Abstract 2841: A simplified approach to measuring thymidine kinase activity in cells, tumors, and blood as a biomarker of tumor proliferative potential

Abstract

Understanding tumor size and growth is essential for preclinical oncology drug development, but costs and technical hurdles can prevent accurate, repeated measurements of cell proliferation and tumor growth. Many investigators instead measure the accumulation of nucleosides and analogs (e.g., [18F]-FLT; [3H]-Thy; BrdU) in cells and tissues, a manifestation of thymidine kinase 1 activity (TK1a). These methods can be costly and laborious, and can generate significant hazardous waste, promoting the use of even less-direct measures of cell division. Seeking further insights into the role of TK1 in cancer, we developed rapid, robust, non-isotopic methods that measure TK1a in tissue extracts and blood. In this approach, samples are exposed to FLT in vivo or in vitro, promoting its phosphorylation. Subsequently, nucleosides and their phosphorylated metabolites are isolated for analysis by a simple organic extraction. Using these methods, TK1a was measured in primary and transformed cells, and showed excellent comparability with established proliferation assays including 3H-thymidine incorporation, ATP generation, and fluorescent dye dilution flow cytometry. In drug screens, our method was as sensitive as commercial viability assays, and typically detected drug effects much sooner than those less-specific measures. We also demonstrated chemotherapy-induced alterations in TK1a in xenografted tumor tissue that were comparable to results obtained using radioisotopes. Finally, this approach was adapted to measure TK1a in blood samples. Consistent with published reports, circulating TK1a correlated well with tumor burden in solid tumor models and primary canine hematological cancer. These methods require only small sample volumes (<50μL serum or other biofluids, or a few thousand cells), allowing serial blood collection for monitoring tumor growth kinetics in rodents. These translational studies are underway, and should greatly inform our understanding of the role of TK1 in tumor responses to treatment in vitro and in vivo. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2841. doi:1538-7445.AM2012-2841