Design, synthesis, and biological evaluation of third-generation anticancer tubulin inhibitors incorporating beta and unnatural amino acids

Abstract

Tubulysins are highly potent microtubule-depolymerizing tetrapeptides with remarkable anticancer activity; however, their complex structures, poor metabolic stability, and susceptibility to proteolytic degradation limit their therapeutic application. In this work, we report the design and synthesis of third-generation tubulin inhibitors that incorporate β-amino acids and unnatural amino acids to enhance protease resistance and improve overall peptide stability. The central fragments of thiazole-containing amino acids were synthesized using solution-phase reaction. Reductive amination was employed to access the N-terminal N-alkyl pipecolic acid derivatives. The novel tubulin inhibitors were synthesized using solid-phase peptide synthesis (SPPS) on glutamic acid and asparagine resins, followed by purification using RP-HPLC. The potency of the newly synthesized third-generation tubulin inhibitors was evaluated against the skin cancer cell line, A431, and human breast cancer cell line, MCF7, to establish detailed structure–activity relationships and identify a potential lead compound for anticancer drug development.