John Maga

Unravelling the kinetoplastid paraflagellar rod

Researchers who study human pathogens are often interested in unique and essential aspects of the biology of the pathogen. Recent progress has been made in understanding such a target in kinetoplastid parasites. The paraflagellar rod is a unique cytoskeletal structure that plays a key role in the life-cycle of these fascinating organisms. This review discusses the protein components and structure of the paraflagellar rod and its function in cell motility.

Glycosylation-independent Lysosomal Targeting of Acid α-Glucosidase Enhances Muscle Glycogen Clearance in Pompe Mice

Background: Acid α-glucosidase, an enzyme replacement therapy for Pompe disease, is poorly targeted to lysosomes when relying on phosphomannose residues. Results: Fusing IGF-II to acid α-glucosidase resulted in more efficient uptake and glycogen clearance from muscle of Pompe mice. Conclusion: Enhanced binding to the cation-independent mannose 6-phosphate receptor (CI-MPR) enabled improved glycogen clearance in Pompe mice. Significance: BMN 701 is now being tested for Pompe disease in human clinical studies. We have used a peptide-based targeting system to improve lysosomal delivery of acid α-glucosidase (GAA), the enzyme deficient in patients with Pompe disease. Human GAA was fused to the glycosylation-independent lysosomal targeting (GILT) tag, which contains a portion of insulin-like growth factor II, to create an active, chimeric enzyme with high affinity for the cation-independent mannose 6-phosphate receptor. GILT-tagged GAA was taken up by L6 myoblasts about 25-fold more efficiently than was recombinant human GAA (rhGAA). Once delivered to the lysosome, the mature form of GILT-tagged GAA was indistinguishable from rhGAA and persisted with a half-life indistinguishable from rhGAA. GILT-tagged GAA was significantly more effective than rhGAA in clearing glycogen from numerous skeletal muscle tissues in the Pompe mouse model. The GILT-tagged GAA enzyme may provide an improved enzyme replacement therapy for Pompe disease patients.

BMN 701-mediated receptor redistribution is responsible for increased uptake

BMN 701 is a novel chimeric fusion protein of Insulin-like Growth Factor 2 (IGF-2) and acid alpha-glucosidase (GAA) that is now being tested in the clinic for the treatment of Pompe disease. BMN 701 was designed to improve delivery of GAA to the lysosome of muscle cells by fusing a high affinity ligand for the cation independent mannose-6phosphate receptor (CI-MPR) to GAA. In uptake experiments with rat L6 myoblasts, the Kuptake improvedmarkedly compared to untagged rhGAA, a predicted consequence of higher affinity binding to the CI-MPR. Additionally, the total amount of BMN 701 taken up by cells (system capacity) was doubled compared to rhGAA. This surprising observation could be explained by a phenomenon reported previously, namely that IGF-1 and IGF-2 can induce a signaling cascade resulting in redistribution of the CI-MPR so as to increase the quantity of the CI-MPR on the cell surface. We have investigated the ability of BMN 701 to do the same. In vitro, we find: i) BMN 701 can bind to the IGF-1 receptor; ii) BMN 701 can transduce a signaling cascade from the IGF-1 receptor; iii) the elevated system capacity of BMN701 is dependent on this signaling from the IGF-1 receptor; and iv) system capacity for uptake of untagged rhGAA can be increased by IGF-1. These results are consistent with the higher system capacity resulting from tag-dependent signaling. This novel attribute of BMN 701 may contribute to its enhanced ability to reverse glycogen storage in Pompe mice.