Liposomal Formulations for Efficient Delivery of a Novel, Highly Potent Pyrimidine-Based Anticancer Drug.

Background/Objectives: Cancer is one of the deadliest diseases worldwide. Despite the existing treatments, the adverse side effects and the increasing drug resistance to the current therapies lead to a reduced quality of life for patients and poor prognosis. The pyrimido[5,4-d]pyrimidine compound (PP) was identified as a promising new anticancer drug due to its potent activity against colorectal and triple-negative breast cancers; however it showed poor aqueous solubility and safety profile. This study aimed the synthesis of compound PP, its encapsulation in liposomal formulations based on phosphatidylcholines (PC), the characterization of liposomal formulations and its biological evaluation. Methods: A new synthesis method for PP was developed. The compound was incorporated into different liposomal formulations. The hydrodynamic size, polydispersity, and zeta potential of loaded and non-loaded formulations were measured by DLS. The cytotoxic effects of compound PP, placebo nanoformulations, and PP-loaded nanoformulations were assessed in colorectal (HCT 116) and triple-negative breast cancer (MDA-MB-231) cell lines, as well as in non-tumor BJ-5ta cells. Results: The PP compound was efficiently synthesized. The PP-loaded liposomal formulations exhibit sizes below 150 nm, low polydispersity, and long-time stability upon storage at 4 °C. The antitumor compound was encapsulated with excellent efficiency, and sustained release profiles were obtained. The PP compound showed high activity against HCT 116 (IC(50) = 2.04 ± 0.45 µM) and MDA-MB-231 (IC(50) = 5.24 ± 0.24 µM) cell lines. DPPC-containing formulations were effective against cancer cells, but showed toxicity comparable to free PP in BJ-5ta normal cells. Conversely, PP-EggPC-Chol-L formulation displayed strong anticancer activity with residual toxicity to normal cells. Conclusions: The PP-loaded liposomal formulation, composed of 70% PC from egg yolk (EggPC) and 30% cholesterol (Chol), designated as PP-EggPC-Chol-L, was the most promising formulation, showing effective anticancer activity in both cancer cell lines and a significant improvement in the safety profile which is of utmost importance to progress to the next phase of drug development.