Chimeric Virus-like Particle-Based COVID-19 Vaccine Confers Strong Protection against SARS-CoV-2 Viremia in K18-hACE2 Mice

Vaccines

2022 May 16

Kaewborisuth, C;Wanitchang, A;Koonpaew, S;Srisutthisamphan, K;Saenboonrueng, J;Im-Erbsin, R;Inthawong, M;Sunyakumthorn, P;Thaweerattanasinp, T;Tanwattana, N;Jantraphakorn, Y;Reed, MC;Lugo-Roman, LA;Hunsawong, T;Klungthong, C;Jones, AR;Fernandez, S;Teeravechyan, S;Lombardini, ED;Jongkaewwattana, A;
PMID: 35632541 | DOI: 10.3390/vaccines10050786

Virus-like particles (VLPs) are highly immunogenic and versatile subunit vaccines composed of multimeric viral proteins that mimic the whole virus but lack genetic material. Due to the lack of infectivity, VLPs are being developed as safe and effective vaccines against various infectious diseases. In this study, we generated a chimeric VLP-based COVID-19 vaccine stably produced by HEK293T cells. The chimeric VLPs contain the influenza virus A matrix (M1) proteins and the SARS-CoV-2 Wuhan strain spike (S) proteins with a deletion of the polybasic furin cleavage motif and a replacement of the transmembrane and cytoplasmic tail with that of the influenza virus hemagglutinin (HA). These resulting chimeric S-M1 VLPs, displaying S and M1, were observed to be enveloped particles that are heterogeneous in shape and size. The intramuscular vaccination of BALB/c mice in a prime-boost regimen elicited high titers of S-specific IgG and neutralizing antibodies. After immunization and a challenge with SARS-CoV-2 in K18-hACE2 mice, the S-M1 VLP vaccination resulted in a drastic reduction in viremia, as well as a decreased viral load in the lungs and improved survival rates compared to the control mice. Balanced Th1 and Th2 responses of activated S-specific T-cells were observed. Moderate degrees of inflammation and viral RNA in the lungs and brains were observed in the vaccinated group; however, brain lesion scores were less than in the PBS control. Overall, we demonstrate the immunogenicity of a chimeric VLP-based COVID-19 vaccine which confers strong protection against SARS-CoV-2 viremia in mice.

Histologic, viral, and molecular correlates of heart disease in fatal COVID-19

Annals of Diagnostic Pathology

2022 Oct 01

Mezache, L;Nuovo, G;Suster, D;Tili, E;Awad, H;Radwański, P;Veeraraghavan, R;
| DOI: 10.1016/j.anndiagpath.2022.151983

Cardiac manifestations are common in severe COVID-19. This study compared the histologic, viral, and molecular findings in cardiac tissue in fatal COVID-19 (n = 11) and controls (n = 11). In situ hybridization (SARS-CoV2 RNA) and immunohistochemistry for viral proteins and the host response were quantified for the samples and compared with qRTPCR and Western blot data. Control hearts showed a high resident population of macrophages that had variable ACE2 expression. Cardiac ACE2 expression was 10× greater in the heart tissues of cases and controls with obesity or type II diabetes. Multifocal endothelial cell swelling and degeneration, perivascular edema plus microvascular thrombi were unique to the cases. SARS-CoV2 RNA and nucleocapsid protein were rarely detected in situ in any COVID-19 heart. However, in each case abundant SARS-CoV-2 spike protein was evident. Co-expression experiments showed that the spike protein localized mostly to the ACE2+ interstitial macrophages/pericytes that were activated as evidenced by increased IL6 and TNFα expression. Western blots confirmed the presence of the viral spike protein, but not the nucleocapsid protein, in the cardiac homogenates. The intercalated disc proteins connexin 43, the primary cardiac gap junction protein, and NaV1.5, the predominant cardiac sodium channel, each showed marked lateral migration in the myocytes in the cases, which would increase the risk of reentrant arrhythmias. It is concluded that the viral spike protein, endocytosed by macrophages/pericytes, can induce a myocarditis with the possibility of conduction dysfunction due to abnormal localization of key intercalated disc proteins.

Co-administration of Favipiravir and the Remdesivir Metabolite GS-441524 Effectively Reduces SARS-CoV-2 Replication in the Lungs of the Syrian Hamster Model

mBio

2022 Feb 01

Chiba, S;Kiso, M;Nakajima, N;Iida, S;Maemura, T;Kuroda, M;Sato, Y;Ito, M;Okuda, M;Yamada, S;Iwatsuki-Horimoto, K;Watanabe, T;Imai, M;Armbrust, T;Baric, RS;Halfmann, PJ;Suzuki, T;Kawaoka, Y;
PMID: 35100870 | DOI: 10.1128/mbio.03044-21

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has spread worldwide since December 2019, causing coronavirus disease 2019 (COVID-19). Although vaccines for this virus have been developed rapidly, repurposing drugs approved to treat other diseases remains an invaluable treatment strategy. Here, we evaluated the inhibitory effects of drugs on SARS-CoV-2 replication in a hamster infection model and in in vitro assays. Favipiravir significantly suppressed virus replication in hamster lungs. Remdesivir inhibited virus replication in vitro, but was not effective in the hamster model. However, GS-441524, a metabolite of remdesivir, effectively suppressed virus replication in hamsters. Co-administration of favipiravir and GS-441524 more efficiently reduced virus load in hamster lungs than did single administration of either drug for both the prophylactic and therapeutic regimens; prophylactic co-administration also efficiently inhibited lung inflammation in the infected animals. Furthermore, pretreatment of hamsters with favipiravir and GS-441524 effectively protected them from virus transmission via respiratory droplets upon exposure to infected hamsters. Repurposing and co-administration of antiviral drugs may help combat COVID-19. IMPORTANCE During a pandemic, repurposing drugs that are approved for other diseases is a quick and realistic treatment option. In this study, we found that co-administration of favipiravir and the remdesivir metabolite GS-441524 more effectively blocked SARS-CoV-2 replication in the lungs of Syrian hamsters than either favipiravir or GS-441524 alone as part of a prophylactic or therapeutic regimen. Prophylactic co-administration also reduced the severity of lung inflammation. Moreover, co-administration of these drugs to naive hamsters efficiently protected them from airborne transmission of the virus from infected animals. Since both drugs are nucleotide analogs that interfere with the RNA-dependent RNA polymerases of many RNA viruses, these findings may also help encourage co-administration of antivirals to combat future pandemics.

SARS-CoV-2 colonization of maternal and fetal cells of the human placenta promotes alteration of local renin-angiotensin system

Med (New York, N.Y.)

2021 Apr 13

Verma, S;Joshi, CS;Silverstein, RB;He, M;Carter, EB;Mysorekar, IU;
PMID: 33870242 | DOI: 10.1016/j.medj.2021.04.009

SARS-CoV-2 infection appears to increase the risk of adverse pregnancy outcomes such as preeclampsia in pregnant women. The mechanism(s) by which this occurs remains unclear. We investigated the pathophysiology of SARS-CoV-2 at maternal-fetal interface in pregnant women who tested positive for the virus using RNA in situ hybridization (viral RNA), immunohistochemistry, and hematoxylin and eosin staining. To investigate whether viral infection alters the renin angiotensin system (RAS) in placenta which controls blood pressure, we treated human trophoblasts with recombinant Spike protein or a live modified virus with a vesicular stomatitis viral backbone expressing Spike protein (VSV-S). Viral colonization was highest in maternal decidua, fetal trophoblasts, Hofbauer cells, and in placentas delivered prematurely. We localized SARS-CoV-2 to cells expressing Angiotensin-converting enzyme 2 (ACE2), and demonstrate that infected placentas had significantly reduced ACE2. In response to both Spike protein and VSV-S, cellular ACE2 decreased while Angiotensin II receptor type 1 (AT1R) increased with concomitant increase in soluble fms-like tyrosine kinase-1(sFlt1). Viral infection decreased pro-angiogenic factors, AT2R and Placental growth factor, which competitively binds to sFlt1. Sera from infected pregnant women had elevated levels of sFlt1 and Angiotensin II type 1-Receptor Autoantibodies prior to delivery, both signatory markers of preeclampsia. SARS-CoV-2 colonizes ACE2-expressing maternal and fetal cells in the placenta. Infection in pregnant women correlates with alteration of placental RAS. As RAS regulates blood pressure, SARS-CoV-2 infection may thus increase adverse hemodynamic outcomes such as preeclampsia in pregnant women. NIH/NICHD grants R01 HD091218 and 3R01HD091218-04S1 (RADx-UP Supplement).

Sars Cov-2 Spike Protein-ACE receptor link, Graphene and Electro-Magnetic Field: toxicological aspetcs HYPOTESYS OF WORK

Trends Gen Med

2023 Jan 01

M, L;N, A;K, E;R, CB;G, T;R, A;Rasool, MG;C, F;Yurevich, CL;

Related the profile of some toxicological aspect of some COVID -19 vaccine and in specifc way to the rare effect like pericarditis or trombosys are reported in this work some relevant literature involved in the effect played by the SPIKE PROTEINS and its link on epithelial tissue ACE receptor, the Graphene (if present) and under some magnetic field or electrical condition. An experimental hypotesys is submitted to the reseacher in order to produce a global conclusion of toxicological interest.

A series of COVID-19 autopsies with clinical and pathologic comparisons to both seasonal and pandemic influenza

The journal of pathology. Clinical research

2021 May 07

McMullen, P;Pytel, P;Snyder, A;Smith, H;Vickery, J;Brainer, J;Guzy, R;Wu, D;Schoettler, N;Adegunsoye, A;Sperling, A;Hart, J;Alpert, L;Chang, A;Gurbuxani, S;Krausz, T;Husain, AN;Mueller, J;
PMID: 33960723 | DOI: 10.1002/cjp2.220

Autopsies of patients who have died from COVID-19 have been crucial in delineating patterns of injury associated with SARS-CoV-2 infection. Despite their utility, comprehensive autopsy studies are somewhat lacking relative to the global burden of disease, and very few comprehensive studies contextualize the findings to other fatal viral infections. We developed a novel autopsy protocol in order to perform postmortem examinations on victims of COVID-19 and herein describe detailed clinical information, gross findings, and histologic features observed in the first 16 complete COVID-19 autopsies. We also critically evaluated the role of ancillary studies used to establish a diagnosis of COVID-19 at autopsy, including immunohistochemistry (IHC), in situ hybridization (ISH), and electron microscopy (EM). IHC and ISH targeting SARS-CoV-2 were comparable in terms of the location and number of infected cells in lung tissue; however, nonspecific staining of bacteria was seen occasionally with IHC. EM was unrevealing in blindly sampled tissues. We then compared the clinical and histologic features present in this series to six archival cases of fatal seasonal influenza and six archival cases of pandemic influenza from the fourth wave of the 'Spanish Flu' in the winter of 1920. In addition to routine histology, the inflammatory infiltrates in the lungs of COVID-19 and seasonal influenza victims were compared using quantitative IHC. Our results demonstrate that the clinical and histologic features of COVID-19 are similar to those seen in fatal cases of influenza, and the two diseases tend to overlap histologically. There was no significant difference in the composition of the inflammatory infiltrate in COVID-19 and influenza at sites of acute lung injury at the time of autopsy. Our study underscores the relatively nonspecific clinical features and pathologic changes shared between severe cases of COVID-19 and influenza, while also providing important caveats to ancillary methods of viral detection.

SARS-CoV-2 Infection in Unvaccinated High Risk Pregnant Women in the Bronx, NY is Associated with Placental Abnormalities, Shorter Gestation and Lower Apgar Scores

Preprint

2022 Sep 06

Reznik, S;Vuguin, P;Khoury, R;Loudig, O;Balakrashnian, R;Fineberg, S;Hughes, F;Harigopal, M;Charron, M;
| DOI: 10.20944/preprints202209.0063.v1

. Babies born to severe acute respiratory syndrome corona virus-2 (SARS-CoV-2) infected mothers are at greater risk for perinatal morbidity and more likely to receive a neurodevelopmental diagnosis in the first year of life. However, the effect of maternal infection on placental function and neonatal outcomes varies depending upon the patient population. We set out to test our hypothesis that maternal SARS-CoV-2 infection in our underserved, socioeconomically disadvantaged, predominantly African American and Latina population in the Bronx, NY would have effects evident at birth. Fifty-five SARS-CoV-2 positive and 61 negative third trimester patients were randomly selected from Montefiore Medical Center (MMC), Bronx, NY. In addition, two positive cases from Yale New Haven Hospital, CT were included as controls. All 55 placentas delivered by SARS-CoV-2 positive mothers were uninfected by the virus, based on immunohistochemistry, in-situ hybridization, and qPCR analysis. However, placental villous infarcts, mild preeclampsia, shortened gestational periods and lower Apgar scores were observed in the infected cases. These findings suggest that even without entering the placenta, SARS-CoV-2 can affect various systemic pathways culminating in altered placental development and function, which may adversely affect the fetus, especially in a high-risk patient population such as ours. These results underline the importance of vaccination among pregnant women, particularly in low resource areas.

The differential immune response in mild versus fatal SARS-CoV2 infection

Annals of diagnostic pathology

2022 Sep 02

Suster, D;Tili, E;Nuovo, GJ;
PMID: 36113259 | DOI: 10.1016/j.anndiagpath.2022.152032

This study compared the immune response in mild versus fatal SARS-CoV2 infection. Forty nasopharyngeal swabs with either productive mild infection (n = 20) or negative for SARS-CoV2 (n = 20) were tested along with ten lung sections from people who died of COVID-19 which contained abundant SARS-CoV2 and ten controls. There was a 25-fold increase in the CD3+T cell numbers in the viral positive nasopharyngeal swabs compared to the controls (p < 0.001) and no change in the CD3+T cell count in the fatal COVID-19 lungs versus the controls. CD11b + and CD206+ macrophage counts were significantly higher in the mild versus fatal disease (p = 0.002). In situ analysis for SARS-CoV2 RNA found ten COVID-19 lung sections that had no/rare detectable virus and also lacked the microangiopathy typical of the viral positive sections. These viral negative lung tissues when compared to the viral positive lung samples showed a highly significant increase in CD3+ and CD8 T cells (p < 0.001), equivalent numbers of CD163+ cells, and significantly less PDL1, CD11b and CD206+ cells (p = 0.002). It is concluded that mild SARS-CoV2 infection is marked by a much stronger CD3/CD8 T cell, CD11b, and CD206 macrophage response than the fatal lung disease where viral RNA is abundant.

The cGAS-STING pathway drives type I IFN immunopathology in COVID-19

Nature

2022 Jan 19

Di Domizio, J;Gulen, MF;Saidoune, F;Thacker, VV;Yatim, A;Sharma, K;Nass, T;Guenova, E;Schaller, M;Conrad, C;Goepfert, C;De Leval, L;von Garnier, C;Berezowska, S;Dubois, A;Gilliet, M;Ablasser, A;
PMID: 35045565 | DOI: 10.1038/s41586-022-04421-w

Coronavirus disease 2019 (COVID-19), caused by infection with Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), is characterized by significant lung pathology and extrapulmonary complications1,2. Type I interferons (IFNs) play an essential role in the pathogenesis of COVID-193-5. While rapid induction of type I IFNs limits virus propagation, sustained elevation of type I IFNs in the late phase of the infection is associated with aberrant inflammation and poor clinical outcome5-17. Here, we identify the cyclic GMP-AMP synthase (cGAS)-Stimulator of interferon genes (STING)-pathway, which controls immunity to cytosolic DNA, as a critical driver of aberrant type I IFN responses in COVID-1918. Profiling COVID-19 skin manifestations, we uncover a STING-dependent type I IFN signature primarily mediated by macrophages adjacent to areas of endothelial cell damage. Moreover, cGAS-STING activity was detected in lung samples of COVID-19 patients with prominent tissue destruction and associated with type I IFN responses. A lung-on-chip model revealed that, in addition to macrophages, SARS-CoV-2 infection activates cGAS-STING signalling in endothelial cells through mitochondrial DNA release, leading to cell death and type I IFN production. In mice, pharmacological inhibition of STING reduces severe lung inflammation induced by SARS-CoV-2 and improves disease outcome. Collectively, our study establishes a mechanistic basis of pathological type I IFN responses in COVID-19 and reveals a novel principle for the development of host-directed therapeutics.

Long-term stability and protection efficacy of the RBD-targeting COVID-19 mRNA vaccine in nonhuman primates

Signal transduction and targeted therapy

2021 Dec 24

Zhao, H;Wang, TC;Li, XF;Zhang, NN;Li, L;Zhou, C;Deng, YQ;Cao, TS;Yang, G;Li, RT;Huang, YJ;Li, YG;Zhang, YM;Li, FX;Zhou, YR;Jiang, YH;Lu, XS;Sun, SH;Cheng, ML;Gu, KP;Zhang, M;Ma, QQ;Yang, X;Ying, B;Gao, YW;Qin, CF;
PMID: 34952914 | DOI: 10.1038/s41392-021-00861-4

Messenger RNA (mRNA) vaccine technology has shown its power in preventing the ongoing COVID-19 pandemic. Two mRNA vaccines targeting the full-length S protein of SARS-CoV-2 have been authorized for emergency use. Recently, we have developed a lipid nanoparticle-encapsulated mRNA (mRNA-LNP) encoding the receptor-binding domain (RBD) of SARS-CoV-2 (termed ARCoV), which confers complete protection in mouse model. Herein, we further characterized the protection efficacy of ARCoV in nonhuman primates and the long-term stability under normal refrigerator temperature. Intramuscular immunization of two doses of ARCoV elicited robust neutralizing antibodies as well as cellular response against SARS-CoV-2 in cynomolgus macaques. More importantly, ARCoV vaccination in macaques significantly protected animals from acute lung lesions caused by SARS-CoV-2, and viral replication in lungs and secretion in nasal swabs were completely cleared in all animals immunized with low or high doses of ARCoV. No evidence of antibody-dependent enhancement of infection was observed throughout the study. Finally, extensive stability assays showed that ARCoV can be stored at 2-8 °C for at least 6 months without decrease of immunogenicity. All these promising results strongly support the ongoing clinical trial.

Infection of lung megakaryocytes and platelets by SARS-CoV-2 anticipate fatal COVID-19

Cellular and molecular life sciences : CMLS

2022 Jun 16

Zhu, A;Real, F;Capron, C;Rosenberg, AR;Silvin, A;Dunsmore, G;Zhu, J;Cottoignies-Callamarte, A;Massé, JM;Moine, P;Bessis, S;Godement, M;Geri, G;Chiche, JD;Valdebenito, S;Belouzard, S;Dubuisson, J;Lorin de la Grandmaison, G;Chevret, S;Ginhoux, F;Eugenin, EA;Annane, D;Bordé, EC;Bomsel, M;
PMID: 35708858 | DOI: 10.1007/s00018-022-04318-x

SARS-CoV-2, although not being a circulatory virus, spread from the respiratory tract resulting in multiorgan failures and thrombotic complications, the hallmarks of fatal COVID-19. A convergent contributor could be platelets that beyond hemostatic functions can carry infectious viruses. Here, we profiled 52 patients with severe COVID-19 and demonstrated that circulating platelets of 19 out 20 non-survivor patients contain SARS-CoV-2 in robust correlation with fatal outcome. Platelets containing SARS-CoV-2 might originate from bone marrow and lung megakaryocytes (MKs), the platelet precursors, which were found infected by SARS-CoV-2 in COVID-19 autopsies. Accordingly, MKs undergoing shortened differentiation and expressing anti-viral IFITM1 and IFITM3 RNA as a sign of viral sensing were enriched in the circulation of deadly COVID-19. Infected MKs reach the lung concomitant with a specific MK-related cytokine storm rich in VEGF, PDGF and inflammatory molecules, anticipating fatal outcome. Lung macrophages capture SARS-CoV-2-containing platelets in vivo. The virus contained by platelets is infectious as capture of platelets carrying SARS-CoV-2 propagates infection to macrophages in vitro, in a process blocked by an anti-GPIIbIIIa drug. Altogether, platelets containing infectious SARS-CoV-2 alter COVID-19 pathogenesis and provide a powerful fatality marker. Clinical targeting of platelets might prevent viral spread, thrombus formation and exacerbated inflammation at once and increase survival in COVID-19.

Surface Proteins of SARS-CoV-2 Drive Airway Epithelial Cells to Induce IFN-Dependent Inflammation

Journal of immunology (Baltimore, Md. : 1950)

2021 Jun 02

Anand, G;Perry, AM;Cummings, CL;Raymond, E;Clemens, RA;Steed, AL;
PMID: 34078711 | DOI: 10.4049/jimmunol.2001407

SARS-CoV-2, the virus that has caused the COVID-19 pandemic, robustly activates the host immune system in critically ill patients. Understanding how the virus engages the immune system will facilitate the development of needed therapeutic strategies. In this study, we demonstrate both in vitro and in vivo that the SARS-CoV-2 surface proteins spike (S) and envelope (E) activate the key immune signaling IFN pathway in both human and mouse immune and epithelial cells independent of viral infection and replication. These proteins induce reactive oxidative species generation and increases in human- and murine-specific, IFN-responsive cytokines and chemokines, similar to their upregulation in critically ill COVID-19 patients. Induction of IFN signaling is dependent on canonical but discrepant inflammatory signaling mediators, as the activation induced by S is dependent on IRF3, TBK1, and MyD88, whereas that of E is largely MyD88 independent. Furthermore, these viral surface proteins, specifically E, induced peribronchial inflammation and pulmonary vasculitis in a mouse model. Finally, we show that the organized inflammatory infiltrates are dependent on type I IFN signaling, specifically in lung epithelial cells. These findings underscore the role of SARS-CoV-2 surface proteins, particularly the understudied E protein, in driving cell specific inflammation and their potential for therapeutic intervention.

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sense Example: Hs-LAG3-sense	Standard probes for RNA detection are in antisense. Sense probe is reverse complent to the corresponding antisense probe.
Intron# Example: Mm-Htt-intron2	Probe targets the indicated intron in the target gene, commonly used for pre-mRNA detection
Pool/Pan Example: Hs-CD3-pool (Hs-CD3D, Hs-CD3E, Hs-CD3G)	A mixture of multiple probe sets targeting multiple genes or transcripts
No-XSp Example: Hs-PDGFB-No-XMm	Does not cross detect with the species (Sp)
XSp Example: Rn-Pde9a-XMm	designed to cross detect with the species (Sp)
O# Example: Mm-Islr-O1	Alternative design targeting different regions of the same transcript or isoforms
CDS Example: Hs-SLC31A-CDS	Probe targets the protein-coding sequence only
EnEm	Probe targets exons n and m
En-Em	Probe targets region from exon n to exon m
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tvn Example: Hs-LEPR-tv1	Designed to target transcript variant n
ORF Example: Hs-ACVRL1-ORF	Probe targets open reading frame
UTR Example: Hs-HTT-UTR-C3	Probe targets the untranslated region (non-protein-coding region) only
5UTR Example: Hs-GNRHR-5UTR	Probe targets the 5' untranslated region only
3UTR Example: Rn-Npy1r-3UTR	Probe targets the 3' untranslated region only
Pan Example: Pool	A mixture of multiple probe sets targeting multiple genes or transcripts

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