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Molecular profiling of circulating tumor cells predicts clinical outcome in head and neck squamous cell carcinoma

多田, 紘恵 タダ, ヒロエ Tada, Hiroe 群馬大学

2020.09.30

概要

末梢血循環癌細胞(circulating tumor cells: CTCs)は、腫瘍組織から末梢血中に漏出した癌細胞で、従来の観血的Biopsyより非侵襲的に施行できると期待されているリキッドバイオプシーのバイオマーカー候補として注目されている。今回、頭頸部扁平上皮癌と診断された44名の患者から治療前に末梢血7.5mlを採血し、Cell Sieve社のmicrofilterで低圧濾過し、CTCsを分離、RNAを抽出したのちRT-qPCR法を用いて、上皮系マーカー(EPCAM, MET, KRT19, EGFR)の発現によるCTCsの同定と、予後との関連について検討を行った。さらにCTCsが陽性であった場合、同様にRT-qPCR法を用いcell growth関連遺伝子(PIK3CA, CCND1)、EMT関連遺伝子(SNAI1, VIM )、cancer stemness関連遺伝子(CD44, NANOG, ALDH1A1 )、immune regulation関連遺伝子(CD47, CD274 ,PDCD1LG2)の発現を解析し、臨床因子との関連について検討を行った。

44名中28名 (63.6%) で少なくとも1つ以上の上皮系マーカーの発現が認められ、頭頸部癌においてもCTCs が同定できた。各上皮系マーカーについては、44名中EPCAM 陽性患者が6名 (13.6 %) 、MET 陽性患者が12名 (27.3%) 、KRT19 陽性患者が21名 (47.7%) 、EGFR 陽性患者が7名 (15.9%) であった。4つの上皮系マーカーすべてが陽性であったのは44名中3名 (6.8%) であった。

CTCsが陽性であった患者は初期治療に抵抗性であり(p=0.0363)、局所再発しやすく(p=0.0151)、遠隔転移も多い傾向があった(p=0.0891)。無病増悪期間(PFS)、全生存期間(OS)に関してはCTCs陽性患者において有意に無病増悪期間が短かった(p=0.0107)。特にMET 陽性CTCsを有する患者は、有意に短い無増悪生存期間であったことから(p=0.0426)、頭頸部癌においてはMET 陽性CTCsが予後不良因子になることが示唆された。

CTCsにおける各癌関連遺伝子の発現には各患者間でのheterogeneityを認めた。各癌関連遺伝子の発現と予後 について検討すると、CD274 陽性CTCsを有する患者は、CD274 陰性CTCsを有する患者より、有意にPFSとOSが長 かった(PFS, p=0.0346 ; OS, p=0.0378)。頭頸部扁平上皮癌患者のCTCsにおけるCD274 (PD-L1) 発現は予後 良好因子であり、CTCsが原発巣のtumor immune microenvironmentを反映している可能性が示唆された。つまり、 CTCs上のPD-L1の発現は、腫瘍組織における免疫細胞の浸潤とPD-L1の過剰発現を部分的に反映している可能性が 考えられた。またCD274 ,PDCD1LG2 を除く8遺伝子発現を基にk-meansクラスタリング法を用いて高発現群、 低発現群の2群に分けて臨床因子との関係を解析した。PFSやOSには有意差を認めなかったが(PFS, p=0.3327 ;OS, p=0.9802)、進行病期(stage III-IV)では遺伝子高発現群が有意に多かった(p=0.0228)。

以上のことから、頭頸部扁平上皮癌においてCTCsの存在は予後不良因子であり、CTCsの遺伝子発現プロファイルは、いくつかの臨床因子と関連することが示唆された。CTCsの同定とともに、CTCsの遺伝子発現プロファイリングは、頭頸部扁平上皮癌においても、癌の個別化治療、治療効果判定、予後予測に活用できる可能性があり、 precision medicineの実現を目指す1つの手段となり得ると考えられた。

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