We previously reported interim results of a phase 1 trial in patients with chronic lymphocytic leukemia (CLL) whereby flavopiridol was administered intravenously as a 30-minute bolus followed by 4-hour infusion. We now report full pharmacokinetic (PK) data, correlations of PK with clinical outcomes, and final response and progression-free survival (PFS). Twenty-one (40%) of 52 patients with relapsed CLL achieved a partial response (PR) with a median PFS of 12 months. Responders included 17 (40%) of 43 fludarabine refractory patients, 7 (39%) of 18 patients with del(17p13), and 14 (74%) of 19 patients with del(11q22). Six responders received repeat therapy at relapse, and 5 responded again with a second median PFS of 10 months. Noncompartmental analysis and nonlinear mixed effects modeling was used to estimate PK parameters and evaluate covariates. Two-compartment population parameter estimates were 31.4 L/h, 65.8 L, 8.49 L/h, and 157 L for CL, V1, Q, and V2, respectively. Flavopiridol area under the plasma concentration-time curve (AUC) correlated with clinical response and cytokine release syndrome, and glucuronide metabolite AUC correlated with tumor lysis syndrome. These composite results confirm high activity of this pharmacokinetically derived schedule in relapsed, genetically high-risk CLL. Furthermore, PK describes some, but not all, variability in response and toxicity.

BackgroundFlavopiridol is a cyclin-dependent kinase inhibitor in phase II clinical development for treatment of various forms of cancer. When administered with a pharmacokinetically (PK)-directed dosing schedule, flavopiridol exhibited striking activity in patients with refractory chronic lymphocytic leukemia. This study aimed to evaluate pharmacogenetic factors associated with inter-individual variability in pharmacokinetics and outcomes associated with flavopiridol therapy.Methodology/Principal FindingsThirty-five patients who received single-agent flavopiridol via the PK-directed schedule were genotyped for 189 polymorphisms in genes encoding 56 drug metabolizing enzymes and transporters. Genotypes were evaluated in univariate and multivariate analyses as covariates in a population PK model. Transport of flavopiridol and its glucuronide metabolite was evaluated in uptake assays in HEK-293 and MDCK-II cells transiently transfected with SLCO1B1. Polymorphisms in ABCC2, ABCG2, UGT1A1, UGT1A9, and SLCO1B1 were found to significantly correlate with flavopiridol PK in univariate analysis. Transport assay results indicated both flavopiridol and flavopiridol-glucuronide are substrates of the SLCO1B1/OATP1B1 transporter. Covariates incorporated into the final population PK model included bilirubin, SLCO1B1 rs11045819 and ABCC2 rs8187710. Associations were also observed between genotype and response. To validate these findings, a second set of data with 51 patients was evaluated, and overall trends for associations between PK and PGx were found to be consistent.Conclusions/SignificancePolymorphisms in transport genes were found to be associated with flavopiridol disposition and outcomes. Observed clinical associations with SLCO1B1 were functionally validated indicating for the first time its relevance as a transporter of flavopiridol and its glucuronide metabolite. A second 51-patient dataset indicated similar trends between genotype in the SLCO1B1 and other candidate genes, thus providing support for these findings. Further study in larger patient populations will be necessary to fully characterize and validate the clinical impact of polymorphisms in SLCO1B1 and other transporter and metabolizing enzyme genes on outcomes from flavopiridol therapy.

Lenalidomide, an immunomodulatory agent, and flavopiridol, a broad cyclin-dependent kinase inhibitor, are both active therapies for clinical use in genomic high risk chronic lymphocytic leukemia (CLL). A high-performance liquid chromatographic assay with tandem mass spectrometric detection has been developed to simultaneously quantify lenalidomide and flavopiridol in human and mouse plasma to facilitate their combined clinical development. Samples were prepared by liquid-liquid extraction with acetonitrile- (ACN) containing internal standard (IS), genistein, followed by evaporation of solvent and reconstitution in 95/5 H2O/ACN. Lenalidomide and IS were separated by reverse phase liquid chromatography on a C-18 column using a gradient of H2O and ACN, each with 0.1% formic acid. Atmospheric pressure chemical ionization (APCI) in positive-ion mode with single reaction monitoring on a triple quadrupole mass spectrometer were applied to detect transitions of lenalidomide (260.06 > 149.10) and flavopiridol (402.09 > 341.02). Lower limits of quantification (LLOQ) of lenalidomide and flavopiridol were 1nM and 0.3nM respectively. Recoveries of lenalidomide and flavopiridol from human plasma ranged from 99% to 116% throughout their linear ranges. Within and between-run precision and accuracy of replicate samples were all less than 15%. This is the most sensitive analytical method reported to date for both lenalidomide and flavopiridol. This sensitivity will enable late terminal phase concentration measurements and accurate pharmacokinetic parameter estimation in a planned clinical trial with lenalidomide and flavopiridol in CLL patients.

A high-performance liquid chromatographic assay with tandem mass spectrometric detection was developed and validated for quantitation of the broad spectrum kinase inhibitor, flavopiridol, in human plasma. Sample preparation conditions included liquid-liquid extraction in acetonitrile (ACN), drying, and reconstitution in 20/80 water/ACN. Flavopiridol and the internal standard (IS), genistein, were separated by reversed phase chromatography using a C-18 column and a gradient of water with 25 mM ammonium formate and ACN. Electrospray ionization and detection of flavopiridol and genistein were accomplished with single reaction monitoring of m/z 402.09>341.02 and 271.09>152.90, respectively in positive-ion mode [M+H](+) on a triple quadrupole mass spectrometer. Recovery was greater than 90% throughout the linear range of 3-1000 nM. Replicate sample analysis indicated within- and between-run accuracy and precision to be less than 13% throughout the linear range. This method has the lowest lower limit of quantitation (LLOQ) reported to date for flavopiridol, and it allows for more accurate determination of terminal phase concentrations and improved pharmacokinetic parameter estimation in patients receiving an active dosing schedule of flavopiridol.

It is essential that clinical physicians, medical personnel, medical examiners, and law enforcement agencies understand the types of injuries seen and demographics of children affected by intentional blunt force as this understanding can be crucial to the death and/or criminal investigations. An understanding of the injuries can also assist in drawing conclusions regarding how those injuries could have been sustained. This study discusses the types and patterns of injuries seen in blunt force homicides in children younger than 6 years. The study found that male infants are more often intentionally injured than are female infants and that fatal head injuries most frequently occur in the first year of life, whereas most fatal thoracoabdominal injuries occur in the first 3 years of life. In children with head injuries, subdural hemorrhage was the most common finding, followed by subarachnoid hemorrhage. In 2.5% of deaths due to head injury, concurrent neck injury was seen, a percentage far lower than previous literature would suggest if shaking was the primary mechanism of injury. Twelve legal confessions were also reviewed, none of which disclosed a pure mechanism of shaking the infant.