Use of designer recombinant mitochondrial antigens in the diagnosis of primary biliary cirrhosis

TLDR

Autoantibodies against mitochondrial E2 subunits of the pyruvate dehydrogenase and branched‑chain 2‑oxo‑acid dehydrogenase complexes have been recognized for over 25 years, but crude extracts caused nonspecificity until cloning identified these subunits as the major immunodominant antigens. The authors aimed to create a dual‑headed recombinant antigen displaying the immunodominant epitopes of both E2 subunits to enable a sensitive, reproducible assay for anti‑mitochondrial antibodies in primary biliary cirrhosis. They engineered a single protein that co‑expresses the key epitopes of the pyruvate dehydrogenase and branched‑chain 2‑oxo‑acid dehydrogenase E2 subunits, allowing precise epitope mapping and assay development. Using this designer antigen, the assay detected antibodies in over 90 % of PBC patients, including the ~10 % who recognize only the branched‑chain E2 subunit, and represents the first example of a designer molecule for immunodiagnosis. Hepatology 1992; 15:367‑372.

Abstract

The appearance of autoantibodies against mitochondria in patients with primary biliary cirrhosis has been known for more than 25 yr. In the past, based on the biochemical complexity of the mitochondrion and the use of crude extracts for immunodiagnosis, a degree of nonspecificity in assaying for antibodies to mitochondria has been present. This problem has been largely circumvented by the cloning of the mitochondrial antigens and the identification of the E2 subunits of the pyruvate dehydrogenase complex and the branched chain 2-oxo-acid dehydrogenase complex as the major and immunodominant autoantigens of primary biliary cirrhosis. More than 90% of patients with primary biliary cirrhosis have been shown to react with one or both of these enzymes using either recombinant antigen or purified native protein. Approximately 10% of patients recognize only E2 subunits of branched chain 2-oxo-acid dehydrogenase complex and not pyruvate dehydrogenase complex. Such patients would be missed by diagnostic assay that has a low sensitivity to antibodies against E2 subunits of branched chain 2-oxo-acid dehydrogenase complex. The use of recombinant and biochemically pure antigens has permitted structural and conformational analysis of epitope mapping. We have taken advantage of the antigenic mapping studies of both primary biliary cirrhosis and branched chain 2-oxo-acid dehydrogenase complex E2 subunits and designed a molecule that expresses the immunodominant epitopes of both. Using this dual-headed molecule that coexpresses the epitope of two different antigens, we report herein a sensitive and reproducible assay for antibodies to mitochondria in patients with primary biliary cirrhosis. We believe that this recombinant protein is the first example of the use of designer molecules for immunodiagnosis. (Hepatology 1992; 15:367-372).