Development of a platform process for the production and purification of single-domain antibodies.

Autor:	Crowell LE; Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.; Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA., Goodwine C; Department of Chemical and Biological Engineering, Rensselaer Polytechnic Institute, Troy, New York, USA.; Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York, USA., Holt CS; Instituto de Virología, Investigación en Ciencias Veterinarias y Agronómicas, Instituto Nacional de Tecnología Agropecuaria, Buenos Aires, Argentina., Rocha L; Instituto de Virología, Investigación en Ciencias Veterinarias y Agronómicas, Instituto Nacional de Tecnología Agropecuaria, Buenos Aires, Argentina., Vega C; Instituto de Virología, Investigación en Ciencias Veterinarias y Agronómicas, Instituto Nacional de Tecnología Agropecuaria, Buenos Aires, Argentina., Rodriguez SA; Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA., Dalvie NC; Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.; Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA., Tracey MK; Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA., Puntel M; Instituto de Virología, Investigación en Ciencias Veterinarias y Agronómicas, Instituto Nacional de Tecnología Agropecuaria, Buenos Aires, Argentina., Wigdorovitz A; Instituto de Virología, Investigación en Ciencias Veterinarias y Agronómicas, Instituto Nacional de Tecnología Agropecuaria, Buenos Aires, Argentina., Parreño V; Instituto de Virología, Investigación en Ciencias Veterinarias y Agronómicas, Instituto Nacional de Tecnología Agropecuaria, Buenos Aires, Argentina., Love KR; Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA., Cramer SM; Department of Chemical and Biological Engineering, Rensselaer Polytechnic Institute, Troy, New York, USA.; Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York, USA., Love JC; Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.; Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
Jazyk:	angličtina
Zdroj:	Biotechnology and bioengineering [Biotechnol Bioeng] 2021 Sep; Vol. 118 (9), pp. 3348-3358. Date of Electronic Publication: 2021 Mar 25.
DOI:	10.1002/bit.27724
Abstrakt:	Single-domain antibodies (sdAbs) offer the affinity and therapeutic value of conventional antibodies, with increased stability and solubility. Unlike conventional antibodies, however, sdAbs do not benefit from a platform manufacturing process. While successful production of a variety of sdAbs has been shown in numerous hosts, purification methods are often molecule specific or require affinity tags, which generally cannot be used in clinical manufacturing due to regulatory concerns. Here, we have developed a broadly applicable production and purification process for sdAbs in Komagataella phaffii (Pichia pastoris) and demonstrated the production of eight different sdAbs at a quality appropriate for nonclinical studies. We developed a two-step, integrated purification process without the use of affinity resins and showed that modification of a single process parameter, pH of the bridging buffer, was required for the successful purification of a variety of sdAbs. Further, we determined that this parameter can be predicted based only on the biophysical characteristics of the target molecule. Using these methods, we produced nonclinical quality sdAbs as few as 5 weeks after identifying the product sequence. Nonclinical studies of three different sdAbs showed that molecules produced using our platform process conferred protection against viral shedding of rotavirus or H1N1 influenza and were equivalent to similar molecules produced in Escherichia coli and purified using affinity tags. (© 2021 The Authors. Biotechnology and Bioengineering published by Wiley Periodicals LLC.)
Databáze:	MEDLINE
Externí odkaz:	Zobrazit plný text záznamu Plný text