局部麻醉劑利多卡因和布比卡因藉由增加MCF-7細胞中的miR-187-5p來抑制MYB和DANCR lncRNA

Lidocaine and Bupivacaine Downregulate MYB and DANCR lncRNA by Upregulating miR-187-5p in MCF-7 Cells

林巧宜(Chiao-Yi Lin)

指導教授 ： 賴亮全

國立臺灣大學/醫學院/生理學研究所/博士(2022年)

https://doi.org/10.6342/NTU202203050

摘要

乳腺癌是最常見的惡性腫瘤疾病，是女性死亡的主要原因。大多數患者需要手術，而回顧性研究顯示了手術期間的麻醉技術與臨床預後之間的關聯。局部麻醉劑可能通過非編碼核糖核酸(noncoding RNA)相互作用來導致致癌作用。然而，詳細的作用機轉仍不清楚。在這項研究中，使用兩種最常用的局部麻醉劑，利多卡因和布比卡因，在乳腺癌 MCF-7 細胞上進行了研究。功能分析數據顯示局部麻醉劑會抑制癌細胞的生殖。接著，利用次世代定序技術檢驗局部麻醉劑治療的乳腺癌 MCF-7 細胞所產生微小核糖核酸(microRNAs)的表現量。其中，總共有131個微小核糖核酸(microRNAs)的表現量在兩種局部麻醉劑治療的乳腺癌MCF-7細胞中是同步表現上升的。接著使用聚合酶連鎖反應(PCR)驗證，選擇了miR-187-5p這個生物標誌候選物進行進一步分析。生物資訊學預測和螢光素酶基因檢測顯示MYB是miR-187-5p的直接標靶基因。此外，基於長片段非編碼核糖核酸 (lncRNA) 會抑制微小核糖核酸(miRNA sponge)的假設，DIANA-LncBase v2預測了miR-187-5p的標靶基因長片段非編碼核糖核酸DANCR，並通過螢光素酶基因檢測進行了驗證。此外，我們發現了DANCR和miR-187-5p之間也有相互抑制作用。總結以上的實驗結果，這些結果證明了兩種局部麻醉劑，利多卡因和布比卡因，通過 DANCR-miR-187-5p-MYB 軸而呈現的抗腫瘤機轉。我們的研究結果呈現了局部麻醉劑抑制乳腺癌腫瘤細胞的另一種新分子機制。

關鍵字

利多卡因 ； 布比卡因 ； 微小核糖核酸 ； MYB ； 長片段非編碼核糖核酸 ； DANCR ； 乳癌 ； 微小核糖核酸海綿

並列摘要

Breast cancer is the most prevalent malignant disease and the main reason for deaths in women. The majority of patients require surgery, and retrospective studies have demonstrated an association between types of anesthesia during operation and clinical outcomes. By interacting with non-coding RNAs (ncRNAs), local anesthetics (LAs) have an effect on carcinogenesis. However, the detailed mechanisms underlying the interaction between LAs and ncRNAs remain unclear. In this dissertation, I report the mechanisms which two commonly used LAs, bupivacaine and lidocaine, affected the malignancy of MCF-7 breast cancer cells. Next-generation sequencing was used to investigate the expression profiles of the microRNAs (miRNAs) that responded to LA treatments. Results from the functional assay revealed that the LAs inhibited the proliferation of MCF-7 cells. Following treatment with the LAs, next-generation sequencing results showed that 131 miRNAs were upregulated. The putative biomarker miR-187-5p was identified through validation using the polymerase chain reaction (PCR), and it was selected for further examination. Primary prediction with bioinformatics tools and following experiments with luciferase reporter assays demonstrated that MYB is a direct target gene of miR-187-5p. Based on the hypothesis that lncRNAs served as sponges for miRNAs, the target lncRNA, DANCR, of miR-187-5p was predicted using DIANA-LncBase v2 and validated using luciferase reporter assays. Furthermore, the reciprocal inhibitory effect between DANCR and miR-187-5p was discovered. This study suggests that the DANCR-miR-187-5p-MYB axis has a role in one of the anti-tumor mechanisms of lidocaine and bupivacaine. This noval molecular pathway for suppressing breast tumor may open an avenue in treating breast cancer.

並列關鍵字

lidocaine ； bupivacaine ； miRNA ； MYB ； long non-coding RNA ； DANCR ； breast cancer ； miRNA sponge

參考文獻

1. Cizmarikova, R., Cizmarik, J., Valentova, J. et al. Chiral aspects of local anesthetics. Molecules 25, 2738, doi:10.3390/molecules25122738 (2020).

2. Taqi, A., Hong, X., Mistraletti, G. et al. Thoracic epidural analgesia facilitates the restoration of bowel function and dietary intake in patients undergoing laparoscopic colon resection using a traditional, nonaccelerated, perioperative care program. Surg Endosc 21, 247, doi:10.1007/s00464-006-0069-5 (2007).

3. Weibel, S., Jokinen, J., Pace, N. L. et al. Efficacy and safety of intravenous lidocaine for postoperative analgesia and recovery after surgery: A systematic review with trial sequential analysis. Br J Anaesth 116, 770, doi:10.1093/bja/aew101 (2016).

4. Becker, D. E. Reed, K. L. Local anesthetics: Review of pharmacological considerations. Anesth Prog 59, 90, doi:10.2344/0003-3006-59.2.90 (2012).

5. Tedore, T. Regional anaesthesia and analgesia: Relationship to cancer recurrence and survival. Br J Anaesth 115 Suppl 2, ii34, doi:10.1093/bja/aev375 (2015).