Phytochemicals and micronutrients in suppressing infectivity caused by SARS-CoV-2 virions and seasonal coronavirus HCoV-229E in vivo.

SARS-CoV-2 infection still poses health threats especially to older and immunocompromised individuals. New emerging variants of SARS-CoV-2, including Omicron and Arcturus, have been challenging the effectiveness of humoral immunity resulting from repeated vaccination and infection. With recent study implying a wave of new mutants in vaccinated people making them more susceptible to the newest variants and fueling a rapid viral evolution, there is a need for alternative or adjunct approaches against coronavirus infections other than vaccines. Our earlier work indicated that a specific combination of micronutrients and phytochemicals can inhibit key infection mechanisms shared by SARS-CoV-2 and its variants in vitro. Here we demonstrate in vivo that an intake of this micronutrient combination before and during infection of mice with engineered SARS-CoV-2 virions and HCoV-229E virus results in a significant decrease in viral load and level of spike protein in the lungs. This was accompanied by decreased inflammatory response, including TNFα, IL1ß, ILα, and IL17. These and our earlier results confirm that by targeting multiple mechanisms simultaneously by a combination treatment we can effectively and safely challenge SARS-CoV-2 and HCoV-229E virus. If clinically confirmed, such an approach could complement already in-use preventive and therapeutic strategies against coronavirus infections.

Linoleic acid binds to SARS-CoV-2 RdRp and represses replication of seasonal human coronavirus OC43.

Fatty acids belong to a group of compounds already acknowledged for their broad antiviral efficacy. However, little is yet known about their effect on replication of human coronaviruses. To shed light on this subject, we first screened 15 fatty acids, three lipid-soluble vitamins, and cholesterol, on SARS-CoV-2 RdRp, and identified the four fatty acids with the highest RdRp inhibitory potential. Among them, linoleic acid was found to have the greatest interaction with SARS-CoV-2 RdRp, with its direct binding to the cavity formed by the RNA double helix and protein. Linoleic acid forms hydrophobic interactions with multiple residues, and at the same time forms electrostatic interactions including the hydrogen bond with Lys593 and Asp865. In line with these results, a dose-dependent inhibition of HCoV-OC43 replication in vitro was observed, additionally strengthened by data from in vivo study, which also confirmed anti-inflammatory potential of linoleic acid. Based on these results, we concluded that our study provides a new understanding of the antiviral properties of fatty acids against human coronaviruses including the SARS-CoV-2 strain. Particularly, they lays down a new prospect for linoleic acid's RdRp-inhibitory activity, as a candidate for further studies, which are warranted to corroborate the results presented here.

Hereditary Breast and Ovarian Cancer Service in Sparsely Populated Western Pomerania.

The German Consortium Hereditary Breast and Ovarian Cancer (GC-HBOC) consists of 23 academic centers striving to provide high-quality regional care for affected individuals and healthy at-risk family members. According to the standard operating procedures defined by the GC-HBOC, a Familial Breast and Ovarian Cancer Center was implemented at the University Medicine Greifswald over a four-year period from 2018 to 2021, despite the COVID-19 pandemic. Genetic analyses were performed in a total of 658 individuals, including 41 males, which paved the way to local annual risk-adapted breast cancer surveillance for 91 women and prophylactic surgery for 34 women in 2021. Our experience in the North Eastern part of Germany demonstrates that it is possible to establish a high-risk breast and ovarian cancer service even in a sparsely populated region. Major facilitators are the interdisciplinary collaboration of dedicated local experts, the support of the GC-HBOC, fruitful clinical and scientific cooperations and the use of technical improvements. As a blueprint, our project report may help to further expand the network of specialized and knowledge-generating care for HBOC families.

Composition of naturally occurring compounds decreases activity of Omicron and SARS-CoV-2 RdRp complex.

Naturally-occurring compounds are acknowledged for their broad antiviral efficacy. Little is however known about their mutual cooperation. Here, we evaluated in vitro efficacy of the defined mixture of agents against the RdRp complex of the original SARS-CoV-2 and Omicron variant. This composition of vitamin C, N-acetylcysteine, resveratrol, theaflavin, curcumin, quercetin, naringenin, baicalin, and broccoli extract showed to inhibit activity of RdRp/nsp7/nsp8 both these variants. In vitro exposure of recombinant RdRp complex to individual compounds of this composition pointed to quercetin as the driving inhibitory compound. The outcome of this study supports the motion of antiviral efficacy of natural compounds against SARS-CoV-2 and Omicron and implies that their reciprocal or mutual interaction may augment antiviral action through simultaneous effect on different mechanisms. Consequently, this makes it more difficult for an infectious agent to evade all these mechanisms at the same time. Considering the urgency in finding effective prevention, but also side-effects free treatment of COVID-19 our results call for clinical affirmation of the benefits of this micronutrient combination in both preventive and therapeutic aspects. Whether observed effects can be achieved, by concentrations of the active agents used in these in vitro experiments, in in vivo or clinical setting warrants further study.

Inhibitory effects of specific combination of natural compounds against SARS-CoV-2 and its Alpha, Beta, Gamma, Delta, Kappa, and Mu variants.

Despite vaccine availability, the global spread of COVID-19 continues, largely facilitated by emerging SARS-CoV-2 mutations. Our earlier research documented that a specific combination of plant-derived compounds can inhibit SARS-CoV-2 binding to its ACE2 receptor and controlling key cellular mechanisms of viral infectivity. In this study, we evaluated the efficacy of a defined mixture of plant extracts and micronutrients against original SARS-CoV-2 and its Alpha, Beta, Gamma, Delta, Kappa, and Mu variants. The composition containing vitamin C, N-acetylcysteine, resveratrol, theaflavin, curcumin, quercetin, naringenin, baicalin, and broccoli extract demonstrated a highest efficacy by inhibiting the receptor-binding domain (RBD) binding of SARS-CoV-2 to its cellular ACE2 receptor by 90%. In vitro exposure of test pseudo-typed variants to this formula for 1 h before or simultaneously administrated to human pulmonary cells resulted in up to 60% inhibition in their cellular entry. Additionally, this composition significantly inhibited other cellular mechanisms of viral infectivity, including the activity of viral RdRp, furin, and cathepsin L. These findings demonstrate the efficacy of natural compounds against SARS-CoV-2 including its mutated forms through pleiotropic mechanisms. Our results imply that simultaneous inhibition of multiple mechanisms of viral infection of host cells could be an effective strategy to prevent SARS-CoV-2 infection.

Phenolic compounds disrupt spike-mediated receptor-binding and entry of SARS-CoV-2 pseudo-virions.

In the pursuit of suitable and effective solutions to SARS-CoV-2 infection, we investigated the efficacy of several phenolic compounds in controlling key cellular mechanisms involved in its infectivity. The way the SARS-CoV-2 virus infects the cell is a complex process and comprises four main stages: attachment to the cognate receptor, cellular entry, replication and cellular egress. Since, this is a multi-part process, it creates many opportunities to develop effective interventions. Targeting binding of the virus to the host receptor in order to prevent its entry has been of particular interest. Here, we provide experimental evidence that, among 56 tested polyphenols, including plant extracts, brazilin, theaflavin-3,3'-digallate, and curcumin displayed the highest binding with the receptor-binding domain of spike protein, inhibiting viral attachment to the human angiotensin-converting enzyme 2 receptor, and thus cellular entry of pseudo-typed SARS-CoV-2 virions. Both, theaflavin-3,3'-digallate at 25 µg/ml and curcumin above 10 µg/ml concentration, showed binding with the angiotensin-converting enzyme 2 receptor reducing at the same time its activity in both cell-free and cell-based assays. Our study also demonstrates that brazilin and theaflavin-3,3'-digallate, and to a still greater extent, curcumin, decrease the activity of transmembrane serine protease 2 both in cell-free and cell-based assays. Similar pattern was observed with cathepsin L, although only theaflavin-3,3'-digallate showed a modest diminution of cathepsin L expression at protein level. Finally, each of these three compounds moderately increased endosomal/lysosomal pH. In conclusion, this study demonstrates pleiotropic anti-SARS-CoV-2 efficacy of specific polyphenols and their prospects for further scientific and clinical investigations.

Polyunsaturated ω-3 fatty acids inhibit ACE2-controlled SARS-CoV-2 binding and cellular entry.

The strain SARS-CoV-2, newly emerged in late 2019, has been identified as the cause of COVID-19 and the pandemic declared by WHO in early 2020. Although lipids have been shown to possess antiviral efficacy, little is currently known about lipid compounds with anti-SARS-CoV-2 binding and entry properties. To address this issue, we screened, overall, 17 polyunsaturated fatty acids, monounsaturated fatty acids and saturated fatty acids, as wells as lipid-soluble vitamins. In performing target-based ligand screening utilizing the RBD-SARS-CoV-2 sequence, we observed that polyunsaturated fatty acids most effectively interfere with binding to hACE2, the receptor for SARS-CoV-2. Using a spike protein pseudo-virus, we also found that linolenic acid and eicosapentaenoic acid significantly block the entry of SARS-CoV-2. In addition, eicosapentaenoic acid showed higher efficacy than linolenic acid in reducing activity of TMPRSS2 and cathepsin L proteases, but neither of the fatty acids affected their expression at the protein level. Also, neither reduction of hACE2 activity nor binding to the hACE2 receptor upon treatment with these two fatty acids was observed. Although further in vivo experiments are warranted to validate the current findings, our study provides a new insight into the role of lipids as antiviral compounds against the SARS-CoV-2 strain.

Inhibition of ACE2 Expression by Ascorbic Acid Alone and its Combinations with Other Natural Compounds.

BACKGROUND: Angiotensin-converting enzyme II or ACE2 is an integral membrane protein present on many types of cells, including vascular endothelial cells and lung alveolar epithelial cells. This receptor serves as the entry point for SARS-coronaviruses (SARS-CoVs), including a novel coronavirus 2019-nCoV. Limited availability of these receptors can thwart cellular entry of this virus. METHODS: We tested the effects of ascorbic acid (vitamin C) on cellular expression of ACE2 at the protein and RNA levels in human small alveolar epithelial cells and microvascular endothelial cells. In addition, we investigated whether combinations of ascorbic acid with other natural compounds can affect ACE2 expression. RESULTS: The results show that ascorbic acid itself has moderate but consistent lowering effects on ACE2 expression at the cellular, protein, and RNA levels. Some natural compounds were effective in lowering ACE2 cellular expression, with the highest inhibitory effects observed for baicalin (75%) and theaflavin (50%). Significantly, combinations of these and other test compounds with ascorbic acid further decreased ACE2 expression. The highest impact of ascorbate on ACE2 expression was noted when combined with theaflavin (decrease from 50% to 87%), zinc (decrease from 22% to 62%), and with 10-undecenoic acid (from 18% to 53%). Ascorbic acid showed moderate additional benefits in decreasing ACE2 expression when combined with N-acetylcysteine and baicalin. CONCLUSION: Our study provides valuable experimental confirmation of the efficacy of micronutrients in controlling ACE2 expression-the coronavirus cellular "entry" point. It further validates the importance of nutrient interactions in various aspects of cellular metabolism and in considering potential therapeutic applications of nutrient-based approaches. The study shows that ascorbic acid and its combination with some natural compounds could be included in developing preventive and therapeutic approaches toward the current pandemic.