Carbohydrate-targeted optimization of therapeutic gonadotropins - Carbohydrate Chemistry: Chemical and Biological Approaches - page 467

Table 2 Distribution of acidic and complex glycoforms of LH during the ovarian cycle.
Acidic glycoforms are determined as variants with pIr6.55 and complex glycoforms as
non-retained material from Concanavalin A chromatography. Adapted from Anobile et al.,
1998. 33

Early to mid-follicular Late-follicular Midcycle Luteal Menopause

Acidic 64% 57% 47% 68% 87%

Complex 18% 20% 13% 24% 4%

i) hCG contains predominantly sialylated mono- and biantennary N-
linked glycans 40, 42-45 but also triantennary N-glycans 46 and sialylated tri-,
tetra- and hexasaccharide O-linked glycans. 47-49 Serum hCG is found to
be more glycosylated than placental or urinary forms, as well as first
trimester hCG is more heavily glycosylated than other forms of hCG. 50
ii) Hyperglycosylated hCG secreted by choriocarcinoma cells displays
no sialic acid, 50% of core-fucosylated glycans 46,51,52 together with an
increased content in hybrid, triantennary N-glycans and tetrasaccharide
O-linked glycans compared to hCG. 46,49,53,54 The biantennary structures
typical of hCG are not detected at all. 53 Altered glycosylation of hyper-
glycosylated hCG seems to prevent the complete folding of the ab dimer. 3
Hyperglycosylated hCG is the principal variant of hCG produced in early
pregnancy with an average of 87% of the total hCG produced in serum
during the third week, 51% during the fourth week and 43% during the
fifth week of gestation. Hyperglycosylated hCG levels then dwindles to
o 1% of total hCG during the second and third trimesters of pregnancy. 3
Interestingly, high levels of hyperglycosylated hCG were correlated with
term pregnancies while insucient levels of hyperglycosylated hCG were
correlated with pregnancy failure 55,56 inferring that miscarriages are due
to insucient production of hyperglycosylated hCG leading to a failure of
egg implantation. Moreover, increased hyperglycosylated hCG in urine
(9.5-fold) compared with hCG was correlated with Down's syndrome. 57
iii) The oligosaccharide structure of free b-subunits are similar to that
of hyperglycosylated hCG with a high proportion of triantennary N-linked
glycans and tetrasaccharide O-linked glycans, and a markedly elevated
inner fucosylation. This finding may indicate that large glycans are able
to inhibit the association of b-subunit with the a-subunit. 54
iv) Pituitary hCG differs from pregnancy hCG by a variable portion of
sulfated oligosaccharides. These sulfated groups replace galactose and
sialic acid residues in N- and O-linked glycans. 58

3.2.4 Free alpha subunit. Free a-subunit contains complex multi-
antennary structures with 2.5-fold higher amounts of sialic acid and
galactose as well as a higher amount of GlcNAc and 6-fold higher amount
of fucose than combined a-subunit. 45 The size of such glycans may
prevent early combination with the b-subunit. 59 Little 60,61 or no 62
O-glycosylation is found in combined a-subunit. It was however hy-
pothesized that when a-subunits fail to combine with b-subunit in the
rough endoplasmic reticulum, they become substrates for further add-
ition of O-linked carbohydrates, presumably in the Golgi apparatus. 61