Figure 9. Silencing of Igf2bp3 and Yap1 sensitizes TICs to drug-induced cell death.

(A) Silencing of Igf2bp3 and Yap1 in TICs from HCV Ns5a Tg mice promotes growth inhibition of tumor upon chemotherapeutic drug treatment (rapa and sorafenib) in immune-competent C57BL/6 mice, as monitored by GFP imaging after splenic injection and engraftment enrichment procedures with retrorsine and CCl₄. (B) The tumor volume measurement performed at day 90 after transplantation shows a maximal growth retardation achieved by the combination of the drugs and Igf2bp3/Yap1 silencing (*P < 0.05). (C) TIC proliferation, as determined by BrdU incorporation in vivo at day 90, is most inhibited by the combination of the chemotherapeutic drugs and silencing of Igf2bp3 and Yap1. Concomitantly, the incidence of apoptotic cell death increases conspicuously by the combined treatment. *P < 0.05. (D) Xenograft tumor growth by TICs from patient HCC in NOG mice is most conspicuously suppressed by rapa and sorafanib treatment only in the presence of IGF2BP3 and/or YAP1 silencing (*P < 0.05). (E) Improved survival of mice injected with human TICs transduced with shRNA targeting YAP1 and IGF2BP3 and subjected to injection of the chemotherapeutic drugs, as compared with other groups with single therapies. (F) Immunoblot analysis demonstrates that silencing of both YAP1 and IGF2BP3 induces p-SMAD3 levels, while reducing NANOG expression in xenograft tissue of mice treated with rapa and sorafenib.