Research Article

Characterization of IXINITY® (Trenonacog Alfa), a Recombinant Factor IX with Primary Sequence Corresponding to the Threonine-148 Polymorph

Table 1

Posttranslational modifications in trenonacog alfa, nonacog alfa, and plasma-derived factor IX.

Posttranslational modification Trenonacog alfaNonacog alfaPlasma-derived factor IX

Disulfide bridgingEleven disulfide bridgesEleven disulfide bridges Eleven disulfide bridges [4]

γ-Carboxylation≥11.7 mol γ-carboxyglutamic acid/mol factor IX11.5 mol γ-carboxyglutamic acid/mol factor IX [4]12 mol γ-carboxyglutamic acid/mol factor IX [5]

β-Hydroxylation50%  β-hydroxylated at Asp6446%  β-hydroxylated at Asp64~40%  β-hydroxylated at Asp64 [4]

SulfationTyr155: low degree of sulfation observed (5%)Tyr155: low degree of sulfation observedTyr155: high degree of sulfation observed [4]

PhosphorylationSer158: low degree of phosphorylation observed (3%)Ser158: low degree of phosphorylation observedSer158: ~90% phosphorylated [6]

O-linked glycosylation
 EGF domainSer53: -Glc
Ser61: Neu5Ac-Hex-HexNAc-Fuc
Ser53: -Glc [21]
Ser61: Neu5Ac-Gal-GlcNAc-Fuc [21]
Ser53: -Glc [7]
Ser61: Neu5Ac-Gal-GlcNAc-Fuc [7, 8]
 Activation peptideLow degree of site occupancy observed (13%)Low degree of site occupancy observed [4]Low degree of site occupancy observed [9]

N-linked glycosylationN-glycans are complex, predominantly tri- and tetraantennary, and predominantly sialylated; 80% of N-glycans are either tetrasialylated or trisialylatedN-glycans are complex, predominantly tri- and tetraantennary, and predominantly sialylated [4]N-glycans are complex, predominantly tri- and tetraantennary, and predominantly sialylated [10]

Total sialylation8.8 mol sialic acid/mol factor IX6.5 mol sialic acid/mol factor IX [9]8.8 mol sialic acid/mol factor IX [9]