Proliferation markers, such as proliferating cell nuclear antigen (PCNA), Ki-67, and thymidine kinase 1 (TK1), have potential as diagnostic tools and as prognostic factors in assessing cancer treatment and disease progression. TK1 is involved in cellular proliferation through the recovery of the nucleotide thymidine in the DNA salvage pathway. TK1 upregulation has been found to be an early event in cancer development. In addition, serum levels of TK1 have been shown to be tied to cancer stage, so that higher levels of TK1 indicate a more serious prognosis. As a result of these findings and others, TK1 is not only a potentially viable biomarker for cancer recurrence, treatment monitoring, and survival, but is potentially more advantageous than current biomarkers. Compared to other proliferation markers, TK1 levels during S phase more accurately determine the rate of DNA synthesis in actively dividing tumors. Several reviews of TK1 elaborate on various assays that have been developed to measure levels in the serum of cancer patients in clinical settings. In this review, we include a brief history of important TK1 discoveries and findings, a comprehensive overview of TK1 regulation at DNA to protein levels, and recent findings that indicate TK1’s potential role in cancer pathogenesis and its growing potential as a tumor biomarker and therapeutic target.