The Effect of the Expression of the Antiapoptotic BHRF1 Gene on the Metabolic Behavior of a Hybridoma Cell Line

Abstract

One of the most important limitations of mammalian cells-based bioprocesses, and particularly hybridoma cell lines, is the accelerated metabolism related to glucose and glutamine consumption. The high uptake rates of glucose and glutamine (i.e., the main sources of carbon, nitrogen and energy) lead to the production and accumulation of large amounts of lactate and ammonia in culture broth. Lactate and/or ammonia accumulation, together with the depletion of the main nutrients, are the major causes of apoptosis in hybridoma cell cultures. The KB26.5 hybridoma cell line, producing an IgG3, was engineered with BHRF1 (KB26.5-BHRF1), an Epstein–Barr virus-encoded early protein homologous to the antiapoptotic protein Bcl-2, with the aim of protecting the hybridoma cell line from apoptosis. Surprisingly, besides achieving effective protection from apoptosis, the expression of BHRF1 modified the metabolism of the hybridoma cell line. Cell physiology and metabolism analyses of the original KB26.5 and KB26.5-BHRF1 revealed an increase of cell growth rate, a reduction of glucose and glutamine consumption, as well as a decrease in lactate secretion in KB26.5-BHRF1 cells. A flux balance analysis allowed us to quantify the intracellular fluxes of both cell lines. The main metabolic differences were identified in glucose consumption and, consequently, the production of lactate. The lactate production flux was reduced by 60%, since the need for NADH regeneration in the cytoplasm decreased due to a more than 50% reduction in glucose uptake. In general terms, the BHRF1 engineered cell line showed a more efficient metabolism, with an increase in biomass volumetric productivity under identical culture conditions.