Inhibition of CD44 expression in hepatocellular carcinoma cells enhances apoptosis, chemosensitivity, and reduces tumorigenesis and invasion.
OBJECTIVE
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.


METHODS
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.


RESULTS
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.


CONCLUSIONS
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.
Xie
Zhigang
Z
Department of Medicine, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.
Choong
Pei Feng
PF
Poon
Lai Fong
LF
Zhou
Jianbiao
J
Khng
Jiaying
J
Jasinghe
Viraj Janakakumara
VJ
Palaniyandi
Senthilnathan
S
Chen
Chien-Shing
CS
D017209:Apoptosis / Q000187:drug effects*; D006528:Carcinoma, Hepatocellular / Q000473:pathology*; D045744:Cell Line, Tumor / Q000187:drug effects; D004317:Doxorubicin / Q000008:administration & dosage / Q000494:pharmacology*; D019008:Drug Resistance, Neoplasm / Q000187:drug effects*; D004357:Drug Synergism; D006801:Humans; D018960:Hyaluronan Receptors / Q000096:biosynthesis* / Q000235:genetics; D008113:Liver Neoplasms / Q000473:pathology*; D009361:Neoplasm Invasiveness; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000235:genetics; D016376:Oligonucleotides, Antisense / Q000008:administration & dosage / Q000494:pharmacology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis; D012334:RNA, Neoplasm / Q000096:biosynthesis; D014162:Transfection; D014410:Tumor Stem Cell Assay
C497483:CD44 protein, human; D018960:Hyaluronan Receptors; D009363:Neoplasm Proteins; D016376:Oligonucleotides, Antisense; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D004317:Doxorubicin