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KIAA1114, a full-length protein encoded by the trophinin gene, is a novel surface marker for isolating tumor-initiating cells of multiple hepatocellular carcinoma subtypes.

Kim SW, Yang HG, Kang MC, Lee S, Namkoong H, Lee SW, Sung YC - Oncotarget (2014)

Bottom Line: Notably, KIAA1114 expression was strongly detected in primary hepatic tumor, but neither in the adjacent non-tumorous tissue from the same patient nor normal liver tissue.KIAA1114high cells isolated from HCC cell lines displayed TIC-like features with superior functional and phenotypic traits compared to their KIAA1114low counterparts, including tumorigenic abilities in xenotransplantation model, in vitro colony- and spheroid-forming capabilities, expression of stemness-associated genes, and migratory capacity.Our findings not only address the value of a novel antigen, KIAA1114, as a potential diagnostic factor of human liver cancer, but also as an independent biomarker for identifying TIC populations that could be broadly applied to the heterogeneous HCC subtypes.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Sciences, Pohang University of Science and Technology (POSTECH), Pohang, Gyungbuk, Republic of Korea.

ABSTRACT
Identification of novel biomarkers for tumor-initiating cells (TICs) is of critical importance for developing diagnostic and therapeutic strategies against cancers. Here we identified the role of KIAA1114, a full-length translational product of the trophinin gene, as a distinctive marker for TICs in human liver cancer by developing a DNA vaccine-induced monoclonal antibody targeting the putative extracellular domain of KIAA1114. Compared with other established markers of liver TICs, KIAA1114 was unique in that its expression was detected in both alpha fetoprotein (AFP)-positive and AFP-negative hepatocellular carcinoma (HCC) cell lines with the expression levels of KIAA1114 being positively correlated to their tumorigenic potentials. Notably, KIAA1114 expression was strongly detected in primary hepatic tumor, but neither in the adjacent non-tumorous tissue from the same patient nor normal liver tissue. KIAA1114high cells isolated from HCC cell lines displayed TIC-like features with superior functional and phenotypic traits compared to their KIAA1114low counterparts, including tumorigenic abilities in xenotransplantation model, in vitro colony- and spheroid-forming capabilities, expression of stemness-associated genes, and migratory capacity. Our findings not only address the value of a novel antigen, KIAA1114, as a potential diagnostic factor of human liver cancer, but also as an independent biomarker for identifying TIC populations that could be broadly applied to the heterogeneous HCC subtypes.

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Identification and localization of KIAA1114 with a novel anti-KIAA1114 mAb, Kiatomab(A) Schematic structures of human MAGE-D3 gene, transcript, and protein products – KIAA1114 and TROPHININ (MAGPHININ proteins are not shown). Regions recognized by Kiatomab and anti-TROPHININ mAb (3–11) are indicated. (B) Schematic diagram showing putative transmembrane topology of KIAA1114 protein. Predicted locations of TROPHININ domain, MAGE homology domain (MHD), and partial N-terminal domain encoded by DNA vaccine used for Kiatomab generation are indicated. (C) Western blot and flow cytometric analysis for recognition of human and mouse KIAA1114 by Kiatomab and 3–11 mAb using 293T cells and mouse brain tissue. β-actin was used as a loading control for Western blot. In flow cytometry, KIAA1114 expression is indicated as open histogram and isotype controls are represented as shaded histogram. (D) Analysis of KIAA1114 expression in various murine tissues by flow cytometry and Western blot. Isotype control and Kiatomab stainings are indicated as describe above for flow cytometry. Tubulin was used as an internal control for Western blot assay.
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Figure 1: Identification and localization of KIAA1114 with a novel anti-KIAA1114 mAb, Kiatomab(A) Schematic structures of human MAGE-D3 gene, transcript, and protein products – KIAA1114 and TROPHININ (MAGPHININ proteins are not shown). Regions recognized by Kiatomab and anti-TROPHININ mAb (3–11) are indicated. (B) Schematic diagram showing putative transmembrane topology of KIAA1114 protein. Predicted locations of TROPHININ domain, MAGE homology domain (MHD), and partial N-terminal domain encoded by DNA vaccine used for Kiatomab generation are indicated. (C) Western blot and flow cytometric analysis for recognition of human and mouse KIAA1114 by Kiatomab and 3–11 mAb using 293T cells and mouse brain tissue. β-actin was used as a loading control for Western blot. In flow cytometry, KIAA1114 expression is indicated as open histogram and isotype controls are represented as shaded histogram. (D) Analysis of KIAA1114 expression in various murine tissues by flow cytometry and Western blot. Isotype control and Kiatomab stainings are indicated as describe above for flow cytometry. Tubulin was used as an internal control for Western blot assay.

Mentions: The nomenclature for a full-length protein encoded by the trophinin gene has not been well-defined [11, 14], as experimental evidence for the expression of KIAA1114 in vitro or in vivo has never been provided by earlier studies. In the present study, we used the term “KIAA1114” to describe the full-length product translated from trophinin mRNA (Figure 1A). Hydropathy analysis using the topology prediction program TMpred [15] revealed that KIAA1114 is a transmembrane protein with the N-terminus outside the cell (Supplementary Figure 1). Moreover, as previously suggested [16], KIAA1114 is predicted to contain an intracellular MAGE-homology domain and a trophinin domain that traverses the plasma membrane. Although hydropathy analyses performed in the present and previous studies proposed that a trophinin domain spans the lipid bilayer multiple times [17], a recent review raised a possibility that TROPHININ is a single-pass, type II transmembrane protein that utilizes the majority of its extracellular decapeptide repeats for homophilic adhesion [18]. Accordingly, we proposed that KIAA1114 is a double-pass, type III transmembrane protein with N- and C-termini facing the outside of the cell (Figure 1B). Although thorough structural analysis is required to determine exact location of transmembrane regions, TMPred suggested that the first membrane-spanning segment lies within a MAGE-homology domain and the second one locates near the N-terminus of a trophinin domain (Supplementary Figure 1).


KIAA1114, a full-length protein encoded by the trophinin gene, is a novel surface marker for isolating tumor-initiating cells of multiple hepatocellular carcinoma subtypes.

Kim SW, Yang HG, Kang MC, Lee S, Namkoong H, Lee SW, Sung YC - Oncotarget (2014)

Identification and localization of KIAA1114 with a novel anti-KIAA1114 mAb, Kiatomab(A) Schematic structures of human MAGE-D3 gene, transcript, and protein products – KIAA1114 and TROPHININ (MAGPHININ proteins are not shown). Regions recognized by Kiatomab and anti-TROPHININ mAb (3–11) are indicated. (B) Schematic diagram showing putative transmembrane topology of KIAA1114 protein. Predicted locations of TROPHININ domain, MAGE homology domain (MHD), and partial N-terminal domain encoded by DNA vaccine used for Kiatomab generation are indicated. (C) Western blot and flow cytometric analysis for recognition of human and mouse KIAA1114 by Kiatomab and 3–11 mAb using 293T cells and mouse brain tissue. β-actin was used as a loading control for Western blot. In flow cytometry, KIAA1114 expression is indicated as open histogram and isotype controls are represented as shaded histogram. (D) Analysis of KIAA1114 expression in various murine tissues by flow cytometry and Western blot. Isotype control and Kiatomab stainings are indicated as describe above for flow cytometry. Tubulin was used as an internal control for Western blot assay.
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Related In: Results  -  Collection

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Figure 1: Identification and localization of KIAA1114 with a novel anti-KIAA1114 mAb, Kiatomab(A) Schematic structures of human MAGE-D3 gene, transcript, and protein products – KIAA1114 and TROPHININ (MAGPHININ proteins are not shown). Regions recognized by Kiatomab and anti-TROPHININ mAb (3–11) are indicated. (B) Schematic diagram showing putative transmembrane topology of KIAA1114 protein. Predicted locations of TROPHININ domain, MAGE homology domain (MHD), and partial N-terminal domain encoded by DNA vaccine used for Kiatomab generation are indicated. (C) Western blot and flow cytometric analysis for recognition of human and mouse KIAA1114 by Kiatomab and 3–11 mAb using 293T cells and mouse brain tissue. β-actin was used as a loading control for Western blot. In flow cytometry, KIAA1114 expression is indicated as open histogram and isotype controls are represented as shaded histogram. (D) Analysis of KIAA1114 expression in various murine tissues by flow cytometry and Western blot. Isotype control and Kiatomab stainings are indicated as describe above for flow cytometry. Tubulin was used as an internal control for Western blot assay.
Mentions: The nomenclature for a full-length protein encoded by the trophinin gene has not been well-defined [11, 14], as experimental evidence for the expression of KIAA1114 in vitro or in vivo has never been provided by earlier studies. In the present study, we used the term “KIAA1114” to describe the full-length product translated from trophinin mRNA (Figure 1A). Hydropathy analysis using the topology prediction program TMpred [15] revealed that KIAA1114 is a transmembrane protein with the N-terminus outside the cell (Supplementary Figure 1). Moreover, as previously suggested [16], KIAA1114 is predicted to contain an intracellular MAGE-homology domain and a trophinin domain that traverses the plasma membrane. Although hydropathy analyses performed in the present and previous studies proposed that a trophinin domain spans the lipid bilayer multiple times [17], a recent review raised a possibility that TROPHININ is a single-pass, type II transmembrane protein that utilizes the majority of its extracellular decapeptide repeats for homophilic adhesion [18]. Accordingly, we proposed that KIAA1114 is a double-pass, type III transmembrane protein with N- and C-termini facing the outside of the cell (Figure 1B). Although thorough structural analysis is required to determine exact location of transmembrane regions, TMPred suggested that the first membrane-spanning segment lies within a MAGE-homology domain and the second one locates near the N-terminus of a trophinin domain (Supplementary Figure 1).

Bottom Line: Notably, KIAA1114 expression was strongly detected in primary hepatic tumor, but neither in the adjacent non-tumorous tissue from the same patient nor normal liver tissue.KIAA1114high cells isolated from HCC cell lines displayed TIC-like features with superior functional and phenotypic traits compared to their KIAA1114low counterparts, including tumorigenic abilities in xenotransplantation model, in vitro colony- and spheroid-forming capabilities, expression of stemness-associated genes, and migratory capacity.Our findings not only address the value of a novel antigen, KIAA1114, as a potential diagnostic factor of human liver cancer, but also as an independent biomarker for identifying TIC populations that could be broadly applied to the heterogeneous HCC subtypes.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Sciences, Pohang University of Science and Technology (POSTECH), Pohang, Gyungbuk, Republic of Korea.

ABSTRACT
Identification of novel biomarkers for tumor-initiating cells (TICs) is of critical importance for developing diagnostic and therapeutic strategies against cancers. Here we identified the role of KIAA1114, a full-length translational product of the trophinin gene, as a distinctive marker for TICs in human liver cancer by developing a DNA vaccine-induced monoclonal antibody targeting the putative extracellular domain of KIAA1114. Compared with other established markers of liver TICs, KIAA1114 was unique in that its expression was detected in both alpha fetoprotein (AFP)-positive and AFP-negative hepatocellular carcinoma (HCC) cell lines with the expression levels of KIAA1114 being positively correlated to their tumorigenic potentials. Notably, KIAA1114 expression was strongly detected in primary hepatic tumor, but neither in the adjacent non-tumorous tissue from the same patient nor normal liver tissue. KIAA1114high cells isolated from HCC cell lines displayed TIC-like features with superior functional and phenotypic traits compared to their KIAA1114low counterparts, including tumorigenic abilities in xenotransplantation model, in vitro colony- and spheroid-forming capabilities, expression of stemness-associated genes, and migratory capacity. Our findings not only address the value of a novel antigen, KIAA1114, as a potential diagnostic factor of human liver cancer, but also as an independent biomarker for identifying TIC populations that could be broadly applied to the heterogeneous HCC subtypes.

Show MeSH
Related in: MedlinePlus