af-353 and Disease-Models--Animal

When Zika virus emerged as a public health emergency there were no drugs or vaccines approved for its prevention or treatment. We used a high-throughput screen for Zika virus protease inhibitors to identify several inhibitors of Zika virus infection. We expressed the NS2B-NS3 Zika virus protease and conducted a biochemical screen for small-molecule inhibitors. A quantitative structure-activity relationship model was employed to virtually screen ∼138,000 compounds, which increased the identification of active compounds, while decreasing screening time and resources. Candidate inhibitors were validated in several viral infection assays. Small molecules with favorable clinical profiles, especially the five-lipoxygenase-activating protein inhibitor, MK-591, inhibited the Zika virus protease and infection in neural stem cells. Members of the tetracycline family of antibiotics were more potent inhibitors of Zika virus infection than the protease, suggesting they may have multiple mechanisms of action. The most potent tetracycline, methacycline, reduced the amount of Zika virus present in the brain and the severity of Zika virus-induced motor deficits in an immunocompetent mouse model. As Food and Drug Administration-approved drugs, the tetracyclines could be quickly translated to the clinic. The compounds identified through our screening paradigm have the potential to be used as prophylactics for patients traveling to endemic regions or for the treatment of the neurological complications of Zika virus infection.

Topics: Animals; Antiviral Agents; Artificial Intelligence; Chlorocebus aethiops; Disease Models, Animal; Drug Evaluation, Preclinical; High-Throughput Screening Assays; Immunocompetence; Inhibitory Concentration 50; Methacycline; Mice, Inbred C57BL; Protease Inhibitors; Quantitative Structure-Activity Relationship; Small Molecule Libraries; Vero Cells; Zika Virus; Zika Virus Infection

To evaluate the role that intravesical P2X2/3 purinergic receptors (P2X2/3Rs) play in early and advanced neurogenic lower urinary tract (LUT) dysfunction after contusion spinal cord injury (SCI) in female rats.. Female Sprague-Dawley rats received a thoracic Th8/Th9 spinal cord contusion with either force of 100 kDy (cN); moderate) or 150 kDy (cN; severe); Sham rats had no injury. Evaluations on urethane-anesthetised rats were conducted at either 2 or 4 weeks after SCI. LUT electrical signals and changes in bladder pressure were simultaneously recorded using cystometry and a set of custom-made flexible microelectrodes, before and after intravesical application of the P2X2/3R antagonist AF-353 (10 μM), to determine the contribution of P2X2/3R-mediated LUT modulation.. Severe SCI significantly increased bladder contraction frequency, and reduced both bladder pressure amplitude and intraluminal-pressure high-frequency oscillations (IPHFO). Intravesical P2X2/3R inhibition did not modify bladder pressure or IPHFO in the Sham and moderate-SCI rats, although did increase the intercontractile interval (ICI). At 2 weeks after SCI, the Sham and moderate-SCI rats had significant LUT electromyographic activity during voiding, with a noticeable reduction in LUT electrical signals seen at 4 weeks after SCI. Intravesical inhibition of P2X2/3R increased the ICI in the Sham and moderate-SCI rats at both time-points, but had no effect on rats with severe SCI. The external urethral sphincter (EUS) showed strong and P2X2/3R-independent electrical signals in the Sham and moderate-SCI rats in the early SCI stage. At 4 weeks after SCI, the responsiveness of the EUS was significantly attenuated, independently of SCI intensity.. This study shows that electrophysiological properties of the LUT are progressively impaired depending on SCI intensity and that intravesical P2X2/3R inhibition can attenuate electrical activity in the neurogenic LUT at early, but not at semi-chronic SCI. This translational study should be useful for planning clinical evaluations.

Topics: Administration, Intravesical; Animals; Disease Models, Animal; Electromyography; Female; Phenyl Ethers; Purinergic P2X Receptor Antagonists; Pyrimidines; Rats; Rats, Sprague-Dawley; Signal Transduction; Spinal Cord Injuries; Urethra; Urinary Bladder, Neurogenic; Urination