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1.
Nat Cancer ; 3(6): 681-695, 2022 06.
Artículo en Inglés | MEDLINE | ID: mdl-35437317

RESUMEN

Despite advances in precision medicine, the clinical prospects for patients with ovarian and uterine cancers have not substantially improved. Here, we analyzed genome-scale CRISPR-Cas9 loss-of-function screens across 851 human cancer cell lines and found that frequent overexpression of SLC34A2-encoding a phosphate importer-is correlated with sensitivity to loss of the phosphate exporter XPR1, both in vitro and in vivo. In patient-derived tumor samples, we observed frequent PAX8-dependent overexpression of SLC34A2, XPR1 copy number amplifications and XPR1 messenger RNA overexpression. Mechanistically, in SLC34A2-high cancer cell lines, genetic or pharmacologic inhibition of XPR1-dependent phosphate efflux leads to the toxic accumulation of intracellular phosphate. Finally, we show that XPR1 requires the novel partner protein KIDINS220 for proper cellular localization and activity, and that disruption of this protein complex results in acidic "vacuolar" structures preceding cell death. These data point to the XPR1-KIDINS220 complex and phosphate dysregulation as a therapeutic vulnerability in ovarian cancer.


Asunto(s)
Proteínas de la Membrana , Proteínas del Tejido Nervioso , Neoplasias Ováricas , Femenino , Humanos , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/genética , Neoplasias Ováricas/metabolismo , Fosfatos/farmacología , Receptores Acoplados a Proteínas G/genética , Receptores Virales/genética , Receptor de Retrovirus Xenotrópico y Politrópico/genética , Receptor de Retrovirus Xenotrópico y Politrópico/metabolismo
2.
Sci Signal ; 14(711): eabc4520, 2021 Nov 30.
Artículo en Inglés | MEDLINE | ID: mdl-34846918

RESUMEN

Aberrant activation of the RAS family of guanosine triphosphatases (GTPases) is prevalent in lung adenocarcinoma, with somatic mutation of KRAS occurring in ~30% of tumors. We previously identified somatic mutations and amplifications of the gene encoding RAS family GTPase RIT1 in lung adenocarcinomas. To explore the biological pathways regulated by RIT1 and how they relate to the oncogenic KRAS network, we performed quantitative proteomic, phosphoproteomic, and transcriptomic profiling of isogenic lung epithelial cells in which we ectopically expressed wild-type or cancer-associated variants of RIT1 and KRAS. We found that both mutant KRAS and mutant RIT1 promoted canonical RAS signaling and that overexpression of wild-type RIT1 partially phenocopied oncogenic RIT1 and KRAS, including induction of epithelial-to-mesenchymal transition. Our findings suggest that RIT1 protein abundance is a factor in its pathogenic function. Therefore, chromosomal amplification of wild-type RIT1 in lung and other cancers may be tumorigenic.


Asunto(s)
Oncogenes , Transducción de Señal , Proteínas ras , Células HEK293 , Humanos , Proteínas ras/genética
3.
Nat Commun ; 12(1): 4789, 2021 08 09.
Artículo en Inglés | MEDLINE | ID: mdl-34373451

RESUMEN

CRISPR-based cancer dependency maps are accelerating advances in cancer precision medicine, but adequate functional maps are limited to the most common oncogenes. To identify opportunities for therapeutic intervention in other rarer subsets of cancer, we investigate the oncogene-specific dependencies conferred by the lung cancer oncogene, RIT1. Here, genome-wide CRISPR screening in KRAS, EGFR, and RIT1-mutant isogenic lung cancer cells identifies shared and unique vulnerabilities of each oncogene. Combining this genetic data with small-molecule sensitivity profiling, we identify a unique vulnerability of RIT1-mutant cells to loss of spindle assembly checkpoint regulators. Oncogenic RIT1M90I weakens the spindle assembly checkpoint and perturbs mitotic timing, resulting in sensitivity to Aurora A inhibition. In addition, we observe synergy between mutant RIT1 and activation of YAP1 in multiple models and frequent nuclear overexpression of YAP1 in human primary RIT1-mutant lung tumors. These results provide a genome-wide atlas of oncogenic RIT1 functional interactions and identify components of the RAS pathway, spindle assembly checkpoint, and Hippo/YAP1 network as candidate therapeutic targets in RIT1-mutant lung cancer.


Asunto(s)
Neoplasias Pulmonares/genética , Oncogenes/genética , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Ciclo Celular/genética , Línea Celular Tumoral , Receptores ErbB/genética , Femenino , Técnicas de Inactivación de Genes , Ensayos Analíticos de Alto Rendimiento , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Masculino , Ratones , Terapia Molecular Dirigida , Mutación , Células 3T3 NIH , Proteínas Proto-Oncogénicas p21(ras)/genética , Factores de Transcripción/genética , Ensayos Antitumor por Modelo de Xenoinjerto , Proteínas Señalizadoras YAP , Proteínas ras
4.
Nat Genet ; 53(4): 529-538, 2021 04.
Artículo en Inglés | MEDLINE | ID: mdl-33753930

RESUMEN

Exciting therapeutic targets are emerging from CRISPR-based screens of high mutational-burden adult cancers. A key question, however, is whether functional genomic approaches will yield new targets in pediatric cancers, known for remarkably few mutations, which often encode proteins considered challenging drug targets. To address this, we created a first-generation pediatric cancer dependency map representing 13 pediatric solid and brain tumor types. Eighty-two pediatric cancer cell lines were subjected to genome-scale CRISPR-Cas9 loss-of-function screening to identify genes required for cell survival. In contrast to the finding that pediatric cancers harbor fewer somatic mutations, we found a similar complexity of genetic dependencies in pediatric cancer cell lines compared to that in adult models. Findings from the pediatric cancer dependency map provide preclinical support for ongoing precision medicine clinical trials. The vulnerabilities observed in pediatric cancers were often distinct from those in adult cancer, indicating that repurposing adult oncology drugs will be insufficient to address childhood cancers.


Asunto(s)
Mapeo Cromosómico/métodos , Regulación Neoplásica de la Expresión Génica , Genoma Humano , Mutación , Proteínas de Neoplasias/genética , Neoplasias/genética , Adulto , Proteína 9 Asociada a CRISPR/genética , Proteína 9 Asociada a CRISPR/metabolismo , Sistemas CRISPR-Cas , Línea Celular Tumoral , Niño , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas , Edición Génica , Perfilación de la Expresión Génica , Predisposición Genética a la Enfermedad , Humanos , Proteínas de Neoplasias/clasificación , Proteínas de Neoplasias/metabolismo , Neoplasias/metabolismo , Neoplasias/patología , ARN Guía de Kinetoplastida/genética , ARN Guía de Kinetoplastida/metabolismo
5.
Exp Ther Med ; 8(5): 1351-1356, 2014 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-25289022

RESUMEN

In the present study, the tetracycline-off and Cre/loxP systems were combined to gain temporal and spatial control of transgene expression. Mice were generated that carried three transgenes: Tie2-tTA, tet-O-Cre and either the ZEG or ZAP reporter. Tie2-tTA directs expression of tetracycline-controlled transactivator (tTA) in endothelial and hematopoietic cells under the control of the Tie2 promoter. Tet-O-Cre produces Cre recombinase from a minimal promoter containing the tet-operator (tetO). ZEG or ZAP contains a strong promoter and a loxP-flanked stop sequence, followed by an enhanced green fluorescence protein (EGFP) or human placental alkaline phosphatase (hPLAP) reporter. In the presence of tetracycline, the tTA transactivator produced by Tie-2-tTA is disabled and Cre is not expressed. In the absence of tetracycline, the tTA binds tet-O-Cre to drive the expression of Cre, which recombines the loxP sites of the ZEG or ZAP transgene and results in reporter gene expression. In the present study, the expression of the ZEG or ZAP reporter genes in embryos and adult animals with and without tetracycline treatment was examined. In the presence of tetracycline, no reporter gene expression was observed. When tetracycline was withdrawn, Cre excision was activated and the reporter genes were detected in endothelial and hematopoietic cells. These results demonstrate that this system may be used to bypass embryonic lethality and access adult phenotypes.

6.
Int J Oncol ; 45(5): 1997-2004, 2014 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-25175815

RESUMEN

The Notch1 signaling pathway is essential for hematopoietic development. However, the effects of postnatal activation of Notch1 signaling on hematopoietic system is not yet fully understood. We previously generated ZEG­IC­Notch1 transgenic mice that have a floxed ß­geo/stop signal between a CMV promoter and intracellular domain of Notch1 (IC­Notch1). Constitutively active IC­Notch1 is silent until the introduction of Cre recombinase. In this study, endothelial/hematopoietic specific expression of IC­Notch1 in double transgenic ZEG­IC­Notch1/Tie2­Cre embryos induced embryonic lethality at E9.5 with defects in vascular system but not in hematopoietic system. Inducible IC­Notch1 expression in adult mice was achieved by using tetracycline regulated Cre system. The ZEG­IC­Notch1/Tie2­tTA/tet­O­Cre triple transgenic mice survived embryonic development when maintained on tetracycline. Post­natal withdrawal of tetracycline induced expression of IC­Notch1 transgene in hematopoietic cells of adult mice. The triple transgenic mice displayed extensive T­cell infiltration in multiple organs and T­cell malignancy of lymph nodes. In addition, the protein levels of p53 and alternative reading frame (ARF) were decreased in lymphoma­like neoplasms from the triple transgenic mice while their mRNA expression remained unchanged, suggesting that IC­Notch1 might repress ARF­p53 pathway by a post­transcriptional mechanism. This study demonstrated that activation of constitutive Notch1 signaling after embryonic development alters adult hematopoiesis and induces T­cell malignancy.


Asunto(s)
Transformación Celular Neoplásica/genética , Ganglios Linfáticos/patología , Receptor Notch1/genética , Linfocitos T/inmunología , Animales , Transformación Celular Neoplásica/patología , Embrión de Mamíferos , Regulación del Desarrollo de la Expresión Génica , Humanos , Integrasas , Ganglios Linfáticos/crecimiento & desarrollo , Ratones , Ratones Transgénicos , Regiones Promotoras Genéticas , Receptor Notch1/biosíntesis , Transducción de Señal/genética , Linfocitos T/patología
7.
Contraception ; 69(6): 493-6, 2004 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-15157795

RESUMEN

OBJECTIVE: To gain a better understanding of ethnic Chinese women's perceptions and experiences of using barrier and rhythm methods of contraception in order to improve contraceptive counseling at abortion clinics. DESIGN: Qualitative descriptive study. SETTING: Urban abortion clinic. PARTICIPANTS: Forty ethnic Chinese women presenting for abortion. METHOD: Data were collected in semi-structured interviews by one interviewer who is fluent in English, Mandarin and Cantonese. Transcribed interviews were systematically analyzed to identify salient themes. MAIN FINDINGS: All of the women interviewed had used condoms (none with spermicide), 20 had used rhythm and 17 withdrawal, usually a combination of two or three of these methods. Many women noted that these methods are under male control and talked about the difficulty negotiating their use with partners. The majority of women using rhythm were unable to correctly identify "safe periods."


Asunto(s)
Anticoncepción/psicología , Conocimientos, Actitudes y Práctica en Salud , Pueblo Asiatico , Colombia Británica , Femenino , Humanos , Entrevistas como Asunto
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