Discovery of a Potent (4 R,5 S)-4-Fluoro-5-methylproline Sulfonamide Transient Receptor Potential Ankyrin 1 Antagonist and Its Methylene Phosphate Prodrug Guided by Molecular Modeling.

Transient receptor potential ankyrin 1 (TRPA1) is a non-selective cation channel expressed in sensory neurons where it functions as an irritant sensor for a plethora of electrophilic compounds and is implicated in pain, itch, and respiratory disease. To study its function in various disease contexts, we sought to identify novel, potent, and selective small-molecule TRPA1 antagonists. Herein we describe the evolution of an N-isopropylglycine sulfonamide lead (1) to a novel and potent (4 R,5 S)-4-fluoro-5-methylproline sulfonamide series of inhibitors. Molecular modeling was utilized to derive low-energy three-dimensional conformations to guide ligand design. This effort led to compound 20, which possessed a balanced combination of potency and metabolic stability but poor solubility that ultimately limited in vivo exposure. To improve solubility and in vivo exposure, we developed methylene phosphate prodrug 22, which demonstrated superior oral exposure and robust in vivo target engagement in a rat model of AITC-induced pain.

First report of Rahnella aquatilis infection in crucian carp Carassius auratus in China.

Rahnella aquatilis infection is rare in aquaculture. Here, a Gram-negative rod-shaped bacterium was isolated from diseased crucian carp Carassius auratus in Xuzhou City, Jiangsu Province, eastern China. The isolate was tentatively named strain KCL-5, and subsequently identified as R. aquatilis by biochemical properties and molecular techniques. The results showed that the isolate KCL-5 was most closely related to the type strain ATCC33071 (= DSM4594) of R. aquatilis, which shared 99.67, 96.26 and 99.58% nucleotide sequence identities for 16S rDNA, gyrB and toxin yhaV genes, respectively. Experimental challenges were conducted which demonstrated pathogenicity of the isolate in crucian carp. Antimicrobial susceptibility testing showed that the isolated strain was susceptible to piperacillin, gentamicin, kanamycin, nalidixic acid, norfloxacin, ofloxacin, azithromycin and erythromycin. To our knowledge, this is the first report on R. aquatilis infection in crucian carp, and the first evidence of pathogenicity in fish.

[Two-Dimensional Hetero-Spectral Near-Infrared and Mid-Infrared Correlation Spectroscopy for Discrimination Adulterated Milk].

New approach for discriminant analysis of adulterated milk is proposed based on combining hetero-spectral two-dimensional (2D) near-infrared (NIR) and mid-infrared (IR) correlation spectroscopy along with multi-way partial least squares discriminant analysis (NPLS-DA). Firstly, 36 pure milk samples were collected and 36 adulterated milk with starch samples (0.01 to 1 g · L(-1)) were prepared by adding appropriate mass of starch into pure milk. Then, one-dimensional NIR transmittance spectra and IR attenuated total reflection spectra of pure milk and adulterated milk with starch were measured at room temperature. And the synchronous 2D NIR-IR (4200~4800 vs. 900~1700 cm(-1)) correlation spectra of all samples were calculated. Due to the trace of adulterants, the synchronous 2D IR-NIR correlation spectral differences between adulterated milk with starch and pure milk are very subtle. Consequently, it was impossible to directly distinguish whether the sample was pure milk or adulterated milk. Finally, 2D IR-NIR correlation spectra were to build a discriminant model to classify adulterated milk and pure milk. The classification accuracy rates of samples in calibration set and in prediction set were 95.8% and 100% respectively. Also, the NPLS-DA models were built based on 2D NIR and 2D IR correlation spectra, respectively. The classification accuracy rates of samples in prediction set were 95.8%. Comparison results showed that the NPLS-DA model could provide better results using 2D NIR-IR correlation spectra than using 2D NIR, and 2D IR correlation spectra. The proposed method can not only effectively extract the feature information of adulterants in milk, but also explores a new perspective method for detection of adulterated food.

Multi-substituted N-phenyl-2, 2-dichloroacetamide analogues as anti-cancer drugs: design, synthesis and biological evaluation.

Our earlier research has shown that mono-substituted N-phenyl-2, 2-dichloroacetamide exhibited much higher anti-cancer activity than the lead compound sodium dichloroacetate (DCA). In this paper, a variety of multi-substituted N-phenyl-2, 2-dichloroacetamides were synthesized and biologically evaluated. The results showed that 3, 5-disubstituted N-phenyl-2, 2-dichloroacetamide analogues had satisfactory potency. Among them, N-(3, 5-diiodophenyl)-2, 2-dichloroacetamide had an IC50 of 2.84 micromol x L(-1) against non-small cell lung cancer cell line A549 and could induce cancer cell apoptosis.

[Raman imaging based on morphological model for human breast cancer tissues].

Raman Mapping spectra were collected on the samples of the normal human breast duct epithelia and the infiltrating duct carcinoma with the excitation wavelength of 633 nm and fitted with the morphological model which was presented before by the author. The normalized coefficients were calculated and used to make Raman images. Examining these images, the authors find that they show quite clearly the distribution of the chemicals which the basis spectrum was mainly derived from and that the DNA coefficient image also indicates the positions and scales of the cell nucleus, while the bright DNA area is bigger and brighter for the infiltrating duct carcinoma than for the normal breast dust epithelia, suggesting the cell nucleus in cancer tissue are larger than those in normal tissue. The correlation coefficient images were also made on these mapping spectra and examined in comparison with these model images, and it was showed that the model images have higher resolution and sensitivity although they are very similar. This work is helpful to understanding the morphological basis of the breast tissue Raman spectra and to developing the Raman diagnostic method for breast tumor.

Protolimonoids from Melia toosendan.

Toosendanone A (1), a new euphane (tirucallane)-type triterpene bearing a five-membered ring in the side chain and the first cyclopentanyl protolimonoid, was isolated from the bark of Melia toosendan, along with two new tirucallanes, toosendanic acids A (2) and B (3). The structure and absolute configuration of compound 1 was elucidated by spectroscopic data interpretation and X-ray diffraction analysis. Compounds 1-3 were evaluated for cytotoxicity against a small panel of cancer cell lines.

3D-QSAR studies of boron-containing dipeptides as proteasome inhibitors with CoMFA and CoMSIA methods.

Three-dimensional quantitative structure-activity relationship (3D-QSAR) studies were performed for a series of dipeptide boronate proteasome inhibitors using comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) techniques. A training set containing 46 molecules served to establish the models. The optimum CoMFA and CoMSIA models obtained for the training set were all statistically significant with cross-validated coefficients (q(2)) of 0.676 and 0.630 and conventional coefficients (r(2)) of 0.989 and 0.956, respectively. The predictive capacities of both models were successfully validated by calculating a test set of 13 molecules that were not included in the training set. The predicted correlation coefficients (r(2)(pred)) of CoMFA and CoMSIA are 0.963 and 0.919, respectively. The CoMFA and CoMSIA field contour maps agree well with the structural characteristics of the binding pocket of beta5 subunit of 20S proteasome, which suggests that the 3D-QSAR models constructed in this paper can be used to guide the development of novel dipeptide boronate inhibitors of 20S proteasome.

[Study on Raman linear model of human breast tissue].

A linear regression model for Raman spectra of human breast tissue was developed with 10 basis spectra which were derived from the spectra of fat, cell cytoplasm, collagen, beta-carotene, DNA and cholesterol etc. This model was tested statistically with more than 2 000 spectra collected from both normal and tumor breast tissue samples. The confidence of F-test for the model significance was 1 to all the spectra and the average of multiple determination coefficients measuring the fit goodness was 0.95. The normalized fit coefficients reflected, to a certain extent, the relative quantities of the Raman-active components in tissue represented by the basis spectra. In comparison with normal breast tissue, the coefficients of the cell cytoplasm and DNA basis spectra increased apparently for the tumor one, and the coefficient of fat basis spectrum changed reversely, which was easily understood with the known pathological changes in the tissue. Mapping spectra were also fitted with this model. This work is helpful to understanding the biochemical/morphological changes in the breast tumor tissues and to developing the Raman diagnostic method for breast tumor.

Characterization and expression analysis of Paralichthys olivaceus voltage-dependent anion channel (VDAC) gene in response to virus infection.

Voltage-dependent anion channel (VDAC, also known as mitochondrial porin) is acknowledged to play an important role in stress-induced mammalian apoptosis. In this study, Paralichthys olivaceus VDAC (PoVDAC) gene was identified as a virally induced gene from Scophthalmus Maximus Rhabdovirus (SMRV)-infected flounder embryonic cells (FEC). The full length of PoVDAC cDNA is 1380 bp with an open reading frame of 852 bp encoding a 283 amino acid protein. The deduced PoVDAC contains one alpha-helix, 13 transmembrane beta-strands and one eukaryotic mitochondrial porin signature motif. Constitutive expression of PoVDAC was confirmed in all tested tissues by real-time PCR. Further expression analysis revealed PoVDAC mRNA was upregulated by viral infection. We prepared fish antiserum against recombinant VDAC proteins and detected the PoVDAC in heart lysates from flounder as a 32 kDa band on western blot. Overexpression of PoVDAC in fish cells induced apoptosis. Immunofluoresence localization indicated that the significant distribution changes of PoVDAC have occurred in virus-induced apoptotic cells. This is the first report on the inductive expression of VDAC by viral infection, suggesting that PoVDAC might be mediated flounder antiviral immune response through induction of apoptosis.

3D-QSAR study of 20 (S)-camptothecin analogs.

AIM: To build up a quantitative structure-activity relationship (QSAR) model of 20 (S)-camptothecin (CPT) analogs for the prediction of the activity of new CPT analogs for drug design. METHODS: A training set of 43 structurally diverse CPT analogs which were inhibitors of topoisomerase I were used to construct a quantitative structure-activity relationship model with a comparative molecular field analysis (CoMFA). The QSAR model was optimized using partial least squares (PLS) analysis. A test set of 10 compounds was evaluated using the model. RESULTS: The CoMFA model was constructed successfully, and a good cross-validated correlation was obtained in which q(2) was 0.495. Then, the analysis of the non-cross-validated PLS model in which r(2) was 0.935 was built and permitted demonstrations of high predictability for the activities of the 10 CPT analogs in the test set selected in random. CONCLUSION: The CoMFA model indicated that bulky negative-charged group at position 9, 10 and 11 of CPT would increase activity, but excessively increasing bulky group at position 10 is adverse to inhibitory activity; substituents that occupy position 7 with the bulky positive group will enhance the inhibitive activity. The model can be used to design new CPT analogs and understand the mechanism of action.

[Clinical observation on point implantation of slowly-releasing medication for treatment of functional ventricular premature].

OBJECTIVE: To probe into a safe and effective method for treatment of functional ventricular premature, and develop a new preparation of Chinese herbs with high science and technique contents. METHODS: Two hundred and thirty-eight cases were randomly divided into a slowly-releasing medication group (n = 118) and a western medicine group (n = 120). The slowly-control needle group were treated with intramuscular injection of slowly-releasing medication new type preparation made by proved recipe of Chinese herbs with replenishing and strengthening pectoral qi, and nourishing yin and tranquillization into Tanzhong (CV 17), Neiguan (PC 6), Xinshu (BL 15), Pishu (BL 20) and Feishu (BL 13) by a trocar to slowly release and control releasing of the medicine so as to maintain lasting stimulation; the western medicine group with oral administration of Rythmol, thrice daily, 150 mg each time. RESULTS: The cured rate and the cured and markedly effective rate for the ventricular premature was 51.7% and 71.2% in the slowly-releasing medication group, which were significantly higher than 28.3% and 50.8% in the western medicine group (P < 0.001). CONCLUSION: The slowly-releasing medication new type preparation has a better therapeutic effect on functional ventricular premature, with high safety.

Molecular characterisation and inductive expression of a fish protein arginine methyltransferase 1 gene in response to virus infection.

Protein arginine methyltransferase 1 (PRMT1) is currently thought as an effector to regulate interferon (IFN) signalling. Here Paralichthys olivaceus PRMT1 (PoPRMT1) gene was identified as a virally induced gene from UV-inactivated Scophthalmus maximus Rhabdovirus (SMRV)-infected flounder embryonic cells (FEC). PoPMRT1 encodes a 341-amino-acid protein that shares the conserved domains including post-I, motif I, II and III. Homology comparisons show that the putative PoPMRT1 protein is the closest to zebrafish PMRT1 and belongs to type I PRMT family (including PRMT1, PRMT2, PRMT3, PRMT4, PRMT6, PRMT8). Expression analyses revealed an extensive distribution of PoPMRT1 in all tested tissues of flounder. In vitro induction of PoPRMT1 was determined in UV-inactivated SMRV-infected FEC cells, and under the same conditions, flounder Mx was also transcriptionally up-regulated, indicating that an IFN response might be triggered. Additionally, live SMRV infection of flounders induced an increased expression of PoPRMT1 mRNA and protein significantly in spleen, and to a lesser extent in head kidney and intestine. Immunofluorescence analysis revealed a major cyptoplasmic distribution of PoPRMT1 in normal FEC but an obvious increase occurred in nucleus in response to UV-inactivated SMRV. This is the first report on in vitro and in vivo expression of fish PRMT1 by virus infection, suggesting that PoPRMT1 might be implicated in flounder antiviral immune response.

[Exploring relationship between traditional effects of traditional Chinese medicine and modern pharmacological activities by "co-effect compounds"].

The compound that distributes in the herbs with one common effect was named as "co-effect compound" (CEC). The CECs of three traditional Chinese medicine(TCM) effects, purgative, relieving pain and clearing heat, had been found and studied. A strong corresponding relationship was found between the pharmacological activities of CECs and the TCM effect they belong to. The study shows that it may be a feasible method to connect traditional effect of TCM with modem pharmacological activity.