간암의 DNA 복제수 의존성 유전 발현 변이 발굴
Identification of DNA Copy Number-Dependent Transcriptional Deregulation in Hepatocellular Carcinoma (HCC)
- 주제(키워드) HCC (Hepatocellular carcinoma) , DNA copy number , integrative analysis , IER3 , biomarker , HCC subtypes , 6p21-24 , transcriptional deregulation
- 발행기관 아주대학교
- 지도교수 백은주, 우현구
- 발행년도 2013
- 학위수여년월 2014. 2
- 학위명 박사
- 학과 및 전공 일반대학원 의학계열
- 실제URI http://www.dcollection.net/handler/ajou/000000016188
- 본문언어 영어
- 저작권 아주대학교 논문은 저작권에 의해 보호받습니다.
초록/요약
In recent years, cancer heterogeneity, which is essentially inherent in various types of cancer, has been of interest to the cancer genome research. Many studies using various approaches have tried to solve the conundrum of so-called cancer heterogeneity. However, even though many successes have been earned in this area using the genomic analysis, the identification of precise cancer subtypes, which can be informative and useful from the biological and clinical point of view, still remains a challenge. Among the many trials, the multi-layered genomic profiles analysis, in which the genomic copy numbers and gene expression profiles are analyzed by the integrative way to define the chromosomal regions with both genomic copy number variation and concomitant transcriptional deregulation, is posited to provide a promising strategy to identify driver targets. Here, the integrative analysis of the DNA copy numbers and gene expression profiles of hepatocellular carcinoma (HCC) was performed. By comparing DNA copy numbers in HCC subtypes, which have been previously defined based on gene expression pattern, it was found that the HCC subtype showing aggressive phenotype without expressing stemness-related genes had DNA copy number alteration with concordant gene expression changes in the specific chromosomal area at 6p21-24. Among the genes residing at 6p21-24, IER3 was identified as a potential driver. The clinical utility of IER3 copy numbers was demonstrated by validating its clinical correlation in the independent cohorts. In addition, short hairpin RNA-mediated knock-down experiment revealed the functional relevance of IER3 in liver cancer progression. In conclusion, the results of this study suggest that genomic copy number alterations with transcriptional deregulation at 6p21-24 identify an aggressive HCC phenotype and a novel functional biomarker.
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TABLE OF CONTENTS
ABSTRACT i
TABLE OF CONTENTS iii
LIST OF FIGURES v
LIST OF TABLES vi
ABBREVIATIONS vii
I. INTORDUCTION 1
A. Background of study 1
B. Cancer genomic study of HCC 3
C. Aims of study 7
II. MATERIALS AND METHODS 10
A. MATERIALS 10
1. Cell-lines 10
2. Human tissues and FFPE samples 10
3. Preparation of RNA and genomic DNA 10
B. METHODS 11
1. Comparative genomic hybridization (CGH) profiling 11
2. Data pre-processing 12
3. DNA copy number profiling based on the T-statistic map (TM) 13
4. cDNA Microarray profiling 14
5. Determination of DNA copy number-dependent transcriptional deregulation 14
6. Validation of the prognostic relevance in the independent data set 15
7. Estimation of genomic DNA copy number by quantitative PCR (qPCR) 16
8. Gene set enrichment analysis 17
9. Short hairpin RNA (shRNA)-mediated knock-down experiment 17
10. Estimation of mRNA expression level using quantitative PCR (qPCR) 20
11. Cell viability and proliferation assay 21
12. Cell invasion assay 22
13. Western Blot analysis 22
14. Statistical analysis 23
III. RESULTS 25
A. Identification of subtype-specific DNA copy number alteration 25
B. Region of Interest at 6p showed subtype-specific DNA copy number alteration and concomitant transcriptional deregulation 32
C. IER3 is a putative biomarker for the ROI at 6p amplicon 33
IV. DISCUSSION 70
V. CONCLUSION 78
VI. REFERENCES 79
국문 요약 91
