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DIPG里面的Super-enhancers

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发表于 2017-6-5 20:49:44 | 显示全部楼层 |阅读模式
背景:
Diffuse intrinsic pontine glioma (DIPG) is a universally fatal pediatric cancer.
A histone-3 K27M mutation affects ∼80% of DIPGs and drives aberrant transcription.

早在2015年的83个药物对14个DIPG细胞系的筛选实验中,就发现哪怕是效果最好的multi-HDAC inhibitor panobinostat也会被某些DIPG细胞系产生耐药性。(http://www.biotrainee.com/thread-1599-1-1.html

预示着需要联合用药,或者采取其它策略。
有据可考的有效例子是干扰 RNA polymerase II (RNAPII) transcription 转录活性
这时候需要引入两个基因的背景知识。
BRD4 a key activator of RNAPII transcription at active chromatin marks
CDK7, a member of the cyclin-dependent kinase family involved in regulation of RNAPII phosphorylation, controlling transcriptional initiation, pausing, and elongation
而且它们的抑制剂是:
THZ1 is a highly specific CDK7 inhibitor effective
BET (bromodomain and extra-terminal) protein inhibition, targeting family member BRD4
JQ1也被广泛使用来抑制 bromodomain activity


因为 transcriptional dysregulation in DIPG is chiefly driven by the H3K27M mutation  
提出假说:we hypothesized that DIPG may be vulnerable to transcriptional disruption.

实验材料是8个DIPG细胞系,七个是K27M,一个是WT:
Eight patient-derived DIPG cultures and one pediatric cortical glioblastoma culture (SU-pcGBM2) were used in this study;
seven of the eight DIPGs exhibit the H3K27M mutation and one is histone wild-type (WT)
(H3.3K27M: SU-DIPG-VI, SU-DIPG-XIII-P, SU-DIPG-XVII, SU-DIPG-XXV, SF7761, and JHH-DIPG1; H3.1K27M: SU-DIPG-IV; H3WT and MYCN amplified: VUMC-DIPG-10;).
SU-pcGBM2 is histone-3 WT and exhibits a TP53 mutation and EGFR amplification

结果,经过了JQ1处理后,WT的DIPG的受影响最小。

然后测试了两个目前临床批准的 bromodomain inhibitor drugs ,分别是 iBET762 和  OTX015
然后用targeting BRD4 的shRNA处理,看看变化,结果knockdown后,DIPG严重被破坏了。
同样用THZ1来抑制DIPG细胞系体内的CDK7,Disruption of RNAPII transcription through CDK7 inhibition thus appears to potently disrupt DIPG cell viability.
所以可以用HDAC inhibition together with JQ1 or THZ1  联合治疗。
这样,即使有些DIPG细胞系对HDAC inhibition 耐受了,这些细胞系仍然可以被bromodomain or CDK7 inhibition来杀死。
而且作者对panobinostat, JQ1, and THZ1 这3种处理都做了RNA-seq。(14个数据,第6/8细胞系的3种药物单独处理+联合处理,还有对照)
然后有做了它们3个的ChIP-seq来看看它们的target genes的overlap情况咋样。(8个数据,第6/8细胞系的3个药物处理的H3K27ac,共有一个INPUT)
数据都在:https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE94259

首先是药物处理的表达数据差异分析结果是:
THZ1-treated cells showed preferential disruption of genes related to transcription and gene regulation,
such as ETS1, ELF4, MGA, SOX10, and HES5.
In contrast, both JQ1 and panobinostat disrupted key regulators of nervous system development,
including NTRK3, LINGO1, ASCL1, SYT4, SYT17, MYT1, MYRF, and SALL3.
In addition, both drugs disrupted genes that enriched for synapse organization and structure,
with one panobinostat target being NLGN3, a key mechanism mediating neuronal activity-regulated glioma growth

最后对3个药物处理的H3K27ac数据一直在分析super enhancer,我表示完全蒙圈了,搞不懂写的些什么鬼画符。






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