Evaluation of hollow fiber bioreactors as an alternative to murine ascites production for small scale monoclonal antibody production

Abstract

Monoclonal antibody (MAb) production via the growth of ascitic tumors in mice has been the time-honored technique for producing small scale, research laboratory quantities of MAbs. There are many disadvantages associated with MAb production in murine ascites, including animal welfare concerns. Small scale, commercially available, hollow fiber bioreactor systems, which appear to have advantages over other in vitro cell culture techniques, have recently been introduced. To the author's knowledge, these bioreactor systems have not been independently evaluated as a potential alternative to the in vivo production of MAbs in murine ascites. The first objective of this study was to characterize the clinicopathologic changes in mice associated with the in vivo production of MAbs. Five different hybridoma cell lines were grown in groups of 20 mice. Mice were primed with 0.5 ml pristane intraperitoneally 14 days prior to inoculation of 1 x 106 hybridoma cells.Abdominal paracentesis was performed a maximum of 3 times for collection of ascites from each mouse; ascites volumes were recorded. Clinical observations, body weight measurements, and postmortem examinations were performed. Incidence and severity of clinical abnormalities increased with time. Disseminated intraabdominal seeding of tumor and/or solid tumor masses were observed at postmortem examination. The second objective of the study was to compare MAb production in mice versus hollow fiber bioreactor systems to determine the feasibility of these systems as potential alternatives to the use of mice. Three hybridoma cell lines were grown in each of three different commercially available hollow fiber bioreactor systems and in groups of 20 mice. Bioreactors were harvested 3 times weekly for 65 days and were monitored by cell counts, cell viability and media glucose consumption. Time and materials logs were maintained. Time spent in labor and materials costs were greater for the bioreactors than the mice. The mean antibody concentration, as determined by ELISAs, in mouse ascites versus the range of mean concentrations for the 3 bioreactor systems for each cell line were as follows: cell line 2B11, 4.22 mg/ml vs. 0.71 to 1.31 mg/ml; cell line 3C9, 4.07 mg/ml vs. 1023.06 mg. Significant clinicopathologic changes in mice were associated with MAb production in ascites. Although time and materials costs were greater for the bioreactors, these results suggest that hollow fiber bioreactor systems merit further investigation as potentially viable alternatives to the use of mice for MAb production.2 ACKNOWLEDGEMENTS

Industrial Implementation of in Vitro Production of Monoclonal Antibodies

“…Various in vitro MAb production systems that have been evaluated between 2000 and 2003 at GSK Bio, based on suspension and hollow fiber technologies (Jackson et al 1996), are described below. In suspension systems, the gas exchange occurs by simple diffusion, which may be enhanced by rotation of the culture flask.…”

Monoclonal antibody production: viability improvement of RC1 hybridoma cell in different types of bioreactor

“…The commonly used culture systems for the production of mAb by hybridoma cells are the Stirred Tank (ST) bioreactor and airlift fermenter (Ozturk and Palsson 1991;Guez et al 2004) and in some occasions the hollow-fibre and ceramic matrix module has been used (Butler 1996;Jackson et al 1996;Racher et al 1990;Heilmann et al 2005). Among those bioreactor systems, the ST bioreactor which is the traditionally used fermenter with the internal mechanical agitation is preferable in improving the mAb production because it is highly flexible and it can provide high volumetric mass-transfer coefficient (k L a) values for gas transfer.…”

Specific detection and quantitation of bovine IgG in bioreactor derived mouse mAb preparations

“…Apart from their recombinant forms which are generated in high producer CHO or other type of cells, most of the antibodies are produced using hybridomas, either in animals (in vivo) or in cell culture (in vitro) (Schirrmann et al 2008;Liu 2014). In vivo antibody manufacturing has been a widespread method of mAb production because of its reliability, high antibody yield and cost effectiveness (Jackson et al 1996;Peterson and Peavey 1998). However, because of ethical concerns and because antibodies have to meet strict quality control requirements (purity, similarity, identity) the number of animals used for antibody production for diagnostic and scientific purposes is continuously decreasing.…”