Research in Pharmaceutical Sciences
Volume:15 Issue: 3, Jun 2020

> α-D-ribofuranose analogues are reported to have multifarious biological properties such as analgesic, anti-inflammatory, and antiviral activities. The present study aims to synthesize some α-D-ribofuranose derivatives and investigate their biological properties.

 Four derivatives (2a, 2b, 3, and 4) were synthesized from the starting material 3-O-benzyl-4-C-(hydroxymethyl)-1,2-O-isopropylidene-α-D-ribofuranose via> subsequent benzylation, tosylation, and acetylation reactions in good yields. The compounds were confirmed by spectroscopic methods such as Fourier-transform infrared (FTIR) and proton nuclear magnetic resonance (1</sup>HNMR), and then evaluated for various pharmacological activities using standard in vitro >and in vivo> procedures.

 Compound 2a (50 mg/kg) exhibited both central and peripheral analgesic activity in the tail immersion test (2.52 ± 0.14 min tail flicking reaction time after 30 min from administration, P> < 0.001) and the acetic acid-induced writhing test (65.33 ± 2.06% reduction in abdominal writhing, P> < 0.001) respectively. In the anti-inflammatory assay, percent paw edema inhibition of carrageenan-induced rats for compounds 2a and 4 (100 mg/kg) after 4 h of administration were 82.6% (P> < 0.001) and 87.6% (P> < 0.001), respectively. The compounds were also tested for antioxidant activity in 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay, antimicrobial property in disk diffusion assay, and cytotoxicity in HeLa cell line; however, no significant results were observed in any of those tests.

 Our study indicated that some of the synthesized compounds exhibited promising analgesic and anti-inflammatory effects and may serve as potential lead compounds.

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a cytokine with a wide range of therapeutic applications although expression of GM-CSF in Escherichia coli (E. coli) usually leads to formation of insoluble aggregates mostly lack biological activity. The aim of this study was to compare the soluble expression level of GM-CSF in three E. coli strains, BL21 (DE3), SHuffle®</sup> T7 and OrigamiTM</sup> 2 (DE3).

The effect of different temperatures and inducer concentrations on soluble expression of GM-CSF was evaluated. The soluble GM-CSF was subjected to endotoxin removal and purification using nickel-nitrilotriacetic acid (Ni-NTA) affinity chromatography, ultrafiltration. The biological activity of produced GM-CSF was evaluated based on its growth promotion effect on TF-1 cell lines by MTT assay method.

A significant improvement of the soluble yield of GM-CSF (about 30% of GM-CSF was expressed as soluble proteins) was observed when protein expression was induced at 30 °C with 0.5 mM isopropyl β- d-1-thiogalactopyranoside (IPTG) in E. coli Shuffle T7. The soluble GM-CSF with a high purity up to 95 % and specific activity of 1.25 × 104</sup> IU/μg was obtained.

The proposed strategy here can be used to improve the soluble expression of other hard-to-express proteins with similar structural properties (i.e., containing disulfide binds or cysteine).  

Aberrant signaling by oncogenic RAS proteins occurs in almost all human tumors. One of the promising strategies to overcome such cancers is the inhibition of KRAS protein, a subtype of RAS family involved in cell growth, differentiation, and apoptosis, through preventing its effector, SOS1, from being attached to the protein.

Herein, a virtual screening process was performed using pharmacophore search, molecular docking, and molecular dynamic simulations. A pharmacophore model was created to indicate essential features for a KRAS inhibitor and used for screening the National Cancer Institution (NCI) database to retrieve similar compounds to the pharmacophore model with more than 70% similarity. Chosen compounds were then docked into KRAS and four compounds were selected based on the highest binding scores. Next, a similarity search was done in the whole PubChem database to increase the number of potential inhibitors. The filtered compounds were docked again into KRAS and three of them were selected for molecular dynamic simulation.

Compounds 1a, 2d, and 3a can inhibit SOS-iKRASG12D</sup> interaction due to the higher number of interactions with the protein. Moreover, they achieved the equilibrium faster than the approved inhibitor.

Auriculasin, a polyphenol flavonoid, can be considered as a potential inhibitor of SOS1-KRAS interaction. This compound seems to be a stronger anticancer than 9LI, a known inhibitor of KRAS, due to its better docking scores. Moreover, this compound can be an appropriate candidate to be formulated as an oral drug. 

 Neurobiological changes in memory processes seem to play a role in the pathophysiology of post-traumatic stress disorder (PTSD). Memory itself is influenced by PTSD, too. Histone deacetylase inhibitors (HDAIs) have shown promising results in the extinction of fear-related memories in animals and hence they seem to be important for the treatment of PTSD. Data are scarce about the effect of HDAIs in spatial memory formation/extinction in PTSD models. The main goal of the present work is to find the effect of sodium butyrate (NaBu), as an HDAI, on spatial memory and spatial memory extinction in rats exposed to single prolonged stress procedure (SPS).

Different doses of NaBu were administered subcutaneously for 7 days in different groups of rats after SPS procedure. Learning, memory, and extinction of memory were evaluated in the Morris water maze test of spatial memory in 6 consecutive days.

The results show that NaBu (0.5 mg/kg) alleviates impaired learning and memory in SPS rats. It also facilitates the extinction of newly formed memory in the animals.

 Our data suggest that the administration of HDAIs after a traumatic experience can prevent the aversive effects of SPS on spatial memory. It also reinforces the notion that extinction of spatial memory involves the same or similar brain circuitry that is involved in the extinction of fear memories in PTSD patients.

Cancer is the leading cause of death in today's world, therefore the efforts to achieve anticancer drugs with higher potency and fewer side effects have always been conducted by researchers in the field of pharmaceutical chemistry.Monastrol, a cytotoxic small molecule, from dihydropyrimidinone scaffold, is an inhibitor of the kinesin-5 protein. So, efforts to identify more derivatives of this molecule have been of interest.

Some of monastrol's analogs as Eg5 inhibitors with different substitution patterns were analyzed, synthesized, and their cytotoxic effects were evaluated on MCF-7 and HeLa cancerous cells in vitro</em> using the MTT assay. The structure-activity relationship (SAR) was studied in silico</em> by molecular docking.

Among all proposed structures, in ducking study, those with hydrophobic moieties on the C2-N3 region, those with a hydroxyl group on the phenyl on C4 position, and those with a carboxylic group on C5 were the best candidates. In vitro</em> studies, on the other side, emphasized that monastrol still was the most potent derivative. Another finding was the more moderate activity of synthesized compounds on the HeLa cell compared to the MCF-7 cell line. During different challenges for substitution at 5-position, some earlier reports around the dihydropyrimidinone reactions were questioned. It seems that the change at the position 5 is not merely accessible, as earlier reports claimed. Also, we could not achieve any better cell cytotoxicity by the larger group in the thiourea region or position 5; nonetheless, it seems that the introduction of a methylene group at this position could be beneficial.

The initial results of this study were valuable in terms of design and synthesis and will be useful for future investigations.  

Today, improving rheumatoid arthritis (RA) as a chronic inflammatory disease is attributed to the proper status of the gut microbiota. Although some supplements containing beneficial live microorganisms (probiotics) can reduce inflammation by altering the bacterial composition of the gut, there is limited information on the effect of synbiotic (probiotics mixed with prebiotics) supplements on RA. Therefore, this study aimed to evaluate the anti-inflammatory effects of a synbiotic supplement as an adjuvant therapy in rheumatic patients. Moreover, for the first time, it was attempted to investigate whether addition of a synbiotic (1000 mg/day) to the combination of methotrexate and prednisolone increases the effectiveness of these antirheumatic drugs.

 Eligible patients (186 subjects) were randomly divided into two groups. Both groups received their standard routine antirheumatic drugs, methotrexate and prednisolone. Moreover, the first group received a daily oral synbiotic supplement (1000 mg) for 3 months while the second group received a placebo. Various parameters indicating RA status were evaluated at baseline (time 0) and 3 months after the treatment.

The results showed the changes in the level of RA indicators, including tender joint count with a range of 0 to 28 joints, swollen joint count with a range of 0 to 28 joints, visual analog scale, erythrocyte sedimentation rate, CRP, and disease activity score based on 28 joints, after 3 months.

Overall, no significant differences in the measured parameters were observed between synbiotic and placebo groups probably due to the short duration of the treatment period, and it is suggested to extend the treatment period to six months.

Artemisia is one of the well-known herbal medicinal plants for antimicrobial, insecticidal, antioxidant, and antimalarial activities. The antiproliferative effects of dichloromethane extracts of Artemisia biennis (A. biennis) and A. ciniformis and the petroleum ether extract of A. ciniformis have been demonstrated previously on human cancerous cell lines. In the current study, further fractionation was carried out on the aforementioned extracts and their cytotoxic effects were evaluated on three human cancer cell lines; B16/F10, PC3, and MCF7. F1 to F16, F1’ to F11’, and F1” to F10” were resulted from the fractionation of dichloromethane extracts of A. biennis, A. ciniformis, and petroleum ether extract of A. ciniformis, respectively.

The cytotoxic effects of 16 (F1-F16), 11 (F1’-F11’) and 10 (F1”-F10”) fractions, on B16/F10, PC3, and MCF7 cell lines were assessed using resazurin to measure viability and propidium iodide staining (sub G1) and flow cytometry to detect apoptosis.

The results showed that, some fractions at 100 µg/mL decreased cell viability. F2” in B16/F10 cells, F2, F4-F6, F10’, F11’, and F2” in PC3 cells, and F10’, F11’, and F2” in MCF7 significantly decreased cell viability in a concentration-dependent manner (12.5-50 μg/mL). Among different fractions, F2” demonstrated the most potent cytotoxic effects on cancer cell lines (P < 0.001). All of the mentioned fractions (except F11’ on PC3 cells) increased the number of apoptotic cells and showed the cytotoxic effects on cancer cells compared with the control group.

A. biennis and A. ciniformis are suggested as the potential sources of cytotoxic phytochemicals. The probable presence of terpenoids, steroids, and alkaloids in the selected fractions is proposed based on the preliminary phytochemical study. 

A group of thiosemicarbazones were prepared and their structures were confirmed by spectroscopic methods such as IR and H-NMR, mass spectrometry and also analytical method like elemental analysis. The synthesized semicarbazones were then assessed for their inhibitory activity against bacterial strains including Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Staphylococcus epidermidis, Bacillus cereus, Salmonella species, Enterobacter faecalis, methicillin-resistant Staphylococcus aureus, and fungi such as Candida albicans and Aspergillus niger.

The schiff bases of isatin (2a-j) were prepared by a condensation reaction between thiosemicarbazide and substituted N-aryl isatins leading to the desired thiosemicarbazones with exquisite purity.

The results disclosed that all compounds have noticeable inhibitory activity. Compounds 2a, 2b, 2c, 2g, and 2h were among the most potent derivatives against Gram negative bacteria and fungi. Besides, the activity of theses compounds were tested against Mycobacterium bovis bacillus Calmette-Guerin (M. bovis BCG). The antimycobacterial activity indicated compounds 2e and 2j are highly active against M. bovis BCG (minimum inhibitory concentration < 3.9 µg/mL). Among fluorinated structures, compounds 2a and 2j showed the best activities against M. bovis BCG.

To sum up, amongst the 10 synthesized compounds, fluorinated derivatives exhibited remarkable activities against both gram negative strains and candida albicans microorganism. Therefore, they should be considered as a clue for further modifications in next investigations. Furthermore, inserting a small/medium size halogen atom with electron-withdrawing and lipophilic properties increases anti-salmonella activity of these compounds and moreover 2-halogenated semithiocarbazones presented promising antimycobacterial activity. 

 Aspartyl/asparaginyl β-hydroxylase (ASPH) is abundantly expressed in malignant neoplastic cells. The establishment of a human cell line overexpressing ASPH could provide the native-like recombinant protein needed for developing theranostic probes. In the process of transfection, the obtained cells normally contain a range of cells expressing the different levels of the target of interest. In this paper, we report on our simple innovative approach in the selection of best-transfected cells with the highest expression of ASPH using subclone selection, quantitative real-time polymerase chain reaction, and gradual increment of hygromycin concentration.

To achieve this goal, human embryonic kidney (HEK 293T) cells were transfected with an ASPH-bearing pcDNA3.1/Hygro(+) vector. During antibiotic selection, single accumulations of the resistant cells were separately cultured and the ASPH mRNA levels of each flask were evaluated. The best subclones were treated with a gradually increasing amount of hygromycin. The ASPH protein expression of the obtained cells was finally evaluated using flow cytometry and immunocytochemistry.

The results showed that different selected subclones expressed different levels of ASPH. Furthermore, the gradual increment of hygromycin (up to 400mg/mL) improved the expression of ASPH. The best relative fold change in mRNA levels was 57.59 ± 4.11. Approximately 90.2% of HEKASPH</sup> cells overexpressed ASPH on their surface.

The experiments indicated that we have successfully constructed and evaluated a recombinant human cell line overexpressing ASPH on the surface. Moreover, our innovative selection approach provided an effective procedure for enriching highly expressing recombinant cells. 

A genome-wide clustered regularly interspaced short palindromic repeatsassociated protein 9-based screen has revealed that the cell adhesion molecule matrix remodelling associated protein 8 (Mxra8) acts as an entry mediator for many alphaviruses including chikungunya virus. The first Xray crystal structure reported for Mxra8 a few months ago has a low-resolution of 3.49Å.

Homology modelling of Mxra8 protein was done employing the SWISS-MODEL and PRIME module of Maestro. To design novel Mxra8 inhibitors pharmacophore guided fragment-based drug design and structure-based virtual screening of Food and Drug Administration approved drug libraries were undertaken. Molecular docking and molecular dynamics (MD) simulations study were carried out to validate the findings.

The molecule H1a (dock score: -6.137, binding energy: -48.95 kcal/mol, and PHASE screen score: 1.528816) was identified as the best hit among the fragment-based designed ligands. Structurebased virtual screening suggested histamine, epinephrine, and capreomycin as potential hits which could be repurposed as Mxra8 inhibitor. MD simulations study suggested that only small molecules like histamine could be a potential inhibitor of Mxra8. H-bond interaction with Arg58 and Glu200 amino acid residues seems to be crucial for effective binding.

To the best of our knowledge, this is the first report on the design of novel inhibitors against Mxra8 protein to tackle the menace of alphaviruses infections. This design strategy could be used for structure-based drug design against other apo-proteins. This study also advances the application of in silico tools in the field of drug repurposing.