A population pharmacokinetic meta-analysis of custirsen, an antisense oligonucleotide, in oncology patients and healthy subjects.

AIMS: Custirsen (OGX-011/TV-1011), a second-generation antisense oligonucleotide that reduces clusterin production, is under investigation with chemotherapy in prostate and lung cancer. This meta-analysis evaluated the population pharmacokinetics (PK) of custirsen in cancer patients and healthy subjects. METHODS: The population PK analysis used custirsen plasma concentrations from five Phase 1 studies, one Phase 1/2 study, and one Phase 3 study in two stages. Cancer patients received multiple doses of custirsen (40-640 mg intravenously over 120 min) with chemotherapy; healthy subjects received single or multiple doses (320-640 mg). An interim population PK model was developed using a nonlinear mixed-effect approach incorporating data from four Phase 1 or 1/2 studies, followed by model refinement and inclusion of two Phase 1 and one Phase 3 studies. RESULTS: The final model was developed with 5588 concentrations from 631 subjects with doses of 160-640 mg. Custirsen PK was adequately described by a three-compartment model with first-order elimination. For a representative 66-year-old individual with body weight 82 kg and serum creatinine level 0.933 mg dl-1 , the estimated typical (95% CI) parameter values were clearance (CL) = 2.36 (2.30-2.42) l h-1 , central volume of distribution (V1 ) = 6.08 (5.93-6.23) l, peripheral volume of distribution (V2 ) = 1.13 (1.01-1.25) l, volume of the second peripheral compartment (V3 ) = 15.8 (14.6-17.0) l, inter-compartmental clearance Q2 = 0.0755 (0.0689-0.0821) l h-1 , and Q3 = 0.0573 (0.0532-0.0614) l h-1 . Age, weight and serum creatinine were predictors of CL; age was a predictor of Q3 . CONCLUSION: A population PK model for custirsen was successfully developed in cancer patients and healthy subjects, including covariates contributing to variability in custirsen PK.

Pharmacokinetics and excretion of (14)C-omacetaxine in patients with advanced solid tumors.

Background Omacetaxine mepesuccinate is indicated in adults with chronic myeloid leukemia resistant and/or intolerant to ≥ 2 tyrosine kinase inhibitor treatments. This phase I study assessed the disposition, elimination, and safety of (14)C-omacetaxine in patients with solid tumors. Methods The study comprised a 7-days pharmacokinetic assessment followed by a treatment period of ≤ six 28-days cycles. A single subcutaneous dose of 1.25 mg/m(2) (14)C-omacetaxine was administered to six patients. Blood, urine, and feces were collected through 168 h or until radioactivity excreted within 24 h was <1 % of the dose. Total radioactivity (TRA) was measured in all matrices and concentrations of omacetaxine, 4'-desmethylhomoharringtonine (4'-DMHHT), and cephalotaxine were measured in plasma and urine. For each treatment cycle, patients received 1.25 mg/m(2) omacetaxine twice daily for 7 days. Results Mean TRA recovered was approximately 81 % of the dose, with approximately half of the radioactivity recovered in feces and half in urine. Approximately 20 % of the dose was excreted unchanged in urine; cephalotaxine (0.4 % of dose) and 4' DMHHT (9 %) were also present. Plasma concentrations of TRA were higher than the sum of omacetaxine and known metabolites, suggesting the presence of other (14)C-omacetaxine-derived compounds. Fatigue and anemia were common, consistent with the known toxicity profile of omacetaxine. Conclusion Renal and hepatic processes contribute to the elimination of (14)C-omacetaxine-derived radioactivity in cancer patients. In addition to omacetaxine and its known metabolites, other (14)C-omacetaxine-derived materials appear to be present in plasma and urine. Omacetaxine was adequately tolerated, with no new safety signals.

A thorough QT study to assess the effects of tbo-filgrastim on cardiac repolarization in healthy subjects.

Tbo-filgrastim is a recombinant human granulocyte colony-stimulating factor approved by the US Food and Drug Administration to reduce the duration of severe neutropenia in patients with nonmyeloid malignancies receiving myelosuppressive anticancer drugs associated with a clinically significant incidence of febrile neutropenia. We assessed the effect of tbo-filgrastim on cardiac conduction and repolarization in healthy subjects. A three-arm, parallel-group, active- and placebo-controlled, double-blind study randomized healthy adults to a single 5 µg/kg intravenous tbo-filgrastim infusion, a single intravenous placebo infusion, or a single 400 mg moxifloxacin oral dose. The primary end point was placebo-corrected time-matched change from baseline in QT interval corrected using a QT individual correction (QTcI) method. Secondary end points included heart rate, PR interval, QRS duration, change in electrocardiogram patterns, correlation between QTcI change from baseline (milliseconds) and tbo-filgrastim serum concentrations, and safety variables. A total of 145 subjects were enrolled (50 tbo-filgrastim, 50 placebo, 45 moxifloxacin). Peak placebo-corrected change from baseline for QTcI with tbo-filgrastim was 3.5 milliseconds, with a two-sided 95% upper confidence interval of 7.2 milliseconds, demonstrating no signal for any tbo-filgrastim effect on QTc. Concentration-effect modeling showed no evidence of an effect of tbo-filgrastim on cardiac repolarization. Tbo-filgrastim produced no clinically significant changes in other electrocardiogram parameters. Tbo-filgrastim was well tolerated.

Impact of dosing regimen of custirsen, an antisense oligonucleotide, on safety, tolerability and cardiac repolarization in healthy subjects.

AIMS: Custirsen (OGX-011/TV-1011), a second-generation antisense oligonucleotide (ASO) that reduces clusterin production, is under investigation with chemotherapy in patients with solid tumours. Custirsen is associated with constitutional symptoms (CS) that may interfere with clinical pharmacology investigations, such as QT interval studies. Experience with other ASOs suggests NSAID premedication may ameliorate CS, but we observed suboptimal outcomes in healthy subjects given custirsen and NSAIDs. We sought to establish a custirsen regimen for future clinical pharmacology studies in healthy subjects. METHODS: Subjects received custirsen (640 mg intravenously over 120 min) with dexamethasone premedication or increasing doses (320, 480, 640 mg over 6 days) of custirsen with dexamethasone premedication, then one full custirsen dose without premedication on day 8. Incidence/severity of adverse events (AEs) and extensive electrocardiogram readings were evaluated. Pharmacokinetic parameters were estimated. RESULTS: AEs included CS, elevated transaminases and prolonged activated partial thromboplastin time (aPTT) that were predominantly grade 1/2. Administration of increasing custirsen doses and dexamethasone premedication reduced the incidence of CS associated with full dose custirsen. Transaminase elevation showed a dose-dependent effect (0% at days 2, 4, 27% at day 6) with the highest custirsen doses. Increasing doses of custirsen may have mitigated the severity but not incidence of aPTT prolongation. Neither regimen was associated with cardiac repolarization changes in QT values or concentration-effect analyses. The custirsen pharmacokinetic profile was consistent with previous experience. CONCLUSION: Escalation of custirsen dose combined with dexamethasone premedication reduced CS associated with full dose custirsen and should be considered in future clinical pharmacology studies of custirsen.

Folate transport gene inactivation in mice increases sensitivity to colon carcinogenesis.

Low dietary folate intake is associated with an increased risk for colon cancer; however, relevant genetic animal models are lacking. We therefore investigated the effect of targeted ablation of two folate transport genes, folate binding protein 1 (Folbp1) and reduced folate carrier 1 (RFC1), on folate homeostasis to elucidate the molecular mechanisms of folate action on colonocyte cell proliferation, gene expression, and colon carcinogenesis. Targeted deletion of Folbp1 (Folbp1(+/-) and Folbp1(-/-)) significantly reduced (P < 0.05) colonic Folbp1 mRNA, colonic mucosa, and plasma folate concentration. In contrast, subtle changes in folate homeostasis resulted from targeted deletion of RFC1 (RFC1(+/-)). These animals had reduced (P < 0.05) colonic RFC1 mRNA and exhibited a 2-fold reduction in the plasma S-adenosylmethionine/S-adenosylhomocysteine. Folbp1(+/-) and Folbp1(-/-) mice had larger crypts expressed as greater (P < 0.05) numbers of cells per crypt column relative to Folbp1(+/+) mice. Colonic cell proliferation was increased in RFC1(+/-) mice relative to RFC1(+/+) mice. Microarray analysis of colonic mucosa showed distinct changes in gene expression specific to Folbp1 or RFC1 ablation. The effect of folate transporter gene ablation on colon carcinogenesis was evaluated 8 and 38 weeks post-azoxymethane injection in wild-type and heterozygous mice. Relative to RFC1(+/+) mice, RFC1(+/-) mice developed increased (P < 0.05) numbers of aberrant crypt foci at 8 weeks. At 38 weeks, RFC1(+/-) mice developed local inflammatory lesions with or without epithelial dysplasia as well as adenocarcinomas, which were larger relative to RFC1(+/+) mice. In contrast, Folbp1(+/-) mice developed 4-fold (P < 0.05) more lesions relative to Folbp1(+/+) mice. In conclusion, Folbp1 and RFC1 genetically modified mice exhibit distinct changes in colonocyte phenotype and therefore have utility as models to examine the role of folate homeostasis in colon cancer development.

Renal tubular reabsorption of folate mediated by folate binding protein 1.

Renal tubular reabsorption of filtered folate is essential for the conservation and normal homeostasis of this important vitamin. Different molecular mechanisms have been implicated in epithelial folate transport, including folate receptors. Defective expression or antibody inactivation of these is associated with embryonic defects also correlated with low folate intake; however, their contribution to renal tubular folate reabsorption has not been established. With the use of targeted inactivation of the folate binding protein 1 (folbp1) and folate binding protein 2 (folbp2) genes in mice, the role of folate receptors in renal epithelial folate reabsorption was evaluated during low and normal folate intake. Inactivation of folbp1 was associated with (1) loss of (3)H-folic acid binding to crude kidney membranes, (2) increase in renal folate clearance, and (3) increase in urinary excretion and decrease in renal uptake of injected (3)H-methyltetrahydrofolate. No changes in renal folate handling were observed as a result of folbp2 inactivation. Thus, folbp1 is essential for normal renal tubular folate reabsorption, preventing excessive urinary folate loss. Folbp1 is heavily expressed in choroid plexus, yolk sac, and placenta, supporting a role of folbp1 in folate transport in other tissues. The greatest significance of folbp1 for renal folate uptake was observed at conditions of low folate intake, providing a possible explanation for the ability of folate supplementation to prevent developmental defects associated with folbp1 inactivation.

Developmental consequences of in utero sodium arsenate exposure in mice with folate transport deficiencies.

Previous studies have demonstrated that mice lacking a functional folate binding protein 2 gene (Folbp2-/-) were significantly more sensitive to in utero arsenic exposure than were the wild-type mice similarly exposed. When these mice were fed a folate-deficient diet, the embryotoxic effect of arsenate was further exacerbated. Contrary to expectations, studies on 24-h urinary speciation of sodium arsenate did not demonstrate any significant difference in arsenic biotransformation between Folbp2-/- and Folbp2+/+ mice. To better understand the influence of folate pathway genes on arsenic embryotoxicity, the present investigation utilized transgenic mice with disrupted folate binding protein 1 (Folbp1) and reduced folate carrier (RFC) genes. Because complete inactivation of Folbp1 and RFC genes results in embryonic lethality, we used heterozygous animals. Overall, no RFC genotype-related differences in embryonic susceptibility to arsenic exposure were observed. Embryonic lethality and neural tube defect (NTD) frequency in Folbp1 mice was dose-dependent and differed from the RFC mice; however, no genotype-related differences were observed. The RFC heterozygotes tended to have higher plasma levels of S-adenosylhomocysteine (SAH) than did the wild-type controls, although this effect was not robust. It is concluded that genetic modifications at the Folbp1 and RFC loci confers no particular sensitivity to arsenic toxicity compared to wild-type controls, thus disproving the working hypothesis that decreased methylating capacity of the genetically modified mice would put them at increased risk for arsenic-induced reproductive toxicity.

Effects of dietary folate intake and folate binding protein-1 (Folbp1) on urinary speciation of sodium arsenate in mice.

In most mammalian species, arsenic biotransformation occurs primarily by biomethylation with dimethylarsinic acid being the predominant metabolite excreted in the urine. Folbp1 (folate binding protein-1) mediated intracellular folate uptake is one route by which cells harvest folate cofactors. In light of the likely relationship between folate biochemistry and arsenic biotransformation, our experiments were designed to test: (1) whether Folbp1 is an important determinant in arsenic biotransformation, by performing urinary arsenic speciation in Folbp1 nullizygous (Folbp1(-/-)) and wildtype control mice, and (2) whether dietary folate deficiency alters arsenic biotransformation in these mice. Compared to normal folate intake, folate deficiency caused lower amounts of arsenic to be excreted in the urine of both the wildtype controls and Folbp1(-/-) mice. Folbp1(-/-) mice excreted more dimethylarsinic acid than wildtype control mice during folate deficiency, but not during normal folate intake. The present data suggest that inadequate folate intake may result in decreased biotransformation and excretion of arsenic, which is likely to increase arsenic exposure and related toxicities.

Pathobiology and genetics of neural tube defects.

PURPOSE: Neural tube defects (NTDs), including spina bifida and anencephaly, are common congenital malformations that occur when the neural tube fails to achieve proper closure during early embryogenesis. Based on epidemiological and clinical data obtained over the last few decades, it is apparent that these multifactorial defects have a significant genetic component to their etiology that interacts with specific environmental risk factors. The purpose of this review article is to synthesize the existing literature on the genetic factors contributing to NTD risk. RESULTS: To date, there is evidence that closure of the mammalian neural tube initiates and fuses intermittently at four discrete locations. Disruption of this process at any of these four sites may lead to an NTD, possibly arising through closure site-specific genetic mechanisms. Candidate genes involved in neural tube closure include genes of the folate metabolic pathway, as well as those involved in folate transport. CONCLUSIONS: Although extensive efforts have focused on elucidating the genetic risk factors contributing to the etiology of NTDs, the population burden for these malformations remains unknown. One group at high risk for having children with NTDs is epileptic women receiving antiepileptic medications during pregnancy. Efforts to better understand the genetic factors that may contribute to their heightened risk, as well as the pathogenesis of neural tube closure defects, are reviewed herein.

Gas chromatographic determination of novel valproyl taurinamide derivatives in mouse and dog plasma.

Valproyl taurinamides are a novel group of compounds that possess anticonvulsant activity. In this study a gas chromatographic micromethod was developed for the quantification of selected valproyl taurinamides and some of their metabolites in biological samples. Valproyl taurinamide (VTD), N-methyl valproyl taurinamide (M-VTD), N,N-dimethyl valproyl taurinamide (DM-VTD) and N-isopropyl valproyl taurinamide (I-VTD) were analyzed in mouse and dog plasma and in dog urine using gas chromatography. Flame ionization detection and mass spectrometric detection were compared. The plasma samples were prepared by solid-phase extraction using C(18) cartridges. The urine samples were prepared by liquid-liquid extraction. The sample volume used was 100 microl of dog plasma, 50 microl of mouse plasma and 20 microl of dog or mouse urine. The quantification range of the method was 1.5-50 mg/l in dog plasma (VTD only), 2.5-250 mg/l in mouse plasma (0.7-90 pmol injected) and 0.04-2 mg/ml in dog urine (VTD only). The inter-day precision in plasma and urine samples was around 10% for all quantified concentrations except LOQ (15-20%). The accuracy for all four compounds was between 90 and 110% within the entire concentration range. The developed method was suitable for quantification of a series of CNS-active valproyl taurineamide derivatives in biological samples at relevant in vivo concentrations.

Valproate-induced neural tube defects in folate-binding protein-2 (Folbp2) knockout mice.

BACKGROUND: Folate is an important B vitamin that is transported into cells by way of folate-binding proteins and transporters. Folate-binding protein-2 nullizygous (Folbp2(-/-)) mice develop normally; however, we have found them to be more susceptible to the teratogenic effects of arsenate exposure than wild-type control mice. METHODS: In the current study, we wanted to extend our findings and test the hypothesis that Folbp2(-/-) mice are more susceptible to the teratogenic effects of valproic acid (VPA), a commonly used antiepileptic drug that is known to induce neural tube defects (NTDs) in both humans and laboratory animals. RESULTS: Folbp2(-/-) mice had higher VPA-induced frequencies of embryonic lethality and exencephaly than did the wild-type control mice during folate supplementation and a control diet, respectively. All other differences in response between the two genotypes were short of reaching statistical significance. Folate supplementation of wild-type, but not Folbp2(-/-) dams reduced embryonic lethality of VPA-treated wild-type embryos compared to the folate-deficient diet. CONCLUSIONS: Unlike our previous findings with arsenate, enhanced susceptibility of Folbp2(-/-) mice to in utero VPA exposure was demonstrated in some dietary folate regimens. Thus, our data indicate a relatively frail relationship between Folbp2 and VPA-induced NTDs.

DNA methylation in Folbp1 knockout mice supplemented with folic acid during gestation.

Periconceptional folic acid supplementation has been shown to prevent up to 70% of neural tube and other birth defects in humans; however, the mechanism is still unknown. In this study, we tested whether defective intracellular folate transport, as achieved by inactivation of the murine folate-binding protein 1 (Folbp1), affects global DNA methylation in the liver and brain from gestational day (GD) 15 embryos. Complete Folbp1 inactivation is embryolethal but can be reversed by maternal folinic acid (FA) supplementation, and thus we also tested the effect of FA supplementation on DNA methylation in Folbp1 fetuses. Overall, the extent of global DNA methylation seems to be similar across all genotypes in unsupplemented control Folbp1 mice; however, explicit conclusions regarding Folbp1(-/-) fetuses were not possible because only a single living unsupplemented fetus was viable at GD 15. FA supplementation induced global DNA hypomethylation across all genotypes. FA-induced hypomethylation is most likely due to its ability to inhibit the enzyme glycine hydroxymethyltransferase, thereby inhibiting the homocysteine remethylation cycle necessary to regenerate S-adenosylmethionine, the methyl donor for DNA methyltransferases. Our hypothesis was that due to defective folate transport in Folbp1(-/-) embryos and fetuses, DNA would be hypomethylated, thereby altering the temporal expression of critical genes necessary for normal embryonic development. However, these results suggest that an extended examination of changes in DNA methylation prior to GD 15 is required to unequivocally prove or disprove the hypothesis.