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New Assay Sheds Light on New Strategies for Arthritis Drug

NEW YORK (GenomeWeb) – A team from the Netherlands, the UK, and the US has developed a so called "drug-tolerant assay" to track tumor necrosis factor (TNF) dynamics in rheumatoid arthritis patients treated with the monoclonal antibody adalimumab, a TNF inhibitor.

Using the test, they have garnered evidence that TNF concentrations in the blood do not necessarily reflect response to the drug. 

The researchers relied on their competition enzyme-linked immunosorbent assay (ELISA) to quantify circulating TNF levels in 193 individuals with rheumatoid arthritis and 30 healthy volunteers who received at least one dose of adalimumab (or a biosimilar, in the case of control individuals). The findings, appearing online Wednesday in Science Translational Medicine, pointed to a broad TNF increase in the blood of both rheumatoid arthritis-affected and healthy individuals that was not tied to successful disease control.

The new assay, according to the study's abstract, "accurately measures how arthritis patients respond to treatment with the arthritis drug adalimumab, a finding that highlights potential strategies for predicting the drug’s effectiveness in patients."

Theo Rispens, an immunopathology researcher at Sanquin Research in Amsterdam, and his colleagues wrote, "[L]ongitudinal TNF concentrations are mostly stable during adalimumab treatment and may therefore not predict successful treatment discontinuation."

There was an exception to that pattern, though: the team found that rheumatoid arthritis patients who were adalimumab non-responders — those with reduced remission rates after around three months — tended to have low levels of circulating TNFs and enhanced levels of antibodies developed against the drug. Similarly, low TNF in healthy controls not long after treatment corresponded with antidrug antibody (ADA) production.

"Early (week 4) low TNF concentrations in patients with [rheumatoid arthritis] were associated with less frequent remission after 52 weeks," the authors wrote, noting that TNF concentrations at this stage "may be developed as a biomarker to predict future ADA formation and to identify non-responders in the early phase of treatment."

TNF, a regulatory cytokine that influences inflammation, has become a target in rheumatoid arthritis and other immune-mediated inflammatory diseases, the team explained. But the relationship between blood levels of TNF and disease symptoms has been difficult to discern, in part owing with difficulties in reliably quantifying TNF, which is unstable, found at low concentrations in the blood, and cleared quickly.

Still, past studies hint that TNF levels might offer insights into TNF inhibitor response, the authors added, suggesting it might be a way to identify rheumatoid arthritis patients who can discontinue treatment after achieving remission.

"We expected an overall increase in circulating TNF in the first phase of treatment, followed by a decrease in TNF over [two] years of follow-up in patients who are in clinical remission," the authors explained. "After that, monitoring TNF during anti-TNF treatment could be a potential biomarker in predicting successful treatment discontinuation."

To address these questions, the researchers came up with a quantitative, drug-tolerant, competition ELISA approach that relied on a "biotinylated high-affinity adalimumab mutant" for detecting TNF, including TNF that had been bound to the drug antibodies.

The assay "quantified total TNF concentrations during adalimumab treatment, which mainly comprises inactive, drug-bound TNF," the team explained.

After validating their approach and hammering out TNF detection limits with blood serum samples from dozens of healthy volunteers, the researchers applied it to blood samples collected from 193 rheumatoid arthritis patients at four weeks, 16 weeks, 28 weeks, 40 weeks, one year, 1.5 years, and two years after starting adalimumab treatment. 

The team found relatively stable TNF concentrations in patients following the initial post-treatment TNF jump, for example, though blood concentrations of TNF varied considerably from one individual to the next. Similarly, TNF levels held steady in blood samples from 21 patients who shifted from weekly adalimumab treatment to treatment every three weeks, despite waning concentrations of the drug itself.

On the other hand, the researchers did see declining TNF levels in 11 individuals with rheumatoid arthritis who discontinued adalimumab treatment over the course of the study, though TNF concentrations beyond those observed in early treatment were not associated with disease activity. 

"Overall, these findings indicate that TNF cannot be used as a biomarker for treatment discontinuation," the authors concluded. "However, early, low TNF concentrations can be used as an indicator to predict future ADA formation in the early phase of treatment."