Rational design, synthesis, pharmacophore modeling, and docking studies for identification of novel potent DNA-PK inhibitors.

Drugs of cancer based upon ionizing radiation or chemotherapeutic treatment may affect breaking of DNA double strand in cell. DNA-PK enzyme has emerged as an attractive target for drug discovery efforts toward DNA repair pathways. Hence, the search for potent and selective DNA-PK inhibitors has particularly considered state-of-the art and several series of inhibitors have been designed. In this article, a novel benchmark DNA-PK database of 43 compounds was built and described. Ligand-based approaches including pharmacophore and QSAR modeling were applied and novel models were introduced and analyzed for predicting activity test for DNA-PK drug candidates. Based upon the modeling results, we gave a report of synthesis of fifteen novel 2-((8-methyl-2-morpholino-4-oxo-4H-benzo[e][1,3]oxazin-7-yl)oxy)acetamide derivatives and in vitro evaluation for DNA-PK inhibitory and antiproliferative activities. These fifteen compounds overall are satisfied with Lipinski's rule of five. The biological testing of target compounds showed five promising active compounds 7c, 7d, 7f, 9e and 9f with micromolar DNA-PK activity range from 0.25 to 5µM. In addition, SAR of the compounds activity was investigated and confirmed that the terminal aryl moiety was found to be quite crucial for DNA-PK activity. Moreover flexible docking simulation was done for the potent compounds into the putative binding site of the 3D homology model of DNA-PK enzyme and the probable interaction model between DNA-PK and the ligands was investigated and interpreted.

Modification of an expression vector for efficient recombinant production and purification of mitogillin of Aspergillus fumigatus expressed in Escherichia coli.

The diagnostic potential of secretory proteins of Aspergillus fumigatus is limited by their availability in pure form. We have constructed a vector (pGES-PH-1) to express genes encoding secretory proteins of A. fumigatus as fusion proteins with glutathione S-transferase (GST) in Escherichia coli. The mitogillin, a secretary protein of A. fumigatus, was expressed and purified to homogeneity by using pGES-PH-1. Mitogillin gene was PCR amplified from A. fumigatus DNA, cloned in pGES-PH-1 and expressed in E. coli as fusion protein with GST at N-terminal and 6xHis tag at C-terminal end. Pure mitogillin was obtained by purification on glutathione-Sepharose, cleavage of column-bound fusion protein by PreScission protease and by further purification on Ni-NTA-agarose. Polyclonal anti-mitogillin antibodies were raised in rabbits and were used to study its secretion during in vitro growth of A. fumigatus. The mitogillin was detectable in culture filtrate after 24 h of A. fumigatus growth and thereafter its amount increased progressively until 96 h in both, Sabouraud dextrose broth and potato dextrose broth. However, the secretion of mitogillin in culture medium was slightly delayed when A. fumigatus was grown in a minimal medium as mitogillin was detected only after 36 h of growth. Our results demonstrate the utility of the newly constructed expression vector with two affinity tags and PreScission protease cleavage site for high-level expression and efficient purification of a recombinant A. fumigatus secretory protein expressed in E. coli, which could be used for further studies.

Synthesis and characterization of fluorescent pyrrolyldipyrrinato Sn(IV) complexes.

A series of neutral, 5-coordinate pyrrolyldipyrrinato Sn(IV) complexes have been synthesized via reaction of a pyrrolyldipyrrin, or its corresponding hydrochloride salt, with dibutyltin or diphenyltin oxide. The complexes are structurally unique in that all three nitrogen atoms of the pyrrolyldipyrrinato ligand bind to the tin center, making these complexes the first examples of pyrrolyldipyrrins behaving as LX(2) ligands. The complexes are highly fluorescent, exhibiting fluorescence quantum yields between 0.28 and 0.61, and display interesting preliminary biological activity.

Design, Synthesis, Molecular Modeling, Anticancer Studies, and Density Functional Theory Calculations of 4-(1,2,4-Triazol-3-ylsulfanylmethyl)-1,2,3-triazole Derivatives.

New conjugates of substituted 1,2,3-triazoles linked to 1,2,4-triazoles were synthesized starting from the appropriate S-propargylated 1,2,4-triazoles 7 and 8. Ligation of 1,2,4-triazoles to the 1,2,3-triazole core was performed through Cu(I)-catalyzed cycloaddition of 1,2,4-triazole-based alkyne side chain 7 and/or 8 with several un/functionalized alkyl- and/or aryl-substituted azides 9-15 to afford the desired 1,4-disubstituted 1,2,3-triazoles 16-27, using both classical and microwave methods. After their spectroscopic characterization (infrared, 1H, 13C nuclear magnetic resonance, and elemental analyses), an anticancer screening was carried out against some cancer cell lines including human colon carcinoma (Caco-2 and HCT116), human cervical carcinoma (HeLa), and human breast adenocarcinoma (MCF-7). The outcomes of this exploration revealed that compounds 17, 22, and 25 had a significant anticancer activity against MCF-7 and Caco-2 cancer cell lines with IC50 values of 0.31 and 4.98 µM, respectively, in relation to the standard reference drug, doxorubicin. Enzyme-docking examination was executed onto cyclin-dependent kinase 2; a promising aim for cancer medication. Synthesized compounds acquiring highest potency showcased superior interactions with the active site residue of the target protein and exhibited minimum binding energy. Finally, the density functional theory (DFT) calculations were carried out to confirm the outcomes of the molecular docking and the experimental findings. The chemical reactivity descriptors such as softness (δ), global hardness (η), electronegativity (χ), and electrophilicity were calculated from the levels of the predicted frontier molecular orbitals and their energy gap. The DFT results and the molecular docking calculation results explained the activity of the most expectedly active compounds 17, 22, and 25.

Frequent allelic loss of Dkk-1 locus (10q11.2) is related with low distant metastasis and better prognosis in head and neck squamous cell carcinomas.

Head and neck squamous cell carcinoma (HNSCC) is a frequently occurring cancer worldwide. Dickkopf (Dkk)-1 gene is suggested to function as tumor suppressor gene (TSG) in several kinds of malignancies. In this study, we performed loss of heterozygosity (LOH) analysis of Dkk-1 and examined the correlation between LOH status and clinicopathological parameters for the first time. A pretty high LOH ratio (50%) was detected. Interestingly, in the cases with Dkk-1 retention group showed less distant metastasis and a tendency of longer disease free survival. These results indicate that Dkk-1 can play a role in HNSCC carcinogenesis and it may also be related to distant metastasis.

Formation of vinylic dipyrroles by the deprotonation of meso-alkyl and meso-benzyl dipyrrin HCl salts.

Under basic conditions, dipyrrin salts bearing alkyl and benzyl groups at the meso-position undergo deprotonation to give vinylic dipyrroles, rather than the corresponding free-base dipyrrins. The deprotonation is reversible and quantitatively returns the dipyrrinato framework under acidic conditions.

Fine deletion analysis of 1p36 chromosomal region in oral squamous cell carcinomas.

BACKGROUND: Squamous cell carcinoma is the most common cancer type of the oral cavity and approximately 50% of the patients succumb to the disease. Unfortunately, few are known about the molecular mechanisms involving in the formation of oral squamous cell carcinoma (OSCC). Recently, it has been reported that 1p36 chromosomal region is deleted in various cancer types and is suspected to harbor various tumor suppressor genes (TSGs). However, limited studies exist on genetics alteration on 1p36 in OSCC and the responsible TSG remained unidentified. METHODS: To investigate area susceptible to harbor TSG(s) involved in OSCC on 1p36 region, paired normal and tumor tissues of 27 patients with diagnosis of OSCC have been analyzed for loss of heterozygosity (LOH) using nine microsatellite markers based on recent gene mapping. RESULTS: LOH was found at least in one locus in 85% of the cases (23 of 27). Interestingly, microsatellite instability was also found in 7% (two of 27) of the cases analyzed. The higher LOH frequencies were found with the markers D1S243 (25%), D1S468 (22%), D1S450 (25%), D1S228 (38%), D1S199 (28%), and D1S1676 (23%). CONCLUSIONS: Three preferentially deleted regions have been identified in OSCC: region 1 (D1S468-D1S243), region 2 (D1S450-D1S228), and region 3 (D1S199-D1S1676). Multiple candidate TSGs, such as RIZ1, p73, UBE4B, Rap1GAP, EPHB2, and RUNX3, are located in these three areas. The data obtained in this study can be used for further functional analysis of these genes involved in OSCC carcinogenesis.

Lack of B-RAF mutations in head and neck squamous cell carcinoma.

B-RAF is one of the most commonly mutated oncogenes in human cancer. However, the mutation status of B-RAF has not been established completely in HNSCC. We have analysed the mutation status of the kinase domain of the B-RAF gene (exons 11 and 15) in 91 Japanese HNSCC patients as well as 12 HNSCC cell lines. DNA was extracted and amplified by PCR. Mutations were then analysed by SSCP mutation detection method. Since V600EB-RAF constitutes 90 % of the mutations identified in B-RAF in human cancers, we also used MASA analysis to specifically detect this mutation in exon 15 of B-RAF. Using both methods, no mutation was found in both exon 11 and 15 in all patients and cell lines. Mu tations are absent or rare in the kinase domain of B-RAF in Japanese HNSCC. However, more studies are still needed to determine its usefulness as a target for molecular therapy in these patients.

Highly diastereoselective templated complexation of dipyrromethenes.

[reaction: see text] Bis(dipyrromethene)s with relatively long spacers (>5 atoms) form helical monomeric complexes as the two binding units of the bis(dipyrromethene) chelate around the same tetrahedrally coordinated metal ion. Herein we report the first highly diastereoselective mononuclear helicate-forming complexation reactions of bis(dipyrromethene)s using homochiral binol and tartrate motifs which serve as both linkers and asymmetric templates.