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Copy number gain of granulin-epithelin precursor (GEP) at chromosome 17q21 associates with overexpression in human liver cancer.

Yung MK, Lo KW, Yip CW, Chung GT, Tong CY, Cheung PF, Cheung TT, Poon RT, So S, Fan ST, Cheung ST - BMC Cancer (2015)

Bottom Line: Gain of GEP copy number was observed in 20% (12/60) HCC and associated with hepatitis B virus infection status (P = 0.015).Gain of the GEP gene copy number was observed in 20% HCC and the frequency comparable to literatures reported on the chromosome region 17q.Increased gene copy number contributed to GEP overexpression in subset of HCC.

View Article: PubMed Central - PubMed

Affiliation: Department of Surgery, The University of Hong Kong, Hong Kong, China. hermannyung@gmail.com.

ABSTRACT

Background: Granulin-epithelin precursor (GEP), a secretory growth factor, demonstrated overexpression in various human cancers, however, mechanism remain elusive. Primary liver cancer, hepatocellular carcinoma (HCC), ranks the second in cancer-related death globally. GEP controlled growth, invasion, metastasis and chemo-resistance in liver cancer. Noted that GEP gene locates at 17q21 and the region has been frequently reported to be amplified in subset of HCC. The study aims to investigate if copy number gain would associate with GEP overexpression.

Methods: Quantitative Microsatellite Analysis (QuMA) was used to quantify the GEP DNA copy number, and fluorescent in situ hybridization (FISH) was performed to consolidate the amplification status. GEP gene copy number, mRNA expression level and clinico-pathological features were analyzed.

Results: GEP DNA copy number determined by QuMA corroborated well with the FISH data, and the gene copy number correlated with the expression levels (n = 60, r = 0.331, P = 0.010). Gain of GEP copy number was observed in 20% (12/60) HCC and associated with hepatitis B virus infection status (P = 0.015). In HCC with increased GEP copy number, tight association between GEP DNA and mRNA levels were observed (n = 12, r = 0.664, P = 0.019).

Conclusions: Gain of the GEP gene copy number was observed in 20% HCC and the frequency comparable to literatures reported on the chromosome region 17q. Increased gene copy number contributed to GEP overexpression in subset of HCC.

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Related in: MedlinePlus

GEP gene copy number by FISH analysis with reference to centromere 17 (CEN17) and centromere 3 (CEN3). GEP gene (green) was detected by two flanking probes, RP11-436 J4 (left) and RP11-52 N13 (right), respectively. Control probes (red) included the centromere 17 (CEN17) and centromere 3 (CEN3). DNA copy number for each set was quantified for 100 cells and the scores (signals per cell) presented in Table 1. This case HCC801 showed CEN17 scores ranged 3.37 to 3.51, and GEP scores 3.66 to 3.80. The data indicated an increased chromosome 17 copy number at centromere and GEP locus at 17q21. Nonetheless, CEN3 scores ranged 1.97 to 2.17, indicating approximately two copies of chromosome 3 (diploid) with reference to GEP scores 3.73 by different probes flanking the gene region. GEP copy number for this case HCC801 was 3.60 by FISH analysis (reference to CEN3) compared to 2.56 by QuMA (reference to D3S1609). QuMA is a PCR-based assay method and the extend of underestimation would depend on the percentage of non-tumor cells, e.g. infiltrating lymphocytes etc., within the tumor mass. Details have also been described in Discussion.
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Fig2: GEP gene copy number by FISH analysis with reference to centromere 17 (CEN17) and centromere 3 (CEN3). GEP gene (green) was detected by two flanking probes, RP11-436 J4 (left) and RP11-52 N13 (right), respectively. Control probes (red) included the centromere 17 (CEN17) and centromere 3 (CEN3). DNA copy number for each set was quantified for 100 cells and the scores (signals per cell) presented in Table 1. This case HCC801 showed CEN17 scores ranged 3.37 to 3.51, and GEP scores 3.66 to 3.80. The data indicated an increased chromosome 17 copy number at centromere and GEP locus at 17q21. Nonetheless, CEN3 scores ranged 1.97 to 2.17, indicating approximately two copies of chromosome 3 (diploid) with reference to GEP scores 3.73 by different probes flanking the gene region. GEP copy number for this case HCC801 was 3.60 by FISH analysis (reference to CEN3) compared to 2.56 by QuMA (reference to D3S1609). QuMA is a PCR-based assay method and the extend of underestimation would depend on the percentage of non-tumor cells, e.g. infiltrating lymphocytes etc., within the tumor mass. Details have also been described in Discussion.

Mentions: To further substantiate the gene copy number by QuMA, we have examined the copy number of 17q21 region in primary HCC samples (HCC801 and HCC884) by FISH analysis. Both BAC clones (RP11-436 J4 and RP11-52 N13) flanking GEP gene demonstrated increased DNA copy number (Figure 2; Table 1). Nonetheless, the centromeric probe at chromosome 17 (pEZ17-4) also revealed increased DNA copy number per cell (Table 1). CEN17 scores ranged 2.92 to 3.51 per cell, and GEP scores ranged 3.02 to 3.80 per cell. The data indicated an increased chromosome 17 copy number, at both the centromere and GEP locus in these cases.Figure 2


Copy number gain of granulin-epithelin precursor (GEP) at chromosome 17q21 associates with overexpression in human liver cancer.

Yung MK, Lo KW, Yip CW, Chung GT, Tong CY, Cheung PF, Cheung TT, Poon RT, So S, Fan ST, Cheung ST - BMC Cancer (2015)

GEP gene copy number by FISH analysis with reference to centromere 17 (CEN17) and centromere 3 (CEN3). GEP gene (green) was detected by two flanking probes, RP11-436 J4 (left) and RP11-52 N13 (right), respectively. Control probes (red) included the centromere 17 (CEN17) and centromere 3 (CEN3). DNA copy number for each set was quantified for 100 cells and the scores (signals per cell) presented in Table 1. This case HCC801 showed CEN17 scores ranged 3.37 to 3.51, and GEP scores 3.66 to 3.80. The data indicated an increased chromosome 17 copy number at centromere and GEP locus at 17q21. Nonetheless, CEN3 scores ranged 1.97 to 2.17, indicating approximately two copies of chromosome 3 (diploid) with reference to GEP scores 3.73 by different probes flanking the gene region. GEP copy number for this case HCC801 was 3.60 by FISH analysis (reference to CEN3) compared to 2.56 by QuMA (reference to D3S1609). QuMA is a PCR-based assay method and the extend of underestimation would depend on the percentage of non-tumor cells, e.g. infiltrating lymphocytes etc., within the tumor mass. Details have also been described in Discussion.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
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getmorefigures.php?uid=PMC4403714&req=5

Fig2: GEP gene copy number by FISH analysis with reference to centromere 17 (CEN17) and centromere 3 (CEN3). GEP gene (green) was detected by two flanking probes, RP11-436 J4 (left) and RP11-52 N13 (right), respectively. Control probes (red) included the centromere 17 (CEN17) and centromere 3 (CEN3). DNA copy number for each set was quantified for 100 cells and the scores (signals per cell) presented in Table 1. This case HCC801 showed CEN17 scores ranged 3.37 to 3.51, and GEP scores 3.66 to 3.80. The data indicated an increased chromosome 17 copy number at centromere and GEP locus at 17q21. Nonetheless, CEN3 scores ranged 1.97 to 2.17, indicating approximately two copies of chromosome 3 (diploid) with reference to GEP scores 3.73 by different probes flanking the gene region. GEP copy number for this case HCC801 was 3.60 by FISH analysis (reference to CEN3) compared to 2.56 by QuMA (reference to D3S1609). QuMA is a PCR-based assay method and the extend of underestimation would depend on the percentage of non-tumor cells, e.g. infiltrating lymphocytes etc., within the tumor mass. Details have also been described in Discussion.
Mentions: To further substantiate the gene copy number by QuMA, we have examined the copy number of 17q21 region in primary HCC samples (HCC801 and HCC884) by FISH analysis. Both BAC clones (RP11-436 J4 and RP11-52 N13) flanking GEP gene demonstrated increased DNA copy number (Figure 2; Table 1). Nonetheless, the centromeric probe at chromosome 17 (pEZ17-4) also revealed increased DNA copy number per cell (Table 1). CEN17 scores ranged 2.92 to 3.51 per cell, and GEP scores ranged 3.02 to 3.80 per cell. The data indicated an increased chromosome 17 copy number, at both the centromere and GEP locus in these cases.Figure 2

Bottom Line: Gain of GEP copy number was observed in 20% (12/60) HCC and associated with hepatitis B virus infection status (P = 0.015).Gain of the GEP gene copy number was observed in 20% HCC and the frequency comparable to literatures reported on the chromosome region 17q.Increased gene copy number contributed to GEP overexpression in subset of HCC.

View Article: PubMed Central - PubMed

Affiliation: Department of Surgery, The University of Hong Kong, Hong Kong, China. hermannyung@gmail.com.

ABSTRACT

Background: Granulin-epithelin precursor (GEP), a secretory growth factor, demonstrated overexpression in various human cancers, however, mechanism remain elusive. Primary liver cancer, hepatocellular carcinoma (HCC), ranks the second in cancer-related death globally. GEP controlled growth, invasion, metastasis and chemo-resistance in liver cancer. Noted that GEP gene locates at 17q21 and the region has been frequently reported to be amplified in subset of HCC. The study aims to investigate if copy number gain would associate with GEP overexpression.

Methods: Quantitative Microsatellite Analysis (QuMA) was used to quantify the GEP DNA copy number, and fluorescent in situ hybridization (FISH) was performed to consolidate the amplification status. GEP gene copy number, mRNA expression level and clinico-pathological features were analyzed.

Results: GEP DNA copy number determined by QuMA corroborated well with the FISH data, and the gene copy number correlated with the expression levels (n = 60, r = 0.331, P = 0.010). Gain of GEP copy number was observed in 20% (12/60) HCC and associated with hepatitis B virus infection status (P = 0.015). In HCC with increased GEP copy number, tight association between GEP DNA and mRNA levels were observed (n = 12, r = 0.664, P = 0.019).

Conclusions: Gain of the GEP gene copy number was observed in 20% HCC and the frequency comparable to literatures reported on the chromosome region 17q. Increased gene copy number contributed to GEP overexpression in subset of HCC.

Show MeSH
Related in: MedlinePlus