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NANOG promotes cancer stem cell characteristics and prostate cancer resistance to androgen deprivation.

Jeter CR, Liu B, Liu X, Chen X, Liu C, Calhoun-Davis T, Repass J, Zaehres H, Shen JJ, Tang DG - Oncogene (2011)

Bottom Line: We have recently reported that short-hairpin RNA-mediated knockdown of the embryonic stem cell (ESC) self-renewal gene NANOG significantly reduced the clonogenic and tumorigenic capabilities of various cancer cells.These pro-tumorigenic effects of NANOG were associated with key molecular changes, including an upregulation of molecules such as CXCR4, IGFBP5, CD133 and ALDH1.The present gain-of-function studies, coupled with our recent loss-of-function work, establish the integral role for NANOG in neoplastic processes and shed light on its mechanisms of action.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Carcinogenesis, University of Texas MD Anderson Cancer Center, Science Park-Research Division, Smithville, USA. cjeter@mdanderson.org

ABSTRACT
Cancer cell molecular mimicry of stem cells (SC) imbues neoplastic cells with enhanced proliferative and renewal capacities. In support, numerous mediators of SC self-renewal have been evinced to show oncogenic potential. We have recently reported that short-hairpin RNA-mediated knockdown of the embryonic stem cell (ESC) self-renewal gene NANOG significantly reduced the clonogenic and tumorigenic capabilities of various cancer cells. In this study, we sought to test the potential pro-tumorigenic functions of NANOG, particularly, in prostate cancer (PCa). Using qRT-PCR, we first confirmed that PCa cells expressed NANOG mRNA primarily from the NANOGP8 locus on chromosome 15q14. We then constructed a lentiviral promoter reporter in which the -3.8-kb NANOGP8 genomic fragment was used to drive the expression of green fluorescence protein (GFP). We observed that NANOGP8-GFP(+) PCa cells showed cancer stem cell (CSC) characteristics such as enhanced clonal growth and tumor regenerative capacity. To further investigate the functions and mechanisms of NANOG in tumorigenesis, we established tetracycline-inducible NANOG-overexpressing cancer cell lines, including both PCa (Du145 and LNCaP) and breast (MCF-7) cancer cells. NANOG induction promoted drug resistance in MCF-7 cells, tumor regeneration in Du145 cells and, most importantly, castration-resistant tumor development in LNCaP cells. These pro-tumorigenic effects of NANOG were associated with key molecular changes, including an upregulation of molecules such as CXCR4, IGFBP5, CD133 and ALDH1. The present gain-of-function studies, coupled with our recent loss-of-function work, establish the integral role for NANOG in neoplastic processes and shed light on its mechanisms of action.

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Related in: MedlinePlus

NANOG genomic loci and NANOG mRNA expression in PCa cellsa) Schematic of NANOG1 and NANOGP8 gene structures. Chr, chromosome; E, exon; UTR, untranslated region. The 22-bp region unique to NANOG1 (vertical bar) was used to design primers/probes for NANOG1 and NANOGP8 specific PCR. b) Quantitative PCR (qPCR) detection of NANOGP8 mRNA expression in various cancer cells, normalized to GAPDH. The normalized NANOGP8 mRNA levels in LNCaP cells were set at 1 and NANOGP8 mRNA levels in breast (MCF7), colon (Colo320) and PCa cell lines (LNCaP, Du145 and PC3) and xenografts (LAPC-4 and LAPC-9) were presented relative to LNCaP. c)NANOGP8 mRNA levels in a cohort of primary patient tumors (HPCa) as determined by qPCR. HPCa56P1 and HPCa101P0 are two early-passage (P) xenografts established in our lab.
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Figure 1: NANOG genomic loci and NANOG mRNA expression in PCa cellsa) Schematic of NANOG1 and NANOGP8 gene structures. Chr, chromosome; E, exon; UTR, untranslated region. The 22-bp region unique to NANOG1 (vertical bar) was used to design primers/probes for NANOG1 and NANOGP8 specific PCR. b) Quantitative PCR (qPCR) detection of NANOGP8 mRNA expression in various cancer cells, normalized to GAPDH. The normalized NANOGP8 mRNA levels in LNCaP cells were set at 1 and NANOGP8 mRNA levels in breast (MCF7), colon (Colo320) and PCa cell lines (LNCaP, Du145 and PC3) and xenografts (LAPC-4 and LAPC-9) were presented relative to LNCaP. c)NANOGP8 mRNA levels in a cohort of primary patient tumors (HPCa) as determined by qPCR. HPCa56P1 and HPCa101P0 are two early-passage (P) xenografts established in our lab.

Mentions: Our first aim in the present study was to determine whether cancer cells expressing endogenous NANOG are intrinsically unique compared to their non-expressing (or low-expressing) counterparts. To this end, we conceived a promoter reporter strategy to track and isolate cells differentially expressing NANOG. However, one complication is that at least two highly similar NANOG mRNA species have been reported originating from two distinct loci (Figure 1a), including the NANOG1, located on chromosome 12p13, in ESCs, and its retrogene NANOGP8, located on chromosome 15q14 (Booth and Holland, 2004). Given their remarkable similarity (highly conserved over the entire length of the processed transcripts with only 6 nucleotide substitutions in the coding regions and 99% identical at the amino acid level) (Jeter et al., 2009), theoretically, NANOG1 and/or NANOGP8 could account for the presence of NANOG mRNA in cancer cells. We previously provided preliminary evidence that NANOGP8 was the predominant source of NANOG mRNA in epithelial cancer cells, including PCa (Jeter et al., 2009); however, due to methodological limitations we could not eliminate NANOG1 as an important contributor. Thus, we set out to further investigate the origin of NANOG mRNA using quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis.


NANOG promotes cancer stem cell characteristics and prostate cancer resistance to androgen deprivation.

Jeter CR, Liu B, Liu X, Chen X, Liu C, Calhoun-Davis T, Repass J, Zaehres H, Shen JJ, Tang DG - Oncogene (2011)

NANOG genomic loci and NANOG mRNA expression in PCa cellsa) Schematic of NANOG1 and NANOGP8 gene structures. Chr, chromosome; E, exon; UTR, untranslated region. The 22-bp region unique to NANOG1 (vertical bar) was used to design primers/probes for NANOG1 and NANOGP8 specific PCR. b) Quantitative PCR (qPCR) detection of NANOGP8 mRNA expression in various cancer cells, normalized to GAPDH. The normalized NANOGP8 mRNA levels in LNCaP cells were set at 1 and NANOGP8 mRNA levels in breast (MCF7), colon (Colo320) and PCa cell lines (LNCaP, Du145 and PC3) and xenografts (LAPC-4 and LAPC-9) were presented relative to LNCaP. c)NANOGP8 mRNA levels in a cohort of primary patient tumors (HPCa) as determined by qPCR. HPCa56P1 and HPCa101P0 are two early-passage (P) xenografts established in our lab.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3140601&req=5

Figure 1: NANOG genomic loci and NANOG mRNA expression in PCa cellsa) Schematic of NANOG1 and NANOGP8 gene structures. Chr, chromosome; E, exon; UTR, untranslated region. The 22-bp region unique to NANOG1 (vertical bar) was used to design primers/probes for NANOG1 and NANOGP8 specific PCR. b) Quantitative PCR (qPCR) detection of NANOGP8 mRNA expression in various cancer cells, normalized to GAPDH. The normalized NANOGP8 mRNA levels in LNCaP cells were set at 1 and NANOGP8 mRNA levels in breast (MCF7), colon (Colo320) and PCa cell lines (LNCaP, Du145 and PC3) and xenografts (LAPC-4 and LAPC-9) were presented relative to LNCaP. c)NANOGP8 mRNA levels in a cohort of primary patient tumors (HPCa) as determined by qPCR. HPCa56P1 and HPCa101P0 are two early-passage (P) xenografts established in our lab.
Mentions: Our first aim in the present study was to determine whether cancer cells expressing endogenous NANOG are intrinsically unique compared to their non-expressing (or low-expressing) counterparts. To this end, we conceived a promoter reporter strategy to track and isolate cells differentially expressing NANOG. However, one complication is that at least two highly similar NANOG mRNA species have been reported originating from two distinct loci (Figure 1a), including the NANOG1, located on chromosome 12p13, in ESCs, and its retrogene NANOGP8, located on chromosome 15q14 (Booth and Holland, 2004). Given their remarkable similarity (highly conserved over the entire length of the processed transcripts with only 6 nucleotide substitutions in the coding regions and 99% identical at the amino acid level) (Jeter et al., 2009), theoretically, NANOG1 and/or NANOGP8 could account for the presence of NANOG mRNA in cancer cells. We previously provided preliminary evidence that NANOGP8 was the predominant source of NANOG mRNA in epithelial cancer cells, including PCa (Jeter et al., 2009); however, due to methodological limitations we could not eliminate NANOG1 as an important contributor. Thus, we set out to further investigate the origin of NANOG mRNA using quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis.

Bottom Line: We have recently reported that short-hairpin RNA-mediated knockdown of the embryonic stem cell (ESC) self-renewal gene NANOG significantly reduced the clonogenic and tumorigenic capabilities of various cancer cells.These pro-tumorigenic effects of NANOG were associated with key molecular changes, including an upregulation of molecules such as CXCR4, IGFBP5, CD133 and ALDH1.The present gain-of-function studies, coupled with our recent loss-of-function work, establish the integral role for NANOG in neoplastic processes and shed light on its mechanisms of action.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Carcinogenesis, University of Texas MD Anderson Cancer Center, Science Park-Research Division, Smithville, USA. cjeter@mdanderson.org

ABSTRACT
Cancer cell molecular mimicry of stem cells (SC) imbues neoplastic cells with enhanced proliferative and renewal capacities. In support, numerous mediators of SC self-renewal have been evinced to show oncogenic potential. We have recently reported that short-hairpin RNA-mediated knockdown of the embryonic stem cell (ESC) self-renewal gene NANOG significantly reduced the clonogenic and tumorigenic capabilities of various cancer cells. In this study, we sought to test the potential pro-tumorigenic functions of NANOG, particularly, in prostate cancer (PCa). Using qRT-PCR, we first confirmed that PCa cells expressed NANOG mRNA primarily from the NANOGP8 locus on chromosome 15q14. We then constructed a lentiviral promoter reporter in which the -3.8-kb NANOGP8 genomic fragment was used to drive the expression of green fluorescence protein (GFP). We observed that NANOGP8-GFP(+) PCa cells showed cancer stem cell (CSC) characteristics such as enhanced clonal growth and tumor regenerative capacity. To further investigate the functions and mechanisms of NANOG in tumorigenesis, we established tetracycline-inducible NANOG-overexpressing cancer cell lines, including both PCa (Du145 and LNCaP) and breast (MCF-7) cancer cells. NANOG induction promoted drug resistance in MCF-7 cells, tumor regeneration in Du145 cells and, most importantly, castration-resistant tumor development in LNCaP cells. These pro-tumorigenic effects of NANOG were associated with key molecular changes, including an upregulation of molecules such as CXCR4, IGFBP5, CD133 and ALDH1. The present gain-of-function studies, coupled with our recent loss-of-function work, establish the integral role for NANOG in neoplastic processes and shed light on its mechanisms of action.

Show MeSH
Related in: MedlinePlus