RESUMEN
Chronic lymphocytic leukemia (CLL) is the most common adult leukemia and is characterized by chromosomal aberrations including 13q, 11q, and 17p deletions and a trisomy of chromosome 12 (T12). 13q deletions are often associated with 11q and 17p deletions in aggressive cases. Conversely, T12 CLLs show a variable prognosis, and association with 13q deletions is uncommon. The miR-15a/16-1 cluster is the functional target of 13q deletions, leading to BCL2 overexpression. Chromosomal aberrations in CLL are associated with prognosis, and their identification is carried out by fluorescence in situ hybridization (FISH). Since standard FISH only detects large deletions, we investigated the presence of undetected microdeletions targeting miR-15a/16-1 in CLL cases. We found that â¼34% of CLL samples show an unreported loss of the miR-15a/16-1 locus regardless of their cytogenetic profile. Interestingly, 15 out of 39 (â¼39%) of all CLLs with T12, carry microdeletions of miR-15a/16-1, indicating that, in patients with T12, miR-15a/16-1 are mostly inactivated by microdeletions. In addition, â¼40% of CLL cases bearing T12, 17p-, and 11q- showed unidentified microdeletions of miR-15a/16-1, suggesting that miR-15a/16-1 loss cooperates with such chromosomal alterations in CLL. These data may have clinical relevance for the successful stratification of patients for treatment.
Asunto(s)
Deleción Cromosómica , Cromosomas Humanos Par 11 , Cromosomas Humanos Par 12 , Cromosomas Humanos Par 17 , Leucemia Linfocítica Crónica de Células B/genética , MicroARNs/genética , Trisomía , Variaciones en el Número de Copia de ADN , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Humanos , Hibridación Fluorescente in Situ , Leucemia Linfocítica Crónica de Células B/diagnósticoRESUMEN
The existence of "leukemia-initiating cells" (LICs) in chronic lymphocytic leukemia (CLL) remains controversial due to the difficulty in isolating and identifying the tumor-initiating cells. Here, we demonstrate a microchannel electroporation (MEP) microarray that injects RNA-detecting probes into single live cells, allowing the imaging and characterization of heterogeneous LICs by intracellular RNA expression. Using limited-cell FACS sequencing (LC-FACSeq), we can detect and monitor rare live LICs during leukemogenesis and characterize their differential drug sensitivity. Disease-associated mutation accumulation in developing B lymphoid but not myeloid lineage in CLL patient hematopoietic stem cells (CLL-HSCs), and development of independent clonal CLL-like cells in murine patient-derived xenograft models, suggests the existence of CLL LICs. Furthermore, we identify differential protein ubiquitination and unfolding response signatures in GATA2high CLL-HSCs that exhibit increased sensitivity to lenalidomide and resistance to fludarabine compared to GATA2lowCLL-HSCs. These results highlight the existence of therapeutically targetable disease precursors in CLL.
Asunto(s)
Leucemia Linfocítica Crónica de Células B , Animales , Células Cultivadas , Células Madre Hematopoyéticas/metabolismo , Humanos , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/metabolismo , Ratones , Células Madre Neoplásicas/metabolismo , ARN/metabolismoRESUMEN
Hematopoiesis is hierarchical, and it has been postulated that acute myeloid leukemia (AML) is organized similarly with leukemia stem cells (LSCs) residing at the apex. Limited cells acquired by fluorescence activated cell sorting in tandem with targeted amplicon-based sequencing (LC-FACSeq) enables identification of mutations in small subpopulations of cells, such as LSCs. Leveraging this, we studied clonal compositions of immunophenotypically-defined compartments in AML through genomic and functional analyses at diagnosis, remission and relapse in 88 AML patients. Mutations involving DNA methylation pathways, transcription factors and spliceosomal machinery did not differ across compartments, while signaling pathway mutations were less frequent in putative LSCs. We also provide insights into TP53-mutated AML by demonstrating stepwise acquisition of mutations beginning from the preleukemic hematopoietic stem cell stage. In 10 analyzed cases, acquisition of additional mutations and del(17p) led to genetic and functional heterogeneity within the LSC pool with subclones harboring varying degrees of clonogenic potential. Finally, we use LC-FACSeq to track clonal evolution in serial samples, which can also be a powerful tool to direct targeted therapy against measurable residual disease. Therefore, studying clinically significant small subpopulations of cells can improve our understanding of AML biology and offers advantages over bulk sequencing to monitor the evolution of disease.
Asunto(s)
Biomarcadores de Tumor/genética , Evolución Clonal , Genómica/métodos , Células Madre Hematopoyéticas/patología , Leucemia Mieloide Aguda/patología , Mutación , Células Madre Neoplásicas/patología , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Citometría de Flujo , Estudios de Seguimiento , Células Madre Hematopoyéticas/metabolismo , Humanos , Inmunofenotipificación , Leucemia Mieloide Aguda/genética , Masculino , Persona de Mediana Edad , Células Madre Neoplásicas/metabolismo , Pronóstico , Adulto JovenRESUMEN
Rare, recurrent balanced translocations occur in a variety of cancers but are often not functionally interrogated. Balanced translocations with the immunoglobulin heavy chain locus (IGH; 14q32) in chronic lymphocytic leukemia (CLL) are infrequent but have led to the discovery of pathogenic genes including CCND1, BCL2, and BCL3. Following identification of a t(X;14)(q28;q32) translocation that placed the mature T cell proliferation 1 gene (MTCP1) adjacent to the immunoglobulin locus in a CLL patient, we hypothesized that this gene may have previously unrecognized importance. Indeed, here we report overexpression of human MTCP1 restricted to the B cell compartment in mice produces a clonal CD5+/CD19+ leukemia recapitulating the major characteristics of human CLL and demonstrates favorable response to therapeutic intervention with ibrutinib. We reinforce the importance of genetic interrogation of rare, recurrent balanced translocations to identify cancer driving genes via the story of MTCP1 as a contributor to CLL pathogenesis.
Asunto(s)
Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/metabolismo , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas/metabolismo , Translocación Genética , Adulto , Anciano , Anciano de 80 o más Años , Animales , Proteínas del Linfoma 3 de Células B , Ciclina D1 , Femenino , Regulación de la Expresión Génica , Genes de las Cadenas Pesadas de las Inmunoglobulinas , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Oncogenes/genética , Proteínas Proto-Oncogénicas c-bcl-2RESUMEN
BACKGROUND: The BRAFV600E mutation is the most common driver in papillary thyroid carcinoma (PTC) tumors. In recent years, gene fusions have also been recognized as important drivers of cancer in PTC. Previous studies have suggested that thyroid tumors with fusion genes frequently display an aggressive course. These observations prompted further exploration of gene fusions in PTC tumors. The aim was to search for previously unrecognized gene fusions using thyroid tissue samples from PTC patients. METHODS: Gene fusions were analyzed in RNA sequencing data obtained from 12 PTC tumors and paired unaffected thyroid tissue samples. Candidate fusions were further filtered and validated using reverse transcriptase polymerase chain reaction, Sanger sequencing, and fluorescence in situ hybridization. An Ohio cohort of 148 PTC tumor samples was screened for a LMO7-BRAF fusion and the BRAFV600E mutation. Functional assays were performed to assess the LMO7-BRAF fusion. RESULTS: Two coding fusions (CCDC6-RET and LMO7-BRAF) were found in one tumor sample each. The novel LMO7-BRAF fusion was validated by reverse transcriptase polymerase chain reaction and fluorescence in situ hybridization. The LMO7-BRAF fusion was a recurrent somatic alteration with a frequency of 2.0% (3/148) in PTC tumors, while the BRAFV600E point mutation was found in 63.5% (94/148) of tumors. Enforced expression of LMO7-BRAF fusion protein stimulated endogenous ERK1/2 phosphorylation and promoted anchorage independent cell growth to an extent similar to BRAFV600E. CONCLUSIONS: A novel fusion gene, LMO7-BRAF, was identified in PTC tumors. The results indicate that the LMO7-BRAF fusion behaves as an oncogenic alteration. This observation expands the spectrum of fusion genes involving kinases in thyroid cancer.
Asunto(s)
Proteínas con Dominio LIM/genética , Proteínas de Fusión Oncogénica/genética , Proteínas Proto-Oncogénicas B-raf/genética , Cáncer Papilar Tiroideo/genética , Cáncer Papilar Tiroideo/patología , Factores de Transcripción/genética , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Animales , Carcinoma Papilar/genética , Carcinoma Papilar/patología , Femenino , Humanos , Masculino , Ratones , Persona de Mediana Edad , Mutación , Células 3T3 NIH , Recurrencia Local de Neoplasia , Fosforilación , Mutación Puntual , ARN/análisis , Análisis de Secuencia de ARN , Neoplasias de la Tiroides/genética , Neoplasias de la Tiroides/patología , Adulto JovenRESUMEN
Ibrutinib is a highly effective targeted therapy for chronic lymphocytic leukemia (CLL). However, ibrutinib must be discontinued in a subset of patients due to progressive CLL or transformation to aggressive lymphoma (Richter transformation). Transformation occurs early in the course of therapy and has an extremely poor prognosis. Thus, identification of prognostic markers associated with transformation is of utmost importance. Near-tetraploidy (4 copies of most chromosomes within a cell) has been reported in various lymphomas, but its incidence and significance in CLL has not been described. Using fluorescence in situ hybridization, we detected near-tetraploidy in 9 of 297 patients with CLL prior to beginning ibrutinib treatment on 1 of 4 clinical trials (3.0%; 95% confidence interval [CI], 1.4%-5.7%). Near-tetraploidy was associated with aggressive disease characteristics: Rai stage 3/4 (P = .03), deletion 17p (P = .03), and complex karyotype (P = .01). Near-tetraploidy was also associated with ibrutinib discontinuation due to Richter transformation (P < .0001), but not due to progressive CLL (P = .41). Of the 9 patients with near-tetraploidy, 6 had Richter transformation with diffuse large B-cell lymphoma. In a multivariable model, near-tetraploidy (hazard ratio [HR], 8.66; 95% CI, 3.83-19.59; P < .0001) and complex karyotype (HR, 4.77; 95% CI, 1.42-15.94; P = .01) were independent risk factors for discontinuing ibrutinib due to transformation. Our results suggest that near-tetraploidy is a potential prognostic marker for Richter transformation to assess in patients going on ibrutinib.
Asunto(s)
Adenina/análogos & derivados , Aberraciones Cromosómicas , Cromosomas Humanos Par 2/genética , Dosificación de Gen , Leucemia Linfocítica Crónica de Células B/patología , Piperidinas/uso terapéutico , Adenina/uso terapéutico , Adulto , Anciano , Anciano de 80 o más Años , Estudios de Cohortes , Femenino , Estudios de Seguimiento , Humanos , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Leucemia Linfocítica Crónica de Células B/genética , Masculino , Persona de Mediana Edad , Pronóstico , Tasa de SupervivenciaRESUMEN
A causal role of gene amplification in tumorigenesis is well known, whereas amplification of DNA regulatory elements as an oncogenic driver remains unclear. In this study, we integrated next-generation sequencing approaches to map distant estrogen response elements (DEREs) that remotely control the transcription of target genes through chromatin proximity. Two densely mapped DERE regions located on chromosomes 17q23 and 20q13 were frequently amplified in estrogen receptor-α-positive luminal breast cancer. These aberrantly amplified DEREs deregulated target gene expression potentially linked to cancer development and tamoxifen resistance. Progressive accumulation of DERE copies was observed in normal breast progenitor cells chronically exposed to estrogenic chemicals. These findings may extend to other DNA regulatory elements, the amplification of which can profoundly alter target transcriptome during tumorigenesis.
Asunto(s)
Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Estrógenos/metabolismo , Regulación Neoplásica de la Expresión Génica , Elementos de Respuesta , Tamoxifeno/farmacología , Antineoplásicos Hormonales/farmacología , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Resistencia a Antineoplásicos , Receptor alfa de Estrógeno/genética , Receptor alfa de Estrógeno/metabolismo , Femenino , Amplificación de Genes , Genómica , HumanosRESUMEN
CONTEXT: The family risk ratio for papillary thyroid carcinoma (PTC) is among the highest of all cancers. Collectively, familial cases (fPTC) and sporadic cases (sPTC) are not known to show molecular differences. However, one study reported that telomeres were markedly shorter and the telomerase reverse transcriptase (TERT) gene was amplified and up-regulated in germline DNA from patients with fPTC compared with sPTC. OBJECTIVE: The aim of this study was to evaluate telomere length and TERT gene amplification and expression in blood samples of fPTC and sPTC patients in a genetically distinct population from the previous study. DESIGN: In 42 fPTC and 65 sPTC patients, quantitative real-time PCR was employed to measure the relative telomere length (RTL) and TERT gene copy number and RNA level. To validate the results using alternative methods, we further studied a subset of the original cohort consisting of randomly chosen fPTC (n = 10) and sPTC (n = 14) patients and controls (n = 21) by assessing both telomere length by flow fluorescent in situ hybridization and TERT gene expression by quantitative real-time PCR. RESULTS: RTL and TERT gene copy number did not differ between fPTC and sPTC (P = 0.957 and P = 0.998, respectively). The mean RTL and TERT gene expression were not significantly different among the groups of the validation series (P = 0.169 and P = 0.718, respectively). CONCLUSION: Our data show no difference between familial and sporadic PTC with respect to telomere length, TERT copy number, or expression in our cohort. Further investigations in additional cohorts of patients are desirable.
Asunto(s)
Carcinoma Papilar/genética , Telomerasa/genética , Telómero/patología , Neoplasias de la Tiroides/genética , Adulto , Carcinoma Papilar/patología , Femenino , Dosificación de Gen , Humanos , Masculino , Persona de Mediana Edad , ARN Mensajero/genética , Telómero/genética , Neoplasias de la Tiroides/patologíaRESUMEN
Tumor evasion of T-cell immunity remains a significant obstacle to adoptive T-cell therapy. It is unknown whether the mode of immune evasion is dictated by the cancer cells or by the tumor antigens. Taking advantage of the fact that multiple lineages of tumor cells share the tumor antigen P1A, we adoptively transferred transgenic T cells specific for P1A (P1CTL) into mice with established P1A-expressing tumors, including mastocytoma P815, plasmocytoma J558, and fibrosarcoma Meth A. Although P1CTL conferred partial protection, tumors recurred in almost all mice. Analysis of the status of the tumor antigen revealed that all J558 tumors underwent antigenic drift whereas all P815 tumors experienced antigenic loss. Interestingly, although Meth A cells are capable of both antigenic loss and antigenic drift, the majority of recurrent Meth A tumors retained P1A antigen. The ability of Meth A to induce apoptosis of P1CTL in vivo alleviated the need for antigenic drift and antigenic loss. Our data showed that, in spite of their shared tumor antigen, different lineages of cancer cells use different mechanisms to evade T-cell therapy.