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1.
Ther Adv Urol ; 13: 17562872211022462, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34408788

RESUMO

BACKGROUND: Prostate cancer (PCa) phenotypes vary from indolent to aggressive. Molecular subtyping may be useful in predicting aggressive cancers and directing therapy. One such subtype involving deletions of chromodomain helicase DNA binding protein 1 (CHD1), a tumor suppressor gene, are found in 10-26% of PCa tumors. In this study, we evaluate the functional cellular effects that follow CHD1 deletion. METHODS: CHD1 was knocked out (KO) in the non-tumorigenic, human papillomavirus 16 (HPV16)-immortalized prostate epithelial cell line, RWPE-1, using CRISPR/Cas9. In vitro assays such as T7 endonuclease assay, western blot, and sequencing were undertaken to characterize the CHD1 KO clones. Morphologic and functional assays for cell adhesion and viability were performed. To study expression of extracellular matrix (ECM) and adhesion molecules, a real-time (RT) profiler assay was performed using RWPE-1 parental, non-target cells (NT2) and CHD1 KO cells. RESULT: Compared to parental RWPE-1 and non-target cells (NT2), the CHD1 KO cells had a smaller, rounder morphology and were less adherent under routine culture conditions. Compared to parental cells, CHD1 KO cells showed a reduction in ECM and adhesion molecules as well as a greater proportion of viable suspension cells when cultured on standard tissue culture plates and on plates coated with laminin, fibronectin or collagen I. CHD1 KO cells showed a decrease in the expression of secreted protein acidic and rich in cysteine (SPARC), matrix metalloproteinase 2 (MMP2), integrin subunit alpha 2 (ITGA2), integrin subunit alpha 5 (ITGA5), integrin subunit alpha 6 (ITGA6), fibronectin (FN1), laminin subunit beta-3 precursor (LAMB3), collagen, tenascin and vitronectin as compared to parental and NT2 cells. CONCLUSION: These data suggest that in erythroblast transformation specific (ETS) fusion-negative, phosphatase and tensin homolog (PTEN) wildtype PCa, deletion of CHD1 alters cell-cell and cell-matrix adhesion dynamics, suggesting an important role for CHD1 in the development and progression of PCa.

2.
Proc Natl Acad Sci U S A ; 117(2): 1119-1128, 2020 01 14.
Artigo em Inglês | MEDLINE | ID: mdl-31888983

RESUMO

Reprogramming the tumor microenvironment to increase immune-mediated responses is currently of intense interest. Patients with immune-infiltrated "hot" tumors demonstrate higher treatment response rates and improved survival. However, only the minority of tumors are hot, and a limited proportion of patients benefit from immunotherapies. Innovative approaches that make tumors hot can have immediate impact particularly if they repurpose drugs with additional cancer-unrelated benefits. The seasonal influenza vaccine is recommended for all persons over 6 mo without prohibitive contraindications, including most cancer patients. Here, we report that unadjuvanted seasonal influenza vaccination via intratumoral, but not intramuscular, injection converts "cold" tumors to hot, generates systemic CD8+ T cell-mediated antitumor immunity, and sensitizes resistant tumors to checkpoint blockade. Importantly, intratumoral vaccination also provides protection against subsequent active influenza virus lung infection. Surprisingly, a squalene-based adjuvanted vaccine maintains intratumoral regulatory B cells and fails to improve antitumor responses, even while protecting against active influenza virus lung infection. Adjuvant removal, B cell depletion, or IL-10 blockade recovers its antitumor effectiveness. Our findings propose that antipathogen vaccines may be utilized for both infection prevention and repurposing as a cancer immunotherapy.


Assuntos
Imunoterapia/métodos , Vacinas contra Influenza/administração & dosagem , Vacinas contra Influenza/imunologia , Vacinas contra Influenza/uso terapêutico , Injeções Intralesionais , Neoplasias/tratamento farmacológico , Neoplasias/imunologia , Adjuvantes Imunológicos/administração & dosagem , Animais , Linfócitos B , Fatores de Transcrição de Zíper de Leucina Básica/genética , Linfócitos T CD8-Positivos/imunologia , Humanos , Imunidade Celular , Vírus da Influenza A Subtipo H1N1/imunologia , Influenza Humana , Interleucina-10 , Pulmão/patologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Repressoras/genética , Estações do Ano , Pele , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/patologia , Esqualeno/administração & dosagem , Microambiente Tumoral/efeitos dos fármacos , Vacinação
3.
J Immunother Cancer ; 7(1): 136, 2019 05 22.
Artigo em Inglês | MEDLINE | ID: mdl-31118108

RESUMO

BACKGROUND: ONC201 is a small molecule antagonist of DRD2, a G protein-coupled receptor overexpressed in several malignancies, that has prolonged antitumor efficacy and immunomodulatory properties in preclinical models. The first-in-human trial of ONC201 previously established a recommended phase II dose (RP2D) of 625 mg once every three weeks. Here, we report the results of a phase I study that evaluated the safety, pharmacokinetics (PK), and pharmacodynamics (PD) of weekly ONC201. METHODS: Patients ≥ 18 years old with an advanced solid tumor refractory to standard treatment were enrolled. Dose escalation proceeded with a 3 + 3 design from 375 mg to 625 mg of ONC201. One cycle, also the dose-limiting toxicity (DLT) window, was 21 days. The primary endpoint was to determine the RP2D of weekly ONC201, which was confirmed in an 11-patient dose expansion cohort. RESULTS: Twenty patients were enrolled: three at 375 mg and 17 at 625 mg of ONC201. The RP2D was defined as 625 mg with no DLT, treatment discontinuation, or dose modifications due to drug-related toxicity. PK profiles were consistent with every-three-week dosing and similar between the first and fourth dose. Serum prolactin and caspase-cleaved cytokeratin-18 induction were detected, along with intratumoral integrated stress response activation and infiltration of granzyme B+ Natural Killer cells. Induction of immune cytokines and effectors was higher in patients who received ONC201 once weekly versus once every three weeks. Stable disease of > 6 months was observed in several prostate and endometrial cancer patients. CONCLUSIONS: Weekly, oral ONC201 is well-tolerated and results in enhanced immunostimulatory activity that warrants further investigation. TRIAL REGISTRATION: NCT02250781 (Oral ONC201 in Treating Patients With Advanced Solid Tumors), NCT02324621 (Continuation of Oral ONC201 in Treating Patients With Advanced Solid Tumors).


Assuntos
Antineoplásicos/uso terapêutico , Neoplasias/tratamento farmacológico , Receptores de Dopamina D2/imunologia , Administração Oral , Adulto , Idoso , Idoso de 80 Anos ou mais , Antineoplásicos/farmacologia , Humanos , Pessoa de Meia-Idade
4.
Oncotarget ; 8(27): 44639-44653, 2017 Jul 04.
Artigo em Inglês | MEDLINE | ID: mdl-28591718

RESUMO

Recent evidence suggests that glutamate signaling plays an important role in cancer. Riluzole is a glutamate release inhibitor and FDA-approved drug for the treatment of amyotrophic lateral sclerosis. It has been investigated as an inhibitor of cancer cell growth and tumorigenesis with the intention of repurposing it for the treatment of cancer. Riluzole is thought to act by indirectly inhibiting glutamate signaling. However, the specific effects of riluzole in breast cancer cells are not well understood. In this study, the anti-cancer effects of riluzole were explored in a panel of breast cancer cell lines in comparison to the metabotropic glutamate receptor 1-specific inhibitor BAY 36-7620. While both drugs inhibited breast cancer cell proliferation, there were distinct functional effects suggesting that riluzole action may be metabotropic glutamate receptor 1-independent. Riluzole induced mitotic arrest independent of oxidative stress while BAY 36-7620 had no measurable effect on mitosis. BAY 36-7620 had a more pronounced and significant effect on DNA damage than riluzole. Riluzole altered cellular metabolism as demonstrated by changes in oxidative phosphorylation and cellular metabolite levels. These results provide a better understanding of the functional action of riluzole in the treatment of breast cancer.


Assuntos
Antineoplásicos/farmacologia , Receptores de Glutamato Metabotrópico/antagonistas & inibidores , Riluzol/farmacologia , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/genética , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Dano ao DNA , Relação Dose-Resposta a Droga , Metabolismo Energético , Feminino , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Expressão Gênica , Perfilação da Expressão Gênica , Humanos , Mitose/efeitos dos fármacos , Mitose/genética , Fosforilação Oxidativa/efeitos dos fármacos , Receptores de Glutamato Metabotrópico/genética , Receptores de Glutamato Metabotrópico/metabolismo , Transdução de Sinais/efeitos dos fármacos
5.
Clin Cancer Res ; 22(24): 6176-6191, 2016 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-27307599

RESUMO

PURPOSE: Current prostate cancer management calls for identifying novel and more effective therapies. Self-renewing tumor-initiating cells (TICs) hold intrinsic therapy resistance and account for tumor relapse and progression. As BMI-1 regulates stem cell self-renewal, impairing BMI-1 function for TIC-tailored therapies appears to be a promising approach. EXPERIMENTAL DESIGN: We have previously developed a combined immunophenotypic and time-of-adherence assay to identify CD49bhiCD29hiCD44hi cells as human prostate TICs. We utilized this assay with patient-derived prostate cancer cells and xenograft models to characterize the effects of pharmacologic inhibitors of BMI-1. RESULTS: We demonstrate that in cell lines and patient-derived TICs, BMI-1 expression is upregulated and associated with stem cell-like traits. From a screened library, we identified a number of post-transcriptional small molecules that target BMI-1 in prostate TICs. Pharmacologic inhibition of BMI-1 in patient-derived cells significantly decreased colony formation in vitro and attenuated tumor initiation in vivo, thereby functionally diminishing the frequency of TICs, particularly in cells resistant to proliferation- and androgen receptor-directed therapies, without toxic effects on normal tissues. CONCLUSIONS: Our data offer a paradigm for targeting TICs and support the development of BMI-1-targeting therapy for a more effective prostate cancer treatment. Clin Cancer Res; 22(24); 6176-91. ©2016 AACR.


Assuntos
Antineoplásicos/farmacologia , Autorrenovação Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células-Tronco Neoplásicas/efeitos dos fármacos , Complexo Repressor Polycomb 1/metabolismo , Neoplasias da Próstata/metabolismo , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Transformação Celular Neoplásica/efeitos dos fármacos , Transformação Celular Neoplásica/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Células-Tronco Neoplásicas/metabolismo , Neoplasias da Próstata/tratamento farmacológico , Receptores Androgênicos/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
6.
Oncotarget ; 7(20): 29689-707, 2016 May 17.
Artigo em Inglês | MEDLINE | ID: mdl-27102439

RESUMO

Receptor tyrosine kinases-based autocrine loops largely contribute to activate the MAPK and PI3K/AKT pathways in melanoma. However, the molecular mechanisms involved in generating these autocrine loops are still largely unknown. In the present study, we examine the role of the transcription factor RUNX2 in the regulation of receptor tyrosine kinase (RTK) expression in melanoma. We have demonstrated that RUNX2-deficient melanoma cells display a significant decrease in three receptor tyrosine kinases, EGFR, IGF-1R and PDGFRß. In addition, we found co-expression of RUNX2 and another RTK, AXL, in both melanoma cells and melanoma patient samples. We observed a decrease in phosphoAKT2 (S474) and phosphoAKT (T308) levels when RUNX2 knock down resulted in significant RTK down regulation. Finally, we showed a dramatic up regulation of RUNX2 expression with concomitant up-regulation of EGFR, IGF-1R and AXL in melanoma cells resistant to the BRAF V600E inhibitor PLX4720. Taken together, our results strongly suggest that RUNX2 might be a key player in RTK-based autocrine loops and a mediator of resistance to BRAF V600E inhibitors involving RTK up regulation in melanoma.


Assuntos
Comunicação Autócrina/fisiologia , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Regulação Neoplásica da Expressão Gênica/fisiologia , Melanoma/metabolismo , Receptores Proteína Tirosina Quinases/biossíntese , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/fisiologia , Humanos
7.
Cancer Lett ; 350(1-2): 52-60, 2014 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-24747123

RESUMO

The human homologue of mouse double minute 2 (MDM2) is overexpressed in tumors and contributes to tumorigenesis through inhibition of p53 activity. We investigated the effect of the anti-estrogen fulvestrant on MDM2 expression and sensitivity of estrogen receptor positive human breast cancer cell lines to chemotherapeutics. Fulvestrant down-regulated MDM2 through increased protein turnover. Fulvestrant blocked estrogen-dependent up-regulation of MDM2 and decreased basal expression of MDM2 in the absence of estradiol. As combinations of fulvestrant with doxorubicin, etoposide or paclitaxel were synergistic, altering cell cycle distribution and increasing cell death, this provides rationale for testing combinatorial chemotherapy with fulvestrant as a novel therapeutic strategy for patients with advanced breast cancer.


Assuntos
Antineoplásicos Hormonais/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Estradiol/análogos & derivados , Antagonistas de Estrogênios/uso terapêutico , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Receptores de Estrogênio/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Ciclo Celular/genética , Linhagem Celular Tumoral , Senescência Celular/efeitos dos fármacos , Inibidor de Quinase Dependente de Ciclina p21/biossíntese , Regulação para Baixo/efeitos dos fármacos , Doxorrubicina/farmacologia , Estradiol/uso terapêutico , Etoposídeo/farmacologia , Feminino , Fulvestranto , Humanos , Células MCF-7 , Paclitaxel/farmacologia , Proteínas Proto-Oncogênicas c-mdm2/biossíntese , Proteínas Proto-Oncogênicas c-mdm2/genética , RNA Mensageiro/biossíntese , Proteína Supressora de Tumor p53/biossíntese
8.
PLoS One ; 9(4): e91042, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24722054

RESUMO

Mantle cell lymphoma (MCL) is associated with a significant risk of therapeutic failure and disease relapse, but the biological origin of relapse is poorly understood. Here, we prospectively identify subpopulations of primary MCL cells with different biologic and immunophenotypic features. Using a simple culture system, we demonstrate that a subset of primary MCL cells co-cultured with either primary human mesenchymal stromal cells (hMSC) or murine MS-5 cells form in cobblestone-areas consisting of cells with a primitive immunophenotype (CD19-CD133+) containing the chromosomal translocation t (11;14)(q13;q32) characteristic of MCL. Limiting dilution serial transplantation experiments utilizing immunodeficient mice revealed that primary MCL engraftment was only observed when either unsorted or CD19-CD133+ cells were utilized. No engraftment was seen using the CD19+CD133- subpopulation. Our results establish that primary CD19-CD133+ MCL cells are a functionally distinct subpopulation of primary MCL cells enriched for MCL-initiating activity in immunodeficient mice. This rare subpopulation of MCL-initiating cells may play an important role in the pathogenesis of MCL.


Assuntos
Antígenos CD/metabolismo , Regulação Neoplásica da Expressão Gênica , Glicoproteínas/metabolismo , Linfoma de Célula do Manto/metabolismo , Células-Tronco Neoplásicas/citologia , Peptídeos/metabolismo , Antígeno AC133 , Animais , Antígenos CD19/metabolismo , Técnicas de Cocultura/métodos , Meios de Cultura , Humanos , Imunofenotipagem , Antígenos Comuns de Leucócito/metabolismo , Células-Tronco Mesenquimais/citologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Transplante de Neoplasias , Células Estromais , Translocação Genética
9.
Cancer Biol Ther ; 14(6): 546-55, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23760497

RESUMO

PDCD2 is an evolutionarily conserved eukaryotic protein with unknown function. The Drosophlia PDCD2 ortholog Zfrp8 has an essential function in fly hematopoiesis. Zfrp8 mutants exhibit marked lymph gland hyperplasia that results from increased proliferation of partially differentiated hemocytes, suggesting Zfrp8 may participate in cell growth. Based on the above observations we have focused on the role of PDCD2 in human cancer cell proliferation and hypothesized that aberrant PDCD2 expression may be characteristic of human malignancies. We report that PDCD2 is highly expressed in human acute leukemia cells as well as in normal hematopoietic progenitors. PDCD2 knockdown in cancer cells impairs their proliferation, but not viability relative to parental cells, supporting the notion that PDCD2 overexpression facilitates cancer cell growth. Prospective analysis of PDCD2 in acute leukemia patients indicates PDCD2 RNA expression correlates with disease status and is a significant predictor of clinical relapse. PDCD2's role in cell proliferation and its high expression in human malignancies make it an attractive, novel potential molecular target for new anti-cancer therapies.


Assuntos
Proteínas Reguladoras de Apoptose/metabolismo , Biomarcadores Tumorais/metabolismo , Proliferação de Células , Leucemia Mieloide Aguda/metabolismo , Recidiva Local de Neoplasia/metabolismo , Proteínas Reguladoras de Apoptose/genética , Biomarcadores Tumorais/genética , Linhagem Celular Tumoral , Expressão Gênica , Técnicas de Silenciamento de Genes , Células-Tronco Hematopoéticas/metabolismo , Humanos , Leucemia Mieloide Aguda/patologia , Estudos Prospectivos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
10.
Cancer Biol Ther ; 13(2): 114-22, 2012 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-22336911

RESUMO

Plitidepsin (Aplidin), an antitumor agent of marine origin, presently is undergoing phase II/III clinical trials, and has shown promise for the treatment of lymphoma. Here, we describe the antitumor effects of plitidepsin alone and in combination with rituximab and investigated the effects of each drug and the combination on the cell cycle and mechanism of cell death. Several Diffuse Large Cell Lymphoma (DLCL) lines and Burkitt cell lines were tested for sensitivity to plitidepsin and rituximab. All DLCL and Burkitt lymphoma cell lines were inhibited by plitidepsin in nanomolar concentrations, while rituximab sensitivity varied among different cell lines. Ramos and the RL cell lines proved sensitive to rituximab and were used to test the effects of each of the two drugs. The two agents exhibited synergism at all tested concentrations. For in vivo studies, irradiated athymic nude mice were engrafted with the Ramos lymphoma. Treatment was initiated when the tumors were ~0.5 cm in diameter, and toxic and therapeutic effects were monitored. In the in vivo study, additive effects of the combined two drugs, was demonstrated without an increase in host toxicity. The in vitro synergy and the in vivo additive antitumor effects without an increase in host toxicity with two relatively non-marrow suppressive agents encourages further development of this combination for treatment of aggressive B-cell lymphomas.


Assuntos
Anticorpos Monoclonais Murinos/farmacologia , Antineoplásicos/farmacologia , Linfoma de Burkitt/tratamento farmacológico , Depsipeptídeos/farmacologia , Linfoma Difuso de Grandes Células B/tratamento farmacológico , Animais , Anticorpos Monoclonais Murinos/administração & dosagem , Antígenos CD20/genética , Antígenos CD20/metabolismo , Antineoplásicos/administração & dosagem , Apoptose/efeitos dos fármacos , Linfoma de Burkitt/metabolismo , Linfoma de Burkitt/mortalidade , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Depsipeptídeos/administração & dosagem , Feminino , Humanos , Imunofenotipagem , Linfoma Difuso de Grandes Células B/metabolismo , Linfoma Difuso de Grandes Células B/mortalidade , Camundongos , Camundongos Nus , Peptídeos Cíclicos , Rituximab , Ensaios Antitumorais Modelo de Xenoenxerto
11.
Haematologica ; 97(8): 1255-63, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-22371181

RESUMO

BACKGROUND: There is increasing evidence that stromal cell interactions are required for the survival and drug resistance of several types of B-cell malignancies. There is relatively little information regarding the role of the bone marrow/lymphoid microenvironment in the pathogenesis of mantle cell lymphoma. In this study we investigated the interaction of primary mantle cell lymphoma cells with stromal cells in an ex vivo co-culture system. DESIGN AND METHODS: The murine stromal cell line MS-5 and human bone marrow mesenchymal stromal cells were each co-cultured with primary mantle cell lymphoma cells for up to 7 months. Mantle cell lymphoma cultures alone or combined with human stromal cells were analyzed for cell number, cell migration, nuclear factor-κB activation and drug resistance. RESULTS: Co-culture of mantle cell lymphoma cells and human stromal cells results in the survival and proliferation of primary mantle cell lymphoma cells for at least 7 months compared to mantle cell lymphoma cells cultured alone. Mantle cell lymphoma-human stromal cell interactions resulted in activation of the B-cell activating factor/nuclear factor-κB signaling axis resulting in reduced apoptosis, increased mantle cell lymphoma migration and increased drug resistance. CONCLUSIONS: Direct mantle cell lymphoma-human stromal cell interactions support long-term expansion and increase the drug-resistance of primary mantle cell lymphoma cells. This is due in part to activation of the canonical and non-canonical nuclear factor κB pathways. We also demonstrated the ability of B-cell activating factor to augment CXCL12- and CXCL13-induced cell migration. Collectively, these findings demonstrate that human stromal cell-mantle cell lymphoma interactions play a pivotal role in the pathogenesis of mantle cell lymphoma and that analysis of mantle cell lymphoma-human stromal cell interactions may help in the identification of novel targets for therapeutic use.


Assuntos
Fator Ativador de Células B/metabolismo , Linfoma de Célula do Manto/metabolismo , Células-Tronco Mesenquimais/metabolismo , NF-kappa B/metabolismo , Transdução de Sinais , Animais , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Comunicação Celular , Sobrevivência Celular , Quimiocina CXCL12/metabolismo , Quimiocina CXCL13/metabolismo , Técnicas de Cocultura , Resistencia a Medicamentos Antineoplásicos , Humanos , Linfoma de Célula do Manto/tratamento farmacológico , Camundongos , Células Estromais/fisiologia
12.
Biochem Biophys Res Commun ; 394(3): 606-11, 2010 Apr 09.
Artigo em Inglês | MEDLINE | ID: mdl-20214878

RESUMO

Differentiation agents such as 12-O-tetradecanoylphorbol-13-acetate (TPA) engage cell signaling pathways that activate downstream transcriptional programs necessary for cell differentiation. Recent evidence has indicated microRNAs (miRNAs) are an integral part of these transcriptional programs, which target key proteins and impact cell growth thereby facilitating changes required for differentiation. To further investigate the role of miRNAs in cell growth and differentiation, we focused on miR-22, a miRNA induced by TPA in the HL-60 leukemia cell line model of monocytic differentiation. TPA-induced miR-22 transcription was found to be downstream of the protein kinase c (PKC)-extracellular regulated kinase (ERK) signaling module, a pathway central to the growth and differentiation of many different cell types. Enforced miR-22 expression inhibited the growth of several different cancer cell lines, causing an accumulation of cells in the G1 phase of the cell cycle. The mechanism of miR-22's inhibitory effects involves targeting of the obligate c-Myc binding partner Max. Enforced miR-22 expression presumably lowers Max levels available for Myc binding, which differentially influenced the transcription of downstream targets of the Myc-Max complex. Our study provides additional support for miRNAs targeting key cellular regulatory microcircuits such as those governed by the Myc-Max transcriptional complex as well as their being active participants in cell growth and differentiation.


Assuntos
Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Ciclo Celular/genética , Regulação Leucêmica da Expressão Gênica , MicroRNAs/metabolismo , Ativação Transcricional , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Regulação para Baixo , Células HL-60 , Humanos , MicroRNAs/genética , Monócitos/efeitos dos fármacos , Ésteres de Forbol/farmacologia , Proteína Quinase C , Transcrição Gênica/efeitos dos fármacos
13.
Exp Hematol ; 37(2): 245-55, 2009 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-19135902

RESUMO

OBJECTIVE: MicroRNAs (miRNAs) have been implicated in complex vertebrate developmental systems, such as hematopoiesis, and may play an integral role in the development of human cancers. Based on these observations, we investigated the contribution of miRNAs to acute myelogenous leukemia cell lineage-specific differentiation. MATERIALS AND METHODS: To facilitate the identification of miRNAs and their targets relevant to leukemic cell differentiation, changes miRNA expression were analyzed in the human leukemia cell line HL-60, which historically has been utilized to study lineage-specific changes in response to the differentiation agent 12-O-tetradecanoylphorbol-13-acetate (TPA). RESULTS: Using this approach, we have identified a panel of TPA-induced miRNAs that are expressed coincident with HL-60 stereotypic morphological changes characteristic of monocytic differentiation. The transferrin receptor 1(TfR-1; CD71), whose surface expression is downregulated during TPA-mediated HL-60 cell differentiation, has been identified as a target of the TPA-induced miRNA miR-320. Cell culture experiments indicate that enforced miR-320 expression can suppress TfR-1 expression and cell proliferation. CONCLUSION: TPA induces the expression of several miRNAs in HL-60 cells, one such miRNA (miR-320) contributes to downregulation of TfR-1 surface expression characteristically seen during HL-60 monocytic differentiation. Moreover, TfR-1-targeting miRNAs, such as miR-320, may have potential as novel therapeutic agents for cancer due to their inhibitory effects on cell proliferation.


Assuntos
Antígenos CD/biossíntese , Proliferação de Células , Regulação Leucêmica da Expressão Gênica , Leucemia Mieloide Aguda/metabolismo , MicroRNAs/biossíntese , Receptores da Transferrina/biossíntese , Antígenos CD/genética , Carcinógenos/farmacologia , Diferenciação Celular/genética , Proliferação de Células/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Regulação Leucêmica da Expressão Gênica/genética , Células HL-60 , Hematopoese , Humanos , Leucemia Mieloide Aguda/genética , MicroRNAs/genética , Receptores da Transferrina/genética , Acetato de Tetradecanoilforbol/farmacologia
14.
Clin Cancer Res ; 15(4): 1232-40, 2009 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-19174488

RESUMO

PURPOSE: Small-cell lung cancers (SCLC) are defective in many regulatory mechanisms that control cell cycle progression, i.e., functional retinoblastoma protein (pRb). Flavopiridol inhibits proliferation and induces apoptosis in SCLC cell lines. We hypothesized that the sequence flavopiridol followed by doxorubicin would be synergistic in pRb-deficient SCLC cells. EXPERIMENTAL DESIGN: A H69 pRb-deficient SCLC cell line, H865, with functional pRb and H865 pRb small interfering RNA (siRNA) knockdown cells were used for in vitro and in vivo experiments. The in vivo efficiencies of various sequential combinations were tested using nude/nude athymic mice and human SCLC xenograft models. RESULTS: Flavopiridol then doxorubicin sequential treatment was synergistic in the pRB-negative H69 cell line. By knocking down pRb with specific siRNA, H865 clones with complete pRb knockdown became sensitive to flavopiridol and doxorubicin combinations. pRb-deficient SCLC cell lines were highly sensitive to flavopiridol-induced apoptosis. pRb-positive H865 cells arrested in G0-G1 with flavopiridol exposure, whereas doxorubicin and all flavopiridol/doxorubicin combinations caused a G2-M block. In contrast, pRb-negative SCLC cells did not arrest in G0-G1 with flavopiridol exposure. Flavopiridol treatment alone did not have an in vivo antitumor effect, but sequential flavopiridol followed by doxorubicin treatment provided tumor growth control and a survival advantage in Rb-negative xenograft models, compared with the other sequential treatments. CONCLUSIONS: Flavopiridol and doxorubicin sequential treatment induces potent in vitro and in vivo synergism in pRb-negative SCLC cells and should be clinically tested in tumors lacking functional pRB.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinoma de Células Pequenas/tratamento farmacológico , Doxorrubicina/administração & dosagem , Flavonoides/administração & dosagem , Genes do Retinoblastoma/fisiologia , Neoplasias Pulmonares/tratamento farmacológico , Piperidinas/administração & dosagem , Animais , Apoptose/efeitos dos fármacos , Carcinoma de Células Pequenas/genética , Carcinoma de Células Pequenas/patologia , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Masculino , Camundongos , RNA Interferente Pequeno/genética , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Clin Cancer Res ; 14(22): 7564-8, 2008 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-19010875

RESUMO

PURPOSE: Nuclear factor-kappaB (NF-kappaB) is constitutively expressed in many acute myelogenous leukemia (AML) cells and AML stem cells. Ex vivo treatment of AML cells with inhibitors of NF-kappaB results in diminished AML cell survival and enhances the cytotoxic effects of chemotherapeutic agents. The purpose of this study was to determine if standard anti-inflammatory agents modulate AML cell nuclear NF-kappaB when administered in conjunction with induction chemotherapy. EXPERIMENTAL DESIGN: Patients with newly diagnosed AML were treated with dexamethasone, choline magnesium trisalicylate, or both for 24 hours prior to and 24 hours following initiation of standard induction chemotherapy. AML cell nuclear NF-kappaB was measured at baseline, 24, and 48 hours. RESULTS: Choline magnesium trisalicylate +/- dexamethasone decreased nuclear NF-kappaB, whereas dexamethasone alone was associated with an increase in nuclear NF-kappaB in AML cells. CONCLUSIONS: These results show the feasibility of NF-kappaB modulation in conjunction with induction chemotherapy for patients with AML using inexpensive readily available medications. A follow-up study to determine the effects of NF-kappaB modulation on clinical end points is warranted.


Assuntos
Anti-Inflamatórios/administração & dosagem , Colina/análogos & derivados , Dexametasona/administração & dosagem , Leucemia Mieloide Aguda/tratamento farmacológico , NF-kappa B/biossíntese , Salicilatos/administração & dosagem , Adulto , Idoso , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Colina/administração & dosagem , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , NF-kappa B/efeitos dos fármacos
16.
Biochem Pharmacol ; 76(5): 582-8, 2008 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-18619946

RESUMO

MicroRNAs are short non-coding RNA molecules able to affect stability and/or translation of mRNA, thereby regulating the expression of genes involved in many biological processes. We report here that microRNAs miR-27a and miR-451 are involved in activating the expression of P-glycoprotein, the MDR1 gene product that confers cancer cell resistance to a broad range of chemotherapeutics. We showed that expressions of miR-27a and miR-451 were up-regulated in multidrug resistant (MDR) cancer cell lines A2780DX5 and KB-V1, as compared with their parental lines A2780 and KB-3-1. Treatment of A2780DX5 cells with the antagomirs of miR-27a or miR-451 decreased the expression of P-glycoprotein and MDR1 mRNA. In contrast, the mimics of miR-27a and miR-451 increased MDR1 expression in the parental cells A2780. The sensitivity to and intracellular accumulation of cytotoxic drugs that are transported by P-glycoprotein were enhanced by the treatment with the antagomirs of miR-27a or miR-451. Our results demonstrate for the first time the roles of microRNAs in the regulation of drug resistance mediated by MDR1/P-glycoprotein, and suggest the potential for targeting miR-27a and miR-451 as a therapeutic strategy for modulating MDR in cancer cells.


Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica , MicroRNAs/fisiologia , Neoplasias/genética , Sequência de Bases , Western Blotting , Linhagem Celular Tumoral , Primers do DNA , Doxorrubicina/farmacologia , Humanos , MicroRNAs/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Rodamina 123/metabolismo
17.
Leuk Res ; 32(12): 1842-8, 2008 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-18614230

RESUMO

Allogeneic hematopoietic stem cell transplantation provides curative therapy for some patients with advanced hematologic malignancies. Disease response after allogeneic transplant is, at least in part, mediated by donor immune cells. In this report we describe a cellular therapy using haploidentical peripheral blood stem cells administered after very low dose total body irradiation (TBI) (100cGy). The donor cells were anticipated to be rejected, so no graft-versus-host (GVHD) prophylaxis was used. Patients with persistent disease beyond 8 weeks could be further treated with infusions of irradiated haploidentical donor cells. Of the 10 patients enrolled in the study, durable engraftment of allogeneic cells was seen in one patient. Two patients with resistant relapsed acute myelogenous leukemia (AML) had a disease response. Analysis of T cell reactivity from one patient who achieved a complete response but did not have durable engraftment of donor cells indicated that disease response was associated with the generation of host-derived anti-leukemic cytotoxic CD8+ T cells that reacted with an AML-associated proteinase 3 epitope. Results from this patient suggest that allogeneic therapy induced a host anti-tumor response associated with cytotoxic T cells reactive with a low affinity self-antigen.


Assuntos
Neoplasias Hematológicas/cirurgia , Idoso , Idoso de 80 Anos ou mais , Antígenos CD/sangue , Antígenos CD34/sangue , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Complexo CD3/sangue , Linfócitos T CD8-Positivos/imunologia , Transplante de Células , Feminino , Citometria de Fluxo , Neoplasias Hematológicas/imunologia , Humanos , Leucemia Linfocítica Crônica de Células B/imunologia , Leucemia Linfocítica Crônica de Células B/cirurgia , Leucemia Mieloide Aguda/imunologia , Leucemia Mieloide Aguda/cirurgia , Linfoma não Hodgkin/imunologia , Linfoma não Hodgkin/cirurgia , Masculino , Pessoa de Meia-Idade , Projetos Piloto , Doadores de Tecidos , Expansão de Tecido/métodos , Coleta de Tecidos e Órgãos/métodos , Transplante Homólogo
18.
Cancer Res ; 68(11): 4331-9, 2008 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-18519693

RESUMO

Carcinoma-associated fibroblasts (CAF) have recently been implicated in important aspects of epithelial solid tumor biology, such as neoplastic progression, tumor growth, angiogenesis, and metastasis. However, neither the source of CAFs nor the differences between CAFs and fibroblasts from nonneoplastic tissue have been well defined. In this study, we show that human bone marrow-derived mesenchymal stem cells (hMSCs) exposed to tumor-conditioned medium (TCM) over a prolonged period of time assume a CAF-like myofibroblastic phenotype. More importantly, these cells exhibit functional properties of CAFs, including sustained expression of stromal-derived factor-1 (SDF-1) and the ability to promote tumor cell growth both in vitro and in an in vivo coimplantation model, and expression of myofibroblast markers, including alpha-smooth muscle actin and fibroblast surface protein. hMSCs induced to differentiate to a myofibroblast-like phenotype using 5-azacytidine do not promote tumor cell growth as efficiently as hMSCs cultured in TCM nor do they show increased SDF-1 expression. Furthermore, gene expression profiling revealed similarities between TCM-exposed hMSCs and CAFs. Taken together, these data suggest that hMSCs are a source of CAFs and can be used in the modeling of tumor-stroma interactions. To our knowledge, this is the first report showing that hMSCs become activated and resemble carcinoma-associated myofibroblasts on prolonged exposure to conditioned medium from MDAMB231 human breast cancer cells.


Assuntos
Diferenciação Celular , Células-Tronco Mesenquimais/citologia , Neoplasias/patologia , Animais , Sequência de Bases , Divisão Celular , Linhagem Celular Tumoral , Meios de Cultivo Condicionados , Primers do DNA , Fibroblastos/citologia , Imunofluorescência , Humanos , Camundongos , Camundongos Nus , Reação em Cadeia da Polimerase Via Transcriptase Reversa
19.
J Biol Chem ; 281(14): 9719-27, 2006 Apr 07.
Artigo em Inglês | MEDLINE | ID: mdl-16443928

RESUMO

Overexpression of extracellular matrix metalloproteinase inducer (EMMPRIN or CD147), a member of the immunoglobulin family and a glycoprotein enriched on the surface of tumor cells, promotes invasion, metastasis, and growth and survival of malignant cells and confers resistance to some chemotherapeutic drugs. However, the molecular mechanisms underlying the actions of EMMPRIN are not fully understood. In this study we sought to determine whether EMMPRIN contributes to the malignant phenotype of breast cancer by inhibiting anoikis, a form of apoptosis induced by loss or alteration of cell-cell or cell-matrix anchorage, and to explore the signaling pathways involved. We found that in the absence of attachment, human breast carcinoma cells expressing high levels of EMMPRIN formed less compact aggregates with larger surface area and less fibronectin matrix assembly, had higher viability, and were resistant to anoikis. Knockdown of EMMPRIN expression by RNA interference (small interfering RNA or short hairpin RNA) sensitized cancer cells to anoikis, as demonstrated by activation of caspase-3, increased DNA fragmentation, and decreased cellular viability. Furthermore, we observed that the accumulation of Bim, a proapoptotic BH3-only protein, was reduced in EMMPRIN-expressing cells and that silencing of EMMPRIN expression elevated Bim protein levels and enhanced cellular sensitivity to anoikis. Treatment of cells with a MEK inhibitor (U0126) or proteasome inhibitor (epoxomicin) also up-regulated Bim accumulation and rendered cells more sensitive to anoikis. These results indicated that expression of EMMPRIN protects cancer cells from anoikis and that this effect is mediated at least in part by a MAP kinase-dependent reduction of Bim. Because anoikis deficiency is a key feature of neoplastic transformation and invasive growth of epithelial cancer cells, our study on the role of EMMPRIN in anoikis resistance and the mechanism involved underscores the potential of EMMPRIN expression as a prognostic marker and novel target for cancer therapy.


Assuntos
Anoikis/fisiologia , Proteínas Reguladoras de Apoptose/metabolismo , Basigina/fisiologia , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Proteínas de Membrana/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Reguladoras de Apoptose/biossíntese , Basigina/biossíntese , Proteína 11 Semelhante a Bcl-2 , Caspase 3 , Caspases/metabolismo , Sobrevivência Celular , Dano ao DNA , Regulação para Baixo , Ativação Enzimática , Matriz Extracelular , Feminino , Perfilação da Expressão Gênica , Inativação Gênica , Humanos , Proteínas de Membrana/biossíntese , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Fenótipo , Prognóstico , Proteínas Proto-Oncogênicas/biossíntese , Interferência de RNA
20.
Exp Hematol ; 33(11): 1337-47, 2005 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-16263418

RESUMO

Mantle cell lymphoma (MCL) is a distinct form of non-Hodgkin's lymphoma (NHL) derived from CD5+ B cells. MCL cells overexpress cyclin D1 as a consequence of translocation of the gene into the immunoglobulin heavy-chain gene locus. MCL is an aggressive form of NHL with frequent relapses after standard-dose chemotherapy. In this context, a variety of novel therapies for patients with MCL have been investigated. In this study, we use an expanded panel of attenuated adenoviruses to study adenovirus-mediated cytotoxicity of MCL cells. Our results demonstrate: 1) adenovirus infection of MCL cells despite the absence of receptor/coreceptor molecules known to be important for adenovirus infection of other cells types; 2) cytotoxicity of MCL cells after infection with specific adenovirus mutants; 3) a high degree of cytotoxicity after infection of some patient samples with viruses lacking the E1B 19k "antiapoptotic" gene; and 4) cytotoxicity after infection with viruses containing mutations in E1A pRb or p300 binding. The extent of cytotoxicity with the panel of viruses demonstrated interpatient variability, but 100% cytotoxicity, as determined by molecular analysis, was detected in some samples. These studies provide the foundation for: 1) the development of adenoviruses as cytotoxic agents for MCL and 2) analyses of key regulatory pathways operative in MCL cells.


Assuntos
Infecções por Adenoviridae , Adenoviridae/patogenicidade , Terapia Biológica/métodos , Linfoma de Célula do Manto/terapia , Adenoviridae/genética , Proteínas E1A de Adenovirus/genética , Proteínas E1B de Adenovirus/deficiência , Proteínas E1B de Adenovirus/genética , Morte Celular , Humanos , Linfoma de Célula do Manto/patologia , Mutação , Vacinas Atenuadas/farmacologia , Virulência/genética
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