免疫螢光顯微術用於癌症免疫療法之生物指標研究

Abstract

頭頸癌為世界第七大癌症，每年造成全球四十餘萬人死亡。傳統療法對於末期的病患的成效有限，五年存活率僅約40%，而近期免疫療法的進展為癌症治療帶來一線曙光。免疫檢查點抑制劑 (immune checkpoint inhibitor, ICI) 為常用於頭頸癌的免疫療法藥物，透過阻斷免疫細胞表面的免疫檢查點與癌細胞結合，達到刺激免疫系統清除病患體內腫瘤的效果。然而，並非所有病患對免疫療法皆有反應。在預測病患的投藥後反應方面，當前以PD-L1做為生物指標的檢測方式僅有20~30% 的準確率。因此，尋找能有效預測病患反應的生物指標是當前免疫療法研究的重要課題。 在本研究中，我們試圖從腫瘤微環境(tumor microenvironment, TME)中免疫細胞的表現量尋找潛在的生物指標。從接受免疫療法的頭頸癌病患中，我們取各五位反應者與無反應者在投藥前的腫瘤石蠟切片作為樣品，以多重免疫螢光染色對組織內的6種免疫細胞、9種潛在生物指標進行標記，進而比較兩組病患細胞表現量的差異。我們搭建了四頻道的螢光顯微系統，對樣品進行全切片掃描(Whole Slide Imaging)，並以影像計算細胞數量，分析各種細胞在腫瘤內部的占比。 根據10組樣品的結果顯示，腫瘤內CD8+ T 細胞的比例在兩組病患間有顯著差異，與過往研究的結果相同，而腫瘤相關巨噬細胞 (TAM) 則無如過往研究呈現相關性。 本研究使用的系統尚須許多改進以滿足當前臨床研究的需求，未來將試圖增加頻道數、減少曝光時間以加快篩檢速度。

Head and neck squamous cell carcinoma (HNSCC) is the seventh most common cancer in the world. Due to poor prognosis, it has become the serious health issue in Taiwan. Recently, the progress in the immunotherapy shows effectiveness to the treatment of HNSCC. Patients receiving immune check point inhibitor (ICI) can be treated. However, only around 20 to 30% of the patients respond to the treatment. Therefore, searching for biomarkers that better predict the efficacy of immunotherapy has become an important issue is current research. In this research, we established a four-channel multiplex immunofluorescence microscope to search potential biomarker in the tumor microenvironment (TME). Formalin-fixed, paraffin-embedded samples from 5 responders and 5 non-responders of HNSCC patients were studied. 9 markers were labeled in 3 cycles, and whole slide imaging was performed on 10 samples. The number of six types of immune cells within tumor was calculated through image cytometry. The intratumoral CD8+ T cell ratio shows a significant difference between two groups, which in accordance the result in previous research.