Uniform Gold Nanostructure Formation via Weakly Adsorbed Gold Films and Thermal Annealing for Reliable Localized Surface Plasmon Resonance-Based Detection of DNase-I.

Deoxyribonuclease-I (DNase-I), a representative endonuclease, is an important biomarker for the diagnosis of infectious diseases and cancer progression. However, enzymatic activity decreases rapidly ex vivo, which highlights the need for precise on-site detection of DNase-I. Here, a localized surface plasmon resonance (LSPR) biosensor that enables the simple and rapid detection of DNase-I is reported. Moreover, a novel technique named electrochemical deposition and mild thermal annealing (EDMIT) is applied to overcome signal variations. By taking advantage of the low adhesion of gold clusters on indium tin oxide substrates, both the uniformity and sphericity of gold nanoparticles are increased under mild thermal annealing conditions via coalescence and Ostwald ripening. This ultimately results in an approximately 15-fold decrease in LSPR signal variations. The linear range of the fabricated sensor is 20-1000 ng mL-1 with a limit of detection (LOD) of 127.25 pg mL-1 , as demonstrated by spectral absorbance analyses. The fabricated LSPR sensor stably measured DNase-I concentrations from samples collected from both an inflammatory bowel disease (IBD) mouse model, as well as human patients with severe COVID-19 symptoms. Therefore, the proposed LSPR sensor fabricated via the EDMIT method can be used for early diagnosis of other infectious diseases.

Cleavage-Responsive Biofactory T Cells Suppress Infectious Diseases-Associated Hypercytokinemia.

Severe infectious diseases, such as coronavirus disease 2019 (COVID-19), can induce hypercytokinemia and multiple organ failure. In spite of the growing demand for peptide therapeutics against infectious diseases, current small molecule-based strategies still require frequent administration due to limited half-life and enzymatic digestion in blood. To overcome this challenge, a strategy to continuously express multi-level therapeutic peptide drugs on the surface of immune cells, is established. Here, chimeric T cells stably expressing therapeutic peptides are presented for treatment of severe infectious diseases. Using lentiviral system, T cells are engineered to express multi-level therapeutic peptides with matrix metallopeptidases- (MMP-) and tumor necrosis factor alpha converting enzyme- (TACE-) responsive cleavage sites on the surface. The enzymatic cleavage releases γ-carboxyglutamic acid of protein C (PC-Gla) domain and thrombin receptor agonist peptide (TRAP), which activate endothelial protein C receptor (EPCR) and protease-activated receptor-1 (PAR-1), respectively. These chimeric T cells prevent vascular damage in tissue-engineered blood vessel and suppress hypercytokinemia and lung tissue damages in vivo, demonstrating promise for use of engineered T cells against sepsis and other infectious-related diseases.

Nanodiamond as a Cytokine Sponge in Infectious Diseases.

Cytokine release syndrome (CRS) is a systemic inflammatory response resulting in overexpression of cytokines in serum and tissues, which leads to multiple-organ failure. Due to rapid aggravation of symptoms, timely intervention is paramount; however, current therapies are limited in their capacity to address CRS. Here, we find that the intravenous injection of highly purified detonation-synthesized nanodiamonds (DND) can act as a therapeutic agent for treating CRS by adsorbing inflammatory cytokines. Highly purified DNDs successfully inactivated various key cytokines in plasma from CRS patients with pneumonia, septic shock, and coronavirus disease 2019 pandemic (COVID-19). The intravenous injection of the DND samples in a mouse sepsis model by cecal ligation and puncture significantly improved survival rates and prevented tissue damage by reducing the circulating inflammatory cytokines. The results of this study suggest that the clinical application of highly purified DND can provide survival benefits for CRS patients by adsorbing inflammatory cytokines.

PEGylated nanoparticle albumin-bound steroidal ginsenoside derivatives ameliorate SARS-CoV-2-mediated hyper-inflammatory responses.

The rapid spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on a global scale urges prompt and effective countermeasures. Recently, a study has reported that coronavirus disease-19 (COVID-19), the disease caused by SARS-CoV-2 infection, is associated with a decrease in albumin level, an increase in NETosis, blood coagulation, and cytokine level. Here, we present drug-loaded albumin nanoparticles as a therapeutic agent to resolve the clinical outcomes observed in severe SARS-CoV-2 patients. PEGylated nanoparticle albumin-bound (PNAB) was used to promote prolonged bioactivity of steroidal ginsenoside saponins, PNAB-Rg6 and PNAB-Rgx365. Our data indicate that the application of PNAB-steroidal ginsenoside can effectively reduce histone H4 and NETosis-related factors in the plasma, and alleviate SREBP2-mediated systemic inflammation in the PBMCs of SARS-CoV-2 ICU patients. The engineered blood vessel model confirmed that these drugs are effective in suppressing blood clot formation and vascular inflammation. Moreover, the animal model experiment showed that these drugs are effective in promoting the survival rate by alleviating tissue damage and cytokine storm. Altogether, our findings suggest that these PNAB-steroidal ginsenoside drugs have potential applications in the treatment of symptoms associated with severe SARS-CoV-2 patients, such as coagulation and cytokine storm.

Lung-selective 25-hydroxycholesterol nanotherapeutics as a suppressor of COVID-19-associated cytokine storm.

In response to the coronavirus disease-19 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), global efforts are focused on the development of new therapeutic interventions. For the treatment of COVID-19, selective lung-localizing strategies hold tremendous potential, as SARS-CoV-2 invades the lung via ACE2 receptors and causes severe pneumonia. Similarly, recent reports have shown the association of COVID-19 with decreased 25-hydroxycholesterol (25-HC) and increased cytokine levels. This mechanism, which involves the activation of inflammatory NF-κB- and SREBP2-mediated inflammasome signaling pathways, is believed to play a crucial role in COVID-19 pathogenesis, inducing acute respiratory distress syndrome (ARDS) and sepsis. To resolve those clinical conditions observed in severe SARS-CoV-2 patients, we report 25-HC and didodecyldimethylammonium bromide (DDAB) nanovesicles (25-HC@DDAB) as a COVID-19 drug candidate for the restoration of intracellular cholesterol level and suppression of cytokine storm. Our data demonstrate that 25-HC@DDAB can selectively accumulate the lung tissues and effectively downregulate NF-κB and SREBP2 signaling pathways in COVID-19 patient-derived PBMCs, reducing inflammatory cytokine levels. Altogether, our findings suggest that 25-HC@DDAB is a promising candidate for the treatment of symptoms associated with severe COVID-19 patients, such as decreased cholesterol level and cytokine storm.

Early Hydroxychloroquine Administration for Rapid Severe Acute Respiratory Syndrome Coronavirus 2 Eradication.

There are no proven therapeutics for Coronavirus disease 2019 (COVID-19) pneumonia outbreak. We observed and analyzed the clinical efficacy of the most used hydroxychloroquine (HCQ) for 30 days. In this study, administration of HCQ <5 days from diagnosis (odds ratio: 0.111, 95% confidence interval: 0.034 - 0.367, P = 0.001) was the only protective factor for prolonging of viral shedding in COVID-19 patients. Early administration of HCQ significantly ameliorates inflammatory cytokine secretion by eradicating COVID-19, at discharge. Our findings suggest that patients confirmed of COVID-19 infection should be administrated HCQ as soon as possible.

Long-acting nanoparticulate DNase-1 for effective suppression of SARS-CoV-2-mediated neutrophil activities and cytokine storm.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a new strain of coronavirus not previously identified in humans. Globally, the number of confirmed cases and mortality rates of coronavirus disease 2019 (COVID-19) have risen dramatically. Currently, there are no FDA-approved antiviral drugs and there is an urgency to develop treatment strategies that can effectively suppress SARS-CoV-2-mediated cytokine storms, acute respiratory distress syndrome (ARDS), and sepsis. As symptoms progress in patients with SARS-CoV-2 sepsis, elevated amounts of cell-free DNA (cfDNA) are produced, which in turn induce multiple organ failure in these patients. Furthermore, plasma levels of DNase-1 are markedly reduced in SARS-CoV-2 sepsis patients. In this study, we generated recombinant DNase-1-coated polydopamine-poly(ethylene glycol) nanoparticulates (named long-acting DNase-1), and hypothesized that exogenous administration of long-acting DNase-1 may suppress SARS-CoV-2-mediated neutrophil activities and the cytokine storm. Our findings suggest that exogenously administered long-acting nanoparticulate DNase-1 can effectively reduce cfDNA levels and neutrophil activities and may be used as a potential therapeutic intervention for life-threatening SARS-CoV-2-mediated illnesses.

Bioinspired DNase-I-Coated Melanin-Like Nanospheres for Modulation of Infection-Associated NETosis Dysregulation.

The current outbreak of the beta-coronavirus (beta-Cov) severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) began in December 2019. No specific antiviral treatments or vaccines are currently available. A recent study has reported that coronavirus disease 2019 (COVID-19), the disease caused by SARS-CoV-2 infection, is associated with neutrophil-specific plasma membrane rupture, and release excessive neutrophil extracellular traps (NETs) and extracellular DNAs (eDNAs). This mechanism involves the activation of NETosis, a neutrophil-specific programmed cell death, which is believed to play a crucial role in COVID-19 pathogenesis. Further progression of the disease can cause uncontrolled inflammation, leading to the initiation of cytokine storms, acute respiratory distress syndrome (ARDS), and sepsis. Herein, it is reported that DNase-I-coated melanin-like nanospheres (DNase-I pMNSs) mitigate sepsis-associated NETosis dysregulation, thereby preventing further progression of the disease. Recombinant DNase-I and poly(ethylene glycol) (PEG) are used as coatings to promote the lengthy circulation and dissolution of NET structure. The data indicate that the application of bioinspired DNase-I pMNSs reduce neutrophil counts and NETosis-related factors in the plasma of SARS-CoV-2 sepsis patients, alleviates systemic inflammation, and attenuates mortality in a septic mouse model. Altogether, the findings suggest that these nanoparticles have potential applications in the treatment of SARS-CoV-2-related illnesses and other beta-CoV-related diseases.

Ferritin Nanocage-Based Methyltransferase SETD6 for COVID-19 Therapy

Abstract The transcription factor nuclear factor-?B (NF-?B) signaling is a mediator of viral infection-mediated inflammation and SET-domain containing 6 (SETD6) is known as a methyltransferase that suppresses the activity of NF-?B signaling. However, the downside of the SETD6 is that it cannot be directly utilized as an inflammatory regulator due to the short half-life and poor intracellular delivery. Here, a ferritin nanocage-based delivery system is presented that can maintain the activity of SETD6 in vivo. According to the analysis of severe COVID-19 patients? peripheral blood mononuclear cells (PBMCs), the SETD6 expression is downregulated while that of NF-?B is upregulated. By engineering the structure of ferritin, a protein scaffold is fabricated in which short ferritin is decorated with cell-penetrating peptide and nuclear-localizing TAT-NBD peptide together with SETD6, termed TFS. The TFS enhances the SETD6 level and reduces the NF-?B signaling in PBMCs of severe COVID-19 patients and subsequently suppresses the cytokine storm. When the TFS is intravenously administered in the cytokine storm mouse model, the survival rate is rescued and the lung tissue damage and cytokine expression are also inhibited. These results indicate that the ferritin nanocage-based peptide delivery system allows stable in vivo delivery and efficient suppression of NF-?B signaling-mediated inflammation.

COVID-19-activated SREBP2 disturbs cholesterol biosynthesis and leads to cytokine storm.

Sterol regulatory element binding protein-2 (SREBP-2) is activated by cytokines or pathogen, such as virus or bacteria, but its association with diminished cholesterol levels in COVID-19 patients is unknown. Here, we evaluated SREBP-2 activation in peripheral blood mononuclear cells of COVID-19 patients and verified the function of SREBP-2 in COVID-19. Intriguingly, we report the first observation of SREBP-2 C-terminal fragment in COVID-19 patients' blood and propose SREBP-2 C-terminal fragment as an indicator for determining severity. We confirmed that SREBP-2-induced cholesterol biosynthesis was suppressed by Sestrin-1 and PCSK9 expression, while the SREBP-2-induced inflammatory responses was upregulated in COVID-19 ICU patients. Using an infectious disease mouse model, inhibitors of SREBP-2 and NF-κB suppressed cytokine storms caused by viral infection and prevented pulmonary damages. These results collectively suggest that SREBP-2 can serve as an indicator for severity diagnosis and therapeutic target for preventing cytokine storm and lung damage in severe COVID-19 patients.

Acetylated K676 TGFBIp as a severity diagnostic blood biomarker for SARS-CoV-2 pneumonia.

The outbreak of the highly contagious and deadly severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), also known as coronavirus disease 2019 (COVID-19), has posed a serious threat to public health across the globe, calling for the development of effective diagnostic markers and therapeutics. Here, we report a highly reliable severity diagnostic biomarker, acetylated 676th lysine transforming growth factor-beta-induced protein (TGFBIp K676Ac). TGFBIp K676Ac was consistently elevated in the blood of patients with SARS-CoV-2 pneumonia (n = 113), especially in patients in the intensive care unit (ICU) compared to non-ICU patients. Patients' blood samples showed increased cytokines and lymphopenia, which are exemplary indicators of SARS-CoV-2 pneumonia. Treatment with TGFBIp neutralizing antibodies suppressed the cytokine storm. The increased level of TGFBIp K676Ac in ICU patients suggests the promise of this protein as a reliable severity diagnostic biomarker for severe SARS-CoV-2 disease.

Early Hydroxychloroquine Administration for Rapid Severe Acute Respiratory Syndrome Coronavirus 2 Eradication

There are no proven therapeutics for Coronavirus disease 2019 (COVID-19) pneumonia outbreak. We observed and analyzed the clinical efficacy of the most used hydroxychloroquine (HCQ) for 30 days. In this study, administration of HCQ <5 days from diagnosis (odds ratio: 0.111, 95% confidence interval: 0.034 - 0.367, P = 0.001) was the only protective factor for prolonging of viral shedding in COVID-19 patients. Early administration of HCQ significantly ameliorates inflammatory cytokine secretion by eradicating COVID-19, at discharge. Our findings suggest that patients confirmed of COVID-19 infection should be administrated HCQ as soon as possible.

Prognostic Factors for Severe Coronavirus Disease 2019 in Daegu, Korea.

BACKGROUND: Since its first detection in December 2019, coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 infection has spread rapidly around the world. Although there have been several studies investigating prognostic factors for severe COVID-19, there have been no such studies in Korea. METHODS: We performed a retrospective observational study of 110 patients with confirmed COVID-19 hospitalized at a tertiary hospital in Daegu, Korea. Demographic, clinical, laboratory, and outcome data were collected and analyzed. Severe disease was defined as a composite outcome of acute respiratory distress syndrome, intensive care unit care, or death. RESULTS: Diabetes mellitus (odds ratio [OR], 19.15; 95% confidence interval [CI], 1.90-193.42; P = 0.012), body temperature ≥ 37.8°C (OR, 10.91; 95% CI, 1.35-88.36; P = 0.025), peripheral oxygen saturation < 92% (OR, 33.31; 95% CI, 2.45-452.22; P = 0.008), and creatine kinase-MB (CK-MB) > 6.3 (OR, 56.84; 95% CI, 2.64-1,223.78, P = 0.010) at admission were associated with higher risk of severe COVID-19. The likelihood of development of severe COVID-19 increased with an increasing number of prognostic factors. CONCLUSION: In conclusion, we found that diabetes mellitus, body temperature ≥ 37.8°C, peripheral oxygen saturation < 92%, and CK-MB > 6.3 are independent predictors of severe disease in hospitalized COVID-19 patients. Appropriate assessment of prognostic factors and close monitoring to provide the necessary interventions at the appropriate time in high-risk patients may reduce the case fatality rate of COVID-19.

Prognostic Accuracy of the SIRS, qSOFA, and NEWS for Early Detection of Clinical Deterioration in SARS-CoV-2 Infected Patients.

BACKGROUND: The case fatality rate of coronavirus disease 2019 (COVID-19) is estimated to be between 4.3% and 11.0%. Currently there is no effective antiviral treatment for COVID-19. Thus, early recognition of patients at high risk is important. METHODS: We performed a retrospective observational study of 110 patients with severe acute respiratory syndrome coronavirus 2 infection. We compared the effectiveness of three scoring systems: the Systemic Inflammatory Response Syndrome (SIRS), quick Sequential Organ Failure Assessment (qSOFA), and National Early Warning Score (NEWS) systems, for predicting the prognosis of COVID-19. The area under the receiver operating characteristic curve (AUROC) was used for these assessments, and Kaplan-Meier survival curves were used to identify the cumulative risk for 28-day mortality according to the NEWS stratification. RESULTS: For predicting 28-day mortality, NEWS was superior to qSOFA (AUROC, 0.867 vs. 0.779, P < 0.001), while there was no significant difference between NEWS and SIRS (AUROC, 0.867 vs. 0.639, P = 0.100). For predicting critical outcomes, NEWS was superior to both SIRS (AUROC, 0.918 vs. 0.744, P = 0.032) and qSOFA (AUROC, 0.918 vs. 0.760, P = 0.012). Survival time was significantly shorter for patients with NEWS ≥ 7 than for patients with NEWS < 7. CONCLUSION: Calculation of the NEWS at the time of hospital admission can predict critical outcomes in patients with COVID-19. Early intervention for high-risk patients can thereby improve clinical outcomes in COVID-19 patients.

Clinical Features and Outcomes of 98 Patients Hospitalized with SARS-CoV-2 Infection in Daegu, South Korea: A Brief Descriptive Study.

Although some information on the epidemiology of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and a few selected cases has been reported, data on the clinical characteristics and outcomes of patients hospitalized therewith in South Korea are lacking. We conducted a retrospective single-center study of 98 consecutive hospitalized patients with confirmed SARS-CoV-2 infection at Yeungnam University Medical Center in Daegu, South Korea. Sixty patients were women (61.2%), and the mean age was 55.4±17.1 years. Thirteen patients (13.3%) were treated in the intensive care unit (ICU). The mean interval from symptom onset to hospitalization was 7.7±4.5 days. Patients who received ICU care were significantly older and were more likely to have diabetes mellitus. The National Early Warning Score on the day of admission was significantly higher in patients requiring ICU care. Acute respiratory distress syndrome (13/13 patients; 100%), septic shock (9/13; 69.2%), acute cardiac injury (9/13; 69.2%), and acute kidney injury (8/13; 61.5%) were more common in patients who received ICU care. All patients received antibiotic therapy, and most (97/98 patients; 99.0%) received antiviral therapy (lopinavir/ritonavir). Hydroxychloroquine was used in 79 patients (80.6%), and glucocorticoid therapy was used in 18 patients (18.4%). In complete blood counts, lymphopenia was the most common finding (40/98 patients; 40.8%). Levels of all proinflammatory cytokines were significantly higher in ICU patients. As of March 29, 2020, the mortality rate was 5.1%. Here, we report the clinical characteristics and laboratory findings of SARS-CoV-2 patients in South Korea up to March 29, 2020.