Establishment and Characterization of Genetically Modified Cancer Cells for Experimental Cancer Therapy

目的：為了解不同種類的細胞株在帶有皰疹病毒之胸喀睫激活酵素(HSVtk)或顆粒球巨噬細胞生長因子(GM-CSF)時，能否作為癌症治療之細胞疫苗。材料與方法：我們嘗試將HSVtk基因或老鼠之GM-CSF基因分別或共同轉染至人類子宮頸癌細胞株Hela。經過一系列篩選，因而建立三種細胞株，命名為H-tk, H-gm，及H-tk/gm。我們所測試之腫瘤模式為K-BALB sarcoma及其同源之BALB/C鼠。分別把等量(lx10^6)的K-BALB細胞與H-tk, H-gm，或H-tk/gm細胞混合，而後注射入BALB/C鼠之右腰；每一測試組之BALB/C鼠左腰亦注入1x10^6的K-BALB細胞；再給予ganciclovir (GCV)或PBS注射，每週測量腫瘤之直徑各二次。結果：我們選殖到2株含有HSVtk基因的HeLa細胞，分別命名為H-tk 3-2及H-tk 12-2。隨後建立之H-tk/gm細胞株亦分泌GM-CSF而能支持GM-CSF依賴性細胞株NSF60之生長。隨後之活體內腫瘤測試時顯示，當H-tk與K-BALB細胞混合在植入老鼠後，再給予GCV時，並不能延緩K-BALB 腫瘤之生長。反之，K-BALB細胞與含有GM-CSF之細胞株混合再植入老鼠時，能抑制大部份的老鼠右腰部腫瘤的生長。進一步的實驗顯示H-tk細胞在注射入老鼠3天後仍會表達H-SVtk，但與同源細胞HeLa之問無法測試到旁觀者效應(bystander effect)。然而，每一實驗組之老鼠其左腰部腫瘤之抑制效果不明。結論：H-tk細胞雖然在活體內仍會表達HSVtk基因，但不能引起異種癌細胞的旁觀者效應，可能與HeLa細胞本身對代謝合作具有抵抗性有關。反之，局部之GM-CSF分泌能壓抑異種癌細胞形成腫瘤，但卻不足引起強烈的腫瘤免疫反應以抑制遠端腫的生長。

關鍵字

顆粒球巨噬細胞生長因子；胸嘧啶激活酵素；基因轉染之癌細胞；腫瘤疫苗

並列摘要

Purpose: To study whether a xenogeneic cell line engineered to express the herpes simplex virus thymidine kinase (HSVtk) and/or granulocyte-macrophage colony-stimulating factor (GM-CSF) can be used as cellular vaccine to repress tumor growth. Materials and Methods: The HSVtk gene, murine GM-CSF gene, or both were stably transfected into the human cervix cancer cell line HeLa, thus creating three different cell lines designated as H-tk, H-gm, and H-tk/gm. The tumor model we tested was the KBALB sarcoma cells in syngenic BALB/c mice. For in vivo studies, equal number of KBALB cells and H-tk, H-gm, or H-tk/gm cells were premixed and injected into the right flanks of mice. The tumor growth was monitored after ganciclovir (GCV) administration. Results: Two HSVtk-expressing clones (H-tk 2-3 and H-tk 12-2) were isolated. The established H-tk/gm or H-gm cell line was confirmed to express GM-CSF by supporting the growth of a GM-CSF-dependent cell line NSF6O. The right flank tumors in mice injected with premixed K-BALB and H-tk cells showed no definite growth retardation upon GCV administration. Further studies implied that the H-tk cells expressed HSVtk in the injection sites, but failed to confer bystander effect between homologous cells by in vitro studies. The right flanks of mice injected with premixed K-BALB and H-tk/gm or Hgm showed no detectable or much delayed tumor growth either with or without GCV administration; however, the left flank K-BALB tumor progressed with similar growth curve in each study group. Conclusion: Cur studies suggest that though HeLa cells can express HSVtk gene in vivo, they can not confer bystander killing effect to xenogeneic tumor (sarcoma) cells upon GCV administration. This might be because the HeLa cells are intrinsically resistant to metabolic cooperation with other cells. Local production of GM-CSF is able to suppress local tumor growth, but not potent enough to induce distant antitumor immunity. This inability might be related to insufficient duration or amount of GM-CSF action.

並列關鍵字

GM-CSF ； Thymidine kinase ； Genetically modified cancer cells ； Tumor vaccine

國際替代計量

Establishment and Characterization of Genetically Modified Cancer Cells for Experimental Cancer Therapy

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