Aptamer-directed crossing of BBB therapy of MPS 111B

描述（由申请人提供）：

DESCRIPTION (provided by applicant): MPS III B (Sanfilippo syndrome III B) is a devastating disease caused by mutation of the NAGLU gene and absence of the lysosomal enzyme alpha-N-acetylglucosaminidase. The clinical features include profound mental retardation, behavioral problems and death, usually in adolescence. There is no effective treatment. Enzyme replacement, a therapy becoming available for a growing number of lysosomal storage diseases, is not considered an option for the Sanfilippo syndrome because the blood brain barrier (BBB) prevents therapeutic enzyme from reaching the brain. However, essential proteins such as transferrin are normally transported across the BBB by receptor-mediated transcytosis through capillary endothelial cells. The goal of this application is to develop a novel strategy to ferry alpha-N-acetylglucosaminidase across the BBB in a mouse model of MPS III B. This strategy will make use of aptamers that bind to the transferrin receptor. Aptamers are single stranded nucleic acids that can be selected from large randomized libraries to bind to any desired target; in this respect, they resemble monoclonal antibodies. Specific Aim 1 is to isolate and characterize RNA aptamers that bind to the extracellular domain of the mouse transferring receptor. The aptamers will be made relatively resistant to RNase degradation by incorporation of 2'fluoropydmidines, and their binding affinities to the transferrin receptor will be determined. Specific Aim 2 is to conjugate selected aptamers to proteins - first to eGFP as a model protein, then to recombinant human alpha N-acetylglucosaminidase. Specific Aim 3 is to test the protein aptamer conjugates for transferrin receptor-mediated endocytosis by a mouse cell line and by cultured neurons isolated from brain of MPS III B mice. Specific Aim 4 is to test aptamer-enzyme conjugates in vivo, in order to determine whether they can ferry alpha-N-acetylglucosaminidase into the brain parenchyma and whether the enzyme will be functional in neural cells. The alpha-N-acetylglucosaminidase-aptamer conjugates found useful in the endocytosis test of Aim 3 will be administered to MPS III B mice and the brains subjected to biochemical and morphological examination. Should this strategy show promising results, it could easily be adapted for enzyme delivery for other neurodegenerative lysosomal storage diseases, as well as for drug delivery in common diseases such as Alzheimer's and Parkinson's.