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1.
Nat Commun ; 10(1): 3959, 2019 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-31477729

RESUMO

Successful combinations of radiotherapy and immunotherapy depend on the presence of live T cells within the tumor; however, radiotherapy is believed to damage T cells. Here, based on longitudinal in vivo imaging and functional analysis, we report that a large proportion of T cells survive clinically relevant doses of radiation and show increased motility, and higher production of interferon gamma, compared with T cells from unirradiated tumors. Irradiated intratumoral T cells can mediate tumor control without newly-infiltrating T cells. Transcriptomic analysis suggests T cell reprogramming in the tumor microenvironment and similarities with tissue-resident memory T cells, which are more radio-resistant than circulating/lymphoid tissue T cells. TGFß is a key upstream regulator of T cell reprogramming and contributes to intratumoral Tcell radio-resistance. These findings have implications for the design of radio-immunotherapy trials in that local irradiation is not inherently immunosuppressive, and irradiation of multiple tumors might optimize systemic effects of radiotherapy.

2.
Cancer Res ; 2018 Nov 27.
Artigo em Inglês | MEDLINE | ID: mdl-30482772

RESUMO

Given the integral role of Stimulator of interferon genes (STING, TMEM173) in the innate immune response, its loss or impairment in cancer is thought to primarily affect antitumor immunity. Here we demonstrate a role for STING in the maintenance of cellular homeostasis through regulation of the cell cycle. Depletion of STING in human and murine cancer cells and tumors resulted in increased proliferation compared to wild-type controls. Microarray analysis revealed genes involved in cell cycle regulation are differentially expressed in STINGko compared to WT MEFs. STING-mediated regulation of the cell cycle converged on NF-κB- and p53-driven activation of p21. The absence of STING led to premature activation of cyclin-dependent kinase 1 (CDK1), early onset S phase and mitosis, and increased chromosome instability, which was enhanced by ionizing radiation (IR). These results suggest a pivotal role for STING in maintaining cellular homeostasis and response to genotoxic stress.

3.
Immunity ; 49(3): 490-503.e4, 2018 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-30170810

RESUMO

The NF-κB pathway plays a crucial role in supporting tumor initiation, progression, and radioresistance of tumor cells. However, the role of the NF-κB pathway in radiation-induced anti-tumor host immunity remains unclear. Here we demonstrated that inhibiting the canonical NF-κB pathway dampened the therapeutic effect of ionizing radiation (IR), whereas non-canonical NF-κB deficiency promoted IR-induced anti-tumor immunity. Mechanistic studies revealed that non-canonical NF-κB signaling in dendritic cells (DCs) was activated by the STING sensor-dependent DNA-sensing pathway. By suppressing recruitment of the transcription factor RelA onto the Ifnb promoter, activation of the non-canonical NF-κB pathway resulted in decreased type I IFN expression. Administration of a specific inhibitor of the non-canonical NF-κB pathway enhanced the anti-tumor effect of IR in murine models. These findings reveal the potentially interactive roles for canonical and non-canonical NF-κB pathways in IR-induced STING-IFN production and provide an alternative strategy to improve cancer radiotherapy.

4.
J Clin Oncol ; 36(16): 1611-1618, 2018 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-29437535

RESUMO

Purpose Stereotactic body radiotherapy (SBRT) may stimulate innate and adaptive immunity to augment immunotherapy response. Multisite SBRT is an emerging paradigm for treating metastatic disease. Anti-PD-1-treatment outcomes may be improved with lower disease burden. In this context, we conducted a phase I study to evaluate the safety of pembrolizumab with multisite SBRT in patients with metastatic solid tumors. Patients and Methods Patients progressing on standard treatment received SBRT to two to four metastases. Not all metastases were targeted, and metastases > 65 mL were partially irradiated. SBRT dosing varied by site and ranged from 30 to 50 Gy in three to five fractions with predefined dose de-escalation if excess dose-limiting toxicities were observed. Pembrolizumab was initiated within 7 days after completion of SBRT. Pre- and post-SBRT biopsy specimens were analyzed in a subset of patients to quantify interferon-γ-induced gene expression. Results A total of 79 patients were enrolled; three patients did not receive any treatment and three patients only received SBRT. Patients included in the analysis were treated with SBRT and at least one cycle of pembrolizumab. Most (94.5%) of patients received SBRT to two metastases. Median follow-up for toxicity was 5.5 months (interquartile range, 3.3 to 8.1 months). Six patients experienced dose-limiting toxicities with no radiation dose reductions. In the 68 patients with imaging follow-up, the overall objective response rate was 13.2%. Median overall survival was 9.6 months (95% CI, 6.5 months to undetermined) and median progression-free survival was 3.1 months (95% CI, 2.9 to 3.4 months). Expression of interferon-γ-associated genes from post-SBRT tumor biopsy specimens significantly correlated with nonirradiated tumor response. Conclusion Multisite SBRT followed by pembrolizumab was well tolerated with acceptable toxicity. Additional studies exploring the clinical benefit and predictive biomarkers of combined multisite SBRT and PD-1-directed immunotherapy are warranted.

5.
Nature ; 545(7652): 98-102, 2017 05 04.
Artigo em Inglês | MEDLINE | ID: mdl-28445461

RESUMO

The relative contribution of the effector molecules produced by T cells to tumour rejection is unclear, but interferon-γ (IFNγ) is critical in most of the analysed models. Although IFNγ can impede tumour growth by acting directly on cancer cells, it must also act on the tumour stroma for effective rejection of large, established tumours. However, which stroma cells respond to IFNγ and by which mechanism IFNγ contributes to tumour rejection through stromal targeting have remained unknown. Here we use a model of IFNγ induction and an IFNγ-GFP fusion protein in large, vascularized tumours growing in mice that express the IFNγ receptor exclusively in defined cell types. Responsiveness to IFNγ by myeloid cells and other haematopoietic cells, including T cells or fibroblasts, was not sufficient for IFNγ-induced tumour regression, whereas responsiveness of endothelial cells to IFNγ was necessary and sufficient. Intravital microscopy revealed IFNγ-induced regression of the tumour vasculature, resulting in arrest of blood flow and subsequent collapse of tumours, similar to non-haemorrhagic necrosis in ischaemia and unlike haemorrhagic necrosis induced by tumour necrosis factor. The early events of IFNγ-induced tumour ischaemia resemble non-apoptotic blood vessel regression during development, wound healing or IFNγ-mediated, pregnancy-induced remodelling of uterine arteries. A better mechanistic understanding of how solid tumours are rejected may aid the design of more effective protocols for adoptive T-cell therapy.


Assuntos
Vasos Sanguíneos/crescimento & desenvolvimento , Hipóxia Celular/imunologia , Interferon gama/imunologia , Isquemia/imunologia , Neoplasias/irrigação sanguínea , Neoplasias/imunologia , Remodelação Vascular , Animais , Vasos Sanguíneos/imunologia , Vasos Sanguíneos/metabolismo , Linhagem Celular Tumoral , Células Endoteliais/imunologia , Células Endoteliais/metabolismo , Feminino , Interferon gama/biossíntese , Microscopia Intravital , Isquemia/metabolismo , Isquemia/patologia , Masculino , Camundongos , Necrose , Neoplasias/metabolismo , Neoplasias/patologia , Receptores de Interferon/metabolismo , Células Estromais/imunologia , Células Estromais/metabolismo , Especificidade por Substrato , Cicatrização
6.
Cancer Immunol Res ; 5(2): 127-136, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-28077434

RESUMO

Adoptively transferred CD8+ T cells can stabilize the size of solid tumors over long periods of time by exclusively recognizing antigen cross-presented on tumor stroma. However, these tumors eventually escape T-cell-mediated growth control. The aim of this study was to eradicate such persistent cancers. In our model, the SIYRYYGL antigen is expressed by cancer cells that lack the MHC-I molecule Kb needed for direct presentation, but the antigen is picked up and cross-presented by tumor stroma. A single injection of antigen-specific 2C CD8+ T cells caused long-term inhibition of tumor growth, but without further intervention, tumors started to progress after approximately 3 months. Escape was associated with reduced numbers of circulating 2C cells. Tumor-infiltrating 2C cells produced significantly less TNFα and expressed more of the "exhaustion" markers PD-1 and Tim-3 than T cells from lymphoid organs. High-dose local ionizing radiation, depletion of myeloid-derived suppressor cells, infusions of additional 2C cells, and antibodies blocking PD-L1 did not prevent tumor escape. In contrast, adoptive transfer of allogeneic CD4+ T cells restored the numbers of circulating Ag-specific CD8+ T cells and their intratumoral function, resulting in tumor eradication. These CD4+ T cells had no antitumor effects in the absence of CD8+ T cells and recognized the alloantigen cross-presented on tumor stroma. CD4+ T cells might also be effective in cancer patients when PD-1/PD-L1 blockade does not rescue intratumoral CD8+ T-cell function and tumors persist. Cancer Immunol Res; 5(2); 127-36. ©2017 AACR.


Assuntos
Transferência Adotiva , Antígeno B7-H1/antagonistas & inibidores , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Resistencia a Medicamentos Antineoplásicos , Neoplasias/imunologia , Evasão Tumoral/imunologia , Animais , Antígenos de Neoplasias/imunologia , Antineoplásicos Imunológicos/farmacologia , Antineoplásicos Imunológicos/uso terapêutico , Linhagem Celular Tumoral , Terapia Combinada , Citocinas/biossíntese , Modelos Animais de Doenças , Antígenos de Histocompatibilidade Classe II/genética , Antígenos de Histocompatibilidade Classe II/imunologia , Contagem de Linfócitos , Camundongos , Camundongos Knockout , Neoplasias/patologia , Neoplasias/terapia , Carga Tumoral/genética , Carga Tumoral/imunologia
7.
Oncoimmunology ; 5(6): e1130207, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-27471609

RESUMO

We recently reported that therapeutic vaccination with live tumor antigen-producing Salmonella typhimurium rescues dysfunctional endogenous T cell responses and eradicates long-established tumors refractory to αCTLA-4 and αPD-L1 checkpoint inhibitor blockade. Here, we show that live intravenously injected or heat-killed (HK) intratumorally injected Salmonella typhimurium, even when not producing tumor antigen, synergize with adoptive T cell therapy to eradicate tumors. These data demonstrate that the combination of adoptive T cell transfer with the injection of live or dead Salmonella typhimurium is a promising approach for cancer treatment.

8.
Oncotarget ; 7(28): 43039-43051, 2016 Jul 12.
Artigo em Inglês | MEDLINE | ID: mdl-27343548

RESUMO

The majority of cancer patients respond poorly to either vaccine or checkpoint blockade, and even to the combination of both. They are often resistant to high doses of radiation therapy as well. We examined prognostic markers of immune cell infiltration in pancreatic cancer. Patients with low CD8+ T cell infiltration and high PD-L1 expression (CD8+ TloPD-L1hi) experienced poor outcomes. We developed a mouse tumor fragment model with a trackable model antigen (SIYRYYGL or SIY) to mimic CD8+ TloPD-L1hi cancers. Tumors arising from fragments contained few T cells, even after vaccination. Fragment tumors responded poorly to PD-L1 blockade, SIY vaccination or radiation individually. By contrast, local ionizing radiation coupled with vaccination increased CD8+ T cell infiltration that was associated with upregulation of CXCL10 and CCL5 chemokines in the tumor, but demonstrated modest inhibition of tumor growth. The addition of an anti-PD-L1 antibody enhanced the effector function of tumor-infiltrating T cells, leading to significantly improved tumor regression and increased survival compared to vaccination and radiation. These results indicate that sequential combination of radiation, vaccination and checkpoint blockade converts non-T cell-inflamed cancers to T cell-inflamed cancers, and mediates regression of established pancreatic tumors with an initial CD8+ TloPD-L1hi phenotype. This study has opened a new strategy for shifting "cold" to hot tumors that will respond to immunotherapy.


Assuntos
Imunoterapia/métodos , Neoplasias Pancreáticas/terapia , Radioterapia/métodos , Vacinação/métodos , Animais , Antígeno B7-H1/imunologia , Antígeno B7-H1/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Linhagem Celular Tumoral , Quimiocinas/genética , Quimiocinas/imunologia , Quimiocinas/metabolismo , Terapia Combinada , Regulação Neoplásica da Expressão Gênica/imunologia , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neoplasias Experimentais/genética , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/terapia , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/imunologia , Análise de Sobrevida
9.
Proc Natl Acad Sci U S A ; 113(27): 7551-6, 2016 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-27317748

RESUMO

Fibroblasts are common cell types in cancer stroma and lay down collagen required for survival and growth of cancer cells. Although some cancer therapy strategies target tumor fibroblasts, their origin remains controversial. Multiple publications suggest circulating mesenchymal precursors as a source of tumor-associated fibroblasts. However, we show by three independent approaches that tumor fibroblasts derive primarily from local, sessile precursors. First, transplantable tumors developing in a mouse expressing green fluorescent reporter protein (EGFP) under control of the type I collagen (Col-I) promoter (COL-EGFP) had green stroma, whereas we could not find COL-EGFP(+) cells in tumors developing in the parabiotic partner lacking the fluorescent reporter. Lack of incorporation of COL-EGFP(+) cells from the circulation into tumors was confirmed in parabiotic pairs of COL-EGFP mice and transgenic mice developing autochthonous intestinal adenomas. Second, transplantable tumors developing in chimeric mice reconstituted with bone marrow cells from COL-EGFP mice very rarely showed stromal fibroblasts expressing EGFP. Finally, cancer cells injected under full-thickness COL-EGFP skin grafts transplanted in nonreporter mice developed into tumors containing green stromal cells. Using multicolor in vivo confocal microscopy, we found that Col-I-expressing fibroblasts constituted approximately one-third of the stromal mass and formed a continuous sheet wrapping the tumor vessels. In summary, tumors form their fibroblastic stroma predominantly from precursors present in the local tumor microenvironment, whereas the contribution of bone marrow-derived circulating precursors is rare.


Assuntos
Fibroblastos Associados a Câncer/fisiologia , Neoplasias Experimentais/patologia , Actinas/metabolismo , Animais , Fibroblastos Associados a Câncer/patologia , Linhagem Celular Tumoral , Colágeno Tipo I/metabolismo , Proteínas de Fluorescência Verde , Camundongos Endogâmicos C57BL , Camundongos Knockout
10.
Semin Immunol ; 28(1): 54-63, 2016 02.
Artigo em Inglês | MEDLINE | ID: mdl-26872631

RESUMO

Immune response to tumors can be successfully oriented for therapeutic purposes, as shown by the clinical efficacy of checkpoint blockade in extending the survival of patients with certain solid and hematologic neoplasms. Nonetheless, numerous patients do not benefit from these new treatments. Tumor-specific CD8(+) T lymphocytes, either endogenously revived by checkpoint interference or adoptively transferred after in vitro expansion and retargeting, can be extremely efficient in controlling metastatic disease but have to overcome a number of restraints imposed by growing tumors. This immune escape relies on a profound modification of the tumor environment, which is rendered less permissive to lymphocyte arrival, persistence, and functional activity. We review here emerging findings on the main negative circuits limiting the efficacy of cancer immunotherapy, as well as novel and conventional approaches that can translate into rational combination therapies.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Quimiorradioterapia , Imunoterapia/métodos , Linfócitos do Interstício Tumoral/imunologia , Neoplasias/terapia , Microambiente Tumoral , Animais , Citotoxicidade Imunológica , Perfilação da Expressão Gênica , Humanos , Imunossupressão , Ativação Linfocitária , Neoplasias/imunologia , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/efeitos da radiação
11.
Clin Cancer Res ; 22(11): 2734-43, 2016 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-26667491

RESUMO

PURPOSE: Cancers usually contain multiple unique tumor-specific antigens produced by single amino acid substitutions (AAS) and encoded by somatic nonsynonymous single nucleotide substitutions. We determined whether adoptively transferred T cells can reject large, well-established solid tumors when engineered to express a single type of T-cell receptor (TCR) that is specific for a single AAS. EXPERIMENTAL DESIGN: By exome and RNA sequencing of an UV-induced tumor, we identified an AAS in p68 (mp68), a co-activator of p53. This AAS seemed to be an ideal tumor-specific neoepitope because it is encoded by a trunk mutation in the primary autochthonous cancer and binds with highest affinity to the MHC. A high-avidity mp68-specific TCR was used to genetically engineer T cells as well as to generate TCR-transgenic mice for adoptive therapy. RESULTS: When the neoepitope was expressed at high levels and by all cancer cells, their direct recognition sufficed to destroy intratumor vessels and eradicate large, long-established solid tumors. When the neoepitope was targeted as autochthonous antigen, T cells caused cancer regression followed by escape of antigen-negative variants. Escape could be thwarted by expressing the antigen at increased levels in all cancer cells or by combining T-cell therapy with local irradiation. Therapeutic efficacies of TCR-transduced and TCR-transgenic T cells were similar. CONCLUSIONS: Gene therapy with a single TCR targeting a single AAS can eradicate large established cancer, but a uniform expression and/or sufficient levels of the targeted neoepitope or additional therapy are required to overcome tumor escape. Clin Cancer Res; 22(11); 2734-43. ©2015 AACRSee related commentary by Liu, p. 2602.


Assuntos
Imunoterapia Adotiva , Neoplasias/terapia , Receptores de Antígenos de Linfócitos T/genética , eIF-2 Quinase/genética , Animais , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/imunologia , Antígenos de Neoplasias/metabolismo , Efeito Espectador , Linhagem Celular Tumoral , Apresentação Cruzada , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Epitopos de Linfócito T/metabolismo , Terapia Genética , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neoplasias/genética , Neoplasias/imunologia , Mutação Puntual , Evasão Tumoral , eIF-2 Quinase/imunologia , eIF-2 Quinase/metabolismo
12.
Curr Opin Immunol ; 33: 120-5, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25728992

RESUMO

Novel models of autochthonous tumorigenesis and adoptive T cell therapy (ATT) are providing new clues regarding the pro-tumorigenic and immunosuppressive effects of myeloid-derived suppressor cells (MDSC), and their interaction with T cells. New findings are shifting the perception of the main level at which MDSC act, from direct cell-to-cell suppression to others, such as limiting T cell infiltration. Adoptively transferred, high-avidity T cells recognizing peptides with high-affinity for MHC-I eliminated large tumors. However, low-avidity T cells or low-affinity peptides resulted in failure to eradicate tumors. Manipulation of intratumoral myeloid cells improved the outcome of otherwise unsuccessful ATT. Therefore, therapeutic intervention directed at the tumor stroma might be required when using suboptimal T cells for ATT.


Assuntos
Transferência Adotiva , Comunicação Celular/imunologia , Células Mieloides/imunologia , Células Mieloides/metabolismo , Linfócitos T/imunologia , Linfócitos T/metabolismo , Animais , Humanos , Imunomodulação , Imunoterapia Adotiva/métodos , Neoplasias/imunologia , Neoplasias/metabolismo , Neoplasias/terapia , Resultado do Tratamento , Microambiente Tumoral
13.
Immunity ; 41(5): 843-52, 2014 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-25517616

RESUMO

Ionizing radiation-mediated tumor regression depends on type I interferon (IFN) and the adaptive immune response, but several pathways control I IFN induction. Here, we demonstrate that adaptor protein STING, but not MyD88, is required for type I IFN-dependent antitumor effects of radiation. In dendritic cells (DCs), STING was required for IFN-? induction in response to irradiated-tumor cells. The cytosolic DNA sensor cyclic GMP-AMP (cGAMP) synthase (cGAS) mediated sensing of irradiated-tumor cells in DCs. Moreover, STING was essential for radiation-induced adaptive immune responses, which relied on type I IFN signaling on DCs. Exogenous IFN-? treatment rescued the cross-priming by cGAS or STING-deficient DCs. Accordingly, activation of STING by a second messenger cGAMP administration enhanced antitumor immunity induced by radiation. Thus radiation-mediated antitumor immunity in immunogenic tumors requires a functional cytosolic DNA-sensing pathway and suggests that cGAMP treatment might provide a new strategy to improve radiotherapy.


Assuntos
DNA/imunologia , Proteínas de Membrana/genética , Neoplasias/radioterapia , Nucleotidiltransferases/imunologia , Imunidade Adaptativa , Proteínas Adaptadoras de Transporte Vesicular/genética , Animais , Antineoplásicos/farmacologia , Células Cultivadas , Apresentação Cruzada/imunologia , Células Dendríticas/imunologia , Imunidade Inata , Interferon beta/biossíntese , Interferon beta/imunologia , Interferon beta/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator 88 de Diferenciação Mieloide/genética , Neoplasias/imunologia , Nucleotídeos Cíclicos/farmacologia , Interferência de RNA , RNA Interferente Pequeno , Radiação Ionizante , Receptor de Interferon alfa e beta/genética , Receptor de Interferon alfa e beta/imunologia , Transdução de Sinais/imunologia , Xantonas/farmacologia
14.
Oncoimmunology ; 3: e28464, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25050213

RESUMO

The expansion of cancer-induced myeloid cells is thought to be one of the main obstacles to successful immunotherapy. Nevertheless, in murine tumors undergoing immune-mediated destruction by adoptively transferred T cells, we have recently shown that such cells maintain their immunosuppressive properties. Therefore, adoptive T-cell therapy can, under certain conditions, overcome myeloid cell immunosuppression.

15.
J Immunol ; 192(3): 1286-93, 2014 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-24367029

RESUMO

Myeloid-derived CD11b(+)Gr1(+) suppressor cells (MDSCs) and tumor-associated macrophages (TAMs) are considered a major obstacle for effective adoptive T cell therapy. Myeloid cells suppress naive T cell proliferation ex vivo and can prevent the generation of T cell responses in vivo. We find, however, that adoptively transferred immune T cells eradicate well-established tumors in the presence of MDSCs and TAMs, which are strongly immunosuppressive ex vivo. These MDSCs and TAMs were comparable in numbers and immunosuppressive capacity among different tumor models. Longitudinal microscopy of tumors in vivo revealed that after T cell transfer, tumor vasculature and cancer cells disappeared simultaneously. During T cell-mediated tumor destruction, the tumor stroma contained abundant myeloid cells (mainly TAMs) that retained their suppressive properties. Preimmunized but not naive mice resisted immune suppression caused by an unrelated tumor burden, supporting the idea that in vivo, myeloid immunosuppressive cells can suppress naive but not memory T cell responses.


Assuntos
Imunoterapia Adotiva , Macrófagos/imunologia , Células Mieloides/imunologia , Neoplasias Experimentais/terapia , Subpopulações de Linfócitos T/transplante , Evasão Tumoral/imunologia , Animais , Antígeno CD11b/análise , Vacinas Anticâncer/imunologia , Proteínas de Ligação a DNA/deficiência , Proteínas de Homeodomínio/genética , Imunização , Memória Imunológica , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Modelos Biológicos , Neoplasias Experimentais/irrigação sanguínea , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/patologia , Neovascularização Patológica/imunologia , Neovascularização Patológica/terapia , Receptores de Quimiocinas/análise , Técnica de Janela Cutânea , Subpopulações de Linfócitos T/imunologia , Carga Tumoral
16.
Oncoimmunology ; 2(11): e26677, 2013 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-24482750

RESUMO

A fluorescence-based, high-resolution imaging approach was used to visualize longitudinally the cellular events unfolding during T cell-mediated tumor destruction. The dynamic interplay of T cells, cancer cells, cancer antigen loss variants, and stromal cells-all color-coded in vivo-was analyzed in established, solid tumors that had developed behind windows implanted on the backs of mice. Events could be followed repeatedly within precisely the same tumor region-before, during and after adoptive T cell therapy-thereby enabling for the first time a longitudinal in vivo evaluation of protracted events, an analysis not possible with terminal imaging of surgically exposed tumors. T cell infiltration, stromal interactions, and vessel destruction, as well as the functional consequences thereof, including the elimination of cancer cells and cancer cell variants were studied. Minimal perivascular T cell infiltrates initiated vascular destruction inside the tumor mass eventually leading to macroscopic central tumor necrosis. Prolonged engagement of T cells with tumor antigen-crosspresenting stromal cells correlated with high IFNγ cytokine release and bystander elimination of antigen-negative cancer cells. The high-resolution, longitudinal, in vivo imaging approach described here will help to further a better mechanistic understanding of tumor eradication by T cells and other anti-cancer therapies.

17.
Cancer Immunol Res ; 1(2): 123-33, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24455752

RESUMO

Immunogenic tumors grow progressively even when heavily infiltrated by CD8(+) T cells. We investigated how to rescue CD8(+) T cell function in long-established immunogenic melanomas that contained a high percentage of endogenous PD-1(+) tumor-specific CD8(+) T cells that were dysfunctional. Treatment with αPD-L1 and αCTLA-4 blocking antibodies did not prevent tumors from progressing rapidly. We then tested exogenous tumor-specific antigen delivery into tumors using Salmonella Typhimurium A1-R to increase antigen levels and generate a proinflammatory tumor microenvironment. Antigen-producing A1-R rescued the endogenous tumor-specific CD8(+) T cell response: proliferation was induced in the lymphoid organs and effector function was recovered in the tumor. Treatment with antigen-producing A1-R led to improved mouse survival and resulted in 32% rejection of long-established immunogenic melanomas. Following treatment with antigen-producing A1-R, the majority of tumor-specific CD8(+) T cells still expressed a high level of PD-1 in the tumor. Combining antigen-producing A1-R with αPD-L1 blocking antibody enhanced the expansion of tumor-specific CD8(+) T cells and resulted in 80% tumor rejection. Collectively, these data demonstrate a powerful new therapeutic approach to rescue dysfunctional endogenous tumor-specific CD8(+) T cells and eradicate advanced immunogenic tumors.


Assuntos
Antígenos de Neoplasias/imunologia , Vacinas Bacterianas/imunologia , Linfócitos T CD8-Positivos/imunologia , Vacinas Anticâncer/imunologia , Melanoma Experimental/imunologia , Melanoma Experimental/terapia , Receptor de Morte Celular Programada 1/imunologia , Animais , Apresentação do Antígeno , Antígenos de Neoplasias/genética , Vacinas Bacterianas/genética , Vacinas Bacterianas/farmacologia , Vacinas Anticâncer/genética , Vacinas Anticâncer/farmacologia , Epitopos , Feminino , Ativação Linfocitária , Linfócitos do Interstício Tumoral/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Ovalbumina/imunologia , Ovalbumina/farmacologia , Fragmentos de Peptídeos/imunologia , Fragmentos de Peptídeos/farmacologia , Receptor de Morte Celular Programada 1/genética
18.
Cancer Res ; 72(8): 1964-74, 2012 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-22374983

RESUMO

Natural killer (NK) cells inhibit early stages of tumor formation, recurrence, and metastasis. Here, we show that NK cells can also eradicate large solid tumors. Eradication depended on the massive infiltration of proliferating NK cells due to interleukin 15 (IL-15) released and presented by the cancer cells in the tumor microenvironment. Infiltrating NK cells had the striking morphologic feature of being densely loaded with periodic acid-Schiff-positive, diastase-resistant granules, resembling uterine NK cells. Perforin-mediated killing by these densely granulated NK cells was essential for tumor eradication. Expression of the IL-15 receptor α on cancer cells was needed to efficiently induce granulated NK cells, and expression on host stromal cells was essential to prevent tumor relapse after near complete destruction. These results indicate that IL-15 released at the cancer site induces highly activated NK cells that lead to eradication of large solid tumors.


Assuntos
Interleucina-15/imunologia , Células Matadoras Naturais/imunologia , Ativação Linfocitária/imunologia , Neoplasias Experimentais/imunologia , Microambiente Tumoral/imunologia , Animais , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Imuno-Histoquímica , Corpos de Inclusão , Interleucina-15/metabolismo , Células Matadoras Naturais/metabolismo , Camundongos , Camundongos Knockout , Neoplasias Experimentais/metabolismo
19.
Clin Cancer Res ; 18(9): 2526-33, 2012 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-22415314

RESUMO

PURPOSE: Solid tumors that have grown two weeks or longer in mice and have diameters larger than 1 cm are histologically indistinguishable from autochthonous human cancers. When experimental tumors reach this clinically relevant size, they are usually refractory to most immunotherapies but may be destroyed by adoptive T-cell transfer. However, TCR-transgenic T cells and/or tumor cells overexpressing antigens are frequently used in these experiments. Here we studied the requirements for destroying clinical size, unmanipulated 8101 tumors by adoptive cell therapy. EXPERIMENTAL DESIGN: 8101 arose in an old mouse after chronic exposure to UV light. A cancer line was established, which was never serially transplanted. The immunodominant CD8(+) T cell-recognized antigen of this tumor is caused by a somatic tumor-specific mutation in the RNA helicase p68. 8101 tumors were treated with spleen cells from young naive, or young and old immunized mice to ascertain the characteristics of immune cells that lead to rejection. RESULTS: Here we show that the mutant p68 peptide has an exceptionally high affinity to the presenting MHC class I molecule K(b) and that spleen cells from immunized young syngeneic mice adoptively transferred to Rag(-/-) or cancer-suppressed euthymic mice eradicate 8101 tumors larger than 1 cm in average diameter and established for several weeks. Spleen cells from naive young mice or from old and boosted (reimmunized) mice were ineffective. CONCLUSIONS: Relapse-free destruction of large and long-established tumors expressing a genuine very high-affinity tumor-specific antigen can be achieved by using adoptive transfer of lymphocytes from immunized young individuals.


Assuntos
Memória Imunológica , Imunoterapia Adotiva , Neoplasias Experimentais/etiologia , Neoplasias Experimentais/prevenção & controle , Baço/imunologia , Linfócitos T Citotóxicos/imunologia , Linfócitos T/imunologia , Animais , DNA de Neoplasias/genética , Citometria de Fluxo , Humanos , Imunização , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neoplasias Experimentais/patologia , Reação em Cadeia da Polimerase
20.
PLoS One ; 7(12): e52370, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-23285013

RESUMO

Apolipoprotein A-I (Apo A-I) is a major component of high density lipoproteins (HDL) that transport cholesterol in circulation. We have constructed an expression plasmid encoding a chimeric molecule encompassing interleukin-15 (IL-15) and Apo A-I (pApo-hIL15) that was tested by hydrodynamic injections into mice and was co-administered with a plasmid encoding the sushi domain of IL-15Rα (pSushi) in order to enhance IL-15 trans-presentation and thereby bioactivity. The pharmacokinetics of the Apo A-I chimeric protein were much longer than non-stabilized IL-15 and its bioactivity was enhanced in combination with IL-15Rα Sushi. Importantly, the APO-IL-15 fusion protein was incorporated in part into circulating HDL. Liver gene transfer of these constructs increased NK and memory-phenotype CD8 lymphocyte numbers in peripheral blood, spleen and liver as a result of proliferation documented by CFSE dilution and BrdU incorporation. Moreover, the gene transfer procedure partly rescued the NK and memory T-cell deficiency observed in IL-15Rα(-/-) mice. pApo-hIL15+ pSushi gene transfer to the liver showed a modest therapeutic activity against subcutaneously transplanted MC38 colon carcinoma tumors, that was more evident when tumors were set up as liver metastases. The improved pharmacokinetic profile and the strong biological activity of APO-IL-15 fusion protein holds promise for further development in combination with other immunotherapies.


Assuntos
Técnicas de Transferência de Genes , Terapia Genética , Imunoterapia , Interleucina-15/genética , Interleucina-15/uso terapêutico , Lipoproteínas HDL/sangue , Fígado/metabolismo , Animais , Apolipoproteína A-I/sangue , Western Blotting , Linfócitos T CD8-Positivos/imunologia , Contagem de Células , Linhagem Celular Tumoral , Proliferação de Células , Humanos , Memória Imunológica , Injeções Subcutâneas , Interleucina-15/sangue , Subunidade alfa de Receptor de Interleucina-15/química , Subunidade alfa de Receptor de Interleucina-15/metabolismo , Células Matadoras Naturais/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Metástase Neoplásica , Fenótipo , Estrutura Terciária de Proteína , Proteínas Recombinantes/genética , Proteínas Recombinantes/uso terapêutico , Baço/metabolismo
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