口腔扁平上皮癌に対する癌ワクチン療法の基礎的研究

口腔鳞状细胞癌疫苗治疗的基础研究

• 批准号: 15791178
• 负责人: 宮崎 晃亘
• 金额: $ 1.79万
• 依托单位: Sapporo Medical University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Young Scientists (B)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2005
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-15791178/
• 关键词: 口腔癌; 癌ワクチン療法; CTL

60歳女性の下顎歯肉癌患者および58歳男性の口峡咽頭癌患者の手術材料から癌細胞株POT1、POT2を新たに樹立した。POT1は高分化型扁平上癌細胞株(浸潤様式4D)、POT2は中分化型扁平上皮癌細胞株(浸潤様式4C)で、それぞれSCIDマウスの背部に皮下注射すると、同様の組織所見を示すことが確認された。それぞれの患者末梢血からリンパ球を分離し、AIM-V培養液にて1晩培養後、付着系細胞と非付着系細胞に分離した。付着系細胞はGM-CSF1000U/ml, IL-4 1000U/ml 加AIM-V培養液で1週間培養し、非付着系は10%HS加AIM-V培養液で1週間培養した。その後、非付着系はCD8ビーズによりCD8+ T cellを分離し、MLPCを行った。MLTC当日と翌日は10%HS加AIM-V培養液で培養し、3日目以降はIL-2 50U/ml加AIM-V培養液にて次のMLPC前日まで培養した。MLTC前日は10%HS加AIM-V培養液で培養した。MLTCを4〜5回繰り返し行い、^<51>Cr release assayにより自家癌特異的傷害活性をもつCTL誘導を確認した。現在さらにCTLクローンを樹立して、POT1,POT2細胞株のcDNAライブラリーをスクリーニングして口腔扁平上皮癌抗原遺伝子を同定する予定である。本研究の実施にあたっては倫理的な配慮を慎重にし、患者本人に研究目的・内容を十分に説明し、同意を得た。

The new cancer cell lines POT1 and POT2 were established for surgical materials for 60-year-old female patients with jaw cancer and 58-year-old male patients with throat cancer. POT1 and POT2 were identified by subcutaneous injection of the SCID cells into the back of a well-differentiated squamous cell carcinoma cell line (invasive pattern 4D) and a moderately differentiated squamous cell carcinoma cell line (invasive pattern 4C), respectively. The patient's peripheral blood was separated from the ball, AIM-V culture medium was cultured for 1 night, and the cells were separated from the non-cells. Sub-line cells were cultured in GM-CSF1000U/ml, IL-4 1000U/ml and AIM-V medium for 1 week, while non-subline cells were cultured in 10%HS and AIM-V medium for 1 week. The CD8+ T cell is separated from the MLPC. MLTC was cultured in 10% HS plus AIM-V medium on the first day, IL-2 50U/ml plus AIM-V medium on the third day, and MLPC was cultured on the second day. MLTC was cultured in 10% HS plus AIM-V medium the day before yesterday. MLTC is 4 ~ 5 cycles <51>away, Cr release assay is 4 ~ 5 cycles away, CTL induction is 5 ~ 6 cycles away, and CTL induction is 5 ~ 6 cycles away. Now the CTL gene is established, and the cDNA expression of POT1 and POT2 cell lines is determined. The implementation of this study was carried out with great care due to ethical considerations, the purpose and content of the study were fully explained by the patients themselves, and consent was obtained.