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Rhabdomyosarcoma (RMS) is one of the most common pediatric soft-tissue cancer. Previously, we discovered a gene fusion, MARS-AVIL formed by chromosomal inversion in RMS. Suspecting that forming a fusion with a housekeeping gene may be one of the mechanisms to dysregulate an oncogene, we investigated AVIL expression and its role in RMS. We first showed that MARS-AVIL translates into an in-frame fusion protein, which is critical for RMS cell tumorigenesis. Besides forming a gene fusion with the housekeeping gene, MARS, the AVIL locus is often amplified, and its RNA and protein expression are overexpressed in the majority of RMSs. Tumors with AVIL dysregulation exhibit evidence of oncogene addiction: Silencing MARS-AVIL in cells harboring the fusion, or silencing AVIL in cells with AVIL overexpression, nearly eradicated the cells in culture, as well as inhibited in vivo xenograft growth in mice. Conversely, gain-of-function manipulations of AVIL led to increased cell growth and migration, enhanced foci formation in mouse fibroblasts, and most importantly transformed mesenchymal stem cells in vitro and in vivo. Mechanistically, AVIL seems to serve as a converging node functioning upstream of two oncogenic pathways, PAX3-FOXO1 and RAS, thus connecting two types of RMS associated with these pathways. Interestingly, AVIL is overexpressed in other sarcoma cells as well, and its expression correlates with clinical outcomes, with higher levels of AVIL expression being associated with worse prognosis. AVIL is a bona fide oncogene in RMS, and RMS cells are addicted to its activity.
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Rabdomiossarcoma Alveolar , Rabdomiossarcoma , Humanos , Animais , Camundongos , Fatores de Transcrição Box Pareados/metabolismo , Linhagem Celular Tumoral , Rabdomiossarcoma/genética , Rabdomiossarcoma/patologia , Oncogenes/genética , Feniramina , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Regulação Neoplásica da Expressão Gênica , Rabdomiossarcoma Alveolar/genética , Proteínas dos Microfilamentos/metabolismoRESUMO
Once believed to be unique features of neoplasia, chimeric RNAs are now being discovered in normal physiology. We speculated that some chimeric RNAs may be functional precursors of genes, and that forming chimeric RNA at the transcriptional level may be a 'trial' mechanism before the functional element is fixed into the genome. Supporting this idea, we identified a chimeric RNA, HNRNPA1L2-SUGT1 (H-S), whose sequence is highly similar to that of a 'pseudogene' MRPS31P5. Sequence analysis revealed that MRPS31P5 transcript is more similar to H-S chimeric RNA than its 'parent' gene, MRPS31. Evolutionarily, H-S precedes MRPS31P5, as it can be detected bioinformatically and experimentally in marmosets, which do not yet possess MRPS31P5 in their genome. Conversely, H-S is minimally expressed in humans, while instead, MRPS31P5 is abundantly expressed. Silencing H-S in marmoset cells resulted in similar phenotype as silencing MRPS31P5 in human cells. In addition, whole transcriptome analysis and candidate downstream target validation revealed common signalling pathways shared by the two transcripts. Interestingly, H-S failed to rescue the phenotype caused by silencing MPRS31P5 in human and rhesus cells, whereas MRPS31P5 can at least partially rescue the phenotype caused by silencing H-S in marmoset cells, suggesting that MRPS31P5 may have further evolved into a distinct entity. Thus, multiple lines of evidence support that MRPS31P5 is not truly a pseudogene of MRPS31, but a likely functional descendent of H-S chimera. Instead being a gene fusion product, H-S is a product of cis-splicing between adjacent genes, while MRPS31P5 is likely produced by genome rearrangement.
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Evolução Molecular , Genes , RNA/genética , Recombinação Genética , Genes Mitocondriais , Humanos , Especificidade de Órgãos , Filogenia , Pseudogenes , Transcrição GênicaRESUMO
Pax3 plays an essential role in myogenesis. Previously, we found a tumor-signature chimeric fusion RNA, PAX3-FOXO1 also present during muscle differentiation, raising the possibility of its physiological role. Here we demonstrated that the fusion is needed transiently for muscle lineage commitment. Interestingly, the fusion ortholog was not found in seven mouse muscle differentiation/regeneration systems, nor in other stem cell differentiation systems of another three mammal species. We noticed that Pax3 is expressed at a much lower level in human stem cells, and during muscle differentiation than in other mammals. Given the fact that the fusion and the parental Pax3 share common downstream targets, we reasoned that forming the fusion may be a mechanism for human cells to escape certain microRNA regulation on Pax3. By sequence comparison, we identified 16 candidate microRNAs that may specifically target the human PAX3 3'UTR. We used a luciferase reporter assay, examined the microRNAs expression, and conducted mutagenesis on the reporters, as well as a CRISPR/Cas9 mediated editing on the endogenous allele. Finally, we identified miR-495 as a microRNA that specifically targets human PAX3. Examining several other fusion RNAs revealed that the human-specificity is not limited to PAX3-FOXO1. Based on these observations, we conclude that PAX3-FOXO1 fusion RNA is absent in mouse, or other mammals we tested, the fusion RNA is a mechanism to escape microRNA, miR-495 regulation in humans, and that it is not the only human-specific fusion RNA.
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Diferenciação Celular , Proteína Forkhead Box O1/metabolismo , Regulação da Expressão Gênica , MicroRNAs/genética , Desenvolvimento Muscular , Fator de Transcrição PAX3/metabolismo , Animais , Sistemas CRISPR-Cas , Diferenciação Celular/genética , Linhagem Celular Tumoral , Proteína Forkhead Box O1/genética , Edição de Genes , Técnicas de Silenciamento de Genes , Genes Reporter , Cavalos , Humanos , Camundongos , Desenvolvimento Muscular/genética , Fator de Transcrição PAX3/genética , Interferência de RNA , RNA Interferente Pequeno/genética , Ovinos , SuínosRESUMO
Gene fusions and fusion products were thought to be unique features of neoplasia. However, more and more studies have identified fusion RNAs in normal physiology. Through RNA sequencing of 27 human noncancer tissues, a large number of fusion RNAs were found. By analyzing fusion transcriptome, we observed close clusterings between samples of same or similar tissues, supporting the feasibility of using fusion RNA profiling to reveal connections between biological samples. To put the concept into use, we selected alveolar rhabdomyosarcoma (ARMS), a myogenic pediatric cancer whose exact cell of origin is not clear. PAX3-FOXO1 (paired box gene 3 fused with forkhead box O1) fusion RNA, which is considered a hallmark of ARMS, was recently found during normal muscle cell differentiation. We performed and analyzed RNA sequencing from various time points during myogenesis and uncovered many chimeric fusion RNAs. Interestingly, we found that the fusion RNA profile of RH30, an ARMS cell line, is most similar to the myogenesis time point when PAX3-FOXO1 is expressed. In contrast, full transcriptome clustering analysis failed to uncover this connection. Strikingly, all of the 18 chimeric RNAs in RH30 cells could be detected at the same myogenic time point(s). In addition, the seven chimeric RNAs that follow the exact transient expression pattern as PAX3-FOXO1 are specific to rhabdomyosarcoma cells. Further testing with clinical samples also confirmed their specificity to rhabdomyosarcoma. These results provide further support for the link between at least some ARMSs and the PAX3-FOXO1-expressing myogenic cells and demonstrate that fusion RNA profiling can be used to investigate the etiology of fusion-gene-associated cancers.
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Fusão Gênica , RNA/genética , Rabdomiossarcoma Alveolar/genética , Linhagem Celular Tumoral , Proteína Forkhead Box O1/genética , Perfilação da Expressão Gênica , Humanos , Células-Tronco Mesenquimais , Desenvolvimento Muscular/genética , Fator de Transcrição PAX3/genética , Análise de Sequência de RNARESUMO
Gene fusions and their products (RNA and protein) were once thought to be unique features to cancer. However, chimeric RNAs can also be found in normal cells. Here, we performed, curated and analyzed nearly 300 RNA-Seq libraries covering 30 different non-neoplastic human tissues and cells as well as 15 mouse tissues. A large number of fusion transcripts were found. Most fusions were detected only once, while 291 were seen in more than one sample. We focused on the recurrent fusions and performed RNA and protein level validations on a subset. We characterized these fusions based on various features of the fusions, and their parental genes. They tend to be expressed at higher levels relative to their parental genes than the non-recurrent ones. Over half of the recurrent fusions involve neighboring genes transcribing in the same direction. A few sequence motifs were found enriched close to the fusion junction sites. We performed functional analyses on a few widely expressed fusions, and found that silencing them resulted in dramatic reduction in normal cell growth and/or motility. Most chimeras use canonical splicing sites, thus are likely products of 'intergenic splicing'. We also explored the implications of these non-pathological fusions in cancer and in evolution.
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Fibroblastos/metabolismo , Fusão Gênica , Células-Tronco Mesenquimais/metabolismo , Splicing de RNA , RNA Mensageiro/genética , Animais , Astrócitos/citologia , Astrócitos/metabolismo , Sequência de Bases , Linhagem Celular Transformada , Biologia Computacional , Evolução Molecular , Fibroblastos/citologia , Biblioteca Gênica , Inativação Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Células-Tronco Mesenquimais/citologia , Camundongos , Dados de Sequência Molecular , Cultura Primária de Células , RNA Mensageiro/antagonistas & inibidores , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Análise de Sequência de RNA , Especificidade da EspécieRESUMO
Circular RNAs (circRNAs) are a family of endogenous RNAs that have become a focus of biological research in recent years. Emerging evidence has revealed that circRNAs exert biological functions by acting as transcriptional regulators, microRNA sponges, and binding partners with RNA-binding proteins. However, few studies have identified coding circRNAs, which may lead to a hidden repertoire of proteins. In this study, we unexpectedly discovered a protein-encoding circular RNA circCCDC7(15,16,17,18,19) while we were searching for prostate cancer related chimeric RNAs. circCCDC7(15,16,17,18,19) is derived from exon 19 back spliced to exon 15 of the CCDC7 gene. It is significantly downregulated in patients with high Gleason score. Prostate cancer patients with decreased circCCDC7(15,16,17,18,19) expression have a worse prognosis, while linear CCDC7 had no such association. Overexpressed circCCDC7(15,16,17,18,19) inhibited prostate cancer cell migration, invasion, and viability, supporting classification of circCCDC7(15,16,17,18,19) as a bona fide tumor suppressor gene. We provide evidence that its tumor suppressive activity is driven by the protein it encodes, and that circCCDC7(15,16,17,18,19) encodes a secretory protein. Consistently, conditioned media from circCCDC7(15,16,17,18,19) overexpressing cells has the same tumor suppressive activity. We further demonstrate that the tumor suppressive activity of circCCDC7(15,16,17,18,19) is at least partially mediated by FLRT3, whose expression also negatively correlates with Gleason score and clinical prognosis. In conclusion, circCCDC7(15,16,17,18,19) functions as a tumor suppressor in prostate cancer cells through the circCCDC7-180aa secretory protein it encodes, and is a promising therapeutic peptide for prostate cancer.
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Therapeutic agents targeting cytokine-cytokine receptor (CK-CKR) interactions lead to the disruption in cellular signaling and are effective in treating many diseases including tumors. However, a lack of universal and quick access to annotated structural surface regions on CK/CKR has limited the progress of a structure-driven approach in developing targeted macromolecular drugs and precision medicine therapeutics. Herein we develop CytoSIP (Single nucleotide polymorphisms (SNPs), Interface, and Phenotype), a rich internet application based on a database of atomic interactions around hotspots in experimentally determined CK/CKR structural complexes. CytoSIP contains: (1) SNPs on CK/CKR; (2) interactions involving CK/CKR domains, including CK/CKR interfaces, oligomeric interfaces, epitopes, or other drug targeting surfaces; and (3) diseases and phenotypes associated with CK/CKR or SNPs. The database framework introduces a unique tri-level SIP data model to bridge genetic variants (atomic level) to disease phenotypes (organism level) using protein structure (complexes) as an underlying framework (molecule level). Customized screening tools are implemented to retrieve relevant CK/CKR subset, which reduces the time and resources needed to interrogate large datasets involving CK/CKR surface hotspots and associated pathologies. CytoSIP portal is publicly accessible at https://CytoSIP.biocloud.top , facilitating the panoramic investigation of the context-dependent crosstalk between CK/CKR and the development of targeted therapeutic agents.
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Citocinas , Polimorfismo de Nucleotídeo Único , Receptores de Citocinas , Humanos , Receptores de Citocinas/metabolismo , Receptores de Citocinas/química , Receptores de Citocinas/genética , Citocinas/metabolismo , Citocinas/genética , Citocinas/química , Bases de Dados de Proteínas , FenótipoRESUMO
We recently identified CD46 as a novel prostate cancer cell surface antigen that shows lineage independent expression in both adenocarcinoma and small cell neuroendocrine subtypes of metastatic castration resistant prostate cancer (mCRPC), discovered an internalizing human monoclonal antibody YS5 that binds to a tumor selective CD46 epitope, and developed a microtubule inhibitor-based antibody drug conjugate that is in a multi-center phase I trial for mCRPC (NCT03575819). Here we report the development of a novel CD46-targeted alpha therapy based on YS5. We conjugated 212Pb, an in vivo generator of alpha-emitting 212Bi and 212Po, to YS5 through the chelator TCMC to create the radioimmunoconjugate, 212Pb-TCMC-YS5. We characterized 212Pb-TCMC-YS5 in vitro and established a safe dose in vivo. We next studied therapeutic efficacy of a single dose of 212Pb-TCMC-YS5 using three prostate cancer small animal models: a subcutaneous mCRPC cell line-derived xenograft (CDX) model (subcu-CDX), an orthotopically grafted mCRPC CDX model (ortho-CDX), and a prostate cancer patient-derived xenograft model (PDX). In all three models, a single dose of 0.74 MBq (20 µCi) 212Pb-TCMC-YS5 was well tolerated and caused potent and sustained inhibition of established tumors, with significant increases of survival in treated animals. A lower dose (0.37 MBq or 10 µCi 212Pb-TCMC-YS5) was also studied on the PDX model, which also showed a significant effect on tumor growth inhibition and prolongation of animal survival. These results demonstrate that 212Pb-TCMC-YS5 has an excellent therapeutic window in preclinical models including PDXs, opening a direct path for clinical translation of this novel CD46-targeted alpha radioimmunotherapy for mCRPC treatment.
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Neoplasias de Próstata Resistentes à Castração , Radioimunoterapia , Masculino , Animais , Humanos , Radioimunoterapia/métodos , Chumbo , Partículas alfa , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Radioisótopos de Chumbo/uso terapêutico , Proteína Cofatora de MembranaRESUMO
The key factor of realizing super-resolution optical microscopy at the single-molecule level is to separately position two adjacent molecules. An opportunity to independently localize target molecules is provided by the intermittency (blinking) in fluorescence of a quantum dot (QD) under the condition that the blinking of each emitter can be recorded and identified. Herein we develop a spectral imaging based color nanoscopy which is capable of determining which QD is blinking in the multicolor QD complex through tracking the first-order spectrum, and thus, the distance at tens of nanometers between two QDs is measured. Three complementary oligonucleotides with lengths of 15, 30, and 45 bp are constructed as calibration rulers. QD585 and QD655 are each linked at one end. The measured average distances are in good agreement with the calculated lengths with a precision of 6 nm, and the intracellular dual-color QDs within a diffraction-limited spot are distinguished.
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Microscopia , Pontos Quânticos , Linhagem Celular , Cor , Humanos , Oligonucleotídeos/químicaRESUMO
Transcription factor Foxm1 plays a critical role during embryonic development and its expression is repressed during retinoic acid (RA)-induced differentiation of pluripotent P19 embryonal carcinoma cells at the early stage, correlated with downregulation of expression of pluripotency markers. To study whether Foxm1 participates in the maintenance of pluripotency of stem cells, we knock down Foxm1 expression in P19 cells and identify that Oct4 are regulated directly by Foxm1. Knockdown of Foxm1 also results in spontaneous differentiation of P19 cells to mesodermal derivatives, such as muscle and adipose tissues. Maintaining Foxm1 expression prevents the downregulation of pluripotency-related transcription factors such as Oct4 and Nanog during P19 cell differentiation. Furthermore, overexpression of FOXM1 alone in RA-differentiated P19 cells (4 days) or human newborn fibroblasts restarts the expression of pluripotent genes Oct4, Nanog and Sox2. Together, our results suggest a critical involvement of Foxm1 in maintenance of stem cell pluripotency.
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Células-Tronco de Carcinoma Embrionário/citologia , Fatores de Transcrição Forkhead/fisiologia , Células-Tronco Pluripotentes/citologia , Animais , Diferenciação Celular , Linhagem Celular Tumoral , Células Cultivadas , Regulação para Baixo , Células-Tronco de Carcinoma Embrionário/metabolismo , Fibroblastos/metabolismo , Proteína Forkhead Box M1 , Fatores de Transcrição Forkhead/antagonistas & inibidores , Fatores de Transcrição Forkhead/genética , Técnicas de Silenciamento de Genes , Humanos , Recém-Nascido , Mesoderma/citologia , Camundongos , Camundongos Nus , Fator 3 de Transcrição de Octâmero/genética , Células-Tronco Pluripotentes/metabolismo , Regiões Promotoras GenéticasRESUMO
Arthropod pathogens and other microorganisms have been documented from mass production systems aimed at producing natural enemies for pest control. If losses due to pathogens are encountered in such systems, they could lead to uneconomical production of natural enemies and/or a loss of predator quality for effective field control of target pests. Here, we identify and describe the laboratory transmission of a bacterial pathogen, Acaricomes phytoseiuli, in a Chinese strain of the local predatory mite Neoseiulus californicus following contact with Phytoseiulus persimilis, a predatory mite imported for the control of small sap-sucking pests in greenhouses. Analysis of the 16S rRNA gene and whole-genome sequences of A. phytoseiuli isolated from the Chinese strain of N. californicus showed 99.6 and 99.78% similarity, respectively, to the pathogen isolated from a European population (DSM14247 strain). This is the first report of P. persimilis infected with A. phytoseiuli outside Europe and transmitting to a local predatory mite species. A. phytoseiuli severely damaged local N. californicus, leading to a dorso-ventrally flattened body and reduced prey consumption and reproduction as well as early death. Through fluorescence in situ hybridization, A. phytoseiuli was shown to accumulate in the digestive tract of mites and in the oviductal bulb of adult females. Infected males had no obvious symptoms, but they still were able to pass on the infection to healthy females through contact and mating. The pathogen was transmitted vertically to offspring by either infected parent through adherence to eggshells. A. phytoseiuli could also persist in other herbivorous arthropods from the same habitat, suggesting wider potential risks. Our study highlights pathogen risk to predators in local biocontrol industries through pathogen spread from imported material. IMPORTANCE Predatory mites are important natural enemies for biological control of pests, but mass rearing of the mites can be affected by pathogens. In particular, the mite pathogen Acaricomes phytoseiuli may pose a threat to predatory mite production, and we have now detected this pathogen in China. We explored the pathogen's transmission within species, its ability to transfer to a locally important predatory mite species, and symptoms following transfer. The detection of A. phytoseiuli and its ability to transfer to a local predator where it reduces performance highlight the importance of ongoing monitoring and hygiene in the predatory mite industry.
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Ácaros , Animais , Feminino , Ácaros/microbiologia , RNA Ribossômico 16S/genética , Hibridização in Situ Fluorescente , Controle Biológico de VetoresRESUMO
PURPOSE: Specific gene fusions and their fusion products (chimeric RNA and protein) have served as ideal diagnostic markers and therapeutic targets for cancer. However, few systematic studies for chimeric RNAs have been conducted in neuroendocrine prostate cancer (NEPC). In this study, we explored the landscape of chimeric RNAs in different types of prostate cancer (PCa) cell lines and aimed to identify chimeric RNAs specifically expressed in NEPC. METHODS: To do so, we employed the RNA-seq data of eight prostate related cell lines from Cancer Cell Line Encyclopedia (CCLE) for chimeric RNA identification. Multiple filtering criteria were used and the candidate chimeric RNAs were characterized at multiple levels and from various angles. We then performed experimental validation on all 80 candidates, and focused on the ones that are specific to NEPC. Lastly, we studied the clinical relevance and effect of one chimera in neuroendocrine process. RESULTS: Out of 80 candidates, 15 were confirmed to be expressed preferentially in NEPC lines. Among them, 13 of the 15 were found to be specifically expressed in NEPC, and four were further validated in another NEPC cell line. Importantly, in silico analysis showed that tumor malignancy may be correlated to the level of these chimeric RNAs. Clinically, the expression of TMPRSS2-ERG (e2e4) was elevated in tumor tissues and indicated poor clinical prognosis, whereas the parental wild type transcripts had no such association. Furthermore, compared to the most frequently detected TMPRSS2-ERG form (e1e4), e2e4 encodes 31 more amino acids and accelerated neuroendocrine process of prostate cancer. CONCLUSIONS: In summary, these findings painted the landscape of chimeric RNA in NEPC and supported the idea that some chimeric RNAs may represent additional biomarkers and/or treatment targets independent of parental gene transcripts.
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BACKGROUND: Chemoresistance is the major cause of death in advanced prostate cancer (PCa), especially in metastatic PCa (mPCa). However, the molecular mechanisms underlying the chemoresistance of PCa remain unclear. Understanding the reason behind the drug resistance would be helpful in developing new treatment approaches. METHODS: The Cancer Genome Atlas, Gene Expression Omnibus datasets, and clinical samples were used to examine the correlation between growth arrest and DNA damage-inducible 45 beta (GADD45B) with clinical characteristics and prognosis. Lentiviral transfection was used to construct GADD45B overexpression cell lines. Hypoxic incubator, low serum medium, or docetaxel was used to build environmental stress model or chemotherapy cell model. The MTS assay and colony formation assay were used to test cell viability. Apoptosis and cell cycle were detected by flow cytometry. The RNA and protein levels of related biomarkers were tested by Western blotting and quantitative polymerase chain reaction. Bioinformatics analysis after RNA sequencing was performed to identify the possible mechanism of how GADD45B regulates chemotherapy resistance. RESULTS: GADD45B was related to distant metastasis but not to Gleason score, prostate-specific antigen level, T stage, or lymph node metastasis and indicated a good prognosis. The level of GADD45B increased significantly in PCa cells that faced environmental stress. It was found that a high level of GADD45B significantly enhanced the chemosensitivity. Furthermore, high GADD45B promoted cell apoptosis via mitogen-activated protein kinase (MAPK) pathway. CONCLUSION: GADD45B promoted chemosensitivity of prostate cancer through MAPK pathway. GADD45B could serve as a diagnostic biomarker and therapeutic target for mPCa or chemotherapy-resistant patients.
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Immunotherapy is gradually emerging in the field of tumor treatment. However, because of the complexity of the tumor microenvironment (TME), some patients cannot benefit from immunotherapy. Therefore, we comprehensively analyzed the TME and gene mutations of ccRCC to identify a comprehensive index that could more accurately guide the immunotherapy of patients with ccRCC. We divided ccRCC patients into two groups based on immune infiltration activity. Next, we investigated the differentially expressed genes (DEGs) and constructed a prognostic immune score using univariate Cox regression analysis, unsupervised cluster analysis, and principal component analysis (PCA) and validated its predictive power in both internal and total sets. Subsequently, the gene mutations in the groups were investigated, and patients suitable for immunotherapy were selected in combination with the immune score. The prognosis of the immune score-low group was significantly worse than that of the immune score-high group. The patients with BRCA1-associated protein 1 (BAP1) mutation had a poor prognosis. Thus, this study indicated that establishing an immune score model combined with BAP1 mutation can better predict the prognosis of patients, screen suitable ccRCC patients for immunotherapy, and select more appropriate drug combinations.
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OBJECTIVE: To explore the genetic changes in the progression of castration-resistant prostate cancer (CRPC) and neuroendocrine prostate cancer (NEPC) and the reason why these cancers resist existing therapies. METHODS: We employed our CRPC cell line microarray and other CRPC or NEPC datasets to screen the target gene NEIL3. Lentiviral transfection and RNA interference were used to construct overexpression and knockdown cell lines. Cell and animal models of radiotherapy were established by using a medical electron linear accelerator. Flow cytometry was used to detect apoptosis or cell cycle progression. Western blot and qPCR were used to detect changes in the protein and RNA levels. RESULTS: TCGA and clinical patient datasets indicated that NEIL3 was downregulated in CRPC and NEPC cell lines, and NEIL3 was correlated with a high Gleason score but a good prognosis. Further functional studies demonstrated that NEIL3 had no effect on the proliferation and migration of PCa cells. However, cell and animal radiotherapy models revealed that NEIL3 could facilitate the radiotherapy sensitivity of PCa cells, while loss of NEIL3 activated radiotherapy resistance. Mechanistically, we found that NEIL3 negatively regulated the expression of ATR, and higher NEIL3 expression repressed the ATR/CHK1 pathway, thus regulating the cell cycle. CONCLUSIONS: We demonstrated that NEIL3 may serve as a diagnostic or therapeutic target for therapy-resistant patients.
Assuntos
Carcinoma Neuroendócrino , Neoplasias de Próstata Resistentes à Castração , Animais , Carcinoma Neuroendócrino/genética , Carcinoma Neuroendócrino/metabolismo , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias de Próstata Resistentes à Castração/genética , Neoplasias de Próstata Resistentes à Castração/radioterapia , RNA/uso terapêutico , Interferência de RNARESUMO
Our work in rhabdomyosarcoma led us to the discovery of a novel oncogene, Advillin (AVIL) in glioblastoma. Multiple lines of evidence support that AVIL is an Achilles heel of glioblastoma, with its specific targeting potentially an effective treatment approach for the disease. A new signaling axis was also established.
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Gene fusions and their fusion products have been recognized as ideal biomarkers and drug targets for cancer. However, few recurrent gene fusions were found in colorectal cancer (CRC), despite comprehensive studies. We believe that chimeric RNAs, in the absence of chromosomal rearrangement, may represent a new repertoire of biomarkers and/or therapeutic targets in CRC. In this study, we aim to identify such recurrent chimeric RNAs, and investigate their clinical implications. To do so, we performed extensive data mining for chimeric RNAs using The Cancer Genome Atlas CRC RNA-Seq datasets. Multiple filtering criteria were applied, and the landscape of chimeric RNAs at multiple levels, from various angles, was analyzed. Eleven frequent, cancer biased chimeric RNAs were validated. The expression of RRM2-C2orf48 correlates with poor clinical outcomes, while the expression of parental RRM2 and C2orf48 correlates with positive clinical outcomes. Mechanistically, it is a product of cis-splicing between adjacent genes. Silencing of RRM2-C2orf48 resulted in reduced cellular proliferation in colon cancer cells, whereas overexpressed chimera promoted cell proliferation. These findings suggest that frequent chimeric RNAs are present in CRCs, and that chimeric RNAs may have different expression profiles and functions from parental genes, thus representing a new repertoire of biomarkers and therapeutic targets.
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Biomarcadores Tumorais/genética , Neoplasias Colorretais/genética , Splicing de RNA/genética , RNA Neoplásico/genética , Ribonucleosídeo Difosfato Redutase/genética , HumanosRESUMO
Glioblastoma is a deadly cancer, with no effective therapies. Better understanding and identification of selective targets are urgently needed. We found that advillin (AVIL) is overexpressed in all the glioblastomas we tested including glioblastoma stem/initiating cells, but hardly detectable in non-neoplastic astrocytes, neural stem cells or normal brain. Glioma patients with increased AVIL expression have a worse prognosis. Silencing AVIL nearly eradicated glioblastoma cells in culture, and dramatically inhibited in vivo xenografts in mice, but had no effect on normal control cells. Conversely, overexpressing AVIL promoted cell proliferation and migration, enabled fibroblasts to escape contact inhibition, and transformed immortalized astrocytes, supporting AVIL being a bona fide oncogene. We provide evidence that the tumorigenic effect of AVIL is partly mediated by FOXM1, which regulates LIN28B, whose expression also correlates with clinical prognosis. AVIL regulates the cytoskeleton through modulating F-actin, while mutants disrupting F-actin binding are defective in its tumorigenic capabilities.
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Glioblastoma/metabolismo , Glioblastoma/patologia , Proteínas dos Microfilamentos/metabolismo , Animais , Western Blotting , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Movimento Celular/fisiologia , Proliferação de Células/genética , Proliferação de Células/fisiologia , Citoesqueleto/metabolismo , Imunofluorescência , Glioblastoma/genética , Humanos , Imuno-Histoquímica , Técnicas In Vitro , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Proteínas dos Microfilamentos/genética , Microscopia Confocal , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Ribonucleotide reductase (RNR) catalyzes the rate-limiting step of de novo synthesis of deoxyribonucleotide triphosphates (dNTPs) building blocks for DNA synthesis, and is a well-recognized target for cancer therapy. RNR is a heterotetramer consisting of two large RRM1 subunits and two small RRM2 subunits. RNR activity is greatly stimulated by transcriptional activation of RRM2 during S/G2 phase to ensure adequate dNTP supply for DNA replication. However, little is known about the cell-cycle-dependent regulation of RNR activity through RRM1. Here, we report that RRM1 is phosphorylated at Ser 559 by CDK2/cyclin A during S/G2 phase. And this S559 phosphorylation of RRM1enhances RNR enzymatic activity and is required for maintaining sufficient dNTPs during normal DNA replication. Defective RRM1 S559 phosphorylation causes DNA replication stress, double-strand break, and genomic instability. Moreover, combined targeting of RRM1 S559 phosphorylation and ATR triggers lethal replication stress and profound antitumor effects. Thus, this posttranslational phosphorylation of RRM1 provides an alternative mechanism to finely regulating RNR and therapeutic opportunities for cancer treatment.
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Proteínas Mutadas de Ataxia Telangiectasia/antagonistas & inibidores , Replicação do DNA/genética , Ribonucleosídeo Difosfato Redutase/isolamento & purificação , Ribonucleosídeo Difosfato Redutase/metabolismo , Ciclo Celular , Humanos , FosforilaçãoRESUMO
Biological samples can be grouped into separate clusters based on their gene expression profiles. This approach has yielded meaningful biological insights and facilitated biomarker discoveries. Recently, we developed another approach to study connections between biological samples based on their fusion RNA expression. We have used this approach to provide insights into the cell of origin for a mysterious tumor, alveolar rhabdomyosarcoma.