Synthesis, characterization, X-ray crystallography, and cytotoxicity of a cymantrene keto carboxylic acid for IR labelling of bioactive peptides on a solid support

Abstract

Cym–CO–CH2–CH2–COOH was prepared in good yield by Friedel–Crafts reaction of cymantrene (Cym, CpMn(CO)3) with succinic anhydride for the IR labelling of peptides and fully characterized, including an X-ray structure analysis (monoclinic space group P2(1)/n, a=5.727(3)Å, b=19.865(9)Å, c=10.518(5)Å, β=91.211(9)°). The compound was isolated in pure form without the need for chromatographic work-up and subsequently used for solution-phase synthesis of a bioconjugate with phenylalanine methyl ester to allow a complete spectroscopic characterization of this model system. The cymantrene keto carboxylic acid also turned out to be a very robust marker in automated microwave-assisted solid phase peptide synthesis (SPPS). [Leu5]-enkephalin (Tyr-Gly-Gly-Phe-Leu) was prepared on a Wang resin and labelled with the cymantrene derivative on the solid support under microwave irradiation in all steps. The metal–carbonyl marker stayed intact during cleavage from the resin with concentrated trifluoroacetic acid. After simple precipitation and lyophilization, the cymantrene–enkephalin bioconjugate could be obtained in analytically pure form without the need of HPLC purification. As required, the compound is non-cytotoxic against MCF-7 cells at up to 100μM. This protocol thus allows one to introduce organometallic IR spectroscopic labels to peptides in a very straightforward way.