Targeting ATP2B1 impairs PI3K/Akt/Fox-O3 signaling and reduces SARS-COV-2 replication in vivo.

ATP2B1 is a known regulator of calcium (Ca2+) cellular export and homeostasis. Diminished levels of extra- or intra-cellular Ca2+ content have been suggested to block SARS-CoV-2 replication. Here, we demonstrate that a newly nontoxic caloxin-derivative compound (PI-7) inhibits ATP2B1, reduces the extra- and intra-cellular Ca2+ levels and impairs SARS-CoV-2 replication and propagation (VOCs: Delta and Omicron 2), as also measured by inhibition of syncytia in vitro. Furthermore, a FOXO3 transcriptional site of regulation of expression at the 5 end of the ATP2B1 locus, together with a rare homozygous intronic variant in the ATP2B1 locus (rs11337717; chr12:89643729, T>C), are shown to be associated with severity of COVID19 (symptomatic versus asymptomatic patients). Here, we identify the mechanism of action during SARS-CoV-2 infection, which involves the PI3K/Akt signaling pathway, inactivation of FOXO3 (i.e., phosphorylation), and inhibition of transcriptional control of both membrane and reticulum Ca2+ pumps (ATP2B1 and ATP2A1 [i.e., SERCA1], respectively). The pharmacological action of compound PI-7 on sustaining both ATP2B1 and ATP2A1 expression reduces the intracellular cytoplasmic Ca2+ pool and thus negatively influences SARS-CoV-2 replication and propagation. As compound PI-7 shows a lack of toxicity, its prophylactic use as a therapy against the COVID19 pandemic is here proposed. In briefDe Antonellis et al. shows the importance of the Ca2+ channel pump ATP2B1 in the regulation of extracellular and intracellular Ca2+ levels that positively influence SARS-CoV-2 replication in human cells. Our study identifies the mechanism of action of SARS-CoV-2 in the regulation of the expression of ATP2B1 and ATP2A1 loci during infection via FOXO3 transcriptional factor. Furthermore, a small caloxin-derivative molecule (compound PI-7) can inhibit ATP2B1 activity, thus resulting in SARS-CoV-2 impairment. In further support, we have identified a genetic variant within the noncoding upstream region of ATP2B1 in symtomatic patients affected by severe COVID19, thus indicating this polymorphism as a genetic predisposition factor to SARS-CoV-2 infection. HighlightsO_LIAn anti-viral model of network of action for ATP2B1 against SARS-CoV-2 at the intracellular level that involves the PI3K/Akt signaling pathway, inactivation (i.e., phosphorylation) of FOXO3 and its transcriptional control, and inhibition of both membrane and reticulum Ca2+ pumps (i.e., ATP2B1, ATP2A1, respectively). C_LIO_LIA new drug and its lack of toxicity "compound PI-7", thus envisioning both preventive and therapeutic applications in patients with COVID-19. C_LIO_LIThe specificity of action in the context of Ca2+ homeostasis is one of the strategies that coronaviruses (including SARS-CoV-2 and any new VOC, including Omicron 2) use to infect host cells and promote organ dysfunction. C_LIO_LITherapeutic applications for compound PI-7 against all other viruses belonging to the Coronoviridae family (e.g., SARS-CoV, MERS-CoV), and against the main families of positive sense ssRNA viruses from other hosts (e.g., Nidovirales), as these are all Ca2+ dependent. C_LIO_LIIdentification of a rare homozygous intronic variant in the ATP2B1 locus (rs11337717; chr12:89643729, T>C) that is associated with severity of COVID19 (i.e., symptomatic versus asymptomatic patients). This variant can be used as a marker to identify those patients that might show severe COVID19 following their SARS-COV-2 infection. C_LI

Non-covalent small molecule partnership for redox-active films: Beyond polydopamine technology.

HYPOTHESIS: The possibility to use hexamethylenediamine (HMDA) to impart film forming ability to natural polymers including eumelanins and plant polyphenols endowed with biological activity and functional properties has been recently explored with the aim to broaden the potential of polydopamine (PDA)-based films overcoming their inherent limitations. 5,6-dihydroxyindole-2-carboxylic acid, its methyl ester (MeDHICA) and eumelanins thereof were shown to exhibit potent reducing activity. EXPERIMENTS: MeDHICA and HMDA were reacted in aqueous buffer, pH 9.0 in the presence of different substrates to assess the film forming ability. The effect of different reaction parameters (pH, diamine chain length) on film formation was investigated. Voltammetric and AFM /SEM methods were applied for analysis of the film redox activity and morphology. HPLC, MALDI-MS and 1HNMR were used for chemical characterization. The film reducing activity was evaluated in comparison with PDA by chemical assays and using UV stressed human immortalized keratinocytes (HaCat) cells model. FINDINGS: Regular and homogeneous yellowish films were obtained with moderately hydrophobic properties. Film deposition was optimal at pH 9, and specifically induced by HMDA. The film consisted of HMDA and monomeric MeDHICA accompanied by dimers/small oligomers, but no detectable MeDHICA/HMDA covalent conjugation products. Spontaneous assembly of self-organized networks held together mainly by electrostatic interactions of MeDHICA in the anion form and HMDA as the dication is proposed as film deposition mechanism. The film displayed potent reducing properties and exerted significant protective effects from oxidative stress on HaCaT.

Arsenoplatin-Ferritin Nanocage: Structure and Cytotoxicity.

Arsenoplatin-1 (AP-1), the prototype of a novel class of metallodrugs containing a PtAs(OH)2 core, was encapsulated within the apoferritin (AFt) nanocage. UV-Vis absorption spectroscopy and inductively coupled plasma-atomic emission spectroscopy measurements confirmed metallodrug encapsulation and allowed us to determine the average amount of AP-1 trapped inside the cage. The X-ray structure of AP-1-encapsulated AFt was solved at 1.50 Å. Diffraction data revealed that an AP-1 fragment coordinates the side chain of a His residue. The biological activity of AP-1-loaded AFt was comparatively tested on a few representative cancer and non-cancer cell lines. Even though the presence of the cage reduces the overall cytotoxicity of AP-1, it improves its selectivity towards cancer cells.

Modified denatured lysozyme effectively solubilizes fullerene c60 nanoparticles in water.

Fullerenes, allotropic forms of carbon, have very interesting pharmacological effects and engineering applications. However, a very low solubility both in organic solvents and water hinders their use. Fullerene C60, the most studied among fullerenes, can be dissolved in water only in the form of nanoparticles of variable dimensions and limited stability. Here the effect on the production of C60 nanoparticles by a native and denatured hen egg white lysozyme, a highly basic protein, has been systematically studied. In order to obtain a denatured, yet soluble, lysozyme derivative, the four disulfides of the native protein were reduced and exposed cysteines were alkylated by 3-bromopropylamine, thus introducing eight additional positive charges. The C60 solubilizing properties of the modified denatured lysozyme proved to be superior to those of the native protein, allowing the preparation of biocompatible highly homogeneous and stable C60 nanoparticles using lower amounts of protein, as demonstrated by dynamic light scattering, transmission electron microscopy and atomic force microscopy studies. This lysozyme derivative could represent an effective tool for the solubilization of other carbon allotropes.