Translocator protein 18-kDa, broadly known as TSPO, is a mitochondrial membrane protein, previously identified as the peripheral benzodiazepine receptor (PBR). TSPO involves in a broad number of biochemical events, such as steroidogenesis, mitochondrial cholesterol transport, cell survival and death, cell proliferation, and carcinogenesis. Several investigations have reported the roles of TSPO in various types of cancers, including colorectal cancer, brain cancer, melanoma, breast cancer, prostate cancer, and lung cancer. It was found that TSPO is upregulated in cancer cells, and it appears that its expression is parallel with an aggressive phenotype and/or poor prognosis. As a consequence, there is great potential for developing diagnostic and prognostic tools targeting the TSPO. In this regard, several radioligands targeting the TSPO have been identified, and some of the candidates have advanced to clinical trials. In recent years, image-guided surgery using hybrid probes bearing radioactive and fluorescence molecules has demonstrated promising outcomes in animal and human studies, and thus might serve as a valuable surgical navigator during cancer surgery. In general, current hybrid probes are built from various molecular platforms, including small molecules, nanoparticles, and antibodies. Although several TSPO-targeted imaging probes have been developed, their development for image-guided surgery of cancers is scarce. This review highlights recent findings of the involvement of TSPO in carcinogenesis, and provides a new perspective on the potential application of TSPO-targeted hybrid probes for image-guided surgery.