Development of microcapsules containing enzymes and genetically modified cells for the biochemical degradation of metabolites elevated in renal failure.

摘要

An alginate microcapsule was developed using a vibrational encapsulation process containing three enzymes (urease, uricase, and creatininase) capable of effectively degrading urea, uric acid, and creatinine, which are elevated to pathologic levels in patients with kidney failure. In addition, two strains of genetically modified bacteria expressing the enzymes urease and uricase were identified, combined and dispersed in 600 mum alginate microcapsules suitable for oral administration.; In 24 hour in-vitro experiments, 5 ml of capsules incorporating easily managed and readily identifiable quantities of enzymes effectively degraded all the uric acid, 97 per cent of the urea and 70 per cent of the creatinine from 100 ml of a challenge solution formulated to the concentration of these solutes in a presenting hemodialysis patient. Enzyme degradation of urea followed Michaelis-Menten kinetics. An identical quantity of capsules containing modified cells completely cleared 95% of the urea and >99% of the uric acid from a similar challenge solution. The process of urea degradation was found to be intracellular and each bacterial strain was specific for its substrate, both initially and after conditioning.; In in-vivo trials with Wistar rats, orally administered capsules were found to remain in the gastrointestinal (GI) tract from 12 to 24 hours. A chemically-induced model of acute renal failure was employed to evaluate the ability of orally delivered encapsulated enzymes and cells to degrade uremic toxins. Encapsulated enzyme and cell therapy decreased the severity of azotemia and hyperuricemia by as much as 70%. Reduction of urea concentration in-vivo required co-administration of a cation exchange resin to adsorb ammonia. Preliminary scale up calculations indicated that oral delivery to humans would involve a realistic and practical quantity of enzymes and cells.; Overall, a single delivery vehicle comprising a capsule containing enzymes, cells, or a combination of both was evaluated in-vitro and in-vivo in a rodent model for the degradation of multiple uremic toxins, and shown potential as a possible adjunctive therapy in the treatment of End Stage Renal Disease (ESRD).