Salirasib inhibits the growth of hepatocarcinoma cell lines in vitro and tumor growth in vivo through ras and mTOR inhibition.
BACKGROUND
Dysregulation of epidermal growth factor and insulin-like growth factor signaling play important roles in human hepatocellular carcinoma (HCC), leading to frequent activation of their downstream targets, the ras/raf/extracellular signal-regulated kinase (ERK) and the phosphoinositide 3-kinase (PI3K)/Akt/mammalian Target of Rapamycin (mTOR) pathways. Salirasib is an S-prenyl-cysteine analog that has been shown to block ras and/or mTOR activation in several non hepatic tumor cell lines. We investigated in vitro the effect of salirasib on cell growth as well as its mechanism of action in human hepatoma cell lines (HepG2, Huh7, and Hep3B) and its in vivo effect in a subcutaneous xenograft model with HepG2 cells.


RESULTS
Salirasib induced a time and dose dependent growth inhibition in hepatocarcinoma cells through inhibition of proliferation and partially through induction of apoptosis. A 50 percent reduction in cell growth was obtained in all three cell lines at a dose of 150 μM when they were cultured with serum. By contrast, salirasib was more potent at reducing cell growth after stimulation with EGF or IGF2 under serum-free conditions, with an IC50 ranging from 60 μM to 85 μM. The drug-induced anti-proliferative effect was associated with downregulation of cyclin A and to a lesser extent of cyclin D1, and upregulation of p21 and p27. Apoptosis induction was related to a global pro-apoptotic balance with caspase 3 activation, cytochrome c release, death receptor upregulation, and a reduced mRNA expression of the apoptosis inhibitors cFLIP and survivin. These effects were associated with ras downregulation and mTOR inhibition, without reduction of ERK and Akt activation. In vivo, salirasib reduced tumour growth from day 5 onwards. After 12 days of treatment, mean tumor weight was diminished by 56 percent in the treated animals.


CONCLUSIONS
Our results show for the first time that salirasib inhibits the growth of human hepatoma cell lines through inhibition of proliferation and induction of apoptosis, which is associated with ras and mTOR inhibition. The therapeutic potential of salirasib in human HCC was further confirmed in a subcutaneous xenograft model.
Charette
Nicolas
N
Laboratory of Gastroenterology, Institut de Recherche Expérimentale et Clinique, Université Catholique de Louvain, 1200 Brussels, Belgium.
De Saeger
Christine
C
Lannoy
Valérie
V
Horsmans
Yves
Y
Leclercq
Isabelle
I
Stärkel
Peter
P
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology / Q000627:therapeutic use; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism*; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D019925:Cyclin A / Q000235:genetics; D019938:Cyclin D1 / Q000235:genetics; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000235:genetics; D004815:Epidermal Growth Factor / Q000494:pharmacology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000235:genetics; D005204:Farnesol / Q000031:analogs & derivatives* / Q000494:pharmacology / Q000627:therapeutic use; D005260:Female; D056945:Hep G2 Cells; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins; D007335:Insulin-Like Growth Factor II / Q000494:pharmacology; D008113:Liver Neoplasms / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008819:Mice, Nude; D008869:Microtubule-Associated Proteins / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012459:Salicylates / Q000494:pharmacology / Q000627:therapeutic use; D000077022:Survivin; D058570:TOR Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D023041:Xenograft Model Antitumor Assays; D018631:ras Proteins / Q000235:genetics / Q000378:metabolism*
D000970:Antineoplastic Agents; C107489:BIRC5 protein, human; D019925:Cyclin A; D050759:Cyclin-Dependent Kinase Inhibitor p21; D051034:Inhibitor of Apoptosis Proteins; D008869:Microtubule-Associated Proteins; D012459:Salicylates; D000077022:Survivin; C093323:farnesylthiosalicylic acid; D019938:Cyclin D1; D050760:Cyclin-Dependent Kinase Inhibitor p27; D005204:Farnesol; D004815:Epidermal Growth Factor; D007335:Insulin-Like Growth Factor II; D058570:TOR Serine-Threonine Kinases; D048049:Extracellular Signal-Regulated MAP Kinases; D018631:ras Proteins