硒代胱氨酸通過誘導人肝細胞癌中的 DNA 損傷和抑制 DNA 修復途徑來增強順鉑敏感性;Selenocystine enhances Cisplatin sensitivity by inducing DNA Damage and inhibiting DNA repair pathways in Human Hepatocellular Carcinoma

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题名:	硒代胱氨酸通過誘導人肝細胞癌中的 DNA 損傷和抑制 DNA 修復途徑來增強順鉑敏感性;Selenocystine enhances Cisplatin sensitivity by inducing DNA Damage and inhibiting DNA repair pathways in Human Hepatocellular Carcinoma
作者:	方莎南;HASANAH, ULFAH
贡献者:	生命科學系
关键词:	硒代胱氨酸;順鉑;DNA 損傷;DNA 修復活性;Selenocystine;Cisplatin;DNA damage;DNA repair activity
日期:	2023-02-01
上传时间:	2024-09-19 15:19:54 (UTC+8)
出版者:	國立中央大學
摘要:	順鉑是一種 DNA 損傷劑，是多種癌症的常用化療藥物，已顯示其在提供副作用和產生耐藥性方面的局限性。聯合使用對正常細胞毒性較小的藥物可能會為提高順鉑化療的療效提供解決方案。硒代胱氨酸已被證明具有抗癌作用和靶向癌細胞而不是非癌細胞系的選擇性機制。本研究旨在探討順鉑 (cDDP) 和硒代胱氨酸 (SeC) 的聯合治療是否可以提高對肝癌細胞的治療效果。我們發現在 HepG2 和 Hep3B 細胞中，SeC 預處理後聯合 cDDP 比個別單獨處理表現出更多的抑製作用，而在 L- 02 正常肝細胞中僅顯示出很小的影響。這種顯著降低與 DNA 損傷反應相關，因為 cDDP 和 SeC 已被證明具有靶向 DNA 損傷的能力。在遺傳毒性水平上，10 µM SeC 預處理 1 小時，隨後 5 µM cDDP 預處理 24 小時在鹼性和中性彗星試驗中產生顯著的 DNA 損傷，表明單鍊和雙鏈斷裂。隨後 H2AX 的磷酸化作為 DNA 損傷生物標誌物導致 DNA 修復途徑的激活。為了確定雙鏈斷裂 (DSB) DNA 的反應，我們研究了同源重組 (HR) 和非同源末端連接 (NHEJ) 修復途徑的含義。同源重組(HR) 測定結果和 RAD51 蛋白表達水平表明，單一和聯合藥物治療抑制 HR 修復活性，5 µM 順鉑在 HepG2 中表現出最顯著的抑製作用。另一個 DSB 修復途徑，非同源末端連接 (NHEJ) 顯示 Ku-70 和 Ku-80 的表達水平沒有顯著影響，但參與 NHEJ 的下游蛋白如 Artemis、XRCC4 和 DNA 連接酶 IV 導致聯合治療顯著降低。這些結果表明，SeC 和 cDDP 的聯合處理可以通過 HR 和 NHEJ 阻斷 DNA DSB 修復途徑。總之，我們的研究表明，硒代胱氨酸可以選擇性地增強順鉑敏感性，通過誘導 DNA 損傷和抑制 DNA 修復途徑來促進細胞死亡。;Cisplatin, a DNA damaging agent, is the common chemotherapy drug for various kind of cancer which has been shown its limitation in providing side effects and developing resistance. Combining drugs with less toxicity to normal cells may enhance the efficacy of cisplatin chemotherapy. Selenocystine is proven to have anticancer effect and selective mechanism by targeting cancer cells but not non-cancerous cell lines. The current study is aimed to investigate whether a combination treatment between Cisplatin (cDDP) and Selenocystine (SeC) may improve the therapeutic effect against liver cancer cells. We found out that pretreatment SeC followed by cDDP exhibited more inhibition than HepG2 or Hep3B cells with the treatment of single drug exposure, while there was only little effect shown in L 02 normal liver cells. This significant decrease was correlated with DNA damage response as both cDDP and SeC have been proven their ability to elicit DNA damage. In the genotoxicity level, pretreatment 10 µM SeC for 1 hour, followed by 5 µM cDDP for 24 hours, generated significant DNA damage in alkaline and neutral comet assay, indicating single- and double strand breaks occurrence. Subsequent phosphorylation of H2AX, DNA Double Strand Break (DSB) of biomarker, resulted in activation of DNA repair pathways. To determine the response of DSBs, we investigated the implication of Homologous Recombination (HR) and Non-Homologous End-Joining (NHEJ) repair pathways. We observed that significant increase of RAD51 level treated by cDDP can be suppressed by pretreatment of SeC. Moreover, combination treatment exhibited the most significant blocking of HR repair activity in HepG2 cells compared to each single drug exposure. Another DSB repair pathway, Non-Homologous End-Joining (NHEJ) showed that expression level of Ku-70 and Ku-80 showed no considerable effect, but downstream proteins involved in NHEJ such as Artemis, XRCC4, and DNA Ligase IV resulted in a significant decrease by combination treatment. These results suggested that combination treatment of SeC and cDDP can block DNA DSBs repair pathways via HR and NHEJ. Taken together, our study demonstrated that Selenocystine can selectively enhance Cisplatin sensitivity to promote cell death through induction of DNA damage and inhibition of DNA repair pathways.
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