低氧環境對於標靶藥物治療HER-2過量表現胃癌的影響

Abstract

在台灣，胃癌佔了國人十大死因的第五位。目前治療胃癌的方式主要是以手術輔以化學治療或是放射治療。對於某些會過量表現HER2的胃癌，近年也使用標靶藥物(Herceptin)搭配化學療法，使得癌症治療更具專一性，且減少化療導致的正常細胞受損。然而，使用標靶藥物一段時間後，胃癌常會出現對於Herceptin的抗藥性。有研究指出，腫瘤生長過程所發生的低氧（Hypoxia）情形，對癌細胞抗藥性的發生可能有重要影響。本研究的目的，在探討低氧環境對於(1)胃癌細胞生長趨勢的影響;(2)在過量表現HER2胃癌細胞株N87，使用Herceptin療效的影響，以及其可能的機制。 吾人使用體外細胞株及低氧培養箱，比較胃癌細胞株N87在正常氧及低氧環境時的生長及對Herceptin的藥效。結果顯示，在低氧的環境下，N87的形態呈現非聚合狀且細胞週期滯留於G1期。此外，利用流式細胞儀及MTT assay分析，Herceptin在低氧環境下之抑癌效果較在於正常氧壓下的效果差。 吾人並以西方墨點試驗分析HER2在低氧環境下之訊息傳遞。結果顯示，低氧環境並不影響Herceptin與HER2的聯結，但會減少Herceptin抑制p-HER2表現的現象。 再者，在正常氧壓情形下，Herceptin能有效地抑制pAKT之表現量並觀察到且p27的表現在1, 4, 8, 12小時有增加的現象。反觀，在低氧環境下，AKT與p27的磷酸化則皆不受Herceptin影響。故Herceptin在低氧環境下，對於HER2訊息傳遞並沒有抑制的效果。 從上述實驗結果可以得知，低氧環境會影響N87生長的週期，並導致標靶藥物Herceptin的抑癌效果不彰 其機制可能和低氧改變HER2之訊息傳遞有關。

Gastric cancer ranks the 5th of the 10 leading cause of death in Taiwan in recent years. To date, the main treatment of gastric cancer, is surgery combined with chemotherapy and radiotherapy. Recently, patients with HER2-overexpressed gastric cancer are treated with target therapy (Herceptin) combined with chemotherapy, for a more specific and effective treatment but also to circumvent the cytotoxicity to normal cells. However, drug resistant develops shortly after treatment. Tumor hypoxia has been implicated as one of the underlying factors. The purposes of this study are (i) to observe the growth characteristics of a gastric cancer cell line N87 with overexpressed HER2 under hypoxia, and (ii) the efficacy of Herceptin treatment on N87 under hypoxia and possible mechanisms. We used in vitro gastric cancer cell line N87 and hypoxia chamber to compare the cell growth and Herceptin efficacy differences between N87 cultured under normoxic and hypoxic condition. The results showed that hypoxia altered the confluent growth pattern of N87 cell, reduced proliferation rate and arrest the cell cycle progression at G1 phase. In addition, by using flow cytometry and MTT assay, the cytotoxicity of Herceptin on N87 cells is less evident under hypoxic when compare to that under normoxic condition. The Her2 signaling cascade was analyzed by Western blot. The binding of Herceptin and Her2 was not affected, but Herceptin-induced p-HER2 repression was decreased under hypoxia. In addition, the expression of pAkt, a downstream signal protein of Her2, was reduced after 1, 4, 8, and 12 hours of Herceptin treatment under normoxia; while p27 is increased at the correspondent time points. Under hypoxia, Herceptin treatment did not change the pAKT and p27 levels. The findings of this study clear suggest that hypoxic condition can affect N87 cell growth and mitigate Herceptin’s effectiveness which was related to the alteration of the HER2 signaling under hypoxia.