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1.
Elife ; 82019 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-31268420

RESUMO

Adult neurogenesis in the hippocampus may represent a form of plasticity in brain functions including mood, learning and memory. However, mechanisms underlying neural stem/progenitor cells (NSPCs) proliferation are not well understood. We found that Agrin, a factor critical for neuromuscular junction formation, is elevated in the hippocampus of mice that are stimulated by enriched environment (EE). Genetic deletion of the Agrn gene in excitatory neurons decreases NSPCs proliferation and increases depressive-like behavior. Low-density lipoprotein receptor-related protein 4 (Lrp4), a receptor for Agrin, is expressed in hippocampal NSPCs and its mutation blocked basal as well as EE-induced NSPCs proliferation and maturation of newborn neurons. Finally, we show that Lrp4 interacts with and activates receptor tyrosine kinase-like orphan receptor 2 (Ror2); and Ror2 mutation impairs NSPCs proliferation. Together, these observations identify a role of Agrin-Lrp4-Ror2 signaling for adult neurogenesis, uncovering previously unexpected functions of Agrin and Lrp4 in the brain.


Assuntos
Agrina/metabolismo , Hipocampo/crescimento & desenvolvimento , Proteínas Relacionadas a Receptor de LDL/metabolismo , Células-Tronco Neurais/fisiologia , Neurogênese , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Transdução de Sinais , Agrina/deficiência , Animais , Proliferação de Células , Técnicas de Inativação de Genes , Proteínas Relacionadas a Receptor de LDL/deficiência , Camundongos , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/deficiência
2.
EBioMedicine ; 43: 211-224, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31085100

RESUMO

BACKGROUND: A newly developed drug trastuzumab emtansine (T-DM1) has improved the survival of breast cancer (BC) patients. Despite an impressive initial clinical response, a subgroup of patient develop resistance and present therapeutic challenges. The underlying resistance mechanisms are not fully investigated. We report that T-DM1 treatment modulates the expression of ROR1 (type 1 receptor tyrosine kinase-like orphan receptor) and induces self-renewal of cancer stem cells (CSCs) leading to therapeutic resistance. METHODS: Using BC patient tumor samples, and BC cell lines we gained insight into the T-DM1 treatment induced ROR1 overexpression and resistance. In vitro sphere forming assays and in vivo extreme dilution assays were employed to analyze the stemness and self-renewal capacity of the cells. A series of molecular expression and protein assays including qRT-PCR, FACS-sorting, ELISA, immunostaining, Western blotting were used to provide evidence. FINDINGS: Exposure of cells to T-DM1 shifted ROR1 expression from low to high, enriched within the CSC subpopulation, coincident with increased Bmi1 and stemness factors. T-DM1 induced ROR1 cells showed high spheroid and tumor forming efficiency in vitro and in an animal model exhibiting shorter tumor-free time. Mechanistically, the overexpression of ROR1 is partly induced by the activation of YAP1 and its target genes. Silencing of ROR1 and YAP1 by pharmacologic inhibitors and/or sh/siRNA inhibited spheroid formation, the initiation of tumors and the capacity for self-renewal and ROR1 overexpression. INTERPRETATIONS: The results presented here indicate that simultaneous targeting of ROR1 and YAP1 may suppress CSC self-renewal efficacy and inhibit tumor progression in BC. In this manner such treatments may overcome the T-DM1 mediated therapeutic resistance and improve clinical outcome. FUND: This study was supported by Neurogen Technologies for interdisciplinary research.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Antineoplásicos Imunológicos/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Imunoconjugados/farmacologia , Fosfoproteínas/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/genética , Receptor ErbB-2/antagonistas & inibidores , Biomarcadores , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/mortalidade , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Autorrenovação Celular/genética , Feminino , Citometria de Fluxo , Regulação Neoplásica da Expressão Gênica , Inativação Gênica , Humanos , Imuno-Histoquímica , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Prognóstico , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Fatores de Transcrição
3.
Mol Med Rep ; 19(6): 4663-4672, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30957191

RESUMO

The proliferation and migration of vascular smooth muscle cells (VSMCs) are major cellular events in hypertension­induced vascular remodeling, which is closely involved in the progression of atherosclerosis (AS). Although long non­coding RNAs (lncRNAs) are gaining recognition as novel regulators of VSMCs, their functioning and role in AS remain to be elucidated. In the present study, the role of lncRNA ENST00000430945 (lncRNA 430945) in AS was investigated. VSMCs transfected with a small interfering RNA (siRNA; si­430945) and a negative control (si­NC) were used. Cell Counting Kit­8, wound­healing and Transwell migration arrays were performed to determine whether lncRNA 430945 influenced VSMC proliferation and migration. Furthermore, the study examined whether a correlation exists between lncRNA 430945 and the receptor tyrosine kinase­like orphan receptor 2 (ROR2) signaling pathway. It was found that the expression of lncRNA 430945 was high in human AS tissues, which in turn promoted angiotensin II (AngII)­induced VSMC proliferation. Reverse transcription­quantitative polymerase chain reaction (RT­qPCR) and western blot analyses showed that lncRNA 430945 mediated the AngII­induced upregulation of ROR2. In addition, the microarray and RT­qPCR results showed that the expression of lncRNA 430945 was increased considerably in AS tissues. The downregulation of lncRNA 430945 significantly suppressed AngII­induced VSMC proliferation and migration. In addition, ROR2 levels in VSMCs transfected with si­430945 were markedly lower than those cells transfected with si­NC. Additionally, western blotting showed that lncRNA 430945 activated the signaling pathways associated with ROR2 and Ras homolog gene family member A (RhoA). The upregulation of lncRNA 430945 in AS promoted the proliferation and migration of VSMCs via activation of the ROR2/RhoA signaling pathway. Therefore, targeting ROR2 or RhoA may be a promising technique in developing therapeutic strategies for treating AS.


Assuntos
Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , RNA Longo não Codificante/genética , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/genética , Proteína rhoA de Ligação ao GTP/genética , Animais , Aterosclerose/genética , Aterosclerose/terapia , Movimento Celular , Proliferação de Células , Regulação da Expressão Gênica , Marcação de Genes , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Músculo Liso Vascular/citologia , Miócitos de Músculo Liso/citologia , RNA Longo não Codificante/metabolismo , RNA Interferente Pequeno/metabolismo , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Transdução de Sinais , Transfecção , Remodelação Vascular , Proteína rhoA de Ligação ao GTP/metabolismo
4.
Nature ; 567(7749): 540-544, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30867597

RESUMO

Diversity within or between tumours and metastases (known as intra-patient tumour heterogeneity) that develops during disease progression is a serious hurdle for therapy1-3. Metastasis is the fatal hallmark of cancer and the mechanisms of colonization, the most complex step in the metastatic cascade4, remain poorly defined. A clearer understanding of the cellular and molecular processes that underlie both intra-patient tumour heterogeneity and metastasis is crucial for the success of personalized cancer therapy. Here, using transcriptional profiling of tumours and matched metastases in patient-derived xenograft models in mice, we show cancer-site-specific phenotypes and increased glucocorticoid receptor activity in distant metastases. The glucocorticoid receptor mediates the effects of stress hormones, and of synthetic derivatives of these hormones that are used widely in the clinic as anti-inflammatory and immunosuppressive agents. We show that the increase in stress hormones during breast cancer progression results in the activation of the glucocorticoid receptor at distant metastatic sites, increased colonization and reduced survival. Our transcriptomics, proteomics and phospho-proteomics studies implicate the glucocorticoid receptor in the activation of multiple processes in metastasis and in the increased expression of kinase ROR1, both of which correlate with reduced survival. The ablation of ROR1 reduced metastatic outgrowth and prolonged survival in preclinical models. Our results indicate that the activation of the glucocorticoid receptor increases heterogeneity and metastasis, which suggests that caution is needed when using glucocorticoids to treat patients with breast cancer who have developed cancer-related complications.


Assuntos
Neoplasias da Mama/patologia , Glucocorticoides/efeitos adversos , Glucocorticoides/metabolismo , Metástase Neoplásica/patologia , Animais , Neoplasias da Mama/enzimologia , Linhagem Celular Tumoral , Dexametasona/efeitos adversos , Dexametasona/metabolismo , Progressão da Doença , Feminino , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/secundário , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Quinases/metabolismo , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Receptores de Glucocorticoides/metabolismo , Transdução de Sinais/efeitos dos fármacos , Taxa de Sobrevida
5.
Cancer Cell ; 35(3): 489-503.e8, 2019 03 18.
Artigo em Inglês | MEDLINE | ID: mdl-30889382

RESUMO

Many potential targets for CAR-T cells in solid tumors are expressed in some normal tissues, raising concern for off-tumor toxicity. Following lymphodepletion, CAR-T cells targeting the tumor-associated antigen ROR1 lysed tumors in mice but induced lethal bone marrow failure due to recognition of ROR1+ stromal cells. To improve selectivity, we engineered T cells with synthetic Notch (synNotch) receptors specific for EpCAM or B7-H3, which are expressed on ROR1+ tumor cells but not ROR1+ stromal cells. SynNotch receptors induced ROR1 CAR expression selectively within the tumor, resulting in tumor regression without toxicity when tumor cells were segregated from, but not when co-localized with, normal ROR1+ cells. This strategy, thus, permits safe targeting of tumors that are sufficiently separated from normal cells.


Assuntos
Imunoterapia Adotiva/métodos , Neoplasias/terapia , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/genética , Receptores de Antígenos Quiméricos/imunologia , Animais , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Humanos , Células K562 , Camundongos , Neoplasias/imunologia , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Linfócitos T/imunologia , Ensaios Antitumorais Modelo de Xenoenxerto
6.
Oncogene ; 38(26): 5142-5157, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30894682

RESUMO

The receptor tyrosine kinase-like orphan receptor 1 (ROR1) is a transcriptional target of the lineage-survival oncogene NKX2-1/TTF-1 in lung adenocarcinomas. In addition to its kinase-dependent role, ROR1 functions as a scaffold protein to facilitate interaction between caveolin-1 (CAV1) and CAVIN1, and consequently maintains caveolae formation, which in turn sustains pro-survival signaling toward AKT from multiple receptor tyrosine kinases (RTKs), including epidermal growth factor receptor (EGFR), MET (proto-oncogene, receptor tyrosine kinase), and IGF-IR (insulin-like growth factor receptor 1). Therefore, ROR1 is an attractive target for overcoming EGFR-TKI resistance due to various mechanisms such as EGFR T790M double mutation and bypass signaling from other RTKs. Here, we report that ROR1 possesses a novel scaffold function indispensable for efficient caveolae-dependent endocytosis. CAVIN3 was found to bind with ROR1 at a site distinct from sites for CAV1 and CAVIN1, a novel function required for proper CAVIN3 subcellular localization and caveolae-dependent endocytosis, but not caveolae formation itself. Furthermore, evidence of a mechanistic link between ROR1-CAVIN3 interaction and consequential caveolae trafficking, which was found to utilize a binding site distinct from those for ROR1 interactions with CAV1 and CAVIN1, with RTK-mediated pro-survival signaling towards AKT in early endosomes in lung adenocarcinoma cells was also obtained. The present findings warrant future study to enable development of novel therapeutic strategies for inhibiting the multifaceted scaffold functions of ROR1 in order to reduce the intolerable death toll from this devastating cancer.


Assuntos
Adenocarcinoma de Pulmão/patologia , Cavéolas/fisiologia , Endocitose , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Neoplasias Pulmonares/patologia , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/metabolismo , Animais , Células COS , Cavéolas/metabolismo , Sobrevivência Celular/genética , Células Cultivadas , Endocitose/genética , Células HEK293 , Células HeLa , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Ligação Proteica/fisiologia , Células Sf9 , Transdução de Sinais/genética , Spodoptera
7.
Cells ; 8(3)2019 03 02.
Artigo em Inglês | MEDLINE | ID: mdl-30832318

RESUMO

BACKGROUND: Hepatocellular carcinoma (HCC) is a common and deadly cancer; however, very little improvement has been made towards its diagnosis and prognosis. The expression and functional contribution of the receptor tyrosine kinase ROR1 have not been investigated in HCC before. Hence, we investigated the expression of ROR1 in HCC cells and assessed its involvement in hepatocarcinogenesis. METHODS: Recombinant bacterial ROR1 protein was used as an immunogen to generate ROR1 monoclonal antibodies. ROR1 transcript levels were detected by RT-qPCR and the protein expression of ROR1 in HCC was assessed by Western blotting by using homemade anti-ROR1 monoclonal antibodies. Apoptosis, cell cycle, trans-well migration, and drug efflux assays were performed in shRNA-ROR1 HCC cell clones to uncover the functional contribution of ROR1 to hepatocarcinogenesis. RESULTS: New ROR1 antibodies specifically detected endogenous ROR1 protein in human and mouse HCC cell lines. ROR1-knockdown resulted in decreased proliferation and migration but enhanced resistance to apoptosis and anoikis. The observed chemotherapy-resistant phenotype of ROR1-knockdown cells was due to enhanced drug efflux and increased expression of multi-drug resistance genes. CONCLUSIONS: ROR1 is expressed in HCC and contributes to disease development by interfering with multiple pathways. Acquired ROR1 expression may have diagnostic and prognostic value in HCC.


Assuntos
Carcinoma Hepatocelular/enzimologia , Neoplasias Hepáticas/enzimologia , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Animais , Anoikis/efeitos dos fármacos , Anoikis/genética , Anticorpos Monoclonais/metabolismo , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Apoptose/efeitos dos fármacos , Apoptose/genética , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Fase G1/efeitos dos fármacos , Fase G1/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Camundongos , Fenótipo , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genética
8.
Cell ; 177(1): 8, 2019 03 21.
Artigo em Inglês | MEDLINE | ID: mdl-30901551

RESUMO

Larotrectinib is a small-molecule kinase inhibitor that targets NTRK fusions that occur in multiple types of cancer. Its FDA approval represents the first instance of a treatment indication being designated "tumor-agnostic" from the outset, being based on actionable genomic insights. To view this Bench to Bedside, open or download the PDF.


Assuntos
Pirazóis/metabolismo , Pirazóis/farmacologia , Pirimidinas/metabolismo , Pirimidinas/farmacologia , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Humanos , Neoplasias/genética , Proteínas de Fusão Oncogênica/genética , Inibidores de Proteínas Quinases/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/agonistas , Receptor trkB/metabolismo
9.
Cancer Sci ; 110(4): 1306-1316, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30742741

RESUMO

Collective invasion is an important strategy of cancers of epithelial origin, including colorectal cancer (CRC), to infiltrate efficiently into local tissues as collective cell groups. Within the groups, cells at the invasive front, called leader cells, are highly polarized and motile, thereby providing the migratory traction that guides the follower cells. However, its underlying mechanisms remain unclear. We have previously shown that signaling emanating from the receptor tyrosine kinase Ror2 can promote invasion of human osteosarcoma cells and that intraflagellar transport 20 (IFT20) mediates its signaling to regulate Golgi structure and transport. Herein, we investigated the role of Ror2 and IFT20 in collective invasion of CRC cells, where Ror2 expression is either silenced or nonsilenced. We show by cell biological analyses that IFT20 promotes collective invasion of CRC cells, irrespective of expression and function of Ror2. Intraflagellar transport 20 is required for organization of Golgi-associated, stabilized microtubules, oriented toward the direction of invasion in leader cells. Our results also indicate that IFT20 promotes reorientation of the Golgi apparatus toward the front side of leader cells. Live cell imaging of the microtubule plus-end binding protein EB1 revealed that IFT20 is required for continuous polarized microtubule growth in leader cells. These results indicate that IFT20 plays an important role in collective invasion of CRC cells by regulating organization of Golgi-associated, stabilized microtubules and Golgi polarity in leader cells.


Assuntos
Proteínas de Transporte/metabolismo , Complexo de Golgi/metabolismo , Microtúbulos/metabolismo , Neoplasias/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Expressão Gênica , Humanos , Neoplasias/genética , Neoplasias/patologia , RNA Interferente Pequeno/genética , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/genética , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo
10.
Biofactors ; 45(3): 416-426, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30801854

RESUMO

The receptor-tyrosine-kinase (RTK)-like orphan receptor 1 (ROR1) is a transmembrane glycoprotein regarded as a tumor-associated antigen. ROR1 plays an important role in cancer development, but the detailed function of ROR1 in diffuse large B-cell lymphoma (DLBCL) remains unclear. In this study, we first detected ROR1 expression and evaluated the relationship between ROR1 expression and the clinicopathological characteristics of DLBCL patients. Next we employed shRNA-mediated knockdown of ROR1 in DLBCL cell line to explore the characteristics of ROR1 in DLBCL development both in vitro and in vivo. The results showed a significantly higher level of ROR1 in DLBCL tissues than in lymphatic hyperplasia tissues. High ROR1 expression was correlated with unfavorable prognosis in DLBCL patients. Furthermore, ROR1 knockdown inhibited the growth and induced the apoptosis in DLBCL cells and xenografts. In addition, shROR1 inhibited activation of the PI3K/Akt/mTOR signaling pathway, both in vitro and in vivo. Taken together, our results suggest that ROR1 is a novel prognostic marker for DLBCL survival and ROR1 significantly promotes DLBCL tumorigenesis by regulating the PI3K/Akt/mTOR signaling pathway. Targeting ROR1 may provide a promising strategy for DLBCL treatment. © 2019 BioFactors, 45(3):416-426, 2019.


Assuntos
Linfoma/metabolismo , Linfoma/patologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Animais , Regulação Neoplásica da Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/fisiologia , Humanos , Camundongos Endogâmicos BALB C , Camundongos Nus , Fosfatidilinositol 3-Quinases/genética , Proteínas Proto-Oncogênicas c-akt/genética , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/genética , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Serina-Treonina Quinases TOR/genética , Ensaios Antitumorais Modelo de Xenoenxerto
11.
Oncogene ; 38(19): 3551-3568, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30655605

RESUMO

Although head and neck squamous cell carcinoma (HNSCC) has in the past been largely associated with tobacco use, human papillomavirus (HPV+) oropharynx cancer has in recent years emerged as the fastest growing type of HNSCC. Patients with HPV+ HNSCC have a better prognosis; however, the 5-year survival for both HPV+ and HPV- subtypes with recurrent or metastatic disease is poor. To gain insights into the tumor microenvironments of both HNSCC subtypes and identify potential therapeutic targets, we performed epigenomic deconvolution on 580 HNSCC samples from the TCGA dataset. Deconvolution revealed distinct molecular and histoepigenetic profiles of the two tumor subtypes, including their cellular composition, epigenomic profiles and gene expression for constituent cell types, and potential cancer cell-specific targets. Our analyses show that high abundance of both CD8 T-cells and B-cells explains better prognosis in HPV+ HNSCC. Deconvolution of gene expression profiles revealed higher expression of the immunotherapy target PD-1 in HPV+ immune cells compared to HPV- cells, suggesting that HPV+ tumors may preferentially benefit from PD-1 targeted therapy. Further analyses identified HPV+ and HPV- cancer cell surface proteins that can also serve as potential targets for therapy. Specifically, Wnt pathway receptor ROR2 is preferentially overexpressed in HPV+ subtypes, suggesting opportunities for development of targeted therapy based on HPV status. In summary, the comprehensive molecular and histoepigenetic analysis of tumor microenvironments by epigenomic deconvolution reveals potential novel biomarkers and targets for precision therapy of HNSCC.


Assuntos
Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/virologia , Terapia de Alvo Molecular/métodos , Infecções por Papillomavirus/genética , Fumar/efeitos adversos , Antígenos Ly/metabolismo , Linfócitos B/patologia , Linfócitos T CD8-Positivos/patologia , Antígeno CTLA-4/metabolismo , Metilação de DNA , Epigênese Genética , Proteínas Ligadas por GPI/metabolismo , Regulação Neoplásica da Expressão Gênica , Neoplasias de Cabeça e Pescoço/mortalidade , Neoplasias de Cabeça e Pescoço/patologia , Humanos , Imunoterapia/métodos , Estimativa de Kaplan-Meier , Infecções por Papillomavirus/complicações , Prognóstico , Receptor de Morte Celular Programada 1/metabolismo , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Microambiente Tumoral/imunologia
12.
Oncol Rep ; 41(3): 1531-1538, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30628714

RESUMO

The tumorigenesis and metastasis of tumors are associated with human collagen triple helix repeats containing 1 (CTHRC1). To study the effects and possible impacting mechanisms of CTHRC1 on human cervical carcinoma development, samples of paraffin­embedded cervical carcinoma and HeLa cells were examined. Immunofluorescence, cell wound scratch assay, western blot analysis and Transwell invasion assay were used to evaluate HeLa cells in response to silencing of the CTHRC1 gene in cervical carcinoma. The expression levels of gap­associated proteins of the Wnt/PCP pathway in paraffin­embedded cervical carcinoma samples were also evaluated by immunohistochemical staining. CTHRC1 promoted the migration and invasion of HeLa cells in vitro, downregulated Ror2 and p­c­Jun and activated the Wnt/PCP pathway. Furthermore, the expression of p­c­Jun, Ror2 and Wnt5a was increased after overexpression of CTHRC1 as revealed in HeLa cells compared to control group. The present experiments revealed that CTHRC1 promoted HeLa cell progression by activating the Wnt/PCP signaling pathway and may play a key role in the invasion and metastasis of cervical carcinoma.


Assuntos
Biomarcadores Tumorais/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Neoplasias do Colo do Útero/patologia , Via de Sinalização Wnt , Biomarcadores Tumorais/genética , Movimento Celular , Proliferação de Células , Colo do Útero/patologia , Progressão da Doença , Regulação para Baixo , Proteínas da Matriz Extracelular/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Células HeLa , Humanos , Pessoa de Meia-Idade , Prognóstico , Proteínas Proto-Oncogênicas c-jun/metabolismo , RNA Interferente Pequeno , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Análise de Sobrevida , Regulação para Cima , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/mortalidade , Proteína Wnt-5a/metabolismo
13.
Proc Natl Acad Sci U S A ; 116(4): 1370-1377, 2019 01 22.
Artigo em Inglês | MEDLINE | ID: mdl-30622177

RESUMO

Breast cancers enduring treatment with chemotherapy may be enriched for cancer stem cells or tumor-initiating cells, which have an enhanced capacity for self-renewal, tumor initiation, and/or metastasis. Breast cancer cells that express the type I tyrosine kinaselike orphan receptor ROR1 also may have such features. Here we find that the expression of ROR1 increased in breast cancer cells following treatment with chemotherapy, which also enhanced expression of genes induced by the activation of Rho-GTPases, Hippo-YAP/TAZ, or B lymphoma Mo-MLV insertion region 1 homolog (BMI1). Expression of ROR1 also enhanced the capacity of breast cancer cells to invade Matrigel, form spheroids, engraft in Rag2-/-[Formula: see text] mice, or survive treatment with paclitaxel. Treatment of mice bearing breast cancer patient-derived xenografts (PDXs) with the humanized anti-ROR1 monoclonal antibody cirmtuzumab repressed expression of genes associated with breast cancer stemness, reduced activation of Rho-GTPases, Hippo-YAP/TAZ, or BMI1, and impaired the capacity of breast cancer PDXs to metastasize or reengraft Rag2-/-[Formula: see text] mice. Finally, treatment of PDX-bearing mice with cirmtuzumab and paclitaxel was more effective than treatment with either alone in eradicating breast cancer PDXs. These results indicate that targeting ROR1 may improve the response to chemotherapy of patients with breast cancer.


Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Adenosina Trifosfatases/metabolismo , Animais , Anticorpos Monoclonais , Mama/efeitos dos fármacos , Mama/metabolismo , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Xenoenxertos , Humanos , Camundongos , Células NIH 3T3 , Proteínas Nucleares/metabolismo , Paclitaxel/farmacologia , Complexo Repressor Polycomb 1/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Fatores de Transcrição/metabolismo
14.
Fish Physiol Biochem ; 45(1): 355-363, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30242697

RESUMO

Ror family of receptor tyrosine kinases ROR1 and ROR2 plays crucial roles in animal development by regulating cell proliferation, differentiation, and migration, as well as survival and death by acting as a receptor or co-receptor for Wnt5a and mediating Wnt5a-induced activation. Compared with our extensive understanding of ROR2, our knowledge of ROR1 is limited. In this study, we characterized the zebrafish ror1 gene and determined its temporal and spatial expression and biological activity. Sequence comparison and phylogenetic analyses indicate that its protein structure is similar to its mammalian orthologs. During embryogenesis, the ror1 mRNA levels were relatively low or undetectable at 6 and 9 h postfertilization. In adult fish, ror1 mRNA was most abundantly expressed in the ovary and testis. The levels of ror1 mRNA in non-reproductive system tissues were very low or barely detectable. Spatiotemporal distribution of ror1 and its ligand wnt5a in the ovary was then investigated. Reverse transcription PCR on isolated follicle layers and denuded oocytes demonstrated that both wnt5a and ror1 were exclusively expressed in the oocyte but not in the follicle layers. During oogenesis, the ror1 mRNA levels were relatively low from I to IV stage oocytes and increased dramatically at V stage oocyte. Unlike ror1, the wnt5a mRNA levels were increased gradually from I to V stage oocyte. When Ror1 was co-transfected with Wnt5a and Wnt3a in HEK293T cells, the Wnt3a-induced Wnt reporter activity was inhibited by Ror1 in a dose-dependent manner. Taken together, these results provide new information about the structural and functional conservation, spatial and temporal expression, and biological activity of Ror1 in a fish model organism.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/metabolismo , Sequência de Aminoácidos , Animais , Clonagem Molecular , Filogenia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/genética , Proteínas de Peixe-Zebra/genética
15.
J Invest Surg ; 32(2): 137-142, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29058491

RESUMO

Purpose/aim: To focus on current aspects of primary thyroid lymphoma (PTL), which is a rare clinical entity usually manifested by a rapidly growing mass in the neck that can cause pressure symptoms. MATERIALS AND METHODS: Relevant papers in PubMed published through June 2017 were selected to track updated information about PTL with an emphasis on diagnosis and novel therapeutic management. RESULTS: The most frequent cases include non-Hodgkin lymphoma derived from B-cells, mainly diffuse large B-cell lymphoma (DLBCL) followed by mucosa-associated lymphoid tissue (MALT) lymphoma or a mixed type. Other subtypes are less common. Lymphomas derived from T-cells and Hodgkin lymphomas are extremely rare. Hashimoto's autoimmune thyroiditis has been implicated as a risk factor for lymphoma. At the molecular level, the Wnt5a protein and its receptor Ror2 are involved in the course of the disease. Ultrasonography, fine needle aspiration (FNA) biopsy, and core or open biopsy combined with new diagnostic facilities contribute to an accurate diagnosis. An increased potential exists for a cure without the need for a radical surgical procedure. Modern chemoradiation therapy plus the monoclonal antibody rituximab, which acts against CD20, have limited the need for surgical interventions and provide an excellent outcome in most cases. However, some cases have resulted in treatment failure or recurrence. CONCLUSIONS: A multidisciplinary approach must be used to define the management policy in each case. Future efforts by researchers are likely to be focused on the molecular level.


Assuntos
Linfoma de Zona Marginal Tipo Células B/terapia , Linfoma Difuso de Grandes Células B/terapia , Recidiva Local de Neoplasia/epidemiologia , Neoplasias da Glândula Tireoide/terapia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Biópsia , Quimiorradioterapia/métodos , Ensaios Clínicos como Assunto , Intervalo Livre de Doença , Humanos , Linfoma de Zona Marginal Tipo Células B/diagnóstico , Linfoma de Zona Marginal Tipo Células B/mortalidade , Linfoma Difuso de Grandes Células B/diagnóstico , Linfoma Difuso de Grandes Células B/mortalidade , Recidiva Local de Neoplasia/prevenção & controle , Equipe de Assistência ao Paciente , Prognóstico , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Rituximab/uso terapêutico , Glândula Tireoide/diagnóstico por imagem , Glândula Tireoide/patologia , Neoplasias da Glândula Tireoide/diagnóstico , Neoplasias da Glândula Tireoide/mortalidade , Tireoidectomia , Proteína Wnt-5a/metabolismo
16.
J Exp Med ; 216(2): 428-449, 2019 02 04.
Artigo em Inglês | MEDLINE | ID: mdl-30593464

RESUMO

In a substantial fraction of prostate cancer (PCa) patients, bone metastasis appears after years or even decades of latency. Canonical Wnt/ß-catenin signaling has been proposed to be implicated in dormancy of cancer cells. However, how these tumor cells are kept dormant and recur under control of Wnt/ß-catenin signaling derived from bone microenvironment remains unknown. Here, we report that Wnt5a from osteoblastic niche induces dormancy of PCa cells in a reversible manner in vitro and in vivo via inducing Siah E3 Ubiquitin Protein Ligase 2 (SIAH2) expression, which represses Wnt/ß-catenin signaling. Furthermore, this effect of Wnt5a-induced dormancy of PCa cells depends on receptor tyrosine kinase-like orphan receptor 2 (ROR2), and a negative correlation of ROR2 expression with bone metastasis-free survival is observed in PCa patients. Therefore, these results demonstrate that Wnt5a/ROR2/SIAH2 signaling axis plays a crucial role in inducing and maintaining PCa cells dormancy in bone, suggesting a potential therapeutic utility of Wnt5a via inducing dormancy of PCa cells in bone.


Assuntos
Osso e Ossos/metabolismo , Neoplasias da Próstata/metabolismo , Microambiente Tumoral , Proteína Wnt-5a/metabolismo , Animais , Osso e Ossos/patologia , Humanos , Masculino , Camundongos , Metástase Neoplásica , Proteínas Nucleares , Células PC-3 , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Ratos , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/genética , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Proteína Wnt-5a/genética
17.
Leukemia ; 33(3): 653-661, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30568170

RESUMO

Chronic lymphocytic leukemia cells (CLL) migrate between the blood and lymphoid tissues in response to chemokines. Such migration requires structured cytoskeletal-actin polymerization, which may involve the protein cortactin. We discovered that treatment of CLL cells with Wnt5a causes Receptor tyosin kinase-like orphan receptor 1 (ROR1) to bind cortactin, which undergoes tyrosine phosphorylation at Y421, recruits ARHGEF1, and activates RhoA, thereby enhancing leukemia-cell migration; such effects could be inhibited by cirmtuzumab, a humanized mAb specific for ROR1. We transfected the CLL-cell-line MEC1 with either full-length ROR1 or various mutant forms of ROR1 to examine the structural features required for binding cortactin. We found that the proline-rich domain (PRD) was necessary for ROR1 to recruit cortactin. We generated MEC1 cells that each expressed a mutant form of ROR1 with a single amino-acid substitution of alanine (A) for proline (P) in potential SH3-binding sites in the ROR1-PRD at positions 784, 808, 826, 841, or 850. In contrast to wild-type ROR1, or other ROR1P=>A mutants, ROR1P(841)A failed to complex with cortactin or ARHGEF1 in response to Wnt5a. Moreover, Wnt5a could not induce MEC1-ROR1P(841)A to phosphorylate cortactin or enhance CLL-cell F-actin polymerization. Taken together, these studies show that cortactin plays an important role in ROR1-dependent Wnt5a-enhanced CLL-cell migration.


Assuntos
Movimento Celular/fisiologia , Cortactina/metabolismo , Leucemia Linfocítica Crônica de Células B/metabolismo , Leucemia Linfocítica Crônica de Células B/patologia , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Proteína Wnt-5a/metabolismo , Actinas/metabolismo , Alanina/metabolismo , Substituição de Aminoácidos/efeitos dos fármacos , Anticorpos Monoclonais/farmacologia , Sítios de Ligação/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Humanos , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Fosforilação/efeitos dos fármacos , Fatores de Troca de Nucleotídeo Guanina Rho/metabolismo , Tirosina/metabolismo , Proteína rhoA de Ligação ao GTP/metabolismo
18.
Mol Ther ; 27(2): 287-299, 2019 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-30573301

RESUMO

Chimeric antigen receptor (CAR)-T cell immunotherapy is under intense preclinical and clinical investigation, and it involves a rapidly increasing portfolio of novel target antigens and CAR designs. We established a platform that enables rapid and high-throughput CAR-screening campaigns with reporter cells derived from the T cell lymphoma line Jurkat. Reporter cells were equipped with nuclear factor κB (NF-κB) and nuclear factor of activated T cells (NFAT) reporter genes that generate a duplex output of enhanced CFP (ECFP) and EGFP, respectively. As a proof of concept, we modified reporter cells with CD19-specific and ROR1-specific CARs, and we detected high-level reporter signals that allowed distinguishing functional from non-functional CAR constructs. The reporter data were highly reproducible, and the time required for completing each testing campaign was substantially shorter with reporter cells (6 days) compared to primary CAR-T cells (21 days). We challenged the reporter platform to a large-scale screening campaign on a ROR1-CAR library, and we showed that reporter cells retrieved a functional CAR variant that was present with a frequency of only 6 in 1.05 × 106. The data illustrate the potential to implement this reporter platform into the preclinical development path of novel CAR-T cell products and to inform and accelerate the selection of lead CAR candidates for clinical translation.


Assuntos
Biblioteca Gênica , NF-kappa B/metabolismo , Fatores de Transcrição NFATC/metabolismo , Receptores de Antígenos Quiméricos/metabolismo , Animais , Linhagem Celular Tumoral , Células Cultivadas , Citometria de Fluxo , Proteínas de Fluorescência Verde/metabolismo , Humanos , Imunoterapia/métodos , Células Jurkat , Camundongos , Neoplasias/metabolismo , Neoplasias/terapia , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo
19.
J Gen Virol ; 100(1): 99-104, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30431423

RESUMO

Cytomegalovirus (CMV) infection during pregnancy may lead to adverse pregnancy outcomes and permanent neurological disabilities in infants infected in utero. Congenital CMV disease of the foetus and neonate results from both direct viral cytopathic damage and indirect effects through placental dysfunction. Infection specifically alters Wnt signalling, an essential pathway involved in trophoblast migration and placental development. We examined CMV regulation of trophoblast migration. This virus controls expression of Wnt-binding receptor tyrosine kinase ROR2, but not alternate receptor tyrosine kinases ROR1 or RYK. Ectopic expression of ROR2 reduced Wnt5a-induced trophoblast migration, whilst overexpression of ROR1 or RYK did not affect trophoblast migration. CMV infection increased ROR2 protein expression in trophoblasts, with no effect on ROR1 and RYK expression. These data further support the proposal that specific inhibition of this mechanism may be a target for therapeutic intervention to reduce placental damage and consequent foetal disease due to congenital CMV infection.


Assuntos
Movimento Celular , Citomegalovirus/crescimento & desenvolvimento , Expressão Gênica , Interações Hospedeiro-Patógeno , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Trofoblastos/fisiologia , Trofoblastos/virologia , Linhagem Celular , Humanos
20.
BMC Cardiovasc Disord ; 18(1): 196, 2018 10 20.
Artigo em Inglês | MEDLINE | ID: mdl-30342492

RESUMO

BACKGROUND: Receptor tyrosine kinases (RTK) are potential targets for the treatment of ischemic heart disease. The human RTK family consists of 55 members, most of which have not yet been characterized for expression or activity in the ischemic heart. METHODS: RTK gene expression was analyzed from human heart samples representing healthy tissue, acute myocardial infarction or ischemic cardiomyopathy. As an experimental model, pig heart with ischemia-reperfusion injury, caused by cardiopulmonary bypass, was used, from which phosphorylation status of RTKs was assessed with a phospho-RTK array. Expression and function of one RTK, ROR1, was further validated in pig tissue samples, and in HL-1 cardiomyocytes and H9c2 cardiomyoblasts, exposed to hypoxia and reoxygenation. ROR1 protein level was analyzed by Western blotting. Cell viability after ROR1 siRNA knockdown or activation with Wnt-5a ligand was assessed by MTT assays. RESULTS: In addition to previously characterized RTKs, a group of novel active and regulated RTKs was detected in the ischemic heart. ROR1 was the most significantly upregulated RTK in human ischemic cardiomyopathy. However, ROR1 phosphorylation was suppressed in the pig model of ischemia-reperfusion and ROR1 phosphorylation and expression were down-regulated in HL-1 cardiomyocytes subjected to short-term hypoxia in vitro. ROR1 expression in the pig heart was confirmed on protein and mRNA level. Functionally, ROR1 activity was associated with reduced viability of HL-1 cardiomyocytes in both normoxia and during hypoxia-reoxygenation. CONCLUSIONS: Several novel RTKs were found to be regulated in expression or activity in ischemic heart. ROR1 was one of the most significantly regulated RTKs. The in vitro findings suggest a role for ROR1 as a potential target for the treatment of ischemic heart injury.


Assuntos
Cardiomiopatias/enzimologia , Infarto do Miocárdio/enzimologia , Traumatismo por Reperfusão Miocárdica/enzimologia , Miócitos Cardíacos/enzimologia , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Animais , Cardiomiopatias/tratamento farmacológico , Cardiomiopatias/genética , Cardiomiopatias/patologia , Estudos de Casos e Controles , Linhagem Celular , Sobrevivência Celular , Modelos Animais de Doenças , Regulação Enzimológica da Expressão Gênica , Humanos , Terapia de Alvo Molecular , Infarto do Miocárdio/tratamento farmacológico , Infarto do Miocárdio/genética , Infarto do Miocárdio/patologia , Traumatismo por Reperfusão Miocárdica/tratamento farmacológico , Traumatismo por Reperfusão Miocárdica/genética , Traumatismo por Reperfusão Miocárdica/patologia , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/patologia , Fosforilação , Inibidores de Proteínas Quinases/farmacologia , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/antagonistas & inibidores , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/genética , Transdução de Sinais , Sus scrofa
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