Single-Cell Characterization of 18F-FLT Uptake with Radioluminescence Microscopy

The radiotracer 3'-deoxy-3'-F-18-fluorothymidine (F-18-FLT) is commonly used to measure cell proliferation in vivo. As a marker of cell proliferation, F-18-FLT is expected to be differentially taken up by arrested and actively dividing cells, but PET measures only aggregate uptake by tumor cells and therefore the single-cell distribution of F-18-FLT is unknown. We used a novel in vitro radioluminescence microscopy technique to measure the differential distribution of F-18-FLT radiotracer with single-cell precision. Methods: Using radioluminescence microscopy, we imaged the absolute uptake of F-18-FLT in live MDA-MB-231 cells grown under different serum conditions. We then compared F-18-FLT uptake with a standard measure of cell proliferation, using fluorescence microscopy of 5-ethynyl-2'-deoxyuridine incorporation in fixed cells. Results: According to 5-ethynyl-2'-deoxyuridine staining, few cells (1%) actively cycled under serum deprivation whereas most of them (71%) did under 20% serum. The distribution of F-18-FLT reflected this dynamic. At 0% serum, uptake of F-18-FLT was heterogeneous but relatively low. At 20% serum, a subpopulation of F-18-FLT-avid cells, representing 61% of the total population, emerged. Uptake of F-18-FLT in this population was 5-fold higher than in the remainder of the cells. Such a dichotomous distribution is not typically observed with other radiotracers, such as F-18-FDG. Conclusion: These results suggest that increased F-18-FLT uptake by proliferating cells is due to a greater fraction of F-18-FLT-avid cells rather than a change in F-18-FLT uptake by individual cells. This finding is consistent with the fact that F-18-FLT uptake is mediated by thymidine kinase 1 expression, which is higher in actively dividing cells. Overall, these findings suggest that, within the same patient, changes in F-18-FLT uptake reflect changes in the number of actively dividing cells, provided other parameters remain the same.