Detection and destruction of HER2‐positive cancer cells by Ultra Quenchbody‐siRNA complex

Abstract

Ultra Quenchbody (UQ‐body) is a biosensor that utilizes the quenching behavior of the fluorescent dye linked to the antibody V region. When the corresponding antigen is bound to the UQ‐body, the fluorescence is restored and allows the detection of target molecules easily and sensitively. In this paper, we constructed UQ‐bodies to sensitively detect the human epidermal growth factor receptor 2 (HER2) cancer marker in solution or on cancer cells, which was further used to kill the cancer cells. A synthetic Fab fragment of anti‐HER2 antibody Fab37 with many Trp residues at hypervariable region was prepared and labeled with fluorescent dyes to obtain the UQ‐bodies. The UQ‐body could detect HER2 in solution at concentrations as low as 20 pM with an EC50 of 0.3 nM with a fourfold response. Fluorescence imaging of HER2‐positive cells was successfully performed without any washing steps. To deliver small interfering RNA (siRNA) to cancer cells, a modified UQ‐body with C‐terminal 9R sequence was also prepared. HER2‐positive cancer cells were effectively killed by polo‐like kinase 1 siRNA intracellularly delivered by the UQ‐body‐9R. The novel approach employing siRNA‐empowered UQ‐body could detect and image the HER2 antigen easily and sensitively, and effectively kill the HER2‐positive cancer cells.

Recent Advances in Quenchbody, a Fluorescent Immunosensor

“…The anti-HER2 antibody can block the downstream signal transduction of HER2 positive cells and cause cancer cell death. Dong and Oka et al constructed a Q-body for detecting HER2 in solution or on the surface of cancer cells with high sensitivity, and then to kill tumor cells [47]. They chose Fab37, an anti-HER2 antibody fragment previously selected from a synthetic binary encoding library by phage display.…”

“…(d) Bone Gla protein produced by vitamin D3-induced U2OS osteosarcoma; (e) HER2 on cancer cells with different HER2 expression levels. Reprinted with permission from[47].…”

Isolation of a Monoclonal Antibody and its Derived Immunosensor for Rapid and Sensitive Detection of 17β-Estradiol

“…In contrast, Quenchbody (Q-body), a fluorescent biosensor based on antigen-antibody reactions, is simple to operate, highly specific, and can significantly improve the sensitivity and accuracy of the traditional assay. The Q-body is a recombinant antibody fragment in which the N-terminus is specifically labeled with a fluorescent dye, and the fluorescence is quenched by a tryptophan residue in the variable region of the antibody based on photo-induced electron transfer ( Abe et al, 2011 ; Dong et al, 2020b ; Dong and Ueda, 2021 ). Specific binding of the antigen to the antibody results in quenching of the fluorescent dye inside the antibody.…”

Synthesis of Quenchbodies for One-Pot Detection of Stimulant Drug Methamphetamine

“…Since the fluorescence of the Q-body is quenched in the absence of antigen, its signal-to-background ratio is higher than that of other conventional dye-conjugated antibodies. We have utilized Q-bodies as biosensors for the rapid detection of various targets in a solution, and in some cases on cells, without the need for washing steps [13,14]. Moreover, Q-body assay for detecting BGP peptide was successfully performed not only in the biochemical buffer-based samples but also in 50% plasma with no apparent loss of response [8].…”

“…(underline: tag for labeling or for non-labeling; bold: site for the same; italic: linker). Extract the desired band and purify it using a NucleoSpin Gel Clean-Up Kit according to the manufacturer's instructions 14…”