A Potent, Metabolically Stable Tubulin Inhibitor Targets the Colchicine Binding Site and Overcomes Taxane Resistance.

Antimitotics that target tubulin are among the most useful chemotherapeutic drugs, but their clinical activity is often limited by the development of multidrug resistance. We recently discovered the novel small-molecule DJ101 as a potent and metabolically stable tubulin inhibitor that can circumvent the drug efflux pumps responsible for multidrug resistance of existing tubulin inhibitors. In this study, we determined the mechanism of action of this drug. The basis for its activity was illuminated by solving the crystal structure of DJ101 in complex with tubulin at a resolution of 2.8Å. Investigations of the potency of DJ101 in a panel of human metastatic melanoma cell lines harboring major clinically relevant mutations defined IC50 values of 7-10 nmol/L. In cells, DJ101 disrupted microtubule networks, suppressed anchorage-dependent melanoma colony formation, and impaired cancer cell migration. In melanoma-bearing mice, DJ101 administration inhibited tumor growth and reduced lung metastasis in the absence of observable toxicity. DJ101 also completely inhibited tumor growth in a paclitaxel-resistant xenograft mouse model of human prostate cancer (PC-3/TxR), where paclitaxel was minimally effective. Our findings offer preclinical proof of concept for the continued development of DJ101 as a next-generation tubulin inhibitor for cancer therapy.Significance: These findings offer preclinical proof of concept for the continued development of DJ101 as a next-generation antitubulin drug for cancer therapy. Cancer Res; 78(1); 265-77. ©2017 AACR.

Maternal alcohol exposure during mid-pregnancy dilates fetal cerebral arteries via endocannabinoid receptors.

Prenatal alcohol exposure often results in fetal alcohol syndrome and fetal alcohol spectrum disorders. Mechanisms of fetal brain damage by alcohol remain unclear. We used baboons (Papio spp.) to study alcohol-driven changes in the fetal cerebral artery endocannabinoid system. Pregnant baboons were subjected to binge alcohol exposure via gastric infusion three times during a period equivalent to the second trimester of human pregnancy. A control group was infused with orange-flavored drink that was isocaloric to the alcohol-containing solution. Cesarean sections were performed at a time equivalent to the end of the second trimester of human pregnancy. Fetal cerebral arteries were harvested and subjected to in vitro pressurization followed by pharmacological profiling. During each alcohol-infusion episode, maternal blood alcohol concentrations (BAC) reached 80 mg/dL, that is, equivalent to the BAC considered legal intoxication in humans. Circulating anandamide (AEA) and 2-arachidonoylglycerol (2-AG) remained unchanged. Ultrasound studies on pregnant mothers revealed that fetal alcohol exposure decreased peak systolic blood velocity in middle cerebral arteries when compared to pre-alcohol levels. Moreover, ethanol-induced dilation was observed in fetal cerebral arteries pressurized in vitro. This dilation was abolished by the mixture of AM251 and AM630, which block cannabinoid receptors 1 and 2, respectively. In the presence of AM251, the cannabinoid receptor agonist AEA evoked a higher, concentration-dependent dilation of cerebral arteries from alcohol-exposed fetuses. The difference in AEA-induced cerebral artery dilation vanished in the presence of AM630. CB1 and CB2 receptor mRNA and protein levels were similar in cerebral arteries from alcohol-exposed and control-exposed fetuses. In summary, alcohol exposure dilates fetal cerebral arteries via endocannabinoid receptors and results in an increased function of CB2.

Effectiveness of Rack Sanitation Procedures for Elimination of Bacteria from Automatic Watering Manifolds.

An important responsibility of animal care programs is to protect research animals from exposure to potentially pathogenic microorganisms. To validate the need for steam sterilization of rodent automatic watering racks, we evaluate the post-sanitation microbial contamination of experimentally inoculated racks and of racks that had been used to house conventional mice. We tested three sanitation protocols: rack-washer sanitation without manifold flush, sanitation that included manifold flush, and sanitation that included manifold flush followed by autoclaving. Rack sanitation, with or without manifold flush, did not reliably eliminate microbial flora from the lixits or manifold drainage water. A total of 43% of all non-autoclaved racks were positive for bacterial contamination after sanitation, and racks that had been used for conventional animal housing were more frequently positive than were experimentally inoculated racks (79% vs 18%). These data indicate that steam sterilization is necessary for eliminating bacteria from automatic watering systems. These observations are particularly important in light of increasing numbers of immune-impaired rodents that may be inadvertently and unnecessarily exposed to microbial pathogens via the automatic watering system.