Hitting SARS-CoV-2 in a new spot: A key target for therapeutic antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the spike protein, a trimeric protein complex with each monomer comprising an S1 and an S2 domain that mediate binding to host cells and membrane fusion, respectively. In addition to the receptor binding domain (RBD), S1 has an N-terminal domain (NTD). In searching for neutralizing antibodies, there has been a focus on the RBD. Chi et al. isolated antibodies from 10 convalescent patients and identified an antibody that potently neutralizes the virus but does not bind the RBD. Cryo–electron microscopy revealed the epitope as the NTD. This NTD-targeting antibody may be useful to combine with RBD-targeting antibodies in therapeutic cocktails. Science , this issue p. 650

The global spread of SARS-CoV-2 and its variants poses a serious threat to human health worldwide. Recently, the emergence of Omicron has presented a new challenge to the prevention and control of the COVID-19 pandemic. A convenient and reliable in vitro neutralization assay is an important method for validating the efficiency of antibodies, vaccines, and other potential drugs. Here, we established an effective assay based on a pseudovirus carrying a full-length spike (S) protein of SARS-CoV-2 variants in the HIV-1 backbone, with a luciferase reporter gene inserted into the non-replicate pseudovirus genome. The key parameters for packaging the pseudovirus were optimized, including the ratio of the S protein expression plasmids to the HIV backbone plasmids and the collection time for the Alpha, Beta, Gamma, Kappa, and Omicron pseudovirus particles. The pseudovirus neutralization assay was validated using several approved or developed monoclonal antibodies, underscoring that Omicron can escape some neutralizing antibodies, such as REGN10987 and REGN10933, while S309 and ADG-2 still function with reduced neutralization capability. The neutralizing capacity of convalescent plasma from COVID-19 convalescent patients in Wuhan was tested against these pseudoviruses, revealing the immune evasion of Omicron. Our work established a practical pseudovirus-based neutralization assay for SARS-CoV-2 variants, which can be conducted safely under biosafety level-2 (BSL-2) conditions, and this assay will be a promising tool for studying and characterizing vaccines and therapeutic candidates against Omicron-included SARS-CoV-2 variants.

The Chikungunya virus (CHIKV) is a priority endemic pathogen identified by the World Health Organization and its infection induces an acute febrile illness in humans that is often associated with arthritis and musculoskeletal pain. Therefore, specific vaccines and treatments are urgently needed to prevent or treat Chikungunya disease. Here, we identify a series of CHIKV-specific neutralizing nanobodies (Nbs) from an alpaca which exhibit distinct binding modes compared to those previously reported. Two representative anti-CHIKV Nbs, N033-Fc and N053-Fc, demonstrated significant antiviral activity in Ifnar^−/− mice against lethal challenge. Further studies elucidated the functional mechanisms of N033-Fc and N053-Fc in blocking CHIKV infection at multiple stages of the viral life cycle. This study identifies multiple candidate Nbs that may be suitable for next-generation antibody therapies to combat CHIKV infection.

The Junín virus (JUNV) is one of the New World arenaviruses that cause severe hemorrhagic fever. Human transferrin receptor 1 (hTfR1) has been identified as the main receptor for JUNV for virus entry into host cells. To date, no treatment has been approved for JUNV. Herein, we investigated 12 anti-hTfR1 VHH (variable domain of the heavy chain of heavy-chain antibody) antibodies and confirmed their interaction with hTfR1. Most of them could bind to the hTfR1 apical domain, which is the glycoprotein 1 (GP1) binding domain of JUNV. Among them, 18N18 exhibited neutralizing activity against both the human immunodeficiency virus (HIV)-vectored lentiviral Junín pseudoviruses and the recombinant vesicular stomatitis virus (VSV)-vectored Junín pseudoviruses. We also verified that 18N18 blocked the interaction between hTfR1 and JUNV GP1. In addition, 18N18 could neutralize another New World arenavirus, the Machupo virus. Using AlphaFold 3-based simulation of 18N18–hTfR1 docking, we determined that 18N18’s binding epitope was located at the JUNV GP1 binding epitope. 18N18 represents a candidate for JUNV treatment and provides a potential approach that could be applied to New World arenaviruses.

Rift Valley fever virus (RVFV) is a mosquito-transmitted bunyavirus that can cause substantial morbidity and mortality in livestock and humans, for which there are no currently available licensed human therapeutics or vaccines. Therefore, the development of safe and effective antivirals is both necessary and urgent. The Gc protein is the primary target of the neutralizing antibody response related to Rift Valley fever virus. Here, we report one Gc-specific neutralizing antibody (NA137) isolated from an alpaca and one bispecific antibody (E2-NA137), the protective efficacies of which we evaluated in A129 mice. In this prophylactic study, the survival rates of the NA137 and E2-NA137 groups were both 80%, and in the treatment study, the survival rates were 20% and 60%, respectively. Altogether, our results emphasize that NA137 and E2-NA137 provide a potential approach for treating RVFV either prophylactically or therapeutically.

Background: The interferon-inducible transmembrane protein Ifitm3 (also known as Fragilis) plays an important role in primordial germ cell specification and functions as critical antiviral effector preventing fusion of virion-containing vesicles with endosomal membranes. Results: Here, we identified Ifitm3 as a biomarker of poor clinical outcome in patients with various B-cell malignancies. We found that higher IFITM3 mRNA levels at the time of diagnosis represents a strong predictor of poor clinical outcome for children (COG P9906; P=0.011; n=207) and adults (ECOG E2993; P=0.017; n=55) with B-ALL. Interestingly, IFITM3 is a transcriptional target and strongly repressed by IKZF1 (Ikaros) a potent tumor suppressor in B-ALL and high IFITM3 mRNA levels represents a biomarker for patients with IKZF1-deletion. To study the function of Ifitm3 in a model for human pre-B ALL, pre-B cells from Ifitm3-/- mice were transformed with BCR-ABL1 or oncogenic NRASG12D. Strikingly, deletion of Ifitm3 resulted in destabilization of lipid rafts, loss of CD19 surface expression and loss of PI3K signaling. Ifitm3-/- leukemia cells could not sustain oncogenic signaling from BCR-ABL1 or oncogenic NRASG12D and failed to initiate fatal leukemia in transplant recipient mice. These changes were paralleled by G0/1 cell cycle arrest (P<0.001), loss of colony formation capacity (P=0.0004) and increased propensity to apoptosis. In mechanistic studies, we identified N-terminal phosphorylation at endocytic motif (20YEML23) by Src-kinases induced Ifitm3 cell surface accumulation during normal activation or oncogenic transformation of B-cells. In a lipid-binding assay in vitro, recombinant IFITM3 directly bound to PI(3,4,5)P3 but not any other phospholipids. In the cell membrane, therefore, IFITM3 functioned as a scaffold for PI(3,4,5)P3 and was essential for Src-kinase and PI3K signaling, as well as lipid raft formation and surface expression of multiple raft-associated receptors. Conversely, inducible overexpression of IKZF1 transcriptionally silenced IFITM3, resulting in loss of IFITM3 expression, reduction of lipid rafts and impairment of Src-kinase signaling. In the absence of Ifitm3, resting B-cell populations developed normally. However, consistent with defective Src-kinase and PI3K signaling, Ifitm3-/- B-cells failed to form germinal centers and to give rise to antigen-specific humoral immune responses. Likewise, Ifitm3-/- B-cell precursors were resistant to malignant transformation and lacked the ability to initiate BCR-ABL1- and NRASG12D-driven leukemia. Conversely, an Ifitm3-phosphomimetic of Src-kinase phosphorylation induced constitutive cell membrane localization, triggered oncogenic Src-PI3K signaling and initiated overt leukemia in pre-malignant B-cells. Conclusions: We conclude that Src-kinase-mediated phosphorylation of Ifitm3 induces a dynamic switch from antiviral effector functions in endosomes to lipid raft signaling at the cell membrane. While membrane-bound Ifitm3 is critical for normal B-cell receptor signaling and antigen-specific B-cell responses, its role as signal amplifier can be leveraged by multiple oncogenes for malignant B-cell transformation. Figure Disclosures: Nix: UCSF: Patents & Royalties. Chen:Genovel Biotech Corp: Other: scientific founder and Chairman. Wiita:UCSF: Patents & Royalties; Indapta Therapeutics: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Protocol Intelligence: Equity Ownership, Membership on an entity's Board of Directors or advisory committees.