臺灣博碩士論文加值系統

臺灣地區十大死亡原因中，癌症的死亡率已躍居總死亡人數的第一位。癌症的產生是由於許多致癌基因 (oncogenes) 的活化及抑癌基因 (tumor suppressor genes) 的控制受到擾亂所致，若這種改變發生在生殖細胞 (germline) 上，則會遺傳到下一代。遺傳性癌症的特徵是：發病年齡較早且易再復發，家族成員中罹患此癌症的機率較高，且遺傳到基因變異的可能性大。本實驗研究針對臺灣地區家族性腦瘤 NF2 及乳癌 BRCA2 基因之分子特性的分析研究，了解其基因在腫瘤發展過程中可能扮演的角色，以及基因的變異對基因產物功能和遺傳疾病的影響，希望能釐清遺傳疾病與基因之間的關係。
腦瘤 (brain tumor) 雖不是國人常見的癌症，但對病人卻有極大的傷害性。其中較嚴重的為 Neurofibromatosis type 2 (NF2)，屬中樞神經系統方面的腫瘤，其腫瘤在組織學上屬良性瘤(benign)，但解剖學的位置及腫瘤的多樣性常具危險性，病患存活率低且易再復發為更具侵害性的腫瘤。乳癌是近年來婦女常見的疾病之一，根據衛生署公佈之民國八十五年臺灣地區十大癌症死因，乳癌佔全國癌症死亡率第五位，在女性癌症死亡率中佔第四位。而影響遺傳性腦瘤的 NF2 基因及乳癌的 BRCA2 基因，分別位於chromosome 22q12 及13q12-13，兩者皆為抑癌基因，為體染色體顯性遺傳 (autosomal dominant inheritance) 疾病，其較高的外顯率(highly penetrance)，與遺傳性疾病的發生有很大的關聯性。
在家族性腦瘤或乳癌病人身上已知，生殖細胞的其中一個突變的對偶基因 (alleles) 來自遺傳，而腫瘤的形成是由於局部組織細胞之另一個對偶基因發生體細胞的突變 (somatic mutation) 所造成。生殖細胞上的突變乃存於血液及腫瘤中，而體細胞的突變只能在腫瘤組織細胞發現。因此分析生殖細胞的基因，可瞭解基因的遺傳性與分子特性，有助於癌症遺傳基因的研究。為了發展簡易且準確的基因診斷方法，本實驗藉由聚合酉每連鎖反應 (polymerase chain reaction) 及單股構形多型性分析 (single strand conformation polymorphism, SSCP analysis) 方法，找出遺傳性基因的變異及位置，配合基因選殖及序列分析，以確定突變的結果。
在本研究中，於家族性腦瘤中發現 NF2 基因在 C169T 及 720delG的突變，會導致嚴重的中樞神經系統疾病。BRCA2 基因中2670delC、3073delT 及 6696-7delTC 的突變，在家族性乳癌中也有很大的影響。造成 NF2 基因與 BRCA2 基因 nonsense 或frameshift 的突變，可能使基因產物的功能喪失，導致腫瘤的生成。基因突變分析 (mutation analysis) 的結果中，使用 Phamarcia Phast system 的SSCP 分析中可清楚觀察到 heterozygous 基因。利用基因選殖及 DNA序列分析，比較正常與突變對偶基因的序列，找出基因突變的性質，進而了解其突變的種類，並同時比較二個或更多的家族性遺傳的例子來探討遺傳基因變異的差異性及遺傳基因與表現型的關聯性(genotype-phenotype correlation)。以 DNA 分子診斷來篩選癌症基因的變異，是一種快速、簡易又準確的診斷方法，可用以評估其他家族成員致癌的風險，並有助於臨床醫師對病情的了解，估計疾病遺傳方式、復發的機會並幫助病人採取適當的預防措施。

Neurofibromatosis type 2 (NF2) is an autosomal dominantly inherited disease by bilateral vestibular schwannomas and other tumor of the brain, spinal cord, and central nervous system. NF2 disease is caused by germline mutations in the NF2 tumor suppressor gene on chromosome 22q12. Although the tumors of NF2 are histologically benign, their anatomical location and multiplicity lead to great morbidity and early mortality.
Breast cancer is the most common malignancy among women in developed countries. Recently, the incidence of breast cancer increases n Taiwan and the occurrence of breast cancer tends to be early onset. Familial breast cancer is characterized by early onset, an increased risk of bilateral breast cancer, an increasing risk and numbers of affected family members. The second breast cancer susceptibility gene, BRCA2 on chromosome 13q12-13, has recently been cloned. Germline mutations of BRCA2 are predicted to account for approximately 35% of families with early onset female breast cancer, and they are also associated with an increased risk of male breast cancer.
To identify the nature of genetic mutations in familial brain tumor and breast cancer in Taiwan and develop simple diagnosis procedures for detecting these genetic mutations, we have analyzed germline mutations in the NF2 gene and BRCA2 gene by PCR (polymerase chain reaction) -based SSCP (single strand conformation polymorphism) analysis followed by cloning and sequencing of related PCR products.
In this study, nonsense mutations or frameshift deletions in NF2 and BRCA2 genes had been found, and these mutations are all predicted to lead to the truncated proteins which are usually associated with severe phenotypes. Moreover, polymorphic changes of several missense and silent mutations were also observed among breast cancer patients and normal individuals. The search for genotype-phenotype correlation in NF2 and BRCA2 mutations is important not only to increase understanding of how mutated genes function, but also to improve presymptomatic detection of patients. DNA diagnosis of NF2 gene and BRCA2 gene can improve quality of genetic counseling and clinical management, and possibly reduce psychosocial difficulties in at-risk individuals.