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Cancer Discovery Jun 2018The cornerstone of treatment for advanced ALK-positive lung cancer is sequential therapy with increasingly potent and selective ALK inhibitors. The third-generation ALK...
The cornerstone of treatment for advanced ALK-positive lung cancer is sequential therapy with increasingly potent and selective ALK inhibitors. The third-generation ALK inhibitor lorlatinib has demonstrated clinical activity in patients who failed previous ALK inhibitors. To define the spectrum of mutations that confer lorlatinib resistance, we performed accelerated mutagenesis screening of Ba/F3 cells expressing EML4-ALK. Under comparable conditions, -ethyl--nitrosourea (ENU) mutagenesis generated numerous crizotinib-resistant but no lorlatinib-resistant clones harboring single mutations. In similar screens with EML4-ALK containing single resistance mutations, numerous lorlatinib-resistant clones emerged harboring compound mutations. To determine the clinical relevance of these mutations, we analyzed repeat biopsies from lorlatinib-resistant patients. Seven of 20 samples (35%) harbored compound mutations, including two identified in the ENU screen. Whole-exome sequencing in three cases confirmed the stepwise accumulation of mutations during sequential treatment. These results suggest that sequential ALK inhibitors can foster the emergence of compound mutations, identification of which is critical to informing drug design and developing effective therapeutic strategies. Treatment with sequential first-, second-, and third-generation ALK inhibitors can select for compound mutations that confer high-level resistance to ALK-targeted therapies. A more efficacious long-term strategy may be up-front treatment with a third-generation ALK inhibitor to prevent the emergence of on-target resistance. .
Topics: Aminopyridines; Anaplastic Lymphoma Kinase; Animals; Cell Line, Tumor; Crizotinib; Drug Resistance, Neoplasm; Ethylnitrosourea; Female; Humans; Lactams; Lactams, Macrocyclic; Lung Neoplasms; Mice; Mutation; Oncogene Proteins, Fusion; Protein Kinase Inhibitors; Pyrazoles; Exome Sequencing; Xenograft Model Antitumor Assays
PubMed: 29650534
DOI: 10.1158/2159-8290.CD-17-1256 -
Experimental Animals 2015Considering the suitability of laboratory rats in epilepsy research, we and other groups have been developing genetic models of epilepsy in this species. After epileptic... (Review)
Review
Considering the suitability of laboratory rats in epilepsy research, we and other groups have been developing genetic models of epilepsy in this species. After epileptic rats or seizure-susceptible rats were sporadically found in outbred stocks, the epileptic traits were usually genetically-fixed by selective breeding. So far, the absence seizure models GAERS and WAG/Rij, audiogenic seizure models GEPR-3 and GEPR-9, generalized tonic-clonic seizure models IER, NER and WER, and Canavan-disease related epileptic models TRM and SER have been established. Dissection of the genetic bases including causative genes in these epileptic rat models would be a significant step toward understanding epileptogenesis. N-ethyl-N-nitrosourea (ENU) mutagenesis provides a systematic approach which allowed us to develop two novel epileptic rat models: heat-induced seizure susceptible (Hiss) rats with an Scn1a missense mutation and autosomal dominant lateral temporal epilepsy (ADLTE) model rats with an Lgi1 missense mutation. In addition, we have established episodic ataxia type 1 (EA1) model rats with a Kcna1 missense mutation derived from the ENU-induced rat mutant stock, and identified a Cacna1a missense mutation in a N-Methyl-N-nitrosourea (MNU)-induced mutant rat strain GRY, resulting in the discovery of episodic ataxia type 2 (EA2) model rats. Thus, epileptic rat models have been established on the two paths: 'phenotype to gene' and 'gene to phenotype'. In the near future, development of novel epileptic rat models will be extensively promoted by the use of sophisticated genome editing technologies.
Topics: Animals; Disease Models, Animal; Epilepsy; Ethylnitrosourea; Genetic Engineering; Genetic Predisposition to Disease; Genome; Intercellular Signaling Peptides and Proteins; Kv1.1 Potassium Channel; Models, Genetic; Mutagenesis; Mutation, Missense; NAV1.1 Voltage-Gated Sodium Channel; Phenotype; Proteins; Rats
PubMed: 25312505
DOI: 10.1538/expanim.14-0066 -
Briefings in Functional Genomics &... Nov 2008TILLING, for Targeting Induced Local Lesions in Genomes, is a reverse genetics strategy that identifies mutations in specific genes of interest in chemically mutagenized... (Review)
Review
TILLING, for Targeting Induced Local Lesions in Genomes, is a reverse genetics strategy that identifies mutations in specific genes of interest in chemically mutagenized populations. First described in 2000 for mutation detection in Arabidopsis, TILLING is now used in a wide range of plants including soybean, rice, barley and maize as well as for animal model systems, including Arabidopsis, Drosophila, Caenorhabditis elegans, rat, medaka and zebrafish and for the discovery of naturally occurring polymorphisms in humans. This review summarizes current TILLING methodologies as they have been applied to the zebrafish, ongoing TILLING projects and resources in the zebrafish community, and the future of zebrafish TILLING.
Topics: Animals; Base Pair Mismatch; Ethylnitrosourea; Models, Biological; Mutagenesis; Mutagens; Zebrafish
PubMed: 19028802
DOI: 10.1093/bfgp/eln046 -
Medicine Sep 2022Hepatocellular carcinoma (HCC) is a common malignant tumor worldwide with a poor prognosis. Recent studies have shown that the occurrence, development and prognosis of...
BACKGROUND
Hepatocellular carcinoma (HCC) is a common malignant tumor worldwide with a poor prognosis. Recent studies have shown that the occurrence, development and prognosis of liver cancer are closely related to tumor microenvironment (TME) and tumor immune infiltration.
METHODS
Therefore, important information on various diseases can be obtained from public databases such as The Cancer Gene Atlas (TCGA), and ideas or schemes that may be effective for the treatment of various diseases can be screened and analyzed by screening various conditions. In this study, 424 cases of liver hepatocellular carcinoma (LIHC) in the TCGA database and CIBERSORT algorithm were used to calculate the proportion of tumor-invasive immune cells. Combined with the clinical data from TCGA database, it was concluded that T cells regulatory (Tregs) were correlated with the development and prognosis of HCC. Cox regression analysis was used to screen differentially expressed genes, and survival analysis was performed according to the screened differentially expressed genes to see whether there was a significant association with the prognosis of HCC. Then gene ontology and kyoto encyclopedia of genes and genomes analysis of differentially expressed genes were carried out to explore the possibility of differentially expressed genes becoming potential therapeutic targets of HCC.
RESULTS
Finally, I identified the gene centromere protein o (CENPO), which is associated with immune cells and improve the prognosis of HCC.
CONCLUSION
CENPO may be a potential biological therapeutic target for hepatocellular treatment.
Topics: Carcinoma, Hepatocellular; Cyclic N-Oxides; Ethylnitrosourea; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Humans; Liver Neoplasms; Tumor Microenvironment
PubMed: 36107579
DOI: 10.1097/MD.0000000000030453 -
Journal of Clinical Pathology.... 1977
Review
Topics: Altitude; Animals; Carotid Body; Carotid Body Tumor; Cytoplasm; Erythropoiesis; Ethylnitrosourea; Humans; Hyperplasia; Hypoxia; Microscopy, Electron; Paraganglioma, Extra-Adrenal; Sex Factors; Sudden Infant Death; Time Factors
PubMed: 198435
DOI: 10.1136/jcp.s3-11.1.30 -
Genes Jan 2020Human midfacial clefting is a rare subset of orofacial clefting and in severe cases, the cleft separates the nostrils splitting the nose into two independent structures....
Human midfacial clefting is a rare subset of orofacial clefting and in severe cases, the cleft separates the nostrils splitting the nose into two independent structures. To begin to understand the morphological and genetic causes of midfacial clefting we recovered the mouse line. embryos develop a complete midfacial cleft through the lip, and snout closely modelling human midfacial clefting. The mouse line has ethylnitrosourea (ENU)-induced missense mutations in and . The mutations segregate with the cleft face phenotype. Importantly, the nasal cartilages and surrounding bones are patterned and develop normal morphology, except for the lateral displacement because of the cleft. We conclude that the midfacial cleft arises from the failure of the medial convergence of the paired medial nasal prominences between E10.5 to E11.5 rather than defective cell proliferation and death. Our work uncovers a novel mouse model and mechanism for the etiology of midfacial clefting.
Topics: Aldehyde Oxidoreductases; Animals; Cleft Lip; Cleft Palate; Disease Models, Animal; Ethylnitrosourea; Mice; Mice, Mutant Strains; Mutagenesis; Mutation, Missense; Transcription Factors
PubMed: 31940751
DOI: 10.3390/genes11010083 -
Nucleic Acids Research May 1990The solution conformation of eight leucine tRNAs from Phaseolus vulgaris, baker's yeast and Escherichia coli, characterized by long variable regions, and the interaction...
Solution conformation of several free tRNALeu species from bean, yeast and Escherichia coli and interaction of these tRNAs with bean cytoplasmic Leucyl-tRNA synthetase. A phosphate alkylation study with ethylnitrosourea.
The solution conformation of eight leucine tRNAs from Phaseolus vulgaris, baker's yeast and Escherichia coli, characterized by long variable regions, and the interaction of four of them with bean cytoplasmic leucyl-tRNA synthetase were studied by phosphate mapping with ethylnitrosourea. Phosphate reactivities in the variable regions agree with the existence of RNA helices closed by miniloops. At the junction of these regions with the T-stem, phosphate 48 is strongly protected, in contrast to small variable region tRNAs where P49 is protected. The constant protection of P22 is another characteristics of leucine tRNAs. Conformational differences between leucine isoacceptors concern the anticodon region, the D-arm and the variable region. In several parts of free tRNALeu species, e.g. in the T-loop, phosphate reactivities are similar to those found in tRNAs of other specificities, indicating conformational similarities among tRNAs. Phosphate alkylation of four leucine tRNAs complexed to leucyl-tRNA synthetase indicates that the 3'-side of the anticodon stem, the D-stem and the hinge region between the anticodon and D-stems are in contact with the plant enzyme.
Topics: Alkylation; Amino Acyl-tRNA Synthetases; Base Sequence; Electrophoresis, Polyacrylamide Gel; Escherichia coli; Ethylnitrosourea; Fabaceae; Leucine-tRNA Ligase; Molecular Sequence Data; Nucleic Acid Conformation; Phosphates; Plants, Medicinal; RNA, Transfer, Amino Acid-Specific; RNA, Transfer, Leu; Saccharomyces cerevisiae; Solutions
PubMed: 2187177
DOI: 10.1093/nar/18.9.2589 -
American Journal of Physiology.... Jan 2011Mutagenesis of mice with N-ethyl-N-nitrosourea (ENU) is a phenotype-driven approach to unravel gene function and discover new biological pathways. Phenotype-driven... (Review)
Review
Mutagenesis of mice with N-ethyl-N-nitrosourea (ENU) is a phenotype-driven approach to unravel gene function and discover new biological pathways. Phenotype-driven approaches have the advantage of making no assumptions about the function of genes and their products and have been successfully applied to the discovery of novel gene-phenotype relationships in many physiological systems. ENU mutagenesis of mice is used in many large-scale and more focused projects to generate and identify novel mouse models for the study of gene functions and human disease. This review examines the strategies and tools used in ENU mutagenesis screens to efficiently generate and identify functional mutations.
Topics: Animals; Breeding; Disease Models, Animal; Ethylnitrosourea; Female; Genes; Genome; Genome-Wide Association Study; Inflammatory Bowel Diseases; Male; Mice; Mutagenesis; Phenotype
PubMed: 20947703
DOI: 10.1152/ajpgi.00343.2010 -
Experimental Hematology Nov 2022Therapy-related myeloid neoplasms (t-MNs) share many clinical and molecular characteristics with AML de novo in the elderly. One common factor is that they arise in the...
Therapy-related myeloid neoplasms (t-MNs) share many clinical and molecular characteristics with AML de novo in the elderly. One common factor is that they arise in the setting of chronic inflammation, likely because of advanced age or chemotherapy-induced senescence. Here, we examined the effect of haploinsufficient loss of the del(5q) tumor suppressor gene, EGR1, commonly deleted in high-risk MNs. In mice, under the exogenous stress of either serial transplant or successive doses of the alkylating agent N-ethyl-nitrosourea (ENU), Egr1-haploinsufficient hematopoietic stem cells (HSCs) exhibit a clonal advantage. Complete loss of EGR1 function is incompatible with transformation; mutations of EGR1 are rare and are not observed in the remaining allele in del(5q) patients, and complete knockout of Egr1 in mice leads to HSC exhaustion. Using chromatin immunoprecipitation sequencing (ChIP-seq), we identified EGR1 binding sites in human CD34 cord blood-derived stem and progenitor cells (HSPCs) and found that EGR1 binds genes critical for stem cell differentiation, inflammatory signaling, and the DNA damage response. Notably, in the chromosome 5 sequences frequently deleted in patients, there is a significant enrichment of innate and inflammatory genes, which may confer a fitness advantage in an inflammatory environment. Short hairpin RNA (shRNA)-mediated silencing of EGR1 biases HSPCs toward a self-renewal transcriptional signature. In the absence of EGR1, HSPCs are characterized by upregulated MYC-driven proliferative signals, downregulated CDKN1A (p21), disrupted DNA damage response, and downregulated inflammation-adaptations anticipated to confer a relative fitness advantage for stem cells especially in an environment of chronic inflammation.
Topics: Humans; Mice; Animals; Aged; Haploinsufficiency; Hematopoietic Stem Cells; Antigens, CD34; Ethylnitrosourea; Inflammation; Early Growth Response Protein 1
PubMed: 35995095
DOI: 10.1016/j.exphem.2022.08.003 -
The Journal of Investigative Dermatology Sep 2019The primary goals of modern genetics are to identify disease-causing mutations and to define the functions of genes in biological processes. Two complementary... (Review)
Review
The primary goals of modern genetics are to identify disease-causing mutations and to define the functions of genes in biological processes. Two complementary approaches, reverse and forward genetics, can be used to achieve this goal. Reverse genetics is a gene-driven approach that comprises specific gene targeting followed by phenotypic assessment. Conversely, forward genetics is a phenotype-driven approach that involves the phenotypic screening of organisms with randomly induced mutations followed by subsequent identification of the causative mutations (i.e., those responsible for phenotype). In this article, we focus on how forward genetics in mice can be used to explore dermatologic disease. We outline mouse mutagenesis with the chemical N-ethyl-N-nitrosourea and the strategy used to instantaneously identify mutations that are causative of specific phenotypes. Furthermore, we summarize the types of phenotypic screens that can be performed to explore various aspects of dermatologic disease.
Topics: Animals; Breeding; Disease Models, Animal; Ethylnitrosourea; Gene Expression Regulation; Genetic Testing; Humans; Mice; Mutagenesis; Mutagens; Mutation; Phenotype; Research Design; Signal Transduction; Skin; Skin Diseases
PubMed: 31445571
DOI: 10.1016/j.jid.2019.04.013