To the editor:

B lymphocytes are increasingly assigned an important role in the pathogenesis of auto-and alloimmune diseases including chronic graft-versus-host disease (cGVHD)1-4  where abnormalities of B-cell homeostasis in patients with active cGVHD have been observed.1,4  Extracorporeal photopheresis (ECP) achieves high response rates in steroid-refractory cGVHD.5  At present, no reliable biomarkers are available for prediction of response to ECP and monitoring of patient outcome. We thus investigated the distribution of immature CD19+CD21 B cells and memory CD19+CD27+ B cells in peripheral blood (PB)1  before ECP (n = 19), 6 (n = 28), 12 (n = 34), and 21 months (n = 33) after start of ECP in 49 patients (28 male, 21 female) with moderate (n = 25) or severe (n = 24) cGVHD6  and correlated results with clinical response.7  Informed consent was obtained in accordance with the Declaration of Helsinki. Eighteen patients were included in a prior report.1 

Complete (CR) and partial response (PR) to ECP was observed in 25 of 34 patients (74%) after 12 months. ECP nonresponders at 6 months had a significantly higher (P = .02) percentage of immature CD19+CD21 B lymphocytes (mean 22%, range 4%-52%) in PB before start of ECP compared with CR (mean 8%; range 1.3%-29%) and PR (mean 16%; range 1.6%-55%) patients (Figure 1A). Percentages of memory CD19+CD27+ B cells before first ECP were not significantly different (P = .12) between the groups. The CD21/CD27+ ratio was significantly higher (P = .03) in ECP nonresponders (mean 17.4; range 0.5-50) compared with CR patients (mean 1.6; range 0.3-5).

Figure 1

Comparison of relative amounts of immature CD19+CD21 B lymphocytes between ECP responders and ECP nonresponders. (A) Low percentages of immature CD19+CD21 B cells before therapy correlate significantly with complete resolution of chronic graft-versus-host disease to extracorporeal photopheresis (ECP). Patients were divided into 3 groups, complete responders (CR, n = 6), partial responders (PR, n = 6), and nonresponders (NR, n = 7) 6 months after start of ECP. Immature CD19+CD21 B cells assessed before start of ECP are shown in box plot format. Numbers indicate mean percentages (bold horizontal lines). Comparison of groups was performed using unpaired Student t test. (B) Complete ECP responders have significantly lower percentages of immature CD19+CD21 B cells 6, 12, and 21 months after start of ECP therapy compared with ECP nonresponders. Patients with complete response (CR; black bars), partial response (PR; white bars) and no response (NR; gray bars) to ECP are shown. Results are shown in box plot format. The bold horizontal line indicates the mean percentages. Comparison of groups was performed using unpaired Student t test.

Figure 1

Comparison of relative amounts of immature CD19+CD21 B lymphocytes between ECP responders and ECP nonresponders. (A) Low percentages of immature CD19+CD21 B cells before therapy correlate significantly with complete resolution of chronic graft-versus-host disease to extracorporeal photopheresis (ECP). Patients were divided into 3 groups, complete responders (CR, n = 6), partial responders (PR, n = 6), and nonresponders (NR, n = 7) 6 months after start of ECP. Immature CD19+CD21 B cells assessed before start of ECP are shown in box plot format. Numbers indicate mean percentages (bold horizontal lines). Comparison of groups was performed using unpaired Student t test. (B) Complete ECP responders have significantly lower percentages of immature CD19+CD21 B cells 6, 12, and 21 months after start of ECP therapy compared with ECP nonresponders. Patients with complete response (CR; black bars), partial response (PR; white bars) and no response (NR; gray bars) to ECP are shown. Results are shown in box plot format. The bold horizontal line indicates the mean percentages. Comparison of groups was performed using unpaired Student t test.

Percentages of immature CD19+CD21 B lymphocytes were significantly (P < .001) lower in CR patients compared with ECP nonresponders 6 (mean 5% vs 25%; range 1%-15% vs 14%-43%), 12 (mean 6% vs 24%, range 1.2%-16% vs 1.5%-58%), and 21 (mean 6% vs 26%; range 1%-21% vs 2%-60%) months after start of ECP (Figure 1B). In addition, in ECP responders compared with ECP nonresponders the CD21/CD27+ ratio was significantly lower 6 months (mean 2.2 vs 9.6; range 0.1-6.5 vs 1.2-20; P = .001), 12 (mean 2.7 vs 6; range 0.2-15 vs 0.6-16; P = .006), and 21 months (mean 2 vs 17; range 0.2-9 vs 0.6-76; P = .001) after start of ECP.

To our knowledge, relative amounts of immature CD19+CD21 B lymphocytes represent the first reported cellular biomarker able to predict response to ECP. Whether our findings are ECP-specific has to remain speculative because no other treatment cohorts were analyzed and the mechanisms of action of ECP are still subject to further research.5  CD19+CD21 B lymphocytes could also serve as a novel biomarker for measuring disease activity of cGVHD quantitatively and objectively, which would be highly desirable.8  Of note, CD21 B lymphocytes are increased in proportion in autoimmune diseases such as systemic lupus erythematosus and active cGVHD.1,10  Increased proportions of CD21 B lymphocytes could be part of the autoimmune pathogenesis compatible with inefficient censoring of autoreactive B cells in cGVHD.9  Disrupted B-cell homeostasis and relative rather than absolute preponderance of an activated B-cell pool has recently been reported in cGVHD.4  Our findings warrant larger prospective studies analyzing the predictive value of B-cell subsets for successful ECP treatment of cGVHD.

Authorship

Contribution: H.T.G. and W.F.P. designed the research study, analyzed and interpreted the data, and coauthored the manuscript; Z.K., D.P., and M.K. performed the clinical research, collected and analyzed data; R.W., U.K., and A.R. performed the flow cytometric analyses; R.K. and N.W. performed the ECP treatments; and C.Z. interpreted the data and contributed to the manuscript.

Conflict-of-interest disclosure: H.T.G. and R.K. have served as consultants to Therakos, Inc and participated as lecturers in the speakers bureau for Therakos. The remaining authors declare no competing financial interests.

Acknowledgments: This work was supported by European Commission Grant MCRTN-CT-2004-512253 TRANSNET, by 037703 STEMDIAGNOSTICS and a research grant of the Austrian Society for Transplantation Austrotransplant.

Correspondence: Hildegard T. Greinix, MD, Medizinische Universitaet Wien, Klinik fuer Innere Medizin I, Knochenmarktransplantation, Waehringer Guertel 18-20, A-1090 Vienna, Austria; e-mail: hildegard.greinix@meduniwien.ac.at; or Winfried F. Pickl, MD, Institut fuer Immunologie, Medizinische Universitaet Wien, Borschkegasse 8, A-1090 Vienna, Austria; e-mail: winfried.pickl@meduniwien.ac.at.

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