Sustainable One‐Pot Synthesis of Novel Triazolopyrimidine Derivatives as Selective CDK4 Inhibitors for Breast Cancer Therapy

Abstract

A sustainable and effective one-pot synthesis of new 4,5-dihydro-[1,2,4]triazolo[1,5-a]pyrimidine-6-carbonitrile derivatives was accomplished using Mn₃O₄ nanoparticles as a recyclable nanocatalyst under moderate and environmentally benign conditions. The approach provided outstanding yields and adheres to sustainable chemical principles. Analysis revealed CDK4 as a possible therapeutic target linked to breast cancer growth, therefore elucidating its biological potential. In silico molecular docking validated the robust binding affinities of the synthesized derivatives to the CDK4 active site, especially compound 4, which demonstrated the greatest interaction energy and stable orientation inside the binding pocket. In vitro cytotoxic studies on MCF-7 and MDA-MB-231 breast cancer cell lines demonstrated that compound 4 had the highest antiproliferative activity (IC₅₀ = 5.80 and 6.44 µM, respectively), equivalent to doxorubicin and sorafenib. Compound 3 had significant effects, whereas compounds 9 and 10 showed modest efficacy. Notably, compound 4 exhibited little toxicity to WI-38 normal fibroblasts (IC₅₀ = 51.61 µM), suggesting selectivity for cancer cells. Flow cytometric analyses demonstrated that compound 4 produced G₀/G₁ cell-cycle arrest (82.77%) and initiated apoptosis (35.19%) with little necrosis, therefore validating its CDK4-dependent mechanism. These results together identify compound 4 as a powerful, selective, and environmentally friendly synthetic CDK4 inhibitor, serving as a viable candidate for the future development of tailored anti-breast cancer therapies.