Resident CD8 Research Articles

Preexisting Skin-Resident CD8 and γδ T-cell Circuits Mediate Immune Response in Merkel Cell Carcinoma and Predict Immunotherapy Efficacy.

Merkel cell carcinoma (MCC) is an aggressive neuroendocrine skin cancer with a ∼50% response rate to immune checkpoint blockade (ICB) therapy. To identify predictive biomarkers, we integrated bulk and single-cell RNA sequencing (RNA-seq) with spatial transcriptomics from a cohort of 186 samples from 116 patients, including bulk RNA-seq from 14 matched pairs pre- and post-ICB. In nonresponders, tumors show evidence of increased tumor proliferation, neuronal stem cell markers, and IL1. Responders have increased type I/II interferons and preexisting tissue resident (Trm) CD8 or Vδ1 γδ T cells that functionally converge with overlapping antigen-specific transcriptional programs and clonal expansion of public T-cell receptors. Spatial transcriptomics demonstrated colocalization of T cells with B and dendritic cells, which supply chemokines and costimulation. Lastly, ICB significantly increased clonal expansion or recruitment of Trm and Vδ1 cells in tumors specifically in responders, underscoring their therapeutic importance. These data identify potential clinically actionable biomarkers and therapeutic targets for MCC. Significance: MCC serves as a model of ICB response. We utilized the largest-to-date, multimodal MCC dataset (n = 116 patients) to uncover unique tumor-intrinsic properties and immune circuits that predict response. We identified CD8 Trm and Vδ1 T cells as clinically actionable mediators of ICB response in major histocompatibility complex-high and -low MCCs, respectively.

Polyanhydride nanovaccine against H3N2 influenza A virus generates mucosal resident and systemic immunity promoting protection

Influenza A virus (IAV) causes significant morbidity and mortality worldwide due to seasonal epidemics and periodic pandemics. The antigenic drift/shift of IAV continually gives rise to new strains and subtypes, aiding IAV in circumventing previously established immunity. As a result, there has been substantial interest in developing a broadly protective IAV vaccine that induces, durable immunity against multiple IAVs. Previously, a polyanhydride nanoparticle-based vaccine or nanovaccine (IAV-nanovax) encapsulating H1N1 IAV antigens was reported, which induced pulmonary B and T cell immunity and resulted in cross-strain protection against IAV. A key feature of IAV-nanovax is its ability to easily incorporate diverse proteins/payloads, potentially increasing its ability to provide broad protection against IAV and/or other pathogens. Due to human susceptibility to both H1N1 and H3N2 IAV, several H3N2 nanovaccines were formulated herein with multiple IAV antigens to examine the “plug-and-play” nature of the polyanhydride nanovaccine platform and determine their ability to induce humoral and cellular immunity and broad-based protection similar to IAV-nanovax. The H3N2-based IAV nanovaccine formulations induced systemic and mucosal B cell responses which were associated with antigen-specific antibodies. Additionally, systemic and lung-tissue resident CD4 and CD8 T cell responses were enhanced post-vaccination. These immune responses corresponded with protection against both homologous and heterosubtypic IAV infection. Overall, these results demonstrate the plug-and-play nature of the polyanhydride nanovaccine platform and its ability to generate immunity and protection against IAV utilizing diverse antigenic payloads.

Abstract 3853: T cell ferroptosis attenuates antitumor immune responses in sickle cell disease

Abstract Background: Sickle cell disease (SCD) is the most common inherited blood disorder and arises from homozygosity for the HbS mutation (GAG>GTG: βGlu6Val) leading to the production of abnormal hemoglobin that cause sickle shaped red blood cells (RBCs). Individuals with SCD are at higher risk than the general population for developing hematologic malignancies or solid tumors such as renal medullary carcinoma (RMC). Although relatively rare, RMC is one of the most aggressive kidney cancers and is the third most common kidney cancer among children and young adults who harbor sickle hemoglobinopathies such as SCD or sickle cell trait (SCT) which occurs due to heterozygosity for the HbS mutation. Molecular profiling studies have shown that most cases involve biallelic inactivation of the SMARCB1 gene, either through simultaneous hemizygous loss and translocation or by homozygous loss. The loss of SMARCB1 and subsequent dysregulation of chromatin remodeling are key factors contributing to the development and progression of renal medullary carcinoma. Despite manifesting an inflamed tumor immune microenvironment, RMC consistently demonstrates resistance to conventional immune checkpoint inhibitors (ICI). Ameliorating the efficacy of immunotherapy of RMC is vital for research and potential therapeutic interventions by targeting CD8+ T cells. Methods: CyTOF was performed by using the spleen of WT mice and SCD mouse model. mRNA was isolated from CD8+ T cells of Peripheral Blood Mononuclear Cell (PBMC) collected from healthy donors and SCD patients followed by bulk RNA sequencing. Hi-C was conducted using CD8+ T cells isolated from PBMC of healthy donors and SCD patients. We employed DNA FISH (Fluorescence in Situ Hybridization) to examine alterations in gene architecture within CD8+ T cells under varying conditions. Results: RMC tumors exhibit accelerated tumor growth and reduced numbers of tumor resident CD8+ T cells in SCD mice model compared with WT control. CD8+ T cells in the SCD associated tumor microenvironment (TME) are subjected to ferroptosis by downregulation of SLC7A11; however, SMARCB1 loss promotes the resistance of RMC tumor cells to ferroptosis. Genomic architecture of CD8+ T cells is altered under SCD conditions, potentially contributing to immune dysfunction. Conclusions: Our research has identified significant alterations in the 3D genome architecture of CD8+ T cells in individuals with SCD. These alterations lead to transcriptional inhibition of SLC7A11, resulting in CD8+ T cell ferroptosis and a hindered anti-tumor immune response in RMC. These findings elucidate the mechanistic relationship of SCD with ICI resistance in cancers such as RMC and provide novel avenues for therapeutically sensitizing tumors to immunotherapies in individuals with SCD. Keywords: Sickle Cell Disease, Renal Medullary Carcinoma, SMARCB1, Ferroptosis, Genomic Architecture Alteration, Hi-C, SLC7A11 Citation Format: Zilong Zhao, Benxia Hu, Yalan Deng, Melinda Soeung, Giannicola Genovese, Wenbo Li, Pavlos Msaouel, Liuqing Yang, Chunru Lin. T cell ferroptosis attenuates antitumor immune responses in sickle cell disease [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3853.

Unleashed monocytic engagement in Sézary syndrome during the combination of anti-CCR4 antibody with type I interferon

AbstractSézary syndrome (SS) is an aggressive leukemic expansion of skin-derived malignant CD4+ T cells. Drug monotherapy often results in disease relapse because of the heterogenous nature of malignant CD4+ T cells, but how therapies can be optimally combined remains unclear because of limitations in understanding the disease pathogenesis. We identified immunologic transitions that interlink mycosis fungoides with SS using single-cell transcriptome analysis in parallel with high-throughput T-cell receptor sequencing. Nascent peripheral CD4+ T cells acquired a distinct profile of transcription factors and trafficking receptors that gave rise to antigenically mature Sézary cells. The emergence of malignant CD4+ T cells coincided with the accumulation of dysfunctional monocytes with impaired fragment crystallizable γ-dependent phagocytosis, decreased responsiveness to cytokine stimulation, and limited repertoire of intercellular interactions with Sézary cells. Type I interferon supplementation when combined with a monoclonal antibody targeting the chemokine receptor type 4 (CCR4), unleashed monocyte induced phagocytosis and eradication of Sézary cells in vitro. In turn, coadministration of interferon-α with the US Food and Drug Administration–approved anti-CCR4 antibody, mogamulizumab, in patients with SS induced marked depletion of peripheral malignant CD4+ T cells. Importantly, residual CD4+ T cells after Sézary cell ablation lacked any immunologic shifts. These findings collectively unveil an auxiliary role for augmenting monocytic activity during mogamulizumab therapy in the treatment of SS and underscore the importance of targeted combination therapy in this disease.

Resident Memory-like CD8+ T Cells Are Involved in Chronic Inflammatory and Neurodegenerative Diseases in the CNS.

Resident memory T (Trm) cells are a unique population that can survive and function in a compartmentalized tissue with inflammatory potential. We aim to investigate the alteration of Trm population in acute/chronic inflammatory and neurodegenerative diseases in the CNS. The frequencies of CD4+ and CD8+ T cells expressing both CD69 and CD103, the markers for Trm cells, were quantified in the peripheral blood and CSF (n = 80 and 44, respectively) in a cross-sectional manner. The transcriptional profile of Trm-like population in the CSF was further analyzed using a public single-cell dataset. The frequency of CD69+CD103+CD8+ T cells was strikingly higher in the CSF than in the peripheral blood (among memory fraction, 13.5% vs 0.11%, difference (mean [SE]): 13.4% [2.9]). This CD69+CD103+CD8+ T-cell population was increased in the CSF from patients with chronic inflammatory diseases including multiple sclerosis and with neurodegenerative diseases such as Parkinson disease and Alzheimer disease compared with controls (11.5%, 13.0%, 8.1% vs 2.9%, respectively). By contrast, the frequency was not altered in acute inflammatory conditions in the CNS (4.0%). Single-cell RNAseq analysis confirmed Trm signature in CD69+CD103+CD8+ T cells in the CSF, supporting their Trm-like phenotype, which was not clear in controls. Collectively, an increase in CD69+CD103+CD8+ Trm-like population in the CSF is related with both chronic neuroinflammatory and some neurodegenerative diseases in the CNS, suggesting a partially shared pathology in these diseases.

Fatty acids support the fitness and functionality of tumor resident CD8+ T cells by maintaining SCML4 expression.

CD8+ tissue-resident memory T (Trm) cells and tumor-infiltrating lymphocytes (TILs) regulate tumor immunity and immune surveillance. Characterization of Trm cells and TILs could help identify potential strategies to boost antitumor immunity. Here, we found that the transcription factor SCML4 was required for the progression and polyfunctionality of Trm cells and was associated with a better prognosis in patients with cancer. Moreover, SCML4 maintained multiple functions of TILs. Increased expression of SCML4 in CD8+ cells significantly reduced the growth of multiple types of tumors in mice, while deletion of SCML4 reduced antitumor immunity and promotes CD8+ T-cell exhaustion. Mechanistically, SCML4 recruited the HBO1-BRPF2-ING4 complex to reprogram the expression of T-cell-specific genes, thereby enhancing the survival and effector functions of Trm cells and TILs. SCML4 expression was promoted by fatty acid metabolism through mTOR-IRF4-PRDM1 signaling, and fatty acid metabolism induced epigenetic modifications that promoted tissue-resident and multifunctional gene expression in Trm cells and TILs. SCML4 increased the therapeutic effect of anti-PD-1 treatment by elevating the expression of effector molecules in TILs and inhibiting the apoptosis of TILs, which could be further enhanced by adding an inhibitor of H3K14ac deacetylation. These results provide a mechanistic perspective of functional regulation of tumor-localized Trm cells and TILs and identify an important activation target for tumor immunotherapy.

Socs1-knockout in skin-resident CD4+ T cells in a protracted contact-allergic reaction results in an autonomous skin inflammation with features of early-stage mycosis fungoides

Recent detailed genomic analysis of mycosis fungoides (MF) identified suppressor of cytokine signaling 1 (SOCS1), an inhibitor of JAK/STAT signaling, as one of the frequently deleted tumor suppressors in MF, and one-copy deletion of SOCS1 was confirmed in early-stage MF lesions. To better understand the functional role of SOCS1 in the genesis of MF, we used a genetically engineered mouse model emulating heterozygous SOCS1 loss in skin resident CD4+ T cells. In these mice an experimentally induced contact-allergic reaction was maintained for 20 weeks. Ten weeks after discontinuing contact-allergic challenges, only the skin with locally one-copy deletion of Socs1 in CD4+ T cells still showed high numbers of CD3+/CD4+ Socs1 k.o. cells in the dermis (p ＜ 0.0001) with prevalent Stat3 activation (p ＜0.001). And in one out of 9 mice, this had progressed to far more dramatic increases, including the thickened epidermis, and with an explosive growth of Socs1 k.o. T cells in circulation; indicative of cutaneous lymphoma. Hence, we show that Socs1 mono-allelic loss in CD4+ T cells locally in protractedly inflamed skin results in autonomous skin inflammation with features of early-stage MF.

Cross-presenting Langerhans cells are required for the early reactivation of resident CD8+ memory T cells in the epidermis

Tissue-resident memory CD8+ T cells (TRM) reside at sites of previous infection, providing protection against reinfection with the same pathogen. In the skin, TRM patrol the epidermis, where keratinocytes are the entry site for many viral infections. Epidermal TRM react rapidly to cognate antigen encounter with the secretion of cytokines and differentiation into cytotoxic effector cells, constituting a first line of defense against skin reinfection. Despite the important protective role of skin TRM, it has remained unclear, whether their reactivation requires a professional antigen-presenting cell (APC). We show here, using a model system that allows antigen targeting selectively to keratinocytes in a defined area of the skin, that limited antigen expression by keratinocytes results in rapid, antigen-specific reactivation of skin TRM. Our data identify epidermal Langerhans cells that cross-present keratinocyte-derived antigens, as the professional APC indispensable for the early reactivation of TRM in the epidermal layer of the skin.

IKK2/NFkB signaling controls lung resident CD8+ T cell memory during influenza infection

CD8+ T cell tissue resident memory (TRM) cells are especially suited to control pathogen spread at mucosal sites. However, their maintenance in lung is short-lived. TCR-dependent NFkB signaling is crucial for T cell memory but how and when NFkB signaling modulates tissue resident and circulating T cell memory during the immune response is unknown. Here, we find that enhancing NFkB signaling in T cells once memory to influenza is established, increases pro-survival Bcl-2 and CD122 levels thus boosting lung CD8+ TRM maintenance. By contrast, enhancing NFkB signals during the contraction phase of the response leads to a defect in CD8+ TRM differentiation without impairing recirculating memory subsets. Specifically, inducible activation of NFkB via constitutive active IKK2 or TNF interferes with TGFβ signaling, resulting in defects of lung CD8+ TRM imprinting molecules CD69, CD103, Runx3 and Eomes. Conversely, inhibiting NFkB signals not only recovers but improves the transcriptional signature and generation of lung CD8+ TRM. Thus, NFkB signaling is a critical regulator of tissue resident memory, whose levels can be tuned at specific times during infection to boost lung CD8+ TRM.

O-265 Increased CD8+ T-cell exhaustion due to high NKG2A/HLA-E engagement and IL-15 induction in adenomyosis

Abstract Study question Is there dysregulation of immune cells in the local uterine microenvironment of patients with adenomyosis? If any, what’s the underlying mediators and mechanisms? Summary answer Increased CD8+ T-cell exhaustion due to enhanced HLA-E/NKG2A engagement and IL-15 induction in ectopic adenomyotic lesions existed in human patients and mice models with adenomyosis. What is known already The immunological dysfunction has long been considered in the pathogenesis of adenomyosis. However, high heterogeneity in immune cell alterations and contradictory results were reported due to the technique limitation of immunohistochemistry mainly used in previous studies. A comprehensive picture of the dysregulation of immune cells in the local uterine microenvironment of patients with adenomyosis is still lacking. Specially, increased number of CD8+ T cells have been reported in adenomyotic patients, whether functional deficiency of CD8+ T cells may contribute to the abnormal accumulation and residency of ectopic endometrial cells at pathological sites remains unknown. Study design, size, duration The paired ectopic lesions and eutopic endometrium from 80 patients with adenomyosis, and normal endometrium and myometrium from 62 patients with uterine myoma (controls) were collected between 2021 to 2022. Mice models with adenomyosis by neonatal tamoxifen treatment were established for experiments in 2022. Participants/materials, setting, methods The alteration of immune subsets, NKG2A expression, exhausted phenotypes and CD8+ T-cell dysfunction by flow cytometry, the expression/localization of HLA-E and IL-15 by immunohistochemistry were investigated in uterine tissues of patients and mice models with adenomyosis. CD8+ T cells were sorted, and in-vitro cultured with recombinant human transforming growth factor-β (rhTGF-β) or rhIL-15 to determine the NKG2A expression. Correlation analyses were conducted for association between immune disturbances and disease severity (dysmenorrhea, local/diffuse lesion, CA125 level). Main results and the role of chance We found that an increase in CD8+ T cell number was the predominant alteration in ectopic lesions in patients with adenomyosis, and it was significantly associated with the severity of adenomyosis. For CD8+ T cells in adenomyotic lesions, the expression of NKG2A was positively correlated with CD94 expression, and CD94/NKG2A mainly enriched on CD103+ tissue resident CD8+ T subsets. The NKG2A+CD8+ T cells showed exhausted phenotypes characterized by co-expression of multiple immune inhibitory molecules (PD1, LAG3, TIM3, and TOX) and decreased degranulation capability demonstrated by reduced expression of perforin, granzyme B, and CD107. Importantly, the exhausted NKG2A+CD8+ T-cell subset was associated with disease severity and was significantly increased in ectopic uterine tissues of human patients and mice models with adenomyosis. The expression of NKG2A on CD8+ T cells was markedly upregulated by exogenous rhIL-15 but not rhTGF-β treatment. Additionally, we also found increased co-expression of IL-15 and the NKG2A ligand HLA-E on the glandular epithelial cells, where the NKG2A+CD8+ T cells were closely distributed around the gland, in ectopic adenomyotic microenvironment of patients and mice models with adenomyosis. Limitations, reasons for caution The control uterine samples were collected from patients with uterine myoma. Although they were pathologically confirmed as normal, the potential effects are unknown and cannot be excluded. Further studies in larger independent populations and exploration on the functional significance of CD8+ T-cell exhaustion in adenomyosis pathogenesis is warranted. Wider implications of the findings Our study reveals a previously unrecognized role for CD8+ T-cell exhaustion in the pathogenesis of adenomyosis and suggests that therapeutic interventions targeting and reinvigorating exhausted CD8+ T cells by NKG2A blockade may be beneficial for patients with adenomyosis. Trial registration number not applicable

Cutaneous T cells promote distinct outcomes in human skin structural cells associated with inflammatory disease

Abstract The skin is an immunologically active barrier tissue specialized to deal with a litany of external stresses. Immune cells coordinate with structural cells in the skin to promote appropriate inflammatory responses and subsequent restoration of barrier integrity following insult. Recent single cell analyses of human skin have defined previously unappreciated transcriptional heterogeneity in structural cells, including disease-associated changes in keratinocytes and fibroblasts. Cutaneous T cell activity is implicated in the development of inflammatory skin disease, but the mechanisms by which T cells promote disease-associated changes in the skin remain unclear. We now show that distinct subsets of circulating and resident CD4 +cutaneous T cells promote a diverse array of cytokine-dependent transcriptional outcomes in human keratinocytes and fibroblasts. We used these in vitro generated transcriptional signatures to identify T cell-dependent gene modules associated with inflammatory skin disease in vivo. For example, we find that T helper17 cells regulate the expression of epigenetic modifiers in keratinocytes, genes that are enriched in the skin of patients with psoriasis and normalized in response to anti-IL-17 therapy. Thus, we have identified T cell-dependent cytokine-driven transcriptional changes associated with inflammatory skin disease that can be used to dissect immune mechanisms of anti-cytokine therapeutic activity.

Viral infection in the brain induces priming and maintenance of a novel antigen specific CD8 T cell population within the bone marrow that dysregulates the stem cell niche

Abstract Bone marrow is a primary immune organ responsible for stem cell maintenance and hematopoiesis. While the bone marrow niche is mainly studied in the context of hematopoiesis, aging, and mechanisms underlying stem cell and plasma cell survival, changes of the niche during brain viral infections and primary T cell responses is unknown. Intracranial infection with the neurotropic Theiler’s Murine Encephalomyelitis Virus (TMEV) results in an acute infection in the C57BL/6 mouse which is cleared 30 days post infection due to generation of an antigen specific CD8 T cell response raised against the immunodominant epitope VP2 121–130. We determined that 7–10% of CD8 T cells present in the sternal and femoral bone marrows and 15–20% in the cranial bone marrow were antigen specific 7 days post intracranial TMEV infection. Over 70% of these CD8 T cells were located outside of the vasculature as determined by IV labeling. Importantly, continuous treatment with FTY720, which sequesters T cells outside of the blood, did not eliminate antigen specific CD8 T cells in any bone marrow compartment. CD8 T cells in the bone marrow established durable memory following viral clearance and were reactivated upon cognate antigen reencounter. Both acute TMEV infection and antigen specific reactivation caused an increase in lineage −, Sca-1 +, ckit +(LSK) cells in the bone marrow. This LSK dysregulation was abrogated upon CD8 T cell depletion. We conclude that brain viral infections induce in situ effector and memory T cell responses within the bone marrow compartment. Both acute and memory recall CD8 T cell responses cause stem cell dysregulation. Our data hence paves the way for crucial studies of bone marrow resident antigen specific CD8 T cells in health and disease. Supported by grants from NIH (K99NS117799-01A1 (KA), R01NS103212 (AJJ), and RF1NS122174(AJJ)

Elevated exhausted effector tissue resident CD8+ T-cells in Chronic Graft versus Host Disease patient Oral mucosa unveiled by spatial transcriptomics and scRNA seq analysis

Abstract Chronic graft-versus-host disease (cGVHD), an autoimmune-like disease following allogeneic hematopoietic stem cell transplantation where T cells can attack host tissue. Oral mucosa (OM) is a most common cGVHD effector site in the oral cavity. We studied T cell populations and tissue-specific signals that regulate their differentiation and functionality. Patient OM biopsies from ongoing clinical trials (NCT03602599, NCT00331968) were taken at 6 months post-transplant. We found enrichment of immune cells in the OM mainly in the CD8 T-cell cluster and identified two unique populations in the OM of cGVHD patients: one characterized by expression of proliferative markers and the other more prominent, characterized by exhaustion makers (TIM3, LAG3, TIGIT). The latter had low expression of CD69 but expressed markers of tissue residency; CD103, BLIMP1, PD1, FABP5. Interestingly, this population showed high expression of BATF, IRF4, and RUNX3, transcription factors that induce sustained effector CD8 T-cell programming. These exhausted cells displayed higher expression of effector molecules IFNG, GZMB, PRF1 inducing sustained CD8 T-cell effector function. Ligand-receptor analysis revealed strong interactions between CXCL9-CXCR3 in CD8 T-cells and myeloid cells. Spatial transcriptomic analysis confirmed the CD8 T-cell infiltration in the submucosal region of OM in cGVHD colocalizing with the elevated markers of exhaustion, surrounded by myeloid cells, thus confirming the striking interaction of CXCL9 and CXCR3. These exhausted CD8 T-cells may play an important role in cGVHD pathogenesis causing by driving sustained tissue damage and could serve as a might be used as a novel target for cGVHD therapy. This work was supported by the intramural programs of the NIDCR and NCI.

Naïve T cell activation in the draining lymph nodes is accelerated by airway resident CD4 T cell activation

Abstract Tissue-resident memory CD4 T cells (TRM) protect against pathogens through the early shaping of inflammation at sites of infection. The extent to which their activation can impact subsequent systemic T cell responses is not clear. To ask if lung CD4 TRM recognizing antigen can impact systemic T cell responses against the same antigen presented in secondary lymphoid organs, we generated OVA-specific lung TRM primed by infection with an influenza virus expressing OVA peptide. We isolated the OVA-specific TRM 30 days after infection and transferred one million to naive mice intranasally resulting in an airway-resident TRM cohort similar in magnitude to TRM generated in virus-primed mice. We then recalled the transferred TRM by intranasal administration of FITC-labelled OVA protein to focus on TRM recall in the absence of virus-induced inflammation. One day after OVA challenge more FITC+ APC were present in the draining lymph nodes (dLN) of mice with TRM, suggesting antigen presentation to systemic T cells may be accelerated in the presence of airway TRM. We confirmed this by tracking naive OVA-specific CD4 T cells (OT-II) and CD8 T cell (OT-I) cohorts transferred i.v. to mice harboring OVA-specific TRM or not: more responding naive cells, expressing higher levels of activation markers, were detected 3 days after OVA challenge in mice with TRM. By 4 days, more responding cells were present in the lungs of TRM recipients, indicating that TRM and systemic T cell responses form a circuit via enhanced antigen presentation in the lymph nodes. Our results suggest that enhanced kinetics of systemic T cell activation induced by TRM activation in the airways may represent a novel mechanism of their protection against influenza and other respiratory viruses.

327 The early reactivation of resident CD8+ memory t cells depends on cross-presentation of antigen by langerhans cells in the epidermis

327 The early reactivation of resident CD8+ memory t cells depends on cross-presentation of antigen by langerhans cells in the epidermis

Abstract 4552: 3D-EXplore platform of fresh patient tumoroids with intact TME allows assessment of the efficacy of drugs targeting the tumor stroma on ex vivo tumor immunotherapy

Abstract Introduction: Cancer associated fibroblasts (CAFs) are a major component of the tumormicroenvironment (TME) and exhibit diverse tumorigenic functions such as immunosuppressionand extracellular matrix (ECM) remodeling. Treatment of solid tumors is often hindered by acomplex TME, which persists despite effective tumor-cell directed therapies. Therefore,treatment of solid tumors in combination with stromal-targeting therapies is essential toovercome CAF-facilitated tumor growth, and immunosuppression. Herein we interrogate theimpact of tumor stroma targeting therapeutics, in combination with nivolumab, on the efficacy oftumor cell killing (TCK) in a 3D human tumoroid ex vivo culture model (3D-EXplore). Resultingdata suggests that impairing the recruitment and activation of CAFs within the TME leads toenhanced TCK when combined with immune checkpoint blockade therapy. Materials and Methods: All patients tumor samples were collected with patient consent andrelevant IRB approval. 3D tumoroids measuring 150 microns in size were generated from freshpatient tumors including endometrial, ovarian, and colorectal cancer tissues. Tumoroids werethen treated with stromal targeting strategies for TGF-beta (galunisertib), the FGF pathway(Dovitinib), FAK inhibitors (defactinib), and cell adhesion modulators (plerixafor) alone or incombination with nivolumab for 72-hours ex vivo. Tumor responses to ex vivo treatments wereassessed using a proprietary tumor cell killing assay and 21-color flow cytometry analysis. Results and Summary: Here we evaluated the impact of stromal targeting therapeutics onCAFs and associated tumor cell killing as well as overcoming immune evasion mechanismspreserved in the tumoroid models. Furthermore, we used a multicolor flow panel to analyzewhether combination of stromal targeting drugs enhances nivolumab’s effect on the activation oftumor resident CD4, CD8 T-cells, NKT, and NK cell populations as well as on macrophagepolarization. Treatment-mediated changes in the tumor immune microenvironment was furthercorroborated by a multiplex cytokine release assay detecting GM-CSF, sCD137, IFNγ, sFas,sFasL, Granzyme A, Granzyme B, IL-2, IL-4, IL-5, IL-6, IL-10, IL-13, MIP-1α, MIP-1β, TNF-α,Perforin in tumoroid culture media and correlated with clinicopathologic findings and PD-L1expression for individual tumnors. Further, this 3D-tumoroid platform provides unique insightinto the microenvironment of both treatment responsive and non-responsive tumors and can aidin the development of patient-centered therapeutic regimens. Citation Format: Seth Currlin, Brittney Ruedlinger, Sharon Camacho, Angie Rivera, Jasmin D'Andrea, Jared Ehrhart, Soner Altiok. 3D-EXplore platform of fresh patient tumoroids with intact TME allows assessment of the efficacy of drugs targeting the tumor stroma on ex vivo tumor immunotherapy. [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 4552.

T-cell-B-cell collaboration in the lung.

Collaboration between T and B cells in secondary lymphoid organs is a crucial component of adaptive immunity, but lymphocytes also persist in other tissues. Recent studies have examined T-cell-B-cell interactions in nonlymphoid tissues such as the lung. CD4+ T- resident helper cells (TRH) remain in the lung after influenza infection and support both resident CD8 T cells and B cells. Multiple lung-resident B-cell subsets (B-resident memory (BRM)) that exhibit spatial and phenotypic diversity have also been described. Though not generated by all types of infection, inducible bronchus-associated lymphoid tissue offers a logical place for T and B cells to interact. Perturbations to BRM and TRH cells elicit effects specific to Immunoglobulin A (IgA) production, an antibody isotype with privileged access to mucosa. Understanding the interplay of lymphocytes in mucosal tissues, which can be insulated from systemic immune responses, may improve the design of future vaccines and therapies.

NDC80 status pinpoints mitotic kinase inhibitors as emerging therapeutic options in clear cell renal cell carcinoma

∼30% of clear cell renal cell carcinoma (ccRCC) patients present with metastatic disease at the time of diagnosis, causing a dire 5-year survival rate of 13%. Although anti-PD-1 immunotherapy has improved survival, a strong need remains for new therapeutic options. Using integrated network analysis, we identified the mitotic regulator NDC80 as a predictor of ccRCC progression. Overexpression of NDC80 fosters the malignant phenotype by promoting cell cycle progression through S phase as well as boosting glycolysis and mitochondrial respiration. Despite high levels of immune infiltration, particularly derived from tumor resident CD8+T cells with an exhausted phenotype, NDC80 defines a class of ccRCCs that poorly respond to immune checkpoint blockade. Instead, bioinformatics identified NDC80-high ccRCCs as sensitive to inhibitors of mitotic kinases, PLK1 and AURK, therapeutic approaches we validated in cell lines and mouse xenograft studies. Thus, NDC80 status pinpoints mitotic kinase inhibitors as promising therapeutic options in difficult-to-treat ccRCCs.

The aryl hydrocarbon receptor cell intrinsically promotes resident memory CD8+ T cell differentiation and function

The Aryl hydrocarbon receptor (Ahr) regulates the differentiation and function of CD4+ Tcells; however, its cell-intrinsic role in CD8+ Tcells remains elusive. Herein we show that Ahr acts as a promoter of resident memory CD8+ Tcell (TRM) differentiation and function. Genetic ablation of Ahr in mouse CD8+ Tcells leads to increased CD127-KLRG1+ short-lived effector cells and CD44+CD62L+ T central memory cells but reduced granzyme-B-producing CD69+CD103+ TRM cells. Genome-wide analyses reveal that Ahr suppresses the circulating while promoting the resident memory core gene program. A tumor resident polyfunctional CD8+ Tcell population, revealed by single-cell RNA-seq, is diminished upon Ahr deletion, compromising anti-tumor immunity. Human intestinal intraepithelial CD8+ Tcells also highly express AHR that regulates invitro TRM differentiation and granzyme B production. Collectively, these data suggest that Ahr is an important cell-intrinsic factor for CD8+ Tcell immunity.

Primary oral vaccination followed by a vaginal pull protects mice against genital HSV-2 infection.

HSV-2 infected more than 491 million people aged 15-49 world-wide in 2016. The morbidity associated with recurrent infections and the increased risk of HIV infection make this a major health problem. To date there is no effective vaccine. Because HSV-2 ascends to the dorsal route ganglion within 12-18 hours of infection, an effective vaccine will need to elicit a strong local resident CD8+ T cell response to prevent the infection from becoming life-long. Using a mouse model we investigated the potential of oral immunization with a novel lipid adjuvant (LiporaleTM) followed by local vaginal application of an inflammatory agents to protect against primary HSV-2 infections. Oral vaccination of mice with live-attenuated HSV-2 in LiporaleTM followed by vaginal application of DNFB or CXCL9/10 led to recruitment of tissue-resident CD8+ memory cells into the genital epithelia. This prime and pull vaccination strategy provided complete protection against wild-type HSV-2 challenge and prevented viral dissemination to the spinal cords. Activation of mucosal immunity by oral immunization, combined with induction of transient local genital inflammation can recruit long-lived tissue resident CD8+ T cells into the genital epithelium, providing significant protection against primary HSV-2 infection. This article is protected by copyright. All rights reserved.