title: Mixed-lineage kinase inhibitors require the activation of Trk receptors to maintain long-term neuronal trophism and survival.
abstract: Small-molecule mixed-lineage kinase (MLK) inhibitors, such as CEP-1347 [3,9-bis[(ethylthio)methyl]-(8R*,9S*,11S*)-(-)-9-hydroxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetrahydro-8,11-epoxy-1H,8H, 11H-2,7b,11a-triazadibenzo(a,g)cycloocta(cde)trinden-1-one] and CEP-11004 [3,9-bis-[(isopropylthio)methyl]-(8R*,9S*,11S*)-(-)-9-hydroxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetrahydro-8,11-epoxy-1H,8H,11H-2,7b,11a-triazadibenzo(a,g)cycloocta(cde)trinden-1-one], prevent c-Jun NH(2)-terminal kinase (JNK) pathway activation as well as the consequent neuronal cell death in many cell culture and animal models. In the cell culture model of nerve growth factor (NGF)-deprived sympathetic neurons, we find that CEP-11004 induced a approximately 3-fold increase in the mRNA and protein levels of TrkA, the NGF receptor. This resulted in ligand-independent activation of the TrkA receptor and the downstream phosphatidylinositol 3-kinase (PI3-kinase) pathway. Addition of the Trk inhibitor K252a [(8R*,9S*,11S*)-(-)-9-hydroxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetrahydro-8,11-epoxy-1H,8H,11H-2,7b,11a-triazadibenzo(a,g)cycloocta(cde)-trinden-1-one] or the PI3-kinase inhibitor LY294002 [2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one] significantly decreased the protein synthesis rates, mitochondrial function, and neuronal survival maintained by CEP-11004. In contrast to sympathetic neurons, MLK inhibitors maintain only short-term survival of potassium- and serum-deprived rat cerebellar granule neurons (CGNs), despite continuous inhibition of the JNK pathway. We found that similar to sympathetic neurons, CEP-11004 increased the levels of the Trk receptor expressed in CGNs, TrkB. However, CGNs required the addition of the exogenous ligand brain-derived neurotrophic factor (BDNF) to activate the PI3-kinase pathway and to maintain long-term survival. BDNF activated TrkB, but caused rapid down-regulation of activated receptors and maintained only minimal survival. Therefore, increase in TrkB levels by CEP-11004 mediated a synergism with BDNF resulting in long-term survival in response to the combined treatment of CEP-11004 and BDNF. Taken together, our studies suggest that in addition to the direct inhibition of the JNK pathway, the indirect activation of the PI3-kinase pathway via Trk activation is important for MLK inhibitor-mediated neuronal survival and trophism.
authors.affiliation: Department of Neurology, Washington University School of Medicine, St. Louis, MO 63110, USA.
authors.forename: Leo H
authors.initials: LH
authors.lastname: Wang
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Paden
authors.forename: Eugene M
authors.initials: EM
authors.lastname: Johnson
mesh_terms: D000818:Animals; D019208:Brain-Derived Neurotrophic Factor / Q000494:pharmacology; D002227:Carbazoles / Q000494:pharmacology*; D002470:Cell Survival / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D007211:Indoles / Q000494:pharmacology*; D048031:JNK Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors; D020930:MAP Kinase Kinase Kinases / Q000037:antagonists & inhibitors*; D020932:Nerve Growth Factor / Q000494:pharmacology; D009474:Neurons / Q000166:cytology / Q000187:drug effects*; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D020917:Receptor, trkA / Q000235:genetics / Q000502:physiology*; D020813:Receptor, trkB / Q000502:physiology
chemical_list: D019208:Brain-Derived Neurotrophic Factor; C469298:CEP-11004; D002227:Carbazoles; D004791:Enzyme Inhibitors; D007211:Indoles; D012333:RNA, Messenger; C106592:3,9-bis((ethylthio)methyl)-K-252a; D020932:Nerve Growth Factor; D019869:Phosphatidylinositol 3-Kinases; D020917:Receptor, trkA; D020813:Receptor, trkB; D048031:JNK Mitogen-Activated Protein Kinases; D020930:MAP Kinase Kinase Kinases