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Cholesterol-conjugated let-7a mimics: antitumor efficacy on hepatocellular carcinoma in vitro and in a preclinical orthotopic xenograft model of systemic therapy.

Liu YM, Xia Y, Dai W, Han HY, Dong YX, Cai J, Zeng X, Luo FY, Yang T, Li YZ, Chen J, Guan J - BMC Cancer (2014)

Bottom Line: Furthermore, let-7a abundance in orthotopic xenografts was coincident with a reduction in the expression of 3 human ras mRNAs and RAS proteins.Chol-let-7a exerted significant antitumor effects by down-regulating all human ras genes at the transcriptional and translational levels.Chol-let-7a represents a potential useful modified molecule for systemic HCC therapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Peking Union Medical College (PUMC) Hospital, PUMC & Chinese Academy of Medical Sciences (CAMS), Beijing, China. xhblk@163.com.

ABSTRACT

Background: A major challenge to the clinical utility of let-7 for hepatocellular carcinoma (HCC) therapy is the lack of an effective carrier to target tumours. We confirmed the high transfection efficiency of cholesterol-conjugated let-7a miRNA mimics (Chol-let-7a) in human HCC cells, as well as their high affinity for liver tissue in nude mice. However, their antitumor efficacy via systemic delivery remains unknown.

Methods: We explored the effects of Chol-let-7a on HCC in vitro and in vivo. Cell viability and mobility, let-7a abundance and the target ras genes was measured. Live-cell image and cell ultrastructure was observed. Antitumor efficacy in vivo was analyzed by ultrasonography, hispatholgogy and transmission electronic microscopy in a preclinical model of HCC orthotopic xenografts with systemic therapy.

Results: Chol-let-7a inhibited the viability and mobility of HCC cells. Chol-let-7a was primarily observed in the cytoplasm and induced organelle changes, including autophagy. Mild changes were observed in the cells treated with negative control miRNA. Chol-let-7a reached HCC orthotopic tumours, significantly inhibited tumour growth, and prevented local invasion and metastasis. Compared to control tumours, Chol-let-7a-treated tumours showed more necrosis. Tumour cells showed no significant atypia, and mitoses were very rare after systemic Chol-let-7a therapy. Furthermore, let-7a abundance in orthotopic xenografts was coincident with a reduction in the expression of 3 human ras mRNAs and RAS proteins.

Conclusions: Chol-let-7a exerted significant antitumor effects by down-regulating all human ras genes at the transcriptional and translational levels. Chol-let-7a inhibited cell proliferation, growth, and metastasis, and mainly functioned in the cytoplasm. Chol-let-7a represents a potential useful modified molecule for systemic HCC therapy.

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SystemicChol-let-7atherapy inhibited growth and metastasis of orthotopic HepG2 xenografts. A: Xenograft growth curve. Representative xenografts from all groups are illustrated (N =6 mice per cohort). The Chol-let-7a (green), Chol-miRCtrl (red), and blank control (blue) groups are shown. B: Macroscopic view of the HCC orthotopic tumour and metastasis within the liver of the treatment groups. C: Orthotopic HepG2 xenografts examined by ultrasonography. Xenograft images were recorded 1, 3, and 5 weeks after HepG2 cells were transplanted. D: TEM of HCC orthotopic tumours in vivo. Necrotic and tumour cells in HCC tissues are shown. Short red arrows indicate the microvasculature in orthotopic tumours. Scale bars are 2 μm or 0.5 μm. E: H&E staining of orthotopic HCC tumour tissues. H&E staining was performed at the culmination of Chol-let-7a therapy. Differentiation and mitoses (short yellow arrows) of HCC cells of the 3 treatment groups are shown; all were recorded beside necrotic areas. Scale bars are 20 μm.
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Fig5: SystemicChol-let-7atherapy inhibited growth and metastasis of orthotopic HepG2 xenografts. A: Xenograft growth curve. Representative xenografts from all groups are illustrated (N =6 mice per cohort). The Chol-let-7a (green), Chol-miRCtrl (red), and blank control (blue) groups are shown. B: Macroscopic view of the HCC orthotopic tumour and metastasis within the liver of the treatment groups. C: Orthotopic HepG2 xenografts examined by ultrasonography. Xenograft images were recorded 1, 3, and 5 weeks after HepG2 cells were transplanted. D: TEM of HCC orthotopic tumours in vivo. Necrotic and tumour cells in HCC tissues are shown. Short red arrows indicate the microvasculature in orthotopic tumours. Scale bars are 2 μm or 0.5 μm. E: H&E staining of orthotopic HCC tumour tissues. H&E staining was performed at the culmination of Chol-let-7a therapy. Differentiation and mitoses (short yellow arrows) of HCC cells of the 3 treatment groups are shown; all were recorded beside necrotic areas. Scale bars are 20 μm.

Mentions: To study antitumor efficacy in vivo, we examined the size of HepG2 orthotopic xenografts of different groups by ultrasound weekly after cell transplantation. As shown in Figure 5, the growth of orthotopic tumours was significantly inhibited following Chol-let-7a therapy (Figure 5A-C). One week after HepG2 cell transplantation, the xenografts (Volume, mm3) of the Chol-let-7a group (8.2854 ± 2.122593) were slightly larger than those of the 2 control groups (Chol-miRCtrl, 7.8614 ± 1.69912; blank, 7.0574 ± 1.340323), while the xenografts of the Chol-let-7a-treated group (152.1528 ± 38.43266) were significantly smaller than those of the 2 control groups (Chol-miRCtrl, 424.3472 ± 60.10395; blank, 380.2284 ± 74.83116) at the culmination of therapy. The inhibitory rates produced by Chol-let-7a and Chol-miRCtrl on xenografts were 45.49% (p <0.01) and -7.13% (p >0.05), respectively, in comparison with the blank control group that was treated with saline buffer alone.Figure 5


Cholesterol-conjugated let-7a mimics: antitumor efficacy on hepatocellular carcinoma in vitro and in a preclinical orthotopic xenograft model of systemic therapy.

Liu YM, Xia Y, Dai W, Han HY, Dong YX, Cai J, Zeng X, Luo FY, Yang T, Li YZ, Chen J, Guan J - BMC Cancer (2014)

SystemicChol-let-7atherapy inhibited growth and metastasis of orthotopic HepG2 xenografts. A: Xenograft growth curve. Representative xenografts from all groups are illustrated (N =6 mice per cohort). The Chol-let-7a (green), Chol-miRCtrl (red), and blank control (blue) groups are shown. B: Macroscopic view of the HCC orthotopic tumour and metastasis within the liver of the treatment groups. C: Orthotopic HepG2 xenografts examined by ultrasonography. Xenograft images were recorded 1, 3, and 5 weeks after HepG2 cells were transplanted. D: TEM of HCC orthotopic tumours in vivo. Necrotic and tumour cells in HCC tissues are shown. Short red arrows indicate the microvasculature in orthotopic tumours. Scale bars are 2 μm or 0.5 μm. E: H&E staining of orthotopic HCC tumour tissues. H&E staining was performed at the culmination of Chol-let-7a therapy. Differentiation and mitoses (short yellow arrows) of HCC cells of the 3 treatment groups are shown; all were recorded beside necrotic areas. Scale bars are 20 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4289300&req=5

Fig5: SystemicChol-let-7atherapy inhibited growth and metastasis of orthotopic HepG2 xenografts. A: Xenograft growth curve. Representative xenografts from all groups are illustrated (N =6 mice per cohort). The Chol-let-7a (green), Chol-miRCtrl (red), and blank control (blue) groups are shown. B: Macroscopic view of the HCC orthotopic tumour and metastasis within the liver of the treatment groups. C: Orthotopic HepG2 xenografts examined by ultrasonography. Xenograft images were recorded 1, 3, and 5 weeks after HepG2 cells were transplanted. D: TEM of HCC orthotopic tumours in vivo. Necrotic and tumour cells in HCC tissues are shown. Short red arrows indicate the microvasculature in orthotopic tumours. Scale bars are 2 μm or 0.5 μm. E: H&E staining of orthotopic HCC tumour tissues. H&E staining was performed at the culmination of Chol-let-7a therapy. Differentiation and mitoses (short yellow arrows) of HCC cells of the 3 treatment groups are shown; all were recorded beside necrotic areas. Scale bars are 20 μm.
Mentions: To study antitumor efficacy in vivo, we examined the size of HepG2 orthotopic xenografts of different groups by ultrasound weekly after cell transplantation. As shown in Figure 5, the growth of orthotopic tumours was significantly inhibited following Chol-let-7a therapy (Figure 5A-C). One week after HepG2 cell transplantation, the xenografts (Volume, mm3) of the Chol-let-7a group (8.2854 ± 2.122593) were slightly larger than those of the 2 control groups (Chol-miRCtrl, 7.8614 ± 1.69912; blank, 7.0574 ± 1.340323), while the xenografts of the Chol-let-7a-treated group (152.1528 ± 38.43266) were significantly smaller than those of the 2 control groups (Chol-miRCtrl, 424.3472 ± 60.10395; blank, 380.2284 ± 74.83116) at the culmination of therapy. The inhibitory rates produced by Chol-let-7a and Chol-miRCtrl on xenografts were 45.49% (p <0.01) and -7.13% (p >0.05), respectively, in comparison with the blank control group that was treated with saline buffer alone.Figure 5

Bottom Line: Furthermore, let-7a abundance in orthotopic xenografts was coincident with a reduction in the expression of 3 human ras mRNAs and RAS proteins.Chol-let-7a exerted significant antitumor effects by down-regulating all human ras genes at the transcriptional and translational levels.Chol-let-7a represents a potential useful modified molecule for systemic HCC therapy.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Peking Union Medical College (PUMC) Hospital, PUMC & Chinese Academy of Medical Sciences (CAMS), Beijing, China. xhblk@163.com.

ABSTRACT

Background: A major challenge to the clinical utility of let-7 for hepatocellular carcinoma (HCC) therapy is the lack of an effective carrier to target tumours. We confirmed the high transfection efficiency of cholesterol-conjugated let-7a miRNA mimics (Chol-let-7a) in human HCC cells, as well as their high affinity for liver tissue in nude mice. However, their antitumor efficacy via systemic delivery remains unknown.

Methods: We explored the effects of Chol-let-7a on HCC in vitro and in vivo. Cell viability and mobility, let-7a abundance and the target ras genes was measured. Live-cell image and cell ultrastructure was observed. Antitumor efficacy in vivo was analyzed by ultrasonography, hispatholgogy and transmission electronic microscopy in a preclinical model of HCC orthotopic xenografts with systemic therapy.

Results: Chol-let-7a inhibited the viability and mobility of HCC cells. Chol-let-7a was primarily observed in the cytoplasm and induced organelle changes, including autophagy. Mild changes were observed in the cells treated with negative control miRNA. Chol-let-7a reached HCC orthotopic tumours, significantly inhibited tumour growth, and prevented local invasion and metastasis. Compared to control tumours, Chol-let-7a-treated tumours showed more necrosis. Tumour cells showed no significant atypia, and mitoses were very rare after systemic Chol-let-7a therapy. Furthermore, let-7a abundance in orthotopic xenografts was coincident with a reduction in the expression of 3 human ras mRNAs and RAS proteins.

Conclusions: Chol-let-7a exerted significant antitumor effects by down-regulating all human ras genes at the transcriptional and translational levels. Chol-let-7a inhibited cell proliferation, growth, and metastasis, and mainly functioned in the cytoplasm. Chol-let-7a represents a potential useful modified molecule for systemic HCC therapy.

Show MeSH
Related in: MedlinePlus