Human cell lines: A promising alternative for recombinant FIX production

Autor(es)

Bomfim, Aline de Sousa ^{[1, 2, 3]} ; Correa de Freitas, Marcela Cristina ^{[2, 3, 4]} ; Picanco-Castro, Virginia ^{[2, 3]} ; Soares Neto, Mario de Abreu ^{[2, 3]} ; Swiech, Kamilla ^{[2, 3, 5]} ; Covas, Dimas Tadeu ^{[2, 3, 4]} ; de Sousa Russo, Elisa Maria ^{[1, 2, 3]}

Afiliação do(s) autor(es)

^[1] Univ Sao Paulo, Dept Clin Toxicol & Food Sci Anal, Fac Pharmaceut Sci Ribeirao Preto, 2501 Tenente Catao Roxo St, BR-14051140 Ribeirao Preto, SP - Brazil

^[2] Univ Sao Paulo, Ctr Cell Based Therapy, 2501 Tenente Catao Roxo St, BR-14051140 Ribeirao Preto, SP - Brazil

^[3] Univ Sao Paulo, Reg Blood Ctr, 2501 Tenente Catao Roxo St, BR-14051140 Ribeirao Preto, SP - Brazil

^[4] Univ Sao Paulo, Dept Clin Med, Fac Med Ribeirao Preto, 2501 Tenente Catao Roxo St, BR-14051140 Ribeirao Preto, SP - Brazil

^[5] Univ Sao Paulo, Dept Pharmaceut Sci, Fac Pharmaceut Sci Ribeirao Preto, 2501 Tenente Catao Roxo St, BR-14051140 Ribeirao Preto, SP - Brazil

• Tipo de documento: Artigo Científico
• Fonte: Protein Expression and Purification; v. 121, p. 149-156, MAY 2016.
• Citações Web of Science: 2

Resumo

Factor IX (FIX) is a vitamin K-dependent protein, and it has become a valuable pharmaceutical in the Hemophilia B treatment. We evaluated the potential of recombinant human FIX (rhFIX) expression in 293T and SK-Hep-1 human cell lines. SK-Hep-1-FIX cells produced higher levels of biologically active protein. The growth profile of 293T-FIX cells was not influenced by lentiviral integration number into the cellular genome. SK-Hep-1-FIX cells showed a significantly lower growth rate than SK-Hep-1 cells. gamma-carboxylation process is significant to FIX biological activity, thus we performed a expression analysis of genes involved in this process. The 293T gene expression suggests that this cell line could efficiently carboxylate FIX, however only 28% of the total secreted protein is active. SK-Hep-1 cells did not express high amounts of VKORC1 and carboxylase, but this cell line secreted large amounts of active protein. Enrichment of culture medium with Ca+2 and Mg+2 ions did not affect positively rhFIX expression in SK-Hep-1 cells. In 293T cells, the addition of 0.5 mM Ca+2 and 1 mM Mg+2 resulted in higher rhFIX concentration. SK-Hep-1 cell line proved to be very effective in rhFIX production, and it can be used as a novel biotechnological platform for the production of recombinant proteins. (C) 2016 Elsevier Inc. All rights reserved. (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