Engineering stem cells to make mucopolysaccharidosis IIIB

DESCRIPTION provided by applicant Sanfilippo disease type B also called mucopolysaccharidosis type III or MPS III is a devastating neuro degenerative genetic disorder of childhood that is fatal There is no cure or effective treatment MPS IIIB is caused by the lack of a lysosomal enzyme called NAGLU alpha N acetylglucosaminidase that is required to degrade heparan sulfate glycosaminoglycans GAG Bone marrow transplantation is not effective in treating MPS III because most cells normally synthesize and secrete NAGLU that has mannose phosphate M P sugars attached via post translational modification and M P is essential to achieve adequate cellular uptake of the secreted enzyme via the M P receptor Unfortunately human macrophages the principal therapeutic cells in a bone marrow transplant secrete NAGLU that lacks M P Encouraging preliminary data from the Dickson lab shows that recombinant human NAGLU fused to a small peptide fragment that consists of the receptor binding motif of the insulin like growth factor IGF is able to enter cells using an alternativ uptake pathway the IGF receptor and has kinetic pH and functional characteristics similar to recombinant human NAGLU without the IGF fusion peptide We therefore propose to bioengineer stem cells to secrete NAGLU fused to an IGF peptide fragment that will be secreted and taken up by cells missing NAGLU thus correcting the cause of the disease when these modified stem cells are delivered clinically via injection Feasibility criteria for this phae I STTR will determine whether our approach merits further commercial development for a new therapy to treat MPS IIIB and are defined below in our two proposed AIMS Aim Macrophage secretion of NAGLU IGF Sanfilippo B syndrome is caused by deficiency of the enzyme alpha N acetylglucosaminidase NAGLU NAGLU lacks M P when produced from cells and therefore represents a perfect test case for the IGF technology We will use a lentiviral vector encoding murine Naglu IGF or Naglu and a CMV promoter to overexpress the transgene in murine macrophages We will deem this approach feasible for further development if macrophages are successfully transduced Naglu IGF is secreted in adequate quantities and secreted Naglu IGF is taken up by co cultured Naglu deficient cells via M P receptors traffics to lysosomes and degrades heparan sulfate NAGLUandapos s substrate Aim Modifying murine stem cells to express Naglu IGF This controlled in vivo study in Sanfilippo B mice will test whether IGF can restore uptake of NAGLU from a bone marrow transplant We will perform autologous stem cell transplantation using GFP labeled murine stem cells harvested from bone marrow modified with a lentiviral vector bearing the Naglu IGF coding sequence Feasibility criteria for this aim will be deemed to have been attained if evidence of significant disease correction is observed Our objective is to produce an effective treatment for MPS IIIB as rapidly and efficiently as possible If successful we will immediately proceed with a Phase II project that will propose IND enabling studies PUBLIC HEALTH RELEVANCE We will develop a new treatment for a fatal neurodegenerative disorder of childhood called Sanfilippo B or alternatively mucopolysaccharidosis IIIB or MPS IIIB Sanfilippo B caused by lack of alpha N acetylglucosaminidase NAGLU but delivery of recombinant synthesized NAGLU will not correct the disease because when NAGLU is synthesized in cell culture specific types of sugars that are required for NAGLU to function properly are not attached to the protein To solve this roadblock and create an effective treatment for this catastrophic disease we propose to modify stem cells to express NAGLU fused to the receptor binding motif of the insulin like growth factor NAGLU IGF because our preliminary data shows that this enzyme which we have already synthesized and begun testing can be taken up by cells via an alternative pathway the IGF receptor pathway traffics appropriately to the lysosomes has biochemical and functional properties similar to normal NAGLU and hence could reverse the terrible pathology caused by this disease