RESUMEN
Most cancers exhibit aneuploidy, but its functional significance in tumor development is controversial. Here, we describe ReDACT (Restoring Disomy in Aneuploid cells using CRISPR Targeting), a set of chromosome engineering tools that allow us to eliminate specific aneuploidies from cancer genomes. Using ReDACT, we created a panel of isogenic cells that have or lack common aneuploidies, and we demonstrate that trisomy of chromosome 1q is required for malignant growth in cancers harboring this alteration. Mechanistically, gaining chromosome 1q increases the expression of MDM4 and suppresses p53 signaling, and we show that TP53 mutations are mutually exclusive with 1q aneuploidy in human cancers. Thus, tumor cells can be dependent on specific aneuploidies, raising the possibility that these "aneuploidy addictions" could be targeted as a therapeutic strategy.
Asunto(s)
Proteínas de Ciclo Celular , Edición Génica , Neoplasias , Oncogenes , Trisomía , Proteína p53 Supresora de Tumor , Humanos , Proteínas de Ciclo Celular/genética , Mutación , Neoplasias/genética , Neoplasias/terapia , Proteínas Proto-Oncogénicas/metabolismo , Edición Génica/métodos , Proteína p53 Supresora de Tumor/genética , Carcinogénesis/genéticaRESUMEN
Most cancers exhibit aneuploidy, but its functional significance in tumor development is controversial. Here, we describe ReDACT (Restoring Disomy in Aneuploid cells using CRISPR Targeting), a set of chromosome engineering tools that allow us to eliminate specific aneuploidies from cancer genomes. Using ReDACT, we created a panel of isogenic cells that have or lack common aneuploidies, and we demonstrate that trisomy of chromosome 1q is required for malignant growth in cancers harboring this alteration. Mechanistically, gaining chromosome 1q increases the expression of MDM4 and suppresses TP53 signaling, and we show that TP53 mutations are mutually-exclusive with 1q aneuploidy in human cancers. Thus, specific aneuploidies play essential roles in tumorigenesis, raising the possibility that targeting these "aneuploidy addictions" could represent a novel approach for cancer treatment.
RESUMEN
Aneuploidy is a ubiquitous feature of human tumors, but the acquisition of aneuploidy typically antagonizes cellular fitness. To investigate how aneuploidy could contribute to tumor growth, we triggered periods of chromosomal instability (CIN) in human cells and then exposed them to different culture environments. We discovered that transient CIN reproducibly accelerates the acquisition of resistance to anti-cancer therapies. Single-cell sequencing revealed that these resistant populations develop recurrent aneuploidies, and independently deriving one chromosome-loss event that was frequently observed in paclitaxel-resistant cells was sufficient to decrease paclitaxel sensitivity. Finally, we demonstrated that intrinsic levels of CIN correlate with poor responses to numerous therapies in human tumors. Our results show that, although CIN generally decreases cancer cell fitness, it also provides phenotypic plasticity to cancer cells that can allow them to adapt to diverse stressful environments. Moreover, our findings suggest that aneuploidy may function as an under-explored cause of therapy failure.
Asunto(s)
Aneuploidia , Inestabilidad Cromosómica/genética , Neoplasias/tratamiento farmacológico , Neoplasias/patología , Línea Celular Tumoral , Resistencia a Medicamentos/efectos de los fármacos , Ambiente , Humanos , Neoplasias/genética , Resultado del TratamientoRESUMEN
Aneuploidy has been recognized as a hallmark of tumorigenesis for more than 100 years, but the connection between chromosomal errors and malignant growth has remained obscure. New evidence emerging from both basic and clinical research has illuminated a complicated relationship: despite its frequency in human tumours, aneuploidy is not a universal driver of cancer development and instead can exert substantial tumour-suppressive effects. The specific consequences of aneuploidy are highly context dependent and are influenced by a cell's genetic and environmental milieu. In this Review, we discuss the diverse facets of cancer biology that are shaped by aneuploidy, including metastasis, drug resistance and immune recognition, and we highlight aneuploidy's distinct roles as both a tumour promoter and an anticancer vulnerability.
Asunto(s)
Aneuploidia , Resistencia a Antineoplásicos/genética , Neoplasias/genética , Escape del Tumor/inmunología , Animales , Carcinogénesis/genética , Carcinogénesis/inmunología , Línea Celular Tumoral , Transformación Celular Neoplásica/genética , Transformación Celular Neoplásica/inmunología , Modelos Animales de Enfermedad , Síndrome de Down/complicaciones , Síndrome de Down/genética , Resistencia a Antineoplásicos/inmunología , Humanos , Ratones , Metástasis de la Neoplasia/genética , Metástasis de la Neoplasia/inmunología , Neoplasias/inmunología , Fenotipo , Escape del Tumor/genética , Microambiente Tumoral/genética , Microambiente Tumoral/inmunologíaRESUMEN
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RESUMEN
The factors mediating fatal SARS-CoV-2 infections are poorly understood. Here, we show that cigarette smoke causes a dose-dependent upregulation of angiotensin converting enzyme 2 (ACE2), the SARS-CoV-2 receptor, in rodent and human lungs. Using single-cell sequencing data, we demonstrate that ACE2 is expressed in a subset of secretory cells in the respiratory tract. Chronic smoke exposure triggers the expansion of this cell population and a concomitant increase in ACE2 expression. In contrast, quitting smoking decreases the abundance of these secretory cells and reduces ACE2 levels. Finally, we demonstrate that ACE2 expression is responsive to inflammatory signaling and can be upregulated by viral infections or interferon treatment. Taken together, these results may partially explain why smokers are particularly susceptible to severe SARS-CoV-2 infections. Furthermore, our work identifies ACE2 as an interferon-stimulated gene in lung cells, suggesting that SARS-CoV-2 infections could create positive feedback loops that increase ACE2 levels and facilitate viral dissemination.
Asunto(s)
Células Epiteliales Alveolares/metabolismo , Infecciones por Coronavirus/epidemiología , Interferones/metabolismo , Peptidil-Dipeptidasa A/genética , Neumonía Viral/epidemiología , Mucosa Respiratoria/metabolismo , Contaminación por Humo de Tabaco/efectos adversos , Fumar Tabaco/genética , Adulto , Anciano , Enzima Convertidora de Angiotensina 2 , Animales , COVID-19 , Células CACO-2 , Células Cultivadas , Femenino , Células HCT116 , Humanos , Interferones/genética , Masculino , Ratones , Persona de Mediana Edad , Pandemias , Peptidil-Dipeptidasa A/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , RNA-Seq , Ratas , Transducción de Señal , Análisis de la Célula Individual , Fumar Tabaco/epidemiología , Fumar Tabaco/metabolismo , Regulación hacia ArribaRESUMEN
The CRISPR/Cas9 system is a powerful tool for genome editing, wherein the RNA-guided nuclease Cas9 can be directed to introduce double-stranded breaks (DSBs) at a targeted locus. In mammalian cells, these DSBs are typically repaired through error-prone processes, resulting in insertions or deletions (indels) at the targeted locus. Researchers can use these Cas9-mediated lesions to probe the consequences of loss-of-function perturbations in genes of interest. Here, we describe an optimized protocol to identify specific genes required for cancer cell fitness through a CRISPR-mediated cellular competition assay. Identifying these genetic dependencies is of utmost importance, as they provide potential targets for anti-cancer drug development. This protocol provides researchers with a robust and scalable approach to investigate gene dependencies in a variety of cell lines and cancer types and to validate the results of high-throughput or whole-genome screens.