STABILIZERS AGAINST HEAT-INDUCED AGGREGATION OF RPR-114849, AN ACIDICFIBROBLAST-GROWTH-FACTOR (AFGF)

In an effort to optimize stabilization conditions for RPR 114849, a wi de variety of known stabilizers were screened for their effects on the stability of the protein against thermal denaturation. For the screen ing purpose, the effects of excipients on aggregation sate were examin ed employing UV spectrophotometric turbidity measurements at 50 degree s C and pH 7.4. The protein is sensitive to aggregation near its isoel ectric point. Higher concentrations of the protein promote faster aggr egation. Reducing agents do not decrease the aggregation rate indicati ng that oxidation of thiol groups to intermolecular disulfide bonding is not a rate-limiting factor in the aggregation process. In addition to well-known heparin, a wide variety of sulfated and phosphorylated a nionic polymers have shown to be powerful stabilizers for the protein. The chain length of a polymeric anion is a critical factor in stabili zing the protein aggregation. The stabilizing effect approaches a cons tant value asymptotically as the chain length increases. The combined action of enoxaparin and sodium citrate is additive indicating that th e stabilizers act independently and do not affect each other's mode, d egree, or efficacy of action. High concentrations of non-specific stab ilizers, such as sugars and polyols, are capable of suppressing aggreg ation of the protein to a minor extent. Surfactants, gelling and micro encapsulating agents were found to have no practical utility in stabil izing the protein. (C) 1998 Elsevier Science B.V. All rights reserved.