Background on scientific achievements related to HCU at Kraus Laboratory:
Cystathionine beta-synthase (CBS)-deficient homocystinuria (HCU) is the most common inherited metabolic disease associated with sulfur amino acid metabolism. Biochemically, the disease is characterized by highly elevated levels of homocysteine, methionine and S-adenosylhomocysteine, and low levels of cysteine and cystathionine in blood. Connective tissue problems, vascular disorders, cognitive deficit and skeletal abnormalities are only few of the many manifestations of this disease.
For over 40 years, Dr. Kraus’s laboratory has been in the forefront of homocystinuria research. They were the first to purify the human enzyme to homogeneity, isolate pure CBS mRNA, clone the cDNA and isolate and sequence the entire gene. In addition, they were the first to map the enzyme to human chromosome 21, to express it in bacterial systems, to crystalize it, and to extensively characterize its biochemical properties. Crystallization of the enzyme in both the basal and the activated form provided them with a structural insight into the molecular mechanism of allosteric activation of human CBS by S-adenosylmethionine.
Dr. Kraus’s lab also developed a unique mouse model of homocystinuria and they built and maintain the largest database of CBS homocystinuric alleles (i.e. the specific genetic defects that cause HCU). They have capitalized on this work to develop an enzyme replacement therapy (ERT) destined to enter clinical trials in 2018 (a collaboration between the University of Colorado, Denver and Orphan Technologies Ltd.).
Enzyme replacement therapy is well known for its successful application to lysosomal storage diseases (LSD) in which the administered enzyme is glycosylated to enable uptake to the lysosome itself. This natural uptake to the site of need, unfortunately, cannot be exploited for other metabolic diseases. A question may be raised whether administration of a replacement enzyme into the circulation, rather than to the intracellular environment, may favorably affect intra-cellular metabolism of sulfur amino acids? Dr. Kraus’s lab show for the first time, that administration of a truncated, mutated PEGylated form of the human CBS enzyme to homocystinuric mice greatly and positively affects cysteine, homocysteine and cystathionine metabolism and thus is currently considered for ERT.
Dr. Kraus’s lab is expressing the truncated mutant human CBS in bacteria and have perfected a four column purification process to obtain a highly pure protein that was injected via different routes but failed to exhibit any efficacy due to its fast clearance and loss of activity. They have then devised a PEGylation strategy based on analyses of 10 different PEG molecules conjugated via a variety of chemistries to the enzyme, and selected a lead candidate that showed a powerful positive impact on the equilibrium of sulfur amino acids in circulation and in tissues. The treatment prevented a neonatal lethal phenotype in a knock out (KO) mouse model of the disease and/or reversed facial alopecia, fragile and lean phenotype, liver disease and low bone mass in KO and I278T mouse models. In addition, structurally defective ciliary zonules in the eyes of I278T mice contained low density and/or broken fibers, while administration of ERT from birth partially normalized the ocular phenotype. In conclusion, ERT maintained an improved metabolic pattern and ameliorated many of the clinical complications in three different mouse models of HCU.
Database on CBS and genetic defects maintained by Jan Kraus, along with Viktor Kozich and Miroslav Janosik from Charles University in Prague: http://cbs.lf1.cuni.cz/cbsdata/cbsmain.htm