Pharmacological and molecular characterization of a dorsal root ganglion cell line expressing cannabinoid CB(1) and CB(2) receptors.

The behavioral effects evoked by cannabinoids are primarily mediated by the CB(1) and CB(2) cannabinoid receptor subtypes. In vitro pharmacology of cannabinoid receptors has been elucidated using recombinant expression systems expressing either CB(1) or CB(2) receptors, with limited characterization in native cell lines endogenously expressing both CB(1) and CB(2) receptors. In the current study, we report the molecular and pharmacological characterization of the F-11 cell line, a hybridoma of rat dorsal root ganglion neurons and mouse neuroblastoma (N18TG2) cells, reported to endogenously express both cannabinoid receptors. The present study revealed that both receptors are of mouse origin in F-11 cells, and describes the relative gene expression levels between the two receptors. Pharmacological characterization of the F-11 cell line using cannabinoid agonists and antagonists indicated that the functional responses to these cannabinoid ligands are mainly mediated by CB(1) receptors. The non-selective cannabinoid ligands CP 55,940 and WIN 55212-2 are potent agonists and their efficacies in adenylate cyclase and MAPK assays are inhibited by the CB(1) selective antagonist SR141716A (SR1), but not by the CB(2) selective antagonist SR144528 (SR2). The endocannabinoid ligand 2AG, although not active in adenylate cyclase assays, was a potent activator of MAPK signaling in F-11 cells. The analysis of CB(1) and CB(2) receptor gene expression and the characterization of cannabinoid receptor pharmacology in the F-11 cell line demonstrate that it can be used as a tool for interrogating the endogenous signal transduction of cannabinoid receptor subtypes.

ABT-888, an orally active poly(ADP-ribose) polymerase inhibitor that potentiates DNA-damaging agents in preclinical tumor models.

PURPOSE: To evaluate the preclinical pharmacokinetics and antitumor efficacy of a novel orally bioavailable poly(ADP-ribose) polymerase (PARP) inhibitor, ABT-888. EXPERIMENTAL DESIGN: In vitro potency was determined in a PARP-1 and PARP-2 enzyme assay. In vivo efficacy was evaluated in syngeneic and xenograft models in combination with temozolomide, platinums, cyclophosphamide, and ionizing radiation. RESULTS: ABT-888 is a potent inhibitor of both PARP-1 and PARP-2 with K(i)s of 5.2 and 2.9 nmol/L, respectively. The compound has good oral bioavailability and crosses the blood-brain barrier. ABT-888 strongly potentiated temozolomide in the B16F10 s.c. murine melanoma model. PARP inhibition dramatically increased the efficacy of temozolomide at ABT-888 doses as low as 3.1 mg/kg/d and a maximal efficacy achieved at 25 mg/kg/d. In the 9L orthotopic rat glioma model, temozolomide alone exhibited minimal efficacy, whereas ABT-888, when combined with temozolomide, significantly slowed tumor progression. In the MX-1 breast xenograft model (BRCA1 deletion and BRCA2 mutation), ABT-888 potentiated cisplatin, carboplatin, and cyclophosphamide, causing regression of established tumors, whereas with comparable doses of cytotoxic agents alone, only modest tumor inhibition was exhibited. Finally, ABT-888 potentiated radiation (2 Gy/d x 10) in an HCT-116 colon carcinoma model. In each model, ABT-888 did not display single-agent activity. CONCLUSIONS: ABT-888 is a potent inhibitor of PARP, has good oral bioavailability, can cross the blood-brain barrier, and potentiates temozolomide, platinums, cyclophosphamide, and radiation in syngeneic and xenograft tumor models. This broad spectrum of chemopotentiation and radiopotentiation makes this compound an attractive candidate for clinical evaluation.

Organic anion transporting polypeptide 1B1 activity classified by SLCO1B1 genotype influences atrasentan pharmacokinetics.

OBJECTIVE: Our objective was to learn whether genetic polymorphisms of metabolic enzymes or transport proteins provide a mechanistic understanding of the in vivo disposition of atrasentan, a selective endothelin A receptor antagonist. METHODS: Atrasentan uptake was measured in HeLa cells transfected to express major alleles of organic anion transporting polypeptide 1B1 (OATP1B1). The results were used to classify individuals as extensive, intermediate, or poor OATP1B1 transporters according to their SLCO1B1 genotypes. Analysis of covariance including genotype, study, age, weight, sex, and ethnicity was used to identify factors influencing atrasentan single-dose (n = 44) and steady-state (n = 38) pharmacokinetic parameters. Genotypes for cytochrome P450 3A5, uridine diphosphate-glucuronosyltransferase (UGT) 1A1, UGT2B4, UGT2B15, adenosine triphosphate-binding cassette subfamily B (ABCB) 1, solute carrier organic anion transporter (SLCO) 1B1, and solute carrier family 22 (SLC22) A2 were each assessed. RESULTS: Single-dose atrasentan exposure (P = .0244), steady-state atrasentan exposure (P = .0108), and maximum postdose plasma concentration (P = .0002) were associated with OATP1B1 activity classified by SLCO1B1 genotype. No other tested genotypes were observed to be associated with both single-dose and steady-state atrasentan pharmacokinetics. CONCLUSIONS: OATP1B1 is a meaningful factor for atrasentan disposition. Individuals may be classified as having extensive, intermediate, or poor OATP1B1 transport phenotypes according to SLCO1B1 genotypes. Increased exposures of OATP1B1 substrates might be expected in individuals who have the poor transporter phenotype or are treated with an OATP1B1 inhibitor.

Phase 1 study of ABT-751, a novel microtubule inhibitor, in patients with refractory hematologic malignancies.

PURPOSE: ABT-751 is an oral antimitotic agent that binds to the colchicine site on beta-tubulin. A phase 1 study was conducted to determine the maximum tolerated dose and toxicities of ABT-751 in patients with advanced myelodysplastic syndrome and relapsed or refractory acute leukemias. STUDY DESIGN: Thirty-two patients were treated: nine with 100 (n = 3), 125 (n = 3), or 150 mg/m(2) (n = 3) of ABT-751 given orally once daily for 7 days every 3 weeks and 23 with 75 (n = 3), 100 (n = 3), 125 (n = 5), 150 (n = 5), 175 (n = 3), or 200 mg/m(2) (n = 4) of ABT-751 given orally once daily for 21 days every 4 weeks. Consenting patients had pharmacogenetic sampling and enumeration of circulating endothelial cells (CEC). RESULTS: Dose-limiting toxicity consisted of ileus in one patient at 200 mg/m(2), with a subsequent patient developing grade 2 constipation at the same dose level. One patient with relapsed acute myelogenous leukemia achieved a complete remission that was sustained for 2 months. Four other patients had transient hematologic improvements, consisting of a decrease in peripheral blood blasts and improvements in platelet counts. CEC number was reduced in three patients with a concomitant reduction in peripheral blasts. A previously undescribed nonsynonymous single nucleotide polymorphism, encoding Ala(185)Thr, was identified in exon 4 of the beta-tubulin gene, TUBB, in three other patients. The recommended phase 2 dose in hematologic malignancies is 175 mg/m(2) daily orally for 21 days every 4 weeks. CONCLUSION: Further assessment of ABT-751, especially in combination with other agents, in patients with acute leukemias is warranted.

Impact of CYP2D6 intermediate metabolizer alleles on single-dose desipramine pharmacokinetics.

This study utilized cytochrome P450 2D6 (CYP2D6) genotypes to explain variability of desipramine pharmacokinetics in a cohort of non-poor metabolizer individuals. In an interaction study utilizing desipramine as a probe, genotyping for the CYP2D6*3, *4, *5 and *6 alleles was used to screen out CYP2D6 poor metabolizers. Individuals were categorized according to these and additional alleles (CYP2D6*2, *9, *10, *17, *41 and x2). Genotypes of individuals heterozygous for two or three of *2, *17 and *41 alleles were confirmed by molecular haplotyping. Pharmacokinetic parameters of desipramine were analysed according to CYP2D6 category. Molecular haplotyping was necessary to definitively categorize four of 16 individuals. A subject who had unusually high plasma elimination half-time, exposure and metabolic ratios carried an intermediate metabolizer (IM) *9 allele in combination with a non-functional allele. This combination has a population frequency of less than 1 : 200. Individuals with *1/*1, *1/*2 and *2/*2 genotypes had lower than average plasma elimination half-time, exposure and metabolic ratios. For desipramine, additional genotyping of CYP2D6 IM alleles helped define subgroups of the CYP2D6-positive cohort. This suggests that genotyping for IM alleles will aid in interpretation of clinical trials involving CYP2D6 substrates. Due to the diversity of IM alleles, molecular haplotyping may be necessary to fully characterize CYP2D6 genotype-phenotype relationships.