Melanomas commonly undergo a phenotype switch, from a proliferative to an invasive state. Such tumor cell plasticity contributes to immunotherapy resistance; however, the mechanisms are not ...completely understood and thus are therapeutically unexploited. Using melanoma mouse models, we demonstrated that blocking the MNK1/2-eIF4E axis inhibited melanoma phenotype switching and sensitized melanoma to anti-PD-1 immunotherapy. We showed that phosphoeIF4E-deficient murine melanomas expressed high levels of melanocytic antigens, with similar results verified in patient melanomas. Mechanistically, we identified phospho-eIF4E-mediated translational control of NGFR, a critical effector of phenotype switching. Genetic ablation of phospho-eIF4E reprogrammed the immunosuppressive microenvironment, exemplified by lowered production of inflammatory factors, decreased PD-L1 expression on dendritic cells and myeloid-derived suppressor cells, and increased CD8.sup.+ T cell infiltrates. Finally, dual blockade of the MNK1/2-eIF4E axis and the PD-1/ PD-L1 immune checkpoint demonstrated efficacy in multiple melanoma models regardless of their genomic classification. An increase in the presence of intratumoral stem-like TCF1.sup.+PD-1.sup.+CD8.sup.+ T cells, a characteristic essential for durable antitumor immunity, was detected in mice given a MNK1/2 inhibitor and anti-PD-1 therapy. Using MNK1/2 inhibitors to repress phospho-eIF4E thus offers a strategy to inhibit melanoma plasticity and improve response to anti-PD-1 immunotherapy.
In eukaryotic cells, the rate-limiting component for cap-dependent mRNA translation is the translation initiation factor eIF4E. eIF4E is overexpressed in a variety of human malignancies, but whether ...it has a role in melanoma remains obscure. We hypothesized that eIF4E promotes melanoma cell proliferation and facilitates the development of acquired resistance to the BRAF inhibitor vemurafenib. We show that eIF4E is overexpressed in a panel of melanoma cell lines, compared with immortalized melanocytes. Knockdown of eIF4E significantly repressed the proliferation of a subset of melanoma cell lines. Moreover, in BRAFV600E melanoma cell lines, vemurafenib inhibits 4E-BP1 phosphorylation, thus promoting its binding to eIF4E. Cap-binding and polysome profiling analysis confirmed that vemurafenib stabilizes the eIF4E–4E-BP1 association and blocks mRNA translation, respectively. Conversely, in cells with acquired resistance to vemurafenib, there is an increased dependence on eIF4E for survival; 4E-BP1 is highly phosphorylated and thus eIF4E-4E-BP1 associations are impeded. Moreover, increasing eIF4E activity by silencing 4E-BP1/2 renders vemurafenib-responsive cells more resistant to BRAF inhibition. In conclusion, these data suggest that therapeutically targeting eIF4E may be a viable means of inhibiting melanoma cell proliferation and overcoming vemurafenib resistance.
Melanoma can be stratified into unique subtypes based on distinct pathologies. The acral/mucosal melanoma subtype is characterized by aberrant and constitutive activation of the proto-oncogene ...receptor tyrosine kinase C-KIT, which drives tumorigenesis. Treatment of these melanoma patients with C-KIT inhibitors has proven challenging, prompting us to investigate the downstream effectors of the C-KIT receptor. We determined that C-KIT stimulates MAP kinase-interacting serine/threonine kinases 1 and 2 (MNK1/2), which phosphorylate eukaryotic translation initiation factor 4E (eIF4E) and render it oncogenic. Depletion of MNK1/2 in melanoma cells with oncogenic C-KIT inhibited cell migration and mRNA translation of the transcriptional repressor SNA/1 and the cell cycle gene CCNE1. This suggested that blocking MNK1/2 activity may inhibit tumor progression, at least in part, by blocking translation initiation of mRNAs encoding cell migration proteins. Moreover, we developed an MNK1/2 inhibitor (SEL201), and found that SEL201-treated KIT-mutant melanoma cells had lower oncogenicity and reduced metastatic ability. Clinically, tumors from melanoma patients harboring K/T mutations displayed a marked increase in MNK1 and phospho-eIF4E. Thus, our studies indicate that blocking MNK1/2 exerts potent antimelanoma effects and support blocking MNK1/2 as a potential strategy to treat patients positive for K/T mutations.
Abstract
Objective:
Phosphorylated Eukaryotic translation initiation factor 4E (p-eIF4E) is a critical regulator of protein synthesis and is phosphorylated by MNK1/2 to promote the translation of a ...mRNA subset. We have shown that blocking the eIF4E phosphorylation increased the anti-tumor immune response, especially the CD8 T cell activation. However, the role of p-eIF4E in CD4 T cell subsets, and in particular regulatory T cells (Tregs) is still unknown. The aim of this study was to explore the impact that the absence of p-eIF4E could have on the Treg activity as well as in an inflammatory context.
Methods:
To investigate the role of p-eIF4E on Tregs activity, we used genetically modified mice expressing a non-phosphorylatable form of eIF4E (KI mice). First, we analyzed Treg activity in WT and KI mice and the same mice subjected to dextran sulfate sodium (DSS)-induced colitis. We also analyzed the colonic immune cell infiltration using spectral flow cytometry and CODEX technology.
Results:
Using our mouse models, we observed that the p-eIF4E lack led to a decrease in Treg stability as well as a decrease in their ability to control the helper T cell (Th) proliferation. In the colitis context, we observed an increase in the disease severity in KI compared to WT mice characterized by an increase in colonic immune infiltration. Moreover, our mesenteric lymph node and colon immunophenotyping revealed a significant decrease in Treg, and an increase in Th expressing IFNγ in KI compared to WT mice. Finally, we observed a decrease in KI Treg ability to migrate to the lymph nodes.
Conclusion:
These results demonstrate for the first time the preponderant role of p-eIF4E in the control of Treg stability and Treg migration but also in their ability to regulate inflammation.
Le genre Morbillivirus comprend entre autre le virus de rougeole (MeV) et le virus de la maladie de Carré (CDV), des agents pathogènes majeurs chez leurs hôtes respectifs. L'un des signes ...caractéristiques de cette infection est l'apparition d'une immunosuppression chez l'hôte empêchant l'établissement d'une réponse immunitaire nécessaire pour neutraliser l'infection. Cette immunosuppression est principalement induite par la synthèse de la protéine V, responsable de la virulence. En effet, elle est capable de bloquer les voies de signalisation des interférons de type 1 en interagissant avec I'ARN hélicase MDA5 ou bien les facteurs de transcription STAT1 et STAT2.Au cours de précédant travaux, nous avons observé une induction différentielle des gènes stimulés par les interférons (ISG) en présence de protéines V mutées sur le résidu Y110 incapable d'interagir avec STAT1 ou sur la cystéine C255 empêchant l'interaction avec STAT2 et MDA5. Afin d'identifier des ISG et des voies de signalisations responsables d'une réponse antivirale contre le CDV, nous avons effectué une étude transcriptomique avec les différentes protéines V.Nous observons la présence de deux profils transcriptionnels distincts. Le premier correspond à une forte induction des ISG en absence ou en présence de la protéine V mutée sur le résidu C255 (VC255S) tandis qu'un second profil montre une réduction importante de l'induction des ISGs en présence de la protéine VWT et de la protéine V mutée sur le résidu Y110 (VY110H). Parmi cette diminution, on observe l'absence de protéines majeures de l'immunité innée comme la voie d'induction des IFN ou de la voie des complexes majeurs d'histocompatibilité (CMH). Pour mieux caractériser le rôle de la protéine V sur le CMH, nous avons confirmé par PCR en temps réel l'expression des gènes de la voie du CMH et nous observons une diminution en présence de la protéine V des gènes de la voie classique tels que HLA et TAP. Nous avons de plus montré par cytométrie en flux que l'induction de l'expression à la surface de CMH de classe 1 était diminuée en présence de V.En conclusion, cette étude a contribué à la caractérisation du rôle de la protéine V sur l'immunosuppression. En bloquant les voies des interférons, elle inhibe la synthèse de gènes de la réponse antivirale et de l'immunité innée et en interférant avec la voie de CMH, elle empêcherait la présentation à la surface de la cellule d'antigènes viraux diminuant l'établissement et l'efficacité d'une réponse immunitaire adaptative.
Abstract
Introduction: Melanoma is the deadliest form of skin cancer. Melanoma phenotype switching is characterized by reduced expression of melanocyte lineage transcription factor MITF and its ...downstream targets, leading to increased invasiveness of melanoma cells and resistance to both targeted therapy and immunotherapy, and worse prognosis. In melanoma, MAPK and PI3K pathways ultimately converge upon eukaryotic translation initiation factor 4E (eIF4E) to induce its phosphorylation (p-eIF4E), which is critical for the oncogenicity of eIF4E. Here, we investigate the role of p-eIF4E in melanoma progression and tumor immunity.
Methods: We crossed p-eIF4E deficient (eIF4EKI) mice with an inducible melanoma mouse model. We monitored the primary tumor outgrowth, metastasis, and survival of the eIF4EKI mice versus eIF4EWT mice. Melanoma samples were isolated for further investigation.
Results: Compared to the eIF4EWT mice, eIF4EKI mice exhibit significantly delayed tumor growth, reduced metastasis, and increased survival. Increased expression of MITF and downstream melanoma antigens were observed in eIF4EKI tumors, suggesting a p-eIF4E-mediated phenotype switching. Cytokine array analysis revealed a novel proinvasive cytokine signature in eIF4EWT melanoma primary culture, further supporting a role of phospho-eIF4E in phenotype switching. The cytokine profiling also revealed a pro-myeloid-derived suppressor cell (MDSC) cytokine signature in the eIF4EWT tumor, indicating a p-eIF4E-linked immunosuppression. In support of the immune suppressive cytokine signature associated with phospho-eIF4E expressing melanomas, immune phenotyping of eIF4EWT melanomas showed a significant increase in MDSCs and less cytotoxic T cells, compared to eIF4EKI melanomas. Finally, pharmacologic inhibition of p-eIF4E in combination with anti-PD-1 immunotherapy results in a synergistic delay in primary tumor outgrowth and reduced metastasis.
Conclusions: Here we showed that phosphorylation of eIF4E promotes melanoma phenotype switching, leading to increased invasiveness and reduced expression of tumor-associated antigens. Further, by increasing the secretion of pro-MDSC cytokines, p-eIF4E permits an immunosuppressive microenvironment. Pharmacologic inhibition of p-eIF4E sensitizes melanoma to anti-PD-1 immunotherapy, potentially by increasing melanoma antigen expression and compromising MDSC-mediated immunosuppression. This study provides a novel therapeutic approach for the treatment of melanoma.
Citation Format: Fan Huang, Christophe Gonçalves, Qianyu Guo, Joelle Rémy-Sarrazin, Audrey Emond, William Yang, Dany Plourde, Margarita Bartish, Jie Su, Yao Zhan, Marina G. Gimeno, Elie Khoury, Alexandre Benoit, David Dankort, Wilson H. Miller, Sonia V. del Rincón. Phosphorylation of eIF4E promotes phenotype switching and MDSC-mediated immunosuppression in melanoma abstract. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr A53.
Nos últimos anos tem havido um aumento significativo da procura de frutos vermelhos. Os mirtilos são considerados frutos de boa qualidade, dado o seu elevado teor em compostos fitoquímicos ...biologicamente ativos, associados a efeitos benéficos para a saúde e bem-estar do Homem. A produção em modo biológico é reconhecida pelo consumidor como um processo que melhora a qualidade do produto.No presente trabalho pretendeu-se avaliar o efeito do modo de produção (biológico versus convencional) de três cultivares de mirtilo (Duke, Bluecrop, Ozarkblue) nas suas propriedades físico-químicas, e em particular na sua composição fenólica e atividade antioxidante. Foi ainda estudado o efeito da temperatura de armazenamento (± 5ºC e ± 15-25ºC) sobre essas propriedades. Para tal, as amostras foram analisadas à colheita e após 7 e 14 dias de armazenamento.A atividade antioxidante medida pelos métodos ABTS e DPPH mostrou que não há diferenças significativas entre as cultivares estudadas, sendo elevada em todos os casos. Os resultados obtidos confirmam, por isso, que o mirtilo é uma importante fonte de compostos fenólicos com elevada atividade antioxidante.Foi ainda verificado existirem algumas diferenças significativas em algumas propriedades em função da variedade (nomeadamente teor em matéria seca, cor ou textura). Também se verificara diferenças significativas em função do modo de produção, o qual influencia em particular a acidez e a doçura, o teor em taninos, a cor e a elasticidade dos frutos. Por fim, a temperatura de armazenamento mostrou ter uma influência significativa apenas no que respeita às propriedades físicas, nomeadamente cor e textura.
PDF
