Design, Synthesis and Molecular Dynamics Simulation of Novel Spiropyran Derivatives as Putative CDK6-Targeting Anticancer Agents for Breast Cancer Treatment

Background

Breast cancer treatment with current dual CDK4/6 inhibitors is limited by hematological toxicity and acquired drug resistance. CDK6 is overexpressed in specific breast cancer subtypes and represents a validated molecular target for the development of new anticancer agents. This study aimed to design, synthesize, and evaluate novel spiropyran derivatives as putative CDK6-targeting anticancer agents for breast cancer treatment.

Materials and Methods

Eight spiropyran derivatives (S1-S8) were designed and evaluated through drug-likeness assessment, ADME/toxicity prediction using ForceADME and Geintox-II, and molecular docking against CDK6 (PDB ID: 5L2I) using GeinDock Suite v2.3.1.0. Selected complexes were subjected to 100 ns molecular dynamics simulations with MM-GBSA binding free energy calculations using Desmond/Prime. Compounds were synthesized via multi-step condensation reactions, characterized by IR, ¹H NMR, ¹³C NMR, and MS, and selected derivatives evaluated for in vitro cytotoxicity against MCF-7 breast cancer cells using MTT assay (n=3, triplicates).

Results

Among the eight derivatives, compound S5 showed the highest predicted CDK6 binding affinity (-9.8 kcal/mol), marginally higher than palbociclib (-9.4 kcal/mol), with dual hydrogen bonds to GLY20 and TYR24. Molecular dynamics simulations indicated a binding free energy for S5 (∆Gᵇᵉᵀ = -131.12±7.76 kcal/mol) comparable to palbociclib (-127.66±5.26 kcal/mol). In vitro cytotoxicity revealed an IC₅₀ of 15.58±1.24 μM for S5, comparable to palbociclib (17.33±1.38 μM) in MCF-7 cells.

Conclusion

The observed agreement between computational predictions and in vitro cytotoxicity for the three experimentally evaluated compounds supports the spiropyran scaffold as a candidate framework warranting further investigation. Spiropyran derivative S5 represents an early-stage lead with distinct CDK6 binding interactions, requiring confirmatory kinase inhibition assays, selectivity profiling, and in vivo efficacy studies before its therapeutic potential can be established.