A pH-responsive natural cyclopeptide RA-V drug formulation for improved breast cancer therapy.

The natural plant cyclopeptide RA-V, which was isolated from the roots of Rubia yunnanensis, was discovered to be a novel anti-cancer candidate. However, the cyclic hexapeptide exhibited poor solubility in physiological conditions, limiting its application for cancer therapy in vivo. To solve this problem, pH-sensitive polymers were developed for targeted RA-V delivery into tumor sites and for acid-triggered drug release. The poly(β-amino ester)s (PAE) copolymers self-assembled into micelle-like nanoparticles in an aqueous solution at pH 7.4, and the solubility of RA-V was enhanced by loading the drug into the hydrophobic cores of micelles. The near-infrared (NIR) fluorescent probe squaraine (SQ) dye, as an imaging probe, could also be encapsulated into polymer micelles simultaneously. The diameters of the RA-V/SQ loaded micelles were measured by dynamic light scattering (DLS) and transmission electron microscopy (TEM), which proved that the micelles with sizes of 35-60 nm were suitable as anti-cancer drug nano-vehicles. The drug-loading capacity and drug release profiles of RA-V-loaded micelles were calculated and monitored by high performance liquid chromatography (HPLC) measurements. The RA-V/SQ loaded micelles were stable at a neutral pH, and drug release could be greatly accelerated by the acid-triggered ionization of copolymer chains. Similarly, with free RA-V cyclopeptide, the RA-V/SQ loaded micelles exhibited high anti-cancer efficiency toward MCF-7 cells and Hela cells, while the intact polymer micelles and SQ-loaded micelles are non-toxic. Moreover, the endocytosis pathway and mitochondria-regulated apoptosis of RA-V/SQ loaded micelles were proved by lysosome colocalization and JC-1 assay, respectively. Finally, biodistribution and tumor growth inhibition were evaluated in MCF-7 cell-xenografted nude mice, demonstrating that RA-V/SQ loaded micelles could realize tumor imaging and effectively and simultaneously inhibit tumor growth. Therefore, the RA-V/SQ loaded micelles may find use as potential nano-scaled cancer therapeutics and imaging agents.