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Journal of Biomedicine and Biotechnology
Volume 2012 (2012), Article ID 875383, 10 pages
http://dx.doi.org/10.1155/2012/875383
Research Article

Generation of a Chinese Hamster Ovary Cell Line Producing Recombinant Human Glucocerebrosidase

1Centro de Biotecnologia, Instituto Butantan, Avenue Vital Brasil, 1500, 05503-900 São Paulo, SP, Brazil
2Interunidades em Biotecnologia, Instituto de Ciências Biomédicas, Universidade de São Paulo, 05508-000 São Paulo, SP, Brazil
3Centro de Biotecnologia, Instituto de Pesquisas Energéticas e Nucleares, 055508-000 São Paulo, SP, Brazil
4Laboratório de Biofármacos em Células Animais, Instituto Butantan, Avenue Vital Brasil, 1500, 05503-900 São Paulo, SP, Brazil
5Laboratório Especial de Toxinologia Aplicada, CAT-cepid, Instituto Butantan, Avenue Vital Brasil, 1500, 05503-900 São Paulo, SP, Brazil

Received 9 March 2012; Revised 30 June 2012; Accepted 19 July 2012

Academic Editor: D. M. Clarke

Copyright © 2012 Juliana Branco Novo et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Impaired activity of the lysosomal enzyme glucocerebrosidase (GCR) results in the inherited metabolic disorder known as Gaucher disease. Current treatment consists of enzyme replacement therapy by administration of exogenous GCR. Although effective, it is exceptionally expensive, and patients worldwide have a limited access to this medicine. In Brazil, the public healthcare system provides the drug free of charge for all Gaucher’s patients, which reaches the order of $ 84 million per year. However, the production of GCR by public institutions in Brazil would reduce significantly the therapy costs. Here, we describe a robust protocol for the generation of a cell line producing recombinant human GCR. The protein was expressed in CHO-DXB11 (dhfr) cells after stable transfection and gene amplification with methotrexate. As expected, glycosylated GCR was detected by immunoblotting assay both as cell-associated (~64 and 59 kDa) and secreted (63–69 kDa) form. Analysis of subclones allowed the selection of stable CHO cells producing a secreted functional enzyme, with a calculated productivity of 5.14 pg/cell/day for the highest producer. Although being laborious, traditional methods of screening high-producing recombinant cells may represent a valuable alternative to generate expensive biopharmaceuticals in countries with limited resources.