A new progeroid syndrome reveals that genotoxic stress suppresses the somatotroph axis.

XPF-ERCC1 endonuclease is required for repair of helix-distorting DNA lesions and cytotoxic DNA interstrand crosslinks. Mild mutations in XPF cause the cancer-prone syndrome xeroderma pigmentosum. A patient presented with a severe XPF mutation leading to profound crosslink sensitivity and dramatic progeroid symptoms. It is not known how unrepaired DNA damage accelerates ageing or its relevance to natural ageing. Here we show a highly significant correlation between the liver transcriptome of old mice and a mouse model of this progeroid syndrome. Expression data from XPF-ERCC1-deficient mice indicate increased cell death and anti-oxidant defences, a shift towards anabolism and reduced growth hormone/insulin-like growth factor 1 (IGF1) signalling, a known regulator of lifespan. Similar changes are seen in wild-type mice in response to chronic genotoxic stress, caloric restriction, or with ageing. We conclude that unrepaired cytotoxic DNA damage induces a highly conserved metabolic response mediated by the IGF1/insulin pathway, which re-allocates resources from growth to somatic preservation and life extension. This highlights a causal contribution of DNA damage to ageing and demonstrates that ageing and end-of-life fitness are determined both by stochastic damage, which is the cause of functional decline, and genetics, which determines the rates of damage accumulation and decline.

Paclitaxel in self-micro emulsifying formulations: oral bioavailability study in mice.

The anticancer drug paclitaxel is formulated for i.v. administration in a mixture of Cremophor EL and ethanol.Its oral bioavailability is very low due to the action of P-glycoproteinin the gut wall and CYP450 in gut wall and liver.However, proof-of-concept studies using the i.v. formulation diluted in drinking water have demonstrated the feasibility of the oral route as an alternative when given in combination with inhibitors of P-glycoprotein and CYP450. Because of the unacceptable pharmaceutical properties of the drinking solution, a better formulation for oral application is needed.We have evaluated the suitability of various self-micro emulsifying oily formulations (SMEOF's) of paclitaxel for oral application using wild-type and P-glycoprotein knockout mice and cyclosporin A (CsA) as P-glycoprotein and CYP450 inhibitor. The oral bioavailability of paclitaxel in all SMEOF's without concomitant CsA was low in wild-type mice, showing that this vehicle does not enhance intestinal uptake by itself.Paclitaxel (10 mg/kg) in SMEOF#3 given with CsA resulted in plasma levels that were comparable to the Cremophor ELethanol containing drinking solution plus CsA. Whereas the AUC increased linearly with the oral paclitaxel dose in P-glycoprotein knockout mice, it increased less than proportional in wild-type mice given with CsA. In both strains more unchanged paclitaxel was recovered in the feces at higher doses. This observation most likely reflects more profound precipitation of paclitaxel within the gastro-intestinal tract at higher doses. The resulting absolute reduction in absorption of paclitaxel from the gut was possibly concealed by partial saturation of first-pass metabolism when P-glycoprotein was absent. In conclusion, SMEOF's maybe a useful vehicle for oral delivery of paclitaxel in combination with CsA, although the physical stability within the gastro-intestinal tract remains a critical issue, especially when applied at higher dose levels.

Population pharmacokinetics of intravenously and orally administered docetaxel with or without co-administration of ritonavir in patients with advanced cancer.

AIM: Docetaxel has a low oral bioavailability due to affinity for P-glycoprotein and cytochrome P450 (CYP) 3A4 enzymes. Inhibition of the CYP3A4 enzymes by ritonavir resulted in increased oral bioavailability. The aim of this study was to develop a population pharmacokinetic (PK) model and to evaluate and quantify the influence of ritonavir on the PK of docetaxel. METHODS: Data from two clinical trials were included in the data analysis, in which docetaxel (75 mg m(-2) or 100 mg) had been administered intravenously or orally (10 mg or 100 mg) with or without co-administration of oral ritonavir (100 mg). Population modelling was performed using non-linear mixed effects modelling. A three-compartment model was used to describe the i.v. data. PK data after oral administration, with or without co-administration of ritonavir, were incorporated into the model. RESULTS: Gut bioavailability of docetaxel increased approximately two-fold from 19 to 39% (CV 13%) with ritonavir co-administration. The hepatic extraction ratio and the elimination rate of docetaxel were best described by estimating the intrinsic clearance. Ritonavir was found to inhibit in a concentration dependent manner the intrinsic clearance of docetaxel, which was described by an inhibition constant of 0.028 microg ml(-1) (CV 36%). A maximum inhibition of docetaxel clearance of more then 90% was reached. CONCLUSIONS: A PK model describing both the PK of orally and intravenously administered docetaxel in combination with ritonavir, was successfully developed. Co-administration of ritonavir lead to increased oral absorption and reduced elimination rate of docetaxel.

Organic anion-transporting polypeptide 1B1 mediates transport of Gimatecan and BNP1350 and can be inhibited by several classic ATP-binding cassette (ABC) B1 and/or ABCG2 inhibitors.

Organic anion-transporting polypeptides (OATPs) are important uptake transporters that can have a profound impact on the systemic pharmacokinetics, tissue distribution, and elimination of several drugs. Previous in vivo studies of the pharmacokinetics of the lipophilic camptothecin (CPT) analog gimatecan suggested that the ATP-binding cassette (ABC) B1 (P-glycoprotein) and/or ABCG2 (breast cancer resistance protein) inhibitors elacridar and pantoprazole could inhibit transporters other than ABCB1 and ABCG2. In this study, we tested the possible role of OATP1B1 in this interaction by screening a number of CPT analogs for their transport affinity by human OATP1B1 in vitro. In addition, the impact of several widely used ABCB1 and/or ABCG2 modulators on this OATP1B1-mediated transport was assessed. We identified two novel CPT anticancer drugs, gimatecan and BNP1350, as OATP1B1 substrates, whereas irinotecan, topotecan, and lurtotecan were not transported by OATP1B1. It is interesting to note that transport of 17beta-estradiol 17beta-d-glucuronide (control), gimatecan, and BNP1350 by OATP1B1 could be completely inhibited by the classic ABCB1 and/or ABCG2 inhibitors elacridar, valspodar, pantoprazole, and, to a lesser extent, zosuquidar and verapamil. Therefore, the effect of these ABCB1 and ABCG2 modulators on the plasma pharmacokinetics of gimatecan and BNP1350 (and possibly also other OATP1B1 substrates) may be partly because of inhibition of OATP1B1 besides inhibition of ABCB1 and/or ABCG2. The findings of this study suggest that OATP1B1 polymorphisms or coadministration with one of the ABCB1/ABCG2 inhibitors could affect drug uptake, tissue distribution, and elimination of some CPT anticancer drugs, thereby modifying their efficacy and/or safety profile.

In vitro transport of gimatecan (7-t-butoxyiminomethylcamptothecin) by breast cancer resistance protein, P-glycoprotein, and multidrug resistance protein 2.

Lipophilic camptothecin derivatives are considered to have negligible affinity for breast cancer resistance protein (BCRP; ABCG2). Gimatecan, a new orally available 7-t-butoxyiminomethyl-substituted lipophilic camptothecin derivative, has been previously reported to be not a substrate for BCRP. Using a panel of in vitro models, we tested whether gimatecan is a substrate for BCRP as well as for P-glycoprotein (MDR1) or multidrug resistance protein 2 (MRP2; ABCC2), ATP-binding cassette drug efflux transporters involved in anticancer drug resistance, and able to affect the pharmacokinetics of substrate drugs. Cell survival, drug transport, accumulation, and efflux were studied in IGROV1 and (human BCRP overexpressing) T8 cells, Madin-Darby canine kidney II (MDCKII-WT, MDCKII-Bcrp1, MDCKII-MDR1, and MDCKII-MRP2), and LLCPK (LLCPK-WT and LLCPK-MDR1) cells. Competition with methotrexate uptake was studied in Sf9-BCRP membrane vesicles. In vitro, expression of BCRP resulted in 8- to 10-fold resistance to gimatecan. In Transwell experiments, gimatecan was transported by Bcrp1 and transport was inhibited by the BCRP/P-glycoprotein inhibitors elacridar and pantoprazole. Efflux of gimatecan from MDCKII-Bcrp1 cells was faster than in WT cells. In Sf9-BCRP membrane vesicles, gimatecan significantly inhibited BCRP-mediated transport of methotrexate. In contrast, gimatecan was not transported by MDR1 or MRP2. Gimatecan is transported by BCRP/Bcrp1 in vitro, although to a lesser extent than the camptothecin analogue topotecan. Implications of BCRP expression in the gut for the oral development of gimatecan and the interaction between gimatecan and other BCRP substrate drugs and/or inhibitors warrant further clinical investigation.

The effect of hydroxyurea on P-glycoprotein/BCRP-mediated transport and CYP3A metabolism of imatinib mesylate.

PURPOSE: It has been reported that the combination therapy of imatinib mesylate, a tyrosine kinase inhibitor, plus hydroxyurea, a ribonucleotide reductase inhibitor, is associated with remarkable antitumor activity in patients with recurrent glioblastoma multiforme. However, the mechanism of the added activity of hydroxyurea to imatinib is not known. The purpose of this study was to investigate in vitro, whether hydroxyurea could enhance the central nervous system penetration of imatinib, by inhibition of the ATP-dependent transporter proteins P-glycoprotein (ABCB1; MDR1; Pgp) and Breast Cancer Resistance Protein (ABCG2; BCRP), or by inhibition of cytochrome P450 3A (CYP3A) metabolism of imatinib. METHODS: The effect of hydroxyurea on the Pgp and BCRP mediated transport of imatinib was investigated by the sulforhodamine-B (SRB) drug cytotoxicity assay and transepithelial transport assay. In vitro biotransformation studies with supersomes expressing human CYP3A4 were performed to investigate whether hydroxyurea inhibited CYP3A4. RESULTS: In both in vitro cytotoxicity and transport assays, hydroxyurea did not affect Pgp and BCRP mediated transport of imatinib. In a biotransformation assay, hydroxyurea had no influence on the metabolic degradation of imatinib either. CONCLUSION: The results indicate that hydroxyurea does not interact with imatinib by inhibition of Pgp and BCRP mediated transport or by CYP3A4 mediated metabolism of imatinib.

Targeting cellular adhesion molecules, chemokines and chemokine receptors in rheumatoid arthritis.

The development of specific targeted therapies, such as anti-TNF-alpha treatment, for chronic inflammatory disorders such as rheumatoid arthritis, has significantly improved treatment, although not all patients respond. Targeting cellular adhesion molecules and chemokines/chemokine receptors as regulators of the extravasation and migration of leukocytes may provide a novel approach for the treatment of these diseases. Moreover, the possibility of developing small-molecule antagonists offers an excellent method for the oral delivery of compounds with a short half-life.

The biological and clinical role of drug transporters at the intestinal barrier.

Nowadays, over one fourth of all anticancer drugs are developed as oral formulations and this percentage is expected to increase substantially. Oral administration of drugs is patient convenient and practical and is preferred for many reasons. To enable oral drug therapy adequate oral bioavailability must be achieved. One of the factors that have proven to be important in explaining the often variable and low oral bioavailability of many orally applied anticancer drugs is the presence of ATP-binding Cassette drug transporters (ABC transporters) and solute carrier (SLC) transporters. During the past two decades, significant progress has been made in understanding the pharmacological and physiological role of ABC drug efflux and SLC uptake transporters in the disposition of a broad range of drugs, toxins, endogenous compounds and their metabolites. We focus on the expression of ABC and SLC drug transporters at the intestinal barrier and the impact of these transporters on the absorption and disposition of a wide range of orally administered drugs. Furthermore, preclinical and clinical examples of modulation of the activity of intestinal transporters to increase the systemic exposure of orally administered drugs will be reviewed. Screening of test drugs, nutrients and other molecules for ABC and SLC transporter substrates or inhibitors is a useful way to predict their intestinal absorption. Recognition of the importance of intestinal transporters could guide the design and development of oral drugs.

The effect of P-gp (Mdr1a/1b), BCRP (Bcrp1) and P-gp/BCRP inhibitors on the in vivo absorption, distribution, metabolism and excretion of imatinib.

Imatinib is transported by P-glycoprotein (P-gp) and Breast Cancer Resistance Protein (BCRP), however, the exact impact of these transporters on absorption, distribution, metabolism and excretion (ADME) of imatinib is not fully understood due to incomplete data. We have performed a comprehensive ADME study of imatinib given as single agent or in combination with the well known BCRP/P-gp inhibitors, elacridar and pantoprazole, in wild-type and P-gp and/or BCRP knockout mice. The absence of P-gp and BCRP together resulted in a significantly higher area under the plasma concentration-time curve (AUC) after i.v. administration, whereas the AUC after oral dosing was unaltered. Both elacridar and pantoprazole significantly increased the AUC of orally administered imatinib in wild-type but also in P-gp/BCRP knockout mice. This lower clearance was not due to a (further) reduction in biliary excretion. Fecal excretion was significantly reduced in P-gp and P-gp/BCRP knockout but not in BCRP knockout mice, whereas the brain penetration was significantly higher in P-gp/BCRP knockout mice compared to single P-gp or BCRP knockout or wild-type mice. In conclusion, P-gp and BCRP have only a modest effect on the ADME of imatinib in comparison to metabolic elimination. P-gp is the most prevalent factor for systemic clearance and limiting the brain penetration. The considerable drug-drug interaction observed with elacridar or pantoprazole is only partly mediated by inhibition of P-gp and BCRP and far more by the inhibition of other elimination pathways.

Coadministration of ritonavir strongly enhances the apparent oral bioavailability of docetaxel in patients with solid tumors.

PURPOSE: To enhance the systemic exposure to oral docetaxel by coadministration of ritonavir, an efficacious inhibitor of CYP 3A4 with minor P-glycoprotein inhibiting effects, in patients with cancer. EXPERIMENTAL DESIGN: A proof-of-concept study was carried out in 12 patients with solid tumors. The first cohort of patients (n = 4) received 10 mg and the subsequent cohort (n = 8) 100 mg of oral docetaxel, coadministered with 100 mg oral ritonavir randomized simultaneously or ritonavir given 60 minutes before docetaxel on days 1 and 8. On day 15 or 22, patients received 100 mg i.v. docetaxel. RESULTS: The area under the plasma concentration-time curve in patients who received 10 mg oral docetaxel in combination with ritonavir was low, and the dose could safely be increased to 100 mg. The area under the plasma concentration-time curve in patients who received 100 mg oral docetaxel combined with ritonavir simultaneously or ritonavir given 60 minutes before docetaxel was 2.4 +/- 1.5 and 2.8 +/- 1.4 mg/h/L, respectively, compared with 1.9 +/- 0.4 mg/h/L after i.v. docetaxel. The apparent oral bioavailability of docetaxel combined with ritonavir simultaneously or ritonavir given 60 minutes before docetaxel was 131% +/- 90% and 161% +/- 91%, respectively. The oral combination of docetaxel and ritonavir was well tolerated. CONCLUSION: Coadministration of ritonavir significantly enhanced the apparent oral bioavailability of docetaxel. These data are promising and form the basis for further development of a clinically applicable oral formulation of docetaxel combined with ritonavir.

Determination of imatinib mesylate and its main metabolite (CGP74588) in human plasma and murine specimens by ion-pairing reversed-phase high-performance liquid chromatography.

A sensitive reversed-phase high-performance liquid chromatographic (HPLC) method has been developed and validated for the determination of imatinib, a tyrosine kinase inhibitor, and its main metabolite N-desmethyl-imatinib (CGP74588) in human plasma and relevant murine biological matrices. A simple HPLC assay for the individual quantification of imatinib and CGP74588 in murine specimens has not been reported to date. Sample pre-treatment involved liquid-liquid extraction with tert-butyl-methyl ether. Imatinib, CGP74588 (metabolite) and the internal standard 4-hydroxybenzophenone were separated using a narrow bore (2.1 x 150 mm) stainless steel Symmetry C(18) column and detected by UV at 265 nm. The mobile phase consisted of 28% (v/v) acetonitrile in 50 mM ammonium acetate buffer pH 6.8 containing 0.005 M 1-octane sulfonic acid and was delivered at 0.2 mL/min. The calibration curve was prepared in blank human plasma and was linear over the dynamic range 10 ng/mL to 10 microg/mL). The accuracy was close to 100% and the within-day and between-day precisions were within the generally accepted 15% range. The validation results showed that the assay was selective and reproducible. This method was applied to study the pharmacokinetics of imatinib and its main metabolite in human and mice.