Summary
Prophylaxis with 2–4 times weekly dosing of factor (F)VIII or FIX is established as
an efficacious and safe treatment in haemophilia. Although prophylaxis is not readily
available for the inhibitor patient,recent studies have demonstrated a reduction in
bleeding episodes in inhibitor patients treated with daily infusions of FVIIa. In
order to develop a treatment option comparable to prophylaxis with FVIII or FIX we
looked to PEGylation which is an established method for prolonging the circulatory
half-life of proteins. However, due to the numerous interactions of FVIIa with the
cell surface,TF,FIX and FX there are limited options for unspecific chemical modification
of FVIIa without loss of activity. Consequently, we explored the GlycoPEGylation™
technology for selective PEGylation of the two N-glycans in the FVIIa light chain
and protease domain to generate seven specifically modified derivatives with PEG groups
ranging from 2 to 40 kDa. These derivatives were evaluated in vitro for their ability to interact with small synthetic substrates as well as key molecules
relevant to function in the coagulation pathway. The results demonstrate that modification
of FVIIa using glycoPEGylation has only a very limited effect on the hydrolysis S-2288
and FX activation. However, the modification does to some extend alter the ability
of FVIIa to interact with TF and more importantly, reduces the rate of ATIII inhibition
by up to 50% which could allow for an extended active half-life in circulation.
Keywords
Long-acting - PEG - coagulation - serine protease - factor VIIa - prophylaxis
