Xie Zhigang, Choong Pei Feng, Poon Lai Fong, Zhou Jianbiao, Khng Jiaying, Jasinghe Viraj Janakakumara, Palaniyandi Senthilnathan, Chen Chien-Shing
Department of Medicine, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.
Cancer Chemother Pharmacol. 2008 Nov;62(6):949-57. doi: 10.1007/s00280-008-0684-z. Epub 2008 Feb 8.
CD44 is overexpressed in various tumors including hepatocellular carcinoma (HCC). The purpose of this study was to examine the effects of CD44 antisense oligonucleotide (ASO) alone or combination with doxorubicin on HCC cells in vitro.
Cytotoxicity was measured by use of a cell viability assay in HCC cell line SNU-449. Tumorigenesis and invasion were accessed by colony formation, growth in soft agar and ECMatrix invasion assay. Apoptosis and necrosis were evaluated by using double staining with Hoechst 33342 and propidium iodide. Protein expression and mRNA level were detected by Western blot and RT-PCR.
We have designed novel CD44 ASO, which can effectively down-regulate CD44 expression in SNU-449. Colony formation, growth in soft agar and invasion were significantly impaired after CD44 ASO treatment in SNU-499. In company with CD44 down-regulated by CD44 ASO, MDR-1 and Bcl-2 expression were also greatly reduced. CD44 ASO also increased chemosensitivity to doxorubicin significantly, lowered IC(50 )by one order of magnitude. Apoptosis and necrosis were also induced by CD44 ASO alone or in combination treatment with doxorubicin.
Inhibition of CD44 expression by CD44 ASO significantly induced apoptosis, decreased tumorigenesis and invasion, and increased chemosensitivity. Thus, CD44 ASO is potentially a therapy that is worth investigating in the clinical setting.
CD44在包括肝细胞癌（HCC）在内的多种肿瘤中过表达。本研究的目的是检测CD44反义寡核苷酸（ASO）单独或与阿霉素联合应用对体外HCC细胞的影响。
采用细胞活力测定法检测HCC细胞系SNU - 449的细胞毒性。通过集落形成、软琼脂生长和细胞外基质侵袭试验评估肿瘤发生和侵袭情况。使用Hoechst 33342和碘化丙啶双重染色评估凋亡和坏死情况。通过蛋白质印迹法和逆转录聚合酶链反应检测蛋白质表达和mRNA水平。
我们设计了新型CD44 ASO，其可有效下调SNU - 449中CD44的表达。在SNU - 499中，CD44 ASO处理后集落形成、软琼脂生长和侵袭均受到显著抑制。伴随CD44 ASO下调CD44，多药耐药蛋白1（MDR - 1）和Bcl - 2的表达也大幅降低。CD44 ASO还显著增加了对阿霉素的化疗敏感性，使半数抑制浓度（IC50）降低了一个数量级。单独使用CD44 ASO或与阿霉素联合处理均诱导了凋亡和坏死。
CD44 ASO抑制CD44表达可显著诱导凋亡、降低肿瘤发生和侵袭，并增加化疗敏感性。因此，CD44 ASO可能是一种值得在临床环境中研究的治疗方法。
