Modification of glucose and glutamine metabolism in hybridoma cells through metabolic engineering

The present work describes the genetic modification of a hybridoma cell lin e with the aim to change its metabolic behaviour, particularly reducing the amounts of ammonia and lactate produced by the cells. The cellular excreti on of ammonia was eliminated by transfection of a cloned glutamine syntheta se gene. The metabolic characterisation of the transformed cell line includ es the analysis of the changes introduced in its intracellular metabolic fl uxes by means of a stoichiometric model. Furthermore, the reduction of lact ate accumulation was attempted through an antisense mRNA approach, aiming t o generate a rate limiting step in the glycolytic pathway, thus lowering th e glucose consumption rate. The physiological results obtained with the tra nsformed cells are discussed. A maximum reduction of about 47% in the gluco se consumption rate was obtained for one of the transformations. However a main drawback was the lack of stability of the transformed cells.