Broadly neutralizing human antibodies against Omicron subvariants of SARS-CoV-2.

BACKGROUND: The COVID-19 pandemic continues to pose a significant worldwide threat to human health, as emerging SARS-CoV-2 Omicron variants exhibit resistance to therapeutic antibodies and the ability to evade vaccination-induced antibodies. Here, we aimed to identify human antibodies (hAbs) from convalescent patients that are potent and broadly neutralizing toward Omicron sublineages. METHODS: Using a single B-cell cloning approach, we isolated BA.5 specific human antibodies. We further examined the neutralizing activities of the most promising neutralizing hAbs toward different variants of concern (VOCs) with pseudotyped virus. RESULTS: Sixteen hAbs showed strong neutralizing activities against Omicron BA.5 with low IC50 values (IC50 < 20 ng/mL). Among four of the most promising neutralizing hAbs (RBD-hAb-B22, -B23, -B25 and -B34), RBD-hAb-B22 exhibited the most potent and broad neutralization profiles across Omicron subvariant pseudoviruses, with low IC50 values (7.7-41.6 ng/mL) and a low PRNT50 value (3.8 ng/mL) in plaque assays with authentic BA.5. It also showed potent therapeutic effects in BA.5-infected K18-hACE2 mice. CONCLUSIONS: Thus, our efficient screening of BA.5-specific neutralizing hAbs from breakthrough infectious convalescent donors successfully yielded hAbs with potent therapeutic potential against multiple SARS-CoV-2 variants.

Broadly neutralizing antibodies against Omicron variants of SARS-CoV-2 derived from mRNA-lipid nanoparticle-immunized mice.

The COVID-19 pandemic continues to threaten human health worldwide as new variants of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerge. Currently, the predominant circulating strains around the world are Omicron variants, which can evade many therapeutic antibodies. Thus, the development of new broadly neutralizing antibodies remains an urgent need. In this work, we address this need by using the mRNA-lipid nanoparticle immunization method to generate a set of Omicron-targeting monoclonal antibodies. Five of our novel K-RBD-mAbs show strong binding and neutralizing activities toward all SARS-CoV-2 variants of concern (Alpha, Beta, Gamma, Delta and Omicron). Notably, the epitopes of these five K-RBD-mAbs are overlapping and localized around Y453 and F486 of the spike protein receptor binding domain (RBD). Chimeric derivatives of the five antibodies (K-RBD-chAbs) neutralize Omicron sublineages BA.1 and BA.2 with low IC50 values ranging from 5.7 to 12.9 ng/mL. Additionally, we performed antibody humanization on broadly neutralizing chimeric antibodies to create K-RBD-hAb-60 and -62, which still retain excellent neutralizing activity against Omicron. Our results collectively suggest that these five therapeutic antibodies may effectively combat current and emerging SARS-CoV-2 variants, including Omicron BA.1 and BA.2. Therefore, the antibodies can potentially be used as universal neutralizing antibodies against SARS-CoV-2.

A critical overview of current progress for COVID-19: development of vaccines, antiviral drugs, and therapeutic antibodies.

The novel coronavirus disease (COVID-19) pandemic remains a global public health crisis, presenting a broad range of challenges. To help address some of the main problems, the scientific community has designed vaccines, diagnostic tools and therapeutics for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. The rapid pace of technology development, especially with regard to vaccines, represents a stunning and historic scientific achievement. Nevertheless, many challenges remain to be overcome, such as improving vaccine and drug treatment efficacies for emergent mutant strains of SARS-CoV-2. Outbreaks of more infectious variants continue to diminish the utility of available vaccines and drugs. Thus, the effectiveness of vaccines and drugs against the most current variants is a primary consideration in the continual analyses of clinical data that supports updated regulatory decisions. The first two vaccines granted Emergency Use Authorizations (EUAs), BNT162b2 and mRNA-1273, still show more than 60% protection efficacy against the most widespread current SARS-CoV-2 variant, Omicron. This variant carries more than 30 mutations in the spike protein, which has largely abrogated the neutralizing effects of therapeutic antibodies. Fortunately, some neutralizing antibodies and antiviral COVID-19 drugs treatments have shown continued clinical benefits. In this review, we provide a framework for understanding the ongoing development efforts for different types of vaccines and therapeutics, including small molecule and antibody drugs. The ripple effects of newly emergent variants, including updates to vaccines and drug repurposing efforts, are summarized. In addition, we summarize the clinical trials supporting the development and distribution of vaccines, small molecule drugs, and therapeutic antibodies with broad-spectrum activity against SARS-CoV-2 strains.

Recording and manipulation of the maternal oxytocin neural activities in mice.

Pulsatile release of the hormone oxytocin (OT) mediates uterine contraction during parturition and milk ejection during lactation.1-3 These pulses are generated by the unique activity patterns of the central neuroendocrine OT neurons located in the paraventricular and supraoptic hypothalamus. Classical studies have characterized putative OT neurons by in vivo extracellular recording techniques in rats and rabbits.1,4-10 Due to technical limitations, however, the identity of OT neurons in these previous studies was speculative based on their electrophysiological characteristics and axonal projection to the posterior pituitary, not on OT gene expression. To pinpoint OT neural activities among other hypothalamic neurons that project to the pituitary11,12 and make better use of cell-type-specific neuroscience toolkits,13 a mouse model needs to be developed for the studies of parturition and lactation. We herein introduce viral genetic approaches in mice to characterize the maternal activities of OT neurons by fiber photometry. A sharp photometric peak of OT neurons appeared at approximately 520 s following simultaneous suckling stimuli from three pups. The amplitude of the peaks increased as the mother mice experienced lactation, irrespective of the age of the pups, suggesting the intrinsic plasticity of maternal OT neurons. Based on a mono-synaptic input map to OT neurons, we pharmacogenetically activated the inhibitory neurons in the bed nucleus of the stria terminalis and found the suppression of the activities of OT neurons. Collectively, our study illuminates temporal dynamics in the maternal neural activities of OT neurons and identifies one of its modulatory inputs.

The cytokine-cosmc signaling axis upregulates the tumor-associated carbohydrate antigen Tn.

Tn antigen (GalNAc-α-O-Ser/Thr), a mucin-type O-linked glycan, is a well-established cell surface marker for tumors and its elevated levels have been correlated with cancer progression and prognosis. There are also reports that Tn is elevated in inflammatory tissues. However, the molecular mechanism for its elevated levels in cancer and inflammation is unclear. In the current studies, we have explored the possibility that cytokines may be one of the common regulatory molecules for elevated Tn levels in both cancer and inflammation. We showed that the Tn level is elevated by the conditioned media of HrasG12V-transformed-BEAS-2B cells. Similarly, the conditioned media obtained from LPS-stimulated monocytes also elevated Tn levels in primary human gingival fibroblasts, suggesting the involvement of cytokines and/or other soluble factors. Indeed, purified inflammatory cytokines such as TNF-α and IL-6 up-regulated Tn levels in gingival fibroblasts. Furthermore, TNF-α was shown to down-regulate the COSMC gene as evidenced by reduced levels of the COSMC mRNA and protein, as well as hypermethylation of the CpG islands of the COSMC gene promoter. Since Cosmc, a chaperone for T-synthase, is known to negatively regulate Tn levels, our results suggest elevated Tn levels in cancer and inflammation may be commonly regulated by the cytokine-Cosmc signaling axis.

Reciprocal relationship of Tn/NF-κB and sTn as an indicator of the prognosis of oral squamous cell carcinoma.

AIMS: In order to determine whether the expression of tumour-associated carbohydrate antigens (Tn/sTn) and a representative inflammation marker, nuclear factor-κB (NF-κB), is associated with the invasiveness of oral squamous cell carcinoma (OSCC), this study has attempted to investigate the correlation of the aforementioned markers with the well-established invasive pattern grading score (IPGS) and clinicopathological parameters. METHODS AND RESULTS: Specimens from 143 OSCC patients with classified clinicopathological parameters and IPGS were stained immunohistochemically using anti-Tn, sTn and NF-κB antibodies. Our results showed that the expression of both Tn and NF-κB was correlated positively with staging (P = 0.036; P = 0.015), recurrence (P < 0.001; P < 0.001) and distant metastasis (P = 0.005; P = 0.009), as well as with IPGS, while the expression of sTn was correlated inversely. In addition, poor survival was associated with overexpression of Tn and NF-κB but not with expression of sTn. CONCLUSIONS: Our results indicate that a reciprocal relationship between Tn and sTn expression may serve as a reliable indicator for OSCC prognostic evaluation. In addition, expression of Tn rather than sTn may play an important role in deeply invasive OSCC via regulation of NF-κB signalling.

Mechanisms of ceramide-induced COX-2-dependent apoptosis in human ovarian cancer OVCAR-3 cells partially overlapped with resveratrol.

Ceramide is a member of the sphingolipid family of bioactive molecules demonstrated to have profound, diverse biological activities. Ceramide is a potential chemotherapeutic agent via the induction of apoptosis. Exposure to ceramide activates extracellular-signal-regulated kinases (ERK)1/2- and p38 kinase-dependent apoptosis in human ovarian cancer OVCAR-3 cells, concomitant with an increase in the expression of COX-2 and p53 phosphorylation. Blockade of cyclooxygenase-2 (COX-2) activity by siRNA or NS398 correspondingly inhibited ceramide-induced p53 Ser-15 phosphorylation and apoptosis; thus COX-2 appears at the apex of the p38 kinase-mediated signaling cascade induced by ceramide. Induction of apoptosis by ceramide or resveratrol was inhibited by the endocytosis inhibitor, cytochalasin D (CytD); however, cells exposed to resveratrol showed greater sensitivity than ceramide-treated cells. Ceramide-treated cells underwent a dose-dependent reduction in trans-membrane potential. Although both ceramide and resveratrol induced the expressions of caspase-3 and -7, the effect of inducible COX-2 was different in caspase-7 expression induced by ceramide compared to resveratrol. In summary, resveratrol and ceramide converge on an endocytosis-requiring, ERK1/2-dependent signal transduction pathway and induction of COX-expression as an essential molecular antecedent for subsequent p53-dependent apoptosis. In addition, expressions of caspase-3 and -7 are observed. However, a p38 kinase-dependent signal transduction pathway and change in mitochondrial potential are also involved in ceramide-induced apoptosis.

A novel synthetic bipartite carrier protein for developing glycotope-based vaccines.

Development of successful vaccines against glycotopes remains a major challenge. In the current studies, we have successfully developed a novel carrier protein for glycotopes based on the concept of antigen clustering and specific stimulation of T helper cells to mount strong antibody response to glycotopes. The bipartite carrier protein consists of a tandem repeat of a cysteine-rich peptide for docking of clustered glycotopes to effectively activate B cells and an Fc domain for antigen delivery to antigen presenting cells (APCs). To demonstrate its utility, we conjugated the tumor-specific monosaccharide antigen Tn to this novel carrier protein and successfully developed a Tn vaccine against cancer in animal models. The Tn vaccine effectively elicited high-titer IgG1 antibodies against Tn in immunized mice, and effectively suppressed the development of prostate cancer in Transgenic Adenocarcinoma of the Mouse Prostate (TRAMP) mice. Our results suggest that this novel bipartite carrier protein could be effectively used for developing anti-glycotope vaccines such as the anticancer Tn vaccine.