Abstract: Uncoated ectopic tertiary lymphoid structures (TLSs) are found at sites of chronic inflammation and consist primarily of germinal centers (GCs), peripheral T-cell areas, and high endothelial venules (HEVs). TLSs are typically formed during conditions causing chronic inflammation, such as infections, autoimmune disorders, tissue transplantation, and cancer. With immunotherapy at the forefront of cancer treatment strategies, effective biomarkers for immunotherapy have gained significant research attention. In recent years, several studies have shown that TLSs can improve the efficacy of immunotherapy; therefore, TLSs can be used as specific biomarkers for immune checkpoint inhibitors (ICIs). In this review, we discuss the development of TLSs, their role in the tumor immune microenvironment, and their therapeutic significance as biomarkers.