CATALYTIC ANTIBODIES OR ABZYMES

 

 

 

The year is 2050 - Drugs are being designed with built-in homing devices that seek out their targets like microscopic missiles. Once the target is found, the drug molecules fire until the target is effectively destroyed, without creating so much as a ripple of side effects to the patient – interesting isn’t it? Just read on.

 

The Abzyme or catalytic antibody technology is the offspring of the Monoclonal antibody technology that came up in the 70s. The basic outline of abzyme action came up 28 years back with the proof of concept coming in 1986 with the first descriptions of active abzymes capable of catalyzing ester hydrolysis reactions.

 

       Abzyme = Antibody + Enzyme

 

The basics:

 

        Enzymes are biological catalysts that speed up biochemical reactions without undergoing any change themselves. The mode of action of enzymes is that enzymes stabilize the transition state by bringing down the activation energy (ΔG) thus speeding up the reaction converting it to product. Thus they drive reactions that are, otherwise impossible at biological temperatures. Enzymes have active site that is complimentary, sterically and chemically to the substrate, making them specific (hand in glove hypothesis). Antibodies are specific substances that are produced by the body in response to the presence of any foreign body, the Antigen. The antigen-antibody specificity causes them to bind which then signal the immune system to send in the troops i.e. the Leucocytes that destroy them.

 

Why Abzymes?

 

        The mechanism of antibody action is incomplete as it only targets and catches hold of the intruder, while enzymes actually drive a reaction. So why not combine their actions and make a substance that catches hold of the antigen (analogous to function of antibody) and neutralizes it (analogous to function of enzymes making it inactive by certain chemical reactions) thus was born the idea of combining the functions of antibody and enzymes to create abzymes.

 

The principle:

       

        The generation of abzymes is based on the Transition-state theory of catalysis. Catalysts work by stabilizing the transient form of a molecule (decrease the activation energy ΔG, less energy – more stable), known as the transition state, which occurs as the original molecule changes its shape during a chemical reaction. For example, a molecule will bend and strain just prior to being broken into two pieces. If a mimic of the transition state, i.e. the strained version of the molecule could be generated, then an antibody that would tightly bind that mimic should be able to catalyze the reaction of the original molecule, by causing it to bend and strain and ultimately break. Therefore, by designing a good transition state mimic (Transition State Analog TSA), novel catalysts can be created by harnessing the power of the immune system and directing it towards a new function.

 

The production technique:

 

This hypothesis is made reality by the fact that antibody to almost any molecule can be produced by the use of the immune system. The process of making abzyme goes like this:

    Step 1: the transition state of the specific enzyme reaction is studied.

    Step 2: suitable transition state analogs (TSA) or HAPTENs that mimic the transition states are created.

    Step 3: introduce these HAPTENs into the organism and let the immune system do the rest.

    Step 4: the antibodies are raised against the TSA and are recovered.

These when in action bind to the specific antigen (because they are antibodies) and also destroy them –or whatever they are meant to do (because they resemble the more stable transition state they drive the reaction to completion).

 

The improvements in production:

 

       Although abzymes have been shown to be able to catalyze a wide range of different reactions, the catalytic efficiency of the ones prepared to date is usually low (catalysis rates k_cat >10⁵) when compared with their natural, enzyme counterparts. This has led to efforts to improve the efficiency of abzymes by random mutagenesis methods, such as phage display. In this method, hypervariable regions of the abzyme's binding site are mutated and variants screened for improved binding to substrate and catalytic activity. Previously hybridoma cells of hyper-immunized mice were used to synthesize antibody in vivo which took a longer time (4-6 months) while advances in cell-culture techniques has made it possible to carry out splenocyte immunizations in vitro by incubating the naive spleen cells in medium containing the antigen. This process typically takes 3-5 days, at which stage a cell fusion can be used to obtain hybridomas thus reducing abzyme production time.  

Applications:

Targeting Device: In treatment of cancer to enhance drug delivery

Abzymes have been used as tools to function as homing devices for the site-specific delivery of the prodrug and activators of the prodrug into the cytotoxic form. In cancer treatment one of the major hurdles is that the cytotoxic chemicals destroy both the normal and tumor cells. Hence there is a need for specific targeting of tumor cells and subsequent activation of the prodrug which then destroy the oncogenic cells. This is where the abzymes fit into picture. Prodrug is a pharmacologically inactive compound that converts to the active form of the drug by endogenous enzymes or metabolism. It is generally designed to overcome problems associated with stability, toxicity, lack of specificity, or limited bioavailability. The prodrug is comprised of the active drug compound itself and a chemical masking group that temporarily suppresses activity and appreciably reduces toxicity.

The first approach for site specific targeting is called as ADEPT (Antibody Directed Enzyme Prodrug Therapy). It involves an antibody-enzyme conjugate and prodrug mixture that is injected into the patient. The antibody binds to the specific tumor antigens present on the surface of tumor cells. The antibody-enzyme complex on binding the cell releases the free enzyme which cleaves the prodrug into the drug (active form) and the co-enzyme that suppresses the drug of its function. The released drug then kills the tumor cell by its toxicity. But the problem is that this enzyme should not be present in humans. Also foreign enzymes from bacteria cannot be used due to immunogenicity problems leading to low clinical value. So abzymes are used in the modified ADAPT (Antibody Directed Abzyme Prodrug Therapy). In this approach, antibodies are raised against the appropriate transition state analogues (TSAs) to enable them to catalyze prodrug activation. (Michael Blackburn and colleagues at the University of Sheffield (U.K.). the main drawback is that catalytic rate is slow and cell kill is only 75 -80% as against 90% required to treat tumor else the escaped cells may develop resistance.

New Targets:

    Abzyme-targeted prodrug activation can be used to target HIV-infected cytotoxic T-cells. This method can also be used for other chronic viral diseases, if infected cells display viral protein that can be targeted like in Hepatitis C.

    Correction of inborn metabolic errors, such as those that occur in patients with severe combined immune deficiency. These individuals are highly susceptible to infectious agents because their immune system is unable to produce antibodies. The disease could be managed with a long-lived catalytic antibody.

    In military to destroy chemical and biological weapons (with extensive modifications, of course)

Future promise:

       The growing field of catalytic enzymes or abzymes holds great promise in therapies and if everything goes well the futuristic therapy quoted in the beginning may become a reality soon and revolutionize the way drugs target diseases.

 

 

 

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