Thymidine salvage to reflect tumor cell proliferation: Implications for ¹⁸F-FLT PET as a biomarker in oncology

Objectives The PET tracer 3’-deoxy-3’[¹⁸F]-fluorothymidine (¹⁸F-FLT) potentially serves as a biomarker of proliferation by reporting utilization of thymidine salvage via thymidine kinase 1 (TK1) activity. ¹⁸F-FLT PET has shown promise in some clinical trials, yet it remains unclear what molecular processes affect TK1 regulation in cancer and thus ¹⁸F-FLT uptake in tumors. Therefore, we designed preclinical studies to explore the relationship between ¹⁸F-FLT PET imaging, cellular TK1 levels, and established metrics of proliferation in mouse models of human cancer.

Methods Tumor xenografts were generated in nude mice from human colorectal cancer cell lines (HCT-116, HCT-116^p21-/-, Lim2405, DiFi). ¹⁸F-FLT PET images were acquired in prognostic and therapeutic settings focusing primarily on EGFR inhibition. Tumors were harvested for molecular analysis, including Western blot, qRT-PCR, and immunohistochemistry (IHC). Correlations were noted between tumor ¹⁸F-FLT uptake and TK1 levels at cellular, proteomic and genomic scales and compared to IHC. Molecular regulation of TK1 during EGFR targeted therapies was explored in DiFi cells and xenograft tumors.

Results Tumor uptake of ¹⁸F-FLT agreed well with TK1 protein levels. ¹⁸F-FLT PET and TK1 levels exhibited variable agreement with Ki67. Treatment of DiFi cells and xenografts with mAb-C225 showed that transcriptional and translational regulation of TK1 were required to reduce ¹⁸F-FLT uptake. Despite clinical response to mAb-C225 in vivo at all dosages evaluated, DiFi cells could escape translational regulation of TK1 through an mTOR-driven, cap-dependant mechanism resulting in failure of ¹⁸F-FLT PET to signal decreased tumor proliferation at lower dosages.

Conclusions Molecular determinants affecting TK1 regulation and ¹⁸F-FLT PET are poorly understood across tumor types and therapeutic settings. Further basic validation studies may inform the correct clinical utilization and interpretation of ¹⁸F-FLT PET in oncology