pki-166 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

The epidermal growth factor (EGF) receptor and its ligands are crucially involved in the renal response to ischaemia. We studied the heparin binding-epidermal growth factor (HB-EGF), a major ligand for the EGF receptor, in experimental and human ischaemia/reperfusion injury (IRI). HB-EGF mRNA and protein expression was studied in rat kidneys and cultured human tubular (HK-2) cells that were subjected to IRI and in human donor kidneys during transplantation. The effect of EGF receptor inhibition was investigated in vivo and in vitro. Furthermore, urinary HB-EGF protein excretion was studied after renal transplantation. Finally, HB-EGF KO and WT mice were subjected to IRI to study the role of HB-EGF in renal injury. HB-EGF mRNA was significantly up-regulated in the early phase of IRI in rats, cells, and human donor biopsies. Treatment with PKI-166 reduces macrophage accumulation and interstitial alpha-SMA in the early phase of IRI in rats. In vitro, PKI-166 causes a marked reduction in HB-EGF-induced cellular proliferation. Urinary HB-EGF is increased after transplantation compared with control urines from healthy subjects. HB-EGF KO mice subjected to IRI revealed significantly less morphological damage after IRI, compared with WT mice. We conclude that IRI results in early induction of HB-EGF mRNA and protein in vivo and in vitro. Absence of HB-EGF and inhibition of the EGF receptor in the early phase of IRI has protective effects, suggesting a modulating role for HB-EGF.

Topics: Adult; Aged; Animals; Cells, Cultured; Disease Models, Animal; Epidermal Growth Factor; Female; Heparin-binding EGF-like Growth Factor; Humans; Intercellular Signaling Peptides and Proteins; Kidney; Kidney Transplantation; Male; Mice; Middle Aged; Pyrimidines; Pyrroles; Rats; Rats, Wistar; Reperfusion Injury; RNA, Messenger; Up-Regulation; Young Adult

The aim of this study was to evaluate, whether PET with (18)F-FDG and 3'-deoxy-3'-(18)F-fluorothymidine ((18)F-FLT) may be used to monitor noninvasively the antiproliferative effects of tyrosine kinase inhibitors.. Using a high-resolution small animal scanner, we measured the effect of the ErbB-selective kinase inhibitor PKI-166 on the (18)F-FDG and (18)F-FLT uptake of ErbB1-overexpressing A431 xenograft tumors.. Treatment with PKI-166 markedly lowered tumor (18)F-FLT uptake within 48 h of drug exposure; within 1 wk (18)F-FLT uptake decreased by 79%. (18)F-FLT uptake by the xenografts significantly correlated with the tumor proliferation index as determined by proliferating cell nuclear antigen staining (r = 0.71). Changes in (18)F-FLT uptake did not reflect inhibition of ErbB kinase activity itself but, rather, the effects of kinase inhibition on tumor cell proliferation. Tumor (18)F-FDG uptake generally paralleled the changes seen for (18)F-FLT. However, the baseline signal was significantly lower than that for (18)F-FLT.. These results indicate that (18)F-FLT PET provides noninvasive, quantitative, and repeatable measurements of tumor cell proliferation during treatment with ErbB kinase inhibitors and provide a rationale for the use this technology in clinical trials of kinase inhibitors.

Topics: Animals; Antineoplastic Agents; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Dideoxynucleosides; Disease Models, Animal; Fluorodeoxyglucose F18; Humans; Mice; Mice, SCID; Neoplasm Invasiveness; Positron-Emission Tomography; Protein Kinase Inhibitors; Pyrimidines; Pyrroles; Radiopharmaceuticals; Treatment Outcome

How do myelinated axons signal to the nuclei of cells that enwrap them? The cell bodies of oligodendrocytes and Schwann cells are segregated from axons by multiple layers of bimolecular lipid leaflet and myelin proteins. Conventional signal transduction strategies would seem inadequate to the challenge without special adaptations. Wallerian degeneration provides a model to study axon-to-Schwann cell signaling in the context of nerve injury. We show a hitherto undetected rapid, but transient, activation of the receptor tyrosine kinase erbB2 in myelinating Schwann cells after sciatic nerve axotomy. Deconvolving microscopy using phosphorylation state-specific antibodies shows that erbB2 activation emanates from within the microvilli of Schwann cells, in direct contact with the axons they enwrap. To define the functional role of this transient activation, we used a small molecule antagonist of erbB2 activation (PKI166). The response of myelinating Schwann cells to axotomy is inhibited by PKI166 in vivo. Using neuron/Schwann cell cocultures prepared in compartmentalized cell culture chambers, we show that even transient activation of erbB2 is sufficient to initiate Schwann cell demyelination and that the initiating functions of erbB2 are localized to Schwann cells.

Topics: Analysis of Variance; Animals; Axotomy; Blotting, Western; Bromodeoxyuridine; Cell Proliferation; Cells, Cultured; Coculture Techniques; Demyelinating Diseases; Disease Models, Animal; Embryo, Mammalian; Female; Fluorescent Antibody Technique; Ganglia, Spinal; Gene Expression; Glycoproteins; Immunoprecipitation; Mitogen-Activated Protein Kinase Kinases; Myelin Basic Protein; Myelin Sheath; Neuregulins; Neuroglia; Neurons; Platelet-Derived Growth Factor; Pyrimidines; Pyrroles; Rats; Rats, Sprague-Dawley; Receptor Protein-Tyrosine Kinases; Receptor, ErbB-2; Schwann Cells; Sciatic Neuropathy; Signal Transduction; Sodium Channels; Time Factors; Wallerian Degeneration