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(Referência obtida automaticamente do Web of Science, por meio da informação sobre o financiamento pela FAPESP e o número do processo correspondente, incluída na publicação pelos autores.)

Approaches for recombinant human factor IX production in serum-free suspension cultures

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Autor(es):
Ferraz do Amaral, Robson Luis [1, 2] ; Bomfim, Aline de Sousa [1] ; de Abreu-Neto, Mario Soares [2] ; Picanco-Castro, Virginia [2] ; de Sousa Russo, Elisa Maria [1] ; Covas, Dimas Tadeu [2, 3] ; Swiech, Kamilla [1]
Número total de Autores: 7
Afiliação do(s) autor(es):
[1] Univ Sao Paulo, Sch Pharmaceut Sci Ribeiro Preto, Cafe Ave W-N, BR-14040903 Ribeirao Preto, SP - Brazil
[2] Reg Blood Ctr Ribeirao Preto, Tenente Catao Roxo St 2501, BR-14051140 Ribeirao Preto, SP - Brazil
[3] Univ Sao Paulo, Ribeirao Preto Med Sch, Bandeirantes Ave 3900, BR-14049900 Ribeirao Preto, SP - Brazil
Número total de Afiliações: 3
Tipo de documento: Artigo Científico
Fonte: Biotechnology Letters; v. 38, n. 3, p. 385-394, MAR 2016.
Citações Web of Science: 4
Resumo

Objective To establish a serum-free suspension process for production of recombinant human factor IX (rhFIX) based on the human cell line HEK 293T by evaluating two approaches: (1) serum-free suspension adaptation of previously genetic modified cells (293T-FIX); and (2) genetic modification of cells already adapted to such conditions (293T/SF-FIX). Results After 10 months, 293T-FIX cells had become adapted to FreeStyle 293 serum-free medium (SFM) in Erlenmeyer flasks. After 48 and 72 h of culture, 2.1 A mu g rhFIX/ml and 3.3 A mu g rhFIX/ml were produced, respectively. However, no biological activity was detected. In the second approach, wild-type 293T cells were adapted to the same SFM (adaptation process took only 2 months) and then genetically modified for rhFIX production. After 48 h of culture, rhFIX reached 1.5 A mu g/ml with a biological activity of 0.2 IU/ml, while after 72 h, the production was 2.4 A mu g/ml with a biological activity of 0.3 IU/ml. Conclusion The findings demonstrate that the best approach to establish an rhFIX production process in suspension SFM involves the genetic modification of cells already adapted to the final conditions. This approach is time saving and may better ensure the quality of the produced protein. (AU)

Processo FAPESP: 12/04629-8 - Estabelecimento de uma plataforma de produção de proteínas recombinantes terapêuticas em células humanas
Beneficiário:Kamilla Swiech Antonietto
Linha de fomento: Auxílio à Pesquisa - Apoio a Jovens Pesquisadores
Processo FAPESP: 11/10778-3 - Adaptação para cultivo em suspensão e meio de cultura livre de soro fetal bovino da linhagem celular 293-FIX-GFP para produção de FIX recombinante
Beneficiário:Robson Luis Ferraz do Amaral
Linha de fomento: Bolsas no Brasil - Iniciação Científica