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1.
Cell ; 179(4): 829-845.e20, 2019 Oct 31.
Artigo em Inglês | MEDLINE | ID: mdl-31675496

RESUMO

The immune microenvironment of hepatocellular carcinoma (HCC) is poorly characterized. Combining two single-cell RNA sequencing technologies, we produced transcriptomes of CD45+ immune cells for HCC patients from five immune-relevant sites: tumor, adjacent liver, hepatic lymph node (LN), blood, and ascites. A cluster of LAMP3+ dendritic cells (DCs) appeared to be the mature form of conventional DCs and possessed the potential to migrate from tumors to LNs. LAMP3+ DCs also expressed diverse immune-relevant ligands and exhibited potential to regulate multiple subtypes of lymphocytes. Of the macrophages in tumors that exhibited distinct transcriptional states, tumor-associated macrophages (TAMs) were associated with poor prognosis, and we established the inflammatory role of SLC40A1 and GPNMB in these cells. Further, myeloid and lymphoid cells in ascites were predominantly linked to tumor and blood origins, respectively. The dynamic properties of diverse CD45+ cell types revealed by this study add new dimensions to the immune landscape of HCC.


Assuntos
Carcinoma Hepatocelular/imunologia , Proteínas de Transporte de Cátions/genética , Inflamação/imunologia , Neoplasias Hepáticas/imunologia , Glicoproteínas de Membrana/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Comunicação Celular/genética , Comunicação Celular/imunologia , Linhagem da Célula/genética , Linhagem da Célula/imunologia , Células Dendríticas/imunologia , Células Dendríticas/patologia , Regulação Neoplásica da Expressão Gênica , Humanos , Inflamação/genética , Inflamação/patologia , Antígenos Comuns de Leucócito/imunologia , Fígado/imunologia , Fígado/patologia , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Linfonodos/imunologia , Linfonodos/patologia , Linfócitos/imunologia , Linfócitos/patologia , Proteínas de Membrana Lisossomal/genética , Macrófagos/imunologia , Macrófagos/patologia , Células Mieloides/imunologia , Células Mieloides/patologia , Proteínas de Neoplasias/genética , Análise de Sequência de RNA , Análise de Célula Única , Transcriptoma/genética , Transcriptoma/imunologia , Microambiente Tumoral/genética , Microambiente Tumoral/imunologia
2.
J Mol Diagn ; 21(5): 796-807, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31255795

RESUMO

Clinical biomarker studies are often hindered by the scarcity or suboptimal quality of biological specimens. EdgeSeq, a transcriptomics analysis platform, combines quantitative nuclease protection assay technology with next-generation sequencing, using small amounts of starting material and delivering reproducible gene expression profiles from challenging material, such as formalin-fixed, paraffin-embedded (FFPE) tissue. To evaluate EdgeSeq for analysis of archives of stained FFPE tissue, EdgeSeq was performed on unstained, hematoxylin and eosin (H&E)-stained, and immunohistochemistry-stained slides from patients with small-cell and non-small-cell lung cancer. Pairwise comparisons of gene expression profiles from stained and unstained slides showed higher Pearson correlation coefficients with H&E staining (0.86 to 0.97) than with immunohistochemistry staining (0.21 to 0.56). A 25-gene interferon-γ signature score from unstained slides showed a Pearson correlation coefficient of 0.92 with H&E-stained slides and a significant Spearman correlation (P = 0.0025) with immune scores. To test gene expression profiling in small samples, FFPE sample equivalents were examined from 5.0 to 0.08 mm2 of a section (5 µm thick); sample equivalents ≥0.31 mm2 showed alignment rates >69% and pairwise Pearson correlation coefficients ≥0.87. EdgeSeq can, thus, be used to profile small and H&E-stained FFPE tumor specimens to obtain biomarker data from limited tissue in oncology clinical trials and enable research into tumor microenvironment and immune cell engagement with tumors at the locoregional level.


Assuntos
Biomarcadores Tumorais/genética , Neoplasias Colorretais/genética , Perfilação da Expressão Gênica/métodos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Neoplasias Pulmonares/genética , Inclusão em Parafina/métodos , Carcinoma de Pequenas Células do Pulmão/genética , Neoplasias Colorretais/diagnóstico , Amarelo de Eosina-(YS)/química , Formaldeído/química , Biblioteca Gênica , Hematoxilina/química , Humanos , Neoplasias Pulmonares/diagnóstico , Carcinoma de Pequenas Células do Pulmão/diagnóstico , Fixação de Tecidos
3.
J Immunother ; 39(4): 181-7, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-27070449

RESUMO

Two cancer testis antigens, the New York esophageal squamous cell carcinoma-1 (NY-ESO-1) and the melanoma-antigen family A (MAGE-A), represent promising immunotherapy targets due to the low expression of these antigens in nonmalignant tissue. To assess overexpression patterns in various cancers, we performed a systematic immunohistochemical analysis for NY-ESO-1 and MAGE-A on tissue array samples of 3668 common epithelial carcinomas (CA) and germ cell tumors of high prevalence and mortality. Here, we find significantly higher expression of MAGE-A (>50% on tumor cells) compared with NY-ESO-1 in several CAs including cutaneous squamous cell carcinomas (SCC) (52.8%/2.8%), esophageal SCC (50%/0%), head and neck SCC (41.1%/<1%), bladder urothelial CA (40.4%/8.3%), cervical/anal SCC (37.5%/0%), lung SCC (34%/3.8%), lung adenocarcinomas (27.6%/3.9%), ovarian CA (26.4%/3.6%), endometrial CA (26.3%/1.3%), lung small cell CA (24.4%/2.4%), gastric adenocarcinomas (20%/4%), breast mucinous CA (19.3%/0%), hepatocellular CA (18.8%/1.2%), breast infiltrating ductal CA (16.4%/1.8%), colorectal adenocarcinomas (10.7%/<1%), cholangiocarcinomas (9.8%/0%), thymic CA (9%/4.5%), and mesotheliomas (7.9%/<1%). Furthermore, high expression of MAGE-A, but not NY-ESO-1, was seen in whole slide evaluations of an independent cohort of metastatic SCC (45.5%/3.6%) and metastatic CA (13.5%/0%) of various primaries with significantly higher expression of MAGE-A in metastatic SCC compared with other metastatic CA. MAGE-A is also more highly expressed in germ cell tumors, seminomas (69%/3.5%) and nonseminomas (40.1%/4.7%). In summary, MAGE-A is more highly expressed than NY-ESO-1 in a majority of human malignancies, and targeting MAGE-A may benefit a large number of patients.


Assuntos
Antígenos de Neoplasias/metabolismo , Carcinoma de Células Escamosas/metabolismo , Neoplasias Esofágicas/metabolismo , Neoplasias de Cabeça e Pescoço/metabolismo , Antígenos Específicos de Melanoma/metabolismo , Proteínas de Membrana/metabolismo , Neoplasias Embrionárias de Células Germinativas/metabolismo , Neoplasias Cutâneas/metabolismo , Carcinoma de Células Escamosas/patologia , Estudos de Coortes , Neoplasias Esofágicas/patologia , Feminino , Neoplasias de Cabeça e Pescoço/patologia , Humanos , Imuno-Histoquímica , Masculino , Metástase Neoplásica , Neoplasias Embrionárias de Células Germinativas/patologia , Neoplasias Cutâneas/patologia , Análise Serial de Tecidos
4.
J Immunother ; 39(1): 1-7, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26641256

RESUMO

Melanoma-associated antigen-A (MAGE-A) and New York esophageal squamous cell cancer-1 (NY-ESO-1) are 2 cancer testis antigens (CTA) demonstrating potential for use in targeted immunotherapy. Clinical trials in melanoma and synovial sarcomas targeting these antigens in immune-based therapies have demonstrated durable tumor regression. Although protein expression of NY-ESO-1 has been assessed in a variety of cancer types, the expression of MAGE-A has not been studied in depth. In this study we analyzed MAGE-A and NY-ESO-1 expression in 314 melanoma specimens from 301 melanoma patients, 38 patients with squamous cell cancers and 111 patients with adenocarcinomas. Our results demonstrated higher expression of MAGE-A compared with NY-ESO-1 in melanomas (32% vs. 13%) and squamous cell carcinomas (45% vs. 7.9%), and higher expression of both CTAs in metastatic versus primary tumors. CTA expression in adenocarcinomas was low (MAGE-A: 10%, NY-ESO-1: 0.9%). In addition, we looked at concordance of expression among metastatic melanoma lesions within the same patient and found concordant expression in 38 of 47 patients for MAGE-A and 43 of 47 patients for NY-ESO-1. Our study demonstrated that the MAGE-A family may be of greater utility than NY-ESO-1 for targeted immunotherapy in a variety of cancer histologies, in particular metastatic melanomas and squamous cell carcinomas.


Assuntos
Antígenos de Neoplasias/metabolismo , Proteínas de Membrana/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/imunologia , Biomarcadores Tumorais , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patologia , Feminino , Expressão Gênica , Humanos , Imuno-Histoquímica , Masculino , Melanoma/genética , Melanoma/metabolismo , Melanoma/patologia , Proteínas de Membrana/genética , Proteínas de Membrana/imunologia , Metástase Neoplásica , Estadiamento de Neoplasias , Neoplasias/genética
5.
Clin Cancer Res ; 21(19): 4431-9, 2015 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-26429982

RESUMO

PURPOSE: The E6 and E7 oncoproteins of HPV-associated epithelial cancers are in principle ideal immunotherapeutic targets, but evidence that T cells specific for these antigens can recognize and kill HPV(+) tumor cells is limited. We sought to determine whether TCR gene engineered T cells directed against an HPV oncoprotein can successfully target HPV(+) tumor cells. EXPERIMENTAL DESIGN: T-cell responses against the HPV-16 oncoproteins were investigated in a patient with an ongoing 22-month disease-free interval after her second resection of distant metastatic anal cancer. T cells genetically engineered to express an oncoprotein-specific TCR from this patient's tumor-infiltrating T cells were tested for specific reactivity against HPV(+) epithelial tumor cells. RESULTS: We identified, from an excised metastatic anal cancer tumor, T cells that recognized an HLA-A*02:01-restricted epitope of HPV-16 E6. The frequency of the dominant T-cell clonotype from these cells was approximately 400-fold greater in the patient's tumor than in her peripheral blood. T cells genetically engineered to express the TCR from this clonotype displayed high avidity for an HLA-A*02:01-restricted epitope of HPV-16, and they showed specific recognition and killing of HPV-16(+) cervical, and head and neck cancer cell lines. CONCLUSIONS: These findings demonstrate that HPV-16(+) tumors can be targeted by E6-specific TCR gene engineered T cells, and they provide the foundation for a novel cellular therapy directed against HPV-16(+) malignancies, including cervical, oropharyngeal, anal, vulvar, vaginal, and penile cancers.


Assuntos
Carcinoma/etiologia , Genes Codificadores dos Receptores de Linfócitos T , Papillomavirus Humano 16/imunologia , Proteínas Oncogênicas Virais/imunologia , Proteínas Repressoras/imunologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Neoplasias do Ânus/etiologia , Neoplasias do Ânus/patologia , Carcinoma/metabolismo , Carcinoma/terapia , Linhagem Celular Tumoral , Reações Cruzadas/imunologia , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Feminino , Expressão Gênica , Antígeno HLA-A2/imunologia , Antígeno HLA-A2/metabolismo , Papillomavirus Humano 16/genética , Humanos , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Linfócitos do Interstício Tumoral/patologia , Metástase Neoplásica , Proteínas Oncogênicas Virais/genética , Proteínas Oncogênicas Virais/metabolismo , Ligação Proteica/imunologia , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Especificidade do Receptor de Antígeno de Linfócitos T/imunologia , Transdução Genética , Neoplasias do Colo do Útero/etiologia
6.
J Surg Case Rep ; 2015(7)2015 Jul 14.
Artigo em Inglês | MEDLINE | ID: mdl-26173437

RESUMO

A 61-year-old female with a past medical history significant for von Hippel-Lindau (VHL) syndrome presented with multiple bilateral pulmonary lesions found on surveillance computed tomography scan. Positron emission tomography demonstrated avidity in a lesion in the right upper lobe. After an equivocal biopsy, a lobectomy via a thoracoscopic approach was performed as this lesion was concerning for a primary lung cancer. Pathology revealed a diagnosis of pulmonary mucosa-associated lymphoid tissue (MALT) lymphoma. To our knowledge, this is the first reported case of a pulmonary MALT lymphoma in a patient with VHL.

7.
Semin Cancer Biol ; 35 Suppl: S199-S223, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25865775

RESUMO

Cancer arises in the context of an in vivo tumor microenvironment. This microenvironment is both a cause and consequence of tumorigenesis. Tumor and host cells co-evolve dynamically through indirect and direct cellular interactions, eliciting multiscale effects on many biological programs, including cellular proliferation, growth, and metabolism, as well as angiogenesis and hypoxia and innate and adaptive immunity. Here we highlight specific biological processes that could be exploited as targets for the prevention and therapy of cancer. Specifically, we describe how inhibition of targets such as cholesterol synthesis and metabolites, reactive oxygen species and hypoxia, macrophage activation and conversion, indoleamine 2,3-dioxygenase regulation of dendritic cells, vascular endothelial growth factor regulation of angiogenesis, fibrosis inhibition, endoglin, and Janus kinase signaling emerge as examples of important potential nexuses in the regulation of tumorigenesis and the tumor microenvironment that can be targeted. We have also identified therapeutic agents as approaches, in particular natural products such as berberine, resveratrol, onionin A, epigallocatechin gallate, genistein, curcumin, naringenin, desoxyrhapontigenin, piperine, and zerumbone, that may warrant further investigation to target the tumor microenvironment for the treatment and/or prevention of cancer.


Assuntos
Carcinogênese/efeitos dos fármacos , Neoplasias/tratamento farmacológico , Neovascularização Patológica/tratamento farmacológico , Microambiente Tumoral/genética , Antineoplásicos/uso terapêutico , Carcinogênese/genética , Proliferação de Células/efeitos dos fármacos , Humanos , Terapia de Alvo Molecular , Neoplasias/genética , Neoplasias/prevenção & controle , Neovascularização Patológica/genética , Neovascularização Patológica/prevenção & controle , Transdução de Sinais , Microambiente Tumoral/efeitos dos fármacos
8.
Clin Cancer Res ; 21(10): 2278-88, 2015 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-25695689

RESUMO

PURPOSE: Infusion of interleukin-12 (IL12) can mediate antitumor immunity in animal models, yet its systemic administration to patients with cancer results in minimal efficacy and severe toxicity. Here, we evaluated the antitumor activity of adoptively transferred human tumor-infiltrating lymphocytes (TILs) genetically engineered to secrete single-chain IL12 selectively at the tumor site. EXPERIMENTAL DESIGN: Thirty-three patients with metastatic melanoma were treated in a cell dose-escalation trial of autologous TILs transduced with a gene encoding a single-chain IL12 driven by a nuclear factor of the activated T cells promoter (NFAT.IL12). No IL2 was administered. RESULTS: The administration of 0.001 to 0.1 × 10(9) NFAT.IL12-transduced TILs to 17 patients resulted in a single, objective response (5.9%). However, at doses between 0.3 and 3 × 10(9) cells, 10 of 16 patients (63%) exhibited objective clinical responses. The responses tended to be short, and the administered IL12-producing cells rarely persisted at 1 month. Increasing cell doses were associated with high serum levels of IL12 and IFNγ as well as clinical toxicities, including liver dysfunction, high fevers, and sporadic life-threatening hemodynamic instability. CONCLUSIONS: In this first-in-man trial, administration of TILs transduced with an inducible IL12 gene mediated tumor responses in the absence of IL2 administration using cell doses 10- to 100-fold lower than conventional TILs. However, due to toxicities, likely attributable to the secreted IL12, further refinement will be necessary before this approach can be safely used in the treatment of cancer patients.


Assuntos
Interleucina-12/genética , Linfócitos do Interstício Tumoral/fisiologia , Melanoma/terapia , Neoplasias Cutâneas/terapia , Adulto , Idoso , Células Cultivadas , Feminino , Engenharia Genética , Humanos , Imunoterapia , Interleucina-12/biossíntese , Linfócitos do Interstício Tumoral/transplante , Masculino , Melanoma/imunologia , Melanoma/secundário , Pessoa de Meia-Idade , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/patologia , Ativação Transcricional , Resultado do Tratamento , Adulto Jovem
9.
Cancer Res ; 75(2): 296-305, 2015 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-25432172

RESUMO

Adoptive cell therapy (ACT) using autologous tumor-infiltrating lymphocytes (TIL) results in complete regression of advanced cancer in some patients, but the efficacy of this potentially curative therapy may be limited by poor persistence of TIL after adoptive transfer. Pharmacologic inhibition of the serine/threonine kinase Akt has recently been shown to promote immunologic memory in virus-specific murine models, but whether this approach enhances features of memory (e.g., long-term persistence) in TIL that are characteristically exhausted and senescent is not established. Here, we show that pharmacologic inhibition of Akt enables expansion of TIL with the transcriptional, metabolic, and functional properties characteristic of memory T cells. Consequently, Akt inhibition results in enhanced persistence of TIL after adoptive transfer into an immunodeficient animal model and augments antitumor immunity of CD8 T cells in a mouse model of cell-based immunotherapy. Pharmacologic inhibition of Akt represents a novel immunometabolomic approach to enhance the persistence of antitumor T cells and improve the efficacy of cell-based immunotherapy for metastatic cancer.


Assuntos
Imunoterapia Adotiva/métodos , Linfócitos do Interstício Tumoral/imunologia , Melanoma Experimental/terapia , Melanoma/terapia , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Animais , Humanos , Memória Imunológica , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Linfócitos do Interstício Tumoral/patologia , Melanoma/imunologia , Melanoma Experimental/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos Transgênicos , Proteínas Proto-Oncogênicas c-akt/imunologia , Distribuição Aleatória , Células Tumorais Cultivadas
10.
Int J Surg Case Rep ; 5(10): 677-80, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25194603

RESUMO

INTRODUCTION: Pythiosis is a serious life- and limb-threatening infection endemic to Thailand, but rarely seen in the Western hemisphere. Here, we present a unique case of vascular pythiosis initially managed with limb-sparing vascular bypass grafts complicated by a pseudoaneurysm in our repair. PRESENTATION OF CASE: The patient is a 17 year-old Jamaican male with severe aplastic anemia. He sustained a minor injury to his left leg while fishing in Jamaica, which evolved to become an exquisitely tender inguinal swelling. His physical exam and imaging were significant for arteriovenous fistula with limb ischemia. Pathology obtained during surgery for an extra-anatomic vascular bypass showed extensive invasion by Pythium insidiosum. He later developed a pseudoaneurysm at the site of proximal anastomosis and required urgent intervention. DISCUSSION: This patient presented with a rare, but classic case of vascular pythiosis, which was unrecognized at the time of presentation. A variety of therapeutic modalities have been used to treat this disease, including antibiotics, antifungals, and immunotherapy, but the ultimate management of vascular pythiosis is surgical source control. CONCLUSION: A high index of suspicion in susceptible patients is needed for timely diagnosis of vascular pythiosis to achieve optimal source control.

11.
Immunology ; 143(1): 96-108, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24678989

RESUMO

A growing body of evidence suggests that inflammatory cytokines have a dualistic role in immunity. In this study, we sought to determine the direct effects of interferon-γ (IFN-γ) on the differentiation and maturation of human peripheral blood monocyte-derived dendritic cells (moDC). Here, we report that following differentiation of monocytes into moDC with granulocyte-macrophage colony-stimulating factor and interleukin-4, IFN-γ induces moDC maturation and up-regulates the co-stimulatory markers CD80/CD86/CD95 and MHC Class I, enabling moDC to effectively generate antigen-specific CD4(+) and CD8(+) T-cell responses for multiple viral and tumour antigens. Early exposure of monocytes to high concentrations of IFN-γ during differentiation promotes the formation of macrophages. However, under low concentrations of IFN-γ, monocytes continue to differentiate into dendritic cells possessing a unique gene-expression profile, resulting in impairments in subsequent maturation by IFN-γ or lipopolysaccharide and an inability to generate effective antigen-specific CD4(+) and CD8(+) T-cell responses. These findings demonstrate that IFN-γ imparts differential programmes on moDC that shape the antigen-specific T-cell responses they induce. Timing and intensity of exposure to IFN-γ can therefore determine the functional capacity of moDC.


Assuntos
Diferenciação Celular/imunologia , Células Dendríticas/imunologia , Interferon gama/imunologia , Monócitos/citologia , Monócitos/imunologia , Linhagem Celular , Citocinas/biossíntese , Células Dendríticas/citologia , Citometria de Fluxo , Humanos , Transcriptoma
12.
Front Immunol ; 4: 304, 2013 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-24098300

RESUMO

T lymphocytes first carried foreign genes safely into humans over two decades ago. Since these pioneering studies, scientific techniques to better understand the genomic landscape of cells has directly led to a more sophisticated appreciation of the diversity, functional complexity, and therapeutic potential of T cells. Through the use of mouse models, we now know the function of the many genes that are critical for T cells to recognize foreign, mutated, or self-antigens and the factors responsible for the lineage diversification of T cells that lead to inhibitory or stimulatory immune responses. This knowledge combined with well-established modalities to introduce genes into T cells allows for the design of effector and memory CD8 and CD4 T lymphocytes specific for viral, fungal, bacterial, parasitic, and tumor-antigens and to design regulatory lymphocytes specific for the self-antigens responsible for autoimmune and inflammatory diseases. Here, I review strategies for designing the ideal T cell by introducing genes controlling (1) the secretion of cytokines/chemokines and their receptors, (2) T-cell receptor specificity, (3) chimeric-antigen receptors that enable for the recognition of surface antigens in an MHC-independent fashion, (4) co-stimulatory/inhibitory surface molecules, and (5) disease defining single-gene factors.

13.
Mol Ther ; 21(7): 1369-77, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23568260

RESUMO

Engineering CD8⁺ T cells to deliver interleukin 12 (IL-12) to the tumor site can lead to striking improvements in the ability of adoptively transferred T cells to induce the regression of established murine cancers. We have recently shown that IL-12 triggers an acute inflammatory environment that reverses dysfunctional antigen presentation by myeloid-derived cells within tumors and leads to an increase in the infiltration of adoptively transferred antigen-specific CD8⁺ T cells. Here, we find that local delivery of IL-12 increased the expression of Fas within tumor-infiltrating macrophages, dendritic cells, and myeloid-derived suppressor cells (MDSC), and that these changes were abrogated in mice deficient in IL-12-receptor signaling. Importantly, upregulation of Fas in host mice played a critical role in the proliferation and antitumor activity of adoptively transferred IL-12-modified CD8⁺ T cells. We also observed higher percentages of myeloid-derived cell populations within tumors in Fas-deficient mice, indicating that tumor stromal destruction was dependent on the Fas death receptor. Taken together, these results describe the likely requirement for costimulatory reverse signaling through Fasl on T cells that successfully infiltrate tumors, a mechanism triggered by the induction of Fas expression on myeloid-derived cells by IL-12 and the subsequent collapse of the tumor stroma.


Assuntos
Interleucina-12/uso terapêutico , Melanoma Experimental/metabolismo , Receptor fas/metabolismo , Animais , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Células Dendríticas/metabolismo , Feminino , Citometria de Fluxo , Interleucina-12/administração & dosagem , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Receptor fas/genética
14.
Cancer Res ; 72(13): 3125-30, 2012 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-22721837

RESUMO

Established tumors are complex masses that contain not only neoplastic cells but also nontransformed cellular elements such as stromal cells, the neovasculature, and the full gamut of immune cells. However, evidence suggests that, unlike cells found in lymphoid organs that productively respond to acute infections, immune cells in tumors are dysregulated and functionally impaired. Tumor masses can contain regulatory lymphocytes, myeloid-derived suppressor cells, alternatively activated macrophages, and dendritic cells. Ablation or reprogramming of this aberrant microenvironment might dramatically augment cancer therapies, and this strategy is currently being deployed in a variety of clinical trials. A better understanding of the cellular constituents of tumors and the mechanisms involved in immune evasion may help guide the next generation of innovative cancer immunotherapies.


Assuntos
Evasão Tumoral , Microambiente Tumoral , Humanos , Imunoterapia , Interleucina-12/genética , Ativação de Macrófagos , Neoplasias/imunologia , Neoplasias/terapia
15.
Blood ; 119(18): 4096-7, 2012 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-22555659

RESUMO

In this issue of Blood, Pegram and colleagues describe the ability to improve adoptive T-cell therapies by engineering T cells to secrete the potent proinflammatory cytokine IL-12.1 These exciting findings significantly extend previous observations made in a murine melanoma model targeting naturally occurring tumor antigens.2,3

16.
Clin Cancer Res ; 18(6): 1672-83, 2012 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-22291136

RESUMO

PURPOSE: We investigated the feasibility of delivering the proinflammatory cytokine interleukin (IL)-12 into tumor using T cells genetically engineered to express a chimeric antigen receptor (CAR) against the VEGF receptor-2 (VEGFR-2). EXPERIMENTAL DESIGN: Two different strains of mice bearing five different established subcutaneous tumors were treated with syngeneic T cells cotransduced with an anti-VEGFR-2 CAR and a constitutively expressed single-chain murine IL-12 or an inducible IL-12 gene after host lymphodepletion. Tumor regression, survival of mice, and persistence of the transferred cells were evaluated. RESULTS: Adoptive transfer of syngeneic T cells cotransduced with an anti-VEGFR-2 CAR and a constitutively expressing single-chain IL-12 resulted in the regression of five different established tumors of different histologies without the need for IL-2 administration. T cells transduced with either anti-VEGFR-2 CAR or single-chain IL-12 alone did not alter the tumor growth indicating that both of them had to be expressed in the same cell to mediate tumor regression. Anti-VEGFR-2 CAR and IL-12-cotransduced T cells infiltrated the tumors, expanded, and persisted for prolonged periods. The antitumor effect did not require the presence of host T and B cells but was dependent on host IL-12R-expressing cells. The anti-VEGFR-2 CAR changed the immunosuppressive tumor environment by altering/reducing both the systemic and the intratumoral CD11b(+)Gr1(+) myeloid suppressor cell subsets that expressed VEGFR-2. CONCLUSIONS: These results suggest that targeted delivery of IL-12 into the tumor environment with T cells redirected against VEGFR-2 is a promising approach for treating patients with a variety of solid tumor types.


Assuntos
Imunoterapia Adotiva/métodos , Interleucina-12/administração & dosagem , Interleucina-12/genética , Neoplasias/terapia , Linfócitos T/imunologia , Transdução Genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/imunologia , Animais , Estudos de Viabilidade , Feminino , Humanos , Interleucina-12/metabolismo , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Neoplasias/irrigação sanguínea , Receptores de Antígenos de Linfócitos T/imunologia , Proteínas Recombinantes/metabolismo
17.
Immunity ; 35(6): 972-85, 2011 Dec 23.
Artigo em Inglês | MEDLINE | ID: mdl-22177921

RESUMO

Th17 cells have been described as short lived, but this view is at odds with their capacity to trigger protracted damage to normal and transformed tissues. We report that Th17 cells, despite displaying low expression of CD27 and other phenotypic markers of terminal differentiation, efficiently eradicated tumors and caused autoimmunity, were long lived, and maintained a core molecular signature resembling early memory CD8(+) cells with stem cell-like properties. In addition, we found that Th17 cells had high expression of Tcf7, a direct target of the Wnt and ß-catenin signaling axis, and accumulated ß-catenin, a feature observed in stem cells. In vivo, Th17 cells gave rise to Th1-like effector cell progeny and also self-renewed and persisted as IL-17A-secreting cells. Multipotency was required for Th17 cell-mediated tumor eradication because effector cells deficient in IFN-γ or IL-17A had impaired activity. Thus, Th17 cells are not always short lived and are a less-differentiated subset capable of superior persistence and functionality.


Assuntos
Células-Tronco/metabolismo , Células Th17/imunologia , Animais , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Sobrevivência Celular/genética , Perfilação da Expressão Gênica , Interleucina-17/biossíntese , Camundongos , Camundongos Transgênicos , Neoplasias/imunologia , Células-Tronco/citologia , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Células Th1/citologia , Células Th1/imunologia , Células Th17/citologia , Células Th17/metabolismo , Membro 7 da Superfamília de Receptores de Fatores de Necrose Tumoral/genética , Membro 7 da Superfamília de Receptores de Fatores de Necrose Tumoral/metabolismo
18.
J Clin Invest ; 121(12): 4746-57, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-22056381

RESUMO

Solid tumors are complex masses with a local microenvironment, or stroma, that supports tumor growth and progression. Among the diverse tumor-supporting stromal cells is a heterogeneous population of myeloid-derived cells. These cells are alternatively activated and contribute to the immunosuppressive environment of the tumor; overcoming their immunosuppressive effects may improve the efficacy of cancer immunotherapies. We recently found that engineering tumor-specific CD8(+) T cells to secrete the inflammatory cytokine IL-12 improved their therapeutic efficacy in the B16 mouse model of established melanoma. Here, we report the mechanism underlying this finding. Surprisingly, direct binding of IL-12 to receptors on lymphocytes or NK cells was not required. Instead, IL-12 sensitized bone marrow-derived tumor stromal cells, including CD11b(+)F4/80(hi) macrophages, CD11b(+)MHCII(hi)CD11c(hi) dendritic cells, and CD11b(+)Gr-1(hi) myeloid-derived suppressor cells, causing them to enhance the effects of adoptively transferred CD8(+) T cells. This reprogramming of myeloid-derived cells occurred partly through IFN-γ. Surprisingly, direct presentation of antigen to the transferred CD8(+) T cells by tumor was not necessary; however, MHCI expression on host cells was essential for IL-12-mediated antitumor enhancements. These results are consistent with a model in which IL-12 enhances the ability of CD8(+) T cells to collapse large vascularized tumors by triggering programmatic changes in otherwise suppressive antigen-presenting cells within tumors and support the use of IL-12 as part of immunotherapy for the treatment of solid tumors.


Assuntos
Linfócitos T CD8-Positivos/transplante , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Imunoterapia Adotiva , Interleucina-12/fisiologia , Melanoma Experimental/terapia , Células Mieloides/fisiologia , Evasão Tumoral/imunologia , Microambiente Tumoral/imunologia , Animais , Apresentação de Antígeno , Células Apresentadoras de Antígenos/imunologia , Células Apresentadoras de Antígenos/patologia , Antígenos de Neoplasias/imunologia , Células da Medula Óssea/imunologia , Células da Medula Óssea/patologia , Linfócitos T CD8-Positivos/metabolismo , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica/imunologia , Inflamação/genética , Interferon gama/fisiologia , Interleucina-12/genética , Interleucina-12/metabolismo , Subpopulações de Linfócitos/imunologia , Subpopulações de Linfócitos/patologia , Macrófagos/fisiologia , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Camundongos , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/imunologia , Quimera por Radiação , Proteínas Recombinantes de Fusão/fisiologia , Células Estromais/patologia , Células Estromais/fisiologia
19.
Clin Cancer Res ; 17(16): 5343-52, 2011 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-21737507

RESUMO

PURPOSE: Adoptive cell transfer (ACT) of tumor infiltrating or genetically engineered T cells can cause durable responses in patients with metastatic cancer. Multiple clinically modifiable parameters can comprise this therapy, including cell dose and phenotype, in vivo antigen restimulation, and common gamma-chain (γ(c)) cytokine support. However, the relative contributions of each these individual components to the magnitude of the antitumor response have yet to be quantified. EXPERIMENTAL DESIGN: To systematically and quantitatively appraise each of these variables, we employed the Pmel-1 mouse model treating large, established B16 melanoma tumors. In addition to cell dose and magnitude of in vivo antigen restimulation, we also evaluated the relative efficacy of central memory (T(CM)), effector memory (T(EM)), and stem cell memory (T(SCM)) subsets on the strength of tumor regression as well as the dose and type of clinically available γ(c) cytokines, including IL-2, IL-7, IL-15, and IL-21. RESULTS: We found that cell dose, T-cell differentiation status, and viral vaccine titer each were correlated strongly and significantly with the magnitude of tumor regression. Surprisingly, although the total number of IL-2 doses was correlated with tumor regression, no significant benefit to prolonged (≥6 doses) administration was observed. Moreover, the specific type and dose of γ(c) cytokine only moderately correlated with response. CONCLUSION: Collectively, these findings elucidate some of the key determinants of successful ACT immunotherapy for the treatment of cancer in mice and further show that γ(c) cytokines offer a similar ability to effectively drive antitumor T-cell function in vivo.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Citocinas/imunologia , Imunoterapia Adotiva/métodos , Melanoma Experimental/imunologia , Animais , Linfócitos T CD8-Positivos/transplante , Vacinas Anticâncer/administração & dosagem , Vacinas Anticâncer/imunologia , Diferenciação Celular/imunologia , Linhagem Celular Tumoral , Citocinas/uso terapêutico , Relação Dose-Resposta a Droga , Memória Imunológica/imunologia , Interleucina-15/imunologia , Interleucina-15/farmacologia , Interleucina-2/imunologia , Interleucina-2/farmacologia , Interleucina-7/imunologia , Interleucina-7/farmacologia , Interleucinas/imunologia , Interleucinas/farmacologia , Melanoma Experimental/patologia , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Análise de Regressão , Resultado do Tratamento , Carga Tumoral/efeitos dos fármacos , Carga Tumoral/imunologia
20.
J Immunother ; 34(4): 343-52, 2011 May.
Artigo em Inglês | MEDLINE | ID: mdl-21499127

RESUMO

T-cell receptor (TCR) gene therapy enables for the rapid creation of antigen-specific T cells from mice of any strain and represents a valuable tool for preclinical immunotherapy studies. Here, we describe the superiority of γ-retroviral vectors compared with lentiviral vectors for transduction of murine T cells and surprisingly illustrate robust gene-transfer into phenotypically naive/memory-stem cell like (TN/TSCM; CD62L(hi)/CD44(low)) and central memory (TCM; CD62L(hi)/CD44(hi)) CD8+ T cells using murine stem cell-based γ-retroviral vectors (MSGV1). We created MSGV1 vectors for a major histocompatibility complex-class I-restricted TCR specific for the melanocyte-differentiation antigen, glycoprotein 100 (MSGV1-pmel-1), and a major histocompatibility complex-class II-restricted TCR specific for tyrosinase-related protein-1 (MSGV1-TRP-1), and found that robust gene expression required codon optimization of TCR sequences for the pmel-1 TCR. To test for functionality, we adoptively transferred TCR-engineered T cells into mice bearing B16 melanomas and observed delayed growth of established tumors with pmel-1 TCR engineered CD8+ T cells and significant tumor regression with TRP-1 TCR transduced CD4 T cells. We simultaneously created lentiviral vectors encoding the pmel-1 TCR, but found that these vectors mediated low TCR expression in murine T cells, but robust gene expression in other murine and human cell lines. These results indicate that preclinical murine models of adoptive immunotherapies are more practical using γ-retroviral rather than lentiviral vectors.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Engenharia Genética , Imunoterapia Adotiva , Animais , Técnicas de Transferência de Genes , Vetores Genéticos/genética , Células HEK293 , Humanos , Células Jurkat , Melanoma Experimental/imunologia , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células NIH 3T3 , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia , Retroviridae/genética , Transdução Genética , Antígeno gp100 de Melanoma/genética , Antígeno gp100 de Melanoma/imunologia
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