Modeling Odontohypophosphatasia in the Sheep
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SUMMARY/Hypophosphatasia (HPP) is a rare inherited disorder that affects the development of bones and teeth thecause of which is mutations in the tissue-nonspecific alkaline phosphatase (TNSALP/TNAP) gene (ALPL). Inaddition, dental hard tissues have been hypothesized to be the most sensitive to HPP, as premature loss ofanterior teeth in toddlers is often the first sign of disease in mild cases of HPP, and some individuals fall withinthe subtype odonto-HPP, where dental defects are the only apparent manifestation. Although useful formodeling many important features of HPP, murine models harboring ALPL mutations do not faithfully representthe broad spectrum of human HPP clinical tooth and bone abnormalities. Therefore, in order to gatherimportant insights into the molecular and natural history of the profound tooth and bone phenotypes of HPP, itis essential to develop new models. In this light, the size and basic anatomy of the sheep skeleton andHaversian remodeling is comparable to humans, and sheep are born with baby teeth that are lost naturallybefore being replaced by permanent teeth. Recent advances in mammalian gene editing using theCRISPR/Cas9 system have made disease-specific point mutation knock-in accessible in a variety of species,including sheep. Since the sheep TNAP protein sequence is highly conserved compared to humans (89%identity), two distinct mutations were targeted in the human ALPL gene that have previously beencharacterized, [Alanine (GCC)ïƒ Threonine (ACC) in exon 5 (c.346 G>A) and Isoleucine (ATC)ïƒ Methionine(ATG) in exon 10 (c.1077 C>G). Capitalizing on the advances in gene editing, a sheep model of HPP has beengenerated, for the first time, that accurately recapitulates the tooth phenotype of the exon 10 (c.1077 C>G)human point mutation. In Aim 1, demonstration of the validity of the ALPL knock-in sheep as a model for dentalmanifestations of HPP in both deciduous and permanent teeth will be accomplished. In Aim 2, the efficacy of abone/tooth-targeted recombinant TNAP therapy to reverse the dental manifestations of HPP and improve themineralization for of all dental tissues will be determined. If successful, these studies will represent a newplatform for the accurate evaluation of the phenotype of HPP but also define the sheep as an amenable animalplatform to model other human bone and mineral disorders.
