Chemiluminescent Probe for the In Vitro and In Vivo Imaging of Cancers Over‐Expressing NQO1

Abstract

Activatable (turn‐on) probes that permit the rapid, sensitive, selective, and accurate identification of cancer‐associated biomarkers can help drive advances in cancer research. Herein, a NAD(P)H:quinone oxidoreductase‐1 (NQO1)‐specific chemiluminescent probe 1 is reported that allows the differentiation between cancer subtypes. Probe 1 incorporates an NQO1‐specific trimethyl‐locked quinone trigger moiety covalently tethered to a phenoxy‐dioxetane moiety through a para‐aminobenzyl alcohol linker. Bio‐reduction of the quinone to the corresponding hydroquinone results in a chemiluminescent signal. As inferred from a combination of in vitro cell culture analyses and in vivo mice studies, the probe is safe, cell permeable, and capable of producing a “turn‐on” luminescence response in an NQO1‐positive A549 lung cancer model. On this basis, probe 1 can be used to identify cancerous cells and tissues characterized by elevated NQO1 levels.

An Activatable Photosensitizer Targeting Human NAD(P)H: Quinone Oxidoreductase 1

“…We selected A549 (lung epithelial carcinoma) and H596 (non-smalla denosquamous lung carcinoma) since previously reported Western blots showed high and low (undetectable) hNQO1expression, respectively. [8,54,55] Thus, we expected that if hNQO1w as responsible for 1 activation, we would observea turn-onr esponse in the A549 cells and minimal activation in H596.…”

“…Next, we wanted to identify whether 1 would be selectively activated by intracellular hNQO1 and facilitate the production of 1 O 2 upon irradiation of liberated PN in human cancer cell lines. We selected A549 (lung epithelial carcinoma) and H596 (non‐small adenosquamous lung carcinoma) since previously reported Western blots showed high and low (undetectable) hNQO1 expression, respectively . Thus, we expected that if hNQO1 was responsible for 1 activation, we would observe a turn‐on response in the A549 cells and minimal activation in H596.…”

“…It has primary roles in detoxification, antioxidation and stabilization of p53 against proteasomal degradation . In many cancers, hNQO1 can be significantly overexpressed relative to its healthy tissue counterpart . For example, there is a 10‐ to 100‐fold increase in hNQO1 levels in pancreatic cancer and its activity has been shown to be up to 90 times higher in lung cancers and at least 6 times higher in breast cancer .…”

“…[7] In many cancers, hNQO1c an be significantly overexpressed relative to its healthy tissue counterpart. [8,9] For example, there is a1 0-to 100-fold increase in hNQO1l evels in pancreatic cancer [9][10][11] and its activity has been shownt ob eu p to 90 times higher in lung cancers and at least 6t imes higher in breast cancer. [12] Overexpression of hNQO1 has also been found at levels5 -200 times higheri ns tomach, colon,h ead and neckc ancers, and cholangiocarcinoma, compared to their normal tissues.…”

Persistent Chemiluminescent Glow of Phenoxy‐dioxetane Luminophore Enables Unique CRET‐Based Detection of Proteases

“…[12][13][14] Such probes are useful for non-invasive chemiluminescence imaging of living cells and in animals. [15][16][17][18][19] Despite this considerable progress, the phenoxy-dioxetane probess uffer from an inherent restriction:T hey can be turnedon only by phenol deprotection. For example, in ac lassic dioxetane-based probe for ap rotease, the phenol group of the luminophore is masked with ap eptide substrate througha4aminobenzyl alcohols elf-immolative linker ( Figure 1A).…”