RESUMO
BACKGROUND: Liquid biopsy has the potential to monitor biological effects of treatment. KRAS represents the most commonly mutated oncogene in Caucasian non-small-cell lung cancer (NSCLC). The aim of this study was to explore association of dynamic plasma KRAS genotyping with outcome in advanced NSCLC patients. METHODS: Advanced NSCLC patients were prospectively enrolled. Plasma samples were collected at baseline (T1), after 3 or 4 weeks, according to treatment schedule (T2) and at first radiological restaging (T3). Patients carrying KRAS mutation in tissue were analysed in plasma with droplet digital PCR. Semi-quantitative index of fractional abundance of mutated allele (MAFA) was used. RESULTS: KRAS-mutated cohort included 58 patients, and overall 73 treatments (N = 39 chemotherapy and N = 34 immune checkpoint inhibitors) were followed with longitudinal liquid biopsy. Sensitivity of KRAS detection in plasma at baseline was 48.3% (95% confidence interval (CI): 35.0-61.8). KRAS mutation at T2 was associated with increased probability of experiencing progressive disease as best radiological response (adjusted odds ratio: 7.3; 95% CI: 2.1-25.0, p = 0.0016). Increased MAFA (T1-T2) predicted shorter progression-free survival (adjusted hazard ratio (HR): 2.1; 95% CI: 1.2-3.8, p = 0.0142) and overall survival (adjusted HR: 3.2; 95% CI: 1.2-8.4, p = 0.0168). CONCLUSIONS: Longitudinal analysis of plasma KRAS mutations correlated with outcome: its early assessment during treatment has great potentialities for monitoring treatment outcome in NSCLC patients.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Ácidos Nucleicos Livres/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Idoso , Biomarcadores Tumorais/genética , Carcinoma Pulmonar de Células não Pequenas/sangue , Carcinoma Pulmonar de Células não Pequenas/patologia , Estudos de Coortes , Progressão da Doença , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Mutação/genética , Estadiamento de NeoplasiasRESUMO
Despite substantial progress in treatment of T-cell acute lymphoblastic leukemia (T-ALL), mortality remains relatively high, mainly due to primary or acquired resistance to chemotherapy. Further improvements in survival demand better understanding of T-ALL biology and development of new therapeutic strategies. The Notch pathway has been involved in the pathogenesis of this disease and various therapeutic strategies are currently under development, including selective targeting of NOTCH receptors by inhibitory antibodies. We previously demonstrated that the NOTCH1-specific neutralizing antibody OMP52M51 prolongs survival in TALL patient-derived xenografts bearing NOTCH1/FBW7 mutations. However, acquired resistance to OMP52M51 eventually developed and we used patient-derived xenografts models to investigate this phenomenon. Multi-level molecular characterization of T-ALL cells resistant to NOTCH1 blockade and serial transplantation experiments uncovered heterogeneous types of resistance, not previously reported with other Notch inhibitors. In one model, resistance appeared after 156 days of treatment, it was stable and associated with loss of Notch inhibition, reduced mutational load and acquired NOTCH1 mutations potentially affecting the stability of the heterodimerization domain. Conversely, in another model resistance developed after only 43 days of treatment despite persistent down-regulation of Notch signaling and it was accompanied by modulation of lipid metabolism and reduced surface expression of NOTCH1. Our findings shed light on heterogeneous mechanisms adopted by the tumor to evade NOTCH1 blockade and support clinical implementation of antibody-based target therapy for Notch-addicted tumors.
Assuntos
Leucemia-Linfoma Linfoblástico de Células T Precursoras , Animais , Humanos , Camundongos Endogâmicos NOD , Camundongos SCID , Leucemia-Linfoma Linfoblástico de Células T Precursoras/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Receptor Notch1/genética , Transdução de Sinais , Linfócitos T , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: miR-182-5p (miR-182) is an oncogenic microRNA (miRNA) found in different tumor types and one of the most up-regulated miRNA in colorectal cancer (CRC). Although this microRNA is expressed in the early steps of tumor development, its role in driving tumorigenesis is unclear. METHODS: The effects of miR-182 silencing on transcriptomic profile were investigated using two CRC cell lines characterized by different in vivo biological behavior, the MICOL-14h-tert cell line (dormant upon transfer into immunodeficient hosts) and its tumorigenic variant, MICOL-14tum. Apoptosis was studied by annexin/PI staining and cleaved Caspase-3/PARP analysis. The effect of miR-182 silencing on the tumorigenic potential was addressed in a xenogeneic model of MICOL-14tum transplant. RESULTS: Endogenous miR-182 expression was higher in MICOL-14tum than in MICOL-14h-tert cells. Interestingly, miR-182 silencing had a strong impact on gene expression profile, and the positive regulation of apoptotic process was one of the most affected pathways. Accordingly, annexin/PI staining and caspase-3/PARP activation demonstrated that miR-182 treatment significantly increased apoptosis, with a prominent effect in MICOL-14tum cells. Moreover, a significant modulation of the cell cycle profile was exerted by anti-miR-182 treatment only in MICOL-14tum cells, where a significant increase in the fraction of cells in G0/G1 phases was observed. Accordingly, a significant growth reduction and a less aggressive histological aspect were observed in tumor masses generated by in vivo transfer of anti-miR-182-treated MICOL-14tum cells into immunodeficient hosts. CONCLUSIONS: Altogether, these data indicate that increased miR-182 expression may promote cell proliferation, suppress the apoptotic pathway and ultimately confer aggressive traits on CRC cells.
Assuntos
Apoptose/genética , Carcinogênese/genética , Neoplasias Colorretais/genética , Inativação Gênica , MicroRNAs/genética , Animais , Células CACO-2 , Ciclo Celular/genética , Neoplasias Colorretais/patologia , Modelos Animais de Doenças , Regulação Neoplásica da Expressão Gênica , Células HT29 , Xenoenxertos , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Transfecção , Carga Tumoral/genética , Regulação para Cima/genéticaRESUMO
Loss-of-function mutations and deletions in Wilms tumor 1 (WT1) gene are present in approximately 10% of T-cell acute lymphoblastic leukemia. Clinically, WT1 mutations are enriched in relapsed series and are associated to inferior relapse-free survival in thymic T-cell acute lymphoblastic leukemia cases. Here we demonstrate that WT1 plays a critical role in the response to DNA damage in T-cell leukemia. WT1 loss conferred resistance to DNA damaging agents and attenuated the transcriptional activation of important apoptotic regulators downstream of TP53 in TP53-competent MOLT4 T-leukemia cells but not in TP53-mutant T-cell acute lymphoblastic leukemia cell lines. Notably, WT1 loss positively affected the expression of the X-linked inhibitor of apoptosis protein, XIAP, and genetic or chemical inhibition with embelin (a XIAP inhibitor) significantly restored sensitivity to γ-radiation in both T-cell acute lymphoblastic leukemia cell lines and patient-derived xenografts. These results reveal an important role for the WT1 tumor suppressor gene in the response to DNA damage, and support the view that anti-XIAP targeted therapies could have a role in the treatment of WT1-mutant T-cell leukemia.
Assuntos
Dano ao DNA/efeitos dos fármacos , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Proteína Supressora de Tumor p53/efeitos dos fármacos , Proteínas WT1/deficiência , Animais , Linhagem Celular Tumoral , Relação Dose-Resposta à Radiação , Raios gama , Xenoenxertos , Humanos , Proteínas Inibidoras de Apoptose/antagonistas & inibidores , Proteínas Inibidoras de Apoptose/fisiologia , Camundongos SCID , Leucemia-Linfoma Linfoblástico de Células T Precursoras/radioterapia , Proteína Supressora de Tumor p53/fisiologia , Proteínas WT1/fisiologiaRESUMO
Due to their ability to differentiate into various cell types and to support tissue regeneration, stem cells simultaneously became the holy grail of regenerative medicine and the evil obstacle in cancer therapy. Several studies have investigated niche-related conditions that favor stemness properties and increasingly emphasized their association with an inflammatory environment. Tumor-associated macrophages (TAMs) and myeloid-derived suppressor cells (MDSCs) are major orchestrators of cancer-related inflammation, able to dynamically express different polarized inflammatory programs that promote tumor outgrowth, including tumor angiogenesis, immunosuppression, tissue remodeling and metastasis formation. In addition, these myeloid populations support cancer cell stemness, favoring tumor maintenance and progression, as well as resistance to anticancer treatments. Here, we discuss inflammatory circuits and molecules expressed by TAMs and MDSCs as guiding forces of cancer cell stemness.
Assuntos
Imunoterapia/métodos , Inflamação/imunologia , Macrófagos/imunologia , Células Supressoras Mieloides/imunologia , Neoplasias/imunologia , Células-Tronco Neoplásicas/fisiologia , Animais , Carcinogênese , Regulação Neoplásica da Expressão Gênica , Humanos , Tolerância Imunológica , Inflamação/terapia , Neoplasias/terapia , Microambiente TumoralRESUMO
BACKGROUND: Inference of gene regulation from expression data may help to unravel regulatory mechanisms involved in complex diseases or in the action of specific drugs. A challenging task for many researchers working in the field of systems biology is to build up an experiment with a limited budget and produce a dataset suitable to reconstruct putative regulatory modules worth of biological validation. RESULTS: Here, we focus on small-scale gene expression screens and we introduce a novel experimental set-up and a customized method of analysis to make inference on regulatory modules starting from genetic perturbation data, e.g. knockdown and overexpression data. To illustrate the utility of our strategy, it was applied to produce and analyze a dataset of quantitative real-time RT-PCR data, in which interferon-α (IFN-α) transcriptional response in endothelial cells is investigated by RNA silencing of two candidate IFN-α modulators, STAT1 and IFIH1. A putative regulatory module was reconstructed by our method, revealing an intriguing feed-forward loop, in which STAT1 regulates IFIH1 and they both negatively regulate IFNAR1. STAT1 regulation on IFNAR1 was object of experimental validation at the protein level. CONCLUSIONS: Detailed description of the experimental set-up and of the analysis procedure is reported, with the intent to be of inspiration for other scientists who want to realize similar experiments to reconstruct gene regulatory modules starting from perturbations of possible regulators. Application of our approach to the study of IFN-α transcriptional response modulators in endothelial cells has led to many interesting novel findings and new biological hypotheses worth of validation.
Assuntos
Redes Reguladoras de Genes , Interferon-alfa/genética , Interferência de RNA , RNA Helicases DEAD-box/genética , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Helicase IFIH1 Induzida por Interferon , Modelos Genéticos , Receptor de Interferon alfa e beta/genética , Fator de Transcrição STAT1/genéticaRESUMO
Activation of the NOTCH pathway occurs commonly in T acute lymphoblastic leukemia (T-ALL) mainly due to mutations in NOTCH1 or alterations in FBW7 and is involved in the regulation of cell proliferation and survival. Since mutations hit different domains of the receptor, they are predicted to heterogeneously perturb ligand-induced NOTCH1 activity. Moreover, T-ALL cells also co-express NOTCH3 receptors which could be triggered by different ligands. In this study, we aimed to investigate the role of DLL4 in the regulation of NOTCH signaling in T-ALL cells in the context of different types of NOTCH1 mutation or wild-type NOTCH receptor, as well as the effects of DLL4 neutralization on T-ALL engraftment in mice. We found that NOTCH signaling can be stimulated in T-ALL cells in vitro by either human or murine DLL4 with heterogeneous effects, according to NOTCH1/FBW7 mutation status, and that these effects can be blocked by antibodies neutralizing DLL4, NOTCH1 or NOTCH2/3. In vivo, DLL4 is expressed in the spleen and the bone marrow (BM) of NOD/SCID mice bearing T-ALL xenografts as well as the BM of T-ALL patients. Importantly, DLL4 blockade impaired growth of T-ALL cells in NOD/SCID mice and increased leukemia cell apoptosis. These results show that DLL4 is an important component of the tumor microenvironment which contributes to the early steps of T-ALL cell growth.
Assuntos
Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas de Membrana/metabolismo , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patologia , Receptores Notch/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Animais , Apoptose , Western Blotting , Proteínas de Ligação ao Cálcio , Proliferação de Células , Citometria de Fluxo , Imunofluorescência , Regulação Neoplásica da Expressão Gênica , Humanos , Técnicas Imunoenzimáticas , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/metabolismo , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Receptores Notch/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoAssuntos
Imunoterapia , Neoplasias/terapia , Humanos , Imunoterapia/métodos , Itália , Neoplasias/imunologiaRESUMO
OBJECTIVE: At present, no consensus exists on the beneficial effect of preoperative cisplatin/5-fluorouracil (5-FU)-based chemotherapy versus primary surgery in the management of patients with esophageal cancer. The aim of this study was to evaluate the impact of some relevant genetic polymorphisms, within drug-related and DNA repair genes, on the clinical outcome of esophageal cancer patients subjected to cisplatin/5-FU-based neoadjuvant treatment. METHODS: DNA from 143 esophageal cancer patients, 63 receiving neoadjuvant therapy and 80 receiving primary surgery, was analyzed for the following polymorphisms: the GSTM1 null, GSTT1 null, and GSTP1 Ile105Val (rs16953) in glutathione S-transferase (GST) family, 2 in thymidylate synthase (TS) gene, and the ERCC1 Asn118Asn (rs11615), ERCC1 C8092A (rs3212986), XPD/ERCC2 Asp312Asn (rs1799793), and XPD/ERCC2 Lys751Gln (rs13181) of the nucleotide excision repair pathway. RESULTS: We found that the ERCC1 rs3212986, although not associated with therapeutic response, is an independent predictive marker of better outcome in a cisplatin/5-FU-based neoadjuvant setting (hazard ratio: 0.38, 95% confidence interval: 0.2-0.73, P=0.008). In contrast, no association with clinical outcome was observed for this polymorphism in the primary surgery group. CONCLUSION: Our study indicates the ERCC1 rs3212986 as a predictive marker in the cisplatin/5-FU-based neoadjuvant setting, and also suggests its use as a marker to select the appropriate therapeutic approach in esophageal cancer patients.
Assuntos
Carcinoma de Células Escamosas/genética , Cisplatino/administração & dosagem , Proteínas de Ligação a DNA/genética , Endonucleases/genética , Neoplasias Esofágicas/genética , Fluoruracila/administração & dosagem , Glutationa Transferase/genética , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/genética , Adenocarcinoma/cirurgia , Adulto , Idoso , Carcinoma de Células Escamosas/tratamento farmacológico , Carcinoma de Células Escamosas/cirurgia , Cisplatino/uso terapêutico , Neoplasias Esofágicas/tratamento farmacológico , Neoplasias Esofágicas/cirurgia , Feminino , Fluoruracila/uso terapêutico , Marcadores Genéticos , Genótipo , Glutationa S-Transferase pi/genética , Humanos , Masculino , Pessoa de Meia-Idade , Terapia de Alvo Molecular , Terapia Neoadjuvante , Polimorfismo Genético , Timidilato Sintase/genética , Resultado do Tratamento , Proteína Grupo D do Xeroderma Pigmentoso/genéticaRESUMO
Ontogenesis of T cells in the thymus is a complex process whose molecular control is poorly understood. The present study investigated microRNAs involved in human thymocyte differentiation by comparing the microRNA expression profiles of thymocytes at the double-positive, single-positive CD4(+) and single-positive CD8(+) maturation stages. Microarray analysis showed that each thymocyte population displays a distinct microRNA expression profile that reflects their developmental relationships. Moreover, analysis of small-RNA libraries generated from human unsorted and double-positive thymocytes and from mature peripheral CD4(+) and CD8(+) T lymphocytes, together with the microarray data, indicated a trend toward up-regulation of microRNA expression during T-cell maturation after the double-positive stage and revealed a group of microRNAs regulated during normal T-cell development, including miR-150, which is strongly up-regulated as maturation progresses. We showed that miR-150 targets NOTCH3, a member of the Notch receptor family that plays important roles both in T-cell differentiation and leukemogenesis. Forced expression of miR-150 reduces NOTCH3 levels in T-cell lines and has adverse effects on their proliferation and survival. Overall, these findings suggest that control of the Notch pathway through miR-150 may have an important impact on T-cell development and physiology.
Assuntos
Diferenciação Celular , Regulação da Expressão Gênica , MicroRNAs/metabolismo , Receptores Notch/metabolismo , Linfócitos T/citologia , Linfócitos T/metabolismo , Regiões 3' não Traduzidas , Adulto , Apoptose , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células , Células Cultivadas , Pré-Escolar , Perfilação da Expressão Gênica , Genes Reporter , Humanos , Lactente , Recém-Nascido , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/metabolismo , RNA Mensageiro/metabolismo , Receptor Notch3 , Receptores Notch/antagonistas & inibidores , Receptores Notch/genética , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/metabolismo , Timo/citologia , Timo/metabolismoRESUMO
Nonconsensual distribution of intimate images (NCII), also known as revenge porn, has become a significant social issue in recent years, with severe consequences for victims. The present study aimed to investigate the prevalence and predictors of NCII victimization among young Italian adults, focusing on the role of sexting, intimate partner violence (IPV), impulsivity, and self-disclosure. An online survey was conducted among a sample of 2047 Italian young adults (F = 29.3 %, M = 53.4 %, Not Indicated=16.9 %; mean age = 24.4, SD = 4.4) using a convenience sample recruited through internet. The survey included questions on NCII victimization, sexting behavior, sextortion, and IPV. Our findings showed that 33.9 % of respondents reported engaging in sexting behavior, with females being three times more likely to engage in sexting than males. Furthermore, 3.3 % of participants reported being victims of NCII, with over one-third of victims experiencing three or more types of NCII victimization. Multiple regression analysis revealed that sexting and IPV were significant predictors of NCII victimization, and the interaction effect between self-disclosure and impulsivity was also a significant predictor. This study contributes to the understanding of NCII in Italy and highlights the need for interventions and prevention strategies to address both NCII and IPV, given their strong continuity. The results also suggest that the relationship between self-disclosure, impulsivity, IPV, and NCII victimization is complex and requires further investigation, suggesting a scenario where the climate of violence may impair the victim's decision-making.
RESUMO
Increased Notch1 activity has been observed in intestinal tumours, partially accomplished by ß-catenin-mediated up-regulation of the Notch ligand Jagged-1. Whether further mechanisms of Notch activation exist and other Notch receptors might be involved is unclear. Microarray data indicated that Notch3 transcript levels are significantly up-regulated in primary and metastatic CRC samples compared to normal mucosa. Moreover, Notch3 protein was expressed at strong/moderate levels by 19.7% of 158 CRC samples analysed, and at weak levels by 51.2% of the samples. Intrigued by these findings, we sought to investigate whether Notch3 modulates oncogenic features of CRC cells. By exploiting xenografts of CRC cells with different tumourigenic properties in mice, we found that the aggressive phenotype was associated with altered expression of components of the Notch pathway, including Notch3, Delta-like 4 (DLL4), and Jagged-1 ligands. Stimulation with immobilized recombinant DLL4 or transduction with DLL4-expressing vectors dramatically increased Notch3 expression in CRC cells, associated with accelerated tumour growth. Forced expression of an active form of Notch3 mirrored the effects of DLL4 stimulation and increased tumour formation. Conversely, attenuation of Notch3 levels by shRNA resulted in perturbation of the cell cycle followed by reduction in cell proliferation, clonogenic capacity, and inhibition of tumour growth. Altogether, these findings indicate that Notch3 can modulate the tumourigenic properties of CRC cells and contributes to sustained Notch activity in DLL4-expressing tumours.
Assuntos
Neoplasias Colorretais/patologia , Receptores Notch/fisiologia , Proteínas Adaptadoras de Transdução de Sinal , Animais , Proteínas de Ligação ao Cálcio/metabolismo , Transformação Celular Neoplásica/metabolismo , Neoplasias Colorretais/metabolismo , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteína Jagged-1 , Masculino , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Nus , Camundongos SCID , Proteínas de Neoplasias/metabolismo , Proteínas de Neoplasias/fisiologia , Transplante de Neoplasias , Receptor Notch3 , Receptores Notch/metabolismo , Proteínas Serrate-Jagged , Transdução de Sinais/fisiologia , Transplante Heterólogo , Células Tumorais Cultivadas , Regulação para CimaRESUMO
Although adoptive immunotherapy with CD8(+) CTL is providing clinically relevant results against EBV-driven malignancies, the effector role of CD4(+) T cells has been poorly investigated. We addressed this issue in a lymphoblastoid cell line-induced mouse model of posttransplant lymphoproliferative disease (PTLD) by comparing the therapeutic efficacy of EBV-specific CD4(+) and CD8(+) T cell lines upon adoptive transfer. CD4(+) T cells disclosed a long-lasting and stronger proliferative potential than CD8(+) T cells, had a similar activation and differentiation marker profile, efficiently killed their targets in a MHC class II-restricted manner, and displayed a lytic machinery comparable to that of cognate CD8(+) T cells. A detailed analysis of Ag specificity revealed that CD4(+) T cells potentially target EBV early lytic cycle proteins. Nonetheless, when assessed for the relative therapeutic impact after in vivo transfer, CD4(+) T cells showed a reduced activity compared with the CD8(+) CTL counterpart. This feature was apparently due to a strong and selective downmodulation of MHC class II expression on the tumor cells surface, a phenomenon that could be reverted by the demethylating agent 5-aza-2'-deoxycytidine, thus leading to restoration of lymphoblastoid cell line recognition and killing by CD4(+) T cells, as well as to a more pronounced therapeutic activity. Conversely, immunohistochemical analysis disclosed that HLA-II expression is fully retained in human PTLD samples. Our data indicate that EBV-specific cytotoxic CD4(+) T cells are therapeutic in mice bearing PTLD-like tumors, even in the absence of CD8(+) T cells. These findings pave the way to use cultures of pure CD4(+) T cells in immunotherapeutic approaches for EBV-related malignancies.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/transplante , Testes Imunológicos de Citotoxicidade , Epitopos de Linfócito T/imunologia , Infecções por Vírus Epstein-Barr/imunologia , Infecções por Vírus Epstein-Barr/terapia , Herpesvirus Humano 4/imunologia , Imunoterapia Adotiva , Animais , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/virologia , Callithrix , Linhagem Celular Transformada , Linhagem Celular Tumoral , Testes Imunológicos de Citotoxicidade/métodos , Regulação para Baixo/imunologia , Infecções por Vírus Epstein-Barr/patologia , Antígenos HLA-DR/biossíntese , Antígenos HLA-DR/imunologia , Antígenos HLA-DR/metabolismo , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Imunoterapia Adotiva/métodos , Linfoma de Células B/imunologia , Linfoma de Células B/patologia , Linfoma de Células B/terapia , Camundongos , Camundongos SCIDRESUMO
The peculiar site of development of primary effusion lymphoma (PEL) highlights a specific role of body cavities in the pathogenesis of this neoplasia. We used a xenograft murine model of PEL to characterize the contribution of the host microenvironment to PEL growth. The activity of a murine (ie, host-specific) interferon-alpha(1) (IFN-alpha(1))-expressing lentiviral vector (mIFN-alpha(1)-LV) was compared with that of a human (h) IFN-alpha(2)b-LV. LVs efficiently delivered the transgene to PEL cells and conferred long-term transgene expression in vitro and in vivo. Treatment of PEL-injected severe combined immunodeficiency mice with hIFN-alpha(2)b-LV significantly prolonged mice survival and reduced ascites development. Interestingly, mIFN-alpha(1)-LV showed an antineoplastic activity comparable with that observed with hIFN-alpha(2)b-LV. As mIFN-alpha(1) retained species-restricted activity in vitro, it probably acted in vivo on the intracavitary murine milieu. mIFN-alpha(1)-treated murine mesothelial cells were found to express tumor necrosis factor-related apoptosis-inducing ligand and to significantly trigger apoptosis of cocultured PEL cells in a tumor necrosis factor-related apoptosis-inducing ligand-dependent manner. These data suggest that the interaction between lymphomatous and mesothelial cells lining the body cavities may play a key role in PEL growth control and also indicate that the specific targeting of microenvironment may impair PEL development.
Assuntos
Antineoplásicos/uso terapêutico , Vetores Genéticos , Interferon-alfa/uso terapêutico , Lentivirus/genética , Linfoma de Efusão Primária/tratamento farmacológico , Animais , Células Cultivadas , Citocinas/metabolismo , Avaliação Pré-Clínica de Medicamentos , Feminino , Humanos , Interferon alfa-2 , Rim/citologia , Rim/efeitos dos fármacos , Rim/metabolismo , Linfoma de Efusão Primária/genética , Linfoma de Efusão Primária/patologia , Mesoderma/citologia , Mesoderma/efeitos dos fármacos , Mesoderma/metabolismo , Camundongos , Camundongos SCID , Peritônio/citologia , Peritônio/efeitos dos fármacos , Peritônio/metabolismo , Proteínas RecombinantesRESUMO
T-cell acute lymphoblastic leukemia (T-ALL) is a highly malignant pediatric leukemia, where few therapeutic options are available for patients which relapse. We find that therapeutic targeting of GLI transcription factors by GANT-61 is particularly effective against NOTCH1 unmutated T-ALL cells. Investigation of the functional role of GLI1 disclosed that it contributes to T-ALL cell proliferation, survival, and dissemination through the modulation of AKT and CXCR4 signaling pathways. Decreased CXCR4 signaling following GLI1 inactivation was found to be prevalently due to post-transcriptional mechanisms including altered serine 339 CXCR4 phosphorylation and cortactin levels. We also identify a novel cross-talk between GLI transcription factors and FOXC1. Indeed, GLI factors can activate the expression of FOXC1 which is able to stabilize GLI1/2 protein levels through attenuation of their ubiquitination. Further, we find that prolonged GLI1 deficiency has a double-edged role in T-ALL progression favoring disease dissemination through the activation of a putative AKT/FOXC1/GLI2 axis. These findings have clinical significance as T-ALL patients with extensive central nervous system dissemination show low GLI1 transcript levels. Further, T-ALL patients having a GLI2-based Hedgehog activation signature are associated with poor survival. Together, these findings support a rationale for targeting the FOXC1/AKT axis to prevent GLI-dependent oncogenic Hedgehog signaling.
Assuntos
Fatores de Transcrição Forkhead/metabolismo , Leucemia-Linfoma Linfoblástico de Células T Precursoras/metabolismo , Transdução de Sinais , Proteína GLI1 em Dedos de Zinco/metabolismo , Animais , Apoptose , Biópsia , Pontos de Checagem do Ciclo Celular , Biologia Computacional/métodos , Modelos Animais de Doenças , Progressão da Doença , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Inativação Gênica , Proteínas Hedgehog/metabolismo , Humanos , Imuno-Histoquímica , Camundongos , Mutação , Leucemia-Linfoma Linfoblástico de Células T Precursoras/diagnóstico , Leucemia-Linfoma Linfoblástico de Células T Precursoras/etiologia , Leucemia-Linfoma Linfoblástico de Células T Precursoras/mortalidade , Prognóstico , Ligação Proteica , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptor Notch1/genética , Receptor Notch1/metabolismo , Receptores CXCR4/metabolismo , Fatores de TranscriçãoRESUMO
Several studies strongly suggest that DC differentiated in vitro in the presence of type I IFN acquire more potent immune stimulatory properties, compared with DC differentiated in vitro with IL-4. However, little is known about the molecular mechanisms underlying this phenomenon. To address this question, we compared the Ag-processing machinery (APM) profile in human DC grown in the presence of IFN-alpha ((IFN)DC) or IL-4 ((IL-4)DC). Using a panel of APM component-specific mAb in Western blot experiments, we found that (IFN)DC preferentially express inducible proteasome subunits (LMP2, LMP7, and MECL1) both at immature and mature stages. In contrast, immature (IL-4)DC co-express both constitutive (beta1, beta2, and beta5) and inducible subunits, as shown by Western blotting analysis. In addition, immature (IFN)DC express higher levels of TAP1, TAP2, calnexin, calreticulin, tapasin, and HLA class I molecules than (IL-4)DC. The different proteasome profiles of (IFN)DC and (IL-4)DC were associated with a greater ability of (IFN)DC to present an immunodominant epitope that requires LMP7 expression for its processing. In general, these data show the impact of cytokines on APM component expression and hence the Ag-processing ability of DC.
Assuntos
Diferenciação Celular/imunologia , Monócitos/enzimologia , Complexos Multienzimáticos/biossíntese , Complexo de Endopeptidases do Proteassoma/biossíntese , Transportadores de Cassetes de Ligação de ATP/efeitos dos fármacos , Transportadores de Cassetes de Ligação de ATP/imunologia , Transportadores de Cassetes de Ligação de ATP/metabolismo , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Epitopos/imunologia , Humanos , Fatores Imunológicos/farmacologia , Interferon-alfa/farmacologia , Interleucina-4/farmacologia , Proteínas Sensoras de Cálcio Intracelular/efeitos dos fármacos , Proteínas Sensoras de Cálcio Intracelular/imunologia , Proteínas Sensoras de Cálcio Intracelular/metabolismo , Proteínas de Membrana Transportadoras/efeitos dos fármacos , Proteínas de Membrana Transportadoras/imunologia , Proteínas de Membrana Transportadoras/metabolismo , Monócitos/imunologia , Complexos Multienzimáticos/efeitos dos fármacos , Complexo de Endopeptidases do Proteassoma/efeitos dos fármacosAssuntos
Antineoplásicos/farmacologia , Linfoma de Efusão Primária/tratamento farmacológico , Proteínas de Neoplasias/antagonistas & inibidores , RNA Interferente Pequeno/farmacologia , Proteínas Repressoras/antagonistas & inibidores , Linhagem Celular Tumoral , Feminino , Humanos , Lipossomos , Linfoma de Efusão Primária/genética , Linfoma de Efusão Primária/metabolismo , Masculino , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Fator 1 de Ligação ao Domínio I Regulador Positivo , RNA Interferente Pequeno/genética , Proteínas Repressoras/biossínteseRESUMO
Aim: Clinical features of esophageal cancer (EC) patients have poor prognostic power. Thus, it is paramount to discover biomarkers that can allow a more accurate survival prediction. Methods: To detect genetic variants associated with survival, DNA from 120 patients treated with cisplatin-based neoadjuvant therapy were genotyped using drug metabolism enzymes and transporters array. Results: We identified two variants: the rs2038067 in PPARD (p = 0.0004) and the rs683369 (F160L) in SLC22A1 (p = 0.001). Their prognostic power was greater than that of clinical stage alone (p = 0.017) and comparable to that of response to neoadjuvant therapy (p = 0.71). Interestingly, the prognostic accuracy of response models increased significantly when genetic variables were included (p = 0.003). Conclusion: Our data, though preliminary, strengthen the potential utility of germline variants for a better-tailored management of EC patients.
Assuntos
Antineoplásicos/uso terapêutico , Cisplatino/uso terapêutico , Neoplasias Esofágicas/genética , Variação Genética/genética , Transportador 1 de Cátions Orgânicos/genética , PPAR delta/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias Esofágicas/tratamento farmacológico , Neoplasias Esofágicas/mortalidade , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Terapia Neoadjuvante/métodos , Taxa de Sobrevida/tendênciasRESUMO
Esophageal cancer (EC) is a highly aggressive tumor, and the current monitoring procedures are partially inadequate to evaluate treatment efficacy. The aim of this study was to investigate whether allelic imbalance analysis in liquid biopsy could be used as an additional tool to monitor tumor burden in EC patients. For this purpose, circulating cell-free DNA (cfDNA) from 52 patients with a locally advanced EC, which underwent neoadjuvant treatment and resection, was analyzed. Data from four representative longitudinally followed patients are also reported. Furthermore, 17 DNAs from formalin-fixed paraffin-embedded (FFPE) tumor samples were analyzed and compared to time-matched cfDNAs. To look for allelic imbalance, which is the main genetic alteration in both EC histotypes, we used a panel of five microsatellites (MSs) and three single-nucleotide polymorphisms (SNPs) near genes described as frequently altered. The Fisher exact and Mann-Whitney U tests were used to analyze categorical and continuous data, respectively. The correlation coefficient between cfDNA and FFPE-DNA was calculated with the Pearson's correlation test. We found that the selected tumor-related alterations are present in cfDNA of both adenocarcinoma (EADC) and squamous cell carcinoma (ESCC) with similar frequencies. The only exception were the MSs, one downstream and one upstream, of SMAD4 of which the loss was only observed in EADC (26 vs. 0%, P = 0.018). More interestingly, longitudinal studies disclosed that in patients with disease progression, tumor-related alterations were present in cfDNA before overt clinical or instrumental signs of relapse. In conclusion, our data indicate that the evaluation of tumor-related gene allelic imbalance in cfDNA might be a useful tool to complement the current monitoring procedures for EC patients and to guide their management.
RESUMO
Notwithstanding intensified therapy, a considerable fraction of T-cell acute lymphoblastic leukemia (T-ALL) patients face a dismal prognosis due to primary resistance to treatment and relapse, raising the need for more efficient and targeted therapies. Hedgehog (HH) signaling is a major developmental pathway frequently deregulated in cancer, for which a role in T-ALL is emerging. Mounting evidence suggests that ligand-independent activation of HH pathway occurs in cancer including T-ALL, emphasizing the necessity of dissecting the complex interplay between HH and other signaling pathways regulating activation. In this work, we present a therapeutically relevant crosstalk between HH signaling and the glucocorticoid receptor (NR3C1) pathway acting at the level of GLI1 transcription factor. GLI inhibitor GANT61 and dexamethasone were shown to exert a synergistic anti-leukemic effect in vitro in T-ALL cell lines and patient-derived xenografts. Mechanistically, dexamethasone-activated NR3C1 impaired GLI1 function by dynamically modulating the recruitment of PCAF acetyltransferase and HDAC1 deacetylase. Increased GLI1 acetylation was associated with compromised transcriptional activity and reduced protein stability. In summary, our study identifies a novel crosstalk between GLI1 and NR3C1 signaling pathway which could be exploited in HH-dependent malignancies to increase therapeutic efficacy.