Recall Responses Research Articles

Mucosal vaccines for SARS-CoV-2: triumph of hope over experience.

Currently approved COVID-19 vaccines administered parenterally induce robust systemic humoral and cellular responses. While highly effective against severe disease, there is reduced effectiveness of these vaccines in preventing breakthrough infection and/or onward transmission, likely due to poor immunity elicited at the respiratory mucosa. As such, there has been considerable interest in developing novel mucosal vaccines that engenders more localised immune responses to provide better protection and recall responses at the site of virus entry, in contrast to traditional vaccine approaches that focus on systemic immunity. In this review, we explore the adaptive components of mucosal immunity, evaluate epidemiological studies to dissect if mucosal immunity conferred by parenteral vaccination or respiratory infection drives differential efficacy against virus acquisition or transmission, discuss mucosal vaccines undergoing clinical trials and assess key challenges and prospects for mucosal vaccine development.

Evaluation of the long-term memory T cell in mice after immunization with a live tularemia vaccine

The vaccine strain F. tularensis 15 NIIEG induces long-lived cell-mediated immunity but exhibits a certain reactogenicity and genetic instability. Progress in development of a vaccine against tularemia has been limited by a lack of information regarding the mechanisms required to protect against this disease. The BALB/c mouse is the most commonly used animal to study tularemia due to its relatively low cost, well-characterized genetics, available immunological tools and mouse infection with virulent F. tularensis recapitulates human disease.CD4+ and CD8+T cells are known to be critical for the formation of protective immunity but the relative roles of memory T cell subpopulations in long lived protection against virulent strains of F. tularensis are not well established. We hypothesized that this immunity depends on central (TCM) and effector memory (TEM) T cells and their functional activity. In this study we have dissected the T cell immune response in BALB/c mice 30, 60 and 90 days after subcutaneous vaccination with 15 NIIEG.Multiparametric flow cytometry were used to characterize in vitro recall responses of splenocytes to F. tularensis antigen. TEM cells were identified as CD3+CD4+CD44+CD62L- and CD3+CD8+CD44+CD62L-, TCM cells as CD3+CD4+CD44+CD62L+ and CD3+CD8+CD44+CD62L+, respectively. The functional activity of memory T cells was assessed by the following parameters: the level of expression of the activation marker CD69 and cytokine-producing activity by staining with the intracellular cytokines IFNg and TNFa.Thus, development of a long-lived vaccine directed against F. tularensis is dependent on identifying not only the correlates of immunity present early after vaccination, but also those that persist in the host after the effector phase has ended. The maintenance of long-term protective immunity initiated by vaccination with F. tularensis strain 15 NIIEG has been shown to require the presence of antigen-specific CD4+ and CD8+ memory T cells producing IFNg and TNFa and expressing the activation marker CD69. A decrease in count and functional activity of CD8+TCM and CD8+TEM was detected in the long term after vaccination. The detected parameters of functional activity of memory T cells can be used as criteria for evaluation of protective immunity against virulent strains of F. tularensis.

Viral vector delivered immunogen focuses HIV-1 antibody specificity and increases durability of the circulating antibody recall response.

The modestly efficacious HIV-1 vaccine regimen (RV144) conferred 31% vaccine efficacy at 3 years following the four-shot immunization series, coupled with rapid waning of putative immune correlates of decreased infection risk. New strategies to increase magnitude and durability of protective immunity are critically needed. The RV305 HIV-1 clinical trial evaluated the immunological impact of a follow-up boost of HIV-1-uninfected RV144 recipients after 6-8 years with RV144 immunogens (ALVAC-HIV alone, AIDSVAX B/E gp120 alone, or ALVAC-HIV + AIDSVAX B/E gp120). Previous reports demonstrated that this regimen elicited higher binding, antibody Fc function, and cellular responses than the primary RV144 regimen. However, the impact of the canarypox viral vector in driving antibody specificity, breadth, durability and function is unknown. We performed a follow-up analysis of humoral responses elicited in RV305 to determine the impact of the different booster immunogens on HIV-1 epitope specificity, antibody subclass, isotype, and Fc effector functions. Importantly, we observed that the ALVAC vaccine component directly contributed to improved breadth, function, and durability of vaccine-elicited antibody responses. Extended boosts in RV305 increased circulating antibody concentration and coverage of heterologous HIV-1 strains by V1V2-specific antibodies above estimated protective levels observed in RV144. Antibody Fc effector functions, specifically antibody-dependent cellular cytotoxicity and phagocytosis, were boosted to higher levels than was achieved in RV144. V1V2 Env IgG3, a correlate of lower HIV-1 risk, was not increased; plasma Env IgA (specifically IgA1), a correlate of increased HIV-1 risk, was elevated. The quality of the circulating polyclonal antibody response changed with each booster immunization. Remarkably, the ALVAC-HIV booster immunogen induced antibody responses post-second boost, indicating that the viral vector immunogen can be utilized to selectively enhance immune correlates of decreased HIV-1 risk. These results reveal a complex dynamic of HIV-1 immunity post-vaccination that may require careful balancing to achieve protective immunity in the vaccinated population. Trial registration: RV305 clinical trial (ClinicalTrials.gov number, NCT01435135). ClinicalTrials.gov Identifier: NCT00223080.

A bispecific antibody AP203 targeting PD-L1 and CD137 exerts potent antitumor activity without toxicity

BackgroundBispecific antibody has garnered considerable attention in the recent years due to its impressive preliminary efficacy in hematological malignancies. For solid tumors, however, the main hindrance is the suppressive tumor microenvironment, which effectively impedes the activation of infiltrating T cells. Herein, we designed a bispecific antibody AP203 with high binding affinity to PD-L1 and CD137 and assessed its safety and anti-tumor efficacy, as well as explored the mechanism of action.MethodsThe optimal antibody binders against PD-L1 and CD137 were screened from the OmniMab phagemid library. The binding affinity of the constructed AP203 were evaluated using enzyme-linked immunosorbent assay (ELISA) and biolayer interferometry (BLI). T-cell stimulatory capacity was assessed using the allogeneic mixed lymphocyte reaction (MLR), antigen-specific recall response, and coculture with PD-L1-expressing cells. In vivo antitumor efficacy was evaluated using two models of tumor-xenografted humanized mice with profiling of tumor infiltrating lymphocytes (TILs). The possible toxicity of AP203 was examined using in vitro cytokine release assay by human PBMCs.ResultsAP203, which simultaneously targeted PD-L1 and costimulatory CD137, elicit superior agonistic effects over parental antibodies alone or in combination in terms of T cell activation, enhanced memory recall responses, and overcoming Treg-mediated immunosuppression (P < 0.05). The agonistic activity of AP203 was further demonstrated PD-L1-dependent by coculturing T cells with PD-L1-expressing cells. In vivo animal studies using immunodeficient or immunocompetent mice both showed a dose-related antitumor efficacy superior to parental antibodies in combination (P < 0.05). Correspondingly, AP203 significantly increased tumor infiltrating CD8 + T cells, while decreased CD4 + T cells, as well as Treg cells (P < 0.05), resulting in a dose-dependent increase in the CD8 + /CD4 + ratio. Moreover, either soluble or immobilized AP203 did not induce the production of inflammatory cytokines by human PBMCs.ConclusionsAP203 exerts potent antitumor activity not only by blocking PD-1/PD-L1 inhibitory signaling, but also by activating CD137 costimulatory signaling in effector T cells that consequently counteracts Treg-mediated immunosuppression. Based on promising preclinical results, AP203 should be a suitable candidate for clinical treatment of solid tumors.

Functional CVIDs phenotype clusters identified by the integration of immune parameters after BNT162b2 boosters.

Assessing the response to vaccinations is one of the diagnostic criteria for Common Variable Immune Deficiencies (CVIDs). Vaccination against SARS-CoV-2 offered the unique opportunity to analyze the immune response to a novel antigen. We identify four CVIDs phenotype clusters by the integration of immune parameters after BTN162b2 boosters. We performed a longitudinal study on 47 CVIDs patients who received the 3rd and 4th vaccine dose of the BNT162b2 vaccine measuring the generation of immunological memory. We analyzed specific and neutralizing antibodies, spike-specific memory B cells, and functional T cells. We found that, depending on the readout of vaccine efficacy, the frequency of responders changes. Although 63.8% of the patients have specific antibodies in the serum, only 30% have high-affinity specific memory B cells and generate recall responses. Thanks to the integration of our data, we identified four functional groups of CVIDs patients with different B cell phenotypes, T cell functions, and clinical diseases. The presence of antibodies alone is not sufficient to demonstrate the establishment of immune memory and the measurement of the in-vivo response to vaccination distinguishes patients with different immunological defects and clinical diseases.

Untangling associations between immunoglobulin genotypes, repertoires and function

Immunoglobulin (IG) genes, encoding B cell receptors (BCRs), are fundamental components of the mammalian immune system, which evolved to recognize the diverse antigenic universe present in nature. To handle these myriad inputs, BCRs are generated through combinatorial recombination of a set of highly polymorphic germline genes, resulting in a vast repertoire of antigen receptors that initiate responses to pathogens and regulate commensals. Following antigen recognition and B cell activation, memory B cells and plasma cells form, allowing for the development of anamnestic antibody (Ab) responses. How inherited variation in IG genes impacts host traits, disease susceptibility, and Ab recall responses is a topic of great interest. Here, we consider approaches to translate emerging knowledge about IG genetic diversity and expressed repertoires to inform our understanding of Ab function in health and disease etiology. As our understanding of IG genetics grows, so will our need for tools to decipher preferences for IG gene or allele usage in different contexts, to better understand antibody responses at the population level.

Pro-resolution lipid mediator maresin-1 reduces inflammation, promotes neuroprotection, and prevents disease progression in multiple sclerosis model of autoimmunity

Abstract Multiple sclerosis (MS) is the most common inflammatory neurodegenerative disease in young adults, resulting in neurological defects and disability. The endogenous mechanisms for resolving inflammation are intact but become defective in patients, resulting in a lack of resolution mediators and unresolved inflammation. Because DHA metabolism is impaired in MS, we hypothesize that supplementing its downstream metabolite maresin 1 (MaR1) will alleviate inflammation and demyelination in animal model of MS called experimental allergic encephalomyelitis (EAE). EAE was induced in SJL mice, followed by treatment with MaR1 (300ng, 200ul in PBS/animal/day ip) on day 6 after disease induction. Monitoring the disease course, recall response by ELISA, cytokine expression analysis by qPCR and western blotting, and immune profiling by flow cytometry were used to assess the effect of MaR1 treatment in EAE. The neuroprotective effect of MaR1 was also assessed using single molecule array (SIMOA), histopathology, and IHC. Statistical analysis was done using Graph-Pad Prism. Restoration of MaR1 had a protective effect on neurological deficits, prevented disease progression, and reduced disease severity in EAE by reducing immune cell infiltration (CD4 +IL17 +and CD4 +IFN +) into the CNS (P&amp;lt;0.001). It significantly reduced the proinflammatory cytokine IL17 (P&amp;lt;0.01) and promoted an anti-inflammatory response via IL10 and IL4 (P&amp;lt;0.001). Furthermore, it exerted neuroprotective effects as evidenced by higher MBP expression in the brain from the MaR1 treated group. Overall, MaR1 supplementation has anti-inflammatory and neuroprotective effects in preclinical animal models, implying that it could be a new therapeutic molecule in MS. Supported by grants from National Multiple Sclerosis Society (US) Research Grant (RG-2111-38733, RG-1807–31964 and RG-1508– 05912), the US National Institutes of Health Grant (NS112727 and AI144004), and Henry Ford Hospital Internal Grant (A10270 and A30967) to SG.

Boosting location of COVID-19 mRNA vaccine regulates the germinal center reaction

Abstract Germinal centers (GCs) are the specialized sites for B cells to differentiate into long-lived plasma cells or memory B cells during vaccinations. COVID-19 mRNA vaccines require boosting to mount high levels of neutralizing antibody responses. mRNA vaccines induce persistent GCs in the draining lymph nodes (dLNs), which might change the microenvironment for subsequent immunizations. It is unclear whether the site of booster immunization (ipsilateral versus contralateral) will affect the GC responses in the dLNs. To test this, we compared GC responses induced by ipsilateral and contralateral booster vaccination three weeks post the primary immunization in mice. Although these two boosting strategies induce similar numbers of GC B cells in the dLNs, we discovered that ipsilateral boosting elicits more SARS-CoV-2 RBD-specific GC B cells than contralateral boosting. However, very few RBD-specific GC B cells were differentiating into plasma cells as probed by BCL6 lowIRF4 highPC precursors within the GC B cell population. Serum antibody levels were comparable between these boosting strategies, suggesting the memory recall response was not different. These studies suggest boosting location of COVID19 mRNA vaccines might affect RBD-specific GC B cell responses. Future studies are needed to investigate whether boosting location may impact the persistence of neutralizing antibody response induced by COVID19 mRNA vaccines.

Investigating the Distinct Tissue Repair and Immunosuppressive Roles of Regulatory T cells in HSV-2 Infection

Abstract Herpes Simplex Virus 2 (HSV-2) is the primary cause of genital herpes, affecting 500 million people worldwide of which approximately 64% are women. Despite this disproportionately high burden, the female reproductive tract and its unique mucosal immune environment is surprisingly understudied. Tregs are necessary for CD4 +T cell priming in the draining lymph node (dLN), though their role in shaping tissue-resident memory T cell (T rm) responses during HSV-2 reactivation is unclear. Our preliminary data indicate that activated Tregs accumulate and persist in mouse vaginal tissue (VT) after above baseline out to at least 90 days post-infection (p.i.), thus implicating Tregs in memory recall responses to reinfection. We hypothesize that Tregs restrain robust T rmrecall responses to HSV-2 challenge. Additionally, we hypothesize that vaginal Tregs promote tissue healing in the VT in response to inflammatory alarmins through the expression of the epidermal growth factor ligand, amphiregulin (Areg). Female FoxP3 DTRmice were infected intravaginally with attenuated HSV-2, then systemically depleted of Tregs via intraperitoneal injection of diphtheria toxin and challenged with WT HSV-2 on day 30 p.i. Systemically Treg-depleted mice show increased activation and proliferation of T cells, and increased frequency of cytotoxic HSV-2-specific CD8 +T cells in the VT by day 3 post-challenge. Treg-depleted mice also have significantly higher pathology scores in H&amp;E-stained VT. We also report that VT Tregs from infected mice express more Areg than dLN Tregs via flow cytometry and single cell RNAseq. Ongoing work includes characterization of Areg expressing VT Tregs and further dissection of the T rmrecall responses in Treg-depleted mice. NIH (T32 AI07140, R01 AI141435)

A 47-marker immune profiling flow cytometry assay to enable comprehensive antigen-specific immune analysis

Abstract The complex nature of the immune system requires deep interrogation at the single-cell level. Flow cytometry using CyTOF® technology employs antibodies tagged with isotopically enriched metals instead of fluorophores, resulting in discrete signals that facilitate highly multiparametric characterization on a single-cell level. CyTOF technology enables analysis of over 50 parameters simultaneously without the impediments of spectral overlap or autofluorescence, allowing for a reliable evaluation of immune responses. The Maxpar® Direct™ Immune Profiling Assay™ is a pre-titrated, dried-down, and validated 30-marker antibody cocktail for single-tube staining of human whole blood or PBMC and acquisition on CyTOF systems. Using the assay and Maxpar Pathsetter™ software, stained samples are automatically resolved into 37 immune populations. In this study, we highlight the customizability of the Maxpar® Direct panel by adding an array of 17 surface and intracellular parameters with the Maxpar Direct™ T Cell Activation Expansion Panel and live-cell barcoding. The simultaneous detection of key surface and intracellular markers in this expanded panel enabled comprehensive analysis of immune cell activation and antigen-specific recall responses to PBMC stimulated with a pool of immunodominant microbial peptides. Taken together, this 47-marker panel and automated analysis unlocked deeper immunophenotyping and functional profiling of activated immune cells and provided insight into the cell-mediated adaptive immune response to foreign targets. Such phenomena are hallmarks for research on infection, vaccine development, and immunotherapy. For Research Use Only. Not for use in diagnostic procedures.

Human memory B cells show plasticity and adopt multiple fates upon recall response to SARS-CoV-2

The B cell response to different pathogens uses tailored effector mechanisms and results in functionally specialized memory B (Bm) cell subsets, including CD21+ resting, CD21–CD27+ activated and CD21–CD27– Bm cells. The interrelatedness between these Bm cell subsets remains unknown. Here we showed that single severe acute respiratory syndrome coronavirus 2-specific Bm cell clones showed plasticity upon antigen rechallenge in previously exposed individuals. CD21– Bm cells were the predominant subsets during acute infection and early after severe acute respiratory syndrome coronavirus 2-specific immunization. At months 6 and 12 post-infection, CD21+ resting Bm cells were the major Bm cell subset in the circulation and were also detected in peripheral lymphoid organs, where they carried tissue residency markers. Tracking of individual B cell clones by B cell receptor sequencing revealed that previously fated Bm cell clones could redifferentiate upon antigen rechallenge into other Bm cell subsets, including CD21–CD27– Bm cells, demonstrating that single Bm cell clones can adopt functionally different trajectories.

Remibrutinib Inhibits Neuroinflammation Driven by B Cells and Myeloid Cells in Preclinical Models of Multiple Sclerosis (P2-3.013)

<h3>Objective:</h3> To assess the mechanism of action and efficacy of remibrutinib in experimental autoimmune encephalomyelitis (EAE) mouse models of multiple sclerosis (MS). <h3>Background:</h3> Bruton’s tyrosine kinase (BTK) is a key node in B-cell receptor and fragment crystallizable receptor signaling. BTK inhibitors (BTKis) are an emerging oral option for patients with MS. Remibrutinib (LOU064) is a potent and highly selective covalent BTKi with promising preclinical and clinical profiles for the treatment of MS. <h3>Design/Methods:</h3> Two C57BL/6 mouse EAE models (induced by immunization with human or rat myelin oligodendrocyte glycoprotein [HuMOG and RatMOG EAE, respectively]) were used. Target engagement was assessed in tissue and clinical disease activity was determined. Serum antibody levels, biomarkers, and central nervous system tissue transcriptome were analyzed. <h3>Results:</h3> Remibrutinib inhibited B-cell–dependent HuMOG EAE at daily oral doses of 3 and 30 mg/kg and strongly reduced neurological symptoms. The ex vivo MOG-specific T-cell recall response was inhibited, but the polyclonal T-cell response was not, indicating selective B-cell inhibition. Remibrutinib did not decrease total immunoglobulin G antibody levels. Remibrutinib demonstrated strong BTK occupancy in the peripheral immune organs and brain at the efficacious dose of 30 mg/kg. Remibrutinib also inhibited RatMOG EAE, indicating that myeloid cell and microglia inhibition contributes to its efficacy in MS; this is supported by anti-inflammatory effects detected via single-cell RNA sequencing of the mouse brain and spinal cord. Remibrutinib also significantly reduced neurofilament light chain levels in serum. <h3>Conclusions:</h3> Remibrutinib exhibited dose-dependent efficacy in B-cell–driven EAE models. Potential for efficacy and anti-inflammatory effects on myeloid cells and microglia was observed. Remibrutinib may represent a novel treatment option for patients with MS. <b>Disclosure:</b> Bernd C. Kieseier, MD has nothing to disclose. Mr. Cenni has received personal compensation for serving as an employee of Novartis. Dr. Nuesslein-Hildesheim has received personal compensation for serving as an employee of Novartis Pharma AG. Dr. Nuesslein-Hildesheim has stock in Novartis Pharma AG. Dr. Ferrero has received personal compensation for serving as an employee of Novartis. Catherine Huck has received personal compensation for serving as an employee of Novartis. Mr. Eichlisberger has nothing to disclose. Dr. Ziehn has received personal compensation for serving as an employee of Novartis AG.

An empirical investigation of electronic word-of-mouth: product recall and crisis response strategy in social media

ABSTRACT Product recall crises can damage reputation, result in financial losses, and generate negative word-of-mouth from customers. Appropriate response strategies on social media platforms can help companies minimise losses and save electronic word-of-mouth (eWOM). This study analysed 32,697 customer comments from Facebook to explore customer perceptions and examine the performance of different response strategies. Findings showed that the accommodative response strategy improves overall eWOM compared to the indifferent response strategy. The study provides a comprehensive understanding of the impact of social media crisis response strategies on eWOM and offers evidential managerial insights for better customer perceptions.

Anti-CD3 inhibits circulatory and tissue-resident memory CD4 T cells that drive asthma exacerbations in mice.

Exacerbations of asthma are thought to be strongly dependent on reactivation of allergen-induced lung tissue-resident and circulatory memory CD4 T cells. Strategies that broadly inhibit multiple T cell populations might then be useful to limit asthma. Accordingly, we tested whether targeting CD3 during exposure to inhaled allergen could prevent the accumulation of lung-localized effector memory CD4 T cells and block exacerbations of asthmatic inflammation. House dust mite-sensitized and repetitively challenged BL/6 mice were transiently treated therapeutically with F(ab')2 anti-CD3ε and memory T cell responses and lung inflammation were assessed. PBMCs from HDM-allergic donors were examined for the effect of anti-CD3 on expansion of allergen-reactive T cells. Allergen-sensitized mice undergoing exacerbations of asthma were protected from lung inflammation by transient therapeutic treatment with F(ab')2 anti-CD3. Regardless of whether sensitized mice underwent a secondary or tertiary recall response to inhaled allergen, anti-CD3 inhibited all phenotypes of effector memory CD4 T cells in the lung tissue and lung vasculature by 80%-90%, including those derived from tissue-resident and circulatory memory T cells. This did not depend on Treg cells suggesting it was primarily a blocking effect on memory T cell signaling. Correspondingly, anti-CD3 also strongly inhibited proliferation of human allergen-reactive memory CD4 T cells from allergic individuals. In contrast, the number of surviving tissue-resident memory CD4 T cells that were maintained in the lungs at later times was not robustly reduced by anti-CD3. Anti-CD3 F(ab')2 administration at the time of allergen exposure represents a viable strategy for limiting the immediate activity of allergen-responding memory T cells and asthma exacerbations.

Chitin oligomers promote lymphoid innate and adaptive immune cell activation.

Chitin is a highly abundant N-acetylglucosamine polysaccharide that has been linked to immune responses in the context of fungal infections and allergic asthma, especially to T helper 2 immune responses. Unfortunately, due to the frequent use of crude chitin preparations of unknown purity and degree of polymerization, there is still great uncertainty about how chitin activates different parts of the human immune system. We recently identified chitin oligomers of 6 N-acetylglucosamine units as the smallest immunologically active chitin motif and the innate immune receptor TLR2 as a primary chitin sensor on human and murine myeloid cells, but the response of further immune cells (e.g. lymphoid cells) to oligomeric chitin has not been investigated. Our analysis of primary human immune cells now shows that chitin oligomers activate immune responses of both innate and adaptive lymphocytes: notably, chitin oligomers activated natural killer cells but not B lymphocytes. Moreover, chitin oligomers induced maturation of dendritic cells and enabled potent CD8+ T-cell recall responses. Our results suggest that chitin oligomers not only trigger immediate innate responses in a limited range of myeloid cells but also exert critical activities across the entire human immune system. This highlights chitin oligomer immune activation as an interesting and broadly applicable potential target for both adjuvant development and therapeutic interference in chitin-mediated pathologies.

Abstract LB346: Mapping of shared tumor antigen reactivity with nanoparticle encapsulated mRNA in prostate cancer patients

Abstract Background: Prostate cancer (PC) remains refractory immune checkpoint therapies (CPT) despite having an FDA-approved cancer vaccine. PC is known to have a low mutation burden resulting in low level of neoantigens, so shared tumor associated antigens (TAAs) and translocations presumably represent a dominant source of antigens in this disease. While PC is thought to be a poorly immunogenic tumor, the frequency and breadth of tumor antigen reactivity in PC patients is unclear. Methods: We utilized novel programmable immunogenic mRNA nanoparticle platform and created formulations encoding a range of full-length prostate TAAs (TAAmRNA) to systemically study T cell immune responses to across a range of TAAs in PC patients. We administered 8 PC TAAmRNAs (NY-ESO-1, HK2, STEAP1, TMPRSS2-ERG, PAP, PSA, PSCA and PSMA) to in-vitro cultured peripheral blood mononuclear cells (PBMC) from 43 treatment-naïve localized PC patients compared to 17 age-matched non-cancer control patients. We assessed upregulation of early T cell activation markers and interferon-gamma (INFg) cytokine production through flow cytometry and ELISPOT, respectively. Results: 20% of PC patients possessed TAA-specific CD8+ T cell responses to at least one antigen. NY-ESO-1, HK2 and TMPRSS2-ERG had the highest T cell recall responses at an average of 35%, with NY-ESO-1 demonstrating the strongest responses per patient. Of the patients that respond to any given TAA, 47% of patients responded to three or more antigens, with 11% of PC patients responding to all TAAs. PC patients also showed heightened production of IFNg, with 35% of patients showing pan-responsiveness to three or more antigens. Particularly, HK2 stimulated the highest levels of INFg production. These levels of responsiveness are not seen in the control cohort. Conclusions: Encapsulated mRNA nanoparticles can be used to systematically map antigen reactivity across a cohort of cancer patients and reveal frequent pan-responsiveness to multiple TAAs in what has considered to be an immunologically cold cancer These results also reveal a hierarchy in immunogenicity of candidates that can enable future vaccine development. Continual efforts to assess the immunogenicity of shared PC TAAs in more advanced disease, and how patient responses to TAAs are modulated by immunotherapies, can aid in refining combinatorial therapies for PC. Citation Format: Elena Montauti, Ole Haabeth, Chris Rae, Samuel Deutsch, Lawrence Fong, David Oh. Mapping of shared tumor antigen reactivity with nanoparticle encapsulated mRNA in prostate cancer patients [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr LB346.

Bcl11b sustains multipotency and restricts effector programs of intestinal-resident memory CD8+ T cells.

The networks of transcription factors (TFs) that control intestinal-resident memory CD8+ T (TRM) cells, including multipotency and effector programs, are poorly understood. In this work, we investigated the role of the TF Bcl11b in TRM cells during infection with Listeria monocytogenes using mice with post-activation, conditional deletion of Bcl11b in CD8+ T cells. Conditional deletion of Bcl11b resulted in increased numbers of intestinal TRM cells and their precursors as well as decreased splenic effector and circulating memory cells and precursors. Loss of circulating memory cells was in part due to increased intestinal homing of Bcl11b-/- circulating precursors, with no major alterations in their programs. Bcl11b-/- TRM cells had altered transcriptional programs, with diminished expression of multipotent/multifunctional (MP/MF) program genes, including Tcf7, and up-regulation of the effector program genes, including Prdm1. Bcl11b also limits the expression of Ahr, another TF with a role in intestinal CD8+ TRM cell differentiation. Deregulation of TRM programs translated into a poor recall response despite TRM cell accumulation in the intestine. Reduced expression of MP/MF program genes in Bcl11b-/- TRM cells was linked to decreased chromatin accessibility and a reduction in activating histone marks at these loci. In contrast, the effector program genes displayed increased activating epigenetic status. These findings demonstrate that Bcl11b is a frontrunner in the tissue residency program of intestinal memory cells upstream of Tcf1 and Blimp1, promoting multipotency and restricting the effector program.

Abstract LB339: PARP inhibition reprograms CD8 T cells, enhancing their function and generation of prolonged memory, leading to greater anti-tumor immune response

Abstract Olaparib is the most widely studied third-generation PARP inhibitor (PARPi) in clinical practice with a significant clinical outcome in BRCA deficient tumors such as breast and ovarian cancers. However, innate and adaptive resistance in patients with DNA damage repair (DDR) gene mutations were reported after treatment with PARPi, highlighting the potential implication of other compartments such as the role of immune cells in the mechanism of resistance. Importantly, PARPi is shown to have an immune-modulatory capacity within the tumor microenvironment (TME) by activating the STING pathway and increasing interferon gamma (INF-gamma) and chemokine secretion and hence increasing the recruitment and function of CD8 T cells. In addition, PARPi is shown to affect other cells within the TME such as, macrophages, myeloid-derived suppressor cells (MDSCs) and cancer associated fibroblasts (CAFs), promoting either an immune enhancement or immune suppressor milieu that could indirectly affect CD8 T cell function. However, little is known about the direct effect of PARPi on CD8 T cells. In this study using two cold tumor models, we show that PARPi controls the tumor growth and increases survival in tumor-bearing mice. This effect is due to the enhancement of number and cytotoxicity of antigen-specific CD8 T cells and the maintenance of memory population within the TME. In vitro study shows that PARPi activates the SIRT-1/FOXO1 pathway, leading to the modification of gene expression related to memory and fatty acid oxidation (FAO) metabolism in CD8 T cells. As a result, a unique subpopulation of superior central memory cells with high recall responses and anti-tumor effects are generated after PARPi treatment. Interestingly this unique subpopulation was also generated after PARPi in human cells. Together, our findings highlight the direct effect of PARP inhibition on CD8 T cells and its ability to modulate and reprogram CD8 T cells within the TME, leading to a greater anti-tumor immune response. Citation Format: Wael Traboulsi, Subhadip Kundu, Pankaj Gaur, Zainab Ramlaoui, Dareen Sarhan, Nour Shobaki, Jacob Lee, Nazli Jafarzadeh, Simon T. Barry, Viia E. Valge-Archer, Vivek Verma, Seema Gupta, Samir N. Khleif. PARP inhibition reprograms CD8 T cells, enhancing their function and generation of prolonged memory, leading to greater anti-tumor immune response [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr LB339.

Veterans Health Administration response to 2021 recall of Philips Respironics devices: A case study.

This case study describes, for the time frame of June 2021 through August 2022, the U.S. Veterans Health Administration (VHA) organizational response to a manufacturer's recall of positive airway pressure devices used in the treatment of sleep disordered breathing. VHA estimated it could take over a year for Veterans to receive replacement devices. Veterans awaiting a replacement faced a dilemma. They could continue using the recalled devices and bear the product safety risks that led to the recall, or they could stop using them and bear the risks of untreated sleep disordered breathing. Using a program monitoring approach, we report on the processes VHA put in place to respond to the recall. Specifically, we report on the strategic, service, and operational plans associated with VHA's response to the recall for Veterans needing replacement devices. In program monitoring, the strategic plan reflects the internal process objectives for the program. The service plan articulates how the delivery of services will intersect the customer journey. The operational plan describes how the program's resources and actions must support the service delivery plan. VHA's strategic plan featured a clinician-led, as opposed to primarily legal or administrative response to the recall. The recall response team also engaged with VHA's medical ethics service to articulate an ethical framework guiding the allocation of replacement devices under conditions of scarcity. This framework proposed allocating scarce devices to Veterans according to their clinical need. The service plan invited Veterans to schedule visits with sleep providers who could assess their clinical need and counsel them accordingly. The operational plan distributed devices according to clinical need as they became available. Monitoring our program processes in real time helped VHA launch and adapt its response to a recall affecting more than 700,000 Veterans.

Abstract 3464: Dynamics of resident memory (Trm) cell responses between skin and draining lymph nodes during primary and recall responses to melanoma

Abstract Tissue resident memory (TRM) cells reside durably in tissues where they serve as a vital component of protective anti-tumor immunity. In mice, TRM cells are defined by their expression of tissue retention markers such as CD69 and CD103, and lack of egress markers such as CD62L, CCR7 and S1PR1. We previously showed that depletion of regulatory T cells induces the generation of protective CD8 Trm cells in skin and draining LNs of B16 melanoma tumor-excised mice. While tumor specific Trm cells were shown to stably reside in tissues at steady state, little is known about their migration between tissue compartments and circulation during primary and recall responses to melanoma. Using gp100-specific transgenic pmel CD8 T cells expressing a Kaede-GFP photoconvertible marker, we investigated the trafficking patterns of tumor Ag specific CD8+ T-cells that were labeled in situ by exposure to UV light in the skin. During primary tumor growth, we observed pronounced movement of antigen-experienced pmel cells through skin to tumor-draining LNs. This trafficking behavior could be inhibited by treatment with FTY720, indicating its S1PR-dependence. Such movement was no longer evident at memory timepoints. However, re-challenge with B16 cells in the dermis induced rapid pmel T cell egress to both LNs and circulation. Recallenge-mediated Trm cell skin egress was confirmed using anti-Thy1.1 monoclonal antibody to deplete circulating and lymphoid memory, while preserving skin TRM. Here we found that skin TRM proliferate and transiently increase expression of Tbet, but go on to generate a durable secondary Trm response in lymph nodes. These data highlight T cell movement between tissues during Trm programming and recall responses to cancer, and shed light on a potential mechanism for the generation TRM populations in tumor-draining lymph nodes. Moreover, our results highlight the potential for retrograde movement of Trm responses from peripheral tissue reservoirs to participate more broadly in systemic anti-tumor immunity. Citation Format: Cameron Messier, Nikhil Khatwani, Aaron Hawkes, Yuyang (Poppy) Huang, Mary Jo Turk. Dynamics of resident memory (Trm) cell responses between skin and draining lymph nodes during primary and recall responses to melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3464.