Human Antibodies for Viral Infections.

Antibodies have been used to prevent or treat viral infections since the nineteenth century, but the full potential to use passive immunization for infectious diseases has yet to be realized. The advent of efficient methods for isolating broad and potently neutralizing human monoclonal antibodies is enabling us to develop antibodies with unprecedented activities. The discovery of IgG Fc region modifications that extend antibody half-life in humans to three months or more suggests that antibodies could become the principal tool with which we manage future viral epidemics. Antibodies for members of most virus families that cause severe disease in humans have been isolated, and many of them are in clinical development, an area that has accelerated during the effort to prevent or treat COVID-19 (coronavirus disease 2019). Broad and potently neutralizing antibodies are also important research reagents for identification of protective epitopes that can be engineered into active vaccines through structure-based reverse vaccinology.

Broadly Neutralizing Antibodies to HIV and Their Role in Vaccine Design.

HIV employs multiple means to evade the humoral immune response, particularly the elicitation of and recognition by broadly neutralizing antibodies (bnAbs). Such antibodies can act antivirally against a wide spectrum of viruses by targeting relatively conserved regions on the surface HIV envelope trimer spike. Elicitation of and recognition by bnAbs are hindered by the arrangement of spikes on virions and the relatively difficult access to bnAb epitopes on spikes, including the proximity of variable regions and a high density of glycans. Yet, in a small proportion of HIV-infected individuals, potent bnAb responses do develop, and isolation of the corresponding monoclonal antibodies has been facilitated by identification of favorable donors with potent bnAb sera and by development of improved methods for human antibody generation. Molecular studies of recombinant Env trimers, alone and in interaction with bnAbs, are providing new insights that are fueling the development and testing of promising immunogens aimed at the elicitation of bnAbs.

Respiratory mucosal delivery of next-generation COVID-19 vaccine provides robust protection against both ancestral and variant strains of SARS-CoV-2.

The emerging SARS-CoV-2 variants of concern (VOCs) threaten the effectiveness of current COVID-19 vaccines administered intramuscularly and designed to only target the spike protein. There is a pressing need to develop next-generation vaccine strategies for broader and long-lasting protection. Using adenoviral vectors (Ad) of human and chimpanzee origin, we evaluated Ad-vectored trivalent COVID-19 vaccines expressing spike-1, nucleocapsid, and RdRp antigens in murine models. We show that single-dose intranasal immunization, particularly with chimpanzee Ad-vectored vaccine, is superior to intramuscular immunization in induction of the tripartite protective immunity consisting of local and systemic antibody responses, mucosal tissue-resident memory T cells and mucosal trained innate immunity. We further show that intranasal immunization provides protection against both the ancestral SARS-CoV-2 and two VOC, B.1.1.7 and B.1.351. Our findings indicate that respiratory mucosal delivery of Ad-vectored multivalent vaccine represents an effective next-generation COVID-19 vaccine strategy to induce all-around mucosal immunity against current and future VOC.

Antibody-mediated neutralization of SARS-CoV-2.

Neutralizing antibodies can block infection, clear pathogens, and are essential to provide long-term immunity. Since the onset of the pandemic, SARS-CoV-2 neutralizing antibodies have been comprehensively investigated and critical information on their development, function, and potential use to prevent and treat COVID-19 have been revealed. With the emergence of SARS-CoV-2 immune escape variants, humoral immunity is being challenged, and a detailed understanding of neutralizing antibodies is essential to guide vaccine design strategies as well as antibody-mediated therapies. In this review, we summarize some of the key findings on SARS-CoV-2 neutralizing antibodies, with a focus on their clinical application.

Membrane-bound mRNA immunogens lower the threshold to activate HIV Env V2 apex-directed broadly neutralizing B cell precursors in humanized mice.

Eliciting broadly neutralizing antibodies (bnAbs) is the core of HIV vaccine design. bnAbs specific to the V2-apex region of the HIV envelope acquire breadth and potency with modest somatic hypermutation, making them attractive vaccination targets. To evaluate Apex germline-targeting (ApexGT) vaccine candidates, we engineered knockin (KI) mouse models expressing the germline B cell receptor (BCR) of the bnAb PCT64. We found that high affinity of the ApexGT immunogen for PCT64-germline BCRs was necessary to specifically activate KI B cells at human physiological frequencies, recruit them to germinal centers, and select for mature bnAb mutations. Relative to protein, mRNA-encoded membrane-bound ApexGT immunization significantly increased activation and recruitment of PCT64 precursors to germinal centers and lowered their affinity threshold. We have thus developed additional models for HIV vaccine research, validated ApexGT immunogens for priming V2-apex bnAb precursors, and identified mRNA-LNP as a suitable approach to substantially improve the B cell response.

Vaccination in a humanized mouse model elicits highly protective PfCSP-targeting anti-malarial antibodies.

Repeat antigens, such as the Plasmodium falciparum circumsporozoite protein (PfCSP), use both sequence degeneracy and structural diversity to evade the immune response. A few PfCSP-directed antibodies have been identified that are effective at preventing malaria infection, including CIS43, but how these repeat-targeting antibodies might be improved has been unclear. Here, we engineered a humanized mouse model in which B cells expressed inferred human germline CIS43 (iGL-CIS43) B cell receptors and used both vaccination and bioinformatic analysis to obtain variant CIS43 antibodies with improved protective capacity. One such antibody, iGL-CIS43.D3, was significantly more potent than the current best-in-class PfCSP-directed antibody. We found that vaccination with a junctional epitope peptide was more effective than full-length PfCSP at recruiting iGL-CIS43 B cells to germinal centers. Structure-function analysis revealed multiple somatic hypermutations that combinatorically improved protection. This mouse model can thus be used to understand vaccine immunogens and to develop highly potent anti-malarial antibodies.

Functional convergence of a germline-encoded neutralizing antibody response in rhesus macaques immunized with HCV envelope glycoproteins.

Human IGHV1-69-encoded broadly neutralizing antibodies (bnAbs) that target the hepatitis C virus (HCV) envelope glycoprotein (Env) E2 are important for protection against HCV infection. An IGHV1-69 ortholog gene, VH1.36, is preferentially used for bnAbs isolated from HCV Env-immunized rhesus macaques (RMs). Here, we studied the genetic, structural, and functional properties of VH1.36-encoded bnAbs generated by vaccination, in comparison to IGHV1-69-encoded bnAbs from HCV patients. Global B cell repertoire analysis confirmed the expansion of VH1.36-derived B cells in immunized animals. Most E2-specific, VH1.36-encoded antibodies cross-neutralized HCV. Crystal structures of two RM bnAbs with E2 revealed that the RM bnAbs engaged conserved E2 epitopes using similar molecular features as human bnAbs but with a different binding mode. Longitudinal analyses of the RM antibody repertoire responses during immunization indicated rapid lineage development of VH1.36-encoded bnAbs with limited somatic hypermutation. Our findings suggest functional convergence of a germline-encoded bnAb response to HCV Env with implications for vaccination in humans.

T Cells in Nonlymphoid Tissues Give Rise to Lymph-Node-Resident Memory T Cells.

Immunosurveillance of secondary lymphoid organs (SLO) is performed by central memory T cells that recirculate through blood. Resident memory T (Trm) cells remain parked in nonlymphoid tissues and often stably express CD69. We recently identified Trm cells within SLO, but the origin and phenotype of these cells remains unclear. Using parabiosis of "dirty" mice, we found that CD69 expression is insufficient to infer stable residence of SLO Trm cells. Restimulation of nonlymphoid memory CD8+ T cells within the skin or mucosa resulted in a substantial increase in bona fide Trm cells specifically within draining lymph nodes. SLO Trm cells derived from emigrants from nonlymphoid tissues and shared some transcriptional and phenotypic signatures associated with nonlymphoid Trm cells. These data indicate that nonlymphoid cells can give rise to SLO Trm cells and suggest vaccination strategies by which memory CD8+ T cell immunosurveillance can be regionalized to specific lymph nodes.

Human papillomavirus vaccination 2020 guideline update: American Cancer Society guideline adaptation.

The American Cancer Society (ACS) presents an adaptation of the current Advisory Committee on Immunization Practices recommendations for human papillomavirus (HPV) vaccination. The ACS recommends routine HPV vaccination between ages 9 and 12 years to achieve higher on-time vaccination rates, which will lead to increased numbers of cancers prevented. Health care providers are encouraged to start offering the HPV vaccine series at age 9 or 10 years. Catch-up HPV vaccination is recommended for all persons through age 26 years who are not adequately vaccinated. Providers should inform individuals aged 22 to 26 years who have not been previously vaccinated or who have not completed the series that vaccination at older ages is less effective in lowering cancer risk. Catch-up HPV vaccination is not recommended for adults aged older than 26 years. The ACS does not endorse the 2019 Advisory Committee on Immunization Practices recommendation for shared clinical decision making for some adults aged 27 through 45 years who are not adequately vaccinated because of the low effectiveness and low cancer prevention potential of vaccination in this age group, the burden of decision making on patients and clinicians, and the lack of sufficient guidance on the selection of individuals who might benefit.

Fcγ Receptor Function and the Design of Vaccination Strategies.

Through specific interactions with distinct types of Fcγ receptors (FcγRs), the Fc domain of immunoglobulin G (IgG) mediates a wide spectrum of immunological functions that influence both innate and adaptive responses. Recent studies indicate that IgG Fc-FcγR interactions are dynamically regulated during an immune response through the control of the Fc-associated glycan structure and Ig subclass composition on the one hand and selective FcγR expression on immune cells on the other, which together determine the capacity of IgG to interact in a cell-type-specific manner with specific members of the FcγR family. Here, we present a framework that synthesizes the current understanding of the contribution of FcγR pathways to the induction and regulation of antibody and T cell responses. Within this context, we discuss vaccination strategies to elicit broad and potent immune responses based on the immunomodulatory properties of Fc-FcγR interactions.

Lymphocyte Circadian Clocks Control Lymph Node Trafficking and Adaptive Immune Responses.

Lymphocytes circulate through lymph nodes (LN) in search for antigen in what is believed to be a continuous process. Here, we show that lymphocyte migration through lymph nodes and lymph occurred in a non-continuous, circadian manner. Lymphocyte homing to lymph nodes peaked at night onset, with cells leaving the tissue during the day. This resulted in strong oscillations in lymphocyte cellularity in lymph nodes and efferent lymphatic fluid. Using lineage-specific genetic ablation of circadian clock function, we demonstrated this to be dependent on rhythmic expression of promigratory factors on lymphocytes. Dendritic cell numbers peaked in phase with lymphocytes, with diurnal oscillations being present in disease severity after immunization to induce experimental autoimmune encephalomyelitis (EAE). These rhythms were abolished by genetic disruption of T cell clocks, demonstrating a circadian regulation of lymphocyte migration through lymph nodes with time-of-day of immunization being critical for adaptive immune responses weeks later.

The Kinase mTORC1 Promotes the Generation and Suppressive Function of Follicular Regulatory T Cells.

Follicular regulatory T (Tfr) cells differentiate from conventional regulatory T (Treg) cells and suppress excessive germinal center (GC) responses by acting on both GC B cells and T follicular helper (Tfh) cells. Here, we examined the impact of mTOR, a serine/threonine protein kinase that senses and integrates diverse environmental cues, on the differentiation and functional competency of Tfr cells in response to protein immunization or viral infection. By genetically deleting Rptor or Rictor, essential components for mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2), respectively, we found that mTORC1 but not mTORC2 is essential for Tfr differentiation. Mechanistically, mTORC1-mediated phosphorylation of the transcription factor STAT3 induced the expression of the transcription factor TCF-1 by promoting STAT3 binding to the Tcf7 5'-regulatory region. Subsequently, TCF-1 bound to the Bcl6 promoter to induce Bcl6 expression, which launched the Tfr cell differentiation program. Thus, mTORC1 initiates Tfr cell differentiation by activating the TCF-1-Bcl-6 axis during immunization or infection.

Uncovering social and environmental factors that increase the burden of climate-sensitive diarrheal infections on children.

Climate-sensitive infectious diseases are an issue of growing concern due to global warming and the related increase in the incidence of extreme weather and climate events. Diarrhea, which is strongly associated with climatic factors, remains among the leading causes of child death globally, disproportionately affecting populations in low- and middle-income countries (LMICs). We use survey data for 51 LMICs between 2000 and 2019 in combination with gridded climate data to estimate the association between precipitation shocks and reported symptoms of diarrheal illness in young children. We account for differences in exposure risk by climate type and explore the modifying role of various social factors. We find that droughts are positively associated with diarrhea in the tropical savanna regions, particularly during the dry season and dry-to-wet and wet-to-dry transition seasons. In the humid subtropical regions, we find that heavy precipitation events are associated with increased risk of diarrhea during the dry season and the transition from dry-to-wet season. Our analysis of effect modifiers highlights certain social vulnerabilities that exacerbate these associations in the two climate zones and present opportunities for public health intervention. For example, we show that stool disposal practices, child feeding practices, and immunizing against the rotavirus modify the association between drought and diarrhea in the tropical savanna regions. In the humid subtropical regions, household's source of water and water disinfection practices modify the association between heavy precipitation and diarrhea. The evidence of effect modification varies depending on the type and duration of the precipitation shock.

Guardians of the oral and nasopharyngeal galaxy: IgA and protection against SARS-CoV-2 infection.

In early 2020, a global emergency was upon us in the form of the coronavirus disease 2019 (COVID-19) pandemic. While horrific in its health, social and economic devastation, one silver lining to this crisis has been a rapid mobilization of cross-institute, and even cross-country teams that shared common goals of learning as much as we could as quickly as possible about the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and how the immune system would respond to both the virus and COVID-19 vaccines. Many of these teams were formed by women who quickly realized that the classical model of "publish first at all costs" was maladaptive for the circumstances and needed to be supplanted by a more collaborative solution-focused approach. This review is an example of a collaboration that unfolded in separate countries, first Canada and the United States, and then also Israel. Not only did the collaboration allow us to cross-validate our results using different hands/techniques/samples, but it also took advantage of different vaccine types and schedules that were rolled out in our respective home countries. The result of this collaboration was a new understanding of how mucosal immunity to SARS-CoV-2 infection vs COVID-19 vaccination can be measured using saliva as a biofluid, what types of vaccines are best able to induce (limited) mucosal immunity, and what are potential correlates of protection against breakthrough infection. In this review, we will share what we have learned about the mucosal immune response to SARS-CoV-2 and to COVID-19 vaccines and provide a perspective on what may be required for next-generation pan-sarbecoronavirus vaccine approaches.

Hemoglobin wonders: a fascinating gas transporter dive into molluscs.

Hemoglobin (Hb) has been identified in at least 14 molluscan taxa so far. Research spanning over 130 years on molluscan Hbs focuses on their genes, protein structures, functions, and evolution. Molluscan Hbs are categorized into single-, two-, and multiple-domain chains, including red blood cell, gill, and extracellular Hbs, based on the number of globin domains and their respective locations. These Hbs exhibit variation in assembly, ranging from monomeric and dimeric to higher-order multimeric forms. Typically, molluscan Hbs display moderately high oxygen affinity, weak cooperativity, and varying pH sensitivity. Hb's potential role in antimicrobial pathways could augment the immune defense of bivalves, which may be a complement to their lack of adaptive immunity. The role of Hb as a respiratory protein in bivalves likely originated from the substitution of hemocyanin. Molluscan Hbs demonstrate adaptive evolution in response to environmental changes via various strategies (e.g. increasing Hb types, multimerization, and amino acid residue substitutions at key sites), enhancing or altering functional properties for habitat adaptation. Concurrently, an increase in Hb assembly diversity, coupled with a downward trend in oxygen affinity, is observed during molluscan differentiation and evolution. Hb in Protobranchia, Heteroconchia, and Pteriomorphia bivalves originated from separate ancestors, with Protobranchia inheriting a relative ancient molluscan Hb gene. In bivalves, extracellular Hbs share a common origin, while gill Hbs likely emerged from convergent evolution. In summary, research on molluscan Hbs offers valuable insights into the origins, biological variations, and adaptive evolution of animal Hbs.

Macaque antibodies targeting Marburg virus glycoprotein induced by multivalent immunization.

Marburg virus infection in humans is associated with case fatality rates that can reach up to 90%, but to date, there are no approved vaccines or monoclonal antibody (mAb) countermeasures. Here, we immunized Rhesus macaques with multivalent combinations of filovirus glycoprotein (GP) antigens belonging to Marburg, Sudan, and Ebola viruses to generate monospecific and cross-reactive antibody responses against them. From the animal that developed the highest titers of Marburg virus GP-specific neutralizing antibodies, we sorted single memory B cells using a heterologous Ravn virus GP probe and cloned and characterized a panel of 34 mAbs belonging to 28 unique lineages. Antibody specificities were assessed by overlapping pepscan and binding competition analyses, revealing that roughly a third of the lineages mapped to the conserved receptor binding region, including potent neutralizing lineages that were confirmed by negative stain electron microscopy to target this region. Additional lineages targeted a protective region on GP2, while others were found to possess cross-filovirus reactivity. Our study advances the understanding of orthomarburgvirus glycoprotein antigenicity and furthers efforts to develop candidate antibody countermeasures against these lethal viruses. IMPORTANCE: Marburg viruses were the first filoviruses characterized to emerge in humans in 1967 and cause severe hemorrhagic fever with average case fatality rates of ~50%. Although mAb countermeasures have been approved for clinical use against the related Ebola viruses, there are currently no approved countermeasures against Marburg viruses. We successfully isolated a panel of orthomarburgvirus GP-specific mAbs from a macaque immunized with a multivalent combination of filovirus antigens. Our analyses revealed that roughly half of the antibodies in the panel mapped to regions on the glycoprotein shown to protect from infection, including the host cell receptor binding domain and a protective region on the membrane-anchoring subunit. Other antibodies in the panel exhibited broad filovirus GP recognition. Our study describes the discovery of a diverse panel of cross-reactive macaque antibodies targeting orthomarburgvirus and other filovirus GPs and provides candidate immunotherapeutics for further study and development.

Recombinant parainfluenza virus 5 expressing clade 2.3.4.4b H5 hemagglutinin protein confers broad protection against H5Ny influenza viruses.

The global circulation of clade 2.3.4.4b H5Ny highly pathogenic avian influenza viruses (HPAIVs) in poultry and wild birds, increasing mammal infections, continues to pose a public health threat and may even form a pandemic. An efficacious vaccine against H5Ny HPAIVs is crucial for emergency use and pandemic preparedness. In this study, we developed a parainfluenza virus 5 (PIV5)-based vaccine candidate expressing hemagglutinin (HA) protein of clade 2.3.4.4b H5 HPAIV, termed rPIV5-H5, and evaluated its safety and efficacy in mice and ferrets. Our results demonstrated that intranasal immunization with a single dose of rPIV5-H5 could stimulate H5-specific antibody responses, moreover, a prime-boost regimen using rPIV5-H5 stimulated robust humoral, cellular, and mucosal immune responses in mice. Challenge study showed that rPIV5-H5 prime-boost regimen provided sterile immunity against lethal clade 2.3.4.4b H5N1 virus infection in mice and ferrets. Notably, rPIV5-H5 prime-boost regimen provided protection in mice against challenge with lethal doses of heterologous clades 2.2, 2.3.2, and 2.3.4 H5N1, and clade 2.3.4.4h H5N6 viruses. These results revealed that rPIV5-H5 can elicit protective immunity against a diverse clade of highly pathogenic H5Ny virus infection in mammals, highlighting the potential of rPIV5-H5 as a pan-H5 influenza vaccine candidate for emergency use.IMPORTANCEClade 2.3.4.4b H5Ny highly pathogenic avian influenza viruses (HPAIVs) have been widely circulating in wild birds and domestic poultry all over the world, leading to infections in mammals, including humans. Here, we developed a recombinant PIV5-vectored vaccine candidate expressing the HA protein of clade 2.3.4.4b H5 virus. Intranasal immunization with rPIV5-H5 in mice induced airway mucosal IgA responses, high levels of antibodies, and robust T-cell responses. Importantly, rPIV5-H5 conferred complete protection in mice and ferrets against clade 2.3.4.4b H5N1 virus challenge, the protective immunity was extended against heterologous H5Ny viruses. Taken together, our data demonstrate that rPIV5-H5 is a promising vaccine candidate against diverse H5Ny influenza viruses in mammals.

Comprehensive pan-cancer analysis of the C2ORF40 expression: Infiltration associations and prognostic implications.

In recent years, C2ORF40 has been identified as a tumor suppressor gene with multiple functions, including roles in cell proliferation, migration, and senescence. To explore the role of the C2ORF40 gene in different tumors, we used multiple databases for analysis. Compared to adjacent normal tissues, C2ORF40 is downregulated in a variety of malignant tumors, including tumors such as breast cancer, colorectal cancer, bladder cancer, hepatocellular carcinoma and prostate cancer. Notably, low expression of the gene is significantly associated with poor overall survival and relapse-free survival rates. In specific cancers including colon cancer and prostate cancer, the expression of C2ORF40 is correlated with the infiltration of CAFs. C2ORF40 is also involved in biological processes such as cell apoptosis and regulation of protein stability. In conclusion, C2ORF40 can hold promise as a prognostic marker for pan-cancer analysis.

Systematic pan-cancer analysis insights into ICAM1 as an immunological and prognostic biomarker.

Intercellular adhesion molecule 1 (ICAM1) is a cell surface adhesion glycoprotein in the immunoglobulin supergene family. It is associated with several epithelial tumorigenesis processes, as well as with inflammation. However, the function of ICAM1 in the prognosis of tumor immunity is still unclear. This study aimed to examine the immune function of ICAM1 in 33 tumor types and to investigate the prognostic value of tumors. Using datasets from the Cancer Genome Atlas (TCGA), Genotype Tissue Expression (GTEx), Cancer Cell Lines Encyclopedia (CCLE), Human Protein Atlas (HPA), and cBioPortal, we investigated the role of ICAM1 in tumors. We explored the potential correlation between ICAM1 expression and tumor prognosis, gene mutations, microsatellite instability, and tumor immune cell levels in various cancers. We observed that ICAM1 is highly expressed in multiple malignant tumors. Furthermore, ICAM1 is negatively or positively associated with different malignant tumor prognoses. The expression levels of ICAM1 were correlated with the tumor mutation burden (TMB) in 11 tumors and with MSI in eight tumors. ICAM1 is a gene associated with immune infiltrating cells, such as M1 macrophages and CD8+ T cells in gastric and colon cancer. Meanwhile, the expression of ICAM1 is associated with several immune-related functions and immune-regulation-related signaling pathways, such as the chemokine signaling pathway. Our study shows that ICAM1 can be used as a prognostic biomarker in many cancer types because of its function in tumorigenesis and malignant tumor immunity.

Therapeutic Avenues to Modulate B-Cell Function in Patients With Cardiovascular Disease.

The adaptive immune system plays an important role in the development and progression of atherosclerotic cardiovascular disease. B cells can have both proatherogenic and atheroprotective roles, making treatments aimed at modulating B cells important therapeutic targets. The innate-like B-cell response is generally considered atheroprotective, while the adaptive response is associated with mixed consequences for atherosclerosis. Additionally, interactions of B cells with components of the adaptive and innate immune system, including T cells and complement, also represent key points for therapeutic regulation. In this review, we discuss therapeutic approaches based on B-cell depletion, modulation of B-cell survival, manipulation of both the antibody-dependent and antibody-independent B-cell response, and emerging immunization techniques.