Enzyme replacement with PEGylated cystathionine β-synthase ameliorates homocystinuria in murine model
Erez M. Bublil, … , Viktor Kožich, Jan P. Kraus
Erez M. Bublil, … , Viktor Kožich, Jan P. Kraus
Published May 16, 2016
Citation Information: J Clin Invest. 2016;126(6):2372-2384. https://doi.org/10.1172/JCI85396.
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Research Article Genetics Metabolism

Enzyme replacement with PEGylated cystathionine β-synthase ameliorates homocystinuria in murine model

Abstract

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.

Authors

Erez M. Bublil, Tomas Majtan, Insun Park, Richard S. Carrillo, Helena Hůlková, Jakub Krijt, Viktor Kožich, Jan P. Kraus

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Figure 1

Enzyme retention time in vivo is enhanced following htCBS PEGylation.

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Enzyme retention time in vivo is enhanced following htCBS PEGylation. (A...
(A) CBS activity in plasma taken from C57BL/6J mice that were injected with 5 mg/kg BW of htCBS via the i.p., i.v., or s.c. route. Two experimental arms (n = 5 each) were used for each injection route (due to an IACUC restriction on blood volume collected and time between different bleedings of the same mouse; the 1-hour time point was chosen as a shared point between all groups for comparison). Blood was collected after injection from the mice in group 1 at 0, 1, 8, and 24 hours and from the mice in group 2 at 1, 4, 10, and 48 hours. (B) The htCBS enzyme was incubated in WT or HO plasma at 37°C (160 ng/μl), and activity was measured at the indicated time points. (C) Coomassie-stained SDS-PAGE of the PEGylated and non-PEGylated htCBS with the corresponding specific activity values. (D) Mice (n = 4–5) were injected s.c., as described in A, with ME020MA- or GL4-400MA-PEGylated htCBS as compared with the non-PEGylated htCBS. (E) Plasma samples from mice injected s.c. with non-PEGylated htCBS or GL4-400MA PEGylated htCBS were analyzed by Western blotting using an anti-hCBS Ab. Data in A and D are presented as the mean ± SEM and were compared using ANOVA, followed by Tukey’s post-hoc test. *P ≤ 0.05, **P ≤ 0.01, and ***P ≤ 0.001. PEG, PEGylated; No PEG, non-PEGylated.