Antibodies directed againt transition state analogs

The anti-idiotypic approach: from enzymes to abzymes

Naturally occurring abzymes

Metallo- and Hemoabzymes

           Historically, the first approach for producing abzymes mimicking enzymatic reactions, based on the concept proposed by L. Pauling and developed by W. Jencks, consisted in immunizing mice with stable molecules resembling the transition state structure of the chemical reaction that was expected to be catalyzed.
The first successful results were obtained for hydrolysis of ester and carbonate bonds by the groups of R. Lerner and P. Schultz. They used as immunogens organophosphate compounds that were thought to mimic the tetrahedral structure of the carbon atom during the hydrolytic process (cf. figure n°1).

           Since that time, more than 70 different chemical reactions were described to be catalyzed by antibodies. These reactions include hydrolysis of chemical bonds (esters, carbonates, amides, phosphates,…), stereospecific synthesis of compounds (esters, amides, Diels-Alder addition, …), as well as reactions of isomerization, decarboxylation, oxidation and reduction, ….

           If the first obtained abzymes were characterized by a rather low efficiency, the improvements in hapten design (bait and switch), in the strategies used for immunization (heterologous immunization, reactive immunization), in the site-directed modification of antibodies (mutagenesis) or in the methods developed for screening (catELISA) and selection (phage display) allowed to obtain catalysts with efficiency sufficient for an industrial or a medical application.

           On the other hand the resolution of an increasing number of tridimensional structures of abzymes brings a better understanding of the appearance and evolution of the catalytic function, not only in antibody binding sites but also in enzyme active sites.

           In 1974, Niels Jerne advanced the theory that regarded the immune system as a network of interacting idiotypes. A major postulate of the idiotypic network was that for each immunoglobulin (Ab1) generated against an antigenic determinant there existed a complementary antibody (Ab2) directed against the idiotypic determinants of Ab1.

           When the idiotypic determinant superimpose with the binding site of Ab1, some of the Ab2 may mimic the antigen's determinants and are designed as "internal images" of the original antigen.

           Figure n°2 shows the experimental process which is used to product abzymes.

For producing catalytic antibodies, a first antibody Ab1 is raised that recognizes the active site of an enzyme so that the combining site of Ab1 has structural features complementary to those of the enzyme.
A second set of antibodies (Ab2) is produced against the Ab1 combining site. Among these second-generation, or anti-idiotypic antibodies, some of them may represent a structural internal image of the original enzymatic site, and in some cases may exhibit a catalytic activity.

           Using this approach, antibodies with esterase and amidase activities were characterized. These abzymes usually bear efficient catalytic activities, with a relaxed specificity when compared with the model enzyme.

           Antibodies with catalytic activities were isolated from sera of patients with different diseases. The first natural abzymes were obtained by means of antibody purification from human serum.

           Antibodies with protease activity against vasoactif intestinal peptide (VIP) were first isolated in the serum of patients with asthma. Surprisingly, antibodies exhibiting the same catalytic activity were obtained by immunizing mice with VIP in its ground state. These monoclonal antibodies have allowed to demonstrate that the catalytic activity is borne by the isolated light chain of the antibody molecule.

           This VIP-ase activity was also shown to be present on Bence-Jones proteins, that are monoclonal human light chains found in urine of patients with multiple myeloma.

           Other protease activities were charaterized for cleaving of throglobulin in the serum of patients with Hashimoto thyroiditis, or for hydrolyzing factor VIII in hemophilia patients infused with homologous factor VIII. DNA hydrolyzing autoantibodies were also isolated from the sera of patients with systemic lupus erythematosus or rheumatoid arthritis. The DNA-hydrolyzing activity could be correlated to the presence of high levels of anti-topoisomerase I antibodies in the sera of patients.

           The idea to produce antibodies that mimic metalloenzymes or hemoenzymes was early advanced to generate antibodies able to cleave stable chemical bonds or to catalyze peroxidase-like reactions.

           In 1989, an antibody catalyzing the hydrolysis of a Glycine-Phenylalanine bond was obtained by using as the antigen an hapten that not only induce the selective recognition of the Gly-Phe sequence, but also induce the generation of residues able to complex a metal ion. When complexed with zinc, this antibody exhibits a good catalytic activity.

           The generation of antibodies directed against different metalloporphyrins has allowed to demonstrate that antibodies may mimic metalloproteins. The Hemoabzymes obtained are able to catalyze peroxidase-like activities and could be very interesting tools for the enantioselective oxidation of molecules.