Structural Modification of the Macrolide Brefeldin A to Analogues with Enhanced Cytotoxicity against KB Cells

The macrolide brefeldin A (BFA, 1) exhibited high cytotoxicity against KB cells. However, it was also toxic against non-cancerous cells. In order to lower toxicity against normal cells while maintaining the cytotoxic potency to the cancer cells, structural modification of this compound was undertaken. Starting from compound 1, the analogues 2-13 were synthesized and evaluated for cytotoxicity against KB cells. The analogue 2 exhibited the most potent cytotoxicity against KB cells, with an IC50 value of 0.034 nM, 67-fold more active than its parent compound 1. It was 41764 and 8235 fold more active than the standard drugs ellipticine and doxorubicin, respectively. The higher cytotoxicity against KB cells and lower toxicity against Vero cells of analogue 2 than those of the parent compound 1 contributed to its exceptionally high selectivity index of 9117. The results suggested that this analogue might be utilized to develop a new candidate for an anticancer drug. HIGHLIGHTS Oral cancer is a major worldwide public health issue and may affect any oral cavity region, including the lips, tongue, mouth and throat The ester analogues (2-13) were the first report of the cytotoxic against human epidermoid carcinoma (KB) cells 7-O-acetyl BFA (2) exhibited the most potent cytotoxicity against KB cells, with an IC50 value of 034 nM, 67-fold more active than its parent compound 1 The essentially structural features for the macrolide of BFA-type to exhibit high cytotoxicity against KB cells are the presence of a free hydroxyl group at the 4-position, a free hydroxyl group or the corresponding ester at the 7-position, and unsaturated functions at the 2- and 10-positions GRAPHICAL ABSTRACT

Potent Anticancer Activities of Beauvericin against KB cells In Vitro by Inhibiting the Expression of ACAT1 and Exploring Binding Affinity

Background and Objective: Beauvericin (BEA), a cyclic hexadepsipeptide mycotoxin, is a potent inhibitor of the acyl-CoA: cholesterol acyltransferase enzyme 1 (ACAT1) which involved in multiple tumor-correlated pathways. However, the binding mechanisms between BEA and ACAT1 were not elucidated. Methods: BEA was purified from a mangrove entophytic Fusarium sp. KL11. Single-crystal X-ray diffraction was used to determine the structure of BEA. Wound healing assays of BEA against KB cell line and MDA-MB-231 cell line were evaluated. Inhibitory potency of BEA against ACAT1 was determined by ELISA assays. Molecular docking was carried out to illuminate the bonding mechanism between BEA and ACAT1. Results: The structure of BEA was confirmed by X-ray diffraction, indicating a monoclinic crystal system with P21 space group (α = 90°, β = 92.2216(9)o, γ= 90o). BEA displayed migration-inhibitory activities against KB cells and MDA-MB-231 cells in vitro. ELISA assays revealed the protein expression level of ACAT1 in KB cells was significantly decreased after BEA treatment (P ＜0.05). Molecular docking demonstrated that BEA formed hydrogen bond with His425 and pi-pi staking with Tyr429 in ACAT1. Conclusions: BEA sufficiently inhibited the proliferation and migration of KB cells and MDA-MB-231 cells by downregulating ACAT1 expression. In addition, BEA potentially possessed a strong binding affinity with ACAT1. BEA may serve as a potential lead compound for the development of a new ACAT1-targeted anticancer drug.

Antimalarial activities of ( Z )-2-(nitroheteroarylmethylene)-3(2 H )-benzofuranone derivatives: In vitro , in vivo assessment and β-hematin formation inhibition activity

A series of ( Z )-2-(nitroheteroarylmethylene)-3(2 H )-benzofuranones possessing nitroheteroaryls groups of nitroimidazole, nitrofuran and nitrothiophene moieties was screened for antiplasmodium activity against drug-sensitive (3D7) as well as a chloroquine (CQ) and multi-drug resistant (K1) strains of P. falciparum 5-Nitroimidazole and 4-nitroimidazole analogs were highly selective and active against resistant parasites, while 5-nitrofuran and 5-nitrothiophene derivatives were more potent against the 3D7 strain than the K1 strain. Among the synthetic analogues, ( Z )-6-chloro-2-(1-methyl-5-nitroimidazol-2-ylmethylene)-3(2 H )-benzofuranone ( 5h ) exhibited the highest activity (IC 50 : 0.654 nM) against K1 strain and ( Z )-7-methoxy-2-(5-nitrothiophen-2-ylmethylene)-3(2 H )-benzofuranone ( 10g ) showed the highest activity (IC 50 : 0.28 μM) against the 3D7 strain in comparison with CQ (IC 50 s of 3.13 and 206.3 nM against 3D7 and K1 strains, respectively). The more active compounds with IC 50 s lower than 5 μg/mL (∼20 μM) were further studied for their cytotoxicity responses using KB cells. From these studies, 5-nitroimidazole, 4-nitroimidazole and 5-nitrofuran analogues were shown to be cytotoxic against KB cells, while 5-nitrothiophene analogues were shown to have the least cytotoxic effects. To gain some insight into their potential contributing mechanism of action, three derivatives 10e , 10g and 10h (from nitrothiophene subgroup possessing 6-methoxy, 7-methoxy and 6,7-dimethoxy substituents on their benzofuranone moieties, respectively) showing the least toxicity and highest selectivity indices were assessed for their β-hematin formation inhibition activity. 10g demonstrated the highest inhibition activity (IC 50 : 10.78 μM) in comparison with CQ (IC 50 : 2.63 μM) as the reference drug. Finally, these three analogues ( 10e , 10g and 10h ) were further evaluated for their in vivo activity against P. berghei /albino mice model (Peter’s test). Tested analogues were shown to be active, reducing the percentage of erythrocytes that contained parasites by 53.4, 48.8 and 32.4%, respectively.

Dendrimer-Coated Gold Nanoparticles for Efficient Folate-Targeted mRNA Delivery In Vitro

Messenger RNA (mRNA) is not an attractive candidate for gene therapy due to its instability and has therefore received little attention. Recent studies show the advantage of mRNA over DNA, especially in cancer immunotherapy and vaccine development. This study aimed to formulate folic-acid-(FA)-modified, poly-amidoamine-generation-5 (PAMAM G5D)-grafted gold nanoparticles (AuNPs) and to evaluate their cytotoxicity and transgene expression using the luciferase reporter gene (FLuc-mRNA) in vitro. Nanocomplexes were spherical and of favorable size. Nanocomplexes at optimum nanoparticle:mRNA (w/w) binding ratios showed good protection of the bound mRNA against nucleases and were well tolerated in all cell lines. Transgene expression was significantly (p < 0.0001) higher with FA-targeted, dendrimer-grafted AuNPs (Au:G5D:FA) in FA receptors overexpressing MCF-7 and KB cells compared to the G5D and G5D:FA NPs, decreasing significantly (p < 0.01) in the presence of excess competing FA ligand, which confirmed nanocomplex uptake via receptor mediation. Overall, transgene expression of the Au:G5D and Au:G5D:FA nanocomplexes exceeded that of G5D and G5D:FA nanocomplexes, indicating the pivotal role played by the inclusion of the AuNP delivery system. The favorable properties imparted by the AuNPs potentiated an increased level of luciferase gene expression.

Immunofluorescence Analysis of NF-kB and iNOS Expression in Different Cell Populations during Early and Advanced Knee Osteoarthritis

Synovitis of the knee synovium is proven to be a precursor of knee osteoarthritis (OA), leading to a radiologically advanced stage of the disease. This study was conducted to elucidate the expression pattern of different inflammatory factors—NF-kB, iNOS, and MMP-9 in a subpopulation of synovial cells. Thirty synovial membrane intra-operative biopsies of patients (ten controls, ten with early OA, and ten with advanced OA, according to the Kellgren–Lawrence radiological score) were immunohistochemically stained for NF-kB, iNOS, and MMP9, and for different cell markers for macrophages, fibroblasts, leukocytes, lymphocytes, blood vessel endothelial cells, and blood vessel smooth muscle cells. The total number of CD68+/NF-kB+ cells/mm2 in the intima of early OA patients (median = 2359) was significantly higher compared to the total number of vimentin+/Nf-kB+ cells/mm2 (median = 1321) and LCA+/NF-kB+ cells/mm2 (median = 64) (p < 0.001 and p < 0.0001, respectively). The total number of LCA+/NF-kB+ cells/mm2 in the subintima of advanced OA patients (median = 2123) was significantly higher compared to the total number of vimentin+/NF-kB+ cells/mm2 (median = 14) and CD68+/NF-kB+ cells/mm2 (median = 29) (p < 0.0001). The total number of CD68+/iNOS+ cells/mm2 in the intima of both early and advanced OA patients was significantly higher compared to the total number of vimentin+/iNOS+ cells/mm2 and LCA+/iNOS+ cells/mm2 (p < 0.0001 and p < 0.001, respectively). The total number of CD68+/MMP-9+ cells/mm2 in the intima of both early and advanced OA patients was significantly higher compared to the total number of vimentin+/MMP-9+ cells/mm2 and CD5+/MMP-9+ cells/mm2 (p < 0.0001). Macrophages may have a leading role in OA progression through the NF-kB production of inflammatory factors (iNOS and MMP-9) in the intima, except in advanced OA, where leukocytes could have a dominant role through NF-kB production in subintima. The blocking of macrophageal and leukocyte NF-kB expression is a possible therapeutic target as a disease modifying drug.

Synthesis and Characterization of Radiogallium-Labeled Cationic Amphiphilic Peptides as Tumor Imaging Agents

SVS-1 is a cationic amphiphilic peptide (CAP) that exhibits a preferential cytotoxicity towards cancer cells over normal cells. In this study, we developed radiogallium-labeled SVS-1 (67Ga-NOTA-KV6), as well as two SVS-1 derivatives, with the repeating KV residues replaced by RV or HV (67Ga-NOTA-RV6 and 67Ga-NOTA-HV6). All three peptides showed high accumulation in epidermoid carcinoma KB cells (53–143% uptake/mg protein). Though 67Ga-NOTA-RV6 showed the highest uptake among the three CAPs, its uptake in 3T3-L1 fibroblasts was just as high, indicating a low selectivity. In contrast, the uptake of 67Ga-NOTA-KV6 and 67Ga-NOTA-HV6 into 3T3-L1 cells was significantly lower than that in KB cells. An endocytosis inhibition study suggested that the three 67Ga-NOTA-CAPs follow distinct pathways for internalization. In the biodistribution study, the tumor uptakes were found to be 4.46%, 4.76%, and 3.18% injected dose/g of tissue (% ID/g) for 67Ga-NOTA-KV6, 67Ga-NOTA-RV6, and 67Ga-NOTA-HV6, respectively, 30 min after administration. Though the radioactivity of these peptides in tumor tissue decreased gradually, 67Ga-NOTA-KV6, 67Ga-NOTA-RV6, and 67Ga-NOTA-HV6 reached high tumor/blood ratios (7.7, 8.0, and 3.8, respectively) and tumor/muscle ratios (5.0, 3.3, and 4.0, respectively) 120 min after administration. 67Ga-NOTA-HV6 showed a lower tumor uptake than the two other tracers, but it exhibited very low levels of uptake into peripheral organs. Overall, the replacement of lysine in SVS-1 with other basic amino acids significantly influenced its binding and internalization into cancer cells, as well as its in vivo pharmacokinetic profile. The high accessibility of these peptides to tumors and their ability to target the surface membranes of cancer cells make radiolabeled CAPs excellent candidates for use in tumor theranostics.

Fucoxanthin induced apoptotic cell death in oral squamous carcinoma (KB) cells

Fucoxanthin (Fx) is an active compound commonly found in the many types of seaweed with numerous biological activities. The main goal of this investigation is to explore the effect of Fx against the cell proliferation, apoptotic induction and oxidative stress in the oral squamous (KB) cell line. Cytotoxicity of Fx was determined by MTT assay. The intracellular ROS production, mitochondrial membrane potential (MMP) and apoptosis induction in KB cells were examined through DCFH-DA, Rhodamine-123 and DAPI, and dual staining techniques. Effect of Fx on the antioxidant enzymes and lipid peroxidation in the KB cells was studied through the standard procedures. Fx treated KB cells showed morphological changes and reduced cell survival, which is exhibited by the cytotoxic activity of 50 μM/ml (IC50) Fx against the KB cells. The Fx treatment considerably induced the apoptotosis cells (EB/AO) and decreased the MMP (Rh-123) in KB cells. Further, it was pointed out that there was an increased lipid peroxidation (LPO) with decreased antioxidants (CAT, SOD and GSH). These results concluded that Fx has the cytotoxic effect against KB cells and has the potential to induce the apoptosis via increased oxidative stress. Hence, the Fx can be a promising agent for the treatment of oral cancer and it may lead to the development of cancer therapeutics.

Anticancer potential of sclareol against human KB oral carcinoma cell line invitro

Sclareol, a labdane diterpene phytochemical in the various genus Salvia of the herbaceous plant has been anti-carcinogenic properties on numerous human cancer cell lines. Till known the anticancer properties of sclareol on human oral KB cancer cells are not revealed. Therefore, we scrutinized the viability, nucleate damage associated with apoptosis in KB cells treated with sclareol.The cytotoxicity of KB cells displayed to sclareol was evaluated by hexosaminidase and trypan blue exclusion assay. Apoptosis was noticed by Hoechst staining and affirmed by nucleate damage. We discovered that sclareol treatment for KB cells with expressively lessened cell proliferation or viability and evoked cell death in a dose-dependent manner compared with untreated control. Sclareol stimulated inhibition of growing of oral KB cells came along to be induction nucleate damage. Our data proved that sclareol on stimulated apoptosis in human oral KB cancer cells through nucleate damage. These observations recommend the anti-cancer properties of sclareol.

Nanoparticles decorated with folate based on a site-selective αCD-rotaxanated PEG-b-PCL copolymer for targeted cancer therapy

NPs fabricated from a mixture of PEG-b-PCL and selectively rotaxanated Fol-PEG(αCD)-PCL showed internalisation in KB cells through an active targeting mechanism.