New antigen test speeds drug development

By MELANIE HSU | November 11, 2010

A team of Hopkins researchers led by Scheherazade Sadegh-Nasseri of the Department of Pathology have designed a laboratory test that is able to detect the most suitable binding site, or epitope, on an antigen.

These findings may lead to the improved diagnosis of disorders such as Lyme disease and detection of cancers cells at an earlier stage.

Sadegh-Nasseri’s lab aims to understand how antigenic proteins from pathogens are processed and presented to T-cells of the immune system.

According to Sadegh-Nasseri, T-cells see antigens as a complex of peptide fragments bound to molecules made by the Major Histocompatibility Complex (MHC), a gene family that plays a role in the immune system.

Cellular proteins are constantly being made and broken down as a part of normal processes. MHC proteins take fragments of degraded proteins and display them on the surface of the cell. Immune cells can then recognize if any of these proteins belong to virus or bacteria that have infected the cell.

Identifying the best-fit region on the antigen, the sites where peptides are presented on the surface of these MHC cells, is key to the design effective drug therapies and cancer vaccines.

The test, which models how antigens are processed into peptides and detected by T-cells, not only detects suitable binding sites in a short amount of time but also can be performed entirely in the laboratory.

After seven years of work and building on nearly 20 years of research, the team found that antigen processing and the selection of the best-fit peptide was done by only five essential proteins.

Among the components of the test mix is HLA-DR, a protein produced by the MHC that aids in the peptide selection process, HLA-DM, an accessory molecule that prevents HLA-DR from binding to antigens that it cannot perfectly fit, and three enzymes involved in breaking antigens into protein components.

“The key finding is that different complexes of MHC Class II molecules bound to different peptides are conformationally different, and therefore can be recognized differently by HLA-DM,” said Sadegh-Nasseri.

To confirm the accuracy of the test, the team tested whether a mixture of the immune system proteins could detect two common experimental antigens, type II collagen and a strain of influenza, as well as unknown epitopes for malaria and avian flu.

Mass spectrometry was used to confirm that HLA-DR successfully bonded to an antigen epitope.

The researchers then injected HLA-DR-producing mice with the antigens and collected T-cells that resulted from the immune response.

By exposing the cultured T-cells to the suspected epitopes and matching the T-cell reactions to peaks in the mass spectroscopy, the team was able to pinpoint one dominant binding site for each of the four antigens.

For future experiments, the researchers plan to refine the chemical mixture to analyze other kinds of HLA. According to Sadegh-Nasseri, the team also wishes to extend their technology to cancer and autoimmune antigens.

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