USA HCG REFERENCE SERVICE
SYNTHESIS AND DEGRADATION OF HCG

 

   hCG is glycoprotein hormone produced by normal trophoblast cells of the placenta during pregnancy. It is also is produced by trophoblast cells in hydatidiform mole and choriocarcinoma (trophoblast diseases), and in patients with germ cell tumors (testicular choriocarcinoma, placental site tumors and germ cell carcinomas of the ovary) and sometimes in those with other malignancies. Small amount of hCG may also be produced by the pituitary gland.
    hCG is composed of two subunits, alpha and beta, held together by ionic and hydrophobic forces. The alpha-subunit is a glycopeptide of 92 amino acids, with asparagine-linked (N-linked) sugar moieties attached at residues 52 and 78. It is stabilized by 5 disulfide linkages. The amino acid sequence of the alpha-subunit is identical to that of the alpha subunits of the pituitary glycoprotein hormones, lutenizing hormone (hLH), follicle stimulating hormone (hFSH) and thyroid stimulating hormone (hTSH). The ß-subunit of hCG is a glycopeptide of 145 amino acids, stabilized by 6 disulfide linkages. It has N-linked sugar moieties attached at residues 13 and 30, and 4 serine-linked (O-linked) sugar moieties attached to the C-terminal peptide (ß-subunit residues 121-145). The ß-subunits of the glycoprotein hormones are unique, giving them their different biological characteristics.
    Normal trophoblast hCG molecules (normal pregnancy) have mono and biantennary type N-linked sugar moieties and simple trisaccharide-core O-linked sugar units. Early pregnancy cytotrophoblast cells, and cytotrophoblast cells present in persistent hydatidiform mole and choriocarcinoma cases, and in testicular germ cell cancer cases, make a form of hCG with more-complex sugar side chains, triantennary N-linked sugar moieties and tetrasaccharide core O-linked sugar moieties (reference 1, below). This type of hCG we call hyperglycosylated hCG. A small amount of hCG can be produced by the pituitary gland. The N-linked sugar side chains on pituitary hCG have unusual sulfated sugar moieties (terminating in N-acetylgalactosamine-sulfate rather than  galactose-sialic acid), just like the sugar moieties on pituitary hLH (reference 2, below).
    In addition to hCG, free hCG subunits (free alpha-subunit and free ß-subunit) and hCG and free subunit degradation products are produced in pregnancy, trophoblast disease and cancer. These can be detected in serum, plasma and urine samples. Free subunits can derive from the excess synthesis of alpha or beta subunit or incomplete combination of subunits in cells. Excess alpha subunit which fails to get incorporated in hCG may become hyperglycosylated (like trophoblast disease and cancer hCG) with extra sugar residues on the N-linked sugar moieties (reference 3, below). This is called large free alpha-subunit. This can also be detected in plasma, serum and urine samples. Free subunits also derive from the slow dissociation of hCG, dissociation half-time >40 days at 37 deg, and from the more rapid dissociation of nicked, damaged or hyperglycosylated hCG, dissociation half-time 10-16 days (references 4 and 5, below).
    A nicked or cleaved hCG is produced. Nicked hCG is most evident in serum and urine samples in the later trimesters of pregnancy. It may be the principal form of hCG detected in choriocarcinoma patients, and in germ cell and other cancer cases. It also may be the principal form of hCG in the weeks following evacuation or successful therapy of hydatidiform mole and choriocarcinoma when levels drop below 100 mIU/ml (references 5 and 6, below). As illustrated below (FIGURE 1), nicked hCG is cleaved or cut in the ß-subunit peptide between residues 47 and 48. It can be cleaved between residues 43 and 44 or 44 and 45 in some choriocarcinoma cases (reference 1, below). Although cleaved, the molecules are held together by the intra-peptide disulfide linkages; no amino acids or sugar residues are missing (FIGURE 1). Nicking occurs by the action of proteolytic enzymes in the placental, mole or cancer tissue, or in the circulation. 

FIGURE 1. Nicking of hCG. Yellow ribbon indicates ß-subunit peptide and strings of letters indicate sugar side chains (L=GalNac, G=Gal, S=Sialic acid, M=Mannose, A=GlcNac). White ribbon indicates alpha-subunit peptide.

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    Other forms of degraded hCG are detected in serum and urine samples from individuals with hydatidiform mole, choriocarcinoma and germ cell and other cancers (click here to see sources of hCG). These include hCG missing the ß-subunit C-terminal peptide, and the remnant, the free ß-subunit C-terminal peptide (FIGURE 2) (references 7 and 8, below).

FIGURE 2. Cleavage of hCG ß-subunit C-terminal peptide.

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    Nicked hCG is unstable (dissociation half-time 10-16 days at 37 deg. versus >40 days for non-nicked hCG), splitting apart in the circulation to release a free alpha-subunit and a nicked free ß-subunit (FIGURE 3) (references 5 and 9, below)

FIGURE 3. The dissociation of nicked hCG.  

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   Nicked free ß-subunit is rapidly removed from the circulation. It may be degraded in the kidney to ß-core fragment, and excreted into urine (FIGURE 4) (references 5 and 9, below). ß-core fragment may be the terminal degradation product of hCG. It comprises to short peptides, ß-subunit residues 6 to 40 and ß-subunit residues 55 to 92, held together by 4 disulfide linkages. The 2 N-linked sugar moieties on ß-subunit core fragment are also degraded, each containing only 5 out of 11 (on normal pregnancy hCG) sugar residues. While the molecular weight of hCG is 36,700, the molecular weight of ß-core fragment is only 10,000. From 8 week of gestation until term, ß-core fragment is the principal hCG ß-subunit-related molecule in urine samples. Urine ß-core fragment may be the only hCG-related molecule detectable (in either serum or urine) in certain germ cell cancers, placental-site trophoblastic tumors, in ovarian cancer, bladder cancer and certain other malignancies (click here to see potential sources of hCG in and outside of pregnancy) (reference 10, below).

FIGURE 4. Degradation of nicked free ß-subunit to ß-core fragment in the kidney.     

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REFERENCES

 

1.      Elliott, M., Kardana, A., Lustbader, J. and Cole, L. Carbohydrate and peptide structure of the alpha- and beta-subunits of hCG from normal and aberrant pregnancy and choriocarcinoma. Endocrine 7:15-32, 1997.

2.     Green E,D., and Baenziger, J.U. Asparagine-linked oligosaccharides on lutropin, follitropin, and thyrotropin. II. Distributions of sulfated and sialylated oligosaccharides on bovine, ovine, and human pituitary glycoprotein hormones. [Journal Article] Journal of Biological Chemistry. 263(1):36-44, 1988

3.    Blithe D.L., and Nisula B.C. Variations in the oligosaccharides on free and combined alpha -subunit of human choriogonadotropin in pregnancy. Endocrinology 117:2218-28, 1985.

4.    Butler, S.A., Cole, L.A., Chard, T., and Iles, R.K. Dissociation of hCG into its free subunits is dependent on naturally occurring molecular structural variation, sample matrix and storage conditions. Ann Clin Biochem, 35:754-760, 1998.

5.    Cole, L.A., Kardana, A., Park S-Y., Braunstein, G. The deactivation of hCG by nicking and dissociation. J Clin Endocrinol Metab 76:704-10, 1993.

6.    Nishimura, R., Ide, K., Utsunomiya, T., Kitajima, T., Yuki, Y., Mochizuki, M. Fragmentation of the ß-subunit of human chorionic gonadotropin produced by choriocarcinoma. Endocrinology 123:420-425, 1988.

7.    Amr, S., Rosa, C., Wehmann, S., Birken, S., and Nisula, B. Unusual molecular forms of hCG in gestational trophoblastic neoplasia. Annales Endocrinol (Paris) 45:321-326, 1984.

8.    Cole, L.A., and Kardana, A. Discordant Results in human chorionic gonadotropin assays. Clin. Chem., 38:263-270, 1992

9.    Kardana, A., Cole, L.A. Human chorionic gonadotropin b -subunit nicking enzymes in pregnancy and cancer patient serum. J. Clin. Endocrinol. Metab.,79:761-767, 1994

10.    Cole, L.A., Tanaka, A., Kim, G.S., Park, S-Y., Koh, M.W., Schwartz, P.E., Chambers, J.T., and Nam, J-H. Beta core fragment (b -core / UGF / UGP), a tumor marker: Seven year report. Gynecol. Oncol., 60:264-270, 1996.