-6 did not affect CYP3A6 activity; however, PDTC reduced CYP3A6 activity by 40 and 80% after 4 and 24 h of incubation. In HTIIR, STIIR and IL-6 reduced both CYP1A2 and CYP3A6 activities; this decrease is partially prevented by inhibitors of protein tyrosine kinases, Erk1/2, and NOS2. In HTIIR, SB203580 increased CYP3A6 activity in a dose-dependent manner without changes in protein expression. These results show that the signal transduction pathways mediating the decrease in CYP1A2 and 3A6 activity, produced by STIIR and IL-6, involve JAK, Erk1/2, and NOS2.

Effects of renal impairment on the pharmacokinetics of orally administered deferiprone

Abstract Aims In light of the growing recognition of renal disease in thalassemia, it is important to understand the impact of renal impairment on the pharmacokinetics of iron chelators. This study evaluated the pharmacokinetics and safety of the iron chelator deferiprone (DFP) in subjects with renal impairment in comparison with healthy volunteers (HVs). Methods Thirty‐two subjects were categorized into four groups based on degree of renal impairment: none, mild, moderate or severe, as determined by estimated glomerular filtration rate (eGFR). All subjects received a single oral dose of 33 mg kg−1 DFP, provided serum and urine samples for pharmacokinetic assessment over 24 h and were monitored for safety. Results Renal clearance of DFP decreased as renal impairment increased. However, based on C max, AUC(0,t) and AUC(0,∞), there were no significant group differences in systemic exposure, because less than 4% of the drug was excreted unchanged in the urine. DFP is extensively metabolized to a renally excreted, pharmacologically inactive metabolite, deferiprone 3‐O‐glucuronide (DFP‐G), which exhibited higher C max, AUC(0,t), AUC(0,∞) and longer t max and t 1/2 in the renally impaired groups compared with HVs. The C max and AUCs of DFP‐G increased as eGFR decreased. Overall, 75%–95% of the dose was retrieved in urine, either as DFP or DFP‐G, regardless of severity of renal impairment. With respect to safety, DFP was well tolerated. Conclusions These data suggest that no adjustment of the DFP dosage regimen in patients with renal impairment is necessary, as there were no significant changes in the systemic exposure to the drug.

Randomized, Blinded, Placebo‐ and Positive‐Controlled Crossover Study to Determine the Effect of Deferiprone on the QTc Interval in Healthy Subjects

This study evaluated whether deferiprone, an oral iron chelator, acts to prolong the QT interval. Fifty healthy volunteers received single doses of each of the following: therapeutic dose of deferiprone (33 mg/kg), supratherapeutic dose (50 mg/kg), placebo, or moxifloxacin, a positive control known to significantly prolong QT interval. Following each dose, subjects underwent cardiac monitoring, pharmacokinetics assessments, and safety assessments. Based on the QT interval obtained using the Fridericia correction for heart rate (QTcF), the upper bound of the 1‐sided 95% confidence interval of the mean difference between deferiprone and placebo was <10 milliseconds (the threshold of concern defined by authorities) at all time points for both doses: maximum difference of 3.01 milliseconds for the therapeutic dose and 5.23 milliseconds for the supratherapeutic dose. The difference in dQTcF between moxifloxacin and placebo demonstrated that the study was adequately sensitive to detect a significant prolongation of QTcF. The concentration‐response correlation analyses revealed some weak but statistically significant trends of increase in dQTcF and ddQTcF with increasing exposure to deferiprone, but these trends should have no clinical consequence even at the recommended maximum dosage. In conclusion, there was no clinically meaningful effect on QTc interval following single therapeutic or supratherapeutic doses of deferiprone.