Biology subjects

Rappaport, J.

Publications and source records attributed to Rappaport, J..

16 recordsLinked to original sources

bioRxiv2026

HIV remodels germinal centers through T follicular helper cell loss

T follicular helper cells (Tfh) within germinal centers are among the most permissive cells for HIV infection and serve as a critical part of the viral reservoir. Given the key role of Tfh in affinity maturation and the production of plasma cells and memory B cells, an understanding of this critical viral reservoir is fundamental not only for cure strategies but also for understanding immunodeficiency in people living with HIV (PLWH). Using rare clinical specimens from PLWH, we demonstrate that HIV causes both a change in the cellular composition of germinal centers and changes in their overall architecture in ways that are consistent with the immunodeficiency seen in PLWH, independent of their CD4 count. We note a significant decrease in the number of GC Tfh cells and an increase in the number of follicular CD8 T cells in lymphoid tissues obtained from PLWH. Our data further show that GC size is decreased both by HIV and with increased age. Single-cell RNA sequencing data demonstrate that HIV acutely decreases the expression of Tfh B-cell support pathways and disrupts Tfh expression of cell trafficking receptors. Additionally, in a lymphoid organoid model of acute infection that forms GC-like structures, these findings are recapitulated, suggesting that New Approach Methodologies could be germane to understanding the impact of HIV on the reservoir and immunodeficiency. This study describes previously undocumented remodeling of the germinal center niche in PLWH, providing insight into the immune microenvironment of a key HIV reservoir that may help inform future cure strategies.

microbiology↗

bioRxiv2025

Persistent Activation of Monocytes/Macrophages and Cell Senescence in SIV-Infected Macaques on ART

Despite effective viral suppression with antiretroviral therapy (ART), people living with HIV (PLWH) experience persistent inflammation, immune dysfunction, and premature onset of cardiovascular and aging-related comorbidities. To define the underlying mechanisms, we performed longitudinal transcriptomic profiling in peripheral blood mononuclear cells (PBMCs) from a cohort of simian immunodeficiency virus (SIV)-infected rhesus macaques spanning four key stages: pre-infection, acute infection, short-term ART, and long-term ART. Bulk RNA sequencing revealed dynamic immune remodeling across infection and treatment. Acute SIV infection induced robust antiviral and inflammatory programs, with upregulation of interferon-stimulated genes (ISGs), IL-27, JAK/STAT, and NF-{kappa}B signaling, coupled with suppression of T- and B-cell activation pathways. Short-term ART effectively reversed these transcriptional perturbations, restoring adaptive immune gene expression and reducing innate antiviral responses to near-baseline levels. In contrast, chronic SIV infection on long-term ART maintained viral suppression but was characterized by reactivation of innate immune pathways, including TLR2/TLR4/MYD88, NF-{kappa}B, and inflammasome (NLRP3/or NLRP12, caspase-1) signaling, along with sustained macrophage activation, platelet/coagulation signaling, and senescence-associated secretory phenotype. Protein analyses confirmed persistent CASPASE-1 and NF-{kappa}B activation in spleen tissue. Pathologic evaluation of a carotid artery from an SIV-infected, long-term ART- treated macaque revealed macrophage-rich plaques with p21 senescent cells with intraluminal thrombus formation, recapitulating key features of HIV-associated atherogenesis. Together, these findings demonstrate that while ART normalizes acute infection-induced immune dysregulation, chronic SIV infection sustains a chronic, macrophage- and TLR-driven inflammatory state linked to vascular injury and aging process regardless of long-term suppression of viremia. Targeting inflammasome, NF-{kappa}B, and senescence pathways may mitigate non-AIDS comorbidities in PLWH.

immunology↗

bioRxiv2024

BRAIN-METASTATIC MELANOMA MODELS UNCOVER IMMUNE AND MOLECULAR CORRELATES OF RESPONSE TO IMMUNOTHERAPY

Despite the promising results of immune checkpoint blockade (ICB) therapy, outcomes for patients with brain metastasis (BrM) remain poor. Identifying resistance mechanisms has been hindered by limited access to patient samples and relevant preclinical models. Here, we developed two mouse melanoma BrM models that recapitulate the disparate responses to ICB seen in patients. We demonstrate that these models capture the cellular and molecular complexity of human disease and reveal key factors shaping the tumor microenvironment and influencing ICB response. BR1-responsive tumor cells express inflammatory programs that polarize microglia into reactive states, eliciting robust T cell recruitment. In contrast, BR3-resistant melanoma cells are enriched in neurological programs and exploit tolerance mechanisms to maintain microglia homeostasis and limit T cell infiltration. In humans, BR1 and BR3 expression signatures correlate positively or negatively with T cell infiltration and BrM patient outcomes, respectively. Our study provides clinically relevant models and uncovers mechanistic insights into BrM ICB responses, offering potential biomarkers and therapeutic targets to improve therapy efficacy.

immunology↗

bioRxiv2024

Development of an optimized machine learning approach to enhance brain metastatic burden assessment in preclinical models.

Brain metastases (BrM) occur when malignant cells spread from a primary tumor located in other parts of the body to the brain. BrM is a deadly complication for cancer patients and currently lacks effective therapies. Due to the limited access to patient samples, preclinical models remain a valuable tool for studying metastasis development, progression, and response to therapy. Thus, reliable methods for quantifying metastatic burden in these models are crucial. Here, we describe step by step a new semi-automatic machine-learning approach to quantify metastatic burden on mouse whole-brain stereomicroscope images while preserving tissue integrity. This protocol utilizes the open-source, user-friendly image analysis software QuPath. The method is fast, reproducible, unbiased, and provides access to data points not always obtainable with other existing strategies.

cancer biology↗

bioRxiv2024

High IL-1β and IL-18 Levels Associate with Gut Barrier Disruption and Monocyte Activation During Chronic SIV Infection with Long-Term Suppressive Antiretroviral Therapy

HIV-induced persistent immune activation is a key mediator of inflammatory comorbidities such as cardiovascular disease (CVD) and neurocognitive disorders. While a preponderance of data indicate that gut barrier disruption and microbial translocation are drivers of chronic immune activation, the molecular mechanisms of this persistent inflammatory state remain poorly understood. Here, utilizing the nonhuman primate model of HIV infection with suppressive antiretroviral therapy (ART), we investigated activation of inflammasome pathways and their association with intestinal epithelial barrier disruption and CVD pathogenesis. Longitudinal blood samples obtained from rhesus macaques with chronic SIV infection and long-term suppressive ART were evaluated for biomarkers of intestinal epithelial barrier disruption (IEBD), inflammasome activation (IL-1{beta} and IL-18), inflammatory cytokines, and triglyceride (TG) levels. Activated monocyte subpopulations and glycolytic potential were investigated in peripheral blood mononuclear cells (PBMCs). Higher plasma levels of IL-1{beta} and IL-18 were observed following the hallmark increase in IEBD biomarkers, intestinal fatty acid-binding protein (IFABP) and LPS-binding protein (LBP), during the chronic phase of treated SIV infection. Further, significant correlations of plasma IFABP levels with IL-1{beta} and IL-18 were observed between 10-12 months of ART. Higher levels of sCD14, IL-6, and GM-CSF, among other inflammatory mediators, were also observed only during the long-term SIV+ART phase along with a trend of increase in frequencies of activated CD14+CD16+ intermediate monocyte subpopulations. Lastly, we found elevated levels of blood TG and higher glycolytic capacity in PBMCs of chronic SIV-infected macaques with long-term ART. The increase in circulating IL-18 and IL-1{beta} following IEBD and their significant positive correlation with IFABP suggest a connection between gut barrier disruption and inflammasome activation during chronic SIV infection, despite viral suppression with ART. Additionally, the increase in markers of monocyte activation, along with elevated TG and enhanced glycolytic pathway activity, indicates metabolic remodeling that could accelerate CVD pathogenesis. Further research is needed to understand mechanisms by which gut dysfunction and inflammasome activation contribute to HIV-associated CVD and metabolic complications, enabling targeted interventions in people with HIV.

immunology↗

bioRxiv2023

The Impact of SIV-Induced Immunodeficiency on Clinical Manifestation, Immune Response, and Viral Dynamics in SARS-CoV-2 Coinfection

Persistent and uncontrolled SARS-CoV-2 replication in immunocompromised individuals has been observed and may be a contributing source of novel viral variants that continue to drive the pandemic. Importantly, the effects of immunodeficiency associated with chronic HIV infection on COVID-19 disease and viral persistence have not been directly addressed in a controlled setting. Here we conducted a pilot study wherein two pigtail macaques (PTM) chronically infected with SIVmac239 were exposed to SARS-CoV-2 and monitored for six weeks for clinical disease, viral replication, and viral evolution, and compared to our previously published cohort of SIV-naive PTM infected with SARS-CoV-2. At the time of SARS-CoV-2 infection, one PTM had minimal to no detectable CD4+ T cells in gut, blood, or bronchoalveolar lavage (BAL), while the other PTM harbored a small population of CD4+ T cells in all compartments. Clinical signs were not observed in either PTM; however, the more immunocompromised PTM exhibited a progressive increase in pulmonary infiltrating monocytes throughout SARS-CoV-2 infection. Single-cell RNA sequencing (scRNAseq) of the infiltrating monocytes revealed a less activated/inert phenotype. Neither SIV-infected PTM mounted detectable anti-SARS-CoV-2 T cell responses in blood or BAL, nor anti-SARS-CoV-2 neutralizing antibodies. Interestingly, despite the diminished cellular and humoral immune responses, SARS-CoV-2 viral kinetics and evolution were indistinguishable from SIV-naive PTM in all sampled mucosal sites (nasal, oral, and rectal), with clearance of virus by 3-4 weeks post infection. SIV-induced immunodeficiency significantly impacted immune responses to SARS-CoV-2 but did not alter disease progression, viral kinetics or evolution in the PTM model. SIV-induced immunodeficiency alone may not be sufficient to drive the emergence of novel viral variants.

immunology↗

bioRxiv2023

Immune Correlates of Hyperglycemia and Vaccination in a Non-human Primate Model of Long-COVID

Hyperglycemia, and exacerbation of pre-existing deficits in glucose metabolism, are major manifestations of the post-acute sequelae of SARS-CoV-2 (PASC). Our understanding of lasting glucometabolic disruptions after acute COVID-19 remains unclear due to the lack of animal models for metabolic PASC. Here, we report a non-human primate model of metabolic PASC using SARS-CoV-2 infected African green monkeys (AGMs). Using this model, we have identified a dysregulated chemokine signature and hypersensitive T cell population during acute COVID-19 that correlates with elevated and persistent hyperglycemia four months post-infection. This persistent hyperglycemia correlates with elevated hepatic glycogen, but there was no evidence of long-term SARS-CoV-2 replication in the liver and pancreas. Finally, we report a favorable glycemic effect of the SARS-CoV-2 mRNA vaccine, administered on day 4 post-infection. Together, these data suggest that the AGM metabolic PASC model exhibits important similarities to human metabolic PASC and can be utilized to assess therapeutic candidates to combat this syndrome.

immunology↗

bioRxiv2023

SARS-CoV-2 RNA Persists in the Central Nervous System of Non-Human Primates Despite Clinical Recovery

Adverse neurological and psychiatric outcomes, collectively termed the post-acute sequelae of SARS-CoV-2 infection (PASC), persist in adults clinically recovered from COVID-19. Effective therapeutic interventions are fundamental to reducing the burden of PASC, necessitating an investigation of the pathophysiology underlying the debilitating neurological symptoms associated with the condition. Herein, eight non-human primates (Wild-Caught African Green Monkeys, n=4; Indian Rhesus Macaques, n=4) were inoculated with the SARS-CoV-2 isolate USA-WA1/2020 by either small particle aerosol or via multiple routes. At necropsy, tissue from the olfactory epithelium and pyriform cortex/amygdala of SARS-CoV-2 infected non-human primates were collected for ribonucleic acid in situ hybridization (i.e., RNAscope). First, angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2) mRNA are downregulated in the pyriform cortex/amygdala of non-human primates clinically recovered from SARS-CoV-2 inoculation relative to wildtype controls. Second, abundant SARS-CoV-2 mRNA was detected in clinically recovered non-human primates; mRNA which is predominantly harbored in pericytes. Collectively, examination of post-mortem pyriform cortex/amygdala brain tissue of non-human primates clinically recovered from SARS-CoV-2 infection revealed two early pathophysiological mechanisms potentially underlying PASC. Indeed, therapeutic interventions targeting the downregulation of ACE2, decreased expression of TMPRSS2, and/or persistent infection of pericytes in the central nervous system may effectively mitigate the debilitating symptoms of PASC.

neuroscience↗

bioRxiv2022

Durable protection against SARS-CoV-2 Omicron induced by an adjuvanted subunit vaccine

Despite the remarkable efficacy of COVID-19 vaccines, waning immunity, and the emergence of SARS-CoV-2 variants such as Omicron represents a major global health challenge. Here we present data from a study in non-human primates demonstrating durable protection against the Omicron BA.1 variant induced by a subunit SARS-CoV-2 vaccine, consisting of RBD (receptor binding domain) on the I53-50 nanoparticle, adjuvanted with AS03, currently in Phase 3 clinical trial (NCT05007951). Vaccination induced robust neutralizing antibody (nAb) titers that were maintained at high levels for at least one year after two doses (Pseudovirus nAb GMT: 2207, Live-virus nAb GMT: 1964) against the ancestral strain, but not against Omicron. However, a booster dose at 6-12 months with RBD-Wu or RBD-{beta} (RBD from the Beta variant) displayed on I53-50 elicited equivalent and remarkably high neutralizing titers against the ancestral as well as the Omicron variant. Furthermore, there were substantial and persistent memory T and B cell responses reactive to Beta and Omicron variants. Importantly, vaccination resulted in protection against Omicron infection in the lung (no detectable virus in any animal) and profound suppression of viral burden in the nares (median peak viral load of 7567 as opposed to 1.3x107 copies in unvaccinated animals) at 6 weeks post final booster. Even at 6 months post vaccination, there was significant protection in the lung (with 7 out of 11 animals showing no viral load, 3 out of 11 animals showing ~20-fold lower viral load than unvaccinated controls) and rapid control of virus in the nares. These results highlight the durable cross-protective immunity elicited by the AS03-adjuvanted RBD-I53-50 nanoparticle vaccine platform.

immunology↗

bioRxiv2021

The pigtail macaque (Macaca nemestrina) model of COVID-19 reproduces diverse clinical outcomes and reveals new and complex signatures of disease

The novel coronavirus SARS-CoV-2, the causative agent of COVID-19 disease, has killed over four million people worldwide as of July 2021 with infections rising again due to the emergence of highly transmissible variants. Animal models that faithfully recapitulate human disease are critical for assessing SARS-CoV-2 viral and immune dynamics, for understanding mechanisms of disease, and for testing vaccines and therapeutics. Pigtail macaques (PTM, Macaca nemestrina) demonstrate a rapid and severe disease course when infected with simian immunodeficiency virus (SIV), including the development of severe cardiovascular symptoms that are pertinent to COVID-19 manifestations in humans. We thus proposed this species may likewise exhibit severe COVID-19 disease upon infection with SARS-CoV-2. Here, we extensively studied a cohort of SARS-CoV-2-infected PTM euthanized either 6- or 21-days after respiratory viral challenge. We show that PTM demonstrate largely mild-to-moderate COVID-19 disease. Pulmonary infiltrates were dominated by T cells, including CD4+ T cells that upregulate CD8 and express cytotoxic molecules, as well as virus-targeting T cells that were predominantly CD4+. We also noted increases in inflammatory and coagulation markers in blood, pulmonary pathologic lesions, and the development of neutralizing antibodies. Together, our data demonstrate that SARS-CoV-2 infection of PTM recapitulates important features of COVID-19 and reveals new immune and viral dynamics and thus may serve as a useful animal model for studying pathogenesis and testing vaccines and therapeutics.

immunology↗

bioRxiv2021

An Orally Available Cathepsin L Inhibitor Protects Lungs Against SARS-CoV-2-Induced Diffuse Alveolar Damage in African Green Monkeys

The COVID-19 pandemic resulted from global infection by the SARS-CoV-2 coronavirus and rapidly emerged as an urgent health issue requiring effective treatments. To initiate infection, the Spike protein of SARS-CoV-2 requires proteolytic processing mediated by host proteases. Among the host proteases proposed to carry out this activation is the cysteine protease cathepsin L. Inhibiting cathepsin L has been proposed as a therapeutic strategy for treating COVID-19. SLV213 (K777) is an orally administered small molecule protease inhibitor that exhibits in vitro activity against a range of viruses, including SARS-CoV-2. To confirm efficacy in vivo, K777 was evaluated in an African green monkey (AGM) model of COVID-19. A pilot experiment was designed to test K777 in a prophylactic setting, animals were pre-treated with 100mg/kg K777 (N=4) or vehicle (N=2) before inoculation with SARS-CoV-2. Initial data demonstrated that K777 treatment reduced pulmonary pathology compared to vehicle-treated animals. A second study was designed to test activity in a therapeutic setting, with K777 treatment (33 mg/kg or 100 mg/kg) initiated 8 hours after exposure to the virus. In both experiments, animals received K777 daily via oral gavage for 7 days. Vehicle-treated animals exhibited higher lung weights, pleuritis, and diffuse alveolar damage. In contrast, lung pathology was reduced in K777-treated monkeys, and histopathological analyses confirmed the lack of diffuse alveolar damage. Antiviral effects were further demonstrated by quantitative reductions in viral load of samples collected from upper and lower airways. These preclinical data support the potential for early SLV213 treatment in COVID-19 patients to prevent severe lung pathology and disease progression.

immunology↗

bioRxiv2021

Effective prophylaxis of COVID-19 in rhesus macaques using a combination of two parentally-administered SARS-CoV-2 neutralizing antibodies

SARS-CoV-2 is a respiratory borne pathogenic beta coronavirus that is the source of a worldwide pandemic and the cause of multiple pathologies in man. The rhesus macaque model of COVID-19 was utilized to test the added benefit of combinatory parenteral administration of two high-affinity anti-SARS-CoV-2 monoclonal antibodies (mAbs; C144-LS and C135-LS) expressly developed to neutralize the virus and modified to extend their pharmacokinetics. After completion of kinetics study of mAbs in the primate, combination treatment was administered prophylactically to mucosal viral challenge. Results showed near complete virus neutralization evidenced by no measurable titer in mucosal tissue swabs, muting of cytokine/chemokine response, and lack of any discernable pathologic sequalae. Blocking infection was a dose-related effect, cohorts receiving lower doses (6, 2 mg/kg) resulted in low grade viral infection in various mucosal sites compared to that of a fully protective dose (20 mg/kg). A subset of animals within this cohort whose infectious challenge was delayed 75 days later after mAb administration were still protected from disease. Results indicate this combination mAb effectively blocks development of COVID-19 in the rhesus disease model and accelerates the prospect of clinical studies with this effective antibody combination.

microbiology↗

bioRxiv2021

Safety, immunogenicity and protection provided by unadjuvanted and adjuvanted formulations of recombinant plant-derived virus-like particle vaccine candidate for COVID-19 in non-human primates

Although antivirals are important tools to control the SARS-CoV-2 infection, effective vaccines are essential to control the current pandemic. Plant-derived virus-like particle (VLP) vaccine candidates have previously demonstrated immunogenicity and efficacy against influenza. Here we report the immunogenicity and protection induced in macaques by intramuscular injections of VLP bearing SARS-CoV-2 spike protein (CoVLP) vaccine candidate formulated with or without Adjuvant System 03 (AS03) or cytosine phosphoguanine (CpG) 1018. Although a single dose of unadjuvanted CoVLP vaccine candidate stimulated humoral and cell-mediated immune responses, booster immunization (at 28 days after prime) and adjuvants significantly improved both responses with a higher immunogenicity and protection provided by AS03 adjuvanted CoVLP. Fifteen microgram CoVLP adjuvanted with AS03 induced a balanced IL-2 driven response along with IL-4 expression in CD4 T cells and mobilization of CD4 follicular helper cells (Tfh). Animals were challenged by multiple routes (i.e. intratracheal, intranasal and ocular) with a total viral dose of 106 plaque forming units of SARS-CoV-2. Lower viral replication in nasal swabs and broncho-alveolar lavage (BAL) as well as fewer SARS-CoV-2 infected cells and immune cell infiltrates in the lungs concomitant with reduced levels of pro-inflammatory cytokines and chemotactic factors in BAL were observed in the animals immunized with CoVLP adjuvanted with AS03. No clinical, pathologic or virologic evidences of vaccine associated enhanced disease (VAED) were observed in vaccinated animals. CoVLP adjuvanted with AS03 was therefore selected for vaccine development and clinical trials.

immunology↗

bioRxiv2021

SARS-CoV-2 Variant of Concern Substitutions Alter Spike Glycoprotein Receptor Binding Domain Structure and Stability

The emergence of severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) and the subsequent COVID-19 pandemic has visited a terrible cost on the world in the forms of disease, death, and economic turmoil. The rapid development and deployment of extremely effective vaccines against SARS-CoV-2 have seemingly brought within reach the end of the pandemic. However, the virus has acquired mutations; and emerging variants of concern (VOC) are more infectious and reduce the efficacy of existing vaccines. While promising efforts to combat these variants are underway, the evolutionary pressures leading to these variants are poorly understood. To that end, here we have studied the effects on the structure and function of the SARS-CoV-2 spike glycoprotein receptor-binding domain of three amino-acid substitutions found in several variants of concern, including alpha (B.1.1.7), beta (B.1.351), and gamma (P.1). We found that these substitutions alter the RBD structure, stability, and ability to bind to ACE2, in such a way as to possibly have opposing and compensatory effects. These findings provide new insights into how these VOC may have been selected for infectivity while maintaining the structure and stability of the receptor binding domain.

biochemistry↗

bioRxiv2020

Cellular events of acute, resolving or progressive COVID-19 in SARS-CoV-2 infected non-human primates

We investigated the immune events following SARS-CoV-2 infection, from the acute inflammatory state up to four weeks post infection, in non-human primates (NHP) with heterogeneous pulmonary pathology. The acute phase was characterized by a robust and rapid migration of monocytes expressing CD16 from the blood and concomitant increase in CD16+ macrophages in the lungs. We identified two subsets of interstitial macrophages (HLA-DR+ CD206-), a transitional CD11c+ CD16+ cell population that was directly associated with IL-6 levels in plasma, and one long lasting CD11b+ CD16+ cell population. Strikingly, levels of monocytes were a correlate of viral replication in bronchial brushes and we discovered TARC (CCL17) as a new potential mediator of myeloid recruitment to the lungs. Worse disease outcomes were associated with high levels of cell infiltration in lungs including CD11b+ CD16hi macrophages and CD11b+ neutrophils. Accumulation of macrophages was long-lasting and detectable even in animals with mild or no signs of disease. Interestingly, animals with anti-inflammatory responses including high IL-10:IL-6 and kynurenine to tryptophan ratios had less signs of disease. Our results unravel cellular mechanisms of COVID-19 and suggest that NHP may be appropriate models to test immune therapies.

immunology↗

bioRxiv2020

ARDS and Cytokine Storm in SARS-CoV-2 Infected Caribbean Vervets

SARS-CoV-2 induces a wide range of disease severity ranging from asymptomatic infection, to a life-threating illness, particularly in the elderly and persons with comorbid conditions. Among those persons with serious COVID-19 disease, acute respiratory distress syndrome (ARDS) is a common and often fatal presentation. Animal models of SARS-CoV-2 infection that manifest severe disease are needed to investigate the pathogenesis of COVID-19 induced ARDS and evaluate therapeutic strategies. Here we report ARDS in two aged African green monkeys (AGMs) infected with SARS-CoV-2 that demonstrated pathological lesions and disease similar to severe COVID-19 in humans. We also report a comparatively mild COVID-19 phenotype characterized by minor clinical, radiographic and histopathologic changes in the two surviving, aged AGMs and four rhesus macaques (RMs) infected with SARS-CoV-2. We found dramatic increases in circulating cytokines in three of four infected, aged AGMs but not in infected RMs. All of the AGMs showed increased levels of plasma IL-6 compared to baseline, a predictive marker and presumptive therapeutic target in humans infected with SARS-CoV-2 infection. Together, our results show that both RM and AGM are capable of modeling SARS-CoV-2 infection and suggest that aged AGMs may be useful for modeling severe disease manifestations including ARDS.

pathology↗