Gene therapy using lympho-epithelial Kazal-type-inhibitor (LEKTI) for head and neck cancer

使用淋巴上皮 Kazal 型抑制剂 (LEKTI) 治疗头颈癌的基因治疗

• 批准号: 16591991
• 负责人: MITSUDO Kenji
• 金额: $ 2.11万
• 依托单位: Nagoya University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2004
• 资助国家: 日本
• 起止时间: 2004 至 2005
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-16591991/
• 关键词: gene therapy; LEKTI; serine protease inhibitor; Squamous cell carcinoma; Head and neck; 頭頸部癌; セリンプロテアーゼインヒビター; アデノウイルスベクター; アデノ随伴ウイルスベクター

1.efficacy of adenovirus vector transduction into human squamous cell lines and cytotoxicity in vitroFull length LEKTI cDNA was subcloned into adenovirus vector (Ad-lacZ-LEKTI). Human oral squamous cell lines, SAS, HSC-2, 3, 4, were used. Twenty thousand cells were inoculated into each well of an 8-well glass slide. 24 hours later, Ad-lacZ-LEKTI vectors were added to each well at varying multiplicities of infection (MOI). A solution of X-gal was added to each well, and transduction efficacy was determined. Experiments for toxicity of the adenovirus vector were performed in 24-well plates. 24 hours later, Ad-lacZ-LEKTI vectors were added to each well at varying MOI. Numbers of living cells were counted 96 hours after Ad-lacZ-LEKTI vector infection, using trypan blue dye exclusion method. Experiments for toxicity of GCV were performed similar to the experiments with adenovirus vector. The transducted cells were stained blue by X-gal staining. By 24hours after transduction, nearly 100% of … More all cells were transducted at a MOI of 10. Transduction efficacies at an MOI 1 were almost 20%. Ad-lacZ-LEKTI showed toxicity in all cell lines at an MOI of more than 100, but there was no evident toxicity at an MOI of 10 or less. The cell toxicity to GCV was evaluated in all cell lines, and found to be 25 μg/ml or more.2.effects on cancer cell death in vitroFifty thousand cells were inoculated into each well of a 24-well plate. 24 hours later, Ad-lacZ-LEKTI vectors were added to each well at varying MOI. 10 μg of GCV was added to each well. Numbers of living cells were counted 72 hours after GCV administration, using trypan blue dye exclusion method. More than 60% cell death was induced at an MOI of 1, and almost 100% cancer cell death was obtained at an MOI of 10.3.morphological changes in oral squamous cell lines treated with Ad-lacZ-LEKTI and GCVThe cells were transduced at an MOI of 10 with Ad-lacZ-LEKTI by GCV. At 24 hours after administration of GCV, the cultured cells showed signed apoptosis, such as chromatin condensation, cell shrinkage, blebbing of cell membrane, and ballooning formation. Less

1.腺病毒载体转导人鳞状细胞的效果及体外细胞毒作用全长Lekti基因亚克隆入腺病毒载体(Ad-lacZ-lekti)。采用人口腔鳞状细胞系SAS、HSC-2、HSC-3、HSC-4。将两万个细胞接种到一个8孔玻璃片的每一孔中。24小时后，以不同的感染复数(MOI)将Ad-LacZ-lekti载体加入到每个孔中。每孔加入X-半乳糖溶液，测定转导效率。在24孔板上进行了腺病毒载体的毒性实验。24小时后，将Ad-Lacz-lekti载体以不同的MOI加入到每个孔中。用台盼蓝拒染法计数Ad-LacZ-lekti载体感染96h后活细胞数。GCV的毒性实验与腺病毒载体实验类似。转导细胞经X-Gal染色后呈蓝色。到转导后24小时，几乎100%的…更多的ALL细胞在MOI为10时被转导，MOI为1时的转导效率几乎为20%。在MOI大于100时，Ad-Lacz-lekti对所有细胞株均有毒性作用，而在MOI小于等于10时，则无明显毒性。对所有细胞株的细胞毒性进行评估，发现25μg/ml或更多。2.体外对癌细胞死亡的影响24孔板每孔接种5万个细胞。24小时后，将Ad-Lacz-lekti载体以不同的MOI加入到每个孔中。每孔加入更昔洛韦10μg。用台盼蓝拒染法计数GCV给药后72小时的活细胞数。在MOI为1时，可诱导60%以上的细胞死亡，在MOI为10.3时，癌细胞几乎100%死亡。给药后24小时，培养的细胞出现明显的细胞凋亡，如染色质凝集、细胞皱缩、细胞膜起泡、气球形成等。较少

项目成果

Consequences of C-terminal domains and N-terminal signal peptide deletions on LEKTI secretion, stability, and subcellular distribution

• DOI: 10.1016/j.abb.2004.12.012
• 发表时间: 2005-03-01
• 期刊: ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS
• 影响因子: 3.9
• 作者: Jayakumar, A;Kang, Y;Clayman, GL
• 通讯作者: Clayman, GL

New superselective intra-arterial infusion via superficial temporary artery for oral cancer -Methods of catherization using a catheter with a curvature and P-U catheter-

新型超选择性经临时浅动脉动脉内输注治疗口腔癌 -使用弯曲导管和 P-U 导管进行导管插入的方法 -

• 发表时间: 2005
• 期刊: Japanese J Head Neck 31(1)
• 作者: Tohnai;I.;Fuwa;N.;Mitsudo;K.;et al.
• 通讯作者: et al.

浅側頭動脈よりの超選択的動注化学療法と放射線療法の連日同時併用療法-Stage III,IV口腔癌に対する術前治療-

每日同时进行颞浅动脉超选择性动脉内化疗和放疗的联合治疗 - III、IV期口腔癌的术前治疗 -

• 发表时间: 2005
• 期刊: 頭頸部癌 31(3)
• 作者: 藤内 祝;光藤 健司;他
• 通讯作者: 他

Daily concurrent chemoradiotherapy with docetaxel (DOC) and cisplatin (CDDP) using superselective intra-arterial infusion via superficial temporal artery for stage III and IV oral cancer -Possibility of organ preservation in advanced oral cancer-

每日同步放化疗采用多西他赛 (DOC) 和顺铂 (CDDP)，通过颞浅动脉进行超选择性动脉内输注治疗 III 期和 IV 期口腔癌 -晚期口腔癌器官保存的可能性 -

• 期刊: Japanese J Head Neck 32(1)(in press)
• 作者: Mitsudo;K.;Tohnai;I.;et al.
• 通讯作者: et al.

Gene-environment interaction involved in oral carcinogenesis : molecular epidermiological study for metabolic and DNA repair gene polymorphisms

口腔癌发生中涉及的基因-环境相互作用：代谢和DNA修复基因多态性的分子流行病学研究

• 发表时间: 2006
• 期刊: J Oral Pathol Med 35
• 作者: Sugimura T;Kumimoto H;Tohnai I;Fukui T;Matsuo K;Tsurusako S;Mitsudo K;et al.
• 通讯作者: et al.