Fluorescence Lifetime Imaging for Quantification of Targeted Drug Delivery in Varying Tumor Microenvironments.

RATIONALE

Trastuzumab (TZM) is a monoclonal antibody that targets the human epidermal growth factor receptor (HER2) and is clinically used for the treatment of HER2-positive breast tumors. However, the tumor microenvironment can limit the access of TZM to the HER2 targets across the whole tumor and thereby compromise TZM's therapeutic efficacy. An imaging methodology that can non-invasively quantify the binding of TZM-HER2, which is required for therapeutic action, and distribution within tumors with varying tumor microenvironments is much needed.

METHODS

We performed near-infrared (NIR) fluorescence lifetime (FLI) Forster Resonance Energy Transfer (FRET) to measure TZM-HER2 binding, using microscopy and widefield macroscopy, in HER2 overexpressing breast and ovarian cancer cells and tumor xenografts, respectively. Immunohistochemistry was used to validate imaging results.

RESULTS

NIR FLI FRET microscopy data show variations in intracellular distribution of bound TZM in HER2-positive breast AU565 and AU565 tumor-passaged XTM cell lines in comparison to SKOV-3 ovarian cancer cells. Macroscopy FLI (MFLI) FRET imaging data show that SKOV-3 tumors display reduced TZM binding compared to AU565 and XTM tumors, as validated by immunohistochemistry. Moreover, AU565/XTM and SKOV-3 tumor xenografts display different amounts and distributions of TME components, such as collagen and vascularity. Therefore, these results suggest that SKOV-3 tumors are refractory to TZM delivery due to their disrupted vasculature and increased collagen content.

CONCLUSION

Our study demonstrates that FLI is a powerful analytical tool to monitor the delivery of antibody drug tumor both in cell cultures and in vivo live systems. Especially, MFLI FRET is a unique imaging modality that can directly quantify target engagement with potential to elucidate the role of the TME in drug delivery efficacy in intact live tumor xenografts.