Auxin-like effects of the natural coumarin scopoletin on Arabidopsis cell structure and morphology.

The mode of action and phytotoxic potential of scopoletin, a natural compound belonging to the group of coumarins, has been evaluated in detail. Analysis conducted by light and electron transmission microscopy showed strong cell and tissue abnormalities on treated roots, such as cell wall malformations, multi-nucleated cells, abnormal nuclei and tissue disorganization. Scopoletin compromised root development by inducing wrong microtubule assembling, mitochondrial membrane depolarization and ultimate cell death, in a way similar to auxin herbicides. The structural similarities of the natural compound scopoletin and the auxin herbicide 2,4-D, as well as the ability of scopoletin to fit into the auxin-binding site TIR1, were analyzed, suggesting that the phytotoxic activity of scopoletin matches with that exhibited by auxinic herbicides.

3D-QSAR and Molecular Docking Studies on the TcPMCA1-Mediated Detoxification of Scopoletin and Coumarin Derivatives.

The carmine spider mite, Tetranychus cinnabarinus (Boisduval), is an economically important agricultural pest that is difficult to prevent and control. Scopoletin is a botanical coumarin derivative that targets Ca2+-ATPase to exert a strong acaricidal effect on carmine spider mites. In this study, the full-length cDNA sequence of a plasma membrane Ca2+-ATPase 1 gene (TcPMCA1) was cloned. The sequence contains an open reading frame of 3750 bp and encodes a putative protein of 1249 amino acids. The effects of scopoletin on TcPMCA1 expression were investigated. TcPMCA1 was significantly upregulated after it was exposed to 10%, 30%, and 50% of the lethal concentration of scopoletin. Homology modeling, molecular docking, and three-dimensional quantitative structure-activity relationships were then studied to explore the relationship between scopoletin structure and TcPMCA1-inhibiting activity of scopoletin and other 30 coumarin derivatives. Results showed that scopoletin inserts into the binding cavity and interacts with amino acid residues at the binding site of the TcPMCA1 protein through the driving forces of hydrogen bonds. Furthermore, CoMFA (comparative molecular field analysis)- and CoMSIA (comparative molecular similarity index analysis)-derived models showed that the steric and H-bond fields of these compounds exert important influences on the activities of the coumarin compounds.Notably, the C3, C6, and C7 positions in the skeletal structure of the coumarins are the most suitable active sites. This work provides insights into the mechanism underlying the interaction of scopoletin with TcPMCA1. The present results can improve the understanding on plasma membrane Ca2+-ATPase-mediated (PMCA-mediated) detoxification of scopoletin and coumarin derivatives in T. cinnabarinus, as well as provide valuable information for the design of novel PMCA-inhibiting acaricides.

Design, synthesis and cytotoxic activities of scopoletin-isoxazole and scopoletin-pyrazole hybrids.

12 novel scopoletin-isoxazole and scopoletin-pyrazole hybrids were designed, synthesized and their chemical structures were confirmed by HR-MS, IR, 1H NMR and 13C NMR spectra. The anticancer activities of the newly synthesized compounds were evaluated in vitro against three human cancer cell lines including HCT-116, Hun7 and SW620 by MTT assay. The screening results showed that six compounds (9a, 9c, 9d, 12a, 18b and 18d) exhibited potent cytotoxic activities with IC50 values below 20µM. Besides, we have further evaluated the growth inhibitory activities of six compounds against the human normal tissue cell lines HFL-1. Especially, compound 9d displayed significant anti-proliferative activity with IC50 values ranging from 8.76µM to 9.83µM and weak cytotoxicity with IC50 value of 90.9µM on normal cells HFL-1, which suggested that isoxazole-based hybrids of scopoletin were an effective chemical modification to improve the anticancer activity of scopoletin.

Synthesis and in vitro antitumor activity of novel scopoletin derivatives.

Twenty scopoletin derivatives were developed by a systematic combinatorial chemical approach and their chemical structures were confirmed by MS, IR, (1)H NMR spectra and elemental analysis. Primary screening against mammary (MCF-7 and MDA-MB 231) and colon (HT-29) carcinoma cells indicated that five compounds (8d, 8g, 8j, 11b and 11g) displayed high antitumor potencies with IC(50) values below 20 µM whereas scopoletin showed IC(50) values above 100 µM. Moreover, the most promising compound 11g was more active than 5-fluorouracil. These results clearly indicated that the modification of the scopoletin structure could greatly increase its antitumor activity in vitro.

Scopoletin induces apoptosis in human promyeloleukemic cells, accompanied by activations of nuclear factor kappaB and caspase-3.

Scopoletin (6-methoxy-7-hydroxycoumarin) is a phenolic coumarin and a member of the phytoalexins. In this study we investigated whether scopoletin caused apoptosis in HL-60 promyelocytic cells and, if so, by what mechanisms. We found that scopoletin induced apoptosis as confirmed by a characteristic ladder pattern of discontinuous DNA fragments in a dose-dependent manner. The signal cascade activated by scopoletin included the heterodimeric redox-sensitive transcription factor NF-kappaB, which exhibited an upregulation of nuclear factor-kappa B (NF-kappaB) translocation to the nucleus by increase of IkappaBalpha degradation. In addition, scopoletin activated caspase-3 as was evidenced by both the proteolytic cleavage of the proenzyme and increased protease activity. Activation of caspase-3 resulted in the cleavage of 116 kDa poly(ADP-ribose) polymerase (PARP) to 85 kDa cleavage product in time-and dose-dependent fashions. Prior treatment of the cells with pyrrolidine dithiocarbamate, a potent inhibitor of NF-kappaB activation, or Ac-DEVD-CHO, a specific caspase-3 inhibitor, prevented scopoletin-induced caspase-3 activation, PARP cleavage, and finally DNA fragmentation. Taken together, these results suggest that scopoletin induces NF-kappaB activation, which, in turn, causes activation of caspase-3, degradation of PARP, and eventually leads to apoptotic cell death in HL-60 cells.

Structure-cytotoxicity relationships of a series of natural and semi-synthetic simple coumarins as assessed in two human tumour cell lines.

The cytotoxicity of 22 natural and semi-synthetic simple coumarins was evaluated in GLC4, a human small cell lung carcinoma cell line, and in COLO 320, a human colorectal cancer cell line, using the microculture tetrazolium (MTT) assay. With IC50 values > 100 microM, following a continuous (96 h) incubation, most coumarins exhibited only low cytotoxicity. Several compounds, however, displayed significant potencies. As far as the structure--cytotoxicity relationship is concerned, it is conspicuous that all the potentially active natural compounds possess at least two phenolic groups in either the 6,7- or 6,8-positions. In addition, the 5-formyl-6-hydroxy substituted semi-synthetic analogue was found to be potent, reflecting the importance of at least two polar functions for high cytotoxicity.

Non-mutagenicity of some wood-related compounds in the bacterial/microsome plate incorporation and microsuspension assays.

Seven commercially available wood-related compounds have been tested for mutagenicity by the use of the Ames and fluctuation test-systems. All compounds were found to be non-mutagenic. Among these compounds, 2,6-dimethoxy-p-benzoquinone showed a very weak and questionable mutagenic activity against Salmonella typhimurium TA 100. In this connection, probable causes of nasal adenocarcinoma of woodworkers are briefly discussed.