Enzyme replacement therapy ameliorates multiple symptoms of murine homocystinuria
Tomas Majtan,Wendell Jones Jr., Jakub Krijt, Insun Park, Warren D. Kruger, Viktor Kožich, Steven Bassnett, Erez M. Bublil, Jan P. Kraus
Classical homocystinuria (HCU) is the most common inherited disorder of sulfur amino acid metabolism caused by deficiency in cystathionine beta-synthase (CBS) activity and characterized by severe elevation of homocysteine in blood and tissues. Treatment with dietary methionine restriction is not optimal and poor compliance leads to serious complications. We developed an enzyme replacement therapy (ERT) and studied its efficacy in a severe form of HCU in mouse (the I278T model). Treatment was initiated before or after the onset of clinical symptoms in an effort to prevent or reverse the phenotype. ERT substantially reduced and sustained plasma homocysteine concentration at around 100 μM and normalized plasma cysteine for up to 9 months of treatment. Biochemical balance was also restored in the liver, kidney and brain. Furthermore, ERT corrected liver glucose and lipid metabolism. The treatment prevented or reversed facial alopecia, fragile and lean phenotype and low bone mass. 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 rescued the ocular phenotype. In conclusion, ERT maintained an improved metabolic pattern and ameliorated many of the clinical complications in the I278T mouse model of HCU
Majtan T, Park I, Bublil EM, Kraus JP
Skeletal and connective tissue defects are the most striking symptoms in patients suffering from classical homocystinuria (HCU). Here, we determined body composition and bone mass in three mouse models of HCU and assessed whether a long-term administration of enzyme replacement therapy (ERT) corrected the phenotype. The mouse models of HCU were analyzed using dual-energy X-ray absorptiometry and the data were complemented by plasma biochemical profiles. Both the mouse model lacking CBS (KO) and the one expressing human CBS mutant transgene on a mouse CBS null background (I278T) showed marked bone loss and decreased weight mostly due to a lower fat content compared to negative controls. In contrast, the HO mouse expressing the human CBS WT transgene on a mouse CBS null background showed no such phenotype despite similar plasma biochemical profile to the KO and I278T mice. More importantly, administration of ERT rescued bone mass and changes in body composition in the KO mice treated since birth and reversed bone loss and improved fat content in the I278T mice injected after the development of clinical symptoms. Our study suggests that ERT for HCU may represent an effective way of preventing the skeletal problems in patients without a restricted dietary regime. This article is protected by copyright. All rights reserved.
08|16|2017 Enzyme replacement prevents neonatal death, liver damage, and osteoporosis in murine homocystinuria.
Majtan T, Hůlková H, Park I, Krijt J, Kožich V, Bublil EM, Kraus JP.
Classical homocystinuria (HCU) is an inborn error of sulfur amino acid metabolism caused by deficient activity of cystathionine β-synthase (CBS), resulting in an accumulation of homocysteine and a concomitant decrease of cystathionine and cysteine in blood and tissues. In mice, the complete lack of CBS is neonatal lethal. In this study, newborn CBS knockout (KO) mice were treated with recombinant polyethyleneglycolylated human truncated CBS (PEG-CBS). Full survival of the treated KO mice, along with a positive impact on metabolite levels in plasma, liver, brain, and kidneys, was observed. The PEG-CBS treatment prevented an otherwise fatal liver disease characterized by steatosis, death of hepatocytes, and ultrastructural abnormalities of endoplasmic reticulum and mitochondria. Furthermore, treatment of the KO mice for 5 mo maintained the plasma metabolite balance and completely prevented osteoporosis and changes in body composition that characterize both the KO model and human patients. These findings argue that early treatment of patients with HCU with PEG-CBS may prevent clinical symptoms of the disease possibly without the need of dietary protein restriction.-Majtan, T., Hůlková, H., Park, I., Krijt, J., Kožich, V., Bublil, E. M., Kraus, J. P. Enzyme replacement prevents neonatal death, liver damage, and osteoporosis in murine homocystinuria.
06|12|2017 Engineering and Characterization of an Enzyme Replacement Therapy for Classical Homocystinuria.
Homocystinuria due to loss of cystathionine beta-synthase (CBS) causes accumulation of homocysteine and depletion of cysteine. Current treatments are suboptimal, and thus the development of an enzyme replacement therapy based on PEGylated human truncated CBS (PEG-CBS) has been initiated. Attenuation of potency was observed, which necessitated a screen of several PEG-CBS conjugates for their efficacy to correct and maintain the plasma metabolite profile of murine homocystinuria after repeated administrations interrupted with washouts. We found that CBS coupling with maleimide PEG inconsistently modified the enzyme. In contrast, the PEG-CBS conjugate with 20 kDa N-hydroxysuccinimide-PEG showed very little loss of potency likely due to a reproducible PEGylation resulting in species modified with five PEGs per subunit on average. We developed assays suitable for monitoring the extent of CBS PEGylation and demonstrated a sustainable partial normalization of homocystinuria upon continuous PEG-CBS administration via osmotic pumps. Taken together, we identified the PEG-CBS conjugate suitable for manufacturing and clinical development.
06|01|2016 Enzyme replacement with PEGylated cystathionine β-synthase ameliorates homocystinuria in murine model.
Homocystinuria, which typically results from cystathionine β-synthase (CBS) deficiency, is the most common defect of sulfur amino acid metabolism. CBS condenses homocysteine and serine to cystathionine that is then converted to cysteine. Individuals with homocystinuria have markedly elevated plasma levels of homocysteine and methionine and reduced concentrations of cystathionine and cysteine. Clinical disease manifestations include thromboembolism and neuropsychiatric, ocular, and skeletal complications. Here, we have shown that administration of PEGylated CBS into the circulation of homocystinuria model mice alters the extra- and intracellular equilibrium of sulfur amino acids, resulting in a decrease of approximately 75% in plasma total homocysteine (tHcy) and normalization of cysteine concentrations. Moreover, the decrease in homocysteine and the normalization of cysteine in PEGylated CBS-treated model mice were accompanied by improvement of histopathological liver symptoms and increased survival. Together, these data suggest that CBS enzyme replacement therapy (ERT) is a promising approach for the treatment of homocystinuria and that ERT for metabolic diseases may not necessitate introduction of the deficient enzyme into its natural intracellular compartment.