Antibody-
cytokine fusion
proteins are a class of
biopharmaceuticals, with the potential to modulate the activity of the immune system at the site of disease. The molecular format used to connect antibody moiety and
cytokine payload can have a profound influence on
biological activity and pharmacokinetic properties. The optimization of fusion
protein format is particularly challenging for heterodimeric
cytokines, since various molecular arrangements can be considered. In this article, we have explored the role of linker in a
tumor-targeting
IL12 fusion
protein, based on the L19 antibody, specific to the extra-domain B of
fibronectin. In biodistribution studies performed in
tumor-bearing mice using radioiodinated
protein preparations, fusion of human
IL12 at the N-terminus of the L19 antibody in tandem-diabody format led to higher
tumor uptake and improved
tumor-to-organ ratios, compared to a similar fusion
protein featuring L19 in
IgG1 format. Moreover, optimization of the
amino acid composition in eight variants of the linker connecting the
IL12 moiety to the tandem-diabody revealed that a 15-amino
acid linker (GSADGGSSAGGSDAG) displayed the best
tumor targeting characteristics, with a long residence time at the
tumor site and a rapid clearance from blood and normal organs. The product is being developed for industrial and clinical applications.