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Rheumatoid arthritis biomarkers

A plethora of biomarkers is available for the early diagnosis of rheumatoid arthritis and associated tissue damage and disorders. In addition, new biomarkers that improve the diagnosis and treatment of patients suffering from this common destructive autoimmune disease are constantly being discovered.

Hands affected by rheumatoid arthritis. Joints become swollen and stiffness restricts movement. © German Society of Rheumatology

Around 1% of the world's population is afflicted by rheumatoid arthritis (RA), which means around 800,000 sufferers in Germany. RA is the most common inflammatory joint disorder in which autoantibodies, i.e. antibodies formed by a person's immune system against the body's own substances, attack the synovial tissue that lines the body's joints. If left untreated, the disease leads to inflammation, swelling, pain and erosion of the bone and cartilage and deformation of the joint. The disease can also drastically increase the risk of cardiovascular disease.

Since the discovery of RA (often previously referred to as primary chronic polyarthritis, PCP) as an autoimmune disease around 25 years ago, the detection of blood autoantibodies using ELISA tests, immunoblotting or immunofluorescence has been standard diagnostic practice. Rheumatoid factor (RF) was the first autoreactive element identified in the sera of RA patients in 1930; in the 1980s this was shown to be an autoantibody specific for several mmunoglobulins (IgG, IgM, IgA), which is why the plural, i.e. rheumatoid factors, is usually used. The RF test is one of the classification criteria of the American College of Rheumatology, a leading medical society focused on rheumatic diseases. However, it is only possible to detect RF in around 70 - 80% of RA patients; it is detected even less frequently in other rheumatic diseases. RF is not RA-specific either, and is also positive in other autoimmune diseases, including lupus erythematodes, as well as infectious diseases such as tuberculosis and chronic viral hepatitis.

Immunodiagnostics using cytoskeleton components

The reliable diagnosis of RA requires the development of many other more specific biomarkers. Amongst other factors, the diagnosis of RA involves autoantibodies that target keratin and an autoantibody known as anti-Sa, which was originally identified in a French Canadian RA patient.

Neurofibroma stained with antibodies directed against vimentin. © DKFZ

The anti-Sa antibody was identified in 2004 as an anti-vimentin antibody and turned out to be a promising RA marker. Keratins (especially cytokeratins) and vimentin are structural proteins with a diameter of around 10 nm that form intermediate filaments (IF) of the cytoskeleton. They are called "intermediate" because they are located between the thin actin microfilaments and the microtubules. The cytoskeleton inside cells is thus protected against attacks by the immune system; autoantibodies against the cytoskeleton can only be generated when the cells have been destroyed as a result of inflammation, for example.

Prof. Dr. Werner W. Franke © DKFZ

Vimentin was discovered in 1978 by Professor Dr. Werner Franke, a cell biologist at the German Cancer Research Centre (DKFZ) in Heidelberg. Franke and his team carried out comprehensive studies and were able to show that cytokeratins, vimentin and other factors shown by the studies to be cytoskeleton components are tissue-specific biomarkers that provide information about a tumour's original tissue. Cytokeratins are typically found in epithelia where they lead to epithelial cancer, while vimentin is found in mesenchymal cells and hence in mesenchymal sarcomas, fibromas, lymphomas, etc. Immunodiagnoses involving antibodies against cytoskeleton proteins are now an integral part of all molecular pathology laboratories around the world.

Citrullinated peptide antigens

The RA-characteristic anti-Sa autoantibody later turned out to be directed not against the normal form of vimentin but against its citrullinated form that contains mutations. Protein citrullination occurs naturally in the body. Citrulline is a nonessential amino acid that is formed by way of the post-translational modification of arginine. It is formed through the removal of amino groups from arginine, which loses its positive charge and alters its three-dimensional structure as a result. The altered citrullinated protein is considered foreign by the immune system, which results in the formation of autoreactive antibodies. In the meantime, the process of citrullination has also been discovered in a number of peptides (protein fragments) that act as autoantibody antigens. They are found, for example, in myelin-basic protein that is associated with multiple sclerosis (MS) and also in physiological processes such as the keratinisation of skin and the modification of histones, which has been implicated in the regulation of gene expression.

Conversion of peptide-bound arginine into citrulline by enzymes known as calcium-dependent peptidylarginine deiminases (PADI) © German Society of Rheumatology

The process of citrullination is thus far from being specific to RA. However, autoantibodies directed against naturally occurring protein epitopes such as anti-citrullinated protein antibodies (ACPA), including autoantibodies against mutated citrullinated vimentin (MCV) and others, have turned out to be more powerful and reliable biomarkers of RA and other autoimmune diseases than rheumatoid factors. Standardised tests (e.g. MCV-ELISA) have since been developed, which provide information about the early stages of disease and help doctors establish effective therapies aimed at preventing the irreversible destruction of bones and joints.

Cardiovascular risk biomarkers in rheumatoid arthritis

At a congress of the German Society of Internal Medicine held in May 2011, Roche Diagnostics (Mannheim) ran a symposium on cardiovascular risk biomarkers in rheumatoid arthritis. The symposium ("Biomarker bei rheumatologischen Erkrankungen - hilfreich in Therapie und Prävention?", see Roche Diagnostics press release of 3rd May 2011) presented biomarkers that provide information on RA-related metabolic processes. Patients with rheumatoid arthritis are at increased risk of developing cardiovascular diseases, a risk that becomes even greater in patients using inflammation inhibitors and painkillers such as diclofenac and other COX inhibitors. Such patients run the risk of developing acute, chronic cardiac insufficiency and therefore have to be closely monitored. Clinical trials have shown that the N-terminal (type B) natriuretic peptide (NT-proBNP), which is released into the blood by the heart cells when there is high strain, is a reliable biomarker for predicting the risk of cardiovascular adverse events in RA patients. It is generally assumed that patients with NT-proBNP values greater than 100 pg/ml have a considerably higher risk of adverse cardiovascular events than patients with lower concentrations.

Roche Diagnostics has developed a fully automated test that reliably reports the concentration of NT-proBNP in just 18 minutes. The immunoassay is marketed as Elecsys®, and involves a table-top analysis device based on electrochemiluminescence and streptavidine-biotin reactions.

In addition to "Elecsys®proBNP", Roche will shortly launch a fully automated, total vitamin D assay (Elecsys® vitamin D test) for the quantitative measurement of vitamin D2 and D3 concentrations in human serum and blood plasma. Vitamin D promotes the incorporation of calcium in bones. Studies have shown a link between vitamin D and the risk for rheumatoid arthritis. The new assay enables vitamin D levels to be easily measured and helps prevent the development of vitamin D deficiencies in RA patients.

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