A Synthetic Approach to Poly-γ-Glutamyl Analogs of Methotrexate

Summary

Methotrexate poly-γ-glutamates bearing two and three glutamate units above that present in methotrexate have been synthesized by extension of a previously described route by which the lower congener bearing one added glutamate unit was synthesized. Key steps in the sequence are the peptide coupling of N-[4-[[(benzyloxy)-carbonyl]methylamino]benzoyl]-L-glutamic acid α-benzyl ester (5) with oligo-γ-L-glutamate benzyl esters, removal of blocking groups by catalytic hydrogenolysis, and introduction of the (2,4-diamino-6-pteridinyl)methyl grouping by alkylation with (6-bromomethyl)-2,4-pteridinediamine hydrobromide. Elaboration of the required oligo-γ-L-glutamate chain was achieved one unit at a time beginning with the coupling of L-glutamic acid dibenyl ester with t-butyloxy-carbonyl-L-glutamic acid α-benzyl ester (7) followed by selective removal of the t-butyloxycarbonyl grouping and another coupling step with 5 or 7 as required. Diphenylphosphoryl azide was used as the coupling reagent in each conversion producing a peptide linkage. Intermediates were obtained in pure form according to thin-layer chromatography and elemental analysis results, and the final target compounds were obtained in high purity as judged by thin-layer chromatography, high-performance liquid chromatography, ¹H NMR and mass spectral data, and elemental analysis results.

This work was supported by the National Cancer Institute under Contract Number 263-MD-104307.