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K. V. Sukhoverkov, N. N. Sokolov, O. J. Abakumova, O. V. Podobed, E. V. Kudryashova
Formation of conjugates
with a PEG-chitosan improves biocatalytical efficiency and antitumor activity of
L-asparaginase from Erwinia carotovora
Abstract
Conjugation with new
branched co-polymers, PEG-chitosan, was suggested to improve therapeutic
properties of L-asparaginase from Erwinia
carotovora (EwA). Composition
of EwA conjugates with PEG-chitosan co-polymers was optimized for maximal
catalytic efficiency (kcat/KM) at
physiological conditions, yielding an improvement by the factor of 4–6 in
comparison with native enzyme. This effect is mainly attributed to the shift of
unfavorable pH activity profile towards physiological values. Catalytic
activity of EwA-PEG-chitosan conjugates with L-glutamine, which is known to be responsible for side effects, appears 4–5 times
lower as compared to the native enzyme. The thermostability and proteolytic
stability of EwA conjugates has also been considerably improved (by the factor
of two). In cell assays EwA-PEG-chitosan conjugates demonstrate 20–40%
increased cytostatic activity against human myeloid leukemia K562 cells and
human breast adenocarcinoma cells MCF7, while the activity against Burkitt lymphoma
Raji cells and human acute lymphoid leukemia Jurkat cells is generally at the
same level with native enzyme added in the same IU dose. Taking into account
the higher IU/mg activity of the conjugates, the same therapeutic effect is
achieved with 4–6 times lower amount of enzyme (in mg) compared to native EwA,
suggesting reduced immunogenicity in the case of conjugates. Therefore,
ChitoPEGylation is suggested as a perspective approach for the development of
improved forms of therapeutic enzymes, especially when their pH-optima is
substantially different from physiological pH.
Key words: L-Asparaginase, PEG–chitosan, branched
copolymers, catalytic activity, thermostability, antitumor activity.
Copyright (C) Chemistry Dept., Moscow State University, 2002
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