Tyrosyl-DNA phosphodiesterase 1 (TDP1) is an enzyme that plays a crucial role in repairing DNA lesions caused by the entrapment of DNA topoisomerase IB (TOP1)-DNA break-associated crosslinks. TDP1 inhibitors exhibit synergistic effects with TOP1 inhibitors in cancer cells, effectively overcoming resistance to TOP1 inhibitors. Therefore, this approach presents a promising strategy for reversing tumor resistance to TOP1 inhibitors. This review comprehensively outlines the structural and biological features of TDP1, the substrates involved in its catalytic hydrolysis, and its potential as a therapeutic target in oncology. Additionally, we summarize the various screening methods used to identify TDP1 inhibitors, alongside the latest advancements in TDP1 inhibitor research.