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1.
Breast Cancer Res Treat ; 174(1): 143-155, 2019 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-30484104

RESUMEN

PURPOSE AND METHODS: In human basal-like breast cancer, mutations and deletions in TP53 and BRCA1 are frequent oncogenic events. Thus, we interbred mice expressing the CRE-recombinase with mice harboring loxP sites at TP53 and BRCA1 (K14-Cre; p53f/f Brca1f/f) to test the hypothesis that tissue-specific deletion of TP53 and BRCA1 would give rise to tumors reflective of human basal-like breast cancer. RESULTS: In support of our hypothesis, these transgenic mice developed tumors that express basal-like cytokeratins and demonstrated intrinsic gene expression features similar to human basal-like tumors. Array comparative genomic hybridization revealed a striking conservation of copy number alterations between the K14-Cre; p53f/f Brca1f/f mouse model and human basal-like breast cancer. Conserved events included MYC amplification, KRAS amplification, and RB1 loss. Microarray analysis demonstrated that these DNA copy number events also led to corresponding changes in signatures of pathway activation including high proliferation due to RB1 loss. K14-Cre; p53f/f Brca1f/f also matched human basal-like breast cancer for a propensity to have immune cell infiltrates. Given the long latency of K14-Cre; p53f/f Brca1f/f tumors (~ 250 days), we created tumor syngeneic transplant lines, as well as in vitro cell lines, which were tested for sensitivity to carboplatin and paclitaxel. These therapies invoked acute regression, extended overall survival, and resulted in gene expression signatures of an anti-tumor immune response. CONCLUSION: These findings demonstrate that this model is a valuable preclinical resource for the study of human basal-like breast cancer.


Asunto(s)
Modelos Animales de Enfermedad , Neoplasias Mamarias Experimentales/genética , Neoplasias Mamarias Experimentales/patología , Proteína p53 Supresora de Tumor/genética , Proteínas Supresoras de Tumor/genética , Animales , Proteína BRCA1 , Femenino , Humanos , Ratones , Ratones Transgénicos
2.
Nature ; 448(7155): 807-10, 2007 Aug 16.
Artículo en Inglés | MEDLINE | ID: mdl-17676035

RESUMEN

Germline mutation in serine/threonine kinase 11 (STK11, also called LKB1) results in Peutz-Jeghers syndrome, characterized by intestinal hamartomas and increased incidence of epithelial cancers. Although uncommon in most sporadic cancers, inactivating somatic mutations of LKB1 have been reported in primary human lung adenocarcinomas and derivative cell lines. Here we used a somatically activatable mutant Kras-driven model of mouse lung cancer to compare the role of Lkb1 to other tumour suppressors in lung cancer. Although Kras mutation cooperated with loss of p53 or Ink4a/Arf (also known as Cdkn2a) in this system, the strongest cooperation was seen with homozygous inactivation of Lkb1. Lkb1-deficient tumours demonstrated shorter latency, an expanded histological spectrum (adeno-, squamous and large-cell carcinoma) and more frequent metastasis compared to tumours lacking p53 or Ink4a/Arf. Pulmonary tumorigenesis was also accelerated by hemizygous inactivation of Lkb1. Consistent with these findings, inactivation of LKB1 was found in 34% and 19% of 144 analysed human lung adenocarcinomas and squamous cell carcinomas, respectively. Expression profiling in human lung cancer cell lines and mouse lung tumours identified a variety of metastasis-promoting genes, such as NEDD9, VEGFC and CD24, as targets of LKB1 repression in lung cancer. These studies establish LKB1 as a critical barrier to pulmonary tumorigenesis, controlling initiation, differentiation and metastasis.


Asunto(s)
Diferenciación Celular , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Quinasas de la Proteína-Quinasa Activada por el AMP , Proteínas Quinasas Activadas por AMP , Animales , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Modelos Animales de Enfermedad , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Genes Relacionados con las Neoplasias/genética , Genes Supresores de Tumor/fisiología , Genes p16 , Genes p53/genética , Genes ras/genética , Humanos , Ratones , Metástasis de la Neoplasia/genética , Proteínas Serina-Treonina Quinasas/deficiencia
3.
Am J Pathol ; 173(2): 536-44, 2008 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-18599611

RESUMEN

Endometrial cancer has been generally categorized into two broad groups of tumors, type I (TI) and type II (TII), with distinct epidemiological/clinical features and genetic alterations. Because telomere attrition appears to trigger genomic instability in certain cancers, we explored the role of telomere dysfunction in endometrial cancer by analyzing telomeres and other markers of telomere status in both tumor types. We describe a new method, telomere chromogenic in situ hybridization, which permitted us to detect cells with short telomeres relative to control (stromal) cells within the same tissue section. Using this method, we found that both types of tumor cells had short telomeres. However, only TII tumors were significantly associated with critical telomere shortening in adjacent, morphologically normal epithelium, suggesting that telomere shortening contributes to the initiation of TII but not TI tumors. To explore this hypothesis, we analyzed mice with critically short telomeres and documented distinctive endometrial lesions that histologically resembled the in situ precursor of TII serous carcinomas; these lesions have not been observed previously in TI mouse models of endometrial cancer. Based on this and previous studies, we propose a model in which telomere attrition contributes to the initiation of TII and progression of TI endometrial cancers.


Asunto(s)
Transformación Celular Neoplásica , Neoplasias Endometriales/patología , Endometrio/patología , Telómero/fisiología , Anciano , Aneuploidia , Animales , Femenino , Humanos , Hibridación in Situ/métodos , Ratones
4.
Genetics ; 177(1): 179-94, 2007 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-17660561

RESUMEN

Female infertility syndromes are among the most prevalent chronic health disorders in women, but their genetic basis remains unknown because of uncertainty regarding the number and identity of ovarian factors controlling the assembly, preservation, and maturation of ovarian follicles. To systematically discover ovarian fertility genes en masse, we employed a mouse model (Foxo3) in which follicles are assembled normally but then undergo synchronous activation. We developed a microarray-based approach for the systematic discovery of tissue-specific genes and, by applying it to Foxo3 ovaries and other samples, defined a surprisingly large set of ovarian factors (n = 348, approximately 1% of the mouse genome). This set included the vast majority of known ovarian factors, 44% of which when mutated produce female sterility phenotypes, but most were novel. Comparative profiling of other tissues, including microdissected oocytes and somatic cells, revealed distinct gene classes and provided new insights into oogenesis and ovarian function, demonstrating the utility of our approach for tissue-specific gene discovery. This study will thus facilitate comprehensive analyses of follicle development, ovarian function, and female infertility.


Asunto(s)
Factores de Transcripción Forkhead/fisiología , Genes/fisiología , Genoma , Infertilidad Femenina/metabolismo , Folículo Ovárico/fisiología , Animales , Northern Blotting , Femenino , Proteína Forkhead Box O3 , Factores de Transcripción Forkhead/genética , Perfilación de la Expresión Génica , Hibridación in Situ , Infertilidad Femenina/patología , Rayos Láser , Meiosis , Ciclo Menstrual , Ratones , Ratones Noqueados , Microdisección , Análisis de Secuencia por Matrices de Oligonucleótidos , Oocitos/fisiología , Oocitos/ultraestructura , Oogénesis/fisiología , Folículo Ovárico/ultraestructura , Sondas ARN , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
5.
Mol Immunol ; 44(10): 2719-28, 2007 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-17207856

RESUMEN

B-1 cells are important players in the first line of defense against pathogens. According to current models for the origin of B-1 cells, they either represent a separate lineage from conventional B-2 cells or differentiate from conventional B-2 cells via an intermediate, B-1(int), in response to positive selection by antigen. Here we show that Btk, a Tec family kinase that mediates B cell antigen receptor (BCR) signaling, is required at multiple stages of B-1 cell development. VH12 anti-phosphatidylcholine (PtC) IgH transgenic mice provide a model for the induced differentiation of B-1 cells. This transgene selects for PtC-reactive cells and induces them to adopt a B-1 phenotype. Both processes have been shown to depend on Btk. To determine whether this is secondary to a requirement for Btk in the development of mature B-2 cells, we crossed VH12 transgenic mice to mice expressing low levels of Btk. B-2 cell development occurs normally in Btk(lo) mice despite reduced responsiveness to BCR crosslinking. Analysis of VH12.Btk(lo) mice reveals that Btk regulates the B-1(int) to B-1 transition and/or the survival of splenic B-1 cells, in part via a mechanism independent of its role in BCR signaling. We also show that Btk mediates the survival of, and expression of IL-10 by, those B-1 cells that do develop and migrate to the peritoneum. Multiple roles for Btk in B-1 cell development and maintenance may explain the particular sensitivity of this population to mutations in components of Btk signaling pathways.


Asunto(s)
Linfocitos B/citología , Linfocitos B/inmunología , Linaje de la Célula , Proteínas Tirosina Quinasas/fisiología , Agammaglobulinemia Tirosina Quinasa , Animales , Linfocitos B/enzimología , Linaje de la Célula/genética , Supervivencia Celular , Ratones , Ratones Endogámicos , Ratones Transgénicos , Peritoneo/inmunología , Proteínas Tirosina Quinasas/genética , Receptores de Antígenos de Linfocitos B/inmunología , Receptores de IgE/inmunología
6.
Dis Model Mech ; 3(3-4): 181-93, 2010.
Artículo en Inglés | MEDLINE | ID: mdl-20142330

RESUMEN

Endometrial cancer--the most common malignancy of the female reproductive tract--arises from the specialized epithelial cells that line the inner surface of the uterus. Although significant advances have been made in our understanding of this disease in recent years, one significant limitation has been the lack of a diverse genetic toolkit for the generation of mouse models. We identified a novel endometrial-specific gene, Sprr2f, and developed a Sprr2f-Cre transgene for conditional gene targeting within endometrial epithelium. We then used this tool to generate a completely penetrant Lkb1 (also known as Stk11)-based mouse model of invasive endometrial cancer. Strikingly, female mice with homozygous endometrial Lkb1 inactivation did not harbor discrete endometrial neoplasms, but instead underwent diffuse malignant transformation of their entire endometrium with rapid extrauterine spread and death, suggesting that Lkb1 inactivation was sufficient to promote the development of invasive endometrial cancer. Mice with heterozygous endometrial Lkb1 inactivation only rarely developed tumors, which were focal and arose with much longer latency, arguing against the idea--suggested by some prior studies--that Lkb1 is a haploinsufficient tumor suppressor. Lastly, the finding that endometrial cancer cell lines were especially sensitive to the mTOR (mammalian target of rapamycin) inhibitor rapamycin prompted us to test its efficacy against Lkb1-driven endometrial cancers. Rapamycin monotherapy not only greatly slowed disease progression, but also led to striking regression of pre-existing tumors. These studies demonstrate that Lkb1 is a uniquely potent endometrial tumor suppressor, but also suggest that the clinical responses of some types of invasive cancers to mTOR inhibitors may be linked to Lkb1 status.


Asunto(s)
Neoplasias Endometriales/tratamiento farmacológico , Neoplasias Endometriales/enzimología , Péptidos y Proteínas de Señalización Intracelular/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/uso terapéutico , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/metabolismo , Sirolimus/uso terapéutico , Proteínas Quinasas Activadas por AMP , Animales , Línea Celular Tumoral , Clonación Molecular , Proteínas Ricas en Prolina del Estrato Córneo/genética , Modelos Animales de Enfermedad , Ensayos de Selección de Medicamentos Antitumorales , Neoplasias Endometriales/patología , Endometrio/efectos de los fármacos , Endometrio/metabolismo , Endometrio/patología , Activación Enzimática/efectos de los fármacos , Epitelio/efectos de los fármacos , Epitelio/metabolismo , Epitelio/patología , Femenino , Integrasas/metabolismo , Ratones , Ratones Transgénicos , Invasividad Neoplásica , Especificidad de Órganos/efectos de los fármacos , Penetrancia , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/deficiencia , Elementos de Respuesta/genética , Sirolimus/farmacología , Serina-Treonina Quinasas TOR
7.
PLoS One ; 4(4): e5137, 2009.
Artículo en Inglés | MEDLINE | ID: mdl-19340305

RESUMEN

Human Papilloma Virus (HPV) is the etiologic agent for cervical cancer. Yet, infection with HPV is not sufficient to cause cervical cancer, because most infected women develop transient epithelial dysplasias that spontaneously regress. Progression to invasive cancer has been attributed to diverse host factors such as immune or hormonal status, as no recurrent genetic alterations have been identified in cervical cancers. Thus, the pressing question as to the biological basis of cervical cancer progression has remained unresolved, hampering the development of novel therapies and prognostic tests. Here we show that at least 20% of cervical cancers harbor somatically-acquired mutations in the LKB1 tumor suppressor. Approximately one-half of tumors with mutations harbored single nucleotide substitutions or microdeletions identifiable by exon sequencing, while the other half harbored larger monoallelic or biallelic deletions detectable by multiplex ligation probe amplification (MLPA). Biallelic mutations were identified in most cervical cancer cell lines; HeLa, the first human cell line, harbors a homozygous 25 kb deletion that occurred in vivo. LKB1 inactivation in primary tumors was associated with accelerated disease progression. Median survival was only 13 months for patients with LKB1-deficient tumors, but >100 months for patients with LKB1-wild type tumors (P = 0.015, log rank test; hazard ratio = 0.25, 95% CI = 0.083 to 0.77). LKB1 is thus a major cervical tumor suppressor, demonstrating that acquired genetic alterations drive progression of HPV-induced dysplasias to invasive, lethal cancers. Furthermore, LKB1 status can be exploited clinically to predict disease recurrence.


Asunto(s)
Mutación , Proteínas Serina-Treonina Quinasas/genética , Neoplasias del Cuello Uterino/patología , Quinasas de la Proteína-Quinasa Activada por el AMP , Adulto , Anciano , Anciano de 80 o más Años , Alelos , Progresión de la Enfermedad , Femenino , Eliminación de Gen , Células HeLa , Humanos , Persona de Mediana Edad , Neoplasias del Cuello Uterino/genética
8.
Cancer Res ; 68(3): 759-66, 2008 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-18245476

RESUMEN

Mutations in the LKB1 tumor suppressor gene result in the Peutz-Jeghers syndrome, an autosomal dominant condition characterized by hamartomatous polyps of the gastrointestinal tract and a dramatically increased risk of epithelial malignancies at other sites, including the female reproductive tract. Here we show that female mice heterozygous for a null Lkb1 allele spontaneously develop highly invasive endometrial adenocarcinomas. To prove that these lesions were indeed due to Lkb1 inactivation, we introduced an adenoviral Cre vector into the uterine lumen of mice harboring a conditional allele of Lkb1. This endometrial-specific deletion of the Lkb1 gene provoked highly invasive and sometimes metastatic endometrial adenocarcinomas closely resembling those observed in Lkb1 heterozygotes. Tumors were extremely well differentiated and histopathologically distinctive and exhibited alterations in AMP-dependent kinase signaling. Although Lkb1 has been implicated in the establishment of cell polarity, and loss of polarity defines most endometrial cancers, Lkb1-driven endometrial cancers paradoxically exhibit (given their highly invasive phenotype) normal cell polarity and apical differentiation. In human endometrial cancers, Lkb1 expression was inversely correlated with tumor grade and stage, arguing that Lkb1 inactivation or down-regulation also contributes to endometrial cancer progression in women. This study shows that Lkb1 plays an important role in the malignant transformation of endometrium and that Lkb1 loss promotes a highly invasive phenotype.


Asunto(s)
Adenocarcinoma/genética , Transformación Celular Neoplásica/genética , Neoplasias Endometriales/genética , Proteínas Serina-Treonina Quinasas/deficiencia , Quinasas de la Proteína-Quinasa Activada por el AMP , Proteínas Quinasas Activadas por AMP , Adenocarcinoma/enzimología , Adenocarcinoma/patología , Animales , Polaridad Celular/genética , Transformación Celular Neoplásica/metabolismo , Transformación Celular Neoplásica/patología , Neoplasias Endometriales/enzimología , Neoplasias Endometriales/patología , Femenino , Regulación Enzimológica de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Silenciador del Gen , Genes Supresores de Tumor , Ratones , Complejos Multienzimáticos/metabolismo , Invasividad Neoplásica , Proteínas Serina-Treonina Quinasas/biosíntesis , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo
9.
Eur J Immunol ; 37(4): 1033-42, 2007 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-17372989

RESUMEN

The pre-BCR and the BCR regulate B cell development via a signalosome nucleated by the adaptor protein B cell linker protein (BLNK). Formation of this complex facilitates activation of phospholipase C (PLC) gamma2 by Bruton's tyrosine kinase (Btk). To determine whether Btk and PLCgamma2 also have separate functions, we generated Btk(-/-)PLCgamma2(-/-) mice. They demonstrated a block in development at the pre-B stage and increased pre-BCR surface expression. This phenotype was more severe than that of Btk(-/-) or PLCgamma2(-/-) mice. Although both Btk and PLCgamma2 were required for proliferation of splenic B cells in response to BCR cross-linking, they contributed differently to anti-IgM-induced phosphorylation of ERK. Btk(-/-) and PLCgamma2(-/-) mice each had a reduced frequency of Iglambda-expressing B cells and impaired migration of pre-B cells towards stromal cell-derived factor 1. However, the increase in pre-B cell malignancy that occurs in BLNK(-/-) mice in the absence of Btk was not observed in the absence of PLCgamma2. Thus, Btk and PLCgamma2 act both in concert and independently throughout B cell development.


Asunto(s)
Linfocitos B/citología , Linfocitos B/enzimología , Diferenciación Celular/inmunología , Fosfolipasa C gamma/fisiología , Proteínas Tirosina Quinasas/fisiología , Agammaglobulinemia Tirosina Quinasa , Animales , Células Cultivadas , Isoenzimas/deficiencia , Isoenzimas/genética , Isoenzimas/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Fosfolipasa C gamma/deficiencia , Fosfolipasa C gamma/genética , Proteínas Tirosina Quinasas/deficiencia , Proteínas Tirosina Quinasas/genética
10.
J Immunol ; 171(4): 1850-8, 2003 Aug 15.
Artículo en Inglés | MEDLINE | ID: mdl-12902486

RESUMEN

The development of autoimmunity is correlated with heightened sensitivity of B cells to B cell Ag receptor (BCR) cross-linking. BCR signals are down-regulated by Lyn, which phosphorylates inhibitory receptors. lyn(-/-) mice have reduced BCR signaling thresholds and develop autoantibodies, glomerulonephritis, splenomegaly due to myeloid hyperplasia, and increased B-1 cell numbers. Bruton's tyrosine kinase (Btk), a critical component of BCR signaling pathways, is required for autoantibody production in lyn(-/-) mice. It is unclear whether Btk mediates autoimmunity at the level of BCR signal transduction or B cell development, given that lyn(-/-)Btk(-/-) mice have a severe reduction in conventional B and B-1 cell numbers. To address this issue, we crossed a transgene expressing a low dosage of Btk (Btk(low)) in B cells to lyn(-/-)Btk(-/-) mice. Conventional B cell populations were restored to levels similar to those in lyn(-/-) mice. These cells were as hypersensitive to BCR cross-linking as lyn(-/-) B cells as measured by proliferation, Ca(2+) flux, and activation of extracellular signal-regulated kinase and Akt. However, lyn(-/-)Btk(low) mice did not produce anti-ssDNA, anti-dsDNA, anti-histone, or anti-histone/DNA IgM or IgG. They also lacked B-1 cells and did not exhibit splenomegaly. Thus, B cell hyperresponsiveness is insufficient for autoimmunity in lyn(-/-) mice. These studies implicate B-1 and/or myeloid cells as key contributors to the lyn(-/-) autoimmune phenotype.


Asunto(s)
Subgrupos de Linfocitos B/enzimología , Subgrupos de Linfocitos B/inmunología , Dosificación de Gen , Activación de Linfocitos/genética , Linfopenia/genética , Proteínas Tirosina Quinasas/deficiencia , Proteínas Tirosina Quinasas/genética , Familia-src Quinasas/deficiencia , Agammaglobulinemia Tirosina Quinasa , Animales , Autoanticuerpos/biosíntesis , Subgrupos de Linfocitos B/patología , División Celular/genética , División Celular/inmunología , Reactivos de Enlaces Cruzados/metabolismo , Linfopenia/enzimología , Linfopenia/inmunología , Linfopenia/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Proteínas Oncogénicas Virales/deficiencia , Proteínas Oncogénicas Virales/genética , Proteínas Oncogénicas Virales/fisiología , Proteínas Tirosina Quinasas/fisiología , Receptores de Antígenos de Linfocitos B/inmunología , Receptores de Antígenos de Linfocitos B/metabolismo , Transducción de Señal/genética , Transducción de Señal/inmunología , Esplenomegalia/genética , Esplenomegalia/inmunología , Transgenes/inmunología , Familia-src Quinasas/genética , Familia-src Quinasas/fisiología
11.
Int Immunol ; 15(3): 383-92, 2003 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-12618482

RESUMEN

Current models of lymphocyte activation suggest that formation of a signaling complex, or "signalosome", composed of Syk, Bruton's tyrosine kinase (Btk), phospholipase gamma2 and the adaptor protein B cell linker protein (BLNK) is critical for transmission of signals from the BCR. However, impaired B cell development in mice lacking each individual signalosome component has made it difficult to study the functional consequences of the formation of this complex in mature B cells. Sensitized genetic systems, commonly used in Drosophila, define signaling pathways by combining partial loss of function mutations in the components of interest. This allows genetic interactions to be observed in the absence of pleiotropic or lethal effects of complete deficiency of either gene. We used this approach to demonstrate that Btk and BLNK are limiting components of a common signaling pathway that mediates the mitogenic response of mature B cells to antigen. B cells from transgenic mice expressing a limiting dosage of Btk (Btk(lo)) have normal numbers of mature B cells that have reduced, but measurable, responses to BCR cross-linking. Haploinsufficiency of BLNK did not affect the development of Btk(lo) B cells. However, it exacerbated their defects in BCR-induced Ca(2+) flux, IkappaB degradation, and up-regulation of cyclin D2, bcl-x(L) and A1 leading to dramatic impairment of B cell mitogenic responses. In contrast, no effect of reduced Btk and BLNK dosage was observed on extracellular signal-regulated kinase activation. These results suggest that the signals regulating the maintenance and activation of mature B cells are differentially sensitive to the strength of the signal emanating from the signalosome.


Asunto(s)
Linfocitos B/fisiología , Fosfoproteínas/deficiencia , Proteínas Tirosina Quinasas/deficiencia , Transducción de Señal/fisiología , Proteínas Adaptadoras Transductoras de Señales , Agammaglobulinemia Tirosina Quinasa , Animales , Proteínas Portadoras , Ratones
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