RESUMO
Despite significant advances in the development and refinement of immunotherapies administered to combat cancer over the past decades, a number of barriers continue to limit their efficacy. One significant clinical barrier is the inability to mount initial immune responses towards the tumor. As dendritic cells are central initiators of immune responses in the body, the elucidation of mechanisms that can be therapeutically leveraged to enhance their functions to drive anti-tumor immune responses is urgently needed. Here, we report that the dietary sugar L-fucose can be used to enhance the immunostimulatory activity of dendritic cells (DCs). L-fucose polarizes immature myeloid cells towards specific DC subsets, specifically cDC1 and moDC subsets. In vitro, L-fucose treatment enhances antigen uptake and processing of DCs. Furthermore, our data suggests that L-fucose-treated DCs increase stimulation of T cell populations. Consistent with our functional assays, single-cell RNA sequencing of intratumoral DCs from melanoma- and breast tumor-bearing mice confirmed transcriptional regulation and antigen processing as pathways that are significantly altered by dietary L-fucose. Together, this study provides the first evidence of the ability of L-fucose to bolster DC functionality and provides rational to further investigate how L-fucose can be used to leverage DC function in order to enhance current immunotherapy.
Assuntos
Células Dendríticas , Fucose , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Animais , Camundongos , Fucose/metabolismo , Apresentação de Antígeno , Feminino , Camundongos Endogâmicos C57BL , Polaridade Celular , Linhagem Celular Tumoral , Linfócitos T/imunologia , Linfócitos T/metabolismo , Melanoma Experimental/imunologia , Ativação Linfocitária/imunologiaRESUMO
Melanoma incidence and mortality rates are historically higher for men than women. Although emerging studies have highlighted tumorigenic roles for the male sex hormone androgen and its receptor (AR) in melanoma, cellular and molecular mechanisms underlying these sex-associated discrepancies are poorly defined. Here, we delineate a previously undisclosed mechanism by which androgen-activated AR transcriptionally upregulates fucosyltransferase 4 (FUT4) expression, which drives melanoma invasiveness by interfering with adherens junctions (AJs). Global phosphoproteomic and fucoproteomic profiling, coupled with in vitro and in vivo functional validation, further reveal that AR-induced FUT4 fucosylates L1 cell adhesion molecule (L1CAM), which is required for FUT4-increased metastatic capacity. Tumor microarray and gene expression analyses demonstrate that AR-FUT4-L1CAM-AJs signaling correlates with pathological staging in melanoma patients. By delineating key androgen-triggered signaling that enhances metastatic aggressiveness, our findings help explain sex-associated clinical outcome disparities and highlight AR/FUT4 and its effectors as potential prognostic biomarkers and therapeutic targets in melanoma.
Assuntos
Melanoma , Molécula L1 de Adesão de Célula Nervosa , Humanos , Masculino , Feminino , Melanoma/metabolismo , Androgênios , Molécula L1 de Adesão de Célula Nervosa/metabolismo , Antígenos CD15/metabolismo , Glicosilação , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Linhagem Celular Tumoral , Fucosiltransferases/genética , Fucosiltransferases/metabolismoRESUMO
The atypical cadherins Fat and Dachsous are key regulators of cell growth and animal development. In contrast to classical cadherins, which form homophilic interactions to segregate cells, Fat and Dachsous cadherins form heterophilic interactions to induce cell polarity within tissues. Here, we determine the co-crystal structure of the human homologs Fat4 and Dachsous1 (Dchs1) to establish the molecular basis for Fat-Dachsous interactions. The binding domains of Fat4 and Dchs1 form an extended interface along extracellular cadherin (EC) domains 1-4 of each protein. Biophysical measurements indicate that Fat4-Dchs1 affinity is among the highest reported for cadherin superfamily members, which is attributed to an extensive network of salt bridges not present in structurally similar protocadherin homodimers. Furthermore, modeling suggests that unusual extracellular phosphorylation modifications directly modulate Fat-Dachsous binding by introducing charged contacts across the interface. Collectively, our analyses reveal how the molecular architecture of Fat4-Dchs1 enables them to form long-range, high-affinity interactions to maintain planar cell polarity.
Assuntos
Caderinas , Polaridade Celular , Proteínas Supressoras de Tumor , Humanos , Caderinas/química , Proteínas Supressoras de Tumor/química , Proteínas Relacionadas a Caderinas/químicaRESUMO
Metabolic reprogramming and metabolic plasticity allow cancer cells to fine-tune their metabolism and adapt to the ever-changing environments of the metastatic cascade, for which lipid metabolism and oxidative stress are of particular importance. NADPH is a central co-factor for both lipid and redox homeostasis, suggesting that cancer cells may require larger pools of NADPH to efficiently metastasize. NADPH is recycled through reduction of NADP+ by several enzymatic systems in cells; however, de novo NADP+ is synthesized only through one known enzymatic reaction, catalyzed by NAD+ kinase (NADK). Here, we show that NADK is upregulated in metastatic breast cancer cells enabling de novo production of NADP(H) and the expansion of the NADP(H) pools thereby increasing the ability of these cells to adapt to the challenges of the metastatic cascade and efficiently metastasize. Mechanistically, we found that metastatic signals lead to a histone H3.3 variant-mediated epigenetic regulation of the NADK promoter, resulting in increased NADK levels in cells with metastatic ability. Together, our work presents a previously uncharacterized role for NADK and de novo NADP(H) production as a contributor to breast cancer progression and suggests that NADK constitutes an important and much needed therapeutic target for metastatic breast cancers.
Assuntos
Neoplasias da Mama , Humanos , Feminino , NADP/metabolismo , Epigênese Genética , Estresse Oxidativo , NAD/metabolismo , Melanoma Maligno CutâneoRESUMO
Brain metastasis cancer-associated fibroblasts (bmCAFs) are emerging as crucial players in the development of breast cancer brain metastasis (BCBM), but our understanding of the underlying molecular mechanisms is limited. In this study, we aim to elucidate the pathological contributions of fucosylation (the post-translational modification of proteins by the dietary sugar L-fucose) to tumor-stromal interactions that drive the development of BCBM. Here, we report that patient-derived bmCAFs secrete high levels of polio virus receptor (PVR), which enhance the invasive capacity of BC cells. Mechanistically, we find that HIF1α transcriptionally upregulates fucosyltransferase 11, which fucosylates PVR, triggering its secretion from bmCAFs. Global phosphoproteomic analysis of BC cells followed by functional verification identifies cell-cell junction and actin cytoskeletal signaling as modulated by bmCAF-secreted, -fucosylated PVR. Our findings delineate a hypoxia- and fucosylation-regulated mechanism by which bmCAFs contribute to the invasiveness of BCBM in the brain.
Assuntos
Neoplasias Encefálicas , Neoplasias da Mama , Fibroblastos Associados a Câncer , Feminino , Humanos , Neoplasias Encefálicas/patologia , Neoplasias da Mama/patologia , Fibroblastos Associados a Câncer/patologia , Fibroblastos/patologia , Receptores ViraisRESUMO
Immunotherapy efficacy is limited in melanoma, and combinations of immunotherapies with other modalities have yielded limited improvements but also adverse events requiring cessation of treatment. In addition to ineffective patient stratification, efficacy is impaired by paucity of intratumoral immune cells (itICs); thus, effective strategies to safely increase itICs are needed. We report that dietary administration of L-fucose induces fucosylation and cell surface enrichment of the major histocompatibility complex (MHC)-II protein HLA-DRB1 in melanoma cells, triggering CD4+ T cell-mediated increases in itICs and anti-tumor immunity, enhancing immune checkpoint blockade responses. Melanoma fucosylation and fucosylated HLA-DRB1 associate with intratumoral T cell abundance and anti-programmed cell death protein 1 (PD1) responder status in patient melanoma specimens, suggesting the potential use of melanoma fucosylation as a strategy for stratifying patients for immunotherapies. Our findings demonstrate that fucosylation is a key mediator of anti-tumor immunity and, importantly, suggest that L-fucose is a powerful agent for safely increasing itICs and immunotherapy efficacy in melanoma.
Assuntos
Fucose , Melanoma , Humanos , Cadeias HLA-DRB1/genética , Cadeias HLA-DRB1/metabolismo , Fucose/metabolismo , Melanoma/tratamento farmacológico , Imunoterapia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/patologiaRESUMO
Traditionally, basic scientists have not been as engaged in the translational continuum when it comes to engagement with patients, caregivers, and other community stakeholders. In order to address this discrepancy, a multi-disciplinary team at Moffitt Cancer Center conceived of and enacted the Patient-Researcher Forum (PRF) to promote a community-engaged research approach through communication, compassion, and bi-directional research insight for both patients/caregivers and researchers. We outline the structure and implementation of the PRF, its participants, and qualitative and quantitative results across 14 sessions. PRF sessions were conducted between July 2018 and October 2019 and included 29 patients/caregivers and close to 200 researcher/staff participants; post participation survey response rates assessing the PRF experience were 27.6% (patients/caregivers) and 60.3% (researchers) on average. Research staff overwhelmingly reported that the PRF was beneficial, citing that it helped them gain new patient-centered perspectives and helped them practice communicating research to lay audiences. Patients/caregivers also reported that the PRF was valuable, indicating that they gained a better understanding of research and that they developed a personal connection with researchers. Our PRF model may provide a strategy for improving basic scientist communication, ethics, and understanding of research impacts on the populations they wish to serve. This innovative model provides a much-needed direct connection between basic scientists and patients/caregivers which creates a 2-way learning platform that fosters understanding and research ideas in the spirit of community-engaged research.
Assuntos
Cuidadores , Família , Modelos Organizacionais , Assistência Centrada no Paciente , Institutos de Câncer/organização & administração , Humanos , PesquisaRESUMO
Melanoma cells have the ability to switch to a dedifferentiated, invasive phenotype in response to multiple stimuli. Here, we show that exposure of melanomas to multiple stresses including BRAF-MEK inhibitor therapy, hypoxia, and UV irradiation leads to an increase in histone deacetylase 8 (HDAC8) activity and the adoption of a drug-resistant phenotype. Mass spectrometry-based phosphoproteomics implicated HDAC8 in the regulation of MAPK and AP-1 signaling. Introduction of HDAC8 into drug-naïve melanoma cells conveyed resistance both in vitro and in vivo. HDAC8-mediated BRAF inhibitor resistance was mediated via receptor tyrosine kinase activation, leading to MAPK signaling. Although HDACs function at the histone level, they also regulate nonhistone substrates, and introduction of HDAC8 decreased the acetylation of c-Jun, increasing its transcriptional activity and enriching for an AP-1 gene signature. Mutation of the putative c-Jun acetylation site at lysine 273 increased transcriptional activation of c-Jun in melanoma cells and conveyed resistance to BRAF inhibition. In vivo xenograft studies confirmed the key role of HDAC8 in therapeutic adaptation, with both nonselective and HDAC8-specific inhibitors enhancing the durability of BRAF inhibitor therapy. Our studies demonstrate that HDAC8-specific inhibitors limit the adaptation of melanoma cells to multiple stresses including BRAF-MEK inhibition. SIGNIFICANCE: This study provides evidence that HDAC8 drives transcriptional plasticity in melanoma cells in response to a range of stresses through direct deacetylation of c-Jun.Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/79/11/2947/F1.large.jpg.
Assuntos
Resistencia a Medicamentos Antineoplásicos/genética , Histona Desacetilases/metabolismo , Melanoma/tratamento farmacológico , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Proteínas Repressoras/metabolismo , Neoplasias Cutâneas/tratamento farmacológico , Acetilação , Animais , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Inibidores de Histona Desacetilases/farmacologia , Histona Desacetilases/genética , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Melanoma/metabolismo , Camundongos Endogâmicos NOD , Panobinostat/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Repressoras/antagonistas & inibidores , Proteínas Repressoras/genética , Neoplasias Cutâneas/metabolismo , Estresse Fisiológico/efeitos dos fármacos , Estresse Fisiológico/fisiologiaRESUMO
The fucose salvage pathway is a two-step process in which mammalian cells transform L-fucose into GDP-L-fucose, a universal fucose donor used by fucosyltransferases to modify glycans. Emerging evidence indicates the fucose salvage pathway and the fucosylation of proteins are altered during melanoma progression and metastasis. However the underlying mechanisms are not completely understood. Here, we report that the fucose salvage pathway inhibits invadopodia formation and extracellular matrix degradation by promoting α-1,2 fucosylation. Chemically or genetically increasing the fucose salvage pathway decreases invadopodium numbers and inhibits the proteolytic activity of invadopodia in WM793 melanoma cells. Inhibiting fucosylation by depleting fucokinase abrogates L-fucose-mediated inhibition of invadopodia, suggesting dependence on the fucose salvage pathway. The inhibition of invadopodium formation by L-fucose or ectopically expressed FUK could be rescued by treatment with α-1,2, but not α-1,3/α-1,4 fucosidase, implicating an α-1,2 fucose linkage-dependent anti-metastatic effect. The expression of FUT1, an α-1,2 fucosyltransferase, is remarkably down-regulated during melanoma progression, and the ectopic expression of FUT1 is sufficient to inhibit invadopodium formation and ECM degradation. Our findings indicate that the fucose salvage pathway can inhibit invadopodium formation, and consequently, invasiveness in melanoma via α-1,2 fucosylation. Re-activation of this pathway in melanoma could be useful for preventing melanoma invasion and metastasis.