Perinatal thymic-derived CD8αß-expressing γδ T cells are innate IFN-γ producers that expand in IL-7R-STAT5B-driven neoplasms.

The contribution of γδ T cells to immune responses is associated with rapid secretion of interferon-γ (IFN-γ). Here, we show a perinatal thymic wave of innate IFN-γ-producing γδ T cells that express CD8αß heterodimers and expand in preclinical models of infection and cancer. Optimal CD8αß+ γδ T cell development is directed by low T cell receptor signaling and through provision of interleukin (IL)-4 and IL-7. This population is pathologically relevant as overactive, or constitutive, IL-7R-STAT5B signaling promotes a supraphysiological accumulation of CD8αß+ γδ T cells in the thymus and peripheral lymphoid organs in two mouse models of T cell neoplasia. Likewise, CD8αß+ γδ T cells define a distinct subset of human T cell acute lymphoblastic leukemia pediatric patients. This work characterizes the normal and malignant development of CD8αß+ γδ T cells that are enriched in early life and contribute to innate IFN-γ responses to infection and cancer.

CD155/PVR determines acute myeloid leukemia targeting by Delta One T cells.

ABSTRACT: Relapsed or refractory acute myeloid leukemia (AML) remains a major therapeutic challenge. We have recently developed a Vδ1+ γδ T cell-based product for adoptive immunotherapy, named Delta One T (DOT) cells, and demonstrated their cytolytic capacity to eliminate AML cell lines and primary blasts in vitro and in vivo. However, the molecular mechanisms responsible for the broad DOT-cell recognition of AML cells remain poorly understood. Here, we dissected the role of natural killer (NK) cell receptor ligands in AML cell recognition by DOT cells. Screening of multiple AML cell lines highlighted a strong upregulation of the DNAM-1 ligands, CD155/pulmonary vascular resistance (PVR), CD112/nectin-2, as well as the NKp30 ligand, B7-H6, in contrast with NKG2D ligands. CRISPR-mediated ablation revealed key nonredundant and synergistic contributions of PVR and B7-H6 but not nectin-2 to DOT-cell targeting of AML cells. We further demonstrate that PVR and B7-H6 are critical for the formation of robust immunological synapses between AML and DOT cells. Importantly, PVR but not B7-H6 expression in primary AML samples predicted their elimination by DOT cells. These data provide new mechanistic insight into tumor targeting by DOT cells and suggest that assessing PVR expression levels may be highly relevant to DOT cell-based clinical trials.

Therapeutic avenues for γδ T cells in cancer.

γδ T cells are regarded as promising effector lymphocytes for next-generation cancer immunotherapies. In spite of being relatively rare in human peripheral blood, γδ T cells are more abundant in epithelial tissues where many tumors develop, and have been shown to actively participate in anticancer immunity as cytotoxic cells or as "type 1" immune orchestrators. A major asset of γδ T cells for tackling advanced cancers is their independence from antigen presentation via the major histocompatibility complex, which clearly sets them apart from conventional αß T cells. Here we discuss the main therapeutic strategies based on human γδ T cells. These include antibody-based bispecific engagers and adoptive cell therapies, either focused on the Vδ1+ or Vδ2+ γδ T-cell subsets, which can be expanded selectively and differentiated or engineered to maximize their antitumor functions. We review the preclinical data that supports each of the therapeutic strategies under development; and summarize the clinical trials being pursued towards establishing γδ T cell-based treatments for solid and hematological malignancies.

The emerging roles of γδ T cells in cancer immunotherapy.

Current cancer immunotherapies are primarily predicated on αß T cells, with a stringent dependence on MHC-mediated presentation of tumour-enriched peptides or unique neoantigens that can limit their efficacy and applicability in various contexts. After two decades of preclinical research and preliminary clinical studies involving very small numbers of patients, γδ T cells are now being explored as a viable and promising approach for cancer immunotherapy. The unique features of γδ T cells, including their tissue tropisms, antitumour activity that is independent of neoantigen burden and conventional MHC-dependent antigen presentation, and combination of typical properties of T cells and natural killer cells, make them very appealing effectors in multiple cancer settings. Herein, we review the main functions of γδ T cells in antitumour immunity, focusing on human γδ T cell subsets, with a particular emphasis on the differences between Vδ1+ and Vδ2+ γδ T cells, to discuss their prognostic value in patients with cancer and the key therapeutic strategies that are being developed in an attempt to improve the outcomes of these patients.

Generation and proof-of-concept for allogeneic CD123 CAR-Delta One T (DOT) cells in acute myeloid leukemia.

BACKGROUND: Chimeric antigen receptor (CAR)-T cells have emerged as a breakthrough treatment for relapse/refractory hematological tumors, showing impressive complete remission rates. However, around 50% of the patients relapse before 1-year post-treatment. T-cell 'fitness' is critical to prolong CAR-T persistence and activity. Allogeneic T cells from healthy donors are less dysfunctional or exhausted than autologous patient-derived T cells; in this context, Delta One T cells (DOTs), a recently described cellular product based on MHC/HLA-independent Vδ1+γδ T cells, represent a promising allogeneic platform. METHODS: Here we generated and preclinically validated, for the first time, 4-1BB-based CAR-DOTs directed against the interleukin-3α chain receptor (CD123), a target antigen widely expressed on acute myeloid leukemia (AML) blasts. RESULTS: CD123CAR-DOTs showed vigorous, superior to control DOTs, cytotoxicity against AML cell lines and primary samples both in vitro and in vivo, even on tumor rechallenge. CONCLUSIONS: Our results provide the proof-of-concept for a DOT-based next-generation allogeneic CAR-T therapy for AML.

Battle of the γδ T cell subsets in the gut.

In a study in Science, Reis et al. describe a temporal segregation of γδ T cell activities in colorectal cancer (CRC). Initially tumor surveillance is orchestrated by interferon-γ (IFN-γ)-producing and cytotoxic γδ T cell subsets, but once the tumor thrives, it becomes infiltrated by interleukin (IL)-17+ γδ T cell subsets that promote its growth.

MicroRNA-181a restricts human γδ T cell differentiation by targeting Map3k2 and Notch2.

γδ T cells are a conserved population of lymphocytes that contributes to anti-tumor responses through its overt type 1 inflammatory and cytotoxic properties. We have previously shown that human γδ T cells acquire this profile upon stimulation with IL-2 or IL-15, in a differentiation process dependent on MAPK/ERK signaling. Here, we identify microRNA-181a as a key modulator of human γδ T cell differentiation. We observe that miR-181a is highly expressed in patients with prostate cancer and that this pattern associates with lower expression of NKG2D, a critical mediator of cancer surveillance. Interestingly, miR-181a expression negatively correlates with an activated type 1 effector profile obtained from in vitro differentiated γδ T cells and miR-181a overexpression restricts their levels of NKG2D and TNF-α. Upon in silico analysis, we identify two miR-181a candidate targets, Map3k2 and Notch2, which we validate via overexpression coupled with luciferase assays. These results reveal a novel role for miR-181a as critical regulator of human γδ T cell differentiation and highlight its potential for manipulation of γδ T cells in next-generation immunotherapies.

Toward a better understanding of T cells in cancer.

T cells mediate anti-tumor immune responses and are the key target of immune checkpoint therapy, but they can also promote immune tolerance. A clear understanding of the specific contributions and biology of different T cell subsets is required to fully harness the curative potential of immunotherapies. Experts discuss the state of the field and key challenges for moving forward.

Brazilian Mobile Phone Applications Related to Hand Hygiene and Their Applicability for Healthcare Professionals.

The objective of this study was to identify available mobile applications regarding education for hand hygiene and their applicability as a resource for nurses and other healthcare professionals. The aim was to assess the quality of the mobile apps for education on hand hygiene for health professionals. A review of mobile apps available from Apple App Store and Google Play Store in Brazil was conducted. The World Health Organization recommendations and the Mobile Application Rating Scale for evaluating quality were used. Six applications were selected, only three presented gamification elements incorporated into the learning method and only two of them-SureWash Pocket and Give Me 5-used the international recommendations to improve hand hygiene compliance in a more substantial and interactive way. The mean quality total score for the five rated apps was 3.41, indicating poor to acceptable quality. SureWash Pocket was the only application that reached Mobile Application Rating Scale ≥4 in all dimensions. These mobile applications can be used as complementary alternatives in addition to other available education strategies to improve the standards of hand hygiene and change the behavior of health professionals.

IL-17 triggers the onset of cognitive and synaptic deficits in early stages of Alzheimer's disease.

Neuroinflammation in patients with Alzheimer's disease (AD) and related mouse models has been recognized for decades, but the contribution of the recently described meningeal immune population to AD pathogenesis remains to be addressed. Here, using the 3xTg-AD model, we report an accumulation of interleukin-17 (IL-17)-producing cells, mostly γδ T cells, in the brain and the meninges of female, but not male, mice, concomitant with the onset of cognitive decline. Critically, IL-17 neutralization into the ventricles is sufficient to prevent short-term memory and synaptic plasticity deficits at early stages of disease. These effects precede blood-brain barrier disruption and amyloid-beta or tau pathology, implying an early involvement of IL-17 in AD pathology. When IL-17 is neutralized at later stages of disease, the onset of short-memory deficits and amyloidosis-related splenomegaly is delayed. Altogether, our data support the idea that cognition relies on a finely regulated balance of "inflammatory" cytokines derived from the meningeal immune system.

Distinct metabolic programs established in the thymus control effector functions of γδ T cell subsets in tumor microenvironments.

Metabolic programming controls immune cell lineages and functions, but little is known about γδ T cell metabolism. Here, we found that γδ T cell subsets making either interferon-γ (IFN-γ) or interleukin (IL)-17 have intrinsically distinct metabolic requirements. Whereas IFN-γ+ γδ T cells were almost exclusively dependent on glycolysis, IL-17+ γδ T cells strongly engaged oxidative metabolism, with increased mitochondrial mass and activity. These distinct metabolic signatures were surprisingly imprinted early during thymic development and were stably maintained in the periphery and within tumors. Moreover, pro-tumoral IL-17+ γδ T cells selectively showed high lipid uptake and intracellular lipid storage and were expanded in obesity and in tumors of obese mice. Conversely, glucose supplementation enhanced the antitumor functions of IFN-γ+ γδ T cells and reduced tumor growth upon adoptive transfer. These findings have important implications for the differentiation of effector γδ T cells and their manipulation in cancer immunotherapy.

Functional and metabolic dichotomy of murine γδ T cell subsets in cancer immunity.

γδ T cells can display a plethora of immune functions, but recent studies have highlighted their importance, in multiple disease models, as sources of the pro-inflammatory cytokines, IL-17A (IL-17), and IFN-γ. These are produced by distinct murine effector γδ T cell subsets that diverge during thymic γδ T cell development. Among the multiple roles these subsets play in peripheral tissues, a striking dichotomy has emerged at tumor sites: whereas IFN-γ+ γδ T cells inhibit tumor cell growth, IL-17+ γδ T cells promote tumor progression and metastasis formation. In this review, we discuss the main lines of evidence, mostly from preclinical studies in mouse models, for this functional dichotomy in cancer immunity. We further highlight very recent advances in our understanding how metabolic sources and pathways can impact on the balance between IFN-γ+ and IL-17+ γδ T cells in the tumor microenvironment, which opens a new exciting avenue to explore toward the application of γδ T cells in cancer immunotherapy.

Antagonistic Inflammatory Phenotypes Dictate Tumor Fate and Response to Immune Checkpoint Blockade.

Inflammation can support or restrain cancer progression and the response to therapy. Here, we searched for primary regulators of cancer-inhibitory inflammation through deep profiling of inflammatory tumor microenvironments (TMEs) linked to immune-dependent control in mice. We found that early intratumoral accumulation of interferon gamma (IFN-γ)-producing natural killer (NK) cells induced a profound remodeling of the TME and unleashed cytotoxic T cell (CTL)-mediated tumor eradication. Mechanistically, tumor-derived prostaglandin E2 (PGE2) acted selectively on EP2 and EP4 receptors on NK cells, hampered the TME switch, and enabled immune evasion. Analysis of patient datasets across human cancers revealed distinct inflammatory TME phenotypes resembling those associated with cancer immune control versus escape in mice. This allowed us to generate a gene-expression signature that integrated opposing inflammatory factors and predicted patient survival and response to immune checkpoint blockade. Our findings identify features of the tumor inflammatory milieu associated with immune control of cancer and establish a strategy to predict immunotherapy outcomes.

MicroRNA-181a regulates IFN-γ expression in effector CD8+ T cell differentiation.

CD8+ T cells are key players in immunity against intracellular infections and tumors. The main cytokine associated with these protective responses is interferon-γ (IFN-γ), whose production is known to be regulated at the transcriptional level during CD8+ T cell differentiation. Here we found that microRNAs constitute a posttranscriptional brake to IFN-γ expression by CD8+ T cells, since the genetic interference with the Dicer processing machinery resulted in the overproduction of IFN-γ by both thymic and peripheral CD8+ T cells. Using a gene reporter mouse for IFN-γ locus activity, we compared the microRNA repertoires associated with the presence or absence of IFN-γ expression. This allowed us to identify a set of candidates, including miR-181a and miR-451, which were functionally tested in overexpression experiments using synthetic mimics in peripheral CD8+ T cell cultures. We found that miR-181a limits IFN-γ production by suppressing the expression of the transcription factor Id2, which in turn promotes the Ifng expression program. Importantly, upon MuHV-4 challenge, miR-181a-deficient mice showed a more vigorous IFN-γ+ CD8+ T cell response and were able to control viral infection significantly more efficiently than control mice. These data collectively establish a novel role for miR-181a in regulating IFN-γ-mediated effector CD8+ T cell responses in vitro and in vivo.

Translating gammadelta (γδ) T cells and their receptors into cancer cell therapies.

Clinical responses to checkpoint inhibitors used for cancer immunotherapy seemingly require the presence of αßT cells that recognize tumour neoantigens, and are therefore primarily restricted to tumours with high mutational load. Approaches that could address this limitation by engineering αßT cells, such as chimeric antigen receptor T (CAR T) cells, are being investigated intensively, but these approaches have other issues, such as a scarcity of appropriate targets for CAR T cells in solid tumours. Consequently, there is renewed interest among translational researchers and commercial partners in the therapeutic use of γδT cells and their receptors. Overall, γδT cells display potent cytotoxicity, which usually does not depend on tumour-associated (neo)antigens, towards a large array of haematological and solid tumours, while preserving normal tissues. However, the precise mechanisms of tumour-specific γδT cells, as well as the mechanisms for self-recognition, remain poorly understood. In this Review, we discuss the challenges and opportunities for the clinical implementation of cancer immunotherapies based on γδT cells and their receptors.

NKp46-expressing human gut-resident intraepithelial Vδ1 T cell subpopulation exhibits high antitumor activity against colorectal cancer.

γδ T cells account for a large fraction of human intestinal intraepithelial lymphocytes (IELs) endowed with potent antitumor activities. However, little is known about their origin, phenotype, and clinical relevance in colorectal cancer (CRC). To determine γδ IEL gut specificity, homing, and functions, γδ T cells were purified from human healthy blood, lymph nodes, liver, skin, and intestine, either disease-free, affected by CRC, or generated from thymic precursors. The constitutive expression of NKp46 specifically identifies a subset of cytotoxic Vδ1 T cells representing the largest fraction of gut-resident IELs. The ontogeny and gut-tropism of NKp46+/Vδ1 IELs depends both on distinctive features of Vδ1 thymic precursors and gut-environmental factors. Either the constitutive presence of NKp46 on tissue-resident Vδ1 intestinal IELs or its induced expression on IL-2/IL-15-activated Vδ1 thymocytes are associated with antitumor functions. Higher frequencies of NKp46+/Vδ1 IELs in tumor-free specimens from CRC patients correlate with a lower risk of developing metastatic III/IV disease stages. Additionally, our in vitro settings reproducing CRC tumor microenvironment inhibited the expansion of NKp46+/Vδ1 cells from activated thymic precursors. These results parallel the very low frequencies of NKp46+/Vδ1 IELs able to infiltrate CRC, thus providing insights to either follow-up cancer progression or to develop adoptive cellular therapies.

Lineage tracing of acute myeloid leukemia reveals the impact of hypomethylating agents on chemoresistance selection.

Chemotherapy-resistant cancer recurrence is a major cause of mortality. In acute myeloid leukemia (AML), chemorefractory relapses result from the complex interplay between altered genetic, epigenetic and transcriptional states in leukemic cells. Here, we develop an experimental model system using in vitro lineage tracing coupled with exome, transcriptome and in vivo functional readouts to assess the AML population dynamics and associated molecular determinants underpinning chemoresistance development. We find that combining standard chemotherapeutic regimens with low doses of DNA methyltransferase inhibitors (DNMTi, hypomethylating drugs) prevents chemoresistant relapses. Mechanistically, DNMTi suppresses the outgrowth of a pre-determined set of chemoresistant AML clones with stemness properties, instead favoring the expansion of rarer and unfit chemosensitive clones. Importantly, we confirm the capacity of DNMTi combination to suppress stemness-dependent chemoresistance development in xenotransplantation models and primary AML patient samples. Together, these results support the potential of DNMTi combination treatment to circumvent the development of chemorefractory AML relapses.