Kinetics of pramlintide degradation in aqueous solution as a function of temperature and pH

Abstract

The stability of the 37− amino acid peptide pramlintide, in aqueous solution, was studied as a function of pH and temperature. Samples of pramlintide formulated as a parenteral product were exposed to elevated temperatures and to realistic storage conditions for as long as 30 months. Pramlintide degradation was monitored by three high-performance liquid chromatography (HPLC) methods: a reversedphase (RP-HPLC) and a strong-cation exchange (SCX-HPLC) method for percentage purity determination by area normalization, plus a second RP-HPLC method for potency determination versus external standards. The pH-rate profile for pramlintide shows increasing degradation rate constants with increasing pH over the range pH = 3.5 to 5.0. The Arrhenius expression for pramlintide degradation at pH = 4.0 over the temperature range 5°C to 50° C is ln(k 0)= 37.39-21.900/RT, where k 0 is the zero-order rate constant (in %/mo) for pramlintide degradation. The pramlintide parenteral product formulated at pH = 4.0 is extremely stable, with percentage purity and percentage potency loss of only approximately 2% over 30 months at 5°C. The formulated pramlintide drug product has acceptable shelf life for long-term storage at 5°C and up to a 30-day patient use when stored at ambient temperature.

Development of Cagrilintide, a Long-Acting Amylin Analogue

“…Since the activation of both receptors might be advantageous for maximal efficacy on food intake and body weight, we prioritized initially non-selectivity. We found that formulation at pH 7 caused chemical instability including deamidation of asparagine residues, as reported for pramlintide, , and this could not be solved by formulations or minor pH adjustments. In order not to deviate too much from h-amylin, formulation at low pH seemed necessary, and we initiated a third branch of the program focused on the chemical stability at neutral pH to secure the option of co-formulation with other peptides.…”

“…An initial concern was that attachment of a lipophilic fatty acid would increase intermolecular attractive forces, − causing an even higher propensity to form amyloid fibrils. Additionally, published data on pramlintide , suggested a need for improvement of chemical stability, primarily targeting deamidation of asparagine residues to enable formulation at neutral pH.…”

Pramlintide injection drug product robustness studies

“…Previous studies identified 5° C as a suitable long-term storage condition for pramlintide injection and 25° C as an appropriate storage condition during a 1-month patient-use period [ 10]. The drug product vials, however, may experience temperature excursions (both above and below the recommended storage conditions) during patient use, or product shipping and warehousing.…”

“…The RP-HPLC method for determining pramlintide potency and m-cresol concentration has also been previously described [10]. Visual inspection of the vials was conducted per procedures in USP 23-NF 18 to detect the presence of any extraneous matter such as cores or fragments from the rubber stoppers.…”

“…The formulation contains pramlintide at strengths of 0.30 to 1.0 mg/mL and m-cresol as an antimicrobial preservative. Previous investigations have(1) identified pramlintide hydrolysis products [8]; (2) demonstrated the performance of stabilityindicating reversed-phase high performance liquid chromatography (RP-HPLC) and strong cation exchange (SCX-HPLC) analytical test methods [ 9]; and (3) established the stability of the drug product under realistic and stress conditions [10].…”