LC/MS Characterization of Impurities and Degradation Products of a Potent Antitumor Peptidic Dimer, CU201

Abstract

Compound CU201 [SUIM-(d-Arg-Arg-Pro-Hyp-Gly-Igl-Ser-d-Igl-Oic-Arg)₂, where SUIM = suberimidyl; Hyp = trans-4-hydroxyproline; Igl = α-(2-indanyl)-glycine; Oic = octahydroindole-2-carboxylic acid], is a dimeric analog of the potent bradykinin antagonist peptide B9430. It blocks the G_αq,11 signal of the heterotrimeric G proteins, stimulates c-Jun kinases, and induces apoptosis in lung cancer cells with neuroendocrine features. CU201 shows potent inhibition for small-cell lung cancer cells in vitro (ED₅₀ = 0.15 μM), as well as for small-cell lung cancer SHP-77 tumor growth in vivo. An HPLC method was developed, as part of a study supported by the National Cancer Institute’s (NCI’s) Rapid Access to Interventional Development (RAID) program, to assess the purity and stability of CU201. Impurities and degradation products were characterized by LC/MS. The identity of a major impurity, with 1 mass unit different from CU201, was confirmed by high resolution LC/MS and the investigation of model compounds. Susceptible linkages in the peptide chains were revealed by the degradation study.

Keywords: CU201, B201, LC–MS, Impurities characterization, Peptidic dimer, Bradykinin antagonist