2011-08-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/649951摘要：雖然HBV 帶原者罹患肝細胞癌的相對危險性為非帶原者的20 倍，但取決於病毒和宿主因素不同，HBV 帶原者進展為肝細胞癌的危險性卻具有個體間很大的差異存在。病毒序列變異在病理變化的過程或影響病毒複製的活性上時常扮演重要的角色。由於HBV 利用reverse transcription 複製，缺乏proofreading 的機制，在慢性感染的過程會有許多基因變異產生，以致長期的演化形成各種基因型、基因亞型、和重組型。有關HBV 病毒序列變異及其和肝細胞癌發展的關係，已受到許多研究關注，但過去大部分研究侷限在特殊病毒基因區域或特殊核甘酸位置的變異，事實上病毒全基因的研究稀少，而且並未使用前瞻型研究設計、或未納入分析其他肝炎血清標記的影響。本研究將在三年期間利用前瞻型研究探討病毒全基因變異和肝細胞癌發展的關係，研究目的包括：1)利用重疊病例對照研究對約263 名由一個HBV 帶原者世代研究長期追蹤鑑定之肝細胞癌病例和263名配對的對照個案進行HBV 全基因定序；2)由上述定序資料鑑定基因亞型和重組型，並探討它們和肝細胞癌發展的關係；3) 由重疊病例對照研究對病毒3.2 kb全基因變異掃瞄的結果，利用資料掘礦和機器學習方法尋找最佳預測肝細胞癌發展的一組病毒基因變異，並評估其敏感度和特異度；4)發展TaqMan probe 以進行上述所搜尋獲得之病毒基因單點核酸變異的測量；5)利用所發展的TaqManprobe 對全世代進行研究，分析上述病毒基因單點核酸變異長期的穩定度及其對健康帶原者發展為慢性肝炎、肝硬化，以致最後肝細胞癌的作用；以及6)利用一個在原世代研究下進行的縱斷世代研究（包括1143 名研究個案具有所有追蹤16 年期間HBV DNA 重複測量結果和其它病毒標記資料，例如HBeAg），分析HBV 全基因變異和長期病毒複製狀態及自然史（不活動期、免疫廓清期及再活化期）的關係。本研究之全世代包括3630 名符合進入研究條件的男性HBV 帶原者，他們在進入研究和追蹤檢查時，均接受問卷調查（項目包括人口學特徵、生活飲食習慣和藥物治療史）和血液檢體採集，研究個案每年定期接受腹部超音波和肝生化指標檢查，追蹤期間長達20 年。本研究預期有助於肝細胞癌多階段致癌過程的瞭解，所鑑定和肝細胞癌或病毒長期複製狀態有關的病毒基因變異，將可實際應用在鑑定高危險群，以進行疾病狀態監測、介入或抗病毒藥物治療之參考。<br> Abstract: Although hepatitis B virus (HBV) carriers are approximately 20 times more likely to develophepatocellular carcinoma (HCC) than HBV noncarriers, the rates of progression to HCC is highlyvariable between carriers of HBV depending on both viral and host factors. Viral sequence variationfrequently plays an essential role in viral replication and pathogenesis. Because HBV utilizes reversetranscription to copy its genome, a multitude of genetic variants can occur in the HBV genome. Thelong-term evolution of this virus therefore leads to the occurrence of various genotypes, subgenotypes,and recombinants. Genomic variations of HBV and their association with HCC have been studied.However, studies on full-length sequencing of HBV are poorly documented, and most of previousstudies were not conducted with a prospective study design. Furthermore, other serological markers ofhepatitis B were not analyzed in these studies. Here we proposed a study to identify genetic variants inthe HBV genome for HCC development by studying the complete genomic sequence of HBV in alongitudinal cohort study. The specific aims are to: 1) Sequence the whole viral genome of HBV inabout 263 cases and 263 matched controls nested within a cohort study of male HBsAg carriers whohave been followed for 20 years; 2) Identify recombinants and novel subgenotypes of HBV andexamine their associations with HCC; 3) Employ data mining and rule learning to seek a predictive testfor HCC with combinations of single nucleotide polymorphisms (SNPs) over the 3.2-kb full length ofthe HBV genome; 4) Develop novel TaqMan probes for identification of HCC-associated viral SNPs; 5)Perform TaqMan assay to test the long-term stability of HCC-associated viral SNPs and theirassociations with hepatitis B progression from healthy carrier state to chronic hepatitis, liver cirrhosis,and HCC with the entire cohort; and finally 6) explore polymorphisms elsewhere in the HBV genomeand time trend for viral load and the natural course of hepatitis B (inactive carriers, immune clearance,and reactivation) in a longitudinal cohort study of 1143 HBsAg carriers who were selected from theentire cohort and had available data on multiple measurements of HBV DNA and other viral factors(including HBeAg) during long-term follow-up. The entire cohort consisted of 3630 eligible HBVcarrier subjects followed for up to 20 years. At recruitment and each follow-up examination, weconducted in-person interviews with a structured questionnaire to obtain information on demographiccharacteristics, lifetime habits, and a history of antiviral therapy. Participants were monitored forincident HCC by follow-up examinations including ultrasound scanning and conventional liverbiochemical tests every year. This project will improve our understanding on the HBV-relatedmultistage hepatocarcinogenesis. The viral variants identified for HCC development and time trend ofviral load may have practical implications for defining high-risk groups for intensive monitoring,intervention, or antiviral therapy.世代研究全長序列基因變異B 型肝炎病毒肝細胞癌cohort studyfull-length sequencegenetic variantshepatitis B virushepatocellular carcinoma