T細胞受體β鏈互補決定區3之頻譜暨高脂飲食研究於突變型表皮生長因子受體誘發之肺腺癌小鼠

腫瘤浸潤淋巴球已知存在於包含肺癌的多種癌症中。參與後天免疫的淋巴球成熟過程所產生的可變、變異、連接片段的重組，構成免疫全面多樣性。透過免疫全面多樣分析，可以用於監測個體的T細胞受體與B細胞受體在疾病中的群殖增生。T細胞受體與抗原的互補決定區(CDR)最具變異性的區域為CDR3，本篇研究假設其會在突變型表皮生長因子受體所誘發的肺癌中受到特殊腫瘤抗原刺激而進行群殖增生，其增生頻譜可望用於新穎抗原依賴的免疫療法。首先，利用雙基因轉殖鼠搭配可誘導轉位子系統，將帶有肺組織特異性的CC10基因，及具酪胺酸激酶抑制劑敏感性的突變型表皮生長因子受體EGFR L858R或具酪胺酸激酶抑制劑抗藥性的EGFR L858R+T790M轉殖入C57BL/6品系小鼠，以含有625ppm四環黴素飼料餵食小鼠八周後，帶有EGFR轉殖基因的實驗組小鼠肺高度表達EGFR而誘導肺腺癌；而在餵食四周後以Iressa(一種酪胺酸激酶抑制劑)治療小鼠至十六周，癌細胞會大幅減少。以流式細胞儀分析肺沖洗液，顯示CD4+輔助性T細胞比例大於CD8+胞殺性T細胞;EGFR L858R+T790M轉殖小鼠的CD4+ T細胞中CD4+調節性T細胞多於EGFR L858R轉殖小鼠。接著，利用次世代定序分析肺臟、脾臟、全血中T細胞CDR3的cDNA，用以區分癌細胞周圍的TCR群殖增生以及全身性的分布。以GeneSpring軟體分析T細胞 CDR3的定序讀值數，發現在帶有肺腺癌小鼠的肺臟有TCR CDR3的群殖增生，而在脾臟則呈現離散型態。肺臟增殖最多的前十名T細胞 CDR3片段序列帶有部分胺基酸的相似性，顯示其可能源自於某種特定的肺癌抗原。結果顯示具有肺癌特異性的T細胞CDR3可能因特定抗原而在肺臟群殖增生，在全面性免疫系統中則無。在誘導表皮生長因子受體以及以Iressa治療後，小鼠全血可用於監測血液TCRβ增值變化，便於作為生物指標。另外，也期待免疫全面策略可成為新的EGFR誘導型肺癌免疫療法，甚至用於尋找酪胺酸激酶抑制劑抗藥性相關的抗原。

關鍵字

T細胞受體互補決定區3 ；免疫全面多樣；次世代定序；突變型表皮生長因子受體L858R誘導肺腺癌；酪胺酸抑制劑抗藥性

並列摘要

Tumor infiltrating lymphocytes (TILs) are implicating to deal with tumor cells, including lung tumors. Immune repertoire, the overall V(D)J rearrangements in lymphocytes, is a potential index to monitor individual’s T cell receptors (TCRs) or B cell receptors (BCRs) clonal expansion in cancer progression and remission. We hypothesized the TCRβ CDR3, the most various region of antigen binding site, would be functionally and specifically expand in EGFR-driven lung cancer due to tumor antigens stimulation or immune-editing. In this study, we established a mice model to feature a TIL-TCRβ expanding profile in lung cancer. First, Tet-On system was applied to the bi-transgenic mice bearing Clara cell promotor CC10 plus mutant EGFR L858R (EGFR TKI sensitive) or mutant EGFR L858R+T790M (EGFR TKI resistance). With 625ppm doxycycline (tetracycline analogue) induction for lung EGFR expression, mice lungs highly expressed adenocarcinoma. In contrast, lung tumors remission occurred with Iressa (EGFR-Tyrosine kinase inhibitor, TKI) administration. Flow cytometry analysis of BALF identified higher CD4+/CD3+ T cells ratio in lungs, and more CD4+ regulatory T cells in lungs of EGFR L858R+T790M mice than EGFR L858R mice. Then, we used next generation sequencing (NGS) to analyze cDNA of the TCRβ CDR3 in lungs and spleens. Total reads of TCRβ CDR3 represented the abundance of expanded clones. The results suggested some expanded TCRβ CDR3 clones in lung but distributed clones in spleen. The top abundant lung TCRβ CDR3 sequences have similar amino acids order, pointing that these may result from specific antigens. Hence, we claimed tumor-specific TCRβ CDR3 would clonally expand in tumor site but not in overall immune system. To clarify whether TCRβ CDR3 would expand as a profile in peripheral blood, whole blood samples will be analyzed. With the whole immune-profile, we expected TCRβ CDR3 as a potential biomarker of cancer progression and remission, and as an indication of novel lymphocyte-dependent immune therapy.

並列關鍵字

T cell receptor CDR3 ； immune repertoire ； NGS ； EGFR L858R-driven lung adenocarcinoma ； TKI-resistance

參考文獻

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Google Scholar

1. Torre, L.A., F. Bray, R.L. Siegel, J. Ferlay, J. Lortet-Tieulent, and A. Jemal, Global cancer statistics, 2012. CA: A Cancer Journal for Clinicians, 2015. 65(2): p. 87-108.

Google Scholar

2. Herbst, R.S., J.V. Heymach, and S.M. Lippman, Lung Cancer. New England Journal of Medicine, 2008. 359(13): p. 1367-1380.

Google Scholar

3. Peters, S., A.A. Adjei, C. Gridelli, M. Reck, K. Kerr, E. Felip, and o.b.o.t.E.G.W. Group, Metastatic non-small-cell lung cancer (NSCLC): ESMO Clinical Practice Guidelines for diagnosis, treatment and follow-up. Annals of Oncology, 2012. 23(suppl 7): p. vii56-vii64.

Google Scholar

4. Pao, W. and N. Girard, New driver mutations in non-small-cell lung cancer. The Lancet Oncology, 2011. 12(2): p. 175-180.

Google Scholar

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T細胞受體β鏈互補決定區3之頻譜暨高脂飲食研究於突變型表皮生長因子受體誘發之肺腺癌小鼠

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