An Optimized Triple Modality Reporter for Quantitative In Vivo Tumor Imaging and Therapy Evaluation

Abstract

We present an optimized triple modality reporter construct combining a far-red fluorescent protein (E2-Crimson), enhanced firefly luciferase enzyme (Luc2), and truncated wild type herpes simplex virus I thymidine kinase (wttk) that allows for sensitive, long-term tracking of tumor growth in vivo by fluorescence, bioluminescence, and positron emission tomography. Two human cancer cell lines (MDA-MB-231 breast cancer and HT-1080 fibrosarcoma cancer) were successfully transduced to express this triple modality reporter. Fluorescence and bioluminescence imaging of the triple modality reporter were used to accurately quantify the therapeutic responses of MDA-MB-231 tumors to the chemotherapeutic agent monomethyl auristatin E in vivo in athymic nude mice. Positive correlation was observed between the fluorescence and bioluminescence signals, and these signals were also positively correlated with the ex vivo tumor weights. This is the first reported use of both fluorescence and bioluminescence signals from a multi-modality reporter construct to measure drug efficacy in vivo.

Non-Invasive In Vivo Imaging and Quantification of Tumor Growth and Metastasis in Rats Using Cells Expressing Far-Red Fluorescence Protein

“…The absence of real-time information on the cellular makeup of the tumor, and true tumor volume and dimensions precludes the gathering of clinically relevant information relating tumor stage to responsiveness to therapy. Although, an E2-crimson, enzyme firefly luciferase, herpes simplex virus-1 thymidine kinase, triple modality reporter system has been recently described by Tsien an co-workers in mice subjects, the NIR reporter was not used to gather information on tumor cellularity or volume [ 27 ]. Therefore, the primary objectives of this study were to develop and validate E2-Crimson expressing cancer cells in order to image cellularity and tumor volume in a rat subject.…”

Fluorophore-NanoLuc BRET Reporters Enable Sensitive In Vivo Optical Imaging and Flow Cytometry for Monitoring Tumorigenesis

“…However, the signal intensities of these reporters have limited sensitivity in vivo , and the cassettes encoding these reporters are large, thus restricting their application when using viral delivery methods that require space to encode transgenes or shRNAs (10–14). Accordingly, a compact multi-modal reporter having enhanced signal intensity is needed for preclinical cancer studies.…”

Improved facial nerve identification during parotidectomy with fluorescently labeled peptide

“…Murine salivary gland cancer cells (SCA-9 clone 15, ATCC CRL-1734; American Type Culture Collection, Manassas, VA) were transfected with firefly luciferase enzyme (Luc2) as described in detail elsewhere. 18 Luc2 is a codon-optimized luciferase gene with an increased expression in mammalian cells; cancer cell detection has been reported to be possible at a single cell level. 19 There were 6 3 10 6 murine Luc2-transfected salivary gland cancer cells injected into the right parotid gland of every mouse.…”