Durable Mixed Chimerism May Permit Subsequent Immunosuppression-Free Intestinal Transplantation - a Proof-of-Principle Study.

Intestinal transplantation (ITx) is the definitive treatment for intestinal failure but has the highest rejection rate among solid organ transplants, requiring high doses of immunosuppression with high rates of infection, graft-versus-host disease, and malignancy. Transplant tolerance would overcome the need for long-term immunosuppression. Using non-myeloablative conditioning, our laboratory has developed a novel swine model of hematopoietic stem cell transplantation (HSCT) that produces durable mixed chimerism (MC) and immune tolerance without toxicity. We investigated whether durable MC would promote tolerance of subsequently transplanted donor-matched intestinal allografts without immunosuppression. Using miniature swine with defined MHC, we performed HSCT across an MHC-Class-I haplotype mismatch. Immunosuppression was stopped by day 45. MC was evaluated by flow cytometry, and mixed lymphocyte reaction (MLR) assays were used to evaluate cellular responses. Subsequently, orthotopic ITx was performed without immunosuppression using a donor that was MHC-matched to the HSCT donor. Recipients were observed for four weeks and euthanized for tissue collection and mechanistic assays. After HSCT, the recipients developed durable multilineage MC and apparent deletional tolerance. After ITx, recipients showed no clinical or histological signs of rejection, and chimerism was unchanged. These results demonstrate the potential value of generating durable MC to achieve transplant tolerance.

Discrimination of primary and chronic cytomegalovirus infection based on humoral immune profiles in pregnancy.

BACKGROUNDMost humans have been infected with cytomegalovirus (CMV) by midlife without clinical signs of disease. However, in settings in which the immune system is undeveloped or compromised, the virus is not adequately controlled and consequently presents a major infectious cause of both congenital disease during pregnancy as well as opportunistic infection in children and adults. With clear evidence that risk to the fetus varies with gestational age at the time of primary maternal infection, further research on humoral responses to primary CMV infection during pregnancy is needed.METHODSHere, systems serology tools were applied to characterize antibody responses to CMV infection in pregnant and nonpregnant women experiencing either primary or chronic infection.RESULTSWhereas strikingly different antibody profiles were observed depending on infection status, limited differences were associated with pregnancy status. Beyond known differences in IgM responses used clinically for identification of primary infection, distinctions observed in IgA and FcγR-binding antibodies and among antigen specificities accurately predicted infection status. Machine learning was used to define the transition from primary to chronic states and predict time since infection with high accuracy. Humoral responses diverged over time in an antigen-specific manner, with IgG3 responses toward tegument decreasing over time as typical of viral infections, while those directed to pentamer and glycoprotein B were lower during acute and greatest during chronic infection.CONCLUSIONIn sum, this work provides insights into the antibody response associated with CMV infection status in the context of pregnancy, revealing aspects of humoral immunity that have the potential to improve CMV diagnostics.FUNDINGCYMAF consortium and NIH NIAID.

Intranasal M2SR (M2-deficient Single Replication) Influenza Vaccine Induces Broadly Reactive Mucosal Antibody Production in Adults.

Intranasal M2SR (M2-deficient Single Replication influenza virus) vaccine induces robust immune responses in animal models and human subjects. A high-throughput multiplexed platform was used to analyze hemagglutinin-specific mucosal antibody responses in adults after a single dose of H3N2 M2SR. Nasal swab specimens were analyzed for total and hemagglutinin-specific IgA. Significant, dose-dependent increases in mucosal antibody responses to vaccine-matched and drifted H3N2 hemagglutinin were observed in M2SR vaccinated subjects regardless of baseline serum and mucosal immune status. These data suggest that M2SR induces broadly cross-reactive mucosal immune responses which may provide better protection against drifted and newly emerging influenza strains.

Development of a large animal orthotopic intestinal transplantation model with long-term survival for study of immunologic outcomes.

Introduction: Intestinal transplantation (ITx) is the last remaining therapy for patients with intestinal failure once parenteral nutrition is no longer an option, however its use is limited by immunological complications, including high rates of rejection and morbidity associated with immunosuppression, such as infection and malignancy. We aimed to develop a large animal model of ITx with which to study the immune response to ITx and to design and test tolerance induction regimens. Methods: Learning from prior complications, we developed and progressively improved both surgical methods for the donor and recipient as well as postoperative management strategies. Methods of stoma generation, bowel positioning, vessel preparation, and fluid management were optimized. The immunosuppression strategy mirrored our clinical regimen. Results: As a result of our modifications, results improved from survival less than 1 month to consistent long-term survival with good graft function. We review several techniques that were developed to avoid pitfalls that were encountered, which can be used to optimize outcomes in this model. Discussion: Achieving long-term survival after swine orthotopic ITx permits immunological analysis and pre-clinical trials in a large animal model of ITx.

Update on Maintenance Immunosuppression in Intestinal Transplantation.

Outcomes in intestinal transplantation remain hampered by higher rates of rejection than any other solid organs. However, maintenance immunosuppression regimens have largely remained unchanged despite advances in therapies for induction and treatment of rejection and graft-versus-host disease. Recently, there have been a small number of new maintenance therapies attempted, and older agents have been used in new ways to achieve better outcomes. The authors herein review the traditional maintenance therapies and their mechanisms and then consider updates in new therapies and new ways of using old therapies for maintenance immunosuppression after intestinal transplantation.

Dynamic establishment and maintenance of the human intestinal B cell population and repertoire following transplantation in a pediatric-dominated cohort.

Introduction: It is unknown how intestinal B cell populations and B cell receptor (BCR) repertoires are established and maintained over time in humans. Following intestinal transplantation (ITx), surveillance ileal mucosal biopsies provide a unique opportunity to map the dynamic establishment of recipient gut lymphocyte populations in immunosuppressed conditions. Methods: Using polychromatic flow cytometry that includes HLA allele group-specific antibodies distinguishing donor from recipient cells along with high throughput BCR sequencing, we tracked the establishment of recipient B cell populations and BCR repertoire in the allograft mucosa of ITx recipients. Results: We confirm the early presence of naïve donor B cells in the circulation (donor age range: 1-14 years, median: 3 years) and, for the first time, document the establishment of recipient B cell populations, including B resident memory cells, in the intestinal allograft mucosa (recipient age range at the time of transplant: 1-44 years, median: 3 years). Recipient B cell repopulation of the allograft was most rapid in infant (<1 year old)-derived allografts and, unlike T cell repopulation, did not correlate with rejection rates. While recipient memory B cell populations were increased in graft mucosa compared to circulation, naïve recipient B cells remained detectable in the graft mucosa for years. Comparisons of peripheral and intra-mucosal B cell repertoires in the absence of rejection (recipient age range at the time of transplant: 1-9 years, median: 2 years) revealed increased BCR mutation rates and clonal expansion in graft mucosa compared to circulating B cells, but these parameters did not increase markedly after the first year post-transplant. Furthermore, clonal mixing between the allograft mucosa and the circulation was significantly greater in ITx recipients, even years after transplantation, than in deceased adult donors. In available pan-scope biopsies from pediatric recipients, we observed higher percentages of naïve recipient B cells in colon allograft compared to small bowel allograft and increased BCR overlap between native colon vs colon allograft compared to that between native colon vs ileum allograft in most cases, suggesting differential clonal distribution in large intestine vs small intestine. Discussion: Collectively, our data demonstrate intestinal mucosal B cell repertoire establishment from a circulating pool, a process that continues for years without evidence of stabilization of the mucosal B cell repertoire in pediatric ITx patients.

Characteristics and Functions of Infection-enhancing Antibodies to the N-terminal Domain of SARS-CoV-2.

Background: Fcγ-receptor (FcγR)-independent enhancement of SARS-CoV-2 infection mediated by N-terminal domain (NTD)-binding monoclonal antibodies (mAbs) has been observed in vitro, but the functional significance of these antibodies in vivo is less clear. Methods: We characterized 1,213 SARS-CoV-2 spike (S)-binding mAbs derived from COVID-19 convalescent patients for binding specificity to the SARS-CoV-2 S protein, VH germ-line usage, and affinity maturation. Infection enhancement in a vesicular stomatitis virus (VSV)-SARS-CoV-2 S pseudovirus (PV) assay was characterized in respiratory and intestinal epithelial cell lines, and against SARS-CoV-2 variants of concern (VOC). Proteomic deconvolution of the serum antibody repertoire was used to determine functional attributes of secreted NTD-binding mAbs. Results: We identified 72/1213 (5.9%) mAbs that enhanced SARS-CoV-2 infection in a 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 5 months post-infection. Infection enhancement by NTD-binding mAbs was not observed in intestinal and respiratory epithelial cell lines and was diminished or lost against SARS-CoV-2 VOC. Proteomic deconvolution of the serum antibody repertoire from 2 of the convalescent patients identified, for the first time, NTD-binding, infection-enhancing mAbs among the circulating immunoglobulins directly isolated from serum. Functional analysis of these mAbs demonstrated robust activation of FcγRIIIa associated with antibody binding to recombinant S proteins. Conclusions: Functionally active NTD-specific mAbs arise frequently during natural infection and can last as major serum clonotypes during convalescence. These antibodies display functional attributes that include FcγR activation, and may be selected against by mutations in NTD associated with SARS-CoV-2 VOC.

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 γ-Protocadherin (γ-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 γ-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 versus shared domains regulate distinct γ-Pcdh functions remains incompletely understood. Our previous in vitro studies identified protein kinase C (PKC) phosphorylation of a serine residue within a shared C-terminal motif as a mechanism through which γ-Pcdh promotion of dendrite arborization via myristoylated alanine-rich C-kinase substrate (MARCKS) is abrogated. Here, we used CRISPR/Cas9 genome editing to generate two new mouse lines expressing only non-phosphorylatable γ-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 remain unchanged in both PcdhgS/A and PcdhgCTD cortex. 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 γ-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 γ-Pcdh negative regulation point and contributes to a converging understanding of γ-Pcdh family function in which distinct roles are played by both individual isoforms and discrete protein domains.

Nasal and pharyngeal mucosal immunity to poliovirus in children following routine immunization with inactivated polio vaccine (IPV) in the United States of America.

BACKGROUND: Although polioviruses (PVs) replicate in lymphoid tissue of both the pharynx and ileum, research on polio vaccine-induced mucosal immunity has predominantly focused on intestinal neutralizing and binding antibody levels measured in stool. METHODS: To investigate the extent to which routine immunization with intramuscularly injected inactivated polio vaccine (IPV) may induce nasal and pharyngeal mucosal immunity, we measured PV type-specific neutralization and immunoglobulin (Ig) G, IgA, and IgM levels in nasal secretions, adenoid cell supernatants, and sera collected from 12 children, aged 2 to 5 years, undergoing planned adenoidectomies. All participants were routinely immunized with IPV and had no known contact with live PVs. RESULTS: PV-specific mucosal neutralization was detected in nasal and adenoid samples, mostly from children who had previously received four IPV doses. Across the three PV serotypes, both nasal (Spearman's rho ≥ 0.87, p≤0.0003 for all) and adenoid (Spearman's rho ≥0.57, p≤0.05 for all) neutralization titers correlated with serum neutralization titers. In this small study sample, there was insufficient evidence to determine which Ig isotype(s) was correlated with neutralization. CONCLUSIONS: Our findings provide policy-relevant evidence that routine immunization with IPV may induce nasal and pharyngeal mucosal immunity. The observed correlations of nasal and pharyngeal mucosal neutralization with serum neutralization contrast with previous observations of distinct intestinal and serum responses to PV vaccines. Further research is warranted to determine which antibody isotype(s) correlate with polio vaccine-induced nasal and pharyngeal mucosal neutralizing activity and to understand the differences from intestinal mucosal immunity.

Selection of positive controls and their impact on anti-drug antibody assay performance.

Development of assays to reliably identify and characterize anti-drug antibodies (ADAs) depends on positive control anti-idiotype (anti-id) reagents, which are used to demonstrate that the standards recommended by regulatory authorities are met. This work employs a set of therapeutic antibodies under clinical development and their corresponding anti-ids to investigate how different positive control reagent properties impact ADA assay development. Positive controls exhibited different response profiles and apparent assay analytical sensitivity values depending on assay format. Neither anti-id affinity for drug, nor sensitivity in direct immunoassays related to sensitivity in ADA assays. Anti-ids were differentially able to detect damage to drug conjugates used in bridging assays and were differentially drug tolerant. These parameters also failed to relate to assay sensitivity, further complicating selection of anti-ids for use in ADA assay development based on functional characteristics. Given this variability among anti-ids, alternative controls that could be employed across multiple antibody drugs were investigated as a more uniform means to define ADA detection sensitivity across drug products and assay protocols, which could help better relate assay results to clinical risks of ADA responses. Overall, this study highlights the importance of positive control selection to reliable detection and clinical interpretation of the presence and magnitude of ADA responses.

Astrocyte morphogenesis requires self-recognition.

Self-recognition is a fundamental cellular process across evolution and forms the basis of neuronal self-avoidance1-4. Clustered protocadherins (Pcdh), comprising a large family of isoform-specific homophilic recognition molecules, play a pivotal role in neuronal self-avoidance required for mammalian brain development5-7. The probabilistic expression of different Pcdh isoforms confers unique identities upon neurons and forms the basis for neuronal processes to discriminate between self and non-self5,6,8. Whether this self-recognition mechanism exists in astrocytes, the other predominant cell type of the brain, remains unknown. Here, we report that a specific isoform in the Pcdhγ cluster, γC3, is highly enriched in human and murine astrocytes. Through genetic manipulation, we demonstrate that γC3 acts autonomously to regulate astrocyte morphogenesis in the mouse visual cortex. To determine if γC3 proteins act by promoting recognition between processes of the same astrocyte, we generated pairs of γC3 chimeric proteins capable of heterophilic binding to each other, but incapable of homophilic binding. Co-expressing complementary heterophilic binding isoform pairs in the same γC3 null astrocyte restored normal morphology. By contrast, chimeric γC3 proteins individually expressed in single γC3 null mutant astrocytes did not. These data establish that self-recognition is essential for astrocyte development in the mammalian brain and that, by contrast to neuronal self-recognition, a single Pcdh isoform is both necessary and sufficient for this process.

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 γ-Protocadherin (γ-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 γ-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 γ-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 γ-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 γ-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 PcdhgCTD cortex. 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 γ-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 γ-Pcdh negative regulation point, and contributes to a converging understanding of γ-Pcdh family function in which distinct roles are played by both individual isoforms and discrete protein domains.

Dynamic establishment of recipient resident memory T cell repertoire after human intestinal transplantation.

BACKGROUND: Understanding formation of the human tissue resident memory T cell (TRM) repertoire requires longitudinal access to human non-lymphoid tissues. METHODS: By applying flow cytometry and next generation sequencing to serial blood, lymphoid tissue, and gut samples from 16 intestinal transplantation (ITx) patients, we assessed the origin, distribution, and specificity of human TRMs at phenotypic and clonal levels. FINDINGS: Donor age ≥1 year and blood T cell macrochimerism (peak level ≥4%) were associated with delayed establishment of stable recipient TRM repertoires in the transplanted ileum. T cell receptor (TCR) overlap between paired gut and blood repertoires from ITx patients was significantly greater than that in healthy controls, demonstrating increased gut-blood crosstalk after ITx. Crosstalk with the circulating pool remained high for years of follow-up. TCR sequences identifiable in pre-Tx recipient gut but not those in lymphoid tissues alone were more likely to populate post-Tx ileal allografts. Clones detected in both pre-Tx gut and lymphoid tissue had distinct transcriptional profiles from those identifiable in only one tissue. Recipient T cells were distributed widely throughout the gut, including allograft and native colon, which had substantial repertoire overlap. Both alloreactive and microbe-reactive recipient T cells persisted in transplanted ileum, contributing to the TRM repertoire. INTERPRETATION: Our studies reveal human intestinal TRM repertoire establishment from the circulation, preferentially involving lymphoid tissue counterparts of recipient intestinal T cell clones, including TRMs. We have described the temporal and spatial dynamics of this active crosstalk between the circulating pool and the intestinal TRM pool. FUNDING: This study was funded by the National Institute of Allergy and Infectious Diseases (NIAID) P01 grant AI106697.

The clustered gamma protocadherin PcdhγC4 isoform regulates cortical interneuron programmed cell death in the mouse cortex.

Cortical inhibitory interneurons (cINs) are born in the ventral forebrain and migrate into the cortex where they make connections with locally produced excitatory glutamatergic neurons. Cortical function critically depends on the number of cINs, which is also key to establishing the appropriate inhibitory/excitatory balance. The final number of cINs is determined during a postnatal period of programmed cell death (PCD) when ~40% of the young cINs are eliminated. Previous work shows that the loss of clustered gamma protocadherins (Pcdhgs), but not of genes in the Pcdha or Pcdhb clusters, dramatically increased BAX-dependent cIN PCD. Here, we show that PcdhγC4 is highly expressed in cINs of the mouse cortex and that this expression increases during PCD. The sole deletion of the PcdhγC4 isoform, but not of the other 21 isoforms in the Pcdhg gene cluster, increased cIN PCD. Viral expression of the PcdhγC4, in cIN lacking the function of the entire Pcdhg cluster, rescued most of these cells from cell death. We conclude that PcdhγC4 plays a critical role in regulating the survival of cINs during their normal period of PCD. This highlights how a single isoform of the Pcdhg cluster, which has been linked to human neurodevelopmental disorders, is essential to adjust cIN cell numbers during cortical development.

Application of enzyme-linked immunosorbent assay to detect antimicrobial peptides in human intestinal lumen.

Intestinal transplantation is the definitive treatment for intestinal failure. However, tissue rejection and graft-versus-host disease are relatively common complications, necessitating aggressive immunosuppression that can itself pose further complications. Tracking intraluminal markers in ileal effluent from standard ileostomies may present a noninvasive and sensitive way to detect developing pathology within the intestinal graft. This would be an improvement compared to current assessments, which are limited by poor sensitivity and specificity, contributing to under or over-immunosuppression, respectively, and by the need for invasive biopsies. Herein, we report an approach to reproducibly analyze ileal fluid obtained through stoma sampling for antimicrobial peptide/protein concentrations, reasoning that these molecules may provide an assessment of intestinal homeostasis and levels of intestinal inflammation over time. Concentrations of lysozyme (LYZ), myeloperoxidase (MPO), calprotectin (S100A8/A9) and ß-defensin 2 (DEFB2) were assessed using adaptations of commercially available enzyme-linked immunosorbent assays (ELISAs). The concentration of α-defensin 5 (DEFA5) was assessed using a newly developed sandwich ELISA. Our data support that with proper preparation of ileal effluent specimens, precise and replicable determination of antimicrobial peptide/protein concentrations can be achieved for each of these target molecules via ELISA. This approach may prove to be reliable as a clinically useful assessment of intestinal homeostasis over time for patients with ileostomies.

Plasticity of intragraft alloreactive T cell clones in human gut correlates with transplant outcomes.

The site of transition between tissue-resident memory (TRM) and circulating phenotypes of T cells is unknown. We integrated clonotype, alloreactivity, and gene expression profiles of graft-repopulating recipient T cells in the intestinal mucosa at the single-cell level after human intestinal transplantation. Host-versus-graft (HvG)-reactive T cells were mainly distributed to TRM, effector T (Teff)/TRM, and T follicular helper compartments. RNA velocity analysis demonstrated a trajectory from TRM to Teff/TRM clusters in association with rejection. By integrating pre- and post-transplantation (Tx) mixed lymphocyte reaction-determined alloreactive repertoires, we observed that pre-existing HvG-reactive T cells that demonstrated tolerance in the circulation were dominated by TRM profiles in quiescent allografts. Putative de novo HvG-reactive clones showed a transcriptional profile skewed to cytotoxic effectors in rejecting grafts. Inferred protein regulon network analysis revealed upstream regulators that accounted for the effector and tolerant T cell states. We demonstrate Teff/TRM interchangeability for individual T cell clones with known (allo)recognition in the human gut, providing novel insight into TRM biology.

Dynamic establishment and maintenance of the human intestinal B cell population and repertoire following transplantation.

It is unknown how intestinal B cell populations and B cell receptor (BCR) repertoires are established and maintained over time in humans. Following intestinal transplantation (ITx), surveillance ileal mucosal biopsies provide a unique opportunity to map the dynamic establishment of gut lymphocyte populations. Using polychromatic flow cytometry that includes HLA allele group-specific mAbs distinguishing donor from recipient cells along with high throughput BCR sequencing, we tracked the establishment of recipient B cell populations and BCR repertoire in the allograft mucosa of ITx recipients. We confirm the early presence of naïve donor B cells in the circulation and, for the first time, document the establishment of recipient B cell populations, including B resident memory cells, in the intestinal allograft mucosa. Recipient B cell repopulation of the allograft was most rapid in infant (<1 year old)-derived allografts and, unlike T cell repopulation, did not correlate with rejection rates. While recipient memory B cell populations were increased in graft mucosa compared to circulation, naïve recipient B cells remained detectable in the graft mucosa for years. Comparisons of peripheral and intra-mucosal B cell repertoires in the absence of rejection revealed increased BCR mutation rates and clonal expansion in graft mucosa compared to circulating B cells, but these parameters did not increase markedly after the first year post-transplant. Furthermore, clonal mixing between the allograft mucosa and the circulation was significantly greater in ITx recipients, even years after transplantation, than in healthy control adults. Collectively, our data demonstrate intestinal mucosal B cell repertoire establishment from a circulating pool, a process that continues for years without evidence of establishment of a stable mucosal B cell repertoire.

Adults on pre-exposure prophylaxis (tenofovir-emtricitabine) have faster clearance of anti-HIV monoclonal antibody VRC01.

Broadly neutralizing monoclonal antibodies (mAbs) are being developed for HIV-1 prevention. Hence, these mAbs and licensed oral pre-exposure prophylaxis (PrEP) (tenofovir-emtricitabine) can be concomitantly administered in clinical trials. In 48 US participants (men and transgender persons who have sex with men) who received the HIV-1 mAb VRC01 and remained HIV-free in an antibody-mediated-prevention trial (ClinicalTrials.gov #NCT02716675), we conduct a post-hoc analysis and find that VRC01 clearance is 0.08 L/day faster (p = 0.005), and dose-normalized area-under-the-curve of VRC01 serum concentration over-time is 0.29 day/mL lower (p < 0.001) in PrEP users (n = 24) vs. non-PrEP users (n = 24). Consequently, PrEP users are predicted to have 14% lower VRC01 neutralization-mediated prevention efficacy against circulating HIV-1 strains. VRC01 clearance is positively associated (r = 0.33, p = 0.03) with levels of serum intestinal Fatty Acid Binding protein (I-FABP), a marker of epithelial intestinal permeability, which is elevated upon starting PrEP (p = 0.04) and after months of self-reported use (p = 0.001). These findings have implications for the evaluation of future HIV-1 mAbs and postulate a potential mechanism for mAb clearance in the context of PrEP.