Repeated peripheral infusions of anti-EGFRvIII CAR T cells in combination with pembrolizumab show no efficacy in glioblastoma: a phase 1 trial.

We previously showed that chimeric antigen receptor (CAR) T-cell therapy targeting epidermal growth factor receptor variant III (EGFRvIII) produces upregulation of programmed death-ligand 1 (PD-L1) in the tumor microenvironment (TME). Here we conducted a phase 1 trial (NCT03726515) of CAR T-EGFRvIII cells administered concomitantly with the anti-PD1 (aPD1) monoclonal antibody pembrolizumab in patients with newly diagnosed, EGFRvIII+ glioblastoma (GBM) (n = 7). The primary outcome was safety, and no dose-limiting toxicity was observed. Secondary outcomes included median progression-free survival (5.2 months; 90% confidence interval (CI), 2.9-6.0 months) and median overall survival (11.8 months; 90% CI, 9.2-14.2 months). In exploratory analyses, comparison of the TME in tumors harvested before versus after CAR + aPD1 administration demonstrated substantial evolution of the infiltrating myeloid and T cells, with more exhausted, regulatory, and interferon (IFN)-stimulated T cells at relapse. Our study suggests that the combination of CAR T cells and PD-1 inhibition in GBM is safe and biologically active but, given the lack of efficacy, also indicates a need to consider alternative strategies.

Treg-targeted IL-2/anti-IL-2 complex controls graft-versus-host disease and supports anti-tumor effect in allogeneic hematopoietic stem cell transplantation.

Modulating an immune response in opposite directions represents the holy grail in allogeneic hematopoietic stem cell transplantation (allo-HSCT) to avoid insufficient reactivity of donor T cells and hematologic malignancy relapse while controlling the potential development of graft-versus-host disease (GVHD), in which donor T cells attack the recipient's tissues. IL-2/anti-IL-2 complexes (IL-2Cx) represent a therapeutic option to selectively accentuate or dampen the immune response. In dedicated experimental models of allo-HSCT, including also human cells injected in immunodeficient NSG mice, we evaluated side-by-side the therapeutic effect of two IL-2Cx designed either to boost regulatory T cells (Treg) or alternatively to activate effector T cells (Teff), on GVHD occurrence and tumor relapse. We also evaluated the effect of the complexes on the phenotype and function of immune cells in vivo. Unexpectedly, both pro-Treg and pro-Teff IL-2Cx prevented GVHD development. They both induced Treg expansion and reduced CD8+ T-cell numbers, compared to untreated mice. However, only mice treated with the pro-Treg IL-2Cx, showed a dramatic reduction of exhausted CD8+ T cells, consistent with a potent anti-tumor effect. When evaluated on human cells, pro-Treg IL-2Cx also preferentially induced Treg expansion in vitro and in vivo, while allowing the development of a potent anti-tumor effect in NSG mice. Our results demonstrate the clinical relevance of using a pro-Treg, but not a pro-Teff IL2Cx to modulate alloreactivity after HSCT, while promoting a graft-versus-leukemia effect.

Breast cancer remotely imposes a myeloid bias on haematopoietic stem cells by reprogramming the bone marrow niche.

Myeloid cell infiltration of solid tumours generally associates with poor patient prognosis and disease severity1-13. Therefore, understanding the regulation of myeloid cell differentiation during cancer is crucial to counteract their pro-tumourigenic role. Bone marrow (BM) haematopoiesis is a tightly regulated process for the production of all immune cells in accordance to tissue needs14. Myeloid cells differentiate during haematopoiesis from multipotent haematopoietic stem and progenitor cells (HSPCs)15-17. HSPCs can sense inflammatory signals from the periphery during infections18-21 or inflammatory disorders22-27. In these settings, HSPC expansion is associated with increased myeloid differentiation28,29. During carcinogenesis, the elevation of haematopoietic growth factors supports the expansion and differentiation of committed myeloid progenitors5,30. However, it is unclear whether cancer-related inflammation also triggers demand-adapted haematopoiesis at the level of multipotent HSPCs. In the BM, HSPCs reside within the haematopoietic niche which delivers HSC maintenance and differentiation cues31-35. Mesenchymal stem cells (MSCs) are a major cellular component of the BM niche and contribute to HSC homeostasis36-41. Modifications of MSCs in systemic disorders have been associated with HSC differentiation towards myeloid cells22,42. It is unknown if MSCs are regulated in the context of solid tumours and if their myeloid supportive activity is impacted by cancer-induced systemic changes. Here, using unbiased transcriptomic analysis and in situ imaging of HSCs and the BM niche during breast cancer, we show that both HSCs and MSCs are transcriptionally and spatially modified. We demonstrate that breast tumour can distantly remodel the cellular cross-talks in the BM niche leading to increased myelopoiesis.

Selective control of transposable element expression during T cell exhaustion and anti-PD-1 treatment.

In chronic infections and cancer, T cells are exposed to prolonged antigen stimulation, resulting in loss of function (or exhaustion) and impairment of effective immunological protection. Exhausted T cells are heterogeneous and include early progenitors (Tpex) and terminally exhausted cells (Tex). Here, we used bulk and single-cell transcriptomics to analyze expression of transposable elements (TEs) in subpopulations of mouse and human CD8+ tumor-infiltrating T lymphocytes (TILs). We show that in mice, members of the virus-like murine VL30 TE family (mostly intact, evolutionary young ERV1s) are strongly repressed in terminally exhausted CD8+ T cells in both tumor and viral models of exhaustion. Tpex expression of these VL30s, which are mainly intergenic and transcribed independently of their closest gene neighbors, was driven by Fli1, a transcription factor involved in progression from Tpex to Tex. Immune checkpoint blockade (ICB) in both mice and patients with cancer increased TE expression (including VL30 in mice), demonstrating that TEs may be applicable as ICB response biomarkers. We conclude that expression of TEs is tightly regulated in TILs during establishment of exhaustion and reprogramming by ICB. Analyses of TE expression on single cells and bulk populations open opportunities for understanding immune cell identity and heterogeneity, as well as for defining cellular gene expression signatures and disease biomarkers.

CD39+ conventional CD4+ T cells with exhaustion traits and cytotoxic potential infiltrate tumors and expand upon CTLA-4 blockade.

Conventional CD4+ T (Tconv) lymphocytes play important roles in tumor immunity; however, their contribution to tumor elimination remains poorly understood. Here, we describe a subset of tumor-infiltrating Tconv cells characterized by the expression of CD39. In several mouse cancer models, we observed that CD39+ Tconv cells accumulated in tumors but were absent in lymphoid organs. Compared to tumor CD39- counterparts, CD39+ Tconv cells exhibited a cytotoxic and exhausted signature at the transcriptomic level, confirmed by high protein expression of inhibitory receptors and transcription factors related to the exhaustion. Additionally, CD39+ Tconv cells showed increased production of IFNγ, granzyme B, perforin and CD107a expression, but reduced production of TNF. Around 55% of OVA-specific Tconv from B16-OVA tumor-bearing mice, expressed CD39. In vivo CTLA-4 blockade induced the expansion of tumor CD39+ Tconv cells, which maintained their cytotoxic and exhausted features. In breast cancer patients, CD39+ Tconv cells were found in tumors and in metastatic lymph nodes but were less frequent in adjacent non-tumoral mammary tissue and not detected in non-metastatic lymph nodes and blood. Human tumor CD39+ Tconv cells constituted a heterogeneous cell population with features of exhaustion, high expression of inhibitory receptors and CD107a. We found that high CD4 and ENTPD1 (CD39) gene expression in human tumor tissues correlated with a higher overall survival rate in breast cancer patients. Our results identify CD39 as a biomarker of Tconv cells, with characteristics of both exhaustion and cytotoxic potential, and indicate CD39+ Tconv cells as players within the immune response against tumors.

Imaging and multi-omics datasets converge to define different neural progenitor origins for ATRT-SHH subgroups.

Atypical teratoid rhabdoid tumors (ATRT) are divided into MYC, TYR and SHH subgroups, suggesting diverse lineages of origin. Here, we investigate the imaging of human ATRT at diagnosis and the precise anatomic origin of brain tumors in the Rosa26-CreERT2::Smarcb1flox/flox model. This cross-species analysis points to an extra-cerebral origin for MYC tumors. Additionally, we clearly distinguish SHH ATRT emerging from the cerebellar anterior lobe (CAL) from those emerging from the basal ganglia (BG) and intra-ventricular (IV) regions. Molecular characteristics point to the midbrain-hindbrain boundary as the origin of CAL SHH ATRT, and to the ganglionic eminence as the origin of BG/IV SHH ATRT. Single-cell RNA sequencing on SHH ATRT supports these hypotheses. Trajectory analyses suggest that SMARCB1 loss induces a de-differentiation process mediated by repressors of the neuronal program such as REST, ID and the NOTCH pathway.

Interleukin-17 signaling influences CD8+ T cell immunity and tumor progression according to the IL-17 receptor subunit expression pattern in cancer cells.

IL-17 immune responses in cancer are controversial, with both tumor-promoting and tumor-repressing effects observed. To clarify the role of IL-17 signaling in cancer progression, we used syngeneic tumor models from different tissue origins. We found that deficiencies in host IL-17RA or IL-17A/F expression had varying effects on the in vivo growth of different solid tumors including melanoma, sarcoma, lymphoma, and leukemia. In each tumor type, the absence of IL-17 led to changes in the expression of mediators associated with inflammation and metastasis in the tumor microenvironment. Furthermore, IL-17 signaling deficiencies in the hosts resulted in decreased anti-tumor CD8+ T cell immunity and caused tumor-specific changes in several lymphoid cell populations. Our findings were associated with distinct patterns of IL-17A/F cytokine and receptor subunit expression in the injected tumor cell lines. These patterns affected tumor cell responsiveness to IL-17 and downstream intracellular signaling, leading to divergent effects on cancer progression. Additionally, we identified IL-17RC as a critical determinant of the IL-17-mediated response in tumor cells and a potential biomarker for IL-17 signaling effects in tumor progression. Our study offers insight into the molecular mechanisms underlying IL-17 activities in cancer and lays the groundwork for developing personalized immunotherapies.

Phase Ib/II trial of tipapkinogene sovacivec, a therapeutic human papillomavirus16-vaccine, in combination with avelumab in patients with advanced human papillomavirus16-positive cancers.

PURPOSE: To evaluate tipapkinogene sovacivec (TG4001), a viral immunotherapeutic vaccine expressing human papillomavirus (HPV)16 E6/E7 non-oncogenic proteins and IL-2, in combination with avelumab in HPV16+ cancer patients. PATIENTS AND METHODS: In this open-label, phase Ib/II, multicenter study, HPV16+ advanced cancer patients received subcutaneous TG4001 at two dose levels (DL) in phase Ib and at the recommended phase II dose (RP2D) in phase II weekly for 6 weeks, then every 2 weeks (q2Wk) until 6 months, thereafter every 12 weeks, in combination with avelumab q2Wk starting from day 8. Exploratory end-points included immunomonitoring from sequential tumour and blood samples. RESULTS: Forty-three patients, mainly heavily pretreated (88% ≥ 1 previous line), were included in the safety analysis, with a majority of anal cancer (44%). No dose-limiting toxicities were reported, and DL2 (5 × 107 Plaque forming units (PFU)) was selected as the RP2D. Treatment-related adverse events to TG4001 occurred in 93% of patients, mostly grade 1/2, with grade 3 anaemia in one patient and no grade 4/5. Overall response rate (ORR) was 22% (8/36) and 32% (8/25) in all and patients without liver metastases, respectively. Median progression-free survival (PFS) and Overall Survival (OS) were 2.8 months (95% CI: 1.4-5.6) and 11.0 months (95% CI:7.5-16.7) in the total population and 5.6 months (95% CI:1.6-9.6) and 13.3 months (95% CI:8.7-32.7) in patients without liver metastases. Antigen-specific T-cell response was identified in 7/11 patients by IFNγ ELISpot. CONCLUSIONS: TG4001 in combination with avelumab is safe, demonstrated antitumour activity in heavily pre-treated HPV16+ cancer patients, and is currently being evaluated in a randomised phase II trial in patients with incurable anogenital cancer and limited hepatic involvement. GOV IDENTIFIER: NCT03260023.

Selective STING stimulation in dendritic cells primes antitumor T cell responses.

T cells that recognize tumor antigens are crucial for mounting antitumor immune responses. Induction of antitumor T cells in immunogenic tumors depends on STING, the intracellular innate immune receptor for cyclic guanosine monophosphate-adenosine monophosphate (cGAMP) and related cyclic dinucleotides (CDNs). However, the optimal way to leverage STING activation in nonimmunogenic tumors is still unclear. Here, we show that cGAMP delivery by intratumoral injection of virus-like particles (cGAMP-VLP) led to differentiation of circulating tumor-specific T cells, decreased tumor regulatory T cells (Tregs), and antitumoral responses that synergized with PD1 blockade. By contrast, intratumoral injection of the synthetic CDN ADU-S100 led to tumor necrosis and systemic T cell activation but simultaneously depleted immune cells from injected tumors and induced minimal priming of circulating tumor-specific T cells. The antitumor effects of cGAMP-VLP required type 1 conventional dendritic cells (cDC1), whereas ADU-S100 eliminated cDC1 from injected tumors. cGAMP-VLP preferentially targeted STING in dendritic cells at a 1000-fold smaller dose than ADU-S100. Subcutaneous administration of cGAMP-VLP showed synergy when combined with PD1 blockade or a tumor Treg-depleting antibody to elicit systemic tumor-specific T cells and antitumor activity, leading to complete and durable tumor eradication in the case of tumor Treg depletion. These findings show that cell targeting of STING stimulation shapes the antitumor T cell response and identify a therapeutic strategy to enhance T cell-targeted immunotherapy.

CD8+T cell responsiveness to anti-PD-1 is epigenetically regulated by Suv39h1 in melanomas.

Tumor-infiltrating CD8 + T cells progressively lose functionality and fail to reject tumors. The underlying mechanism and re-programing induced by checkpoint blockers are incompletely understood. We show here that genetic ablation or pharmacological inhibition of histone lysine methyltransferase Suv39h1 delays tumor growth and potentiates tumor rejection by anti-PD-1. In the absence of Suv39h1, anti-PD-1 induces alternative activation pathways allowing survival and differentiation of IFNγ and Granzyme B producing effector cells that express negative checkpoint molecules, but do not reach final exhaustion. Their transcriptional program correlates with that of melanoma patients responding to immune-checkpoint blockade and identifies the emergence of cytolytic-effector tumor-infiltrating lymphocytes as a biomarker of clinical response. Anti-PD-1 favors chromatin opening in loci linked to T-cell activation, memory and pluripotency, but in the absence of Suv39h1, cells acquire accessibility in cytolytic effector loci. Overall, Suv39h1 inhibition enhances anti-tumor immune responses, alone or combined with anti-PD-1, suggesting that Suv39h1 is an "epigenetic checkpoint" for tumor immunity.

Pseudocowpox virus, a novel vector to enhance the therapeutic efficacy of antitumor vaccination.

Objective: Antitumor viral vaccines, and more particularly poxviral vaccines, represent an active field for clinical development and translational research. To improve the efficacy and treatment outcome, new viral vectors are sought, with emphasis on their abilities to stimulate innate immunity, to display tumor antigens and to induce a specific T-cell response. Methods: We screened for a new poxviral backbone with improved innate and adaptive immune stimulation using IFN-α secretion levels in infected PBMC cultures as selection criteria. Assessment of virus effectiveness was made in vitro and in vivo. Results: The bovine pseudocowpox virus (PCPV) stood out among several poxviruses for its ability to induce significant secretion of IFN-α. PCPV produced efficient activation of human monocytes and dendritic cells, degranulation of NK cells and reversed MDSC-induced T-cell suppression, without being offensive to activated T cells. A PCPV-based vaccine, encoding the HPV16 E7 protein (PCPV-E7), stimulated strong antigen-specific T-cell responses in TC1 tumor-bearing mice. Complete regression of tumors was obtained in a CD8+ T-cell-dependent manner after intratumoral injection of PCPV-E7, followed by intravenous injection of the cancer vaccine MVA-E7. PCPV also proved active when injected repeatedly intratumorally in MC38 tumor-bearing mice, generating tumor-specific T-cell responses without encoding a specific MC38 antigen. From a translational perspective, we demonstrated that PCPV-E7 effectively stimulated IFN-γ production by T cells from tumor-draining lymph nodes of HPV+-infected cancer patients. Conclusion: We propose PCPV as a viral vector suitable for vaccination in the field of personalised cancer vaccines, in particular for heterologous prime-boost regimens.

Innate lymphoid cells: NK and cytotoxic ILC3 subsets infiltrate metastatic breast cancer lymph nodes.

Innate lymphoid cells (ILCs) - which include cytotoxic Natural Killer (NK) cells and helper-type ILC - are important regulators of tissue immune homeostasis, with possible roles in tumor surveillance. We analyzed ILC and their functionality in human lymph nodes (LN). In LN, NK cells and ILC3 were the prominent subpopulations. Among the ILC3s, we identified a CD56+/ILC3 subset with a phenotype close to ILC3 but also expressing cytotoxicity genes shared with NK. In tumor-draining LNs (TD-LNs) and tumor samples from breast cancer (BC) patients, NK cells were prominent, and proportions of ILC3 subsets were low. In tumors and TD-LN, NK cells display reduced levels of NCR (Natural cytotoxicity receptors), despite high transcript levels and included a small subset CD127- CD56- NK cells with reduced function. Activated by cytokines CD56+/ILC3 cells from donor and patients LN acquired cytotoxic capacity and produced IFNg. In TD-LN, all cytokine activated ILC populations produced TNFα in response to BC cell line. Analyses of cytotoxic and helper ILC indicate a switch toward NK cells in TD-LN. The local tumor microenvironment inhibited NK cell functions through downregulation of NCR, but cytokine stimulation restored their functionality.

Tissue-resident FOLR2+ macrophages associate with CD8+ T cell infiltration in human breast cancer.

Macrophage infiltration is a hallmark of solid cancers, and overall macrophage infiltration correlates with lower patient survival and resistance to therapy. Tumor-associated macrophages, however, are phenotypically and functionally heterogeneous. Specific subsets of tumor-associated macrophage might be endowed with distinct roles on cancer progression and antitumor immunity. Here, we identify a discrete population of FOLR2+ tissue-resident macrophages in healthy mammary gland and breast cancer primary tumors. FOLR2+ macrophages localize in perivascular areas in the tumor stroma, where they interact with CD8+ T cells. FOLR2+ macrophages efficiently prime effector CD8+ T cells ex vivo. The density of FOLR2+ macrophages in tumors positively correlates with better patient survival. This study highlights specific roles for tumor-associated macrophage subsets and paves the way for subset-targeted therapeutic interventions in macrophages-based cancer therapies.

Autoimmunity affecting the biliary tract fuels the immunosurveillance of cholangiocarcinoma.

Cholangiocarcinoma (CCA) results from the malignant transformation of cholangiocytes. Primary sclerosing cholangitis (PSC) and primary biliary cholangitis (PBC) are chronic diseases in which cholangiocytes are primarily damaged. Although PSC is an inflammatory condition predisposing to CCA, CCA is almost never found in the autoimmune context of PBC. Here, we hypothesized that PBC might favor CCA immunosurveillance. In preclinical murine models of cholangitis challenged with syngeneic CCA, PBC (but not PSC) reduced the frequency of CCA development and delayed tumor growth kinetics. This PBC-related effect appeared specific to CCA as it was not observed against other cancers, including hepatocellular carcinoma. The protective effect of PBC was relying on type 1 and type 2 T cell responses and, to a lesser extent, on B cells. Single-cell TCR/RNA sequencing revealed the existence of TCR clonotypes shared between the liver and CCA tumor of a PBC host. Altogether, these results evidence a mechanistic overlapping between autoimmunity and cancer immunosurveillance in the biliary tract.

Polyfunctional KLRG-1+CD57+ Senescent CD4+ T Cells Infiltrate Tumors and Are Expanded in Peripheral Blood From Breast Cancer Patients.

Senescent T cells have been described during aging, chronic infections, and cancer; however, a comprehensive study of the phenotype, function, and transcriptional program of this T cell population in breast cancer (BC) patients is missing. Compared to healthy donors (HDs), BC patients exhibit an accumulation of KLRG-1+CD57+ CD4+ and CD8+ T cells in peripheral blood. These T cells infiltrate tumors and tumor-draining lymph nodes. KLRG-1+CD57+ CD4+ and CD8+ T cells from BC patients and HDs exhibit features of senescence, and despite their inhibitory receptor expression, they produce more effector cytokines and exhibit higher expression of Perforin, Granzyme B, and CD107a than non-senescent subsets. When compared to blood counterparts, tumor-infiltrating senescent CD4+ T cells show similar surface phenotype but reduced cytokine production. Transcriptional profiling of senescent CD4+ T cells from the peripheral blood of BC patients reveals enrichment in genes associated with NK or CD8+-mediated cytotoxicity, TCR-mediated stimulation, and cell exhaustion compared to non-senescent T cells. Comparison of the transcriptional profile of senescent CD4+ T cells from peripheral blood of BC patients with those of HDs highlighted marked similarities but also relevant differences. Senescent CD4+ T cells from BC patients show enrichment in T-cell signaling, processes involved in DNA replication, p53 pathways, oncogene-induced senescence, among others compared to their counterparts in HDs. High gene expression of CD4, KLRG-1, and B3GAT1 (CD57), which correlates with increased overall survival for BC patients, underscores the usefulness of the evaluation of the frequency of senescent CD4+ T cells as a biomarker in the follow-up of patients.

Developmental bifurcation of human T follicular regulatory cells.

Germinal centers (GCs) are anatomic structures where B cells undergo affinity maturation, leading to production of high-affinity antibodies. The balance between T follicular helper (TFH) and regulatory (TFR) cells is critical for adequate control of GC responses. The study of human TFH and TFR cell development has been hampered because of the lack of in vitro assays reproducing in vivo biology, along with difficult access to healthy human lymphoid tissues. We used a single-cell transcriptomics approach to study the maturation of TFH and TFR cells isolated from human blood, iliac lymph nodes (LNs), and tonsils. As independent tissues have distinct proportions of follicular T cells in different maturation states, we leveraged the heterogeneity to reconstruct the maturation trajectory for human TFH and TFR cells. We found that the dominant maturation of TFR cells follows a bifurcated trajectory from precursor Treg cells, with one arm of the bifurcation leading to blood TFR cells and the other leading to the most mature GC TFR cells. Overall, our data provide a comprehensive resource for the transcriptomics of different follicular T cell populations and their dynamic relationship across different tissues.

Prognostic value of intratumoral Fusobacterium nucleatum and association with immune-related gene expression in oral squamous cell carcinoma patients.

Changes in the oral microbiome, particularly Fusobacterium nucleatum, are associated with oral squamous cell carcinoma (OSCC). F. nucleatum has been reported to modulate local immunity in cancers. We aimed to assess the association between intratumoral F. nucleatum and clinico-pathological features, relapse, and overall survival (OS) in two independent cohorts of patients with OSCC, and to explore the interplay with immune-related genes. We retrospectively analyzed tissue samples from a first cohort of 122 patients with head and neck squamous cell carcinoma, including 61 OSCC (cohort #1), and a second cohort of 90 additional OSCC (cohort #2). We then performed a sensitivity analysis on the merged cohort of OSCC patients (N = 151). F. nucleatum 16S rRNA gene sequences were quantified using real-time quantitative PCR. The presence of gram-negative bacteria and macrophages was confirmed by LPS and CD163 immunostainings, respectively. F. nucleatum positivity was associated with older age, less alcohol and combined alcohol plus tobacco consumption, and less frequent lymph node invasion. There was a trend for a lower recurrence rate in F. nucleatum-positive cases, with less metastatic relapses compared to F. nucleatum-negative tumors, and significantly longer OS, relapse-free and metastasis-free survival. F. nucleatum status was independently associated with OS in multivariate analysis. Immune-related gene and immunohistochemistry analyses showed that gram-negative bacteria load inversely correlated with M2 macrophages. F. nucleatum-associated OSCC has a specific immune microenvironment, is more frequent in older, non-drinking patients, and associated with a favorable prognosis.

Transcriptional and Functional Analysis of CD1c+ Human Dendritic Cells Identifies a CD163+ Subset Priming CD8+CD103+ T Cells.

Dendritic cells (DCs) are antigen-presenting cells controlling T cell activation. In humans, the diversity, ontogeny, and functional capabilities of DC subsets are not fully understood. Here, we identified circulating CD88-CD1c+CD163+ DCs (called DC3s) as immediate precursors of inflammatory CD88-CD14+CD1c+CD163+FcεRI+ DCs. DC3s develop via a specific pathway activated by GM-CSF, independent of cDC-restricted (CDP) and monocyte-restricted (cMoP) progenitors. Like classical DCs but unlike monocytes, DC3s drove activation of naive T cells. In vitro, DC3s displayed a distinctive ability to prime CD8+ T cells expressing a tissue homing signature and the epithelial homing alpha-E integrin (CD103) through transforming growth factor ß (TGF-ß) signaling. In vivo, DC3s infiltrated luminal breast cancer primary tumors, and DC3 infiltration correlated positively with CD8+CD103+CD69+ tissue-resident memory T cells. Together, these findings define DC3s as a lineage of inflammatory DCs endowed with a strong potential to regulate tumor immunity.