10PL8 and 2fPL8 inhibit gastric cancer cell proliferation by targeting degradation of CDK6 proteins to arrest the cell cycle.

Existing studies on PROTAC technology frequently rely on modifications and validation of already marketed drugs, which limits the exploration of non-drug-forming proteins and targets that do not have existing therapeutic agents. In contrast, utilizing easily synthesized combinatorial chemical databases that offer a broad selection of compounds presents a promising strategy for discovering new PROTAC molecules. In this study, virtual screening was conducted on a small molecule database created using combinatorial chemistry, which contains a total of 12,904 active molecules. A potential CDK6 inhibitor, compound 10, was identified and subsequently validated for its activity. The primary aim of the study was to explore its anti-tumor mechanism and explore its potential for PROTAC modification. The results showed that compound 10 effectively targeted CDK6, leading to the modification of its derivative 10PL8, which had a DC50 value of 26.67 ± 5.09 μM for targeting CDK6. Further structural modifications led to the synthesis of additional derivatives and PROTAC molecules, ultimately identifying a more potent CDK6 inhibitor, compound 2f, with an IC50 of 0.56 ± 0.06 μM in AGS cells. Moreover, the PROTAC molecule 2fPL8 demonstrated a DC50 value of 4.80 ± 0.79 μM for CDK6. The experimental findings above suggest that employing virtual screening of target proteins through combinatorial chemical databases to identify potential ligands and degradation agents is a feasible and valuable strategy for future PROTAC studies.