Use of pyrazolo[1,5a]pyrimidin-7-yl amine derivatives in the treatment of neurological disorders
A pyrazolo and barrier technology, applied to regulate the activity of Eph receptors in cells, stimulate and improve nerve regeneration, and can solve problems such as glial scar reduction, axonal damage, and hyperplasia
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[0326] In the following preferred embodiments, general expressions may be replaced by corresponding more specific definitions provided in the context to yield more preferred embodiments of the invention.
[0327] Preferred is the use of a compound of formula (I), a tautomer or a pharmaceutically acceptable salt thereof, wherein the Eph receptor-associated (e.g. neurological) injuries and disorders to be treated are dependent on ephrin receptors Neurological disorders or impairments of kinases (eg, EphA4 kinase).
[0328] The present invention particularly relates to the use of the compound of formula (I) or a pharmaceutically acceptable salt thereof and the use of the compound of formula (I) in the treatment of kinase-dependent diseases or for the preparation of pharmaceutical preparations for the treatment of kinase-dependent diseases,
[0329]
[0330] in:
[0331] R 2 is H, substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, substituted or unsub...
Embodiment 1
[0375] Example 1: EphA4 mode and mechanism of action
[0376] To distinguish between forward and bidirectional signaling that ephrins are capable of in the context of axon regeneration, lentiviral expression vectors for wild-type and kinase-dead EphA4 were generated and overexpressed in purified astrocytes. Cortical layer neurons form a layer over two astrocyte populations, and axonal outgrowths are analyzed and compared. Biological peptides that have been shown to block the interaction of EphA4 with related ligands and subsequently inhibit receptor activation (Murai, K.K. et al., (2003) Mol Cell Neurosci24(4):p.1000), were tested in star EphA4 inhibitory activity in morphocyte / cortical neuron culture system. Identification of neuronal ligand / ephrin-mediated inhibition of EphA4 by systematically blocking expression of candidate ephrins in neurons using RNA interference to knock down ephrins or using dominant negative ephrin constructs and A coat then forms on wild-type astro...
Embodiment 2
[0378] Example 2: Autophosphorylation and Ligand-Dependent Phosphorylation Assays
[0379] Primary astrocyte cultures were constructed from neonatal mouse cortex and purified to obtain approximately 95-98 pure astrocyte cultures. To detect autophosphorylation, cells were incubated in the presence or absence of pharmacological inhibitors and directly thereafter lysed and subjected to immunoprecipitation and Western blot analysis (eg, Figure 1A shown). For ligand-dependent phosphorylation (such as Figure 1B indicated), the cultures were then serum starved for 36 hours to induce basal receptor phosphorylation, and then treated with a soluble form of the cognate ligand in the presence of a candidate kinase inhibitor or blocking peptide (which was added at various concentrations) in combination with In the absence, stimuli were given for different lengths of time. Cells were lysed and lysates were subjected to EphA4 immunoprecipitation followed by Western blotting to analyze re...
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