埼玉医科大学雑誌 第34巻 第1号別頁 (2007年10月) T27-T36頁 ◇論文(図表を含む全文)は,PDFファイルとなっています.

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Thesis
SYBR GreenIを用いたWT1のReal Time PCR標準化曲線の立ち上げ
―SYBR Green法を用いた膵臓癌におけるWT1の発現解析等―

小田 慶太郎
埼玉医科大学 総合医療センター 外科

医学博士 甲第1029号 平成18年6月23日 (埼玉医科大学)


The establishment of standardized curve for Real Time PCR of WT1 using SYBR Green I
- To analyze WT1 expression in pancreatic cancer using SYBR Green I-

Keitaro Oda (Department of Surgery, Saitama Medical Center, Saitama Medical University, 1981 Tsujido-machi, Kamoda, Kawagoe-shi, Saitama 350-8550, Japan)

 Due to difficulties in early detection of pancreatic cancer as well as rapid progression of the disease, it is extremely difficult to completely resect the tumor surgically. The improvement of prognosis may be expected with chemotherapy and radiation therapy for the remaining cancer cells or post-op adjuvant therapy, but there still has not been any effective treatment.
 In recent years, tumor vaccine therapy using tumor antigen peptides are receiving attention. Especially, the WT1 antigen has been known to be expressed in many kinds of cancer, and is receiving attention as the possible universal vaccine for tumors. In Japan, promising results have been reported by H.Sugiyama et al for trial studies of treatment using tumor vaccine derived from WT1 based antigen peptides.
 In order for the vaccine treatment targeted against tumor antigens to be effective, one must confirm that the antigen is expressed by the targeted tumors. In general, when enough cancerous tissue from surgical excision or biopsy can be obtained, immunohistochemical staining is the standard method. On the other hand, in hematological disease and in tumors of difficult anatomic access, allowing only endoscopic or aspiration biopsies, one must rely solely on method utilizing molecular biology.
 The Real Time PCR has been known to be the method to quantitatively analyze the expression of mRNA in tissues and cells. Already, clinical laboratories have established the method to quantitatively analyze several of the protein expressions. Currently, the most common method adopted is Taqman. However, there are many problems with this method when using in research laboratories, since it is troublesome and its running cost is high. In this study, we established the Real Time PCR for WT1 expression, using the SYBR-Green method, which is easier and more cost effective than Taqman method. We then attempted to detect the expression of WT1 gene in clinical specimens of pancreatic cancer using this established method and showed the usefulness of our method.
Keywords: WT1, standardized curve of Real Time PCR, SYBR Green, Taqman, tumor antigen peptide, pancreatic cancer

 


(C) 2007 The Medical Society of Saitama Medical School