Synthesis of Diastereomerically Pure Cetrorelix Acetate by Using Fmoc Solid-Phase Peptide Synthesis (SPPS) Strategy:...

Area of Science:

• Medicinal Chemistry
• Organic Synthesis
• Analytical Chemistry

Background:

• Solid-phase peptide synthesis (SPPS) is a key technique for producing peptide drugs like cetrorelix.
• Controlling stereochemical purity, particularly minimizing racemization of amino acids, is critical for peptide drug efficacy and safety.
• The Fmoc (fluorenylmethyloxycarbonyl) strategy is widely used in SPPS but can be prone to racemization of certain amino acids.

Purpose of the Study:

• To investigate and minimize the racemization of L-arginine and L-serine during the Fmoc-based SPPS of cetrorelix.
• To develop and optimize a robust analytical method for separating closely related cetrorelix isomers, including D-amino acid impurities.
• To establish impurity control strategies essential for the commercial production and therapeutic application of cetrorelix.

Main Methods:

• Solid-phase peptide synthesis (SPPS) utilizing the Fmoc strategy.
• Employing specific coupling agents (HATU), additives (HOBt or HOAt), and bases (TMP) to mitigate racemization.
• Development of a gradient reversed-phase high-performance liquid chromatography (RP-HPLC) method for isomer separation.

Main Results:

• Racemization of L-arginine and L-serine was successfully reduced to below 0.5% during cetrorelix synthesis.
• The optimized RP-HPLC method effectively separated structurally similar cetrorelix isomers.
• D-arginine and D-serine impurities were identified as the most challenging to separate from the main cetrorelix peak.

Conclusions:

• The combination of HATU, HOBt/HOAt, and TMP effectively suppresses racemization in Fmoc-SPPS of cetrorelix.
• A sensitive RP-HPLC method is established for monitoring critical isomeric impurities.
• Strict control over D-amino acid impurities is paramount for the successful commercial manufacturing and therapeutic use of cetrorelix.