Effective antiviral therapy is important for tackling the coronavirus disease 2019 (COVID-19) pandemic. We assessed the efficacy and safety of combined interferon beta-1b, lopinavir–ritonavir, and ...ribavirin for treating patients with COVID-19.
This was a multicentre, prospective, open-label, randomised, phase 2 trial in adults with COVID-19 who were admitted to six hospitals in Hong Kong. Patients were randomly assigned (2:1) to a 14-day combination of lopinavir 400 mg and ritonavir 100 mg every 12 h, ribavirin 400 mg every 12 h, and three doses of 8 million international units of interferon beta-1b on alternate days (combination group) or to 14 days of lopinavir 400 mg and ritonavir 100 mg every 12 h (control group). The primary endpoint was the time to providing a nasopharyngeal swab negative for severe acute respiratory syndrome coronavirus 2 RT-PCR, and was done in the intention-to-treat population. The study is registered with ClinicalTrials.gov, NCT04276688.
Between Feb 10 and March 20, 2020, 127 patients were recruited; 86 were randomly assigned to the combination group and 41 were assigned to the control group. The median number of days from symptom onset to start of study treatment was 5 days (IQR 3–7). The combination group had a significantly shorter median time from start of study treatment to negative nasopharyngeal swab (7 days IQR 5–11) than the control group (12 days 8–15; hazard ratio 4·37 95% CI 1·86–10·24, p=0·0010). Adverse events included self-limited nausea and diarrhoea with no difference between the two groups. One patient in the control group discontinued lopinavir–ritonavir because of biochemical hepatitis. No patients died during the study.
Early triple antiviral therapy was safe and superior to lopinavir–ritonavir alone in alleviating symptoms and shortening the duration of viral shedding and hospital stay in patients with mild to moderate COVID-19. Future clinical study of a double antiviral therapy with interferon beta-1b as a backbone is warranted.
The Shaw-Foundation, Richard and Carol Yu, May Tam Mak Mei Yin, and Sanming Project of Medicine.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Epstein-Barr virus (EBV) is a herpesvirus associated with nasopharyngeal carcinoma (NPC), gastric carcinoma (GC), and other malignancies. EBV is the first human virus found to express microRNAs ...(miRNAs), the functions of which remain largely unknown. We report on the regulation of a cellular protein named p53 up-regulated modulator of apoptosis (PUMA) by an EBV miRNA known as miR-BART5, which is abundantly expressed in NPC and EBV-GC cells. Modulation of PUMA expression by miR-BART5 and anti-miR-BART5 oligonucleotide was demonstrated in EBV-positive cells. In addition, PUMA was found to be significantly underexpressed in approximately 60% of human NPC tissues. Although expression of miR-BART5 rendered NPC and EBV-GC cells less sensitive to proapoptotic agents, apoptosis can be triggered by depleting miR-BART5 or inducing the expression of PUMA. Collectively, our findings suggest that EBV encodes an miRNA to facilitate the establishment of latent infection by promoting host cell survival.
Abstract
Background
Olfactory dysfunction (OD) has been reported in coronavirus disease 2019 (COVID-19). However, there are knowledge gaps about the severity, prevalence, etiology, and duration of OD ...in COVID-19 patients.
Methods
Olfactory function was assessed in all participants using questionnaires and the butanol threshold test (BTT). Patients with COVID-19 and abnormal olfaction were further evaluated using the smell identification test (SIT), sinus imaging, and nasoendoscopy. Selected patients received nasal biopsies. Systematic review was performed according to PRISMA guidelines. PubMed items from January 1, 2020 to April 23, 2020 were searched. Studies that reported clinical data on olfactory disturbances in COVID-19 patients were analyzed.
Results
We included 18 COVID-19 patients and 18 controls. Among COVID-19 patients, 12 of 18 (67%) reported olfactory symptoms and OD was confirmed in 6 patients by BTT and SIT. Olfactory dysfunction was the only symptom in 2 patients. Mean BTT score of patients was worse than controls (P = .004, difference in means = 1.8; 95% confidence interval, 0.6–2.9). Sinusitis and olfactory cleft obstruction were absent in most patients. Immunohistochemical analysis of nasal biopsy revealed the presence of infiltrative CD68+ macrophages harboring severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antigen in the stroma. Olfactory dysfunction persisted in 2 patients despite clinical recovery. Systematic review showed that the prevalence of olfactory disturbances in COVID-19 ranged from 5% to 98%. Most studies did not assess olfaction quantitatively.
Conclusions
Olfactory dysfunction is common in COVID-19 and may be the only symptom. Coronavirus disease 2019-related OD can be severe and prolonged. Mucosal infiltration by CD68+ macrophages expressing SARS-CoV-2 viral antigen may contribute to COVID-19-related OD.
COVID-19-related olfactory dysfunction is common (67%, 12/18 patients). Recovery can be slow (>3 weeks). Sinusitis and olfactory cleft obstruction were absent in most patients. Immunohistochemical analysis of nasal biopsy specimens demonstrated infiltrative CD68+ macrophages expressing SARS-CoV-2 antigen in the stroma.
Nasopharyngeal carcinoma (NPC) is an Epstein‐Barr virus (EBV)‐associated epithelial malignancy. The high expression of BART‐miRNAs (miR‐BARTs) during latent EBV infection in NPC strongly supports ...their pathological importance in cancer progression. Recently, we found that several BART‐miRNAs work co‐operatively to modulate the DNA damage response (DDR) by reducing Ataxia‐telangiectasia‐mutated (ATM) activity. In this study, we further investigated the role of miR‐BARTs on DDR. The immunohistochemical study showed that the DNA repair gene, BRCA1, is consistently down‐regulated in primary NPCs. Using computer prediction programs and a series of reporter assays, we subsequently identified the negative regulatory role of BART2‐3p, BART12, BART17‐5p and BART19‐3p in BRCA1 expression. The ectopic expression of these four miR‐BARTs suppressed endogenous BRCA1 expression in EBV‐negative epithelial cell lines, whereas BRCA1 expression was enhanced by repressing endogenous miR‐BARTs activities in C666‐1 cells. More importantly, suppressing BRCA1 expression in nasopharyngeal epithelial cell lines using miR‐BART17‐5p and miR‐BART19‐3p mimics reduced the DNA repair capability and increased the cell sensitivity to the DNA‐damaging chemotherapeutic drugs, cisplatin and doxorubicin. Our findings suggest that miR‐BARTs play a novel role in DDR and may facilitate the development of effective NPC therapies.
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FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Experience from influenza pandemics suggested that convalescent plasma treatment given within 4 to 5 days of symptom onset might be beneficial. However, robust treatment data are lacking.
This is a ...multicenter, prospective, double-blind, randomized controlled trial. Convalescent plasma from patients who recovered from the 2009 pandemic influenza A(H1N1) (AH1N1) infection was fractionated to hyperimmune IV immunoglobulin (H-IVIG) by CSL Biotherapies (now BioCSL). Patients with severe A(H1N1) infection on standard antiviral treatment requiring intensive care and ventilatory support were randomized to receive H-IVIG or normal IV immunoglobulin manufactured before 2009 as control. Clinical outcome and adverse effects were compared.
Between 2010 and 2011, 35 patients were randomized to receive H-IVIG (17 patients) or IV immunoglobulin (18 patients). One defaulted patient was excluded from analysis. No adverse events related to treatment were reported. Baseline demographics and viral load before treatment were similar between the two groups. Serial respiratory viral load demonstrated that H-IVIG treatment was associated with significantly lower day 5 and 7 posttreatment viral load when compared with the control (P = .04 and P = .02, respectively). The initial serum cytokine level was significantly higher in the H-IVIG group but fell to a similar level 3 days after treatment. Subgroup multivariate analysis of the 22 patients who received treatment within 5 days of symptom onset demonstrated that H-IVIG treatment was the only factor that independently reduced mortality (OR, 0.14; 95% CI, 0.02-0.92; P = .04).
Treatment of severe A(H1N1) infection with H-IVIG within 5 days of symptom onset was associated with a lower viral load and reduced mortality.
ClinialTrials.gov; No.: NCT01617317; URL: www.clinicaltrials.gov.
Infection with the Epstein-Barr virus (EBV) is a strong predisposing factor in the development of nasopharyngeal carcinoma (NPC). Many viral gene products including EBNA1, LMP1, and LMP2 have been ...implicated in NPC tumorigenesis, although the de novo control of these viral oncoproteins remains largely unclear. The recent discovery of EBV-encoded viral microRNA (miRNA) in lymphoid malignancies has prompted us to examine the NPC-associated EBV miRNA. Using large-scale cloning analysis on EBV-positive NPC cells, two novel EBV miRNA, now named miR-BART21 and miR-BART22, were identified. These two EBV-encoded miRNA are abundantly expressed in most NPC samples. We found two nucleotide variations in the primary transcript of miR-BART22, which we experimentally confirmed to augment its biogenesis in vitro and thus may underline the high and consistent expression of miR-BART22 in NPC tumors. More importantly, we determined that the EBV latent membrane protein 2A (LMP2A) is the putative target of miR-BART22. LMP2A is a potent immunogenic viral antigen that is recognized by the cytotoxic T cells; down-modulation of LMP2A expression by miR-BART22 may permit escape of EBV-infected cells from host immune surveillance. Taken together, we demonstrated that two newly identified EBV-encoded miRNA are highly expressed in NPC. Specific sequence variations on the prevalent EBV strain in our locality might contribute to the higher miR-BART22 expression level in our NPC samples. Our findings emphasize the role of miR-BART22 in modulating LMP2A expression, which may facilitate NPC carcinogenesis by evading the host immune response.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Epstein-Barr virus (EBV) was the first human virus found to encode microRNAs (miRNAs), but the function of these miRNAs has been obscure. Nasopharyngeal carcinoma (NPC) is associated with EBV ...infection, and the EBV-encoded LMP1 is believed to be a key factor in NPC development. However, detection of LMP1 protein in NPC is variable. Here, we report that EBV-encoded BART miRNAs target the 3' UTR of the LMP1 gene and negatively regulate LMP1 protein expression. These miRNAs also modulate LMP1-induced NF-κB signaling and alleviate the cisplatin sensitivity of LMP1-expressing NPC cells. Consistent with a previous study on the NPC C666-1 cell line and C15 xenograft, we found abundant expression of BART miRNAs in NPC tissues. Furthermore, DNA sequencing revealed that the 3' UTR of LMP1 is highly conserved in NPC-derived EBV isolates. The data provide insight into the discrepancy between LMP1 transcript and protein detection in NPC and highlight the role of the EBV miRNAs in regulating LMP1 downstream signaling to promote cancer development.
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
Cancer cells are defined genetically by the mutations they harbor, commonly single nucleotide substitutions. Therapeutic approaches which specifically target cancer cells by recognizing these ...defining genetic aberrations are expected to exhibit minimal side-effects. However, current protein-based targeted therapy is greatly limited by the range of genes that can be targeted, as well as by acquired resistance. We hypothesized that a therapeutic oligonucleotide-based strategy may address this need of specific cancer targeting. We used CRISPR/Cas9 system to target a commonly occurring EGFR point mutation, L858R, with an oligonucleotide guide that recognizes L858R as the suitable protospacer-adjacent motif (PAM) sequence for DNA cleavage. We found that this strategy, which utilized PAM to differentiate cancer mutation from normal, afforded high specificity to the extent of a single nucleotide substitution. The anti-L858R vehicle resulted in selective genome cleavage only in L858R mutant cells, as detected by Sanger sequencing and T7 Endonuclease I assay. Wild-type cells were unaffected by the same treatment. Digital PCR revealed 37.9 ± 8.57% of L858R gene copies were targeted in mutant. Only treated mutant cells, but not wild-type cells, showed reduction in EGFR expression and decreased cell proliferation. Treated mutant cells also formed smaller tumor load in vivo. This targeting approach is expected to be able to target a significant subset of the 15–35% cancer mutations with C > G, A > G, and T > G point mutations. Thus, this strategy may serve as a useful approach to target cancer-defining mutations with specificity, to the extent of differentiating the change of a single nucleotide.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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