Quantitative proteomics identification of phosphoglycerate mutase 1 as a novel therapeutic target in hepatocellular carcinoma.
BACKGROUND
Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide with poor prognosis due to resistance to conventional chemotherapy and limited efficacy of radiotherapy. There is an urgent need to develop novel biomarkers for early diagnosis, as well as to identify new drug targets for therapeutic interventions.


METHODS
54 paired HCC samples and 21 normal liver tissues were obtained from West China Hospital of Sichuan University. Informed consent was obtained from all the patients or their relatives prior to analysis, and the project was approved by the Institutional Ethics Committee of Sichuan University. Stable Isotope Labeling with Amino Acids in Cell Culture (SILAC)-based proteomics was employed to profile the differentially expressed proteins between a HepG2 human hepatoma cell line and an immortal hepatic cell line L02. Validation of PGAM1 expression was performed by semi-quantitative RT-PCR, immunoblot and immunohistochemistry using clinical samples. shRNA expressing plasmids specifically targeting PGAM1 were designed and constructed by GenePharma Corporation (Shanghai, China), and were utilized to silence expression of PGAM1 in vitro and in vivo. Cell proliferation was measured by a combination of colony formation assay and Ki67 staining. Apoptosis was examined by flow cytometry and TUNEL assay.


RESULTS
A total of 63 dysregulated proteins were identified, including 51 up-regulated proteins, and 12 down-regulated proteins (over 2-fold, p < 0.01). Phosphoglycerate mutase 1 (PGAM1) was found markedly upregulated. Clinico-pathological analysis indicated that overexpression of PGAM1 was associated with 66.7% HCC, and strongly correlated with poor differentiation and decreased survival rates (p < 0.01). shRNAs-mediated repression of PGAM1 expression resulted in significant inhibition in liver cancer cell growth both in vitro and in vivo.


CONCLUSIONS
Our studies suggested that PGAM1 plays an important role in hepatocarcinogenesis, and should be a potential diagnostic biomarker, as well as an attractive therapeutic target for hepatocellular carcinoma.
Ren
Fenglian
F
The State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China.
Wu
Hong
H
Lei
Yunlong
Y
Zhang
Haiyuan
H
Liu
Rui
R
Zhao
Yong
Y
Chen
Xiancheng
X
Zeng
Dequan
D
Tong
Aiping
A
Chen
Lijuan
L
Wei
Yuquan
Y
Huang
Canhua
C
D000328:Adult; D000818:Animals; D014408:Biomarkers, Tumor / Q000032:analysis*; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D002469:Cell Separation; D005260:Female; D005434:Flow Cytometry; D020869:Gene Expression Profiling; D006801:Humans; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling; D008113:Liver Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D051379:Mice; D008819:Mice, Nude; D008875:Middle Aged; D009367:Neoplasm Staging; D010736:Phosphoglycerate Mutase / Q000096:biosynthesis / Q000235:genetics*; D040901:Proteomics / Q000379:methods; D020133:Reverse Transcriptase Polymerase Chain Reaction; D023041:Xenograft Model Antitumor Assays
D014408:Biomarkers, Tumor; D010736:Phosphoglycerate Mutase
J Hepatol. 2009 May;50(5):1019-28
19303656
Br J Cancer. 2009 May 5;100(9):1369-72
19352381
Gastroenterology. 2008 May;134(6):1752-63
18471552
Hepatology. 2007 Jul;46(1):200-8
17596871
J Bioenerg Biomembr. 2007 Jun;39(3):251-7
17624581
Cell. 2003 Nov 26;115(5):577-90
14651849
Oncogene. 2006 Aug 7;25(34):4633-46
16892078
Proc Natl Acad Sci U S A. 2003 May 27;100(11):6343-6
12730368
Cancer Res. 2005 Jan 1;65(1):177-85
15665293
Protein Sci. 2001 Sep;10(9):1835-46
11514674
J Biol Chem. 1988 Nov 15;263(32):16899-905
2846553
Int J Cancer. 2008 Nov 15;123(10):2377-83
18729184
Mol Cell Proteomics. 2009 Jan;8(1):70-85
18723843
Dig Surg. 2007;24(5):342-8
17785977
Nat Rev Cancer. 2004 Nov;4(11):891-9
15516961
Oncogene. 2006 Jun 26;25(27):3866-84
16799628
Proc Natl Acad Sci U S A. 2003 Nov 11;100(23):13537-42
14573703
Gene. 2001 Feb 21;264(2):273-9
11250083
Proteomics. 2008 May;8(10):2012-23
18491315
Biochim Biophys Acta. 2005 Jun 10;1740(3):403-10
15949708
Hepatology. 2008 Mar;47(3):897-907
18176954
Int J Cancer. 2001 Dec 1;94(5):662-8
11745460
FEBS Lett. 1999 Jul 23;455(3):344-8
10437801
Int J Oncol. 2008 Oct;33(4):725-31
18813785
Cancer Lett. 2009 Apr 18;276(2):125-35
18790562
Nat Biotechnol. 2005 Oct;23(10):1303-7
16200062
Electrophoresis. 2006 Apr;27(7):1417-23
16568407
J Biomed Sci. 2006 Jul;13(4):453-68
16633742
Biochem J. 1990 Jan 15;265(2):503-9
2302181
Cancer Detect Prev. 2004;28(6):443-52
15582268
Front Biosci (Landmark Ed). 2009 Jan 01;14:19-44
19273052
Liver Int. 2009 Jan;29(1):10-7
19120940
Cell. 2000 Jan 7;100(1):57-70
10647931
Gastroenterology. 2007 Jun;132(7):2557-76
17570226
J Biol Chem. 2008 Aug 15;283(33):22700-8
18541534
Mol Biol Evol. 2002 May;19(5):654-63
11961099
Expert Rev Proteomics. 2005 Aug;2(4):589-601
16097891
Curr Drug Targets. 2004 May;5(4):375-82
15134220
Carcinogenesis. 2004 Jun;25(6):865-71
14764456
J Biol Chem. 2004 Aug 20;279(34):35803-12
15181008
Cancer Immunol Immunother. 1995 Feb;40(2):73-8
7882385
Proteomics. 2006 Nov;6(22):6095-106
17051635
Br J Cancer. 2000 Jan;82(1):20-7
10638961
Horm Res. 2004;62(4):191-6
15375329
Trends Biochem Sci. 1999 Feb;24(2):68-72
10098401
Proteomics. 2006 Jan;6(2):547-58
16342241
Mol Cancer Ther. 2003 Jun;2(6):573-80
12813137
Toxicol Sci. 2008 Oct;105(2):286-94
18567617
Gastroenterology. 2004 Nov;127(5 Suppl 1):S5-S16
15508102
FEBS J. 2008 Jul;275(13):3454-69
18510554
Oncology. 2009;77(6):385-94
20130421
Cell Biol Int. 2008 Oct;32(10):1215-22
18675922
Cancer Lett. 2001 Nov 8;173(1):83-91
11578813
J Exp Med. 2000 May 15;191(10):1675-86
10811861
Science. 1956 Feb 24;123(3191):309-14
13298683