Endoplasmic Reticulum-Targeting Near-Infrared Fluorescent Probe for CYP2J2 Activity and Its Imaging Application in Endoplasmic Reticulum Stress and Tumor

Abstract

CYP2J2 as an endoplasmic reticulum (ER)-expressed vital cytochrome P450 isoform participates in the metabolism of endogenous polyunsaturated fatty acids. Its abnormal expression and function are closely related to the progress of cancer and cardiovascular diseases. Herein, an ERtargeting near-infrared (NIR) fluorescent probe ER-BnXPI was developed for monitoring CYP2J2 activity, which possessed a high selectivity and sensitivity toward CYP2J2 among various CYP450 isoforms and exhibited excellent subcellular localization for ER. Then, the CYP2J2 variation behavior under the ER stress model was imaged by ER-BnXPI in living cells and successfully used for the in vivo imaging in different tumors that well distinguished tumor tissues from para-cancerous tissues. All these findings fully demonstrated that ER-BnXPI could be used as a promising tool for exploring the physiological function of CYP2J2 and provided some novel approach for the diagnosis and therapy of CYP2J2-related vascular inflammation and cancer.

Spatial Chemoproteomics for Mapping the Active Proteome

“…For example, to localize in lysosomes, chemical probes functionalized with morpholine groups were developed and used to monitor lysosomal carboxylesterase activity in live cells and zebrafish models [29] . To monitor enzyme activity in the ER, the Ma lab developed a near‐infrared (NIR) fluorescent probe, ER−BnXPI, for quantitative detection of cytochrome P450 isoform CYP2J2 activity [28,30] . An ER‐targeting methyl sulfonamide moiety conjugated to an O ‐alkyl caged quenched dye resulted in strong accumulation in the ER and activatable fluorescence upon demethylation by CYP2J2.…”

“…[25] As a result, many research groups have turned to activatable "turnon" probes to quantify spatially defined enzyme activities. [11b] Using various approaches, chemical probes to target specific enzymes or families of enzymes have emerged for localizing to the plasma membrane, [26] mitochondria, [27] ER, [28] and lysosomes. [29] For example, to localize in lysosomes, chemical probes functionalized with morpholine groups were developed and used to monitor lysosomal carboxylesterase activity in live cells and zebrafish models.…”

“…[29] To monitor enzyme activity in the ER, the Ma lab developed a near-infrared (NIR) fluorescent probe, ERÀ BnXPI, for quantitative detection of cytochrome P450 isoform CYP2J2 activity. [28,30] An ERtargeting methyl sulfonamide moiety conjugated to an O-alkyl caged quenched dye resulted in strong accumulation in the ER and activatable fluorescence upon demethylation by CYP2J2. This probe proved useful for monitoring the regulation of CYP2J2 activity under ER stress models in live cells and in the differentiation of tumor vs paracancerous tissue in vivo.…”

“…This probe uses another advantage of the covalent modification by employing an alkyne tag for analysis, allowing not only in situ characterization by fluorescence but also by LC-MS/MS and gel electrophoresis to confirm target engagement. Other organelle-targeting moieties, such as lipophilic cations to target mitochondria, [32] Hoechst dye to target the nucleus, [33] and sulfonamides to target the ER, [28,34] may find further utility to covalently profile active proteins and protein families in organelle-specific contexts. In fact, many of these have been and are currently being used for global identification of organellar proteomes [24] but have not yet been adapted to activity-based proteomic profiling.…”

A near-infrared fluorescent probe based on a hemi-cyanine skeleton for detecting CES1 activity and evaluating pesticide toxicity

“…5 Over the past decades, many fluorescent probes with excellent fluorescent properties have been developed for monitoring enzymatic activity, although their application has been limited by factors such as their low fluorescence quantum yield, short emission wavelength, and background signal interference, all of which require the development of new probes for the detection of enzymes. 6 Hemi-cyanine dyes are promising candidates for the nearinfrared (NIR) fluorescence imaging of bio-samples since they possess an emission wavelength of greater than 650 nm, 7 and they have been used for the activity monitoring of various enzymes, such as CYP2J2, 8,9 MAO-A, 10 UGT1A1, 11 pantetheinase 12 and so on. 1,13 Carboxylesterase 1 (CES1), as a broad-spectrum serine hydrolase that is mainly distributed in the liver, and participates in the metabolism of endogenous substances, the activation of exogenous drugs and the elimination of environmental toxicants.…”