In recent years, a novel treatment method for cancer emerged, which is based on the starvation of tumors of amino acids like arginine. The deprivation of arginine in serum is based on enzymatic degradation and can be realized by arginine deaminases like the L-amino acid oxidase found in the ink toxin of the sea hare Aplysia punctata. Previously isolated from the ink, the L amino acids oxidase was described to oxidate the essential amino acid L-lysine and L arginine to their corresponding deaminated alpha-keto acids. Here, we present the recombinant production and functionalization of amino acid oxidase Aplysia Punctata ink toxin (APIT). PEGylated APIT (APIT-PEG) increased the blood circulation time. APIT-PEG treatment of patient-derived xenografted mice shows a significant dose dependent reduction of tumor growth over time mediated by amino acid starvation of the tumor. Treatment of mice with APIT-PEG which lead to deprivation of arginine was well tolerated.

Vascular endothelial growth factor A-165 (VEGF-A 165 ) positively modulates neointimal hyperplasia, lumen stenosis, and neovascularization. One challenge for the use of VEGF-A 165 for potential therapy is its short serum half-life. Therefore, we are designing VEGF-A 165 bioconjugates carrying polyethylene glycol (PEG). The purity of the recombinantly expressed human VEGF-A 165 exceeded 90%. The growth factor had a half-maximal effective concentration of 0.9 ng/mL (EC50) and induced tube formation of human umbilical vein endothelial cells. PEGylation was conducted by Schiff base reaction followed by reductive amination. After purification, two species were obtained, with one or two PEG attached per VEGF-A 165 dimer. Both resulting bioconjugates had a purity exceeding 90%, wild-type bioactivity, and increased hydrodynamic radii as required for prolonging the half-life.