Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 109
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Recent Results Cancer Res ; 214: 93-128, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31473850

RESUMO

As a specifically programmable, living immunotherapeutic drug, chimeric antigen receptor (CAR)-modified T cells are providing an alternative treatment option for a broad variety of diseases including so far refractory cancer. By recognizing a tumor-associated antigen, the CAR triggers an anti-tumor response of engineered patient's T cells achieving lasting remissions in the treatment of leukemia and lymphoma. During the last years, significant progress was made in optimizing the CAR design, in manufacturing CAR-engineered T cells, and in the clinical management of patients showing promise to establish adoptive CAR T cell therapy as an effective treatment option in the forefront.


Assuntos
Imunoterapia Adotiva , Neoplasias/terapia , Receptores de Antígenos Quiméricos , Antígenos de Neoplasias/imunologia , Ensaios Clínicos como Assunto , Humanos , Receptores de Antígenos de Linfócitos T , Linfócitos T/imunologia
2.
Mol Ther ; 27(10): 1825-1835, 2019 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-31331813

RESUMO

Chimeric antigen receptor (CAR)-engineered T cells are efficacious in controlling advanced leukemia and lymphoma, however, they fail in the treatment of solid cancer, which is thought to be due to insufficient T cell activation. We revealed that the immune response of CAR T cells with specificity for carcinoembryonic antigen (CEA) was more efficacious against CEA+ cancer cells when simultaneously incubated with an anti-CD30 immunotoxin or anti-CD30 CAR T cells, although the targeted cancer cells lack CD30. The same effect was achieved when the anti-CD30 single-chain variable fragment (scFv) was integrated into the extracellular domain of the anti-CEA CAR. Improvement in T cell activation was due to interfering with the T cell CD30-CD30L interaction by the antagonistic anti-CD30 scFv HRS3; an agonistic anti-CD30 scFv or targeting the high-affinity interleukin-2 (IL-2) receptor was not effective. T cells with the anti-CD30/CEA CAR showed superior immunity against established CEA+ CD30- tumors in a mouse model. The concept is broadly applicable since anti-CD30/TAG72 CAR T cells also showed improved elimination of TAG72+ CD30- cancer cells. Taken together, targeting CD30 on CAR T cells by the HRS3 scFv within the anti-tumor CAR improves the redirected immune response against solid tumors.

3.
Sci Rep ; 9(1): 8410, 2019 Jun 10.
Artigo em Inglês | MEDLINE | ID: mdl-31182802

RESUMO

The demand for recombinant proteins for analytic and therapeutic purposes is increasing; however, most currently used bacterial production systems accumulate the recombinant proteins in the intracellular space, which requires denaturating procedures for harvesting and functional testing. We here present a novel FimH-based expression system that enables display of fully functional eukaryotic proteins while preventing technical difficulties in translocating, folding, stabilizing and isolating the displayed proteins. As examples, Gaussia Luciferase (GLuc), epidermal growth factor (EGF), transforming growth factor-α (TGF-α) and epiregulin (EPRG) were expressed as FimH fusion proteins on the surface of E. coli bacteria. The fusion proteins were functionally active and could be released from the bacterial surface by specific proteolytic cleavage into the culture supernatant allowing harvesting of the produced proteins. EGFR ligands, produced as FimH fusion proteins and released by proteolytic cleavage, bound to the EGF receptor (EGFR) on cancer cells inducing EGFR phosphorylation. In another application of the technology, GLuc-FimH expressed on the surface of bacteria was used to track tumor-infiltrating bacteria by bioluminescence imaging upon application to mice, thereby visualizing the colonization of transplanted tumors. The examples indicate that the FimH-fusion protein technology can be used in various applications that require functionally active proteins to be displayed on bacterial surfaces or released into the culture supernatant.

4.
Mol Ther ; 26(8): 1906-1920, 2018 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-30078440

RESUMO

Autologous T cells genetically modified with a chimeric antigen receptor (CAR) redirected at CD19 have potent activity in the treatment of B cell leukemia and B cell non-Hodgkin's lymphoma (B-NHL). Immunotherapies to treat multiple myeloma (MM) targeted the B cell maturation antigen (BCMA), which is expressed in most cases of MM. We developed a humanized CAR with specificity for BCMA based on our previously generated anti-BCMA monoclonal antibody. The targeting single-chain variable fragment (scFv) domain exhibited a binding affinity in the low nanomolar range, conferring T cells with high functional avidity. Redirecting T cells by this CAR allowed us to explore BCMA as an alternative target for mature B-NHLs. We validated BCMA expression in diffuse large B cell lymphoma, follicular lymphoma, mantle cell lymphoma, and chronic lymphocytic leukemia. BCMA CAR T cells triggered target cell lysis with an activation threshold in the range of 100 BCMA molecules, which allowed for an efficient eradication of B-NHL cells in vitro and in vivo. Our data corroborate BCMA is a suitable target in B cell tumors beyond MM, providing a novel therapeutic option for patients where BCMA is expressed at low abundance or where anti-CD19 immunotherapies have failed due to antigen loss.

5.
Stem Cells ; 36(11): 1752-1763, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30063808

RESUMO

The trabecular extracellular matrix (ECM) forms a three-dimensional scaffold to stabilize the bone marrow, provide substrates for cell-matrix interactions and retain, present or release signals to modulate hematopoietic stem and progenitor cell development. However, the impact of trabecular ECM components on hematopoiesis has been poorly studied. Using collagen IX alpha1 - deficient (Col9a1(-/-) ) mice, we revealed that a lack of collagen IX alpha1 results in a disorganized trabecular network enriched in fibronectin, and in a reduction in myeloid cells, which was accompanied by a decrease in colony-stimulating factor 1 receptor expression on monocytes from the bone marrow. In contrast, B-cell numbers in the bone marrow and T-cell numbers in the thymus remained unchanged. Alterations in the bone marrow microenvironment may not only reduce myeloid cell numbers, but also have long-term implications for myeloid cell function. Mice were infected with Listeria moncytogenes to analyze the function of myeloid cells. In this case, an inadequate macrophage-dependent clearance of bacterial infections was observed in Col9a1(-/-) mice in vivo. This was mainly caused by an impaired interferon-gamma/tumor necrosis factor-alpha-mediated activation of macrophages. The loss of collagen IX alpha1 therefore destabilizes the trabecular bone network, impairs myeloid cell differentiation, and affects the innate immune response against Listeria. Stem Cells 2018;36:1752-1763.

6.
Mol Ther ; 26(9): 2218-2230, 2018 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-30055872

RESUMO

Adoptive cell therapy with chimeric antigen receptor (CAR)-redirected T cells induced spectacular regressions of leukemia and lymphoma, however, failed so far in the treatment of solid tumors. A cause is thought to be T cell repression through TGF-ß, which is massively accumulating in the tumor tissue. Here, we show that T cells with a CD28-ζ CAR, but not with a 4-1BB-ζ CAR, resist TGF-ß-mediated repression. Mechanistically, LCK activation and consequently IL-2 release and autocrine IL-2 receptor signaling mediated TGF-ß resistance; deleting the LCK-binding motif in the CD28 CAR abolished both IL-2 secretion and TGF-ß resistance, while IL-2 add-back restored TGF-ß resistance. Other γ-cytokines like IL-7 and IL-15 could replace IL-2 in this context. This is demonstrated by engineering IL-2 deficient CD28ΔLCK-ζ CAR T cells with a hybrid IL-7 receptor to provide IL-2R ß chain signaling upon IL-7 binding. Such modified T cells showed improved CAR T cell activity against TGF-ß+ tumors. Data draw the concept that an autocrine loop resulting in IL-2R signaling can make CAR T cells more potent in staying active against TGF-ß+ solid tumors.

7.
Hum Gene Ther ; 29(5): 559-568, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29620951

RESUMO

The adoptive transfer of chimeric antigen receptor (CAR)-modified T cells is attracting growing interest for the treatment of malignant diseases. Early trials with anti-CD19 CAR T cells have achieved spectacular remissions in B-cell leukemia and lymphoma, so far refractory, very recently resulting in the Food and Drug Administration approval of CD19 CAR T cells for therapy. With further applications and increasing numbers of patients, the reproducible manufacture of high-quality clinical-grade CAR T cells is becoming an ever greater challenge. New processing techniques, quality-control mechanisms, and logistic developments are required to meet both medical needs and regulatory restrictions. This paper summarizes the state-of-the-art in manufacturing CAR T cells and the current challenges that need to be overcome to implement this type of cell therapy in the treatment of a variety of malignant diseases and in a greater number of patients.

8.
Int J Mol Sci ; 19(2)2018 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-29385713

RESUMO

Recent clinical trials have shown that adoptive chimeric antigen receptor (CAR) T cell therapy is a very potent and possibly curative option in the treatment of B cell leukemias and lymphomas. However, targeting a single antigen may not be sufficient, and relapse due to the emergence of antigen negative leukemic cells may occur. A potential strategy to counter the outgrowth of antigen escape variants is to broaden the specificity of the CAR by incorporation of multiple antigen recognition domains in tandem. As a proof of concept, we here describe a bispecific CAR in which the single chain variable fragment (scFv) is replaced by a tandem of two single-antibody domains or nanobodies (nanoCAR). High membrane nanoCAR expression levels are observed in retrovirally transduced T cells. NanoCARs specific for CD20 and HER2 induce T cell activation, cytokine production and tumor lysis upon incubation with transgenic Jurkat cells expressing either antigen or both antigens simultaneously. The use of nanobody technology allows for the production of compact CARs with dual specificity and predefined affinity.


Assuntos
Receptores de Antígenos de Linfócitos T , Proteínas Recombinantes de Fusão , Anticorpos de Domínio Único , Linfócitos T/imunologia , Humanos , Células Jurkat , Leucemia de Células B/genética , Leucemia de Células B/imunologia , Leucemia de Células B/patologia , Leucemia de Células B/terapia , Linfoma de Células B/genética , Linfoma de Células B/imunologia , Linfoma de Células B/patologia , Linfoma de Células B/terapia , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Anticorpos de Domínio Único/genética , Anticorpos de Domínio Único/imunologia , Linfócitos T/patologia , Transdução Genética
9.
Matrix Biol ; 65: 59-74, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-28797711

RESUMO

After skin injury fibroblasts migrate into the wound and transform into contractile, extracellular matrix-producing myofibroblasts to promote skin repair. Persistent activation of myofibroblasts can cause excessive fibrotic reactions, but the underlying mechanisms are not fully understood. We used SMA-GFP transgenic mice to study myofibroblast recruitment and activation in skin wounds. Myofibroblasts were initially recruited to wounds three days post injury, their number reached a maximum after seven days and subsequently declined. Expression profiling showed that 1749 genes were differentially expressed in sorted myofibroblasts from wounds seven days post injury. Most of these genes were linked with the extracellular region and cell periphery including genes encoding for extracellular matrix proteins. A unique panel of core matrisome and matrisome-associated genes was differentially expressed in myofibroblasts and several genes not yet known to be linked to myofibroblast-mediated wound healing were found (e.g. Col24a1, Podnl1, Bvcan, Tinagl1, Thbs3, Adamts16, Adamts19, Cxcl's, Ccl's). In addition, a complex network of G protein-coupled signaling events was regulated in myofibroblasts (e.g. Adcy1, Plbc4, Gnas). Hence, this first characterization of a myofibroblast-specific expression profile at the peak of in situ granulation tissue formation provides important insights into novel target genes that may control excessive ECM deposition during fibrotic reactions.


Assuntos
Actinas/genética , Perfilação da Expressão Gênica/métodos , Redes Reguladoras de Genes , Pele/lesões , Actinas/metabolismo , Animais , Diferenciação Celular , Modelos Animais de Doenças , Regulação da Expressão Gênica , Camundongos , Camundongos Transgênicos , Miofibroblastos/química , Miofibroblastos/citologia , Análise de Sequência com Séries de Oligonucleotídeos , Especificidade de Órgãos , Pele/citologia , Pele/metabolismo
10.
Cell Rep ; 21(11): 3205-3219, 2017 Dec 12.
Artigo em Inglês | MEDLINE | ID: mdl-29241547

RESUMO

Adoptive therapy with chimeric antigen receptor (CAR)-redirected T cells has achieved remarkable efficacy in the treatment of hematopoietic malignancies. However, eradicating large solid tumors in advanced stages of the disease remains challenging. We explored augmentation of the anti-tumor immune reaction by establishing an acute inflammatory reaction. Systematic screening indicates that IL-18 polarizes CAR T cells toward T-bethigh FoxO1low effectors with an acute inflammatory response. CAR T cells engineered with inducible IL-18 release exhibited superior activity against large pancreatic and lung tumors that were refractory to CAR T cells without cytokines. IL-18 CAR T cell treatment was accompanied by an overall change in the immune cell landscape associated with the tumor. More specifically, CD206- M1 macrophages and NKG2D+ NK cells increased in number, whereas Tregs, suppressive CD103+ DCs, and M2 macrophages decreased, suggesting that "iIL18 TRUCKs" can be used to sensitize large solid tumor lesions for successful immune destruction.


Assuntos
Proteína Forkhead Box O1/imunologia , Imunoterapia Adotiva/métodos , Interleucina-18/imunologia , Neoplasias Pulmonares/terapia , Neoplasias Pancreáticas/terapia , Proteínas com Domínio T/imunologia , Linfócitos T/imunologia , Animais , Antígeno Carcinoembrionário/genética , Antígeno Carcinoembrionário/imunologia , Engenharia Celular , Células Dendríticas/imunologia , Células Dendríticas/patologia , Modelos Animais de Doenças , Proteína Forkhead Box O1/genética , Expressão Gênica , Humanos , Interleucina-18/genética , Interleucina-18/metabolismo , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/patologia , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/mortalidade , Neoplasias Pulmonares/patologia , Macrófagos/imunologia , Macrófagos/patologia , Camundongos , Proteínas Mutantes Quiméricas/genética , Proteínas Mutantes Quiméricas/imunologia , Neoplasias Pancreáticas/imunologia , Neoplasias Pancreáticas/mortalidade , Neoplasias Pancreáticas/patologia , Cultura Primária de Células , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia , Análise de Sobrevida , Proteínas com Domínio T/genética , Linfócitos T/patologia , Linfócitos T/transplante , Transgenes , Células Tumorais Cultivadas
11.
Hum Gene Ther ; 28(12): 1147-1157, 2017 12.
Artigo em Inglês | MEDLINE | ID: mdl-29207878

RESUMO

The treatment of leukemia/lymphoma by chimeric antigen receptor (CAR) redirected T cells with specificity for CD19 induced complete remissions in the majority of patients, with a realistic hope for cure. However, recent follow-up data revealed a substantial risk of relapse through leukemic cells that lack the CAR targeted antigen. In this situation, a bispecific CAR with binding domains for CD19 and CD20 is aimed at recognizing leukemic cells with only one cognate antigen. The anti-CD20-CD19 bispecific CAR induced a full T-cell response upon engagement of CD19 or CD20 on target cells showing a true "OR" gate recognition in redirecting T-cell activation. T cells with the anti-CD20-CD19 CAR efficiently killed patients' chronic lymphocytic leukemia cells in vitro. The bispecific CAR T cells cleared pediatric acute lymphocytic leukemia with a mixed CD19+CD20+/CD20- phenotype from the blood and bone marrow of transplanted mice, while anti-CD20 CAR T cells left CD20- leukemic cells behind without curing the disease. Data indicate the superior anti-leukemic activity in the control of leukemia, implying that the anti-CD20-CD19 bispecific CAR T cells may reduce the risk of relapse through antigen-loss leukemic cells in the long term.


Assuntos
Antígenos CD19/imunologia , Antígenos CD20/imunologia , Imunoterapia Adotiva/métodos , Linfoma de Células B/imunologia , Linfoma de Células B/terapia , Receptores de Antígenos de Linfócitos T , Proteínas Recombinantes de Fusão , Linfócitos T , Linhagem Celular Tumoral , Feminino , Células HEK293 , Humanos , Linfoma de Células B/genética , Masculino , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Linfócitos T/imunologia , Linfócitos T/transplante
12.
Front Immunol ; 8: 1125, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28955341

RESUMO

Cellular therapy with chimeric antigen receptor (CAR)-redirected cytotoxic T cells has shown impressive efficacy in the treatment of hematologic malignancies. We explored a regulatory T cell (Treg)-based therapy in the treatment of allergic airway inflammation, a model for asthma, which is characterized by an airway hyper-reactivity (AHR) and a chronic, T helper-2 (Th2) cell-dominated immune response to allergen. To restore the immune balance in the lung, we redirected Tregs by a CAR toward lung epithelia in mice upon experimentally induced allergic asthma, closely mimicking the clinical situation. Adoptively transferred CAR Tregs accumulated in the lung and in tracheobronchial lymph nodes, reduced AHR and diminished eosinophilic airway inflammation, indicated by lower cell numbers in the bronchoalveolar lavage fluid and decreased cell infiltrates in the lung. CAR Treg cells furthermore prevented excessive pulmonary mucus production as well as increase in allergen-specific IgE and Th2 cytokine levels in exposed animals. CAR Tregs were more efficient in controlling asthma than non-modified Tregs, indicating the pivotal role of specific Treg cell activation in the affected organ. Data demonstrate that lung targeting CAR Treg cells ameliorate key features of experimental airway inflammation, paving the way for cell therapy of severe allergic asthma.

13.
Cancers (Basel) ; 9(9)2017 Aug 29.
Artigo em Inglês | MEDLINE | ID: mdl-28850063

RESUMO

Evidences are accumulating that CD4⁺ T cells can physiologically mediate antigen specific target cell lysis. By circumventing major histocompatibility complex (MHC)-restrictions through an engineered chimeric antigen receptor (CAR), CD4⁺ T cells lyse defined target cells as efficiently as do CD8⁺ T cells. However, the cytolytic capacity of redirected CD4⁺CD25- T cells, in comparison with CD4⁺CD25⁺ regulatory T (Treg) cells was so far not thoroughly defined. Treg cells require a strong CD28 signal together with CD3ζ for activation. We consequently used a CAR with combined CD28-CD3ζ signalling for redirecting CD4⁺CD25- T cells and CD4⁺CD25⁺ Treg cells from the same donor. CAR redirected activation of these T cell subsets and induced a distinct cytokine pattern with high IL-10 and a lack of IL-2 release by Treg cells. Despite strong antigen-specific activation, CAR Treg cells produced only weak target cell lysis, whereas CD4⁺CD25- CAR T cells were potent killers. Cytolysis did not correlate with the target cell sensitivity to Fas/FasL mediated killing; CD4⁺CD25- T cells upregulated perforin and granzyme B upon CAR activation, whereas Treg cells did less. The different cytolytic capacities of CAR redirected conventional CD4⁺ cells and Treg cells imply their use for different purposes in cell therapy.

14.
Hum Gene Ther ; 28(11): 1047-1060, 2017 11.
Artigo em Inglês | MEDLINE | ID: mdl-28810803

RESUMO

Adoptive therapy with chimeric antigen receptor (CAR) redirected T cells achieved lasting remissions in hematologic malignancies, even in terminal stages of the disease. Exploring CAR T cell therapy in the treatment of solid tumors has just begun, balancing efficacy versus toxicity in early phase trials. In contrast to leukemia/lymphoma, solid tumors display a tremendously variable biology demanding different strategies to make a T cell attack successful in the long term. This article summarizes current developments, discusses the hurdles, and considers some modifications to improve the CAR T cell therapy in the treatment of solid tumors.


Assuntos
Imunoterapia Adotiva/tendências , Neoplasias/terapia , Receptores de Antígenos de Linfócitos T/uso terapêutico , Linfócitos T/imunologia , Terapia Baseada em Transplante de Células e Tecidos/tendências , Humanos , Neoplasias/genética , Neoplasias/imunologia , Receptores de Antígenos de Linfócitos T/imunologia
15.
Cancer Immunol Res ; 5(9): 730-743, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28778961

RESUMO

Cancer immunotherapy by therapeutic activation of T cells has demonstrated clinical potential. Approaches include checkpoint inhibitors and chimeric antigen receptor T cells. Here, we report the development of an alternative strategy for cellular immunotherapy that combines induction of a tumor-directed T-cell response and antibody secretion without the need for genetic engineering. CD40 ligand stimulation of murine tumor antigen-specific B cells, isolated by antigen-biotin tetramers, resulted in the development of an antigen-presenting phenotype and the induction of a tumor antigen-specific T-cell response. Differentiation of antigen-specific B cells into antibody-secreting plasma cells was achieved by stimulation with IL21, IL4, anti-CD40, and the specific antigen. Combined treatment of tumor-bearing mice with antigen-specific CD40-activated B cells and antigen-specific plasma cells induced a therapeutic antitumor immune response resulting in remission of established tumors. Human CEA or NY-ESO-1-specific B cells were detected in tumor-draining lymph nodes and were able to induce antigen-specific T-cell responses in vitro, indicating that this approach could be translated into clinical applications. Our results describe a technique for the exploitation of B-cell effector functions and provide the rationale for their use in combinatorial cancer immunotherapy. Cancer Immunol Res; 5(9); 730-43. ©2017 AACR.


Assuntos
Antígenos de Neoplasias/imunologia , Antígenos CD40/imunologia , Imunoterapia , Neoplasias/imunologia , Linfócitos T/imunologia , Animais , Células Apresentadoras de Antígenos/imunologia , Linfócitos B/imunologia , Células Dendríticas/imunologia , Humanos , Imunidade Celular , Interleucina-4/imunologia , Interleucinas/imunologia , Ativação Linfocitária/imunologia , Camundongos , Neoplasias/patologia , Neoplasias/terapia , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia
16.
Hum Gene Ther Methods ; 28(6): 302-309, 2017 12.
Artigo em Inglês | MEDLINE | ID: mdl-28741380

RESUMO

Redirected T cells genetically modified with a chimeric antigen receptor (CAR) have induced spectacular remissions of refractory leukemia/lymphoma in early phase trials, attracting interest to use CAR T cells in a variety of other applications including solid cancer and nonmalignant diseases. However, extensive preclinical explorations demand highly effective and robust procedures for the genetic modification of blood T cells; the same applies for engineering with a recombinant T cell receptor. We present laboratory procedures in a step-by-step protocol to engineer human and mouse T cells with a CAR by γ-retro- or lentiviral transduction for further preclinical testing.


Assuntos
Lentivirus/genética , Receptores de Antígenos de Linfócitos T/genética , Linfócitos T/metabolismo , Transdução Genética/métodos , Animais , Células Cultivadas , Terapia Genética/métodos , Vetores Genéticos/genética , Humanos , Camundongos , Receptores de Antígenos de Linfócitos T/metabolismo
18.
Mol Ther Methods Clin Dev ; 5: 116-129, 2017 Jun 16.
Artigo em Inglês | MEDLINE | ID: mdl-28480311

RESUMO

The role of striatin interacting protein 2 (Strip2) in differentiation of embryonic stem cells (ESCs) is still under debate. Strip2-silenced murine (KD) ESCs were differentiated for 4, 8, 12, and 16 days. We show that Strip2 is distributed in the perinucleus or nuclei of wild-type (WT) undifferentiated ESCs, but is localized in high-density nuclear bodies in differentiated cells. CellNet analysis of microarray gene expression data for the KD and scrambled control (SCR) embryoid bodies (EBs), as well as immunostainings of key pluripotent factors, demonstrated that differentiation of KD ESCs is repressed. This occurs even in 16-day-old EBs, which possessed a high tumorigenic potential. Correlated with very high expression levels of epigenetic regulator genes, Hat1 and Dnmt3, enzymatic activities of the histone acetyltransferase type B (Hat1) and DNA (cytosine-5)-methyltransferase 3 beta (Dnmt3b) were higher in differentiated 16-day-old KD EBs than in SCR or WT EBs. The expression levels of let-7, 290, and 302 microRNA families were opposed in KD ESCs, while KD EBs had levels comparable to WT and SCR ESCs during differentiation. Strip2 is critical for the regular differentiation of ESCs. Moreover, Strip2 deficient ESCs showed a dysregulation of epigenetic regulators and microRNAs regulating pluripotency.

19.
Oncoimmunology ; 6(3): e1283460, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28405508

RESUMO

Recent clinical studies indicate that adoptive T-cell therapy and especially chimeric antigen receptor (CAR) T-cell therapy is a very potent and potentially curative treatment for B-lineage hematologic malignancies. Currently, autologous peripheral blood T cells are used for adoptive T-cell therapy. Adoptive T cells derived from healthy allogeneic donors may have several advantages; however, the expected occurrence of graft versus host disease (GvHD) as a consequence of the diverse allogeneic T-cell receptor (TCR) repertoire expressed by these cells compromises this approach. Here, we generated T cells from cord blood hematopoietic progenitor cells (HPCs) that were transduced to express an antigen receptor (AR): either a CAR or a TCR with or without built-in CD28 co-stimulatory domains. These AR-transgenic HPCs were culture-expanded on an OP9-DL1 feeder layer and subsequently differentiated to CD5+CD7+ T-lineage precursors, to CD4+ CD8+ double positive cells and finally to mature AR+ T cells. The AR+ T cells were largely naive CD45RA+CD62L+ T cells. These T cells had mostly germline TCRα and TCRß loci and therefore lacked surface-expressed CD3/TCRαß complexes. The CD3- AR-transgenic cells were mono-specific, functional T cells as they displayed specific cytotoxic activity. Cytokine production, including IL-2, was prominent in those cells bearing ARs with built-in CD28 domains. Data sustain the concept that cord blood HPC derived, in vitro generated allogeneic CD3- AR+ T cells can be used to more effectively eliminate malignant cells, while at the same time limiting the occurrence of GvHD.

20.
J Leukoc Biol ; 101(2): 577-587, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-27534894

RESUMO

Traditionally, B cells have been best known for their role as producers of antibodies. However, in recent years, a growing body of evidence has accumulated showing that B cells fulfill a range of other immunologic functions. One of the functions that has attracted increasing attention is the capacity of B cells to induce antigen-specific activation of T cells through presentation of antigens. However, the analysis of this B cell function has been hampered by the lack of a phenotypically well-defined antigen-presenting B cell subset. Here, we report the identification of a human antigen-presenting B cell subset with strong immunostimulatory properties. This B cell subset is characterized by low expression of CD21 and high expression of the activation marker CD86 and exhibits strong T cell-stimulatory activity, as demonstrated by means of an autologous mixed-lymphocyte reaction. Phenotypically, CD21lowCD86pos immunostimulatory B cells (BAPC) represented CD27+ class-switched IgMnegIgDneg B lymphocytes and displayed a higher expression of cell surface receptors, which mediate the migration from peripheral blood to sites of inflammation. Flow cytometric analysis of peripheral blood obtained from individuals with inflammatory conditions revealed that the BAPC subset was expanded following vaccination and in patients with rheumatoid arthritis. Taken together, our work shows that BAPC represents a strongly immunostimulatory B cell subset, which could be a promising target for immunotherapeutic intervention in inflammatory diseases.


Assuntos
Apresentação do Antígeno/imunologia , Células Apresentadoras de Antígenos/imunologia , Linfócitos B/imunologia , Linfócitos B/patologia , Inflamação/imunologia , Inflamação/patologia , Adulto , Subpopulações de Linfócitos B/imunologia , Antígeno B7-2/metabolismo , Ligante de CD40/metabolismo , Proliferação de Células , Regulação para Baixo , Humanos , Imunofenotipagem , Ativação Linfocitária/imunologia , Receptores de Antígenos de Linfócitos B/metabolismo , Receptores de Quimiocinas/metabolismo , Receptores de Complemento 3d/metabolismo , Transdução de Sinais , Vacinação
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA