Molecular Therpary and Mechanism Study in Spinal Cord Injury

Valproate is a multi-faceted drug due to its diverse actions, exhaustively tested clinically, and which may have new potential as a therapeutic in some other central nervous system disorders ranging from schizophrenia to neurodegeneration. This is largely mediated by its more recently discovered activity as a broad acting, histone deacetylase (HDAC) inhibitor modifying chromatin structure and neuronal gene expression. Indeed, the chronic antiepileptic effects of valproate may be due in part to HDAC-mediated regulation of the GABA synthetic enzyme, glutamate decarboxylase (10). We speculate that the development of more selective HDAC inhibitors could form a new class of therapeutics in neurodegenerative and spinal cord disorders, and we will investigate the potential application and mechanism of valproate in spinal cord injury (SCI). The dynamics of chromatin provides the major regulatory factor underlying gene expression and the major epigenetic modifications of chromatin involve the reversible acetylation and methylation of core histones on key lysine residues and DNA methylation at CpG islands. Both these modifications have been implicated in a range of neurodegenerative and spinal cord disorders. The reversible acetylation of histone lysine residues by histone acetyltransferases (HAT) is hence a key regulator of gene expression with acetylation leading to an open chromatin structure facilitating RNA polymerase binding and transcription. HDACs have multiple functions, in particular they are major components of repressive complexes and act to compact the chromatin and repress transcription. This has promoted an intensive search for inhibitors of HDACs as drugs to modulate gene transcription through their effects on the chromatin modifications. Thus, in this proposal, we intend to use our novel patented “indole” compound to conjugated with hydroxamic acid in histone deacetylases inhibitor, VA and further modify its structure to generate novel small-molecule neuroprotective compounds. By screening cortical neuron cultures, we will look for compounds that have shown potential for future drug development. In our preliminary studies, we have identified novel compounds, MPT0E014 and MPT0E028, having marked neuroprotective activity. We will test the lead compounds using in vivo rat SCI model for its efficacy and to perform the animal organ distribution studies. Using this approach, we hope to develop novel compounds with HDAC inhibition that are able to be used in future SCI therapy. To delineate the nature and mechanism of VA, MPT0E014 and MPT0E028 in SCI, we will evaluate effects of intraperitoneal VA, MPT0E014 and MPT0E028 injection on rats after spinal cord hemisection in vivo. Furthermore, we will evaluate the effects of VA, MPT0E014 and MPT0E028 on primary culture rat cortical neurons upon excitotoxic insult in vitro. The specific aims were to answer the following questions: (1) Do alterations in motor function occur after VA, MPT0E014 or MPT0E028 treatment? (2) How about the changes of neuron survival by VA, MPT0E014 or MPT0E028 on cord hemisection? (3) Could VA, MPT0E014 or MPT0E028 inhibit glial scar and microglia activation after SCI? (4) Do p21, p53, cleavage of caspase-3, Bcl-2, brain-derived neurotrophic factor (BDNF) and apoptosis change in VA, MPT0E014 or MPT0E028 treated group compared with vehecle controls? The overall objective of this study is to determine the neuroprotective, neurotrophic and anti-inflammatory effects of VA, MPT0E014 and MPT0E028 in SCI