On the Far Side of Telomeres: The Many Roles of Telomerase in the Acquisition and Retention of Cancer Stemness

It is well recognised that upregulation/reactivation of telomerase, the telomere-lengthening enzyme, is the sine qua non of cellular immortalisation and malignant transformation. But there is also convincing evidence that telomerase stands at a crossroads where several developmental signalling pathways meet and that its upregulation/reactivation has effects beyond maintaining telomere length, such as altering energy metabolism and modulating gene expression. We believe that it is important to realise that, in a pathological context, such extratelomeric effects of telomerase are related to the emergence and persistence of the cancer stem cell (CSC) phenotype. Given the common conception of cancer stemness as a major contributor to therapy resistance and tumour relapse, a more complete annotation of biological mechanisms for its regulation by telomerase will provide the opportunity to develop telomerase-targeted anticancer therapies which kill or differentiate CSCs effectively

Graphene Oxide Nanosheets Interact and Interfere with SARS‐CoV‐2 Surface Proteins and Cell Receptors to Inhibit Infectivity

From Wiley via Jisc Publications RouterHistory: received 2021-03-13, pub-electronic 2021-05-14Article version: VoRPublication status: PublishedFunder: University of PaduaFunder: UKRI EPSRC; Grant(s): EP/P00119X/1Funder: Turkish Academy of Sciences (TUBA)Funder: Scientific and Technology Council of Turkey; Grant(s): 18AG020Funder: Türkiye Bilimler Akademisi; Id: http://dx.doi.org/10.13039/501100004412; Grant(s): GEBIP 2018Funder: Türkiye Bilimsel ve Teknolojik Araştirma Kurumu; Id: http://dx.doi.org/10.13039/501100004410; Grant(s): 18AG020Funder: Engineering and Physical Sciences Research Council; Id: http://dx.doi.org/10.13039/501100000266; Grant(s): EP/P00119X/1Abstract: Nanotechnology can offer a number of options against coronavirus disease 2019 (COVID‐19) acting both extracellularly and intracellularly to the host cells. Here, the aim is to explore graphene oxide (GO), the most studied 2D nanomaterial in biomedical applications, as a nanoscale platform for interaction with SARS‐CoV‐2. Molecular docking analyses of GO sheets on interaction with three different structures: SARS‐CoV‐2 viral spike (open state – 6VYB or closed state – 6VXX), ACE2 (1R42), and the ACE2‐bound spike complex (6M0J) are performed. GO shows high affinity for the surface of all three structures (6M0J, 6VYB and 6VXX). When binding affinities and involved bonding types are compared, GO interacts more strongly with the spike or ACE2, compared to 6M0J. Infection experiments using infectious viral particles from four different clades as classified by Global Initiative on Sharing all Influenza Data (GISAID), are performed for validation purposes. Thin, biological‐grade GO nanoscale (few hundred nanometers in lateral dimension) sheets are able to significantly reduce copies for three different viral clades. This data has demonstrated that GO sheets have the capacity to interact with SARS‐CoV‐2 surface components and disrupt infectivity even in the presence of any mutations on the viral spike. GO nanosheets are proposed to be further explored as a nanoscale platform for development of antiviral strategies against COVID‐19

Cancer cell lines involving cancer stem cell populations respond to oxidative stress

Cancer cells may be more prone to the accumulation of reactive oxygen species (ROS) than normal cells; therefore increased oxidative stress can specifically kill cancer cells including cancer stem cells (CSCs). In order to generate oxidative stress in various cancer cell lines including A549, G361 and MCF-7, cultured cells were exposed to H2O2. Incubation of cancer cells with H2O2 results in concentration-dependent cell death in A549 and G361-7 cells, whereas MCF-7 cells showed higher sensitivity even at a lower H2O2 concentration. H2O2 treatment decreased the number of cells in G2/M phase and increased the number of apoptotic cells. Both CD24 negative/CD44 positive cells and CD146 positive cells were found to be present in all tested cancer cell lines, indicating that CSC populations may play role in the cellular response to oxidative stress. This study showed that inducing oxidative stress through ROS can offer a promising approach for anti-cancer therapy

Reasons for success and lessons learnt from nanoscale vaccines against COVID-19.

From PubMed via Jisc Publications RouterPublication status: ppublis

Development of Dual-Activity Vectors by Co-Envelopment of Adenovirus and SiRNA in Artificial Lipid Bilayers.

Gene therapy with human adenovirus type 5 (Ad5) has been extensively explored for the treatment of diseases resistant to traditional therapies. Intravenous administration leads to rapid clearance from blood circulation and high liver accumulation, which restrict the use of Ad-based vectors in clinical gene therapy protocols that involve systemic administration. We have previously proposed that such limitations can be improved by engineering artificial lipid envelopes around Ad and designed a variety of artificial lipid bilayer envelopes around the viral capsid. In this study, we sought to explore further opportunities that the artificially enveloped virus constructs could offer, by designing a previously unreported gene therapy vector by simultaneous envelopment of Ad and siRNA within the same lipid bilayer. Such a dual-activity vector can offer efficacious therapy for different genetic disorders where both turning on and switching off genes would be needed. Dynamic light scattering, transmission electron microscopy and atomic force microscopy were used to characterize these vectors. Agarose gel electrophoresis, Ribo green and dot blot assays showed that siRNA and Ad virions can be enveloped together within lipid bilayers at high envelopment efficiency. Cellular uptake and in vitro transfection experiments were carried out to show the feasibility of combining siRNA-mediated gene silencing with viral gene transfer using these newly designed dual-activity vectors

Characterization of co-enveloped Ad-siRNA vectors by DLS, TEM and AFM.

<p>The naked Ad, enveloped Ad, co-enveloped Ad-siRNA and enveloped siRNA are the vectors examined in this study. (a) The mean average diameter (nm), polydispersity index and surface charge (mV) for each vector were obtained by DLS. (b) Samples were analyzed by TEM imaging. Scale bars represent 100 nm. Red arrows indicate enveloped Ad vectors. (c) Samples were analyzed by AFM imaging.</p

In vivo reprogramming of adult somatic cells to pluripotency by overexpression of yamanaka factors

Induced pluripotent stem (iPS) cells that result from the reprogramming of somatic cells to a pluripotent state by forced expression of defined factors are offering new opportunities for regenerative medicine. Such clinical applications of iPS cells have been limited so far, mainly due to the poor efficiency of the existing reprogramming methodologies and the risk of the generated iPS cells to form tumors upon implantation. We hypothesized that the reprogramming of somatic cells towards pluripotency could be achieved in vivo by gene transfer of reprogramming factors. In order to efficiently reprogram cells in vivo, high levels of the Yamanaka (OKSM) transcription factors need to be expressed at the target tissue. This can be achieved by using different viral or nonviral gene vectors depending on the target tissue. In this particular study, hydrodynamic tail-vein (HTV) injection of plasmid DNA was used to deliver the OKSM factors to mouse hepatocytes. This provided proof-of-evidence of in vivo reprogramming of adult, somatic cells towards a pluripotent state with high efficiency and fast kinetics. Furthermore no tumor or teratoma formation was observed in situ. It can be concluded that reprogramming somatic cells in vivo may offer a potential approach to induce enhanced pluripotency rapidly, efficiently, and safely compared to in vitro performed protocols and can be applied to different tissue types in the future

The comparison of dual-activity vectors with other gene delivery systems.

<p>(a) Dual-activity vectors were compared with different systems in terms of transgene expression and downregulation. (b) A549-luc-A9 cells were transfected with these vectors and after 24 h, cells were lysed. Luciferase assay was performed in order to measure the percentage of luciferase expression. (c) β-Gal expression was assessed by β-Gal assay. * p<0.05 versus naive.</p

“on and off” gene expression <i>in vitro</i> with Ad-GFP transfection and siGFP silencing.

<p>(a) Schematic representation of two-vector treatment protocols. For the two-vector approach <i>in vitro</i> cell transfections, there are three methods: pre-, co- and post- silencing. In pre-silencing, cells are treated firstly with siRNAs complexed to DOTAP:Chol liposomes and after 24 h Ads are added to the cells. In co-silencing protocol, cells are co-incubated with liposome-siRNA complexes and Ad. For post –silencing, cells are treated with liposome-siRNA complexes 24 h after Ad transfection. Cells are lysed after the last treatment in each silencing group and analyzed for gene expression. (b) C33-a cells were subjected to Ad-CMV-GFP (108 particles/mL) transfection and DOTAP:Chol liposomes complexed to siRNA (L-siGFP or L-siNeg, at 60 nM concentration) treatments at different time points as described. The cells were lysed 24 h after the last treatment and fluorescence per mg protein was measured to plot GFP expression as percentage of Ad transfected group. (c) Cell lysates were also run on SDS-PAGE electrophoresis and blotted to reveal the differences in protein levels in each treatment group. * p<0.05, ** p<0.01, *** p<0.001 versus Ad.</p