Diagnostic and clinical utility of antibody testing in rheumatoid arthritis

Rheumatoid Arthritis (RA) is a chronic autoimmune disease that causes joint pain, stiffness, swelling and decreased joint motility. RA affects an estimated 1.28-1.36 million Americans.¹ RA mostly affects women at the age of 40-50 years.² Symptoms typically appear on both the left and right sides of the body symmetrically in the extremities such as hand and finger joints. While there is no cure for RA, treatment is possible to minimize symptoms. Common treatments aim to reduce inflammation and pain. Physical and occupational therapy can be used to increase mobility and strength of joints.³ Without treatment, deformation of the joints may lead to serious limitations.

The presence of specific autoantibodies is a hallmark of RA.⁴ RA is characterized by the production of rheumatoid factors (RFs) and antibodies against citrullinated proteins, or anti-citrullinated protein antibodies (ACPAs). Antibody production results in the formation of immune complexes and subsequent joint inflammation. RA-associated antibodies precede the onset of symptoms and predict disease course.⁴ RF is found in serum and synovial fluid (SF) of most RA patients and is associated with a more aggressive disease course. ACPAs are highly specific for RA and are associated with a severe, erosive destructive disease course.⁴

As a majority of RA patients exhibit antibodies against RFs and ACPAs, they represent useful markers of the disease.⁴ In this article, we will describe how the detection of RA-specific antibodies can be used for the diagnosis of RA. Additionally, we will discuss the advantages of antibody detection for disease subtyping and treatment monitoring.

Diagnostic criteria for RA

Since the prognosis of RA is dependent on prompt treatment, early diagnosis is critical. In 2010, the American College of Rheumatology (ACR) and the European League Against Rheumatism (EULAR) developed an international classification criterion for RA.5 These criteria recommend the determination of both RFs and ACPAs, along with clinical symptoms. Clinical and laboratory findings are weighted according to a point system and the diagnosis RA is either excluded, uncertain or definitive, depending on the score.⁵

Role of ACPA and RF levels in the diagnosis of RA

RF was the first antibody to be discovered in RA patients. RF exists as IgM, IgG and IgA subtypes but is most commonly measured as the IgM form.⁶ RF testing has a high sensitivity (60-90%) and a variable specificity (48-92%).⁷ RF specificity is limited because RF can be found in healthy populations and other autoimmune diseases.⁷ The specificity of RF testing can be increased at high titers. The role of RF in diseases pathogenesis has been demonstrated previously.⁷ Several proposed mechanisms of RF-induced pathogenesis involve the potentiation of inflammation and antigen trapping in joints.⁸

ACPA levels are detected in two thirds of RA patients and have a high diagnostic specificity (98%).⁸ ACPA levels increase during the transition from early undifferentiated arthritis to RA and can be detected in patient serum up to 14 years prior to the onset of symptoms.⁹ In RA, ACPAs can be present in different isotypes including IgG, IgA, IgM and IgE but are mainly IgG. To detect ACPA diagnostically, artificial cyclic citrullinated peptides (CCPs) are used as target antigens. Second-generation ELISAs have been developed for the detection of antibodies directed against citrullinated epitopes (CCP2) and demonstrate high sensitivities (69.6%-77.5%) and specificities (87.8%-96.4%) for RA.¹⁰ Antibodies against citrullinated vimentin, a member of the ACPA family, can be measured using an ELISA with the Sa antigen.¹¹ Anti-Sa antibodies have been proposed to be a promising marker of RA disease severity.¹²

Anti-CCP antibodies exhibit a higher predictive value for RA compared to RF (79% vs 56%).¹³ Both antibodies are useful predictive indicators for the development of erosions and the degree of radiological progression in RA.¹⁴ One study found a mechanistic link by which RF enhances the pathogenicity of ACPA-immune complexes in RA.¹⁵ The combined presence of ACPA and IgM-RF was found to mediate increased proinflammatory cytokine production in vitro and is associated with increased systemic inflammation and disease activity in RA.¹⁵

An additional use for antibody testing is for monitoring RA risk in patients with Porphyromonas gingivalis infection. Porphyromonas gingivalis mediates citrullination of host peptides and, therefore, the generation of RA-associated antibodies such as anti-CEP-1 (discussed below).¹⁶

Other novel target antigens in RA

Additional antigenic targets have been identified in RA. Antibodies against citrullinated -enolase peptide-1 (anti-CEP-1) have been observed in 37%–62 % of patients with RA.¹⁷ CEP-1 testing has been shown to have a high specificity (97.6%) for RA. Anti-CEP-1 antibodies are associated with an erosive disease course and with interstitial lung diseases.¹⁸ While CEP-1 testing may not be able to replace CCP2 testing, it may be useful as a supplementary test for confirmation of serological findings and for disease subtyping. Anti-CEP-1 antibodies occur in a subtype of RA associated with smoking and genetic factors.¹⁹ Anti-CEP-1 is an early marker that can predict the onset of symptoms ≥10.5 years.²⁰

Carbamylation is a post-translational modification responsible for the conversion of lysine into homocitrulline. In some people, extensive carbamylation can trigger an autoimmune response against carbamylated proteins. Antibodies against proteins that contain homocitrulline residues (anti-CarP) have been detected in 45% of RA patients and can occur in patients who are negative for ACPAs.²¹ Anti-CarP antibodies are highly specific for RA as they are not found in healthy patients or those with other inflammatory rheumatic conditions.²² Anti-CarP antibodies can be detected more than 10 years before disease onset and, therefore, are useful for early disease identification.⁴

Several additional RA-associated antibodies have been described including anti-PAD4, anti-BRAF, anti-RA-33, and others directed at post-translationally modified proteins.⁴ PAD4 is responsible for the conversion of arginine into citrulline. PAD4 antibodies have been found to occur in RA patients and are associated with severe disease. PAD4 antibodies occur along with ACPA and can be found in 22%-45% of RA patients.²³ Anti-BRAF antibodies are present in 21%-32% of RA patients but have also been found in similar frequency in other autoimmune disorders indicating that anti-BRAF antibodies are not specific for RA.⁴

Conclusions and ongoing research

As of 2021, several autoantibodies have been identified in RA, many of which are directed against post-translationally modified proteins. In 2010, the international classification criteria for RA recognized the importance of RF and ACPAs in identifying RA disease.⁵ The presence of RF and ACPA levels provides information about the disease course and gives other pathophysiological information. ACPA testing is highly sensitive for RA and has a high predictive value for disease onset and severity. The identification of antibodies other novel target antigens such as CEP-1, CarP, BRAF etc., may be useful as a complement to RF an ACPA testing.

RA management has been improved through disease-modifying antirheumatic drugs (DMARDs). ACPA status predicts response to therapy and with treatment, ACPA titers have been shown to decrease. DMARD treatment has been found to cause a 25% or more reduction in ACPA in 50% of patients.²⁴ Changes in RF in response to treatment has been previously demonstrated and research has shown that RF positivity predicts better response to rituximab and tocilizumab treatment.^7,25 Still more research is needed to demonstrate the usefulness of RF testing in treatment monitoring.

As a small percentage of RA patients may be seronegative for the typical RA-associated antibodies, it is important for scientists to continue to search for new biomarkers in RA. As an early and accurate diagnosis is important for patient prognosis, routine testing for a spectrum of biomarkers is advisable so that RA cases are not missed.

References

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