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Master's Dissertation
DOI
10.11606/D.85.2011.tde-20062011-110202
Document
Author
Full name
Fernanda dos Santos Arthuso
E-mail
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2011
Supervisor
Committee
Soares, Carlos Roberto Jorge (President)
Bartolini, Paolo
Paraiba, Diane Belchior
Title in Portuguese
Adaptação de células CHO secretoras de prolactina humana e seus antagonistas para o crescimento em suspensão
Keywords in Portuguese
antagonistas
CHO
prolactina
suspensão
Abstract in Portuguese
O Grupo de Hormônios do Centro de Biotecnologia do IPEN desenvolveu várias linhagens de células de ovário de hamster chinês (CHO) modificadas geneticamente e comprovadamente eficientes na expressão de proteínas heterólogas, dentre elas a prolactina humana (hPRL) e os análogos antagonistas de prolactina (S179D-hPRL e G129R-hPRL). No entanto, todas as linhagens para expressão são cultivadas em monocamadas e dependentes da presença de soro fetal bovino (SFB) no meio de cultivo para um crescimento eficiente. As células em suspensão apresentam um grande interesse industrial-farmacêutico, tanto pela facilidade de cultivo e ampliação de escala, como pela produtividade volumétrica. Desenvolvemos um protocolo para adaptação de células CHO para o crescimento em suspensão e também processos de produção em frascos spinners. Nesse trabalho foi realizada a adaptação das linhagens produtoras de hPRL; S179D-hPRL e G129R-hPRL para o crescimento em suspensão e em meio livre de SFB. Realizamos também a produção em escala laboratorial com as três linhagens adaptadas, assim como a correspondente purificação e caracterização de quatro proteínas heterólogas, incluindo a prolactina humana glicosilada (G-hPRL).
Title in English
Adaptation of CHO cells secreting human prolactin and their antagonists to growth in suspension
Keywords in English
antagonists
CHO
prolactin
suspension
Abstract in English
The Hormone Group of the Biotechnology Center of IPEN has developed different cells lines of genetically modified chinese hamster ovary cells (CHO) for the expression of heterologus protein like human prolactin (hPRL) and its analogs/antagonists (S179D-hPRL and G129R-hPRL). All cell lines for expression are however cultured in monolayer culture dish and depend on fetal bovine serum (FBS) in the medium for an efficient growth. Cells in suspension show a great industrial-pharmaceutical interest, especially for the cultivation facility and scale enlargement as well as for volumetric productivity. We developed a protocol for adapting CHO cells to suspension growth, in spinner flasks. The adaption of our cell lines producing hPRL; S179D-hPRL and G129R-hPRL to suspension growth and in serum-free medium was obtained. We also carried out laboratory scale production with the three suspension-adapted culture line cells and the corresponding purification and characterization of four heterologous proteins, including glycosylated human prolactin (G-hPRL).
 
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Publishing Date
2011-06-21
 
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