RESUMO
BACKGROUND: The histone variant H3.3 K27M mutation is a defining characteristic of diffuse intrinsic pontine glioma (DIPG)/diffuse midline glioma (DMG). This histone mutation is responsible for major alterations to histone H3 post-translational modification (PTMs) and subsequent aberrant gene expression. However, much less is known about the effect this mutation has on chromatin structure and function, including open versus closed chromatin regions as well as their transcriptomic consequences. RESULTS: Recently, we developed isogenic CRISPR-edited DIPG cell lines that are wild-type for histone H3.3 that can be compared to their matched K27M lines. Here we show via ATAC-seq analysis that H3.3K27M glioma cells have unique accessible chromatin at regions corresponding to neurogenesis, NOTCH, and neuronal development pathways and associated genes that are overexpressed in H3.3K27M compared to our isogenic wild-type cell line. As to mechanisms, accessible enhancers and super-enhancers corresponding to increased gene expression in H3.3K27M cells were also mapped to genes involved in neurogenesis and NOTCH signaling, suggesting that these pathways are key to DIPG tumor maintenance. Motif analysis implicates specific transcription factors as central to the neuro-oncogenic K27M signaling pathway, in particular, ASCL1 and NEUROD1. CONCLUSIONS: Altogether our findings indicate that H3.3K27M causes chromatin to take on a more accessible configuration at key regulatory regions for NOTCH and neurogenesis genes resulting in increased oncogenic gene expression, which is at least partially reversible upon editing K27M back to wild-type.
Assuntos
Neoplasias do Tronco Encefálico , Glioma , Humanos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Neoplasias do Tronco Encefálico/genética , Neoplasias do Tronco Encefálico/metabolismo , Neoplasias do Tronco Encefálico/patologia , Cromatina/genética , Glioma/genética , Glioma/metabolismo , Glioma/patologia , Histonas/metabolismo , MutaçãoRESUMO
Histone H3.3 mutations are a hallmark of pediatric gliomas, but their core oncogenic mechanisms are not well-defined. To identify major effectors, we used CRISPR-Cas9 to introduce H3.3K27M and G34R mutations into previously H3.3-wildtype brain cells, while in parallel reverting the mutations in glioma cells back to wildtype. ChIP-seq analysis broadly linked K27M to altered H3K27me3 activity including within super-enhancers, which exhibited perturbed transcriptional function. This was largely independent of H3.3 DNA binding. The K27M and G34R mutations induced several of the same pathways suggesting key shared oncogenic mechanisms including activation of neurogenesis and NOTCH pathway genes. H3.3 mutant gliomas are also particularly sensitive to NOTCH pathway gene knockdown and drug inhibition, reducing their viability in culture. Reciprocal editing of cells generally produced reciprocal effects on tumorgenicity in xenograft assays. Overall, our findings define common and distinct K27M and G34R oncogenic mechanisms, including potentially targetable pathways.
Assuntos
Biomarcadores Tumorais/metabolismo , Edição de Genes , Regulação Neoplásica da Expressão Gênica , Glioma/patologia , Histonas/genética , Mutação , Receptores Notch/metabolismo , Animais , Apoptose , Biomarcadores Tumorais/genética , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Proliferação de Células , Criança , Feminino , Glioma/genética , Glioma/metabolismo , Glicina/química , Glicina/genética , Histonas/química , Humanos , Lisina/química , Lisina/genética , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Receptores Notch/genética , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: Bariatric operation is the most effective treatment for diabetes mellitus in the morbidly obese. The purpose of this study is to compare the rate of resolution of diabetes mellitus after three common laparoscopic bariatric procedures: laparoscopic adjustable gastric banding (LAGB), Roux-en-Y gastric bypass (RYGB), and biliopancreatic diversion with or without duodenal switch (BPD/DS). STUDY DESIGN: All data were prospectively collected and entered into an electronic registry. Characteristics evaluated for this study included preoperative age, body mass index, duration of diabetes, race, gender, operative time, length of stay, percent excess weight loss, oral hypoglycemic requirements, and insulin requirements. RESULTS: A total of 282 bariatric patients with diabetes mellitus were analyzed (218 LAGB, 53 RYGB, and 11 BPD/DS). Preoperative age (46 to 50 years), body mass index (46 to 50; calculated as kg/m(2)), race and gender breakdown, and baseline oral hypoglycemic (82% to 87%) and insulin requirements (18% to 28%) were comparable among the three groups (p = NS). Percent excess weight loss at 1, 2, and 3 years was: 43%, 50%, and 45% for LAGB; 66%, 68%, and 66% for RYGB; and 68%, 77%, and 82% for BPD/DS (p < 0.01 LAGB versus RYGB and LAGB versus BPD/DS at all time intervals). At 1 and 2 years, the proportion of patients requiring oral hypoglycemics postoperatively was 39% and 34% for LAGB; 22% and 13% for RYGB; and 11% and 13% for BPD/DS (p = NS). At 1 and 2 years, the proportion of patients requiring insulin postoperatively was 14% and 18% for LAGB; 7% and 13% for RYGB; and 11% and 13% for BPD/DS (p = NS). CONCLUSIONS: Despite the disparity in percent excess weight loss between LAGB, RYGB, and BPD/DS, the rate of resolution of diabetes mellitus is equivalent.