Clinical Pharmacokinetics and the Impact of Genetic Polymorphism on a CYP2C19 Substrate, BMS-823778, in Healthy Subjects.

BMS-823778 is a potent and selective inhibitor of 11ß-HSD1, an enzyme that regulates tissue-specific intracellular glucocorticoid metabolism and is a compelling target for the treatment of metabolic diseases. Metabolism of BMS-823778 was mediated mainly by polymorphic CYP2C19, with minor contributions from CYP3A4/5 and UGT1A4. The clinical pharmacokinetics (PK) of BMS-823778 was first investigated in healthy volunteers after single and multiple ascending doses. BMS-823778 was rapidly absorbed after the oral dose, and systemic exposure at steady state increased proportionally to the dose. Large intersubject variability in BMS-823778 exposure was likely because of the polymorphism of metabolic enzymes. The impact of genetic polymorphism of CYP2C19, UGT1A4, and CYP3A5 on BMS-823778 PK was assessed in healthy Chinese and Japanese subjects, as well as in a human absorption, distribution, metabolism, and excretion study in which all subjects were genotyped either before or after treatment. A clear trend of high exposure and low clearance was seen in poor metabolizers (PMs) of CYP2C19 compared with extensive (EM) and intermediate metabolizer (IM) subjects. The impact of UGT1A4 or CYP3A5 polymorphism on BMS-823778 PK was statistically not significant in CYP2C19 EM and IM subjects; however, in a subject with predicted CYP2C19 PM phenotype, the PK of BMS-823778 was affected significantly by UGT1A4 polymorphism. Overall, BMS-823778 was safe and well tolerated in healthy subjects after single or multiple oral doses. The PK of BMS-823778 was characterized by rapid absorption, and the systemic clearance directly correlated with the genetic polymorphism of CYP2C19.

Physiologically-based pharmacokinetic modelling of a CYP2C19 substrate, BMS-823778, utilizing pharmacogenetic data.

AIMS: Previous studies demonstrated direct correlation between CYP2C19 genotype and BMS-823778 clearance in healthy volunteers. The objective of the present study was to develop a physiologically-based pharmacokinetic (PBPK) model for BMS-823778 and use the model to predict PK and drug-drug interaction (DDI) in virtual populations with multiple polymorphic genes. METHODS: The PBPK model was built and verified using existing clinical data. The verified model was simulated to predict PK of BMS-823778 and significance of DDI with a strong CYP3A4 inhibitor in subjects with various CYP2C19 and UGT1A4 genotypes. RESULTS: The verified PBPK model of BMS-823778 accurately recovered observed PK in different populations. In addition, the model was able to capture the exposure differences between subjects with different CYP2C19 genotypes. PK simulation indicated higher exposures of BMS-823778 in CYP2C19 poor metabolizers who were also devoid of UGT1A4 activity, compared to those with normal UGT1A4 functionality. Moderate DDI with itraconazole was predicted in subjects with wild-type CYP2C19 or UGT1A4. However, in subjects without CYP2C19 or UGT1A4 functionality, significant DDI was predicted when BMS-823778 was coadministered with itraconazole. CONCLUSIONS: A PBPK model was developed using clinical data that accurately predicted human PK in different population with various CYP2C19 phenotypes. Simulations with the verified PBPK model indicated that UGT1A4 was probably an important clearance pathway in CYP2C19 poor metabolizers. DDI with itraconazole is likely to be dependent on the genotypes of CYP2C19 and UGT1A4.

Clinical significance of CYP2C19 polymorphisms on the metabolism and pharmacokinetics of 11ß-hydroxysteroid dehydrogenase type-1 inhibitor BMS-823778.

AIMS: BMS-823778 is an inhibitor of 11ß-hydroxysteroid dehydrogenase type-1, and thus a potential candidate for Type 2 diabetes treatment. Here, we investigated the metabolism and pharmacokinetics of BMS-823778 to understand its pharmacokinetic variations in early clinical trials. METHODS: The metabolism of BMS-823778 was characterized in multiple in vitro assays. Pharmacokinetics were evaluated in healthy volunteers, prescreened as CYP2C19 extensive metabolizers (EM) or poor metabolizers (PM), with a single oral dose of [14 C]BMS-823778 (10 mg, 80 µCi). RESULTS: Three metabolites (<5%) were identified in human hepatocytes and liver microsomes (HLM) incubations, including two hydroxylated metabolites (M1 and M2) and one glucuronide conjugate (M3). As the most abundant metabolite, M1 was formed mainly through CYP2C19. M1 formation was also correlated with CYP2C19 activities in genotyped HLM. In humans, urinary excretion of dosed radioactivity was significantly higher in EM (68.8%; 95% confidence interval 61.3%, 76.3%) than in PM (47.0%; 43.5%, 50.6%); only small portions (<2%) were present in faeces or bile from both genotypes. In plasma, BMS-823778 exposure in PM was significantly (5.3-fold, P = 0.0097) higher than in EM. Furthermore, total radioactivity exposure was significantly higher (P < 0.01) than BMS-823778 exposure in all groups, indicating the presence of metabolites. M1 was the only metabolite observed in plasma, and much lower in PM. In urine, the amount of M1 and its oxidative metabolite in EM was 7-fold of that in PM, while more glucuronide conjugates of BMS-823778 and M1 were excreted in PM. CONCLUSIONS: CYP2C19 polymorphisms significantly impacted systemic exposure and metabolism pathways of BMS-823778 in humans.

The effect of ketoconazole on the pharmacokinetics and pharmacodynamics of ixabepilone: a first in class epothilone B analogue in late-phase clinical development.

PURPOSE: To determine if ixabepilone is a substrate for cytochrome P450 3A4 (CYP3A4) and if its metabolism by this cytochrome is clinically important, we did a clinical drug interaction study in humans using ketoconazole as an inhibitor of CYP3A4. EXPERIMENTAL DESIGN: Human microsomes were used to determine the cytochrome P450 enzyme(s) involved in the metabolism of ixabepilone. Computational docking (CYP3A4) studies were done for epothilone B and ixabepilone. A follow-up clinical study was done in patients with cancer to determine if 400 mg/d ketoconazole (inhibitor of CYP3A4) altered the pharmacokinetics, drug-target interactions, and pharmacodynamics of ixabepilone. RESULTS: Molecular modeling and human microsomal studies predicted ixabepilone to be a good substrate for CYP3A4. In patients, ketoconazole coadministration resulted in a maximum ixabepilone dose administration to 25 mg/m(2) when compared with single-agent therapy of 40 mg/m(2). Coadministration of ketoconazole with ixabepilone resulted in a 79% increase in AUC(0-infinity). The relationship of microtubule bundle formation in peripheral blood mononuclear cells to plasma ixabepilone concentration was well described by the Hill equation. Microtubule bundle formation in peripheral blood mononuclear cells correlated with neutropenia. CONCLUSIONS: Ixabepilone is a good CYP3A4 substrate in vitro; however, in humans, it is likely to be cleared by multiple mechanisms. Furthermore, our results provide evidence that there is a direct relationship between ixabepilone pharmacokinetics, neutrophil counts, and microtubule bundle formation in PBMCs. Strong inhibitors of CYP3A4 should be used cautiously in the context of ixabepilone dosing.

Population pharmacokinetic analysis of topotecan in pediatric cancer patients.

PURPOSE: To characterize the population pharmacokinetics of topotecan lactone in children with cancer and identify covariates related to topotecan disposition. PATIENTS AND METHODS: The study population consisted of 162 children in seven clinical trials receiving single agent topotecan as a 30-min infusion. A population approach via nonlinear mixed effects modeling was used to conduct the analysis. RESULTS: A two-compartment model was fit to topotecan lactone plasma concentrations (n = 1874), and large pharmacokinetic variability was observed among studies, among individuals, and within individuals. We conducted a covariate analysis using demographics, biochemical data, trial effects, and concomitant drugs. The most significant covariate was body surface area, which explained 54% of the interindividual variability for topotecan systemic clearance. Interoccasion variability was considerable in both clearance and volume (20% and 22%, respectively), but was less than interindividual variability in both variables. Other covariates related to clearance were concomitant phenytoin, calculated glomerular filtration rate, and age (<0.5 years). Including them in the model reduced the interindividual variability for topotecan clearance by an additional 48% relative to the body surface area-normalized model. The full covariate model explained 76% and 50% of interindividual variability in topotecan clearance and volume, respectively. CONCLUSIONS: We developed a descriptive and robust population pharmacokinetic model which identified patient covariates that account for topotecan disposition in pediatric patients. Additionally, dosing topotecan based on the covariate model led to a more accurate and precise estimation topotecan systemic exposure compared with a fixed dosing approach, and could be a tool to assist clinicians to individualize topotecan dosing.

Phase I and pharmacokinetic study of gefitinib in children with refractory solid tumors: a Children's Oncology Group Study.

PURPOSE: Epidermal growth factor receptor is expressed in pediatric malignant solid tumors. We conducted a phase I trial of gefitinib, an epidermal growth factor receptor tyrosine kinase inhibitor, in children with refractory solid tumors. PATIENTS AND METHODS: Gefitinib (150, 300, 400, or 500 mg/m2) was administered orally to cohorts of three to six patients once daily continuously until disease progression or significant toxicity. Pharmacokinetic studies were performed during course one (day 1 through 28). RESULTS: Of the 25 enrolled patients, 19 (median age, 15 years) were fully evaluable for toxicity and received 54 courses. Dose-limiting toxicity was rash in two patients treated with 500 mg/m2 and elevated ALT and AST in one patient treated with 400 mg/m2. The maximum-tolerated dose was 400 mg/m2/d. The most frequent non-dose-limiting toxicities were grade 1 or 2 dry skin, anemia, diarrhea, nausea, and vomiting. One patient with Ewing's sarcoma had a partial response. Disease stabilized for 8 to > or = 60 weeks in two patients with Wilms' tumor and two with brainstem glioma (one exophytic). At 400 mg/m2, the median peak gefitinib plasma concentration was 2.2 microg/mL (range, 1.2 to 3.6 microg/mL) and occurred at a median of 2.3 hours (range, 2.0 to 8.3 hours) after drug administration. The median apparent clearance and median half-life were 14.8 L/h/m2 (range, 3.8 to 24.8 L/h/m2) and 11.7 hours (range, 5.6 to 22.8 hours), respectively. Gefitinib systemic exposures were comparable with those associated with antitumor activity in adults. CONCLUSION: Oral gefitinib is well tolerated in children. Development of the drug in combination with cytotoxic chemotherapy will be pursued.

Using plasma topotecan pharmacokinetics to estimate topotecan exposure in cerebrospinal fluid of children with medulloblastoma.

The purpose of this study was to estimate ventricular cerebrospinal fluid (vCSF) topotecan lactone (TPT) exposures in pediatric medulloblastoma patients from plasma concentration-time data by using a pharmacokinetic (PK) model. We studied children with high-risk medulloblastoma who received pharmacokinetically guided TPT (target plasma area under the concentration-time curve [AUC], 120-160 ng/ml-h) and obtained serial vCSF samples to assess TPT exposure. Population pharmacokinetic parameters were determined by using linear mixed-effects modeling via the two-stage approach. We simulated TPT vCSF exposure duration at plasma TPT AUC values of 120 to 200 ng/ml-h and determined percentages of studies meeting or exceeding the vCSF exposure duration threshold (EDT) of 1 ng/ml for 8 h. We then used bootstrap methods to estimate variability in vCSF EDT. Eighteen PK studies were conducted in six patients (median age, 4.8 years). In these patients, seven of nine studies attaining target plasma TPT AUC achieved the vCSF EDT. Given a plasma TPT AUC of 120 ng/ml-h, the median percentage of results meeting or exceeding EDT was 78% (95% CI, 61%-100%). One patient (four studies) with tumor blockage of CSF flow, which can alter CSF pharmacokinetics, was removed, and the bootstrap analysis was repeated. In this subset, a median 93% (95% CI, 79%-100%) of studies achieved vCSF EDT. Increasing plasma TPT AUC values resulted in increased ability to achieve vCSF EDT. We demonstrated that a plasma PK model could estimate vCSF TPT concentrations. Further, our results indicate that the TPT vCSF EDT can be achieved in more than 80% of studies targeted to a plasma TPT AUC of 120 ng/ml-h.

Development and validation of limited sampling models for topotecan lactone pharmacokinetic studies in children.

PURPOSE: To develop and validate a pharmacokinetic limited sampling model (LSM) for intravenous and oral topotecan pharmacokinetic studies in children. METHODS: Topotecan lactone concentration-time data from five trials were used to develop and validate LSM for intravenous and oral topotecan. Based on full sampling from one intravenous study (30 patients; 195 studies), a LSM for intravenous topotecan was determined using a modification of the D-optimality algorithm. For oral topotecan we used full sampling data from one oral topotecan study (27 patients; 47 studies) to develop an LSM. Accuracy and bias of each LSM were determined relative to the full sampling method. Predictive performance of the LSM was validated using additional data and Monte-Carlo simulations based on these data. RESULTS: LSM for intravenous topotecan includes: 5 min, 1.5, and 2.5 h after the end of the 30 min infusion. The median accuracy (absolute predicted error) and bias (predicted error) are < or =8% and < or =6.1%, respectively. For oral topotecan, the optimal LSM includes: 15 min, 1.5, and 6 h. The median accuracy and bias are 6% and 4%, respectively. CONCLUSIONS: Our results indicate that the optimal sampling times for the intravenous LSM for topotecan in children consist of: predose, and 5 min, 1.5, and 2.5 h after the end of infusion. For oral topotecan the sample times are predose, 15 min, 1.5, and 6 h after dose administration. These LSM are invaluable to children receiving topotecan because it minimizes inconvenience and blood collection.

A validated enantioselective LC-MS/MS assay for quantification of a major chiral metabolite of an achiral 11-ß-hydroxysteroid-dehydrogenase 1 inhibitor in human plasma: Application to a clinical pharmacokinetic study.

BMS-823778 is a potent 11-ß-hydroxysteroid-dehydrogenase 1 (11ßHSD-1) inhibitor and a potential therapeutic agent for type 2 diabetes mellitus (T2DM). A high performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay was developed and validated to enable reliable separation and quantification of both enantiomers of a chiral hydroxy metabolite (BMT-094817) in human plasma. Following liquid-liquid extraction in a 96-well plate format, chromatographic separation of the metabolite enantiomers was achieved by isocratic elution on a Chiralpak IA-3 column. Chromatographic conditions were optimized to ensure separation of both metabolite enantiomers. Metabolite enantiomers and stable isotope-labeled (SIL) internal standards were detected by positive ion electrospray tandem mass spectrometry. The LC-MS/MS assay was validated over a concentration range of 0.200-200ng/mL. Intra- and inter-assay precision values for replicate quality control samples were less than 9.9% for both enantiomers during the assay validation. Mean quality control accuracy values were within ±7.3%. Assay recoveries were high (>75%) and consistent across the assay range. The metabolite enantiomers were stable in human blood for 2h on ice. The analytes were also stable in human plasma for 25h at room temperature, 34days at -20°C and -70°C, and following five freeze-thaw cycles. No interconversion of the metabolite enantiomers was detected under any bioanalytical stress conditions, from blood collection/processing through extracted sample storage. The validated assay was successfully applied to the quantification of both metabolite enantiomers in human plasma in support of a human pharmacokinetic study.

Phase I and pharmacokinetic study of topotecan administered orally once daily for 5 days for 2 consecutive weeks to pediatric patients with refractory solid tumors.

PURPOSE: We conducted a phase I trial of the injectable formulation of topotecan given orally once daily for 5 days for 2 consecutive weeks (qd x 5 x 2) in pediatric patients with refractory solid tumors. PATIENTS AND METHODS: Cohorts of two to six patients received oral topotecan at 0.8, 1.1, 1.4, 1.8, and 2.3 mg/m(2)/d every 28 days for a maximum of six courses. Twenty patients (median age, 10.6 years) received a total of 51 courses. Eight patients received topotecan capsules during course 2 only. RESULTS: Dose-limiting toxicity occurred at 2.3 mg/m(2)/d and consisted of prolonged grade 4 neutropenia (n = 2), grade 3 stomatitis as a result of radiation recall (n = 1), grade 3 hemorrhage (epistaxis) in the presence of grade 4 thrombocytopenia (n = 1), and grade 3 diarrhea in the presence of Clostridium difficile infection (n = 1). Dose-limiting, prolonged grade 4 neutropenia and thrombocytopenia occurred in one patient at 1.4 mg/m(2)/d. Infrequent toxicities were mild nausea, vomiting, elevated liver ALT or AST, and rash. The maximum-tolerated dosage was 1.8 mg/m(2)/d; the mean (+/- standard deviation) area under the plasma concentration-time curve for topotecan lactone at this dosage was 20.9 +/- 8.4 ng/mL. h. The population mean (+/- standard error) oral bioavailability of the injectable formulation was 0.27 +/- 0.03; that of capsules was 0.36 +/- 0.06 (P =.16). Disease stabilized in nine of 19 assessable patients for 1.5 to 6 months. CONCLUSION: Oral topotecan (1.8 mg/m(2)/d) on a qd x 5 x 2 schedule is well tolerated and warrants additional testing in pediatric patients.

Results of a phase II upfront window of pharmacokinetically guided topotecan in high-risk medulloblastoma and supratentorial primitive neuroectodermal tumor.

PURPOSE: To assess the antitumor efficacy of pharmacokinetically guided topotecan dosing in previously untreated patients with medulloblastoma and supratentorial primitive neuroectodermal tumors, and to evaluate plasma and CSF disposition of topotecan in these patients. PATIENTS AND METHODS: After maximal surgical resection, 44 children with previously untreated high-risk medulloblastoma were enrolled, of which 36 were assessable for response. The topotecan window consisted of two cycles, administered initially as a 30-minute infusion daily for 5 days, lasting 6 weeks. Pharmacokinetic studies were conducted on day 1 to attain a topotecan lactone area under the plasma concentration-time curve (AUC) of 120 to 160 ng/mL.h. After 10 patients were enrolled, the infusion was modified to 4 hours, with dosage individualization. RESULTS: Of 36 assessable patients, four patients (11.1%) had a complete response and six (16.6%) showed a partial response, and disease was stable in 17 patients (47.2%). Toxicity was mostly hematologic, with only one patient experiencing treatment delay. The target plasma AUC was achieved in 24 of 32 studies (75%) in the 30-minute infusion group, and in 58 of 93 studies (62%) in the 4-hour infusion group. The desired CSF topotecan exposure was achieved in seven of eight pharmacokinetic studies when the topotecan plasma AUC was within target range. CONCLUSION: Topotecan is an effective agent against pediatric medulloblastoma in patients who have received no therapy other than surgery. Pharmacokinetically guided dosing achieved the target plasma AUC in the majority of patients. This drug warrants testing as part of standard postradiation chemotherapeutic regimens. Furthermore, these results emphasize the importance of translational research in drug development, which in this case identified an effective drug.

Activation and antitumor activity of CPT-11 in plasma esterase-deficient mice.

PURPOSE: To examine the antitumor activity and the pharmacokinetics of CPT-11 (irinotecan, 7-ethyl-10-[4-(1-piperidino)-1-piperidino] carbonyloxycamptothecin) in a plasma esterase-deficient scid mouse model, bearing human tumor xenografts. EXPERIMENTAL DESIGN: Plasma carboxylesterase (CE)-deficient mice were bred with scid animals to develop a strain that would allow growth of human tumor xenografts. Following xenotransplantation, the effect of the plasma esterase on antitumor activity following CPT-11 administration was assessed. In addition, detailed pharmacokinetic studies examining plasma and biliary disposition of CPT-11 and its metabolites were performed. RESULTS: In mice lacking plasma carboxylesterase, the mean SN-38 systemic exposures were approximately fourfold less than that observed in control animals. Consistent with the pharmacokinetic data, four to fivefold more CPT-11 was required to induce regressions in human Rh30 xenografts grown in esterase-deficient scid mice, as opposed to those grown in scid animals. Additionally, the route of elimination of CPT-11, SN-38, and SN-38 glucuronide (SN-38G) was principally in the bile. CONCLUSIONS: The pharmacokinetic profile for CPT-11 and its metabolites in the esterase-deficient mice more closely reflects that seen in humans. Hence, these mice may represent a more accurate model for antitumor studies with this drug and other agents metabolized by CEs.

Development of a pharmacokinetic limited sampling model for temozolomide and its active metabolite MTIC.

PURPOSE: To develop a pharmacokinetic limited sampling model (LSM) for temozolomide and its metabolite MTIC in infants and children. METHODS: LSMs consisting of either two or four samples were determined using a modification of the D-optimality algorithm. This accounted for prior distribution of temozolomide and MTIC pharmacokinetic parameters based on full pharmacokinetic sampling from 38 patients with 120 pharmacokinetic studies (dosage range 145-200 mg/m(2) per day orally). Accuracy and bias of each LSM were determined relative to the full sampling method. We also assessed the predictive performance of the LSMs using Monte-Carlo simulations. RESULTS: The four strategies generated from the D-optimality algorithm were as follows: LSM 1=0.25, 1.25, and 3 h; LSM 2=0.25, 1.25, and 6 h; LSM 3=0.25, 0.5, 1.25, and 3 h; LSM 4=0.25, 0.5, 1.25, and 6 h. LSM 2 demonstrated the best combination of low bias [0.1% (-8.9%, 11%) and 11% (4.3%, 15%)] and high accuracy [-1.0% (-12%, 24%) and 14% (7.9%, 37%)] for temozolomide clearance and MTIC AUC, respectively. Furthermore, adding a fourth sample (e.g., LSM 4) did not substantially decrease the bias or increase the accuracy for temozolomide clearance or MTIC AUC. Results from Monte-Carlo simulations also revealed that LSM 2 had the best combination of lowest bias (0.1+/-6.1% and -0.8+/-6.5%), and the highest accuracy (4.5+/-4.1% and 5.0+/-4.3%) for temozolomide clearance and MTIC apparent clearance, respectively. CONCLUSIONS: Using data derived from our population analysis, the sampling times for a limited sample pharmacokinetic model for temozolomide and MTIC in children are prior to the temozolomide dose, and 15 min, 1.25 h and 6 h after the dose.

The importance of pharmacokinetic limited sampling models for childhood cancer drug development.

Since the development of effective chemotherapy for children with cancer, it has been recognized that the response of children to apparently identical therapy, both in terms of efficacy and toxicity, can vary widely. Our understanding of the interindividual differences in drug metabolism and disposition as significant determinants of drug response continues to evolve. An increasing area of clinical investigation is focused on studies to gain a better understanding of the variability in critical drug metabolic and elimination pathways and how this variability translates into varied pharmacological effects. Analyzing how drug metabolism and elimination are affected by patient characteristics such as age, sex, race, organ function, drug interactions, and, perhaps most importantly, genetic polymorphisms, is now a routine component of drug development studies. Recent advances in analytical methodologies, computer hardware, and pharmacokinetic software have improved our ability to conduct studies of the disposition of anticancer drugs in larger, more representative pediatric cancer populations. Along with advances in pharmacogenetics, the advances made in the conduct of pharmacokinetic studies in children with cancer have enabled establishment of sophisticated phenotype-genotype correlations, which may ultimately improve care. However, unique challenges and limitations remain that complicate the performance of pharmacokinetic studies in the child with cancer. This review addresses the need to perform pharmacokinetic studies throughout the drug development process in pediatric oncology patients, methods used to develop and validate limited sampling models, and selected examples of limited sampling models used in pharmacokinetic studies in children with cancer.

Phase I trial of temozolomide and protracted irinotecan in pediatric patients with refractory solid tumors.

PURPOSE: The purpose is to estimate the maximum-tolerated dose (MTD) of temozolomide and irinotecan given on a protracted schedule in 28-day courses to pediatric patients with refractory solid tumors. EXPERIMENTAL DESIGN: Twelve heavily pretreated patients received 56 courses of oral temozolomide at 100 mg/m(2)/day for 5 days combined with i.v. irinotecan given daily for 5 days for 2 consecutive weeks at either 10 mg/m(2)/day (n = 6) or 15 mg/m(2)/day (n = 6). We assessed toxicity, the pharmacokinetics of temozolomide and irinotecan, and the DNA repair phenotype in tumor samples. RESULTS: Two patients experienced dose-limiting toxicity (DLT) at the higher dose level; one had grade 4 diarrhea, whereas the other had bacteremia with grade 2 neutropenia. In contrast, no patient receiving temozolomide and 10 mg/m(2)/day irinotecan experienced DLT. Myelosuppression was minimal and noncumulative. No pharmacokinetic interaction was observed. Drug metabolite exposures at the MTD were similar to exposures previously associated with single-agent antitumor activity. One complete response, two partial responses, and one minor response were observed in Ewing's sarcoma and neuroblastoma patients previously treated with stem cell transplant. Responding patients had low or absent O(6)-methylguanine-DNA methyltransferase expression in tumor tissue. CONCLUSIONS: The MTD using this schedule was temozolomide (100 mg/m(2)/day) and irinotecan (10 mg/m(2)/day), with DLT being diarrhea and infection. Drug clearance was similar to single-agent values, and clinically relevant SN-38 lactone and MTIC exposures were achieved at the MTD. As predicted by xenograft models, this combination and schedule appears to be tolerable and active in pediatric solid tumors. Evaluation of a 21-day schedule is planned.

Reinventing the wheel of cyclic AMP: novel mechanisms of cAMP signaling.

Mechanisms of cAMP signal transduction have been thoroughly investigated for more than 40 years. From the binding of hormonal ligands to their receptors on the outer surface of the plasma membrane to the cytoplasmic activation of effectors, the ensuing cAMP signaling cascades and the nuclear gene regulatory functions, coupled with the structural elucidation of the cAMP-dependent protein kinase (PKA) and in vivo functional characterizations of each of the components of PKA by homologous recombination gene targeting, our understanding of cAMP-mediated signal transduction has reached its pinnacle. Despite this trove of knowledge, some recent findings have emerged that suggest hitherto novel and alternative mechanisms of cAMP action that could increase the signaling bandwidth of cAMP and PKA in cell growth and transcriptional regulation. This article attempts to review some of these novel and unconventional mechanisms of cAMP and PKA signaling, and to generate further enthusiasm in investigating and validating these new frontiers of the cAMP signal transduction pathway.

Determination of plasma topotecan and its metabolite N-desmethyl topotecan as both lactone and total form by reversed-phase liquid chromatography with fluorescence detection.

Topotecan (TPT) undergoes hepatic N-demethylation forming N-desmethyl topotecan (NDS). To evaluate the effect of drug-drug interactions on NDS disposition in children receiving TPT we developed and validated a sensitive and specific HPLC-fluorescence detection method for lactone and total (lactone plus carboxylate) TPT and NDS. Deproteinized plasma is vortexed, centrifuged, and the methanolic extract diluted with water for the lactone form of NDS and TPT or diluted with 1.5% phosphoric acid for NDS and TPT total. A 100 microL sample is injected onto a Varian ChromGuard RP column attached to an Agilent SB-C(18) reversed-phase analytical column held at 50 degrees C. The mobile phase (flow-rate, 0.8 mL/min) consists of methanol-aqueous buffer (27:73, v/v) (75 mM potassium phosphate and 0.2% triethylamine, pH 6.5). TPT and NDS were detected with excitation and emission wavelengths set at 376 and 530 nm, respectively. The standard curves for both forms of TPT ranged from 0.25 to 80 ng/mL, and for NDS ranged from 0.10 to 8.0 ng/mL. Within-day and between-day precision (% RSD) was

Topoisomerase I interactive agents.

Increased insight into the mechanism of interaction of topoisomerase I interactive agents will maximize the therapeutic index and enhance the development of additional agents. Preclinical studies designed to elucidate mechanisms by which the topoisomerase I interactive agents induce cell death will be essential. The role of ABC transporters in resistance to topoisomerase I interactive agents has been recently appreciated and future studies should be directed at circumventing this resistance. The results of preclinical studies must be translated into the design of clinical trials so that these agents can be used rationally. In this regard results of preclinical studies have clearly pointed to the enhanced antitumor activity from protracted dosing of topoisomerase I interactive agents and results of clinical trials are now supporting these preclinical findings. Finally, investigators are trying to understand better the mechanism(s) of the dose-limiting toxicities observed with the currently available topoisomerase I interactive agents in an effort to enable the optimal dosing of these agents. Even though the first priority must be to determine the therapeutic potential of the currently available agents, it is reassuring to know that other topoisomerase I interactive agents are currently under development.

A phase I study of oral ixabepilone in patients with advanced solid tumors.

BACKGROUND: Intravenous infusion of ixabepilone is Food and Drug Administration-approved for treatment of patients with metastatic breast cancer. The aim of this study was to establish the maximum tolerated dose (MTD), dose-limiting toxicities (DLTs), safety, and pharmacokinetics (PK) of a novel oral formulation of ixabepilone in patients with advanced solid tumors. PATIENTS AND METHODS: Forty-four patients received one of six daily doses of oral ixabepilone (5, 10, 15, 20, 25, or 30 mg) on days 1-5 of a 21-day cycle. PK parameters were evaluated in cycle 1 for all treated patients and in cycle 1 and cycle 2 for patients participating in assessments of food and gastric pH effects. RESULTS: The most common DLTs (reported in at least one patient) were neutropenia, neutropenic fever, diarrhea, ileus, and hypokalemia. The MTD of oral ixabepilone was 25 mg. Plasma concentrations of ixabepilone showed high variability; coefficients of variation for the area under the curve and the peak plasma concentration ranged from 61 to 131 % and from 17 to 172 %, respectively. The mean half-life of ixabepilone calculated after day 5 of cycle 1 ranged from 24 to 47 h. Ixabepilone exposure was higher when administered with a low-fat meal compared with the fasted state, and when administered 2 h after the histamine H2 receptor antagonist famotidine. CONCLUSIONS: The MTD of oral ixabepilone when administered once daily for five consecutive days every 21 days was 25 mg. Ixabepilone exposure was highly variable; therefore, safety and efficacy of this novel oral formulation might not be reliably predicted.

Phase I and pharmacokinetic study of CT-322 (BMS-844203), a targeted Adnectin inhibitor of VEGFR-2 based on a domain of human fibronectin.

PURPOSE: To determine the maximum tolerated dose (MTD), safety, pharmacokinetics, pharmacodynamics, immunogenicity, and preliminary antitumor activity of CT-322 (BMS-844203), a VEGFR-2 inhibitor and the first human fibronectin domain-based targeted biologic (Adnectin) to enter clinical studies. EXPERIMENTAL DESIGN: Patients with advanced solid malignancies were treated with escalating doses of CT-322 intravenously (i.v.) weekly (qw), or biweekly (q2w). Plasma samples were assayed for CT-322 concentrations, plasma VEGF-A concentrations, and antidrug antibodies. RESULTS: Thirty-nine patients completed 105 cycles of 0.1 to 3.0 mg/kg CT-322 i.v. either qw or q2w. The most common treatment-emergent grade 1/2 toxicities were fatigue, nausea, proteinuria, vomiting, anorexia, and hypertension. Grade 3/4 toxicities were rare. Reversible proteinuria, retinal artery, and vein thrombosis, left ventricular dysfunction, and reversible posterior leukoencephalopathy syndrome were dose limiting at 3.0 mg/kg. The MTD was 2 mg/kg qw or q2w. CT-322 plasma concentrations increased dose proportionally. Plasma VEGF-A levels increased with dose and plateaued at 2 mg/kg qw. Anti-CT-322 antibodies developed without effects on pharmacokinetics, VEGF-A levels, or safety. Minor decreases in tumor measurements occurred in 4 of 34 evaluable patients and 24 patients had stable disease. CONCLUSIONS: CT-322 can be safely administered at 2 mg/kg i.v. qw or q2w and exhibits promising antitumor activity in patients with advanced solid tumors. The absence of severe toxicities at the MTD, demonstration of plasma drug concentrations active in preclinical models, and clinical pharmacodynamic evidence of VEGFR-2 inhibition warrant further development of CT-322 and suggest strong potential for Adnectin-based targeted biologics.