4.7 Article

Tumor-Specific Detection of an Optically Targeted Antibody Combined with a Quencher-Conjugated Neutravidin Quencher-Chaser: A Dual Quench and Chase Strategy to Improve Target to Nontarget Ratios for Molecular Imaging of Cancer

Journal

BIOCONJUGATE CHEMISTRY
Volume 20, Issue 1, Pages 147-154

Publisher

AMER CHEMICAL SOC
DOI: 10.1021/bc8003765

Keywords

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Funding

  1. NIH
  2. National Cancer Institute
  3. Center for Cancer Research
  4. NATIONAL CANCER INSTITUTE [ZIABC010656] Funding Source: NIH RePORTER

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In vivo molecular cancer imaging with monoclonal antibodies has great potential not only for cancer detection, but also for cancer characterization. However, the prolonged retention of intravenously injected antibody in the blood causes low target tumor-to-background ratio (TBR). Avidin has been used as a chase to clear the unbound, circulating biotinylated antibody and decrease the background signal. Here, we utilize a combined approach of a fluorescence resonance energy transfer (FRET) quenched antibody with an avidin chase to increase TBR. Trastuzumab, a humanized monoclonal antibody against human epidermal growth factor receptor type 2 (HER2), was biotinylated and conjugated with the near-infrared (NIR) fluorophore Alexa680 to synthesize Tra-Alexa680-biotin. Next, the FRET quencher, QSY-21, was conjugated to avidin, neutravidin (nAv), or streptavidin (sAv), thus creating Av-QSY21, nAv-QSY21, or sAv-QSY21 as chasers. The fluorescence was quenched in vitro by binding Tra-Alexa680-biotin to Av-QSY21, nAv-QSY21, or sAv-QSY21. To evaluate if the injection of quencher-conjugated avidin derivatives can improve target TBR by using a dual quench and chase strategy, both target (3T3/HER2+) and nontarget (Balb3T3/ZsGreen) tumor-bearing mice were employed. The FRET quench effect induced by all the QSY21 avidin-based conjugates reduced but did not totally eliminate background signal from the blood pool. The addition of nAv-QSY21 administration increased target TBR mainly because of the chase effect where unbound conjugated antibody was preferentially cleared to the liver. The relatively slow clearance of unbound nAv-QSY21 leads to further reductions in background signal by leaking out of the vascular space and binding to unbound antibodies in the extravascular space of tumors, resulting in decreased nontarget tumor-to-background ratios but increased target TBR due to the FRET quench effect, because target-bound antibodies were internalized and could not bind to nAv-QSY21. In conclusion, the proposed quench-and-chase system combines two strategies, fluorescent quenching and avidin chasing, to improve target TBR and reduce nontarget TBR, which should result in both improved tumor sensitivity and improved specificity.

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