The N501Y spike substitution enhances SARS-CoV-2 infection and transmission.

The B.1.1.7 variant (also known as Alpha) of SARS-CoV-2, the cause of the COVID-19 pandemic, emerged in the UK in the summer of 2020. The prevalence of this variant increased rapidly owing to an increase in infection and/or transmission efficiency1. The Alpha variant contains 19 nonsynonymous mutations across its viral genome, including 8 substitutions or deletions in the spike protein that interacts with cellular receptors to mediate infection and tropism. Here, using a reverse genetics approach, we show that of the 8 individual spike protein substitutions, only N501Y resulted in consistent fitness gains for replication in the upper airway in a hamster model as well as in primary human airway epithelial cells. The N501Y substitution recapitulated the enhanced viral transmission phenotype of the eight mutations in the Alpha spike protein, suggesting that it is a major determinant of the increased transmission of the Alpha variant. Mechanistically, the N501Y substitution increased the affinity of the viral spike protein for cellular receptors. As suggested by its convergent evolution in Brazil, South Africa and elsewhere2,3, our results indicate that N501Y substitution is an adaptive spike mutation of major concern.

Nasal delivery of an IgM offers broad protection from SARS-CoV-2 variants.

Resistance represents a major challenge for antibody-based therapy for COVID-191-4. Here we engineered an immunoglobulin M (IgM) neutralizing antibody (IgM-14) to overcome the resistance encountered by immunoglobulin G (IgG)-based therapeutics. IgM-14 is over 230-fold more potent than its parental IgG-14 in neutralizing SARS-CoV-2. IgM-14 potently neutralizes the resistant virus raised by its corresponding IgG-14, three variants of concern-B.1.1.7 (Alpha, which first emerged in the UK), P.1 (Gamma, which first emerged in Brazil) and B.1.351 (Beta, which first emerged in South Africa)-and 21 other receptor-binding domain mutants, many of which are resistant to the IgG antibodies that have been authorized for emergency use. Although engineering IgG into IgM enhances antibody potency in general, selection of an optimal epitope is critical for identifying the most effective IgM that can overcome resistance. In mice, a single intranasal dose of IgM-14 at 0.044 mg per kg body weight confers prophylactic efficacy and a single dose at 0.4 mg per kg confers therapeutic efficacy against SARS-CoV-2. IgM-14, but not IgG-14, also confers potent therapeutic protection against the P.1 and B.1.351 variants. IgM-14 exhibits desirable pharmacokinetics and safety profiles when administered intranasally in rodents. Our results show that intranasal administration of an engineered IgM can improve efficacy, reduce resistance and simplify the prophylactic and therapeutic treatment of COVID-19.

Spike mutation D614G alters SARS-CoV-2 fitness.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein substitution D614G became dominant during the coronavirus disease 2019 (COVID-19) pandemic1,2. However, the effect of this variant on viral spread and vaccine efficacy remains to be defined. Here we engineered the spike D614G substitution in the USA-WA1/2020 SARS-CoV-2 strain, and found that it enhances viral replication in human lung epithelial cells and primary human airway tissues by increasing the infectivity and stability of virions. Hamsters infected with SARS-CoV-2 expressing spike(D614G) (G614 virus) produced higher infectious titres in nasal washes and the trachea, but not in the lungs, supporting clinical evidence showing that the mutation enhances viral loads in the upper respiratory tract of COVID-19 patients and may increase transmission. Sera from hamsters infected with D614 virus exhibit modestly higher neutralization titres against G614 virus than against D614 virus, suggesting that the mutation is unlikely to reduce the ability of vaccines in clinical trials to protect against COVID-19, and that therapeutic antibodies should be tested against the circulating G614 virus. Together with clinical findings, our work underscores the importance of this variant in viral spread and its implications for vaccine efficacy and antibody therapy.

Loss of furin cleavage site attenuates SARS-CoV-2 pathogenesis.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-a new coronavirus that has led to a worldwide pandemic1-has a furin cleavage site (PRRAR) in its spike protein that is absent in other group-2B coronaviruses2. To explore whether the furin cleavage site contributes to infection and pathogenesis in this virus, we generated a mutant SARS-CoV-2 that lacks the furin cleavage site (ΔPRRA). Here we report that replicates of ΔPRRA SARS-CoV-2 had faster kinetics, improved fitness in Vero E6 cells and reduced spike protein processing, as compared to parental SARS-CoV-2. However, the ΔPRRA mutant had reduced replication in a human respiratory cell line and was attenuated in both hamster and K18-hACE2 transgenic mouse models of SARS-CoV-2 pathogenesis. Despite reduced disease, the ΔPRRA mutant conferred protection against rechallenge with the parental SARS-CoV-2. Importantly, the neutralization values of sera from patients with coronavirus disease 2019 (COVID-19) and monoclonal antibodies against the receptor-binding domain of SARS-CoV-2 were lower against the ΔPRRA mutant than against parental SARS-CoV-2, probably owing to an increased ratio of particles to plaque-forming units in infections with the former. Together, our results demonstrate a critical role for the furin cleavage site in infection with SARS-CoV-2 and highlight the importance of this site for evaluating the neutralization activities of antibodies.

A potent and broad neutralization of SARS-CoV-2 variants of concern by DARPins.

We report the engineering and selection of two synthetic proteins-FSR16m and FSR22-for the possible treatment of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. FSR16m and FSR22 are trimeric proteins composed of DARPin SR16m or SR22 fused with a T4 foldon. Despite selection by a spike protein from a now historical SARS-CoV-2 strain, FSR16m and FSR22 exhibit broad-spectrum neutralization of SARS-CoV-2 strains, inhibiting authentic B.1.351, B.1.617.2 and BA.1.1 viruses, with respective IC50 values of 3.4, 2.2 and 7.4 ng ml-1 for FSR16m. Cryo-EM structures revealed that these DARPins recognize a region of the receptor-binding domain (residues 456, 475, 486, 487 and 489) overlapping a critical portion of the angiotensin-converting enzyme 2 (ACE2)-binding surface. K18-hACE2 transgenic mice inoculated with B.1.617.2 and receiving intranasally administered FSR16m showed less weight loss and 10-100-fold lower viral burden in upper and lower respiratory tracts. The strong and broad neutralization potency makes FSR16m and FSR22 promising candidates for the prevention and treatment of infection by SARS-CoV-2.

Recognition of a highly conserved glycoprotein B epitope by a bivalent antibody neutralizing HCMV at a post-attachment step.

Human cytomegalovirus (HCMV) is one of the main causative agents of congenital viral infection in neonates. HCMV infection also causes serious morbidity and mortality among organ transplant patients. Glycoprotein B (gB) is a major target for HCMV neutralizing antibodies, yet the underlying neutralization mechanisms remain largely unknown. Here we report that 3-25, a gB-specific monoclonal antibody previously isolated from a healthy HCMV-positive donor, efficiently neutralized 14 HCMV strains in both ARPE-19 cells and MRC-5 cells. The core epitope of 3-25 was mapped to a highly conserved linear epitope on antigenic domain 2 (AD-2) of gB. A 1.8 Å crystal structure of 3-25 Fab in complex with the peptide epitope revealed the molecular determinants of 3-25 binding to gB at atomic resolution. Negative-staining electron microscopy (EM) 3D reconstruction of 3-25 Fab in complex with de-glycosylated postfusion gB showed that 3-25 Fab fully occupied the gB trimer at the N-terminus with flexible binding angles. Functionally, 3-25 efficiently inhibited HCMV infection at a post-attachment step by interfering with viral membrane fusion, and restricted post-infection viral spreading in ARPE-19 cells. Interestingly, bivalency was required for HCMV neutralization by AD-2 specific antibody 3-25 but not the AD-4 specific antibody LJP538. In contrast, bivalency was not required for HCMV binding by both antibodies. Taken together, our results reveal the structural basis of gB recognition by 3-25 and demonstrate that inhibition of viral membrane fusion and a requirement of bivalency may be common for gB AD-2 specific neutralizing antibody.

Potent Bispecific Neutralizing Antibody Targeting Glycoprotein B and the gH/gL/pUL128/130/131 Complex of Human Cytomegalovirus.

Human cytomegalovirus (HCMV) is a ubiquitous pathogen that can cause developmental disorders following congenital infection and life-threatening complications among transplant patients. Potent neutralizing monoclonal antibodies (MAbs) are promising drug candidates against HCMV infection. HCMV can infect a broad range of cell types. Therefore, single neutralizing antibodies targeting one HCMV glycoprotein often lack either potency or broad cell-type coverage. We previously characterized two human-derived HCMV neutralizing MAbs. One was the broadly neutralizing MAb 3-25, which targets the antigenic domain 2 of glycoprotein B (gB). The other was the highly potent MAb 2-18, which specifically recognizes the gH/gL/pUL128/130/131 complex (pentamer). To combine the strengths of gB- and pentamer-targeting MAbs, we developed an IgG-single-chain variable fragment (scFv) bispecific antibody by fusing the 2-18 scFv to the heavy-chain C terminus of MAb 3-25. The resulting bispecific antibody showed high-affinity binding to both gB and pentamer. Functionally, the bispecific antibody demonstrated a combined neutralization breadth and potency of the parental MAbs in multiple cell lines and inhibited postinfection viral spreading. Furthermore, the bispecific antibody was easily produced in CHO cells at a yield above 1 g/liter and showed a single-dose pharmacokinetic profile comparable to that of parental MAb 3-25 in rhesus macaques. Importantly, the bispecific antibody retained broadly and potent neutralizing activity after 21 days in circulation. Taken together, our research provides a proof-of-concept study for developing bispecific neutralizing antibody therapies against HCMV infection.

Identification of adipocyte plasma membrane-associated protein as a novel modulator of human cytomegalovirus infection.

Human cytomegalovirus (HCMV) is a ubiquitous pathogen that can cause disability in newborns and serious clinical diseases in immunocompromised patients. HCMV has a large genome with enormous coding potential; its viral particles are equipped with complicated glycoprotein complexes and can infect a wide range of human cells. Although multiple host cellular receptors interacting with viral glycoproteins have been reported, the mechanism of HCMV infection remains a mystery. Here we report identification of adipocyte plasma membrane-associated protein (APMAP) as a novel modulator active in the early stage of HCMV infection. APMAP is necessary for HCMV infection in both epithelial cells and fibroblasts; knockdown of APMAP expression significantly reduced HCMV infection of these cells. Interestingly, ectopic expression of human APMAP in cells refractory to HCMV infection, such as canine MDCK and murine NIH/3T3 cells, promoted HCMV infection. Furthermore, reduction in viral immediate early (IE) gene transcription at 6 h post infection and delayed nucleus translocation of tegument delivered pp65 at 4 h post infection were detected in APMAP-deficient cells but not in the wildtype cells. These results suggest that APMAP plays a role in the early stage of HCMV infection. Results from biochemical studies of APMAP and HCMV proteins suggest that APMAP could participate in HCMV infection through interaction with gH/gL containing glycoprotein complexes at low pH and mediate nucleus translocation of tegument pp65. Taken together, our results suggest that APMAP functions as a modulator promoting HCMV infection in multiple cell types and is an important player in the complex HCMV infection mechanism.

Structure, Immunogenicity, and Protective Mechanism of an Engineered Enterovirus 71-Like Particle Vaccine Mimicking 80S Empty Capsid.

Enterovirus 71 (EV71) is the major causative agent of severe hand, foot, and mouth disease, which affects millions of young children in the Asia-Pacific region annually. In this study, we engineered a novel EV71 virus-like particle (VLP) that lacks VP4 (therefore designated VLPΔVP4) and investigated its structure, antigenicity, and vaccine potential. The cryo-electron microscopy (cryo-EM) structure of VLPΔVP4 was reconstructed to 3.71-Å resolution. Results from structural and biochemical analyses revealed that VLPΔVP4 resembles the end product of the viral uncoating process, the 80S empty capsid. VLPΔVP4 is able to elicit high-titer neutralizing antibodies and to fully protect mice against lethal viral challenge. Mechanistic studies showed that, at the cellular level, the anti-VLPΔVP4 sera exert neutralization effects at both pre- and postattachment stages by inhibiting both virus attachment and internalization, and at the molecular level, the antisera can block multiple interactions between EV71 and its key receptors. Our study gives a better understanding of EV71 capsid assembly and provides important information for the design and development of new-generation vaccines for EV71, and perhaps for other enteroviruses, as well.IMPORTANCE Enterovirus 71 (EV71) infection may lead to severe hand, foot, and mouth disease, with significant morbidity and mortality. Knowledge regarding EV71 particle assembly remains limited. Here, we report the generation and characterization of a novel EV71 virus-like particle that lacks the VP4 capsid subunit protein. This particle, termed VLPΔVP4, structurally mimics the 80S empty capsid, which is the end stage of EV71 uncoating. We further show that VLPΔVP4 exhibits desirable immunogenicity and protective efficacy in proof-of-concept studies. In addition, the inhibitory mechanisms of the VLPΔVP4-induced antibodies are unraveled at both the cellular and molecular levels. Our work provides the first evidence of picornaviral particle assembly in the complete absence of VP4 and identifies VLPΔVP4 as an improved EV71 vaccine candidate with desirable traits. These findings not only enhance our understanding of particle assembly and uncoating of picornaviruses, but also provide important information for structure-guided vaccine design for EV71 and other enteroviruses.

Beta-Propiolactone Inactivation of Coxsackievirus A16 Induces Structural Alteration and Surface Modification of Viral Capsids.

Beta-propiolactone (BPL) is an inactivating agent that is widely used in the vaccine industry. However, its effects on vaccine protein antigens and its mechanisms of action remain poorly understood. Here we present cryo-electron microscopy (cryo-EM) structures of BPL-treated coxsackievirus A16 (CVA16) mature virions and procapsids at resolutions of 3.9 Å and 6.5 Å, respectively. Notably, both particles were found to adopt an expanded conformation resembling the 135S-like uncoating intermediate, with characteristic features including an opened 2-fold channel, the externalization of the N terminus of VP1 capsid protein, and the absence of pocket factor. However, major neutralizing epitopes are very well preserved on these particles. Further biochemical analyses revealed that BPL treatment impairs the abilities of CVA16 particles to bind to the attachment receptor heparan sulfate and to a conformation-dependent monoclonal antibody in a BPL dose-dependent manner, indicating that BPL is able to modify surface-exposed amino acid residues. Taken together, our results demonstrate that BPL treatment may induce alteration of the overall structure and surface properties of a nonenveloped viral capsid, thus revealing a novel mode of action of BPL.IMPORTANCE Beta-propiolactone (BPL) is commonly used as an inactivating reagent to produce viral vaccines. It is recognized that BPL inactivates viral infectivity through modification of viral nucleic acids. However, its effect on viral proteins remains largely unknown. Here, we present high-resolution cryo-EM structures of BPL-treated coxsackievirus A16 (CVA16) mature virions and procapsids, which reveals an expanded overall conformation and characteristic features that are typical for the 135S-like uncoating intermediate. We further show that the BPL concentration affects the binding of inactivated CVA16 particles to their receptor/antibody. Thus, BPL treatment can alter the overall structure and surface properties of viral capsids, which may lead to antigenic and immunogenic variations. Our findings provide important information for future development of BPL-inactivated vaccines.

Structural Basis for Recognition of Human Enterovirus 71 by a Bivalent Broadly Neutralizing Monoclonal Antibody.

Enterovirus 71 (EV71) is the main pathogen responsible for hand, foot and mouth disease with severe neurological complications and even death in young children. We have recently identified a highly potent anti-EV71 neutralizing monoclonal antibody, termed D5. Here we investigated the structural basis for recognition of EV71 by the antibody D5. Four three-dimensional structures of EV71 particles in complex with IgG or Fab of D5 were reconstructed by cryo-electron microscopy (cryo-EM) single particle analysis all at subnanometer resolutions. The most critical EV71 mature virion-Fab structure was resolved to a resolution of 4.8 Å, which is rare in cryo-EM studies of virus-antibody complex so far. The structures reveal a bivalent binding pattern of D5 antibody across the icosahedral 2-fold axis on mature virion, suggesting that D5 binding may rigidify virions to prevent their conformational changes required for subsequent RNA release. Moreover, we also identified that the complementary determining region 3 (CDR3) of D5 heavy chain directly interacts with the extremely conserved VP1 GH-loop of EV71, which was validated by biochemical and virological assays. We further showed that D5 is indeed able to neutralize a variety of EV71 genotypes and strains. Moreover, D5 could potently confer protection in a mouse model of EV71 infection. Since the conserved VP1 GH-loop is involved in EV71 binding with its uncoating receptor, the scavenger receptor class B, member 2 (SCARB2), the broadly neutralizing ability of D5 might attribute to its inhibition of EV71 from binding SCARB2. Altogether, our results elucidate the structural basis for the binding and neutralization of EV71 by the broadly neutralizing antibody D5, thereby enhancing our understanding of antibody-based protection against EV71 infection.

Single Neutralizing Monoclonal Antibodies Targeting the VP1 GH Loop of Enterovirus 71 Inhibit both Virus Attachment and Internalization during Viral Entry.

UNLABELLED: Antibodies play a critical role in immunity against enterovirus 71 (EV71). However, how EV71-specific antibodies neutralize infections remains poorly understood. Here we report the working mechanism for a group of three monoclonal antibodies (MAbs) that potently neutralize EV71. We found that these three MAbs (termed D5, H7, and C4, respectively) recognize the same conserved neutralizing epitope within the VP1 GH loop of EV71. Single MAbs in this group, exemplified by D5, could inhibit EV71 infection in cell cultures at both the pre- and postattachment stages in a cell type-independent manner. Specifically, MAb treatment resulted in the blockade of multiple steps of EV71 entry, including virus attachment, internalization, and subsequent uncoating and RNA release. Furthermore, we show that the D5 and C4 antibodies can interfere with EV71 binding to its key receptors, including heparan sulfate, SCARB2, and PSGL-1, thus providing a possible explanation for the observed multi-inhibitory function of the MAbs. Collectively, our study unravels the mechanism of neutralization by a unique group of anti-EV71 MAbs targeting the conserved VP1 GH loop. The findings should enhance our understanding of MAb-mediated immunity against enterovirus infections and accelerate the development of MAb-based anti-EV71 therapeutic drugs. IMPORTANCE: Enterovirus 71 (EV71) is a major causative agent of hand, foot, and mouth disease (HFMD), which has caused significant morbidities and mortalities in young children. Neither a vaccine nor an antiviral drug is available. Neutralizing antibodies are major protective components in EV71 immunity. Here, we unraveled an unusual mechanism of EV71 neutralization by a group of three neutralizing monoclonal antibodies (MAbs). All of these MAbs bound the same conserved epitope located at the VP1 GH loop of EV71. Interestingly, mechanistic studies showed that single antibodies in this MAb group could block EV71 attachment and internalization during the viral entry process and interfere with EV71 binding to heparan sulfate, SCARB2, and PSGL-1 molecules, which are key receptors involved in different steps of EV71 entry. Our findings greatly enhance the understanding of the interplays among EV71, neutralizing antibodies, and host receptors, which in turn should facilitate the development of an MAb-based anti-EV71 therapy.

Chimeric virus-like particle vaccines displaying conserved enterovirus 71 epitopes elicit protective neutralizing antibodies in mice through divergent mechanisms.

Enterovirus 71 (EV71) is a major causative agent of hand, food, and mouth disease, which frequently occurs in young children. Since there are 11 subgenotypes (A, B1 to B5, and C1 to C5) within EV71, an EV71 vaccine capable of protecting against all of these subgenotypes is desirable. We report here the vaccine potential and protective mechanism of two chimeric virus-like particles (VLPs) presenting conserved neutralizing epitopes of EV71. We show that fusions of hepatitis B core antigen (HBc) with the SP55 or SP70 epitope of EV71, designated HBcSP55 and HBcSP70, respectively, can be rapidly generated and self-assembled into VLPs with the epitopes displayed on the surface. Immunization with the chimeric VLPs induced carrier- and epitope-specific antibody responses in mice. Anti-HBcSP55 and anti-HBcSP70 sera, but not anti-HBc sera, were able to neutralize in vitro multiple genotypes and strains of EV71. Importantly, passive immunization with anti-HBcSP55 or anti-HBcSP70 sera protected neonatal mice against lethal EV71 infections. Interestingly, anti-HBcSP70 sera could inhibit EV71 attachment to susceptible cells, whereas anti-HBcSP55 sera could not. However, both antisera were able to neutralize EV71 infection in vitro at the postattachment stage. The divergent mechanism of neutralization and protection conferred by anti-SP70 and anti-SP55 sera is in part attributed to their respective ability to bind authentic viral particles. Collectively, our study not only demonstrates that chimeric VLPs displaying the SP55 and SP70 epitopes are promising candidates for a broad-spectrum EV71 vaccine but also reveals distinct mechanisms of neutralization by the SP55- and SP70-targeted antibodies.

Blocking Oncostatin M receptor abrogates STAT3 mediated integrin signaling and overcomes chemoresistance in ovarian cancer.

Chemotherapy such as cisplatin is widely used to treat ovarian cancer either before or after surgical debulking. However, cancer relapse due to chemotherapy resistance is a major challenge in the treatment of ovarian cancer. The underlying mechanisms related to chemotherapy resistance remain largely unclear. Therefore, identification of effective therapeutic strategies is urgently needed to overcome therapy resistance. Transcriptome-based analysis, in vitro studies and functional assays identified that cisplatin-resistant ovarian cancer cells express high levels of OSMR compared to cisplatin sensitive cells. Furthermore, OSMR expression associated with a module of integrin family genes and predominantly linked with integrin αV (ITGAV) and integrin ß3 (ITGB3) for cisplatin resistance. Using ectopic expression and knockdown approaches, we proved that OSMR directly regulates ITGAV and ITGB3 gene expression through STAT3 activation. Notably, targeting OSMR using anti-OSMR human antibody inhibited the growth and metastasis of ovarian cancer cells and sensitized cisplatin treatment. Taken together, our results underscore the pivotal role of OSMR as a requirement for cisplatin resistance in ovarian cancer. Notably, OSMR fostered the expression of a distinct set of integrin genes, which in turn resulted into a crosstalk between OSMR and integrins for signaling activation that is critical for cisplatin resistance. Therefore, targeting OSMR emerges as a promising and viable strategy to reverse cisplatin-resistance in ovarian cancer.

Transcriptional signature of durable effector T cells elicited by a replication defective HCMV vaccine.

Human cytomegalovirus (HCMV) is a leading infectious cause of birth defects and the most common opportunistic infection that causes life-threatening diseases post-transplantation; however, an effective vaccine remains elusive. V160 is a live-attenuated replication defective HCMV vaccine that showed a 42.4% efficacy against primary HCMV infection among seronegative women in a phase 2b clinical trial. Here, we integrated the multicolor flow cytometry, longitudinal T cell receptor (TCR) sequencing, and single-cell RNA/TCR sequencing approaches to characterize the magnitude, phenotype, and functional quality of human T cell responses to V160. We demonstrated that V160 de novo induces IE-1 and pp65 specific durable polyfunctional effector CD8 T cells that are comparable to those induced by natural HCMV infection. We identified a variety of V160-responsive T cell clones which exhibit distinctive "transient" and "durable" expansion kinetics, and revealed a transcriptional signature that marks durable CD8 T cells post-vaccination. Our study enhances the understanding of human T-cell immune responses to V160 vaccination.

An ACE2 decamer viral trap as a durable intervention solution for current and future SARS-CoV.

The capacity of SARS-CoV-2 to evolve poses challenges to conventional prevention and treatment options such as vaccination and monoclonal antibodies, as they rely on viral receptor binding domain (RBD) sequences from previous strains. Additionally, animal CoVs, especially those of the SARS family, are now appreciated as a constant pandemic threat. We present here a new antiviral approach featuring inhalation delivery of a recombinant viral trap composed of ten copies of angiotensin-converting enzyme 2 (ACE2) fused to the IgM Fc. This ACE2 decamer viral trap is designed to inhibit SARS-CoV-2 entry function, regardless of viral RBD sequence variations as shown by its high neutralization potency against all known SARS-CoV-2 variants, including Omicron BQ.1, BQ.1.1, XBB.1 and XBB.1.5. In addition, it demonstrates potency against SARS-CoV-1, human NL63, as well as bat and pangolin CoVs. The multivalent trap is effective in both prophylactic and therapeutic settings since a single intranasal dosing confers protection in human ACE2 transgenic mice against viral challenges. Lastly, this molecule is stable at ambient temperature for more than twelve weeks and can sustain physical stress from aerosolization. These results demonstrate the potential of a decameric ACE2 viral trap as an inhalation solution for ACE2-dependent coronaviruses of current and future pandemic concerns.

Overcoming adaptive resistance to anti-VEGF therapy by targeting CD5L.

Antiangiogenic treatment targeting the vascular endothelial growth factor (VEGF) pathway is a powerful tool to combat tumor growth and progression; however, drug resistance frequently emerges. We identify CD5L (CD5 antigen-like precursor) as an important gene upregulated in response to antiangiogenic therapy leading to the emergence of adaptive resistance. By using both an RNA-aptamer and a monoclonal antibody targeting CD5L, we are able to abate the pro-angiogenic effects of CD5L overexpression in both in vitro and in vivo settings. In addition, we find that increased expression of vascular CD5L in cancer patients is associated with bevacizumab resistance and worse overall survival. These findings implicate CD5L as an important factor in adaptive resistance to antiangiogenic therapy and suggest that modalities to target CD5L have potentially important clinical utility.

Recent progress in development of monoclonal antibodies against human cytomegalovirus.

Human cytomegalovirus (HCMV) is a ubiquitous pathogen that can cause permanent childhood disabilities following in utero infection and life threatening diseases in immune-compromised individuals such as those post transplantation. Without an effective vaccine, small molecule antiviral drugs are routinely used in high-risk transplant recipients, but the effectiveness of which is limited by side effects and drug resistance. The potentials of antibody-based passive immune therapies alone or in combination with the small molecule antivirals to treat or prevent HCMV infection have been actively studied. In this review, we focus on the recent publications on identification and characterization of monoclonal antibodies that have the potential to be developed as anti-HCMV therapies. We review the progress in clinical evaluation of antibody-based therapies to prevent HCMV-associated diseases.

Passive Immunotherapy Against SARS-CoV-2: From Plasma-Based Therapy to Single Potent Antibodies in the Race to Stay Ahead of the Variants.

The COVID-19 pandemic is now approaching 2 years old, with more than 440 million people infected and nearly six million dead worldwide, making it the most significant pandemic since the 1918 influenza pandemic. The severity and significance of SARS-CoV-2 was recognized immediately upon discovery, leading to innumerable companies and institutes designing and generating vaccines and therapeutic antibodies literally as soon as recombinant SARS-CoV-2 spike protein sequence was available. Within months of the pandemic start, several antibodies had been generated, tested, and moved into clinical trials, including Eli Lilly's bamlanivimab and etesevimab, Regeneron's mixture of imdevimab and casirivimab, Vir's sotrovimab, Celltrion's regdanvimab, and Lilly's bebtelovimab. These antibodies all have now received at least Emergency Use Authorizations (EUAs) and some have received full approval in select countries. To date, more than three dozen antibodies or antibody combinations have been forwarded into clinical trials. These antibodies to SARS-CoV-2 all target the receptor-binding domain (RBD), with some blocking the ability of the RBD to bind human ACE2, while others bind core regions of the RBD to modulate spike stability or ability to fuse to host cell membranes. While these antibodies were being discovered and developed, new variants of SARS-CoV-2 have cropped up in real time, altering the antibody landscape on a moving basis. Over the past year, the search has widened to find antibodies capable of neutralizing the wide array of variants that have arisen, including Alpha, Beta, Gamma, Delta, and Omicron. The recent rise and dominance of the Omicron family of variants, including the rather disparate BA.1 and BA.2 variants, demonstrate the need to continue to find new approaches to neutralize the rapidly evolving SARS-CoV-2 virus. This review highlights both convalescent plasma- and polyclonal antibody-based approaches as well as the top approximately 50 antibodies to SARS-CoV-2, their epitopes, their ability to bind to SARS-CoV-2 variants, and how they are delivered. New approaches to antibody constructs, including single domain antibodies, bispecific antibodies, IgA- and IgM-based antibodies, and modified ACE2-Fc fusion proteins, are also described. Finally, antibodies being developed for palliative care of COVID-19 disease, including the ramifications of cytokine release syndrome (CRS) and acute respiratory distress syndrome (ARDS), are described.