Microneedles (MNs) are a novel transdermal drug delivery technique that penetrate the skin stratum corneum and deliver drugs to the target site, which are safer and more effective than injection and oral preparation. The materials used for preparing microneedles are directly related to drug delivery, release, and therapeutic efficacy. Currently, polymer materials are widely used in microneedles because of their excellent mechanical properties, biocompatibility, and degradability. However, in the field of therapeutic and diagnostic, there are issues like excessive mechanical properties and uncontrollable drug release, leading to poor treatment effects or a possible risk of drug overdose. Whereas, stimulus-responsive polymer microneedles (SR-pMNs), an emerging on-demand drug release technology relays on physiological changes in the body or stimulation of the external environment, can realize continuous controlled release of microneedles, evenly and effectively encapsulating the active medication, and increasing the drug loading dose. This review sketches out the materials and methods of preparation of SR-pMNs, and their basic types and mechanisms of action. Besides, we emphasize the applications of SR-pMNs in the biomedical field, such as in drug delivery, diagnosis, and treatment of diseases. Furthermore, summarized the patents and clinical trials in the field of extractive diagnostics. Finally, we discussed the challenges and prospects for its clinical translation. We anticipate that SR-pMNs will open up new avenues for the treatment of human diseases by acting as transdermal medication delivery systems.