An Activatable NIR Fluorescent Probe for NAD(P)H and Its Application to the Real-Time Monitoring of p53 Abnormalities In Vivo.

NAD(P)H is crucial for biosynthetic reactions and antioxidant functions. However, the current probes developed for detecting NAD(P)H in vivo require intratumoral injection, which limited their application for animal imaging. To address this issue, we have developed a liposoluble cationic probe, KC8, which exhibits excellent tumor-targeting ability and near-infrared (NIR) fluorescence after reaction with NAD(P)H. By using KC8, it was demonstrated for the first time that the level of NAD(P)H in the mitochondria of living colorectal cancer (CRC) cells was highly related to the abnormality of the p53. Furthermore, KC8 was successfully used to differentiate not only between tumor and normal tissue but also between tumors with p53 abnormality and normal tumors when administered intravenously. Finally, we evaluated tumor heterogeneity through two fluorescent channels after treating a tumor with 5-Fu. This study provides a new tool for real-time monitoring of the p53 abnormality of CRC cells.

Unique assembly of carbonylpyridinium and chromene reveals mitochondrial thiol starvation under ferroptosis and novel ferroptosis inducer.

To reveal the delicate function of mitochondria, spatiotemporally precise detection tools remain highly desirable. However, current probes with positively charged warheads for targeting mitochondria diffuse out of the mitochondria as the potential of the mitochondrial membrane changes, which directly influences the accuracy of the detection. Herein, we assembled carbonylpyridinium and chromene to afford the probe CM-Mit. Following the ultrafast response to thiol and the dissociation of carbonylpyridinium, the formation of o-quinone methide from CM-Mit was proposed to label proteins, thus avoiding diffusion out of the mitochondria. Therefore, the accurate spatiotemporal detection of thiol in mitochondria was realized. With this excellent probe, ferroptosis inducers were proved to stimulate thiol starvation in mitochondria for the first time in cancer cells. Moreover, CM-Mit was used to screen a compound library developed in-house and the stemona alkaloid analog SA-11 was shown to induce ferroptosis in various cancer cell lines, including a drug-resistant one.

Endogenous cysteine fluorescence monitoring and its deployment in tumour demarcation.

A cysteine-specific fluorescent probe with a wide concentration detection range was used to monitor changes in cysteine levels in HeLa cells under stress and to demarcate the boundary of a xenograft tumour.

A sequentially activated bioluminescent probe for observation of cellular H2O2 production induced by cysteine.

We report herein a caged luciferin probe Cy-Hy as a sequentially activated probe to selectively and sensitively sense L-Cys and H2O2. The probe displayed fluorescence and bioluminescence responses toward the two analytes. Utilizing the present probe, cellular excess L-Cys-induced H2O2 up-regulation was observed for the first time in living MDA-MB-231 cells.

Substitution-rearrangement-cyclization strategy to construct fluorescent probe for multicolor discriminative analysis biothiols in cells and zebrafish.

Discriminative detection of biothiols (Cysteine, homocysteine and glutathione) is of great significance to clarificate their complex physiological processes, the occurrence and development of related diseases. However, similar structure and reactivity among such species pose huge challenges in developing fluorescent probes to distinguish among of them. In this work, a dual-site probe CTT reacted with the analytes to regulate molecular conjugation through substitution-rearrangement-cyclization strategy, utilizing a multi-channel signal combination mode to realize the distinguishing detection of the three biothiols. Cell and zebrafish imaging experiments sufficiently demonstrated that CTT could semiquantify biothiols, which will provide valuable chemical tool for elucidating the complex biological functions of biothiols.

Comparing the abundance of HClO in cancer/normal cells and visualizing in vivo using a mitochondria-targeted ultra-fast fluorescent probe.

Organisms are operating and evolving with a highly sophisticated and intelligent defense mechanism to resist bacterial and viral infections. This process involves a variety of reactive oxygen species (ROS), and they coordinate with each other to support different physiological activities. Due to its strong oxidizing properties, hypochlorous acid (HClO), a part of ROS, is a powerful antimicrobial agent in living organisms and exerts a crucial role in the immune system. However, the excessive production of HClO can cause cell damage and even cell death. Herein, we combined benzene-conjugated benzopyrylium as the fluorophore and dimethylthiocarbamoyl chloride as the recognition site to rationally design a probe (BBD). The fluorescence of the probe was quenched based on an effective PET molecular mechanism. Surprisingly, BBD exhibited a turn-on red fluorescence signal for HClO with ultra-fast response (5 s) and high selectivity. Moreover, BBD located mitochondria well and it was found that the abundance of HClO is higher in HeLa cells compared to that in normal cells. Finally, BBD was successfully applied to the visualization of HClO in zebrafish and nude mice.

Discovery of a potent ß-catenin destabilizer for overcoming the resistance of 5-fluorouracil in colorectal cancer.

Wnt/ß-catenin signalling is frequently activated in colorectal cancer, in which nuclear ß-catenin accumulation contributes to tumour initiation and progression. However, therapeutic agents in clinical use targeting this pathway are lacking. In this report, we describe the synthesis of novel stemona alkaloid analogues and their biological evaluation, among which compound 3 was identified to efficiently inhibit various CRC cells, including 5-fluorouracil-resistant CRC cells. Mechanistically, this study revealed that compound 3 reduced the protein level of ß-catenin without affecting its mRNA level, which suggests an alternative mechanism for ß-catenin degradation. The expression of downstream proteins, including c-myc, survivin, and cyclin D1, was also significantly inhibited, even in Wnt-activated CRC cells. Briefly, our data highlight the potential of compound 3 as a destabilizer of ß-catenin for the treatment of CRC patients.

HSA-Lys-161 covalent bound fluorescent dye for in vivo blood drug dynamic imaging and tumor mapping.

The specific combination of human serum albumin and fluorescent dye will endow superior performance to a coupled fluorescent platform for in vivo fluorescence labeling. In this study, we found that lysine-161 in human serum albumin is a covalent binding site and could spontaneously bind a ketone skeleton quinoxaline-coumarin fluorescent dye with a specific turn-on fluorescence signal for the first time. Supported by the abundant drug binding domains in human serum albumin, drugs such as ibuprofen, warfarin and clopidogrel could interact with the fluorescent dye labeled human serum albumin to feature a substantial enhancement in fluorescence intensity (6.6-fold for ibuprofen, 4.5-fold for warfarin and 5-fold for clopidogrel). The drug concentration dependent fluorescence intensity amplification realized real-time, in situ blood drug concentration monitoring in mice, utilizing ibuprofen as a model drug. The non-invasive method avoided continuous blood sample collection, which fundamentally causes suffering and consumption of experimental animals in the study of pharmacokinetics. At the same time, the coupled fluorescent probe can be efficiently enriched in tumors in mice which could map a tumor with a high-contrast red fluorescence signal and could hold great potential in clinical tumor marking and surgical resection.

Thiol "Click" Chromene Ring Opening and Subsequent Cascade Nucleophilic Cyclization NIR Fluorescence Imaging Reveal High Levels of Thiol in Drug-Resistant Cells.

As the structural unit of natural products, chromene derivatives show a wide range of biological activity and pharmacological activity due to their unique photophysical and chemical properties. Ten years ago, our research group discovered the "thiol-chromene" click reaction, which achieved the selective detection of thiols through the change of the optical spectrum. Afterward, we attempted to develop various chromene-based fluorescent probes for imaging including near-infrared (NIR) probe, ratiometric probe, and multifunctional probe. However, how to integrate the fluorophore and reaction sites into the chromene-based skeleton remains challenging. In this work, we connected the chromene motif with the NIR fluorophore methylene blue utilizing a carbamate spacer to provide a new fluorescent probe (CM-NIR), which is triggered by thiols to open the pyran ring followed by attacking the carbamate by phenolate to releases the methylene blue. This novel cascade mechanism avoids the formation of para-quinone methides, which proved to be toxic to normal cells. CM-NIR also showed the specific imaging of thiols in living cells and mice. More importantly, the thiols level in drug-resistant cancer cells was found to be significantly higher than that in the corresponding cancer cell, which indicated that the thiols level may have an important role in cancer cells developing drug resistance.

Mutual correlation evaluation of Cys and Hcy in serum through reaction activity regulated fluorescence quantification.

We report herein a strategy involving using a single fluorescent probe with the assistance of two pH conditions to simultaneously detect cysteine and homocysteine quantitatively. The concentrations of Cys and Hcy in human serum samples were found to display a positive correlation, which might promote a further understanding of thiol-related diseases.

Pyrrole Strategy for the γ-Lactam-Containing Stemona Alkaloids: (±)Stemoamide, (±)Tuberostemoamide, and (±)Sessilifoliamide A.

Stemona alkaloids contain family members with diverse structural scaffolds. Many of them feature a γ-lactam ring embedded in their characteristic 5-7-5 fused tricyclic core. Herein a pyrrole strategy was developed to enable the total syntheses of three Stemona alkaloids: (±)stemoamide, (±)tuberostemoamide, and (±)sessilifoliamide A. In these cases, a substituted pyrrole was used as the γ-lactam precursor. A sequential pyrrole oxidation and enamide reduction were realized to convert the pyrrole to the corresponding γ-lactam in those three natural products. The use of a pyrrole in an early stage of the synthesis offers the advantage of rapid construction of the key intermediates by exploiting its nucleophilicity.

Novel Anthraquinone Compounds Inhibit Colon Cancer Cell Proliferation via the Reactive Oxygen Species/JNK Pathway.

A series of amide anthraquinone derivatives, an important component of some traditional Chinese medicines, were structurally modified and the resulting antitumor activities were evaluated. The compounds showed potent anti-proliferative activities against eight human cancer cell lines, with no noticeable cytotoxicity towards normal cells. Among the candidate compounds, 1-nitro-2-acyl anthraquinone-leucine (8a) showed the greatest inhibition of HCT116 cell activity with an IC50 of 17.80 µg/mL. In addition, a correlation model was established in a three-dimensional quantitative structure-activity relationship (3D-QSAR) study using Comparative Molecular Field Analysis (CoMFA) and comparative molecular similarity index analysis (CoMSIA). Moreover, compound 8a effectively killed tumor cells by reactive oxygen species (ROS)-JNK activation, causing an increase in ROS levels, JNK phosphorylation, and mitochondrial stress. Cytochrome c was then released into cytoplasm, which, in turn activated the cysteine protease pathway and ultimately induced tumor cell apoptosis, suggesting a potential use of this compound for colon cancer treatment.

A novel strategy: A consecutive reaction was used to distinguish in the presence of statins between normal cells and cancer cells.

Statins, as the most commonly drugs could reduce the concentration of low-density lipoprotein cholesterol, have been proved to elevate the H2S generation in cells. Besides, the abnormal levels of biothiols might lead to cancer. Therefore, it is worth considering how to combine the characteristics of the two diseases to realize the detection of cancer cells. Based on this view, we developed a multiresponse fluorescent probe for the detection of hydrogen sulfide (H2S) and biothiols successively based on theoretical calculation. It is interesting that the fluorescence intensity of the probe reacting H2S and biothiols successively was significantly higher than that of probe reacting either of them. Based on this view, we further explored the biological application of the probe and found that the probe had obvious signal response to cancer cells than the normal cells in the presence of fluvastatin. This interesting finding might provide a new insight into cancer cell recognition.

Formal enantioselective total synthesis of bisdehydroneostemoninine.

A formal enantioselective total synthesis of bisdehydroneostemoninine employing L-glutamic acid as the chiral pool is described. The key features of the synthesis include regioselective and enantioselective opening the chiral epoxide with dimethylsulfonium methylide and tandem Friedel-Crafts cyclization followed by lactonization to form the 5-7-5 tricyclic core of the target stemona alkaloids. The synthetic route provides opportunities to explore the biological behavior of enantiopure bisdehydroneostemoninine. [Formula: see text].

Natural product syntheses via carbonylative cyclizations.

Covering: 2000-2018In this review, we highlight recent examples of natural product total syntheses employing transition metal-mediated/catalyzed carbonylative cyclization strategies to build key ring systems. It mainly covers carbonylative cyclizations for the construction of O-heterocycles, N-heterocycles and carbocycles including cyclic ketones and phenols. The reaction types include carbonylation of epoxide to ß-lactones, carbonylative (macro)lactonization/lactamization, the Semmelhack reaction, tandem hydroformylation-cyclization, the Pauson-Khand reaction, carbonylative C-H activation cyclization, the Stille/Suzuki carbonylation, [n + m + 1] carbonylative cycloaddition, the Dötz annulation, and others.

Total Syntheses of Bisdehydroneostemoninine and Bisdehydrostemoninine by Catalytic Carbonylative Spirolactonization.

The first total syntheses of the stemona alkaloids bisdehydroneostemoninine and bisdehydrostemoninine in racemic forms have been achieved. The synthetic strategy features a novel palladium-catalyzed carbonylative spirolactonization of a hydroxycyclopropanol to rapidly construct the oxaspirolactone moiety. It also features a Lewis acid promoted tandem Friedel-Crafts cyclization and lactonization to form the 5-7-5 tricyclic core of the target stemona alkaloids.

Increasing the Level of IRS-1 and Insulin Pathway Sensitivity by Natural Product Carainterol A.

Carainterol A is a eudesmane sesquiterpenoid extracted from Caragana intermedia. We have reported that carainterol A showed potent glucose consumption activity in C2C12 muscle cells and the db/db mouse model. However, the mechanism of the hypoglycemic effect of carainterol A remains elusive. In this article, we present a network pharmacology approach to predict the target and signaling pathway of carainterol A which was subsequently validated in HepG2 cells. It was demonstrated that carainterol A could increase the protein levels of IRS-1 and the downstream protein kinase AKT phosphorylation at a low micromolar level. These findings suggest that carainterol A can be a valuable lead compound and a promising chemical probe for the insulin signaling pathway.

Rational design of 2-pyrrolinones as inhibitors of HIV-1 integrase.

HIV-1 integrase is an essential enzyme for viral replication and a validated target for the development of drugs against AIDS. With an aim to discover new potent inhibitors of HIV-1 integrase, we developed a pharmacophore model based on reported inhibitors embodying structural diversity. Eight compounds of 2-pyrrolinones fitting all the features of the pharmacophore query were found through the screening of an in-house database. These candidates were successfully synthesized, and three of them showed strand transfer inhibitory activity, in which, one compound showed antiviral activity. Further mapping analysis and docking studies affirmed these results.