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Neutrophils are abundant immune cells in the circulation and frequently infiltrate tumors in substantial numbers. However, their precise functions in different cancer types remain incompletely understood, including in the brain microenvironment. We therefore investigated neutrophils in tumor tissue of glioma and brain metastasis patients, with matched peripheral blood, and herein describe the first in-depth analysis of neutrophil phenotypes and functions in these tissues. Orthogonal profiling strategies in humans and mice revealed that brain tumor-associated neutrophils (TANs) differ significantly from blood neutrophils and have a prolonged lifespan and immune-suppressive and pro-angiogenic capacity. TANs exhibit a distinct inflammatory signature, driven by a combination of soluble inflammatory mediators including tumor necrosis factor alpha (TNF-É) and Ceruloplasmin, which is more pronounced in TANs from brain metastasis versus glioma. Myeloid cells, including tumor-associated macrophages, emerge at the core of this network of pro-inflammatory mediators, supporting the concept of a critical myeloid niche regulating overall immune suppression in human brain tumors.
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Immunotherapies have shown remarkable, albeit tumor-selective, therapeutic benefits in the clinic. Most patients respond transiently at best, highlighting the importance of understanding mechanisms underlying resistance. Herein, we evaluated the effects of the engineered immunocytokine PD1-IL2v in a mouse model of de novo pancreatic neuroendocrine cancer that is resistant to checkpoint and other immunotherapies. PD1-IL2v utilizes anti-PD-1 as a targeting moiety fused to an immuno-stimulatory IL-2 cytokine variant (IL2v) to precisely deliver IL2v to PD-1+ T cells in the tumor microenvironment. PD1-IL2v elicited substantial infiltration by stem-like CD8+ T cells, resulting in tumor regression and enhanced survival in mice. Combining anti-PD-L1 with PD1-IL2v sustained the response phase, improving therapeutic efficacy both by reprogramming immunosuppressive tumor-associated macrophages and enhancing T cell receptor (TCR) immune repertoire diversity. These data provide a rationale for clinical trials to evaluate the combination therapy of PD1-IL2v and anti-PD-L1, particularly in immunotherapy-resistant tumors infiltrated with PD-1+ stem-like T cells.
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Linfocitos T CD8-positivos , Inmunoterapia , Macrófagos , Neoplasias , Animales , Ratones , Antígeno B7-H1/inmunología , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Modelos Animales de Enfermedad , Inmunoterapia/métodos , Macrófagos/inmunología , Macrófagos/metabolismo , Neoplasias/terapia , Microambiente Tumoral , Anticuerpos Biespecíficos/inmunología , Interleucina-2 , Receptor de Muerte Celular Programada 1/inmunologíaRESUMEN
The process of cancer immunosurveillance is a mechanism of tumour suppression that can protect the host from cancer development throughout its lifetime1,2. However, it is unknown whether the effectiveness of cancer immunosurveillance fluctuates over a single day. Here we demonstrate that the initial time of day of tumour engraftment dictates the ensuing tumour size across mouse cancer models. Using immunodeficient mice as well as mice lacking lineage-specific circadian functions, we show that dendritic cells (DCs) and CD8+ T cells exert circadian anti-tumour functions that control melanoma volume. Specifically, we find that rhythmic trafficking of DCs to the tumour draining lymph node governs a circadian response of tumour-antigen-specific CD8+ T cells that is dependent on the circadian expression of the co-stimulatory molecule CD80. As a consequence, cancer immunotherapy is more effective when synchronized with DC functions, shows circadian outcomes in mice and suggests similar effects in humans. These data demonstrate that the circadian rhythms of anti-tumour immune components are not only critical for controlling tumour size but can also be of therapeutic relevance.
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Linfocitos T CD8-positivos , Ritmo Circadiano , Células Dendríticas , Melanoma , Animales , Humanos , Ratones , Linfocitos T CD8-positivos/inmunología , Células Dendríticas/inmunología , Inmunoterapia/métodos , Melanoma/inmunología , Melanoma/patología , Melanoma/terapia , Ratones Endogámicos C57BL , Antígeno B7-1 , Antígenos de Neoplasias/inmunología , Ganglios Linfáticos , Ritmo Circadiano/inmunologíaRESUMEN
NOTCH1 is a well-established lineage specifier for T cells and among the most frequently mutated genes throughout all subclasses of T cell acute lymphoblastic leukemia (T-ALL). How oncogenic NOTCH1 signaling launches a leukemia-prone chromatin landscape during T-ALL initiation is unknown. Here we demonstrate an essential role for the high-mobility-group transcription factor Tcf1 in orchestrating chromatin accessibility and topology, allowing aberrant Notch1 signaling to convey its oncogenic function. Although essential, Tcf1 is not sufficient to initiate leukemia. The formation of a leukemia-prone epigenetic landscape at the distal Notch1-regulated Myc enhancer, which is fundamental to this disease, is Tcf1-dependent and occurs within the earliest progenitor stage even before cells adopt a T lymphocyte or leukemic fate. Moreover, we discovered a unique evolutionarily conserved Tcf1-regulated enhancer element in the distal Myc-enhancer, which is important for the transition of preleukemic cells to full-blown disease.
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Leucemia-Linfoma Linfoblástico de Células T Precursoras , Carcinogénesis/genética , Línea Celular Tumoral , Cromatina/genética , Humanos , Oncogenes , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Receptor Notch1/genéticaRESUMEN
BACKGROUND: Whereas the dietary intake of industrial trans fatty acids (iTFA) has been specifically associated with inflammation, cardiovascular disease, and type 2 diabetes, understanding the impact of dietary fats on human health remains challenging owing to their complex composition and individual effects of their lipid components on metabolism. The aim of this study is to profile the composition of blood, measured by the fatty acid (FAs) profile and untargeted metabolome of serum and the transcriptome of blood cells, in order to identify molecular signatures that discriminate dietary fat intakes. METHODS: In a parallel study, the molecular effects of consuming dairy fat containing ruminant TFA (rTFA) or margarine containing iTFA were investigated. Healthy volunteers (n = 42; 45-69 y) were randomly assigned to diets containing margarine without TFA as major source of fat (wTFA control group with 0.4 g TFA per 100 g margarine), margarine with iTFA (iTFA group with 4.1 g TFA per 100 g margarine), or butter with rTFA (rTFA group with 6.3 g TFA per 100 g butter) for 4 weeks. The amounts of test products were individually selected so that fat intake contributed to 30-33% of energy requirements and TFA in the rTFA and iTFA groups contributed to up to 2% of energy intake. Changes in fasting blood values of lipid profiles (GC with flame-ionization detection), metabolome profiles (LC-MS, GC-MS), and gene expression (microarray) were measured. RESULTS: Eighteen FAs, as well as 242 additional features measured by LC-MS (185) and GC-MS (54) showed significantly different responses to the diets (PFDR-adjusted < 0.05), mainly distinguishing butter from the margarine diets while gene expression was not differentially affected. The most abundant TFA in the butter, i.e. TFA containing (E)-octadec-11-enoic acid (C18:1 t11; trans vaccenic acid), and margarines, i.e. TFA containing (E)-octadec-9-enoic acid (C18:1 t9; elaidic acid) were reflected in the significantly different serum levels of TFAs measured after the dietary interventions. CONCLUSIONS: The untargeted serum metabolome differentiates margarine from butter intake although the identification of the discriminating features remains a bottleneck. The targeted serum FA profile provides detailed information on specific molecules differentiating not only butter from margarine intake but also diets with different content of iTFAs in margarine. TRIAL REGISTRATION: ClinicalTrials.gov NCT00933322.
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Diabetes Mellitus Tipo 2 , Ácidos Grasos trans , Mantequilla , Grasas de la Dieta/farmacología , Humanos , MargarinaRESUMEN
Elevated peripheral blood and tumor-infiltrating neutrophils are often associated with a poor patient prognosis. However, therapeutic strategies to target these cells are difficult to implement due to the life-threatening risk of neutropenia. In a genetically engineered mouse model of lung adenocarcinoma, tumor-associated neutrophils (TAN) demonstrate tumor-supportive capacities and have a prolonged lifespan compared to circulating neutrophils. Here, we show that tumor cell-derived GM-CSF triggers the expression of the anti-apoptotic Bcl-xL protein and enhances neutrophil survival through JAK/STAT signaling. Targeting Bcl-xL activity with a specific BH3 mimetic, A-1331852, blocked the induced neutrophil survival without impacting their normal lifespan. Specifically, oral administration with A-1331852 decreased TAN survival and abundance, and reduced tumor growth without causing neutropenia. We also show that G-CSF, a drug used to combat neutropenia in patients receiving chemotherapy, increased the proportion of young TANs and augmented the anti-tumor effect resulting from Bcl-xL blockade. Finally, our human tumor data indicate the same role for Bcl-xL on pro-tumoral neutrophil survival. These results altogether provide preclinical evidence for safe neutrophil targeting based on their aberrant intra-tumor longevity.
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Neoplasias Pulmonares , Neutropenia , Animales , Humanos , Ratones , Envejecimiento , Apoptosis , Proteínas Reguladoras de la Apoptosis/metabolismo , Proteína bcl-X , Línea Celular Tumoral , Neoplasias Pulmonares/patología , Neutropenia/tratamiento farmacológico , Neutropenia/metabolismo , Neutropenia/patología , Neutrófilos/metabolismoRESUMEN
Cancer cell plasticity contributes to therapy resistance and metastasis, which represent the main causes of cancer-related death, including in breast cancer. The tumor microenvironment drives cancer cell plasticity and metastasis, and unraveling the underlying cues may provide novel strategies for managing metastatic disease. Using breast cancer experimental models and transcriptomic analyses, we show that stem cell antigen-1 positive (SCA1+) murine breast cancer cells enriched during tumor progression and metastasis had higher in vitro cancer stem cell-like properties, enhanced in vivo metastatic ability, and generated tumors rich in Gr1hiLy6G+CD11b+ cells. In turn, tumor-educated Gr1+CD11b+ (Tu-Gr1+CD11b+) cells rapidly and transiently converted low metastatic SCA1- cells into highly metastatic SCA1+ cells via secreted oncostatin M (OSM) and IL-6. JAK inhibition prevented OSM/IL-6-induced SCA1+ population enrichment, while OSM/IL-6 depletion suppressed Tu-Gr1+CD11b+-induced SCA1+ population enrichment in vitro and metastasis in vivo. Moreover, chemotherapy-selected highly metastatic 4T1 cells maintained high SCA1+ positivity through autocrine IL-6 production, and in vitro JAK inhibition blunted SCA1 positivity and metastatic capacity. Importantly, Tu-Gr1+CD11b+ cells invoked a gene signature in tumor cells predicting shorter overall survival (OS), relapse-free survival (RFS), and lung metastasis in breast cancer patients. Collectively, our data identified OSM/IL-6/JAK as a clinically relevant paracrine/autocrine axis instigating breast cancer cell plasticity and triggering metastasis.
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Neoplasias de la Mama , Neoplasias Pulmonares , Neoplasias Primarias Secundarias , Ataxias Espinocerebelosas , Humanos , Ratones , Animales , Femenino , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Interleucina-6/genética , Oncostatina M , Plasticidad de la Célula , Línea Celular Tumoral , Recurrencia Local de Neoplasia , Neoplasias Pulmonares/patología , Metástasis de la Neoplasia , Microambiente TumoralRESUMEN
Brain metastasis (BrM) is a common malignancy, predominantly originating from lung, melanoma, and breast cancers. The vasculature is a key component of the BrM tumor microenvironment with critical roles in regulating metastatic seeding and progression. However, the heterogeneity of the major BrM vascular components, namely endothelial and mural cells, is still poorly understood. We perform single-cell and bulk RNA-sequencing of sorted vascular cell types and detect multiple subtypes enriched specifically in BrM compared to non-tumor brain, including previously unrecognized immune regulatory subtypes. We integrate the human data with mouse models, creating a platform to interrogate vascular targets for the treatment of BrM. We find that the CD276 immune checkpoint molecule is significantly upregulated in the BrM vasculature, and anti-CD276 blocking antibodies prolonged survival in preclinical trials. This study provides important insights into the complex interactions between the vasculature, immune cells, and cancer cells, with translational relevance for designing therapeutic interventions.
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Neoplasias Encefálicas , Neoplasias de la Mama , Melanoma , Ratones , Animales , Humanos , Femenino , Neoplasias Encefálicas/patología , Encéfalo/metabolismo , Neoplasias de la Mama/patología , Factores de Transcripción/metabolismo , Microambiente Tumoral , Antígenos B7RESUMEN
Dendritic cells (DCs) are antigen-presenting myeloid cells that regulate T cell activation, trafficking and function. Monocyte-derived DCs pulsed with tumor antigens have been tested extensively for therapeutic vaccination in cancer, with mixed clinical results. Here, we present a cell-therapy platform based on mouse or human DC progenitors (DCPs) engineered to produce two immunostimulatory cytokines, IL-12 and FLT3L. Cytokine-armed DCPs differentiated into conventional type-I DCs (cDC1) and suppressed tumor growth, including melanoma and autochthonous liver models, without the need for antigen loading or myeloablative host conditioning. Tumor response involved synergy between IL-12 and FLT3L and was associated with natural killer and T cell infiltration and activation, M1-like macrophage programming and ischemic tumor necrosis. Antitumor immunity was dependent on endogenous cDC1 expansion and interferon-γ signaling but did not require CD8+ T cell cytotoxicity. Cytokine-armed DCPs synergized effectively with anti-GD2 chimeric-antigen receptor (CAR) T cells in eradicating intracranial gliomas in mice, illustrating their potential in combination therapies.
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Citocinas , Neoplasias , Humanos , Ratones , Animales , Inmunoterapia , Células Dendríticas , Neoplasias/terapia , Interleucina-12RESUMEN
Gain-of-function mutations in NOTCH1 are among the most frequent genetic alterations in T-cell acute lymphoblastic leukemia (T-ALL), highlighting the Notch signaling pathway as a promising therapeutic target for personalized medicine. Yet, a major limitation for long-term success of targeted therapy is relapse due to tumor heterogeneity or acquired resistance. Thus, we performed a genome-wide CRISPR-Cas9 screen to identify prospective resistance mechanisms to pharmacological NOTCH inhibitors and novel targeted combination therapies to efficiently combat T-ALL. Mutational loss of phosphoinositide-3-kinase regulatory subunit 1 (PIK3R1) causes resistance to Notch inhibition. PIK3R1 deficiency leads to increased PI3K/AKT signaling, which regulates cell cycle and the spliceosome machinery, both at the transcriptional and posttranslational level. Moreover, several therapeutic combinations have been identified, in which simultaneous targeting of the cyclin-dependent kinases 4 and 6 (CDK4/6) and NOTCH proved to be the most efficacious in T-ALL xenotransplantation models.
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Leucemia-Linfoma Linfoblástico de Células T Precursoras , Humanos , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Receptor Notch1/genética , Receptor Notch1/metabolismo , Estudios Prospectivos , Linfocitos T/metabolismoRESUMEN
MAGI1 acts as a tumor suppressor in estrogen receptor-positive (ER+) breast cancer (BC), and its loss correlates with a more aggressive phenotype. To identify the pathways and events affected by MAGI1 loss, we deleted the MAGI1 gene in the ER+ MCF7 BC cell line and performed RNA sequencing and functional experiments in vitro. Transcriptome analyses revealed gene sets and biological processes related to estrogen signaling, the cell cycle, and DNA damage responses affected by MAGI1 loss. Upon exposure to TNF-α/IFN-γ, MCF7 MAGI1 KO cells entered a deeper level of quiescence/senescence compared with MCF7 control cells and activated the AKT and MAPK signaling pathways. MCF7 MAGI1 KO cells exposed to ionizing radiations or cisplatin had reduced expression of DNA repair proteins and showed increased sensitivity towards PARP1 inhibition using olaparib. Treatment with PI3K and AKT inhibitors (alpelisib and MK-2206) restored the expression of DNA repair proteins and sensitized cells to fulvestrant. An analysis of human BC patients' transcriptomic data revealed that patients with low MAGI1 levels had a higher tumor mutational burden and homologous recombination deficiency. Moreover, MAGI1 expression levels negatively correlated with PI3K/AKT and MAPK signaling, which confirmed our in vitro observations. Pharmacological and genomic evidence indicate HDACs as regulators of MAGI1 expression. Our findings provide a new view on MAGI1 function in cancer and identify potential treatment options to improve the management of ER+ BC patients with low MAGI1 levels.
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Neoplasias de la Mama , Guanilato-Quinasas , Femenino , Humanos , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Neoplasias de la Mama/patología , Moléculas de Adhesión Celular/metabolismo , Línea Celular Tumoral , Daño del ADN , Guanilato-Quinasas/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismoRESUMEN
The transforming growth factor-ß (TGF-ß) family member activin A (hereafter Activin-A) is overexpressed in many cancer types, often correlating with cancer-associated cachexia and poor prognosis. Activin-A secretion by melanoma cells indirectly impedes CD8+ T cell-mediated anti-tumor immunity and promotes resistance to immunotherapies, even though Activin-A can be proinflammatory in other contexts. To identify underlying mechanisms, we here analyzed the effect of Activin-A on syngeneic grafts of Braf mutant YUMM3.3 mouse melanoma cells and on their microenvironment using single-cell RNA sequencing. We found that the Activin-A-induced immune evasion was accompanied by a proinflammatory interferon signature across multiple cell types, and that the associated increase in tumor growth depended at least in part on pernicious STING activity within the melanoma cells. Besides corroborating a role for proinflammatory signals in facilitating immune evasion, our results suggest that STING holds considerable potential as a therapeutic target to mitigate tumor-promoting Activin-A signaling at least in melanoma.
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Activinas , Melanoma , Factor de Crecimiento Transformador beta , Escape del Tumor , Animales , Ratones , Activinas/metabolismo , Melanoma/inmunología , Transducción de Señal , Factor de Crecimiento Transformador beta/metabolismo , Microambiente TumoralRESUMEN
Interleukin (IL)-26 is a TH17 cytokine with known antimicrobial and pro-inflammatory functions. However, the precise role of IL-26 in the context of pathogenic TH17 responses is unknown. Here we identify a population of blood TH17 intermediates that produce high levels of IL-26 and differentiate into IL-17A-producing TH17 cells upon TGF-ß1 exposure. By combining single cell RNA sequencing, TCR sequencing and spatial transcriptomics we show that this process occurs in psoriatic skin. In fact, IL-26+ TH17 intermediates infiltrating psoriatic skin induce TGF-ß1 expression in basal keratinocytes and thereby promote their own differentiation into IL-17A-producing cells. Thus, our study identifies IL-26-producing cells as an early differentiation stage of TH17 cells that infiltrates psoriatic skin and controls its own maturation into IL17A-producing TH17 cells, via epithelial crosstalk involving paracrine production of TGF-ß1.
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Psoriasis , Factor de Crecimiento Transformador beta1 , Humanos , Interleucina-17/genética , Diferenciación Celular , PielRESUMEN
The immune-specialized environment of the healthy brain is tightly regulated to prevent excessive neuroinflammation. However, after cancer development, a tissue-specific conflict between brain-preserving immune suppression and tumor-directed immune activation may ensue. To interrogate potential roles of T cells in this process, we profiled these cells from individuals with primary or metastatic brain cancers via integrated analyses on the single-cell and bulk population levels. Our analysis revealed similarities and differences in T cell biology between individuals, with the most pronounced differences observed in a subgroup of individuals with brain metastasis, characterized by accumulation of CXCL13-expressing CD39+ potentially tumor-reactive T (pTRT) cells. In this subgroup, high pTRT cell abundance was comparable to that in primary lung cancer, whereas all other brain tumors had low levels, similar to primary breast cancer. These findings indicate that T cell-mediated tumor reactivity can occur in certain brain metastases and may inform stratification for treatment with immunotherapy.
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Neoplasias Encefálicas , Linfocitos T , Humanos , Multiómica , Neoplasias Encefálicas/secundario , Encéfalo , InmunoterapiaRESUMEN
High-grade gliomas, the most common and aggressive primary brain tumors, are characterized by a complex tumor microenvironment (TME). Among the immune cells infiltrating the glioma TME, tumor-associated microglia and macrophages (TAMs) constitute the major compartment. In patients with gliomas, increased TAM abundance is associated with more aggressive disease. Alterations in TAM phenotypes and functions have been reported in preclinical models of multiple cancers during tumor development and after therapeutic interventions, including radiotherapy and molecular targeted therapies. These findings indicate that it is crucial to evaluate TAM abundance and dynamics over time. Current techniques to quantify TAMs in patients rely mainly on histological staining of tumor biopsies. Although informative, these techniques require an invasive procedure to harvest the tissue sample and typically only result in a snapshot of a small region at a single point in time. Fluorine isotope 19 MRI (19F MRI) represents a powerful means to noninvasively and longitudinally monitor myeloid cells in pathological conditions by intravenously injecting perfluorocarbon-containing nanoparticles (PFC-NP). In this study, we demonstrated the feasibility and power of 19F MRI in preclinical models of gliomagenesis, breast-to-brain metastasis, and breast cancer and showed that the major cellular source of 19F signal consists of TAMs. Moreover, multispectral 19F MRI with two different PFC-NP allowed us to identify spatially and temporally distinct TAM niches in radiotherapy-recurrent murine gliomas. Together, we have imaged TAMs noninvasively and longitudinally with integrated cellular, spatial, and temporal resolution, thus revealing important biological insights into the critical functions of TAMs, including in disease recurrence.
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Fluorocarburos , Glioma , Miopatías Estructurales Congénitas , Animales , Ratones , Macrófagos Asociados a Tumores , Flúor , Recurrencia Local de Neoplasia , Tamoxifeno , Glioma/diagnóstico por imagen , Microambiente Tumoral , Melanoma Cutáneo MalignoRESUMEN
Many human cancers manifest the capability to circumvent attack by the adaptive immune system. In this work, we identified a component of immune evasion that involves frequent up-regulation of fragile X mental retardation protein (FMRP) in solid tumors. FMRP represses immune attack, as revealed by cancer cells engineered to lack its expression. FMRP-deficient tumors were infiltrated by activated T cells that impaired tumor growth and enhanced survival in mice. Mechanistically, FMRP's immunosuppression was multifactorial, involving repression of the chemoattractant C-C motif chemokine ligand 7 (CCL7) concomitant with up-regulation of three immunomodulators-interleukin-33 (IL-33), tumor-secreted protein S (PROS1), and extracellular vesicles. Gene signatures associate FMRP's cancer network with poor prognosis and response to therapy in cancer patients. Collectively, FMRP is implicated as a regulator that orchestrates a multifaceted barrier to antitumor immune responses.
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Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil , Evasión Inmune , Tolerancia Inmunológica , Neoplasias , Animales , Humanos , Ratones , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/genética , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil/metabolismo , Neoplasias/inmunología , Quimiocina CCL7/metabolismo , Interleucina-33 , Proteína S/metabolismoRESUMEN
Stem and progenitor cells residing in the intestinal crypts drive the majority of colorectal cancers (CRCs), yet vascular contribution to this niche remains largely unexplored. VEGFA is a key driver of physiological and tumor angiogenesis. Accordingly, current anti-angiogenic cancer therapies target the VEGFA pathway. Here we report that in CRC expansion of the stem/progenitor pool in intestinal crypts requires VEGFA-independent growth and remodeling of blood vessels. Epithelial transformation induced expression of the endothelial peptide apelin, directs migration of distant venous endothelial cells towards progenitor niche vessels ensuring optimal perfusion. In the absence of apelin, loss of injury-inducible PROX1+ epithelial progenitors inhibited both incipient and advanced intestinal tumor growth. Our results establish fundamental principles for the reciprocal communication between vasculature and the intestinal progenitor niche and provide a mechanism for resistance to VEGFA-targeting drugs in CRCs.