In Vitro-Transcribed mRNAs as a New Generation of Therapeutics in the Dawn of Twenty-First Century: Exploitation of Peptides as Carriers for Their Intracellular Delivery

In vitro-transcribed mRNAs (IVT-mRNAs) are easily and rapidly designed in vitro synthesized RNA molecules. After their intracellular delivery through an efficient delivery system, the host cell ribosomes will be recruited to translate and produce the corresponding desired proteins. Nowadays, about 20 years after the first report as for the use of IVT-mRNA, this technology has all the spotlight on it, due to the recently produced vaccines against SARS-CoV-2. All this enthusiasm around IVT-mRNA has pushed a wave of biotech companies to leverage this technology, raising significant investments annually. Thus, IVT-mRNA technology has gained an impressive dynamic, with multidimensional applications [as protein replacement therapy (PRT), cancer immunotherapy, vaccine production, production of antibodies, cytokines and growth factors, gene silencing, cellular reprogramming, and gene editing] with remarkable results. This chapter will emphasize the recent advances in IVT-mRNA delivery. A wide range of in vitro and in vivo transfection reagents have been shown to protect IVT-mRNA from degradation, to escape immunosurveillance and facilitate its intracellular delivery. IVT-mRNA delivery systems can be classified into two broad categories: (i) physical transfection methods, like electroporation, that temporarily disrupt cell membrane barrier function and (ii) chemically formulated nanocarriers, like polymer-based, lipid-based nanovectors, lipid–polymer hybrid nanoparticles, and peptide vectors. As the peptide-based delivery systems are gaining ground due to the flexibility that peptides can offer, this chapter will present this very interesting aspect that combines IVT-mRNA technology with protein transduction domain (PTD) technology. Compared to cationic polymers, the peptides are of low-molecular weight, with degradable amino acid sequences and distinct biological properties, such as cell permeability efficiency and cell and nuclear surface targeting. Either by non-covalent or covalent binding, peptide-based carriers and hybrids are suggested as interesting alternatives to the various existing non-viral vectors for IVT-mRNA delivery. © 2022, The Author(s), under exclusive license to Springer Nature Switzerland AG

Development of a novel PTD-mediated IVT-mRNA delivery platform for potential protein replacement therapy of metabolic/genetic disorders

The potential clinical applications of the powerful in vitro-transcribed (IVT)-mRNAs, to restore defective protein functions, strongly depend on their successful intracellular delivery and transient translation through the development of safe and efficient delivery platforms. In this study, an innovative (international patent-pending) methodology was developed, combining the IVT-mRNAs with the protein transduction domain (PTD) technology, as an efficient delivery platform. Based on the PTD technology, which enables the intracellular delivery of various cargoes intracellularly, successful conjugation of a PTD to the IVT-mRNAs was achieved and evaluated by band-shift assay and NMR spectroscopy. In addition, the PTD-IVT-mRNAs were applied and evaluated in two protein-disease models, including the mitochondrial disorder fatal infantile cardioencephalomyopathy and cytochrome c oxidase (COX) deficiency (attributed to SCO2 gene mutations) and β-thalassemia. The PTD-IVT-mRNA of SCO2 was successfully transduced and translated to the corresponding Sco2 protein inside the primary fibroblasts of a SCO2/COX-deficient patient, whereas the PTD-IVT-mRNA of β-globin was transduced and translated in bone marrow cells, derived from three β-thalassemic patients. The transducibility and the structural stability of the PDT-IVT-mRNAs, in both cases, were confirmed at the RNA and protein levels. We propose that our novel delivery platform could be clinically applicable as a protein therapy for metabolic/genetic disorders. © 2021 The Author

Synthesis, molecular structure determination, and antitumor activity of platinum(II) and palladium(II) complexes of 2-substituted benzimidazole

The complexes of 2-aminomethyl benzimidazole, 2-(β-aminoethyl)benzimidazole, and 2-(α-aminoethyl)benzimidazole with Pt(II) and Pd(II) have been prepared. The molecular structure of the free ligands and their complexes were studied by IR and 1H NMR. It was concluded that the substituted benzimidazole derivatives behave as bidentate ligands, being bound to the metal atoms via the nitrogen of the -N = group and the amino group of the side chain of the benzimidazole ring. The metal complexes were tested for antineoplastic activity both in cultures of neoplastic cells (MEL-745, K-562, Colon 205, IMP-32, SK-N-SH) and in vivo in rodents bearing L-1210 leukemia. The antiproliferative activity of these agents was compared to that of cis-platin. © 1988

Metabolomics assisted fingerprint of Hypericum perforatum chemotypes and assessment of their cytotoxic activity

Hypericum perforatum is known as an important medicinal plant, used for the treatment of several diseases, while its pharmacological properties are attributed to the presence of a wide range of secondary metabolites. Due to the great chemotypic variability of Hypericum species in the nature, and the demand for standardized herbal products, a detailed phytochemical investigation was carried out on different parts (herba, leaf, flowers) from wild collected and cultivated populations, using advanced chromatographic tools. Liquid Chromatographic analysis (LC-MS/MS MRM) revealed significant variability in the secondary metabolites content of the examined methanolic extracts. The most common derivatives belong to 9 groups i.e. benzoic acids, phenylpropanoids, coumarins, flavones, flavonols, flavan-3-ols, anthocyanins, phloroglucinols and naphtodianthrones. The main polyphenolic compounds were catechin, epicatechin, quercetin, quercetin 3-O-rhamnoside, quercetin 3-O-glucoside, neochlorogenic acid, proanthocyanidins (A and B series) and cyanidin-3-O-glucoside. In addition, the content of the characteristic compounds hypericin and hyperforin in herba crude extracts ranged between 0.5 and 1.7 mg/g and 0.6–3.3 mg/g respectively. The cytotoxic activity of the crude extracts was assessed at concentrations ranged between 0.01 and 100 μg/mL, on Caco-2 intestinal cancer cell cultures, and a cytotoxic behavior was shown only at the highest concentration of 100 μg/mL

Aiding in the Treatment of Low Back Pain by a Fuzzy Linguistic Web System

Abstract. Low back pain affects a large proportion of the adult popu-lation at some point in their lives and has a major economic and social impact. To soften this impact, one possible solution is to make use of rec-ommender systems, which have already been introduced in several health fields. In this paper, we present TPLUFIB-WEB, a novel fuzzy linguistic Web system that uses a recommender system to provide personalized exercises to patients with low back pain problems and to offer recom-mendations for their prevention. This system may be useful to reduce the economic impact of low back pain, help professionals to assist pa-tients, and inform users on low back pain prevention measures. A strong part of TPLUFIB-WEB is that it satisfies the Web quality standard