Novel Therapeutic Strategies in Sjögren’s Syndrome: B-Cell Targeting

1

Therapeutic Potential for B-Cell Modulation in Sjögren’s Syndrome

Critical dysregulated immune pathways, including the key role of B cells as well as the ectopic lymphoneogenesis, are involved in primary Sjögren’s syndrome (pSS) pathogenesis. B cells may provide a critical link between the development of tertiary lymphoid tissue within target tissues and the propagation of the autoimmune process. Although the peculiar role of B cells in autoimmunity is the production of autoantibodies, some data suggests that B cells may also exert additional pivotal functions such as antigen presentation and release of specific cytokines with immune regulatory, proinflammatory, polarizing, and tissue-organizing functions. In particular, a growing body of evidence has pointed out that B cells play a central role in the development, maintenance, and progression of the disease, with multiple roles at different points of pSS pathophysiology. B-lymphocyte hyperactivity, minor salivary gland (MSG) infiltration, and the development of B-cell follicles containing germinal center (GC)–like structures, represent the hallmarks of the disease. Excessive B-cell activation is responsible for a number of the extraglandular manifestations and serological features of pSS, including hypergammaglobulinemia, cryoglobulinemia, elevated levels of free light chains and β2-microglobulin, presence of anti-Ro/SSA and anti-La/SSB autoantibodies or rheumatoid factor (RF), hypocomplementemia, hypergammaglobulinemic purpura, arthritis, vasculitis, neuropathy, and glomerulonephritis. Finally, prolonged B-cell survival and aberrant B-cell activity may lead to the development of non-Hodgkin lymphoma in 5% of pSS patients.

Most pSS patients have a clinical pattern mainly dominated by severe dryness, fatigue, and pain, which although not life threatening, seriously impacts the quality of life. In contrast, systemic involvement plays a key role in the disease prognosis. In particular, Ioannidis et al. first proposed a prognostic classification, dividing pSS patients into two groups according to the presence or absence of risk factors, such as palpable purpura and low C4 levels. Recently, Baldini et al. found that low C3/C4, hypergammaglobulinemia, RF, and cryoglobulinemia are markers of severity in pSS, and the prevalence of the high-risk pSS patient subset for severe systemic manifestations was about 15%. Quartuccio et al. demonstrated for the first time that among pSS patients with salivary swelling, only those with low C4, cryoglobulins, anti-La/SSB antibodies, and leukopenia had an increased risk of lymphoma evolution. This data suggests three important concepts: (1) the subset of patients at high risk of systemic complications is specifically characterized by an active serological profile, suggestive for B-cell chronic activation; (2) serological markers of disease severity should be considered in the management of pSS; and (3) patients with this clinical or immunological higher risk pattern should receive closer follow-up observation and an earlier and more aggressive immunosuppressive treatment.

Recent systematic reviews highlighted the lack of evidence-based recommendations for the majority of the drugs commonly employed in the spectrum of extraglandular involvement and, at last, pSS may be still considered an orphan disease. Besides conventional immunosuppressive compounds, efficacy of targeted therapies in other systemic autoimmune diseases such as rheumatoid arthritis and systemic lupus erythematosus (SLE) suggested their possible use in pSS as well. Indeed, the treatment with immunosuppressive and biological agents in pSS is mainly based on their efficacy in the aforementioned conditions, expert opinion, and uncontrolled studies. Although many clinical manifestations may be shared among different systemic autoimmune diseases, it should be kept in mind, however, that the underlying pathogenic mechanisms could be different. This data points out the need for randomized clinical trials to investigate the safety and efficacy of these drugs in pSS in order to provide solid scientific evidence. Taken together, the difficulty in building therapeutic recommendations in pSS may be related to the heterogeneity of the clinical picture, the common failure of first line treatments, the lack of scientific evidence for drugs licensed for other diseases, and, finally, the lack of innovative therapeutic compounds. The need for more effective therapies with less toxic side effects as well as new insights into B-cell roles in disease pathogenesis has propelled interest in targeted biological therapies on the basis of an expanding understanding about disease pathogenesis. In this context, specific therapies with monoclonal antibodies (mAbs) recognizing targets on B-cell membrane (anti-CD20, anti-CD22), interfering with B/T cells, or inhibiting cytokines in B-cell development or activation such as B-cell activating factor (BAFF) and interleukin (IL) 6 or organization of ectopic lymphoid structures as shown for lymphotoxin (LT) β were proposed ( Fig. 18.1 ).

2

Anti-CD20 Antibodies in pSS Treatment: The Role of Rituximab

The CD20 (human B-lymphocyte–restricted differentiation antigen, Bp35) antigen is the most widely studied target for depleting B cells. This transmembrane molecule is found on the surface of most B cells, including pre-B and mature B lymphocytes, but not on stem cells, pro-B cells, normal plasma cells, or other normal tissues. CD20 mediates activation, proliferation, and differentiation of B cells, and may participate in the generation of T-cell independent antibody response. Many therapeutic mAbs targeting CD20 are currently available: rituximab (RTX), ocrelizumab, afutuzumab, ibritumomab tiuxetan, ofatumumab, TRU-015, tositumomab, and veltuzumab. Generally, anti-CD20 mAbs are classified as type I (RTX-like) or type II (tositumomab-like), based on their ability to redistribute CD20 molecules in the plasma membrane and activate various effector functions. Type II mAbs are mostly employed in the treatment of B-cell malignancies owing to their longer depletion of B cells from the peripheral blood (PB) and secondary lymphoid organs. To date, the only anti-CD20 monoclonal antibody that has been tested in pSS is RTX.

RTX, a chimeric (murine/human) mAb directly targeting CD20, prevents B-cell proliferation and induces depletion of circulating B cells from PB, salivary glands, and other target tissues by complement-mediated cytotoxicity, antibody-dependent cell-mediated cytotoxicity, or induction of apoptosis.

Early studies on the efficacy of RTX reported the histological evidence of a B-cell depletion and a reduction in glandular inflammation combined with a disappearance of GCs. It is worth noting that the histological findings paralleled the increase of parotid salivary flow and the normalization of the salivary sodium content, pointing out that the efficacy of B-cell depletion may also induce a potential glandular restoration in pSS.

The recovery of B-cell subsets after RTX treatment in pSS patients seemed to recapitulate B-cell ontogeny. The first B cells appearing in PB are transitional type 1 (T1) B cells (CD19+CD5+IgD+CD38++) and plasmablasts (CD19+CD5−IgD−CD38++), followed by a further increased of naive B cells migrating from the bone marrow (BM). Memory B cells were detected early during repopulation, first in PB and only later in MSGs, although the number of memory B cells remained relatively lower in both PB and MSGs. Sequential MSG biopsies revealed that B cells were absent in these glands for 12 months, but they colonized the affected glands 24 months after RTX treatment. Memory and T1 B cells were the first B cells identified locally, and, interestingly, the GCs previously seen in the MSGs were no longer present after B-cell recovery. The sole difference among the treated patients was in the timing of B-cell reappearance and, in this respect, higher baseline serum levels of BAFF inversely correlated with the duration of B-cell depletion, which resulted in the reconstitution of the preexisting abnormalities in PB. Moreover, after RTX treatment, circulating CD19+ B-cells started to reappear at week 24 and were partially or fully reconstituted 36 to 48 weeks after treatment. The large majority of the B cells that reappeared had a phenotype of transitional B cells and, interestingly, they did not derive from mature naive or memory peripheral B cells that were not depleted by RTX, but were newly generated B cells in the BM. In addition, the percentages and the absolute numbers of regulatory T (Treg) cells and effector T cells, as well as the ratio of effector T/Treg cells, did not significantly change after RTX treatment.

Beyond its direct activity on the B-cell compound, recent studies seem to further highlight the role of RTX in modifying the immunological microenvironment of inflamed salivary glands, suggesting new possible mechanisms: (1) reduction of the local expression of IL-22, interfering with the IL-22/IL-22RA1 axis, an important step in the emergence of T- and B-cell lymphoma ; (2) reduction of the expression of IL-17 and specifically induction of a pronounced apoptotic depletion of mast cells ; (3) reduction of serum levels of different molecules, including granulocyte macrophage colony-stimulating factor, IL-1Ra, IL-6, IL-10, interferon α, tumor necrosis factor (TNF) α, CCL4, and CXCL9, as indirect effects of B-cell depletion ; (4) induction of a significant decrease of inflammation proteins, cytokines, and growth factors released by epithelial cells, decreasing nuclear factor κB (NF-κB) activity and interrupting the NF-κB signaling pathway through the up-regulation of the Raf-1 kinase inhibitor protein ; and (5) interference with ectopic lymphoneogenesis not only by depleting B cells but also by tuning the delicate equilibrium between cells, molecules, and receptors, partially affecting the pro-B-cell inflammatory milieu. Specifically, we recently showed a reduction of lymphocytic foci and in the Chisholm and Mason score in the majority of patients treated with RTX, providing additional insights in the biological effects of RTX. MSGs of these patients presented, after 120 weeks of treatment, a nonspecific chronic sialadenitis pattern or a full restoration of glandular architecture associated to the disappearance of GC structures. Moreover, we observed a consistent reduction at messenger RNA (mRNA) levels of CXCR4 and CXCR5 associated to a parallel increase of the CXCL12 and CXCL13 mRNAs after anti-CD20 therapy. Although LTα and LTβ were markedly reduced by RTX, BAFF was not affected by the biological treatment. Finally, our study showed, at a histological and molecular level, the strong effect of RTX in dissolving the immunological organization of the affected tissues, which was not observed with disease-modifying antirheumatic drugs (DMARDs) therapy.