Biology subjects

Weiner, J. A.

Publications and source records attributed to Weiner, J. A..

13 recordsLinked to original sources

bioRxiv2026

RNA-Encoded PGT121-LS Anti-HIV Antibody: Comprehensive Preclinical Characterization and Translational Pharmacokinetics

Human Immunodeficiency Virus (HIV)-1 broadly neutralizing antibodies (bNAbs) have demonstrated clinical efficacy, but face manufacturing challenges associated with recombinant protein production and purification. Here, we present a ribonucleic acid (RNA)-encoded bNAb (RibobNAb) platform that enables in vivo antibody production of the clinically validated bNAb PGT121 via lipid nanoparticle (LNP) delivery, supporting rapid evaluation of Fc variants (LS, del294, LS-del294) in vitro and in vivo. We confirmed expression, sub-nanomolar HIV-1 Env binding, and potent neutralization across all RibobNAb variants in vitro. In mice, single RNA-LNP administrations yielded in vivo expression of all RibobNAb variants, with PGT121-LS exhibiting a prolonged half-life compared with PGT121. In non-human primates (NHPs), a single intravenous administration of PGT121-LS RNA-LNP was well tolerated without anti-drug antibody (ADA) formation over 180 days and resulted in PGT121-LS half-lives comparable to the reference protein. Single intramuscular administration showed RibobNAb expression but resulted in ADA development from Day 14 onwards and lower bioavailability. In vivo-expressed PGT121-LS RibobNAb retained identical antiviral functionality to PGT121-LS reference protein. An NHP pharmacokinetics model integrating RNA transfection and translation dynamics enabled allometric scaling and first-in-human dose prediction. We highlight RibobNAbs as an alternative to conventional purified protein antibodies for rapid development of bNAb-based therapeutic strategies. O_FIG O_LINKSMALLFIG WIDTH=193 HEIGHT=200 SRC="FIGDIR/small/734219v1_ufig1.gif" ALT="Figure 1"> View larger version (44K): org.highwire.dtl.DTLVardef@141319forg.highwire.dtl.DTLVardef@120caadorg.highwire.dtl.DTLVardef@1da2782org.highwire.dtl.DTLVardef@158064d_HPS_FORMAT_FIGEXP M_FIG Graphical abstract C_FIG

immunology↗

bioRxiv2026

Late induction of IgG4 following SARS-CoV-2 mRNA vaccination in pregnant and non-pregnant individuals includes clonotypes raised early in the response

To better understand how pregnancy impacts humoral immunity, we conducted an in-depth longitudinal analysis of the kinetics and characteristics of vaccine responses in a prospective cohort of pregnant and non-pregnant women. Humoral immune responses observed among pregnant participants who received the mRNA-delivered SARS-CoV-2 vaccination, including their effector functions, were in some cases marginally lower than those among non-pregnant controls, while prior infection was associated with some potentiation in humoral responses. Importantly, vaccine-induced antibodies were efficiently transferred across the placenta, providing the fetus with passive immunity and underscoring the dual benefit of maternal vaccination for both mother and neonate against COVID-19. Delayed induction of spike-specific IgG4 following the primary two-dose vaccination series was observed in vaccine recipients, independent of pregnancy status. In a subset (n=6) of pregnant women whose spike-specific serum IgG repertoires were extensively profiled at the clonotypic level over time as part of another study, we proteomically identified secreted IgG clonotypes that had class-switched to IgG4. Matching of these clonotypes detected as IgG4 to those defined as SARS-CoV-2 spike-specific revealed that, while a minority of total clonotypes, they were elicited early in the immunization series and tended to be more highly mutated, more prevalent, and more persistent than clonotypes in the serological repertoire that were not detected as IgG4. Consistent with the increase in secreted vaccine-specific IgG4 over time, but its poorer placental transfer, these clonotypes were detected at greater levels in maternal but not cord blood at the time of delivery as compared to 28 days post the second vaccine dose. These findings indicate some impact in the kinetics, characteristics, and functions of the humoral response that may be associated with pregnancy-related immune modulation. Conservation of the late class-switch recombination to IgG4 that has previously been associated with mRNA-based SARS-CoV-2 vaccines raises questions about how different immunological states and vaccine components influence short- and long-term characteristics of the humoral immune response.

immunology↗

bioRxiv2026

Intranasal Replicating Adenovirus type 4-SARS-CoV-2 Recombinants Induce Superior Immune Response Durability and Efficacy in Preclinical Testing Compared to Standard Intramuscular Vaccines

The portfolio of next generation COVID-19 vaccines would benefit from candidates that induce durable systemic and mucosal immune responses that would lessen person-to-person transmission. We constructed an intranasal (IN) replication-competent adenovirus type 4 recombinant platform to express SARS-CoV-2 Spike variants (Ad4-S) and assessed immunogenicity and efficacy in the Syrian hamster model. Although both IN Ad4-S and intramuscular (IM) vaccines (Ad26.CoV2.S and mRNA-1273) induced serum binding antibodies, only Ad4-S induced a robust mucosal response in the nasal cavity. IN Ad4-S vaccination induced serum neutralizing titers equivalent to or greater than IM vaccination but more durable up to 6 months. Upon challenge, IN immunization also resulted in less weight loss, greater breadth and durability of restriction of viral replication, and less lung pathology than IM immunization up to 268 days after immunization. These data support the potential of the IN Ad4 vaccine platform to reduce transmission of SARS-CoV-2 and other respiratory viruses with pandemic potential.

immunology↗

bioRxiv2025

Effects and mechanisms of monoclonal and polyclonal human antibodies in protection of humanized mice from HIV-1 challenge

Recent clinical trials of both active and passive immunization demonstrate the barriers to the successful development of efficacious preventative HIV-1 vaccines and prophylactic antibody treatments. More facile means to explore these interventions in preclinical models could fill key gaps in knowledge and contribute to identifying and optimizing the most promising interventions. Here we report simple adaptations to standard virus challenge strategies in the humanized mouse model that support evaluation of antibody-mediated protection from infection with fewer high stakes design choices and demonstrate evaluation of protection afforded by polyclonal human serum IgG antibodies (pAbs) and monoclonal antibodies (mAbs) with variable pharmacokinetic (PK) and functional profiles. Using these adaptations, we observed that both neutralization and Fc-mediated effector functions contribute to the in vivo antiviral activity of broadly-neutralizing antibody VRC01, that confounding of results due to differences in mAb PK can be overcome, and most promisingly, that polyclonal human serum IgG that exhibits potent neutralizing and Fc-effector function can protect from infection. Collectively, this work demonstrates insights into antibody-mediated protection and methods that hold promise in supporting testing the protection from HIV-1 afforded by human pAb responses induced by vaccination.

immunology↗

bioRxiv2025

A new mouse mutant with a discrete mutation in Pcdhgc5 reveals that the Protocadherin γC5 isoform is not essential for dendrite arborization in the cerebral cortex

There are [~]60 clustered protocadherin (cPcdh) isoforms expressed from three gene clusters (Pcdha, Pcdhb, Pcdhg) arrayed in tandem across nearly 1 Mb in mammals. cPcdhs are homophilic cell adhesion molecules (CAMs) critical for a host of neural developmental functions consistent with a role in cell-cell recognition. Indeed, isoforms make recognition modules in combination to generate recognition diversity far exceeding the [~]60 individual CAMs. However, there is also growing evidence for specialized functions for specific isoforms, particularly the C-type isoforms found at the 3 ends of the Pcdha cluster (C1 and C2) and at the 3 end of the Pcdhg cluster ({gamma}C3, {gamma}C4, and {gamma}C5). We have previously described unique roles for {gamma}C3 in dendrite arborization in the cerebral cortex and neural circuit formation in the spinal cord, as well as for {gamma}C4 in neuronal survival. Here we report a new mouse mutant specifically targeting the Pcdhgc5 exon encoding {gamma}C5. Unlike the rest of the Pcdhg cluster, expression of this isoform does not begin until postnatal stages of mouse development, increasing in the second week of life, suggesting specialized roles. We found significant expression changes in gene pathways involved in synaptic activity, learning and memory, and cognition. Despite this, we saw no major disruption in the cerebral cortex in neuronal organization, survival, dendritic arborization, or synaptic protein expression in these mutants. This new model will be an important tool for future studies delineating specific functions for {gamma}C5.

neuroscience↗

bioRxiv2024

PCDHGC3 silencing promotes renal carcinoma metastasis via mTOR/HIF2a and lipid metabolism rewiring

Clustered protocadherins (cPCDH) are widely expressed in the nervous system with known functions, but their roles in cancer, particularly metastasis, are largely unexplored. Our previous research revealed that epigenetic silencing of PCDHGC3 is linked to decreased survival in neuroendocrine cancer patients. This study investigates PCDHGC3s role in clear cell renal cell carcinoma (ccRCC). We found that decreased PCDHGC3 expression is associated with lower survival and advanced disease stage in ccRCC patients. shRNA-mediated PCDHGC3 silencing in renal cancer cell lines significantly increased cell proliferation, invasion, and survival. In orthotopic mouse models, PCDHGC3 silencing promoted metastasis. The mTOR and HIF2 pathways were identified as downstream targets activated by PCDHGC3 loss. Inhibition of these pathways counteracted the effects of PCDHGC3 silencing, highlighting their importance in tumor progression. Proteomic and metabolomic analyses showed that PCDHGC3 silencing led to overexpression of proteins involved in fatty acid and cholesterol synthesis, increasing lipid droplets and shifting lipid metabolism. This metabolic reprogramming characterizes aggressive ccRCC. Our study emphasizes PCDHGC3s impact on ccRCC metastasis and suggests mTOR or HIF2 inhibitors as potential therapies for PCDHGC3-deficient patients.

cancer biology↗

bioRxiv2024

A C-terminal motif containing a PKC phosphorylation site regulates γ-Protocadherin-mediated dendrite arborization in the cerebral cortex in vivo

The Pcdhg gene cluster encodes 22 {gamma}-Protocadherin ({gamma}-Pcdh) cell adhesion molecules that critically regulate multiple aspects of neural development, including neuronal survival, dendritic and axonal arborization, and synapse formation and maturation. Each {gamma}-Pcdh isoform has unique protein domains-a homophilically-interacting extracellular domain and a juxtamembrane cytoplasmic domain- as well as a C-terminal cytoplasmic domain shared by all isoforms. The extent to which isoform-specific vs. shared domains regulate distinct {gamma}-Pcdh functions remains incompletely understood. Our previous in vitro studies identified PKC phosphorylation of a serine residue within a shared C-terminal motif as a mechanism through which {gamma}-Pcdh promotion of dendrite arborization via MARCKS is abrogated. Here, we used CRISPR/Cas9 genome editing to generate two new mouse lines expressing only non-phosphorylatable {gamma}-Pcdhs, due either to a serine-to-alanine mutation (PcdhgS/A) or to a 15-amino acid C-terminal deletion resulting from insertion of an early stop codon (PcdhgCTD). Both lines are viable and fertile, and the density and maturation of dendritic spines remains unchanged in both PcdhgS/A and PcdhgCTDcortex. Dendrite arborization of cortical pyramidal neurons, however, is significantly increased in both lines, as are levels of active MARCKS. Intriguingly, despite having significantly reduced levels of {gamma}-Pcdh proteins, the PcdhgCTD mutation yields the strongest phenotype, with even heterozygous mutants exhibiting increased arborization. The present study confirms that phosphorylation of a shared C-terminal motif is a key {gamma}-Pcdh negative regulation point, and contributes to a converging understanding of {gamma}-Pcdh family function in which distinct roles are played by both individual isoforms and discrete protein domains.

neuroscience↗

bioRxiv2023

Characteristics and functions of infection-enhancing antibodies to the N-terminal domain of SARS-CoV-2

Characterization of functional antibody responses to the N-terminal domain (NTD) of the SARS-CoV-2 spike (S) protein has included identification of both potent neutralizing activity and putative enhancement of infection. Fc{gamma}-receptor (Fc{gamma}R)-independent enhancement of SARS-CoV-2 infection mediated by NTD-binding monoclonal antibodies (mAbs) has been observed in vitro, but the functional significance of these antibodies in vivo is not clear. Here we studied 1,213 S-binding mAbs derived from longitudinal sampling of B-cells collected from eight COVID-19 convalescent patients and identified 72 (5.9%) mAbs that enhanced infection in a VSV-SARS-CoV-2-S-Wuhan pseudovirus (PV) assay. The majority (68%) of these mAbs recognized the NTD, were identified in patients with mild and severe disease, and persisted for at least five months post-infection. Enhancement of PV infection by NTD-binding mAbs was not observed using intestinal (Caco-2) and respiratory (Calu-3) epithelial cells as infection targets and was diminished or lost against SARS-CoV-2 variants of concern (VOC). Proteomic deconvolution of the serum antibody repertoire from two of the convalescent subjects identified, for the first time, NTD-binding, infection-enhancing mAbs among the circulating immunoglobulins directly isolated from serum (i.e., functionally secreted antibody). Functional analysis of these mAbs demonstrated robust activation of Fc{gamma}RIIIa associated with antibody binding to recombinant S proteins. Taken together, these findings suggest functionally active NTD-specific mAbs arise frequently during natural infection and can last as major serum clonotypes during convalescence. These antibodies display diverse attributes that include Fc{gamma}R activation, and may be selected against by mutations in NTD associated with SARS-CoV-2 VOC.

immunology↗

bioRxiv2023

The Clustered Gamma Protocadherin Pcdhγc4 Isoform Regulates Cortical Interneuron Programmed Cell Death in the Mouse Cortex.

Cortical function critically depends on inhibitory/excitatory balance. Cortical inhibitory interneurons (cINs) are born in the ventral forebrain and migrate into cortex, where their numbers are adjusted by programmed cell death. Previously, we showed that loss of clustered gamma protocadherins (Pcdh{gamma}), but not of genes in the alpha or beta clusters, increased dramatically cIN BAX-dependent cell death in mice. Here we show that the sole deletion of the Pcdh{gamma}c4 isoform, but not of the other 21 isoforms in the Pcdh{gamma} gene cluster, increased cIN cell death in mice during the normal period of programmed cell death. Viral expression of the Pcdh{gamma}c4 isoform rescued transplanted cINs lacking Pcdh{gamma} from cell death. We conclude that Pcdh{gamma}, specifically Pcdh{gamma}c4, plays a critical role in regulating the survival of cINs during their normal period of cell death. This demonstrates a novel specificity in the role of Pcdh{gamma} isoforms in cortical development.

neuroscience↗

bioRxiv2022

γ-Protocadherins control synapse formation and peripheral branching of touch sensory neurons

Light touch sensation begins with activation of low-threshold mechanoreceptor (LTMR) endings in the skin and propagation of their signals to the spinal cord and brainstem. We found that the clustered protocadherin gamma (Pcdhg) gene locus, which encodes 22 cell-surface homophilic binding proteins, is required in somatosensory neurons for normal behavioral reactivity to a range of tactile stimuli. Developmentally, distinct Pcdhg isoforms mediate LTMR synapse formation through neuron-neuron interactions and peripheral axonal branching through neuron-glia interactions. The Pcdhgc3 isoform mediates homophilic interactions between sensory axons and spinal cord neurons to promote synapse formation in vivo, and is sufficient to induce postsynaptic specializations in vitro. Moreover, loss of Pcdhgs and somatosensory synaptic inputs to the dorsal horn lead to fewer corticospinal synapses onto dorsal horn neurons. These findings reveal essential roles for Pcdhg isoform diversity in somatosensory neuron synapse formation, peripheral axonal branching, and step-wise assembly of central mechanosensory circuitry.

neuroscience↗

bioRxiv2021

p53-mediated neurodegeneration in the absence of the nuclear protein Akirin2

Proper gene regulation is critical for both neuronal development and maintenance as the brain matures. We previously demonstrated that Akirin2, an essential nuclear protein that interacts with transcription factors and chromatin remodeling complexes, is required for the embryonic formation of the cerebral cortex. Here we show that Akirin2 plays a mechanistically distinct role in maintaining healthy neurons during cortical maturation. Restricting Akirin2 loss to excitatory cortical neurons resulted in progressive neurodegeneration via necroptosis and severe cortical atrophy with age. Comparing transcriptomes from Akirin2-null postnatal neurons and cortical progenitors revealed that targets of the tumor suppressor p53, a regulator of both proliferation and cell death encoded by Trp53, were consistently upregulated. Heterozygous deletion of Trp53 rescued neurodegeneration in Akirin2-null neurons. These data: 1) implicate Akirin2 as a critical neuronal maintenance protein; 2) identify p53 pathways as mediators of Akirin2 functions; and 3) suggest Akirin2 dysfunction may be relevant to neurodegenerative diseases.

neuroscience↗

bioRxiv2020

Serotonin signaling by maternal neurons upon stress ensures progeny survival

Germ cells are vulnerable to stress. Therefore, how organisms protect their future progeny from damage in a fluctuating environment is a fundamental question in biology. We show that in Caenorhabditis elegans, serotonin released by maternal neurons during stress ensures the viability and stress tolerance of future offspring by enabling the transcription factor HSF1 to alter chromatin in soon-to-be fertilized germ cells by recruiting the histone chaperone FACT, displacing histones, and initiating protective gene expression. Without maternal serotonin signaling by neurons, FACT is not recruited by HSF1 in germ cells, transcription occurs but is delayed, and progeny of stressed C. elegans mothers fail to complete development. Serotonin acts through a signal transduction pathway conserved between C. elegans and mammalian cells to facilitate HSF1 to recruit FACT. These studies uncover a novel mechanism by which stress sensing by neurons is coupled to transcription response times of germ cells to protect future offspring.

cell biology↗

bioRxiv2019

CRISPR/Cas9 interrogation of the mouse Pcdhg gene cluster reveals a crucial isoform-specific role for Pcdhgc4

The mammalian Pcdhg gene cluster encodes a family of 22 cell adhesion molecules, the gamma-Protocadherins ({gamma}-Pcdhs), critical for neuronal survival and neural circuit formation. The extent to which isoform diversity-a{gamma}-Pcdh hallmark-is required for their functions remains unclear. We used a CRISPR/Cas9 approach to reduce isoform diversity, targeting each Pcdhg variable exon with pooled sgRNAs to generate an allelic series of 26 mouse lines with 1 to 21 isoforms disrupted via discrete indels at guide sites and/or larger deletions/rearrangements. Analysis of 5 mutant lines indicates that postnatal viability and neuronal survival do not require isoform diversity. Surprisingly, as it is the only {gamma}-Pcdh that cannot independently engage in homophilic trans-interactions, we find that {gamma}C4, encoded by Pcdhgc4, is the only critical isoform. Because the human orthologue is the only PCDHG gene constrained in humans, our results indicate a conserved {gamma}C4 function that likely involves distinct molecular mechanisms.

developmental biology↗