Epstein-Barr virus in bone marrow of rheumatoid arthritis patients predicts response to rituximab treatment

Abstract

Objectives: Viruses may contribute to RA. This prompted us to monitor viral load and response to anti-CD20 therapy in RA patients.

Methods: Blood and bone marrow from 35 RA patients were analysed for CMV, EBV, HSV-1, HSV-2, parvovirus B19 and polyomavirus using real-time PCR before and 3 months after rituximab (RTX) treatment and related to the levels of autoantibodies and B-cell depletion. Clinical response to RTX was defined as decrease in the 28-joint disease activity score (DAS-28) >1.3 at 6 months.

Results: Before RTX treatment, EBV was identified in 15 out of 35 patients (EBV-positive group), of which 4 expressed parvovirus. Parvovirus was further detected in eight patients (parvo-positive group). Twelve patients were negative for the analysed viruses. Following RTX, EBV was cleared, whereas parvovirus was unaffected. Eighteen patients were responders, of which 12 were EBV positive. The decrease in the DAS-28 was significantly higher in EBV-positive group compared with parvo-positive group (P = 0.002) and virus-negative patients (P = 0.04). Most of EBV-negative patients that responded to RTX (75%) required retreatment within the following 11 months compared with only 8% of responding EBV-positive patients. A decrease of RF, Ig-producing cells and CD19(+) B cells was observed following RTX but did not distinguish between viral infections. However, EBV-infected patients had significantly higher levels of Fas-expressing B cells at baseline as compared with EBV-negative groups.

Conclusions: EBV and parvovirus genomes are frequently found in bone marrow of RA patients. The presence of EBV genome was associated with a better clinical response to RTX. Thus, presence of EBV genome may predict clinical response to RTX.

Fig. 1

RTX therapy clears EBV genome from blood and bone marrow. Titre of EBV in bone marrow and blood at baseline and 3 months following start of RTX therapy in RA patients expressing EBV at baseline (n = 15 of 35 RA patients). Individual values are represented by dots and the median by the horizontal line.

Fig. 2

Patients expressing EBV at baseline have a better clinical outcome to RTX therapy than EBV-negative patients. (A) DAS scores [median and interquartile range (IQR)] at baseline, 3 and 6 months post-initiation of RTX in patients expressing EBV (EBV positive), expressing only parvovirus (parvo positive) and expressing none of the analysed viruses (virus negative) in bone marrow (*EBV positive vs virus negative, P = 0.038). (B) Median reduction (and IQR) in DAS at 6 months in EBV-positive vs EBV-negative patients (*P = 0.049). (C) Per cent responders (defined as a decrease in DAS-28 > 1.3 at 6 months after initiation of RTX therapy) and proportion of responders receiving retreatment with RTX within 11 months from baseline in EBV-positive and EBV-negative patients. There is significantly more responders (height of the bars) in the EBV-positive group as compared with the EBV-negative group (**P = 0.0034) and the proportion of responders (the height of the black space in relation to the height of the bar) receiving retreatment within 11 months is significantly higher in the EBV-negative group (*P = 0.020).

Fig. 3

No significant difference in RF or anti-CCP between EBV-positive and EBV-negative patients. Levels of total RF (A) and anti-CCP (B) antibodies at baseline in EBV-positive and EBV-negative group. Individual values are represented by dots and the median by the horizontal line.

Fig. 4

No significant difference in RTX-mediated B-cell depletion efficiency between EBV-positive and EBV-negative patients. Per cent of B cells (of white blood cells) defined as CD19⁺ cells [(A) in blood and (B) in bone marrow] in EBV-positive and EBV-negative group at baseline and 1–3 months after RTX treatment. The line in each box represents the median, whereas the box represents the IQR with whiskers at ±1.5 IQR. Outliers are indicated.

Fig. 5

Higher baseline frequency of CD95-expressing B cells in EBV-positive than EBV-negative patients. Per cent CD95-expressing B cells in bone marrow of all B cells at baseline in EBV-positive and EBV-negative group. The line in each box represents the median, whereas the box represents the IQR with whiskers at ±1.5 IQR. Outliers are indicated. *P = 0.043.