| Autor: |
Yingxia Wen, Megan Leong, Avishek Nandi, Andrea Carfi, Claudio Ciferri, Viola Fühner, Irmgard Hofmann, Andrea Gerber, Rachel Gerrein, Holger Laux, Anders Lilja, Axel Schulze |
| Rok vydání: |
2015 |
| Předmět: |
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| Zdroj: |
Biotechnology and Bioengineering. 112:2505-2515 |
| ISSN: |
0006-3592 |
| Popis: |
Human cytomegalovirus (HCMV) causes significant disease worldwide. Multiple HCMV vaccines have been tested in man but only partial protection has been achieved. The HCMV gH/gL/UL128/UL130/UL131A complex (Pentamer) is the main target of neutralizing antibodies in HCMV seropositive individuals and raises high titers of neutralizing antibodies in small animals and non-human primates (NHP). Thus, Pentamer is a promising candidate for an effective HCMV vaccine. Development of a Pentamer-based subunit vaccine requires expression of high amounts of a functional and stable complex. We describe here the development of a mammalian expression system for large scale Pentamer production. Several approaches comprising three different CHO-originated cell lines and multiple vector as well as selection strategies were tested. Stable cell pools expressed the HCMV Pentamer at a titer of approximately 60 mg/L at laboratory scale. A FACS-based single cell sorting approach allowed selection of a highly expressing clone producing Pentamer at the level of approximately 400 mg/L in a laboratory scale fed-batch culture. Expression in a 50 L bioreactor led to the production of HCMV Pentamer at comparable titers indicating the feasibility of further scale-up for manufacturing at commercial scale. The CHO-produced HCMV Pentamer bound to a panel of human neutralizing antibodies and raised potently neutralizing immune response in mice. Thus, we have generated an expression system for the large scale production of functional HCMV Pentamer at high titers suitable for future subunit vaccine production. Biotechnol. Bioeng. 2015;112: 2505–2515. © 2015 Wiley Periodicals, Inc. |
| Databáze: |
OpenAIRE |
| Externí odkaz: |
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