Paul Kovac, Ph.D., Dr. h.c., Scientist Emeritus

Research Goal

The purpose of my research is to develop/improve methods for chemical attachment of carbohydrates to suitable carriers, and design protocols for making conjugate vaccines for infectious bacterial diseases from synthetic and bacterial carbohydrate antigens. Such vaccines are often free from undesirable traditional bacterial vaccines often have. Some neoglycoconjugates we prepare can be used as diagnostic tools to identify presence of pathogens.

Current Research

Our primary focus is to further the development of conjugate vaccines from synthetic and bacterial carbohydrate antigens. Ultimately, we would like to develop reliable protocols for preparation of neoglycoconjugates that could become substitutes for traditional vaccines based on whole-cell killed or attenuated bacteria. Such vaccines are often pyrogenic or have other undesirable effects. As synthetic carbohydrate antigens, we use oligosaccharides that mimic the structure of polysaccharides present on the surface of bacterial pathogens. As bacterial bacterial antigens we use protective antigen-containing fragments of lipopolysacchardies (LPS). Because there is virtually an infinite number of choice of architectonic details that a synthetic neoglycoconjugate can incorporate, part of our work involves studies of the effects of variables such as size of the carbohydrate antigen, type of linker, linking chemistry, type of carrier, etc., upon immunogenicity and protective capacity. In addition to obtaining potent immunogens, we expect our studies to result in findings of general utility in synthetic vaccine preparation.

With synthetic fragments of LPS, our approach involves five stages:

1. Synthesis of fragments of antigenic polysaccharides and the deoxy and deoxyfluoro analogs of those fragments
2. Studies of binding the above ligands with antibodies to the native antigen and identification of critical hydrogen bonding interactions
3. Identification of the immunologically dominant oligosaccharide sequence in the antigenic polysaccharide
4. Chemical conjugation of such fragment(s) to suitable carrier(s), to obtain neoglycoconjugates
5. Probing the antigenicity, immunogenicity, and protective capacity of neoglycoconjugate(s)

With bacterial LPS, our approach involves:

1. Detoxification of LPS by mild acid hydrolysis to remove lipid A and arrive at O-SP–core fragment (O-SPc) of the LPS
2. Purification of O-SPc
3. Conjugation of O-SPc to a suitable protein carrier
4. Probing the antigenicity, immunogenicity, and protective capacity of neoglycoconjugate(s)

Our section has studied the interaction of carbohydrate antigens and antibodies for many years using the above approach. As a result, we have been able to obtain a great deal of detailed information on binding at the molecular level. Following the same concept, groundwork was laid for development of immunogens for Shigella dysenteria type 1. The current objective of our work is development of synthetic vaccines for cholera and other enteric diseases, and anthrax.

In addition, we often engage in collaborative research, within the United States and internationally, with universities and other scientific institutions.

Applying our Research

Safer vaccines and better diagnostic tools that we aim to develop translate into improved public health.

Need for Further Study

Further areas of study include optimization of the conjugation protocol, identification of the optimal structure of the conjugate vaccine and immunization schedule in order to deliver the optimal amount of antigen necessary to elicit protective level of antibodies.

Research in Plain Language

The primary focus of the Carbohydrates Section in the NIDDK’s Bioorganic Chemistry Laboratory is to develop safer, highly effective vaccines. Instead of using actual disease-causing organisms that have been killed or attenuated, we use in vaccine development antigens that have been synthesized especially for this purpose. Alternatively, we use carbohydrates obtained from chemically modified bacterial lipopolysaccharides. Vaccines made from whole cell killed or weakened disease organisms often have undesirable properties, they may cause high fever (they are pyrogenic) or have other side effects.

Sometimes the outer coat of an actual disease-causing organism is not the optimal antigen (a substance that causes your immune system to produce the desirable antibodies) for a strong, desirable immune response. In order to enhance this response, the carbohydrate portion of this bacterial coating can be combined (or conjugated) with a suitable carrier, often after removal of its toxic part, Lipid A. The material resulting from conjugation is called a glycoconjugate.

Our preferred approach is to create a synthetic carbohydrate molecule—a synthetic oligosaccharide—that is chemically similar to the carbohydrate coat of the actual disease-causing organism and evokes a strong immune response when it is conjugated to a carrier. Because the synthetic material does not contain any, potentially harmful fragments of the bacterial pathogen, the construct resulting from linking together purified synthetic oligosaccharide and a purified carrier does not have pyrogenic or other undesirable side effects. Our current objective is to develop safe conjugate vaccines for cholera and other enteric diseases, and anthrax, using such synthetic components.

There are many ways for synthesizing synthetic oligosaccharides that mimic the carbohydrate bacterial coat and linking them to a carrier. Therefore, part of our work is to study how various factors, such as the size of the carbohydrate fragment and the mode of linking these substances to suitable carriers, affect the ability of the conjugated vaccine to evoke a strong protective immune response in a vaccinated person. For many years, we have used a systematic, staged approach to develop synthetic oligosaccharides and the conjugate vaccines made from them. This approach starts with synthesizing fragments of the weakly antigenic carbohydrate coat of the disease organism, finding the fragments that evoke the strongest production of effective antibodies, and conjugating these fragments to suitable carriers. We then test how well this conjugated compound works as a protective vaccine.