Open Access Peer-reviewed

Glycoregulatory Enzymes as Early Diagnostic Markers during Premalignant Stage in Hepatocellular Carcinoma

NM Abdel-Hamid1,, MF Ramadan2, SW Amgad2

1Departments of Biochemistry, Colleges of Pharmacy, Kafrelsheikh University, Kafrelsheikh

2Departments of Biochemistry, Colleges of Pharmacy, Minia University, Minia, Egypt

American Journal of Cancer Prevention. 2013, 1(2), 14-19. DOI: 10.12691/ajcp-1-2-1
Published online: August 25, 2017


Hepatocellular carcinoma (HCC) is the third leading cause of death and fifth most common malignancy worldwide. Objective: Present study focused on the abnormal tumor cell glucose metabolism, considering the pathways of hexose monophosphate (HMP) shunt enzymes. The key regulatory enzymes of HMP include hexokinase (HK), glyeraldehyde-3-phosphate dehydrogenase (GAPDH) and glucose-6-phosphate dehydrogenase (G6PD). Their perturbations sought to be helpful in the diagnosis and prognosis of HCC in addition to alpha-fetoprotein (AFP). Materials and methods: Diethyl nitrosamine (DENA) plus carbon tetra chloride (CCl4) chemically-induced HCC model was used. Sixteen male albino rats were equally divided into 2 groups. Group I: served as a normal control received single intraperitonial (I.P) injection of saline, 2 weeks later, received subcutaneous (S.C) injection of saline, in equal volumes given for group II animals. Group II animals received single I.P injection of DENA (200mg/kg), 2 weeks later, received S.C injection of CCl4 (3ml/Kg/week) for 6 weeks. Then animals were sacrificed, blood and liver samples were obtained. Results: In HCC group, relative liver weights, serum AFP and HK, GAPDH and G6PD activities in both serum and liver homogenate were significantly increased; subsequent decrease in body weight was also evident. The histopathological examination of liver biopsies revealed the presence of few dysplastic nodules. Such nodules were 1mm or more in diameter on macroscopic examination, indicating carcinogenic features, nuclear and cytoplasmic alterations with clustering of population cells, structurally abnormal portal tracts, supporting serum enzyme and tumor marker assays. Conclusion: Glycolytic alterations can be used for diagnosis and prognosis of carcinogenesis in liver. These biomarkers may be beneficial tools to improve diagnostic performance of conventional tumor markers as AFP.


alpha fetoprotein, diagnosis, hepatocellular carcinoma, glycoregulatory enzymes
[1]  Llovet JM (2005) Updated treatment approach to hepatocellular carcinoma. J Gastroenterol 40, 225-235.View Article  PubMed
[2]  El-Serag HB & Rudolph KL (2007) Hepatocellular carcinoma: epidemiology and molecular carcinogenesis. Gastroenterology 132, 2557-2576.View Article  PubMed
[3]  Llovet JM & Bruix J. (2003) Systematic Review of Randomized Trials for Unresectable Hepatocellular Carcinoma: Chemoembolization Improves Survival. Hepatol 37, 429-442.View Article  PubMed
[4]  Abdel-Hamid NM (2009) Recent insights on risk factors of hepatocellular carcinoma. World J Hepatol 1, 3-7.View Article  PubMed
[5]  Bosch FX, Ribes J & Diaz M CR (2004) Primary liver cancer: worldwide incidence and trends. . Gastroenterology 127, 5-16.View Article
[6]  Lai EC & WY. L (2005) The continuing challenge of hepatic cance rin Asia. Surgeon 3, 210-215.View Article
[7]  Lau W & EC L (2008) Hepatocellular carcinoma: current management and recent advances. Hepatobiliary Pancreat Dis Int 7, 237-257. PubMed
[8]  Brown RS & Wahl RL (1993) Overexpression of Glut-1 glucose transporter in human breast cancer. Cancer 72 2979-2985.View Article
[9]  Guillemin K & Krasnow MA (1997) The hypoxic response: huffing and HIFing. Cell 89, 9-12.View Article
[10]  Wang GL, Jiang BH, EA R & and Semenza GL (1995) Hypoxia-inducible factor 1 is a basic-helix-loop-helix-PAS heterodimer regulated by cellular O2 tension. Proc Natl Acad Sci USA 92, 5510-5514.View Article  PubMed
[11]  Beckner ME, Stracke ML, LA L & and Schiffmann E (1990) Glycolysis as primary energy source in tumor cell chemotaxis. J Natl Cancer Inst 82, 1836-1840.View Article  PubMed
[12]  Greiner EF, M G & Brand K (1994) Glucose is essential for proliferation and the glycolytic enzyme induction that provokes a transition to glycolytic energy production. J Biol Chem 269, 31484-31490. PubMed
[13]  IARC. (2000) N-nitrosodiethanolamine. IARC Monogr Eval Carcinog Risks Hum. 77, 403-438. PubMed
[14]  Loeppky RN (1999) The mechanism of bioactivation of N-nitrosodiethanolamine. Drug Metab Rev 31, 175-193.View Article  PubMed
[15]  Abdel-Hamid NM, Nazmy MH & Abdel-Bakey AI (2011) Polyol profile as an early diagnostic and prognostic marker in natural product chemoprevention of hepatocellular carcinoma in diabetic rats. Diabetes Res Clin Pract.View Article  PubMed
[16]  Parimala R & Sachdanandam P (1993) Effect of Plumbagin on some glucose matabolising enzymes studied in rats in experimental hepatoma. Mol Cell Biochem 125, 59-63.View Article  PubMed
[17]  Harris RA (2004) Glycolysis, Overview. Encyclopedia of Biological Chemistry 2, 266-271.
[18]  Shaw RJ (2006) Glucose metabolism and cancer. Current Opinion in Cell Biology 18, 598-608.View Article  PubMed
[19]  Zhivotovsky B & Orrenius S (2009) TheWarburg Effect returns to the cancer stage. Seminars in Cancer Biology 19, 1-3.View Article  PubMed
[20]  Permalatha B, V S & Sachdanandam (1997) Modulating effect of Semecarpus Anacardium L. nut extract on glucose metabolizing enzymes in aflatoxin B1-induced experimental Hepatocellular carcinoma. Pharmacological Research 36, 187-192.View Article  PubMed
[21]  Sivanesan D & Begum VH (2007) Modulating effect of Gynandropsis gynandra L on glucose metabolizing enzymes in aflatoxin B1-induced experimental Hepatocellular carcinoma in rats. Indian Journal of Biochemistry & Biophysics 44, 477-480. PubMed
[22]  Yuen M & C L (2005) Serological markers of liver cancer. Best Practice & Research Clinical Gastroenterology 19, 91-99.View Article  PubMed
[23]  Dakshayani KB, Subramanian P, Manivasagam T & Essa MM MS (2005) Melatonin modulates the oxidant–antioxidant imbalance during N-nitrosodiethylamine induced hepatocarcinogenesis in rats. J Pharm Sci 8, 316-321.
[24]  Singha BN, Braj R. Singhb, B.K. Sarmaa & H.B. Singha (2009) Potential chemoprevention of N-nitrosodiethylamine-induced hepatocarcinogenesis by polyphenolics from Acacia nilotica bark. Chemico-Biological Interactions 181, 20-28.View Article  PubMed
[25]  Subramanian P, 1 SM, Kadiyala Babu Dakshayani1 & Seithikurippu R. Pandi-Perumal2 ITaDPC (2007) Prevention by melatonin of hepatocarcinogenesis in rats injected with N-nitrosodiethylamine. J Pineal Res 43, 305-312.View Article  PubMed
[26]  Bergmeyer HU, Grassl M & HE W (1983) in Methods of Enzymatic Analysis (Bergmeyer, H.U. ed) 3rd ed., Verlag Chemie, Deerfield. In, pp. 222-223.
[27]  Velick SF (1955) Methods Enzymol. In, pp. 401-406.
[28]  kornberg A (1955) Methods in Enzymology. Acedemic Press, New york, 232.
[29]  Gornall AG, Bardawill CJ & MM D (1949) Determination of serum proteins by means of the biuret reaction. J Biol Chem 177, 751-766. PubMed
[30]  Bergmeyer HU, Grassl, M., and Walter, H.E. (1983) Methods of Enzymatic Analysis Vol. Volume II, Verlag Chemie,, Deerfield Beach, FL.
[31]  Llovet JM (2005) Updated treatment approach to hepatocellular carcinoma. J Gastroenterol 40, 225-235.View Article  PubMed
[32]  Davis GL, Dempster J, Meler JD, Orr DW, Walberg MW, Brown B, Berger BD, O'Connor JK & Goldstein RM (2008) Hepatocellular carcinoma: management of an increasingly common problem. Proc (Bayl Univ Med Cent) 21, 266-280.
[33]  Abdel-Hamid M, El-Daly M, Molnegren V, El-Kafrawy S, Abdel-Latif S, Esmat G, Strickland GT, Loffredo C, Albert J & Widell A (2007) Genetic diversity in hepatitis C virus in Egypt and possible association with hepatocellular carcinoma. J Gen Virol 88, 1526-1531.View Article  PubMed
[34]  Anwar WA, Khaled HM, Amra HA, H E-N & CA L (2008) Changing pattern of hepatocellular carcinoma (HCC) and its risk factors in Egypt: Possibilities for prevention. Mutation Research 659, 176-184.View Article  PubMed
[35]  Al-Rejaie SS, Aleisa AM, Al-Yahya AA, Bakheet SA, Alsheikh A, Fatani AG, Al-Shabanah OA & MM A (2009) Progression of diethylnitrosamine-induced hepatic carcinogenesis in carnitine-depleted rats. World J Gastroenterol 15, 1373-1380.View Article  PubMed
[36]  Sundaresan S & P S (2003) S- allycysteine inhibits circulatory lipid peroxidation and promotes antioxidants in N-nitrosodiethylamine-induced carcinogenesis. polJ Pharmacol 55, 37-42. PubMed
[37]  Theise ND (1995) macroregenerative (dysplastic) nodules and hepatocarcinogenesis; theoretical and clinical consideration. Semin liver Dis 15, 360-371.View Article  PubMed
[38]  Kondo Y (1997) pathology of early hepatocellular carcinoma and preneoplastic lesion in the liver. In: Okudak, Tabor E (eds.); liver cancer. New York, churchill Livingstone. 135-153.
[39]  Lopez J (2005) Recent Developments in the first detection of Hepatocellular carcinoma Clin Biochem 26, 65-75.
[40]  Sivaramakrishnan V, Shilpa PN, Praveen Kumar VR & Niranjali Devaraj S (2008) Attenuation of N-nitrosodiethylamine-induced hepatocellular carcinogenesis by a novel favonol Morin. Chem Biol Interact 171, 79-88.View Article  PubMed
[41]  Xu R, Pelicano H Z, hou Y CJ & S FL, Bhalla KN, Keating MJ and Huang P, (): "I". ;: (2005) Inhibition of Glycolysis in Cancer Cells: A Novel Strategy to Overcome Drug Resistance Associated with Mitochondrial Respiratory Defect and Hypoxia. Cancer Res 65, 613-621. PubMed
[42]  Schafer D, B H-K & Brand K (1997) Glucose regulates the promoter activity of aldolase A and pyruvate kinase M2 via dephosphorylation of Sp1. FEBS Lett 417, 325-328.View Article
[43]  Izuishi K, Kato K, Ogura T & Kinoshita T and Esumi H (2000) Remarkable tolerance of tumor cells to nutrient deprivation: possible new biochemical target for cancer therapy. Cancer Res 60, 6201-6207. PubMed
[44]  Kato K, Ogura T, Kishimoto A & Minegishi Y NN, Miyazaki M and Esumi H, (): "".;(2000) Critical roles of AMP-activated protein kinase in constitutive tolerance of cancer cells to nutrient deprivation and tumor formation. Oncogene 21, 6082-6090.View Article  PubMed
[45]  King CP (2004) Molecular imaging. Hepatocellular carcinoma screening, diagnosis, and management. National Institutes of Health, Bethesda, Maryland, 109-112.
[46]  Dang CV & Semenza GL (1999) Oncogenic alterations of metabolism. Trends Biochem Sci 24, 68-72.View Article
[47]  Selak MA, Armour SM, MacKenzie ED & . ea (2005) Succinate links TCA cycle dysfunction to oncogenesis by inhibiting HIF-alpha prolyl hydroxylase. Cancer Cell 7, 77-85.View Article  PubMed
[48]  Katzen HM & Schimke RT (1965) Multiple forms of hexokinase in the rat: Tissue distribution, age dependency, and properties. Proc NatlAcad Sci USA 54, 1218-1225.View Article  PubMed
[49]  Kuang Y, Schomisch SJ, V C & and Lee Z, (2006) Hexokinase and glucose-6-phosphatase activity in woodchuck model of hepatitis virus-induced hepatocellular carcinoma. Comparative Biochemistry and Physiology, Part C 143, 225-231.
[50]  Mathupala SP, Rempel A & Pedersen PL (1997) Aberrant glycolytic metabolism of cancer cells: a remarkable coordination of genetic, transcriptional, post-translational, and mutational events that lead to a critical role for type II hexokinase. J Bioenerg Biomembr 29, 339-343.View Article  PubMed
[51]  Torizuka T, Tamaki N, Inokuma T & Magata Y SS, Yonekura Y, Tanaka A, Yamaoka Y, Yamamoto K and Konishi J, (): ".1811-1817 (1995) In vivo assessment of glucose metabolism in hepatocellular carcinoma with FDG-PET. J Nucl Med 36, 1811-1817. PubMed
[52]  Mazurek S, Eigenbrodt E, K F & and Steinberg P (1999) Alterations in the glycolytic and glutaminolytic pathways after malignant transformation of rat liver oval cells. J Cell Physiol 181, 136-146.View Article
[53]  Nagashima RA, Paggi MG, LJ S & and Pedersen PL (1988) Purification characterization of a bindable form of mitochondrial bound hexokinase from the highly glycolytic AS-30D rat hepatoma cell line. Cancer Res 48, 913-919.
[54]  Katabi MM, Chan HL, SE K & and Batist G (1999) Hexokinase type II: a novel tumor-specific promoter for gene-targeted therapy differentially expressed and regulated in human cancer cells. Hum Gene Ther 10, 155-164.View Article  PubMed
[55]  Kuzminskaya EV, Asryants RA & Nagradova NK (1993) SH containing compounds as allosteric effectors of glyceraldehyde-3-phosphate dehydrogenase. FEBS Lett 336, 208-210.View Article
[56]  Shalova IN, Cechalova K, and RZ & Dimitrova P (2007) Decrease of dehydrogenase activity of cerebral glyceraldehyde-3-phosphate dehydrogenase in different animal models of Alzheimer's disease. Biochimica et Biophysica Acta 1770, 826-832.View Article  PubMed
[57]  Sirover MA (1997) Role of the Glycolytic Protein, Glyceraldehyde-3-Phosphate Dehydrogenase, in Normal Cell Function and in Cell Pathology. Journal of Cellular Biochemistry 66, 133-140.View Article
[58]  Sirover MA (1999) New insights into an old protein: the functional diversity of mammalian glyceraldehyde-3-phosphate dehydrogenase. Biochim Biophys Acta 1432, 159-184.View Article
[59]  Shibuya A & Ikewaki N (2002) High serum glyceraldehyde-3-phosphate dehydrogenase levels in patients with liver cirrhosis. Hepatology Research 22, 174-179.View Article
[60]  Gordon GB, MC M & Levy HR (1995) On the mechanism of interaction of steroids with human glucose-6-phosphate dehydrogenase. Arch Biochem Biophys 318, 25-29.View Article  PubMed
[61]  Tian W, Braustein LD, Apse K PJ & RM, Tian X and Staton RC,: (1999) Importance of glucose-6-phosphate dehydrogenase activity in cell death. Am J Physiol Cell Physiol 276, 1121-1131.
[62]  Coussens LM & Werb Z (2002) Inflammation and cancer. Nature 420, 860-867.View Article  PubMed
[63]  Homem de Bittencourt PI, Peres CM, Yano MM, Hiratea MH & and Curi R (1993) Pyruvate is a lipid precursor for rat lymphocytes in culture: Evidence for a lipid exporting capacity. Biochem Mol Biol Int 30, 631-641. PubMed
[64]  Kondoh H, Lieonart ME, Bernard D & and Gil J (2007) Protection from oxidative stress by enhanced glycolysis; a possible mechanism of cellular immortalization. Histol Histopathol 22, 85-90. PubMed
[65]  Kondoh H, Lieonart ME, Gil J BD & and Peters G (2005) Glycolysis and cellular immortalization. Drug Discov Today 2, 263-267.View Article
[66]  Peng S, Lai P, Pan H & Hsiao L and Hsu H (2008) Aberrant expression of the glycolytic enzymes aldolase B and type II hexokinase in hepatocellular carcinoma are predictive markers for advanced stage, early recurrence and poor prognosis. ONCOLOGY REPORTS 19, 1045-1053. PubMed