The value, qualification, and regulatory use of surrogate end points in drug development.

The acceptance and use of either surrogate end points (SEPs) or efficient clinical end points are associated with greater and more rapid availability of new medicines as compared with disease situations for which clinical end points are inefficient or no surrogates exist. This review of the history of the development, qualification, and acceptance of key SEPs shows that both successes and failures had three key characteristics: (i) apparent biologic plausibility, (ii) prognostic value for the outcome of the disease, and (iii) an association between changes in the SEP and changes in outcome with therapeutic intervention--the three factors recommended for SEPs in the International Conference on Harmonisation's "Statistical Principles for Clinical Trials." We recommend that only prognostic value be an absolute prerequisite for surrogacy, because therapeutic interventions may not exist a priori, and biological plausibility can be subjective. Ideally, all three of these factors would be traded off against one another in a consistent and transparent risk-management process.

Development of a chiral HPLC method to evaluate in vivo enantiomeric inversion of an unstable, polar radiosensitizer in plasma.

A chiral HPLC method to quantify in vivo enantiomeric inversion of prodrug CI-1010 (IR) or its drug IIR (PD 146923), a radiosensitizer, upon X-irradiation of dosed rats was developed. These polar enantiomers were separated only by using normal-phase chiral HPLC. A Chiralpak AS column provided the best separation. Isolation of analytes from plasma employed solid-phase extraction (SPE), and required conditions that were compatible with normal-phase HPLC. Options for SPE were restricted by the chemically reactive nature of both prodrug and drug, which produced analyte losses as high as 100%. Acceptable recoveries using SPE required evaluation of conditions for analyte chemical stability. The validated method gave a lower-limit of quantitation (LLOQ) of 200 ng/ml for each enantiomer extracted from 0.15 ml of plasma. The LLOQ of the inverted enantiomer could be detected in the presence of 10,000 ng/ml of the dosed enantiomer. Precision (RSD) ranged from 14.2 to 4.4%, and from 24.2 to 5.1% for IIS and IIR, respectively. Accuracy (RE) was +/- 13.1 and +/- 13.2%, respectively. Recoveries ranged from 44.3 to 71.4%, and from 40.7 to 67.9%, for IIS and IIR, respectively.

Validated HPLC/MS/MS assay for CI-1011 in rat plasma and a comparison with an HPLC/UV assay.

A liquid chromatographic/mass spectrometric (LC/MS/MS) method to quantitate CI-1011 in rat plasma has been validated and compared to an LC/UV assay. The analyte and internal standard were isolated from the plasma matrix by using liquid/liquid extraction with diethyl ether. The ether layer was evaporated to dryness and the residue reconstituted in acetonitrile-water (70:30, v/v). A 2.1 x 150 mm x 5 microns Zorbax RX-C18 column with a mobile phase of acetonitrile-ammonium acetate (pH 8.0; 5 mM)-triethylamine (70:30:0.03, v/v/v) delivered at a flow rate of 0.2 ml min-1 was used for chromatography. Analyte and internal standard ion chromatograms were obtained by operating the mass spectrometer in the negative ion multiple reaction monitoring mode to detect the presence of a precursor-product ion pair for both the analyte and the internal standard. Samples were introduced into the mass spectrometer using electrospray ionization. Retention times of CI-1011 and of the internal standard (IS), [13C6]CI-1011, were approximately 4.2 min. No peaks interfering with the quantitation of CI-1011 were observed throughout the validation process. Mean recoveries of CI-1011 from rat plasma ranged from 98.2 to 105%. The recovery of the IS was 100%. Assay precision for CI-1011, based on the percent relative standard deviation of replicate quality controls, was less than or equal to 5.60% with an accuracy of +/- 8.80%. The lower limit of quantitation for CI-1011 was 0.500 ng ml-1 for a 0.2-ml sample aliquot. CI-1011 is stable in rat plasma for 24 h at room temperature and for at least 34 days at -20 degrees C. This assay has been proven suitable for routine quantitation of CI-1011 in rat plasma at concentrations from 0.500 (100 pg on-column) to 500 ng ml-1. The applicability of this method to determine CI-1011 concentrations in rat plasma is reported in this manuscript. CI-1011 concentrations, in plasma samples from cholesterol- and chow-fed rats administered single daily oral doses of CI-1011 in a CMC/Tween suspension, obtained using a validated LC/UV assay were compared to concentrations obtained using the reported LC/MS/MS assay over the concentration range 0.0806-12.3 micrograms ml-1. The concordance correlation coefficient determined for this comparison was 0.9977, suggesting that the CI-1011 concentrations obtained by the two assays are in excellent agreement.

Management of nonmatrix interfering peaks in a chiral high-performance liquid chromatographic assay produced by solid-phase extraction of rat plasma.

PD 146923, under evaluation as an alkylating radiosensitizing drug, contains one chiral center and one chemically reactive aziridine ring. A method was developed to evaluate possible in vivo enantiomeric inversion of PD 146923 in rat plasma. Normal-phase chiral HPLC was necessary to separate the enantiomers, but a typical aqueous-based solid-phase extraction (SPE) was needed to isolate the analytes from plasma. SPE at higher analyte concentrations removed all interfering peaks and gave acceptable recoveries. However, peaks (A-G) from seven new components interfering with analyte detection at lower concentrations were produced by SPE. The interfering peaks overlapped each other, so some were not observed until other, more intense interfering peaks had been managed. The low separation efficiency of the chiral column precluded management of interfering peaks by modifying chromatographic parameters. Chemical reactivity of the analytes forced the use of mild conditions for management of interfering peaks. Peaks A-F were: (A) water from the SPE cartridge; (B) SPE sorbent endcapping; (C, E and F) nonvolatile salts of the SPE elution acid reacting with bases from the injection solvent or with unidentified bases from the SPE cartridge; (D and G) analyte degradation products. This study identifies the nonmatrix peaks coeluting with the analytes, and describes how an aqueous-based SPE method was developed for isolating these very polar, highly reactive analytes in plasma for separation in a normal-phase chiral HPLC assay. Additionally, B, C, E or F probably are present in many other solid-phase extractions, but are not observed because of polarity or solubility properties.

A phase I trial and pharmacokinetic evaluation of CI-980 in patients with advanced solid tumors.

CI-980 is a synthetic mitotic inhibitor that binds to the colchicine binding site of tubulin. It demonstrates broad activity against human and murine tumor models and shows no cross resistance with tumor models whose mechanism of resistance is mediated by P-glycoprotein (MDR-1). A phase I study was completed in 25 patients with solid tumors using a 24-hour infusion schedule, with courses repeated every 3 weeks. Eight dose levels were tested between 1.2 and 15.6 mg/m2. The maximum tolerated dose was 14.4 mg/m2. Neutropenia was dose-related but not dose-limiting; thrombocytopenia was infrequent. CNS toxicities were dose-limiting and consisted of dizziness, headache, loss of coordination, loss of consciousness, nervousness, and other symptoms. These events occurred near the end of the infusion and were reversible, usually within 24 hours. One patient who was to be treated at dose level 8 (intended dose was 19.2 mg/m2; actual dose was 15.6 mg/m2) became encephalopathic prior to completion of the infusion. Other adverse events included gastrointestinal toxicities (nausea, vomiting, anorexia, constipation, stomatitis, dyspepsia, bleeding, cheilitis), IV site erythema, fever, and fatigue. A partial response was observed in one patient with colon cancer and reductions in CA-125 levels were observed in 2 patients with ovarian cancer. Pharmacokinetics were linear and dose-proportional. Results indicate high systemic clearance and wide tissue distribution. Mean pharmacokinetic parameter values: T1/2 = 5.52 hours, plasma clearance 1163 mL/min/m2, and Vdss 376 L/m2.

An HPLC assay utilizing solid-phase extraction for CI-1010, an alkylating radiosensitizer, in rat plasma.

CI-1010, a 2-nitroimidazole, is a chiral prodrug for the active moiety PD 146923 and is under development as an alkylating radiosensitizer to be used as an adjuvant to radiotherapy. Because CI-1010 has an estimated half-life < or = 2 min under physiological conditions its metabolites/degradation products PD 146415, an inactive moiety, and PD 146923 were assayed to support rat toxicology studies. The method involves the processing of plasma samples through phenyl solid-phase extraction cartridges followed by chromatography on CN columns with UV detection at 325 nm. The assay appears linear over the range 0.050-100 micrograms ml-1 for both PD 146415 and PD 146923. Interrun accuracy and precision estimates for PD 146415 and PD 146923 were within +/- 6.50 and < or = 3.27%, respectively, and +/- 12.8 and < or = 4.06%, respectively, for quality controls containing nominal concentrations of 0.400, 4.00 and 40.0 micrograms ml-1. The absolute recovery of CI-1010, PD 146415 and internal standard, PD 126675, were approximately 40, 96 and 95%, respectively. The recovery of PD 146923 appeared concentration dependent and ranged from 68 to 92%. PD 146415 and PD 146923 were both stable in rat plasma at 4 degrees C and -77 degrees C for at least 7 h and 154 days, respectively. CI-1010 was not stable in rat plasma at 4 degrees C. CI-1010, PD 146415 and PD 126675 were stable for at least 63 days in 10 mM phosphate buffer at pH 3.0 and 4 degrees C. Under identical conditions PD 146923 was stable for only 8 days. The applicability of this method to determine concentrations of PD 146415 and PD 146923 in rat plasma is reported in this paper.