Sensitive, Specific Novel Biomarker Developed for Rheumatoid Arthritis

Through comprehensive antibody profiling utilizing a high-density peptide array, investigators were able to successfully construct a novel diagnostic biomarker for rheumatoid arthritis (RA) with high specificity and sensitivity compared with commercially available assays, according to research published in Arthritis and Rheumatology.

Researchers utilized a comprehensive serum antibody profile to create a report that is an “unbiased and comprehensive” profiling of serum antibodies in patients with RA. Using a high-peptide array, investigators profiled against the human proteome, including the citrullinome and the homocitrullinome. Serum samples were collected from both healthy individuals and patients with RA who fulfilled the American College of Rheumatology 2010 RA classification criteria.

Within a cohort of 18 RA serum samples, antibody reactivity against citrullinated peptide probes was detected at high frequency. Nearly 15,000 citrullinated peptide probes (n=14,953; 1.1% of total citrullinated probes) demonstrated significant reactivity at high frequency in RA samples. This, according to investigators, may potentially represent a diagnostic biomarker.

In contrast, antibody reactivity for both native and homocitrullinated peptide probes was observed at a “substantially lower frequency,” with only 10 native and 2 homocitrullinated peptide probes demonstrating a significant reactivity at >50% frequency in samples from patients with RA. Compared with control samples, the mean number of peptide probes with significant reactivity in RA samples was not statistically different for native and homocitrullinated peptide probes.

Hierarchal clustering analysis indicated that 12 of 18 RA samples demonstrated consistent peptide level reactivity against citrullinated peptide probes, which “readily formed a distinctive group of RA patients from controls.”

Researchers also found that composite epitopes may have potential implications for prognosis or diagnosis in RA. Frequently occurring composite epitopes against citrullinated peptide probes range from 20 to 44 amino acids in size (2 and 7 contiguous peptides, respectively), with a mean of 26.7 amino acids.

Investigators also explored the possibility of developing a novel RA diagnostic biomarker based on peptide array–derived data. Peptides were chosen based on the minimum required number to nonredundantly identify RA samples from control samples. With a signal threshold set at >95% specificity, researchers found that the overall diagnostic performance of an 8-epitope biomarker yielded 96.6% specificity and 92.1% sensitivity in an initial cohort of 92 samples (63 RA and 29 controls). Comparatively, a commercially available CCP2 assay yielded specificity and sensitivity of 96.6% and 68.3%, respectively.

To validate this diagnostic performance, an independent cohort of 181 additional serum samples (127 RA and 54 control) was commercially obtained. Overall, the diagnostic performance was 94.4% specificity and 85% sensitivity, compared with 96.3% specificity and 53.5% sensitivity in the CCP2 assay.

Study limitations include potential shortcomings of available peptide synthesis technology, as well as the small number of samples utilized in the development of the 8-epitope biomarker.

“As demonstrated by our 8-epitope diagnostic biomarker, it is evident that whole proteome profiling of linear epitopes has potential utility in revealing novel insights for disease prognosis,” the researchers concluded.

Disclosure: This study was funded by Roche Sequencing Solutions. Several study authors declared affiliations with the pharmaceutical industry. Please see the original reference for a full list of authors’ disclosure.

Reference

Lo KC, Sullivan E, Bannen RM, et al. Comprehensive profiling of rheumatoid arthritis antibody repertoire [published online August 26, 2019]. Arthritis Rheumatol. doi: 10.1002/art.41089