RESUMEN
Disrupting the interplay between programmed cell death protein 1 (PD-1) and programmed death ligand 1 (PD-L1) is a powerful immunotherapeutic approach to cancer treatment. Herein, spherical nucleic acid (SNA) liposomal nanoparticle conjugates that incorporate a newly designed antisense DNA sequence specifically against PD-L1 (immune checkpoint inhibitor SNAs, or IC-SNAs) are explored as a strategy for blocking PD-1/PD-L1 signaling within the tumor microenvironment (TME). Concentration-dependent PD-L1 silencing with IC-SNAs is observed in MC38 colon cancer cells, where IC-SNAs decrease both surface PD-L1 (sPD-L1) and total PD-L1 expression. Furthermore, peritumoral administration of IC-SNAs in a syngeneic mouse model of MC38 colon cancer leads to reduced sPD-L1 expression in multiple cell populations within the TME, including tumor cells, dendritic cells, and myeloid derived suppressor cells. The treatment effectively increases CD8+ T cells accumulation and functionality in the TME, which ultimately inhibits tumor growth and extends animal survival. Taken together, these data show that IC-SNA nanoconstructs are capable of disrupting the PD-1/PD-L1 interplay via gene regulation, thereby providing a promising avenue for cancer immunotherapy.
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Although the strategy of therapeutic vaccination for the treatment of prostate cancer has advanced to and is available in the clinic (Sipuleucel-T), the efficacy of such therapy remains limited. Here, we develop Immunostimulatory Spherical Nucleic Acid (IS-SNA) nanostructures comprised of CpG oligonucleotides as adjuvant and prostate cancer peptide antigens, and evaluate their antitumor efficacy in syngeneic mouse models of prostate cancer. IS-SNAs with the specific structural feature of presenting both antigen and adjuvant CpG on the surface (hybridized model (HM) SNAs) induce stronger cytotoxic T lymphocyte (CTL) mediated antigen-specific killing of target cells than that for IS-SNAs with CpG on the surface and antigen encapsulated within the core (encapsulated model (EM) SNAs). Mechanistically, HM SNAs increase the co-delivery of CpG and antigen to dendritic cells over that for EM SNAs or admixtures of linear CpG and peptide, thereby improving cross-priming of antitumor CD8+ T cells. As a result, vaccination with HM SNAs leads to more effective antitumor immune responses in two prostate cancer models. These data demonstrate the importance of the structural positioning of peptide antigens together with adjuvants within IS-SNAs to the efficacy of IS-SNA-based cancer immunotherapy.
Asunto(s)
Vacunas contra el Cáncer/farmacología , Inmunoterapia/métodos , Nanoestructuras , Oligodesoxirribonucleótidos/farmacología , Neoplasias de la Próstata , Animales , Antígenos de Neoplasias/inmunología , Antígenos de Neoplasias/farmacología , Vacunas contra el Cáncer/inmunología , Reactividad Cruzada/efectos de los fármacos , Reactividad Cruzada/inmunología , Citotoxicidad Inmunológica/efectos de los fármacos , Humanos , Activación de Linfocitos/efectos de los fármacos , Activación de Linfocitos/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Oligodesoxirribonucleótidos/inmunología , Neoplasias de la Próstata/inmunología , Linfocitos T Citotóxicos/efectos de los fármacos , Linfocitos T Citotóxicos/inmunologíaRESUMEN
Plasmacytoid DCs (pDCs), the major producers of type I interferon, are principally recognized as key mediators of antiviral immunity. However, their role in tumor immunity is less clear. Depending on the context, pDCs can promote or suppress antitumor immune responses. In this study, we identified a naturally occurring pDC subset expressing high levels of OX40 (OX40+ pDC) enriched in the tumor microenvironment (TME) of head and neck squamous cell carcinoma. OX40+ pDCs were distinguished by a distinct immunostimulatory phenotype, cytolytic function, and ability to synergize with conventional DCs (cDCs) in generating potent tumor antigen-specific CD8+ T cell responses. Transcriptomically, we found that they selectively utilized EIF2 signaling and oxidative phosphorylation pathways. Moreover, depletion of pDCs in the murine OX40+ pDC-rich tumor model accelerated tumor growth. Collectively, we present evidence of a pDC subset in the TME that favors antitumor immunity.
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Antígenos de Neoplasias/inmunología , Linfocitos T CD8-positivos/inmunología , Células Dendríticas/inmunología , Neoplasias Experimentales/inmunología , Microambiente Tumoral/inmunología , Animales , Linfocitos T CD8-positivos/patología , Línea Celular Tumoral , Células Dendríticas/patología , Factor 2 Eucariótico de Iniciación/inmunología , Femenino , Humanos , Masculino , Ratones , Neoplasias Experimentales/patología , Receptores OX40/inmunologíaRESUMEN
In the case of cancer immunotherapy, nanostructures are attractive because they can carry all of the necessary components of a vaccine, including both antigen and adjuvant. Herein, we explore how spherical nucleic acids (SNAs), an emerging class of nanotherapeutic materials, can be used to deliver peptide antigens and nucleic acid adjuvants to raise immune responses that kill cancer cells, reduce (or eliminate) tumor growth, and extend life in three established mouse tumor models. Three SNA structures that are compositionally nearly identical but structurally different markedly vary in their abilities to cross-prime antigen-specific CD8+ T cells and raise subsequent antitumor immune responses. Importantly, the most effective structure is the one that exhibits synchronization of maximum antigen presentation and costimulatory marker expression. In the human papillomavirus-associated TC-1 model, vaccination with this structure improved overall survival, induced the complete elimination of tumors from 30% of the mice, and conferred curative protection from tumor rechallenges, consistent with immunological memory not otherwise achievable. The antitumor effect of SNA vaccination is dependent on the method of antigen incorporation within the SNA structure, underscoring the modularity of this class of nanostructures and the potential for the deliberate design of new vaccines, thereby defining a type of rational cancer vaccinology.
Asunto(s)
Vacunas contra el Cáncer , Neoplasias Experimentales/prevención & control , Ácidos Nucleicos/química , Animales , RatonesRESUMEN
Agonist antibodies (Ab) directed against costimulatory molecules on the surface of antigen-primed T cells are in various stages of pre-clinical and clinical trials, albeit with limited therapeutic benefit as single agents. The underlying mechanisms of action remain incompletely understood. Here, we demonstrate an inhibitory role of ecto-enzyme CD73 for agonistic anti-4-1BB/CD137 Ab therapy. In particular, anti-4-1BB treatment preferentially drives CD73- effector T cell response for tumor inhibition. Anti-CD73 neutralizing Ab further improves anti-4-1BB therapy associated with enhanced anti-tumor T cell immunity. However, the TGF-ß-rich tumor milieu confers resistance to anti-4-1BB therapy by sustaining CD73 expression primarily on infiltrating CD8+ T cells across several tumor models. TGF-ß blockade results in downregulation of CD73 expression on infiltrating T cells and sensitizes resistant tumors to agonistic anti-4-1BB therapy. Thus, our findings identify a mechanism of action for more effective clinical targeting of 4-1BB or likely other costimulatory molecules.
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Ligando 4-1BB/inmunología , 5'-Nucleotidasa/antagonistas & inhibidores , Anticuerpos Neutralizantes/inmunología , Linfocitos T CD8-positivos/inmunología , Proteína Relacionada con TNFR Inducida por Glucocorticoide/inmunología , Melanoma Experimental/terapia , 5'-Nucleotidasa/genética , 5'-Nucleotidasa/inmunología , Animales , Línea Celular Tumoral , Citotoxicidad Inmunológica , Regulación de la Expresión Génica/inmunología , Inmunoterapia/métodos , Melanoma Experimental/inmunología , Melanoma Experimental/patología , Glicoproteínas de Membrana/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ligando OX40 , Factor de Crecimiento Transformador beta1/metabolismo , Factores de Necrosis Tumoral/inmunologíaRESUMEN
Immunostimulatory spherical nucleic acids (IS-LSNAs) comprised of RNA selective for toll-like receptors (TLRs) 7/8 are synthesized and characterized. These structures consist of liposomal cores functionalized with a dense shell of RNA inserted into the wall of the lipid core via hydrophobic cholesterol moieties. IS-LSNAs potently activate TLR7/8 via NF-κΒ signaling in reporter cell lines and in primary immune cells as evidenced by cytokine production and the upregulation of costimulatory receptors. Importantly, they are preferentially taken up by plasmacytoid dendritic cells, an observation that makes them potentially useful for immunotherapy. In addition, these structures contain a core that can be loaded with antigens and used to prime T cells. In this regard, it is shown that dendritic cells treated with IS-SNAs loaded with ovalbumin peptide can prime ova specific CD8+ T cells. In addition to introducing the first IS-LSNAs consisting of RNA, these experiments show that one can facilitate an antigen-specific T cell response greater than that of free or cationic lipid-transfected RNA of the same sequence selective for TLR7/8. This work points toward the promise of using IS-LSNAs comprised of RNA as potent and highly tunable TLR-specific agents for the development of vaccines and other pharmaceuticals that require selective immunomodulation.
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Vacunas contra el Cáncer/química , Liposomas/química , Nanopartículas/química , Ácidos Nucleicos/química , Receptor Toll-Like 7/química , Receptor Toll-Like 8/química , Animales , Linfocitos T CD8-positivos/metabolismo , Células Cultivadas , Humanos , RatonesRESUMEN
Although a number of studies have recently explored the contribution of the adaptive immunity in IL-33-mediated antitumor effects, innate immune involvement has been poorly characterized. Utilizing Rag1-/- mice (lacking T and B lymphocytes), we show in this study that either systemic administration of recombinant IL-33 or ectopic expression of IL-33 in melanoma cells is sufficient to inhibit tumor growth independent of adaptive antitumor immunity. We have demonstrated that IL-33-mediated antitumor effects depend on expansion and activation of NK cells. Interestingly, IL-33 also promoted the expansion of active type 2 innate lymphoid cells (ILC2s) via its receptor, ST2, which in turn inhibited NK activation and cytotoxicity. This IL-33-induced ILC2 activity coincided with greater expression of the immunosuppressive ectoenzyme CD73. Removal of CD73 from ILC2s in culture with NK cells resulted in markedly increased activation levels in NK cells, offering a potential mechanism by which ILC2s might suppress NK cell-mediated tumor killing. Thus, our data reveal an important contribution of IL-33-induced ILC2 to tumor growth by weakening NK cell activation and tumor killing, regardless of adaptive immunity.
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Interleucina-33/metabolismo , Células Asesinas Naturales/inmunología , Linfocitos/inmunología , Melanoma/inmunología , Células Th2/inmunología , Animales , Línea Celular Tumoral , Proliferación Celular , Citotoxicidad Inmunológica , Genes RAG-1 , Inmunidad Innata , Proteína 1 Similar al Receptor de Interleucina-1/metabolismo , Activación de Linfocitos , Ratones , Ratones NoqueadosRESUMEN
[This corrects the article DOI: 10.3892/ol.2017.6584.].
RESUMEN
Although immunotherapy has been at the forefront of cancer therapy for the last several years, better clinical responses are still desired. Interleukin-33 is perhaps one of the most overlooked antitumor cytokines. Its ability to promote type 1 immune responses, which control tumor growth in preclinical animal models is overshadowed by its association with type 2 immunity and poor prognosis in some human cancers. Accumulating evidence shows that IL-33 is a powerful new tool for restoring and enhancing the body's natural antitumor immunity cycle. Furthermore, the antitumor mechanisms of IL-33 are two-fold, as it can directly boost CD8+ T cell function and restore dendritic cell dysfunction in vivo. Mechanistic studies have identified a novel pathway induced by IL-33 and its receptor ST2 in which dendritic cells avoid dysfunction and retain cross-priming abilities in tumor-bearing conditions. Here, we also comment on IL-33 data in human cancers and explore the idea that endogenous IL-33 may not deserve its reputation for promoting tumor growth. In fact, tumors may hijack the IL-33/ST2 axis to avoid immune surveillance and escape antitumor immunity.
Asunto(s)
Interleucina-33/inmunología , Neoplasias/inmunología , Animales , Humanos , Neoplasias/terapiaRESUMEN
Wee1-like protein kinase (WEE1) contributes to the upstream regulation of the cyclin-dependent kinase (CDK) complexes by mediating the inactivation of CDK1 [corrected]. Increased expression of WEE1 has been associated with the poor prognosis of patients with ovarian cancer. The present study aimed at examining the in vitro and in vivo antitumor activity of MK1775, a potent pharmacological inhibitor of WEE1, as a single agent against ovarian cancer cells. The cytotoxicity of MK1775 was examined in a panel of tumor cells using MTT in vitro. Subsequently, a cell apoptosis assay was performed in ovarian cancer SKOV3 and ID8 cells to characterize the function of MK1775 in tumor cell apoptosis, under either wild-type tumor protein 53 (p53) or null p53 status. In addition, cell cycle analysis and a western blot analysis were performed to validate the effect of MK1775 on cell cycle progression and to elucidate the underlying molecular mechanism of cell death. Finally, the in vivo antitumor efficacy of MK1775 as a single agent at a clinical well-tolerated dose was determined. A dose-dependent inhibitory effect of MK1775 on tumor cell viability was determined in distinct cell lines, including B16F10, LLC1, BPS1, EG7, ID8 and SKOV3. Results from the cell cycle analysis and western blotting indicated that MK1775 abrogated the G2/M checkpoint through inhibiting the phosphorylation of CDK1 and inducing the apoptosis of ovarian cancer cells that lacked mutations in p53 and breast cancer 1 (BRCA1). Additionally, a significant antitumor effect of MK1775 was observed in C57BL/6 mice bearing syngeneic ID8 ovarian tumors. The results of the present study supported the use of MK1775 as a monotherapy agent in ovarian cancer. MK1775 was effective at inducing mitotic catastrophe, independent of p53 and BRCA1 mutations. Therefore, WEE1 inhibition by MK1775 requires additional investigation to identify novel combination approaches in ovarian cancer therapy with the current DNA damaging agents, including irradiation treatment and cell cycle checkpoint inhibitors.
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The role of IL-33, particularly in tumor growth and tumor immunity, remains ill-defined. We show that exogenous IL-33 can induce robust antitumor effect through a CD8+ T cell-dependent mechanism. Systemic administration of rIL-33 alone was sufficient to inhibit growth of established tumors in transplant and de novo melanoma tumorigenesis models. Notably, in addition to a direct action on CD8+ T cell expansion and IFN-γ production, rIL-33 therapy activated myeloid dendritic cells (mDCs) in tumor-bearing mice, restored antitumor T cell activity, and increased Ag cross-presentation within the tumor microenvironment. Furthermore, combination therapy consisting of rIL-33 and agonistic anti-CD40 Abs demonstrated synergistic antitumor activity. Specifically, MyD88, an essential component of the IL-33 signaling pathway, was required for the IL-33-mediated increase in mDC number and upregulation in expression of costimulatory molecules. Importantly, we identified that the IL-33 receptor ST2, MyD88, and STAT1 cooperate to induce costimulatory molecule expression on mDCs in response to rIL-33. Thus, our study revealed a novel IL-33-ST2-MyD88-STAT1 axis that restores mDC activation and maturation in established cancer and, thereby, the magnitude of antitumor immune responses, suggesting a potential use of rIL-33 as a new immunotherapy option to treat established cancer.
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Células Dendríticas/efectos de los fármacos , Interleucina-33/farmacología , Neoplasias Experimentales/inmunología , Animales , Antígenos CD40/antagonistas & inhibidores , Linfocitos T CD8-positivos/inmunología , Citotoxicidad Inmunológica/efectos de los fármacos , Células Dendríticas/fisiología , Proteína 1 Similar al Receptor de Interleucina-1/fisiología , Ratones , Ratones Endogámicos C57BL , Factor 88 de Diferenciación Mieloide/fisiología , Fosfohidrolasa PTEN/fisiología , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas Recombinantes/farmacología , Factor de Transcripción STAT1/fisiología , Células Tumorales CultivadasRESUMEN
Emerging studies suggest that dominant peripheral tolerance is a major mechanism of immune escape in disseminated leukemia. Using an established murine acute myeloid leukemia (AML) model, we here show that systemic administration of recombinant IL-33 dramatically inhibits the leukemia growth and prolongs the survival of leukemia-bearing mice in a CD8+ T cell dependent manner. Exogenous IL-33 treatment enhanced anti-leukemia activity by increasing the expansion and IFN-γ production of leukemia-reactive CD8+ T cells. Moreover, IL-33 promoted dendritic cell (DC) maturation and activation in favor of its cross presentation ability to evoke a vigorous anti-leukemia immune response. Finally, we found that the combination of PD-1 blockade with IL-33 further prolonged the survival, with half of the mice achieving complete regression. Our data establish a role of exogenous IL-33 in reversing T cell tolerance, and suggest its potential clinical implication into leukemia immunotherapy.
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Linfocitos T CD8-positivos/metabolismo , Tolerancia Inmunológica , Proteína 1 Similar al Receptor de Interleucina-1/metabolismo , Interleucina-33/uso terapéutico , Leucemia Mieloide Aguda/metabolismo , Animales , Presentación de Antígeno , Línea Celular Tumoral , Proliferación Celular , Supervivencia Celular , Células Dendríticas/metabolismo , Inmunoterapia , Interferón gamma/metabolismo , Ratones , Ratones Endogámicos C57BL , Receptor de Muerte Celular Programada 1/antagonistas & inhibidores , Proteínas Recombinantes/uso terapéutico , Análisis de Regresión , Resultado del TratamientoRESUMEN
α-Difluoromethylornithine (DFMO) is currently used in chemopreventive regimens primarily for its conventional direct anticarcinogenesic activity. However, little is known about the effect of ornithine decarboxylase (ODC) inhibition by DFMO on antitumor immune responses. We showed in this study that pharmacologic blockade of ODC by DFMO inhibited tumor growth in intact immunocompetent mice, but abrogated in the immunodeficient Rag1(-/-) mice, suggesting that antitumor effect of DFMO is dependent on the induction of adaptive antitumor T cell immune responses. Depletion of CD8(+) T cells impeded the tumor-inhibiting advantage of DFMO. Moreover, DFMO treatment enhanced antitumor CD8(+) T cell infiltration and IFN-γ production and augmented the efficacy of adoptive T cell therapy. Importantly, DFMO impaired Gr1(+)CD11b(+) myeloid-derived suppressor cells (MDSCs) suppressive activity through at least two mechanisms, including reducing arginase expression and activity and inhibiting the CD39/CD73-mediated pathway. MDSCs were one primary cellular target of DFMO as indicated by both adoptive transfer and MDSC-depletion analyses. Our findings establish a new role of ODC inhibition by DFMO as a viable and effective immunological adjunct in effective cancer treatment, thereby adding to the growing list of chemoimmunotherapeutic applications of these agents.
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Eflornitina/farmacología , Células Mieloides/efectos de los fármacos , Neoplasias Experimentales/inmunología , Inhibidores de la Ornitina Descarboxilasa/farmacología , Ornitina Descarboxilasa/inmunología , Escape del Tumor/efectos de los fármacos , Animales , Antineoplásicos/farmacología , Linfocitos T CD8-positivos/inmunología , Ensayo de Immunospot Ligado a Enzimas , Citometría de Flujo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Células Mieloides/inmunología , Reacción en Cadena en Tiempo Real de la Polimerasa , Escape del Tumor/inmunologíaRESUMEN
miR155 is a regulator of immune cell development and function that is generally thought to be immunostimulatory. However, we report here that genetic ablation of miR155 renders mice resistant to chemical carcinogenesis and the growth of several transplanted tumors, suggesting that miR155 functions in immunosuppression and tumor promotion. Host miR155 deficiency promoted overall antitumor immunity despite the finding of defective responses of miR155-deficient dendritic cells and antitumor T cells. Further analysis of immune cell compartments revealed that miR155 regulated the accumulation of functional myeloid-derived suppressive cells (MDSC) in the tumor microenvironment. Specifically, miR155 mediated MDSC suppressor activity through at least two mechanisms, including SOCS1 repression and a reduced ability to license the generation of CD4(+)Foxp3(+) regulatory T cells. Importantly, we demonstrated that miR155 expression was required for MDSC to facilitate tumor growth. Thus, our results revealed a contextual function for miR155 in antitumor immunity, with a role in MDSC support that appears to dominate in tumor-bearing hosts. Overall, the balance of these cellular effects appears to be a root determinant of whether miR155 promotes or inhibits tumor growth.