RESUMO
PURPOSE: Recent advances in immunotherapy of advanced human cancers underscored the need to address and eliminate tumor immune evasion. The myeloid-derived suppressor cells (MDSC) are important inhibitors of T-cell responses in solid tumors, such as prostate cancers. However, targeting MDSCs proved challenging due to their phenotypic heterogeneity. EXPERIMENTAL DESIGN: Myeloid cell populations were evaluated using flow cytometry on blood samples, functional assays, and immunohistochemical/immunofluorescent stainings on specimens from healthy subjects, localized and metastatic castration-resistant prostate cancer patients. RESULTS: Here, we identify a population of Lin(-)CD15(HI)CD33(LO) granulocytic MDSCs that accumulate in patients' circulation during prostate cancer progression from localized to metastatic disease. The prostate cancer-associated MDSCs potently inhibit autologous CD8(+) T cells' proliferation and production of IFNγ and granzyme-B. The circulating MDSCs have high levels of activated STAT3, which is a central immune checkpoint regulator. The granulocytic pSTAT3(+) cells are also detectable in patients' prostate tissues. We previously generated an original strategy to silence genes specifically in Toll-like Receptor-9 (TLR9) positive myeloid cells using CpG-siRNA conjugates. We demonstrate that human granulocytic MDSCs express TLR9 and rapidly internalize naked CpG-STAT3siRNA, thereby silencing STAT3 expression. STAT3 blocking abrogates immunosuppressive effects of patients-derived MDSCs on effector CD8(+) T cells. These effects depended on reduced expression and enzymatic activity of Arginase-1, a downstream STAT3 target gene and a potent T-cell inhibitor. CONCLUSIONS: Overall, we demonstrate the accumulation of granulocytic MDSCs with prostate cancer progression and the feasibility of using TLR9-targeted STAT3siRNA delivery strategy to alleviate MDSC-mediated immunosuppression.
Assuntos
Células Mieloides/metabolismo , Neoplasias da Próstata/genética , Fator de Transcrição STAT3/genética , Receptor Toll-Like 9/genética , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/patologia , Linhagem da Célula/imunologia , Proliferação de Células/genética , Inativação Gênica , Humanos , Terapia de Imunossupressão , Ativação Linfocitária/imunologia , Masculino , Células Mieloides/imunologia , Células Neoplásicas Circulantes/imunologia , Células Neoplásicas Circulantes/patologia , Neoplasias da Próstata/sangue , Neoplasias da Próstata/imunologia , Neoplasias da Próstata/patologia , Neoplasias da Próstata/terapia , Fator de Transcrição STAT3/biossíntese , Receptor Toll-Like 9/biossíntese , Evasão Tumoral/genéticaRESUMO
Tumour progression is associated with immune-suppressive conditions that facilitate the escape of tumour cells from the regimen of immune cells, subsequently paralysing the host defence mechanisms. Induction of CD4(+) CD25(+) FoxP3(+) T regulatory (Treg) cells has been implicated in the tumour immune escape mechanism, although the novel anti-cancer treatment strategies targeting Treg cells remain unknown. The focus of this study is to define the interaction between tumour and immune system, i.e. how immune tolerance starts and gradually leads to the induction of adaptive Treg cells in the tumour microenvironment. Our study identified hyperactivated mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) -signalling as a potential target for reversing Treg cell augmentation in breast cancer patients. In more mechanistic detail, pharmacological inhibitors of MEK/ERK signalling inhibited transforming growth factor-ß (TGF-ß) production in tumour cells that essentially blocked TGF-ß-SMAD3/SMAD4-mediated induction of CD25/interleukin-2 receptor α on CD4(+) T-cell surface. As a result high-affinity binding of interleukin-2 on those cells was prohibited, causing lack of Janus kinase 1 (JAK1)/JAK3-mediated signal transducer and activator of transcription 3 (STAT3)/STAT5 activation required for FoxP3 expression. Finally, for a more radical approach towards a safe MEK inhibitor, we validate the potential of multi-kinase inhibitor curcumin, especially the nano-curcumin made out of pure curcumin with greater bioavailability; in repealing tumour-shed TGF-ß-induced Treg cell augmentation.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Curcumina/farmacologia , MAP Quinase Quinase Quinases/antagonistas & inibidores , Comunicação Parácrina/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Linfócitos T Reguladores/efeitos dos fármacos , Fator de Crescimento Transformador beta/metabolismo , Microambiente Tumoral , Animais , Neoplasias da Mama/enzimologia , Neoplasias da Mama/genética , Neoplasias da Mama/imunologia , Neoplasias da Mama/patologia , Estudos de Casos e Controles , Química Farmacêutica , Técnicas de Cocultura , Relação Dose-Resposta a Droga , Feminino , Fatores de Transcrição Forkhead/metabolismo , Humanos , Subunidade alfa de Receptor de Interleucina-2/metabolismo , MAP Quinase Quinase Quinases/genética , MAP Quinase Quinase Quinases/metabolismo , Camundongos Endogâmicos BALB C , Nanopartículas , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteínas Smad/metabolismo , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Fatores de Tempo , Fator de Crescimento Transformador beta/genética , Células Tumorais Cultivadas , Evasão Tumoral/efeitos dos fármacosRESUMO
Cancer radiotherapy may be immunogenic, but it is unclear why its immunogenic effects are rarely sufficient to prevent tumor recurrence. Here, we report a novel Toll-like receptor 9 (TLR9)-dependent mechanism that initiates tumor regrowth after local radiotherapy. Systemic inhibition of TLR9, but not TLR4, delayed tumor recurrence in mouse models of B16 melanoma, MB49 bladder cancer, and CT26 colon cancer after localized high-dose tumor irradiation. Soluble factors in the microenvironment of regressing tumors triggered TLR9 signaling in freshly recruited myeloid cells appearing within four days of radiotherapy. The tumorigenic effects of TLR9 depended on MyD88/NF-κB-mediated upregulation of interleukin (IL)-6 expression, which in turn resulted in downstream activation of Jak/STAT3 signaling in myeloid cells. In comparing global gene expression in wild-type, TLR9-, or STAT3-deficient myeloid cells derived from irradiated tumors, we identified a unique set of TLR9/STAT3-regulated genes involved in tumor-promoting inflammation and revascularization. Blocking STAT3 function by two myeloid-specific genetic strategies corrected TLR9-mediated cancer recurrence after radiotherapy. Our results suggest that combining localized tumor irradiation with myeloid cell-specific inhibition of TLR9/STAT3 signaling may help eliminate radioresistant cancers.