Objectives: The aim of the current study is to investigate the influence of mechanical stress as well as cleaning agents on the performance of various polyether ether ketone (PEEK) inserts for implant-retained overdentures (IOD). Materials and Methods: Three different standard PEEK inserts were subjected to rapid artificial aging through storage in chemical denture cleaning agents (acetic acid, sodium hypochlorite, or sparkling denture cleaner) as well as demineralized water. The pre-aged PEEK inserts were then placed in unilateral IOD and subjected to 200,000 chewing loads (5 kg similar to 50 N), with 5000 thermal cycles (5/55 degrees C), and 1100 removal/insertion cycles (vertical movement 2 mm). Results: For all the PEEK inserts, the retention forces decreased significantly with an increasing number of mechanical load cycles and after exposure to all the cleaning agents. PEEK inserts aged by exposure to chemical cleaning agents showed a significantly higher decline in retention force than the inserts stored in water. Confocal laser scanning microscopy indicated that the decline in retention force might be caused by wear on the internal insert surface in contact with the patrix. Conclusions: Within the limitations of this study, it can be concluded that the application of chemical cleaning agents accelerates the decline in the retention forces of PEEK retentive inserts in IODs.