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Plasmalemmal Vesicle Associated Protein (PLVAP) as a therapeutic target for treatment of hepatocellular carcinoma.

Wang YH, Cheng TY, Chen TY, Chang KM, Chuang VP, Kao KJ - BMC Cancer (2014)

Bottom Line: Systemic administration did not induce tumor necrosis.The results of this study suggest that anti-PLVAP Fab-TF may be used to treat HCC cases for which transcatheter arterial chemoembolization (TACE) is currently used and potentially avoid the drawback of high viscosity of chemoembolic emulsion for TACE to improve therapeutic outcome.Anti-PLVAP Fab-TF may become a viable therapeutic agent in patients with advanced disease and compromised liver function.

View Article: PubMed Central - PubMed

Affiliation: Department of Research, Koo Foundation Sun Yat-Sen Cancer Center, Lih-Der Road, Taipei, Taiwan. kjkao@kfsyscc.org.

ABSTRACT

Background: Hepatocellular carcinoma (HCC) is a malignancy with poor survival outcome. New treatment options for the disease are needed. In this study, we identified and evaluated tumor vascular PLVAP as a therapeutic target for treatment of HCC.

Methods: Genes showing extreme differential expression between paired human HCC and adjacent non-tumorous liver tissue were investigated. PLVAP was identified as one of such genes with potential to serve as a therapeutic target for treatment of HCC. A recombinant monoclonal anti-PLVAP Fab fragment co-expressing extracellular domain of human tissue factor (TF) was developed. The potential therapeutic effect and toxicity to treat HCC were studied using a Hep3B HCC xenograft model in SCID mice.

Results: PLVAP was identified as a gene specifically expressed in vascular endothelial cells of HCC but not in non-tumorous liver tissues. This finding was confirmed by RT-PCR analysis of micro-dissected cells and immunohistochemical staining of tissue sections. Infusion of recombinant monoclonal anti-PLVAP Fab-TF into the main tumor feeding artery induced tumor vascular thrombosis and extensive tumor necrosis at doses between 2.5 μg and 12 μg. Tumor growth was suppressed for 40 days after a single treatment. Systemic administration did not induce tumor necrosis. Little systemic toxicity was noted for this therapeutic agent.

Conclusions: The results of this study suggest that anti-PLVAP Fab-TF may be used to treat HCC cases for which transcatheter arterial chemoembolization (TACE) is currently used and potentially avoid the drawback of high viscosity of chemoembolic emulsion for TACE to improve therapeutic outcome. Anti-PLVAP Fab-TF may become a viable therapeutic agent in patients with advanced disease and compromised liver function.

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Differential expression of PLVAP between paired HCC tissue and adjacent non-tumorous liver tissue. A: Differential expression of the PLVAP gene according to microarrays of 18 pairs of HCC and adjacent non-tumorous liver tissue. PN: paired non-tumorous liver; PHCC: paired HCC tissue. B: Relative quantities of PLVAP mRNA in the same 18 tissue pairs. One non-tumorous liver tissue sample was chosen as a reference control (relative quantitative expression = 1). C: Immunohistochemical (IHC) staining of PLVAP in four randomly selected HCC cases. IHC staining was performed using GY5 murine anti-human PLVAP monoclonal antibody. Endothelial cells lining blood vessels of HCC showed positive staining for PLVAP in brown color (arrows). IHC staining (panel C) showed that PLVAP was not expressed by the endothelial cells of hepatic central vein (C-II right panel), hepatic sinusoid (CI-IV right panels), and hepatic arterioles (portal tract) (C-III right panel) in the adjacent non-tumorouse liver tissues. The large empty space in the right panel of C-II was lumen of a hepatic central vein which showed absence of PLVAP expression in the lining endothelial cells. We also stained HCC sections including adjacent non-tumorous liver with anti-human CD34 monoclonal antibody. Endothelial cells of hepatic central vein and hepatic areteriole were stained positively for CD34 expression (data not shown). Liver sinusoidal endothelial cells did not express CD34 as expected.
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Fig3: Differential expression of PLVAP between paired HCC tissue and adjacent non-tumorous liver tissue. A: Differential expression of the PLVAP gene according to microarrays of 18 pairs of HCC and adjacent non-tumorous liver tissue. PN: paired non-tumorous liver; PHCC: paired HCC tissue. B: Relative quantities of PLVAP mRNA in the same 18 tissue pairs. One non-tumorous liver tissue sample was chosen as a reference control (relative quantitative expression = 1). C: Immunohistochemical (IHC) staining of PLVAP in four randomly selected HCC cases. IHC staining was performed using GY5 murine anti-human PLVAP monoclonal antibody. Endothelial cells lining blood vessels of HCC showed positive staining for PLVAP in brown color (arrows). IHC staining (panel C) showed that PLVAP was not expressed by the endothelial cells of hepatic central vein (C-II right panel), hepatic sinusoid (CI-IV right panels), and hepatic arterioles (portal tract) (C-III right panel) in the adjacent non-tumorouse liver tissues. The large empty space in the right panel of C-II was lumen of a hepatic central vein which showed absence of PLVAP expression in the lining endothelial cells. We also stained HCC sections including adjacent non-tumorous liver with anti-human CD34 monoclonal antibody. Endothelial cells of hepatic central vein and hepatic areteriole were stained positively for CD34 expression (data not shown). Liver sinusoidal endothelial cells did not express CD34 as expected.

Mentions: To identify genes specifically expressed in HCC and not in non-tumorous liver tissue, we compared the gene expression profiles of 18 pairs of HCC and adjacent non-tumorous liver tissues. HCC-specific gene expression was defined as expression of a gene determined to be “present” in HCC and “absent or marginal” in adjacent non-tumorous liver tissues by both MAS5.0 and dChip softwares in at least 16 of the 18 pairs of tissue samples. Using this stringent approach only two genes met the criteria. One was PVLAP and the other was MELK. After further examining PLVAP expression in all 18 tissue pairs (Figure 3A), we found that all pairs but one showed higher PLVAP expression in the HCC tissues. The differential expression of PLVAP in HCC was confirmed in 16 out of the same 18 tissue pairs using TaqMan real time quantitative RT-PCR (Figure 3B).Figure 3


Plasmalemmal Vesicle Associated Protein (PLVAP) as a therapeutic target for treatment of hepatocellular carcinoma.

Wang YH, Cheng TY, Chen TY, Chang KM, Chuang VP, Kao KJ - BMC Cancer (2014)

Differential expression of PLVAP between paired HCC tissue and adjacent non-tumorous liver tissue. A: Differential expression of the PLVAP gene according to microarrays of 18 pairs of HCC and adjacent non-tumorous liver tissue. PN: paired non-tumorous liver; PHCC: paired HCC tissue. B: Relative quantities of PLVAP mRNA in the same 18 tissue pairs. One non-tumorous liver tissue sample was chosen as a reference control (relative quantitative expression = 1). C: Immunohistochemical (IHC) staining of PLVAP in four randomly selected HCC cases. IHC staining was performed using GY5 murine anti-human PLVAP monoclonal antibody. Endothelial cells lining blood vessels of HCC showed positive staining for PLVAP in brown color (arrows). IHC staining (panel C) showed that PLVAP was not expressed by the endothelial cells of hepatic central vein (C-II right panel), hepatic sinusoid (CI-IV right panels), and hepatic arterioles (portal tract) (C-III right panel) in the adjacent non-tumorouse liver tissues. The large empty space in the right panel of C-II was lumen of a hepatic central vein which showed absence of PLVAP expression in the lining endothelial cells. We also stained HCC sections including adjacent non-tumorous liver with anti-human CD34 monoclonal antibody. Endothelial cells of hepatic central vein and hepatic areteriole were stained positively for CD34 expression (data not shown). Liver sinusoidal endothelial cells did not express CD34 as expected.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Fig3: Differential expression of PLVAP between paired HCC tissue and adjacent non-tumorous liver tissue. A: Differential expression of the PLVAP gene according to microarrays of 18 pairs of HCC and adjacent non-tumorous liver tissue. PN: paired non-tumorous liver; PHCC: paired HCC tissue. B: Relative quantities of PLVAP mRNA in the same 18 tissue pairs. One non-tumorous liver tissue sample was chosen as a reference control (relative quantitative expression = 1). C: Immunohistochemical (IHC) staining of PLVAP in four randomly selected HCC cases. IHC staining was performed using GY5 murine anti-human PLVAP monoclonal antibody. Endothelial cells lining blood vessels of HCC showed positive staining for PLVAP in brown color (arrows). IHC staining (panel C) showed that PLVAP was not expressed by the endothelial cells of hepatic central vein (C-II right panel), hepatic sinusoid (CI-IV right panels), and hepatic arterioles (portal tract) (C-III right panel) in the adjacent non-tumorouse liver tissues. The large empty space in the right panel of C-II was lumen of a hepatic central vein which showed absence of PLVAP expression in the lining endothelial cells. We also stained HCC sections including adjacent non-tumorous liver with anti-human CD34 monoclonal antibody. Endothelial cells of hepatic central vein and hepatic areteriole were stained positively for CD34 expression (data not shown). Liver sinusoidal endothelial cells did not express CD34 as expected.
Mentions: To identify genes specifically expressed in HCC and not in non-tumorous liver tissue, we compared the gene expression profiles of 18 pairs of HCC and adjacent non-tumorous liver tissues. HCC-specific gene expression was defined as expression of a gene determined to be “present” in HCC and “absent or marginal” in adjacent non-tumorous liver tissues by both MAS5.0 and dChip softwares in at least 16 of the 18 pairs of tissue samples. Using this stringent approach only two genes met the criteria. One was PVLAP and the other was MELK. After further examining PLVAP expression in all 18 tissue pairs (Figure 3A), we found that all pairs but one showed higher PLVAP expression in the HCC tissues. The differential expression of PLVAP in HCC was confirmed in 16 out of the same 18 tissue pairs using TaqMan real time quantitative RT-PCR (Figure 3B).Figure 3

Bottom Line: Systemic administration did not induce tumor necrosis.The results of this study suggest that anti-PLVAP Fab-TF may be used to treat HCC cases for which transcatheter arterial chemoembolization (TACE) is currently used and potentially avoid the drawback of high viscosity of chemoembolic emulsion for TACE to improve therapeutic outcome.Anti-PLVAP Fab-TF may become a viable therapeutic agent in patients with advanced disease and compromised liver function.

View Article: PubMed Central - PubMed

Affiliation: Department of Research, Koo Foundation Sun Yat-Sen Cancer Center, Lih-Der Road, Taipei, Taiwan. kjkao@kfsyscc.org.

ABSTRACT

Background: Hepatocellular carcinoma (HCC) is a malignancy with poor survival outcome. New treatment options for the disease are needed. In this study, we identified and evaluated tumor vascular PLVAP as a therapeutic target for treatment of HCC.

Methods: Genes showing extreme differential expression between paired human HCC and adjacent non-tumorous liver tissue were investigated. PLVAP was identified as one of such genes with potential to serve as a therapeutic target for treatment of HCC. A recombinant monoclonal anti-PLVAP Fab fragment co-expressing extracellular domain of human tissue factor (TF) was developed. The potential therapeutic effect and toxicity to treat HCC were studied using a Hep3B HCC xenograft model in SCID mice.

Results: PLVAP was identified as a gene specifically expressed in vascular endothelial cells of HCC but not in non-tumorous liver tissues. This finding was confirmed by RT-PCR analysis of micro-dissected cells and immunohistochemical staining of tissue sections. Infusion of recombinant monoclonal anti-PLVAP Fab-TF into the main tumor feeding artery induced tumor vascular thrombosis and extensive tumor necrosis at doses between 2.5 μg and 12 μg. Tumor growth was suppressed for 40 days after a single treatment. Systemic administration did not induce tumor necrosis. Little systemic toxicity was noted for this therapeutic agent.

Conclusions: The results of this study suggest that anti-PLVAP Fab-TF may be used to treat HCC cases for which transcatheter arterial chemoembolization (TACE) is currently used and potentially avoid the drawback of high viscosity of chemoembolic emulsion for TACE to improve therapeutic outcome. Anti-PLVAP Fab-TF may become a viable therapeutic agent in patients with advanced disease and compromised liver function.

Show MeSH
Related in: MedlinePlus