INTERLEUKIN-1 RECEPTOR (IL-1R) LIQUID FORMULATION DEVELOPMENT USING DIFFERENTIAL SCANNING CALORIMETRY

Purpose. To elucidate the solution conditions that confer stability of aqueous IL-1R using differential scanning calorimetry (DSC). Methods, Optimal pH conditions were determined by monitoring degradation produ cts encountered during accelerated studies (at elevated temperatures) using SDS-PAGE. At the pH optimum, DSC screened for excipients that en hanced thermal stability by shifting the Tm to higher values. Using SE C the relationship between thermal unfolding and stability was investi gated by considering if lower Tm's in the presence of preservatives co rrelated with degradation products at 37 degrees C over time. The degr ee of aggregation relative to that of a control determined the level o f stability achieved. Results. Circular dichroism (CD) measurements co nfirmed molecular modeling studies showing IL-1R to be about 39% beta- sheet. Two major transitions characterized the DSC data with Tm's obse rved near 47 degrees C and 66 degrees C. Among 21 excipients screened, NaCl exhibited the greatest stabilizing influences based on shifting the low temperature transition to 53 degrees C. The low temperature tr ansition was later found to comprise two transitions, yielding a total of three melting transitions for IL-IR. High Tm's arising from the pr esence of preservatives correlated with the order of stability (i.e., 0.065% phenol > 0.1% m-Cresol > 0.9% benzyl alcohol). Conclusions, The three melting transitions are consistent in origin with the cooperati ve unfolding of three unique immunoglobulin-like domains of IL-1R. Opt imal stability was achieved in 20 mM sodium citrate at pH 6 with suffi cient NaCl to attain the tonicity of human serum. A correlation betwee n the predicted ranking of stability and the extent of aggregation was demonstrated using DSC.