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. 2004 Sep;137(3):513–520. doi: 10.1111/j.1365-2249.2004.02559.x

Amelioration of lupus manifestations by a peptide based on the complementarity determining region 1 of an autoantibody in severe combined immunodeficient (SCID) mice engrafted with peripheral blood lymphocytes of systemic lupus erythematosus (SLE) patients

N MAUERMANN *,†, Z STHOEGER †‡, H ZINGER *, E MOZES *
PMCID: PMC1809128  PMID: 15320900

Abstract

A peptide based on the complementarity determining region (CDR)1 of a human monoclonal anti-DNA autoantibody (hCDR1) was shown to either prevent or treat an already established murine lupus in systemic lupus erythematosus (SLE)-prone mice or in mice with induced experimental SLE. The present study was undertaken to determine the therapeutic potential of hCDR1 in a model of lupus in severe combined immunodeficient (SCID) mice engrafted with peripheral blood lymphocytes (PBL) of patients with SLE. To this end, PBL obtained from lupus patients were injected intraperitoneally into two equal groups of SCID mice that were treated either with the hCDR1 (50 µg/mouse) once a week for 8 weeks, or with a control peptide. Mice were tested for human IgG levels, anti-dsDNA autoantibodies, anti-tetanus toxoid antibodies and proteinuria. At sacrifice, the kidneys of the successfully engrafted mice were assessed for human IgG and murine complement C3 deposits. Of the 58 mice transplanted with PBL of SLE patients, 38 (66%) were engrafted successfully. The mice that were treated with the control peptide developed human dsDNA-specific antibodies. Treatment with hCDR1 down-regulated the latter significantly. No significant effect of the treatment on the levels of anti-tetanus toxoid antibodies could be observed. Treatment with hCDR1 resulted in a significant amelioration of the clinical features manifested by proteinuria, human IgG complex deposits as well as deposits of murine complement C3. Thus, the hCDR1 peptide is a potential candidate for a novel specific treatment of SLE patients.

Keywords: lupus, PBL of patients, peptide treatment, SCID mice

INTRODUCTION

Systemic lupus erythematosus (SLE) is a non-organ-specific, T cell-dependent autoimmune disease that is characterized by the production of high-titre affinity-matured IgG anti-dsDNA autoantibodies. Other targets of the autoimmune response include nuclear proteins. The disease affects mainly women of childbearing age. Due to the systemic availability of the autoantigens, many tissues and organs are afflicted in SLE patients (e.g. dermal, haematological, renal, neurological, musculoskeletal manifestations are observed) [1,2]. There are several animal models for this disease, most of which are genetically based, where mouse strains develop spontaneously a SLE-like disease. The SLE-prone mice include (NZB×NZW) F1, MRL-lpr/lpr, Palmerston North (PN) and BXSB [3,4].

SLE can be induced in naive (not lupus-prone) mice by an active immunization with the human monoclonal anti-DNA antibody that bears the 16/6 idiotype (Id) or with the murine monoclonal anti-DNA 16/6Id, 5G12 antibody [5,6]. Immunized mice develop high levels of autoantibodies, and show SLE-related clinical manifestations (leukopenia, thrombocytopenia and renal impairment) [5,6]. It is noteworthy that high homologies were found between anti-dsDNA autoantibodies isolated from SLE-prone mice [(NZB×NZW) F1] and autoantibodies from 16/6Id immunized diseased mice [7].

Two peptides, based on the sequence of the complementarity determining regions (CDR) 1 and 3 of the pathogenic murine anti-DNA 16/6Id were synthesized. The peptides were shown to be immunodominant T cell epitopes in non-autoimmune (e.g. BALB/c, SJL) and in lupus-prone (NZB×NZW) F1 mice [8–10]. Treatment with the peptides ameliorated clinical manifestations and decreased autoantibody production of spontaneous and induced SLE [11–13]. Amelioration of clinical manifestations following treatment with the CDR-based peptides was associated with down-regulation of interferon (IFN)-γ, interleukin (IL)-10 and tumour necrosis factor (TNF)-α (the latter in the induced model of BALB/c mice) and with an up-regulation of the immunosuppressive cytokine transforming growth factor (TGF)-β [11,13].

As a result of the above findings two peptides (hCDR1 and hCDR3), based on the CDRs of the human anti-DNA 16/6Id, were synthesized [14]. All CDR-based peptides (of either murine or human origin) were shown to inhibit the in vitro proliferation of human peripheral blood lymphocytes (PBL) of SLE patients to stimulation with 16/6Id. The inhibition correlated with a reduction in IL-2 secretion and an up-regulated secretion of the immunosuppressive cytokine TGF-β [14], suggesting a mechanism of inhibition similar to that observed for the animal models [11,13].

Several studies have been published in which attempts were made to create a human SLE model by transferring peripheral blood lymphocytes (PBL) of lupus patients into severe combined immunodeficient (SCID) mice [15–17]. We have reported recently the successful development of two reproducible models of human SLE [18]. One model has been of human PBL engrafted severe combined immunodeficient (SCID) mice, whereas the second model of human/mouse chimera was based on the previously reported studies of Lubin et al. [19]. Some of the SLE serological (human anti-DNA antibodies) and clinical manifestations (proteinuria, immune complex deposits in kidneys) of SLE were observed in the successfully engrafted mice of both models [18]. Thus these models allow the evaluation of potential therapies for the treatment of lupus patients.

In the present study we investigated the in vivo immunomodulating effect of the peptide based on the CDR1 of the human anti-DNA 16/6Id (hCDR1) on SLE-like disease in SCID mice transplanted with PBL of SLE patients. We report here the beneficial specific therapeutic effects of weekly injections of the hCDR1 on the serological (human dsDNA-specific antibodies) and clinical (proteinuria, human IgG and mouse complement C3 deposits in the kidney) manifestations.

MATERIALS AND METHODS

Mice

Female SCID mice (BALB/c background) 5–8 weeks old, were obtained from the Jackson Laboratory (Bar Harbor, ME, USA). The Animal Care and Use Committee of the Weizmann Institute of Science approved the study.

Synthetic peptides

A peptide (designated hCDR1) with the amino acid sequence GYYWSWIRQPPGKGEEWIG, based on the complementarity determining region 1 (CDR1) of the human monoclonal anti-DNA autoantibody that bears the 16/6Id, was synthesized (solid phase synthesis by F-moc chemistry) by Polypeptide Laboratories (LA, USA). A peptide with the amino acids of the hCDR1 synthesized in a scrambled order (‘scrambled peptide’), SKGIPQYGGWPWEGWRYEI, was used as a control. hCDR1 (TV4710) is currently under clinical development for human SLE by Teva Pharmaceutical Industries Ltd.

SLE patients

Seven female SLE patients participated in this study. All fulfilled at least four of the ACR revised diagnostic criteria for SLE [20]. Patients were between 24 and 56 years old (mean 40·5 ± 13·4 years). All SLE patients had high levels of antinuclear antibodies (ANA) and anti-dsDNA antibodies in their sera at the time of the study. All patients (100%) had arthritis; six of them (86%) had haematological disturbances (two patients with haemolytic anaemia, two with thrombocytopenia and four with leukopenia). Three of the patients (43%) had renal involvement at some stage of their disease.

At the time of the study the disease activity index (SLEDAI [21]), was between 2 and 14 (mean 5·7 ± 5·12). One patient had active renal disease, two demonstrated lymphopenia, one thrombocytopenia and two had arthritis. Treatment modalities at the time of the study were prednisone (10–30 mg/day) in four patients, Plaquenil (400 mg/day) in two patients and methotrexate (7·5–10 mg/week) in two patients. All patients signed an informed consent form prior to their participation in the study, which was approved by the Ethics Committee of the Kaplan Medical Center, Rehovot, Israel.

Transplantation of human PBL and treatment

PBL obtained from SLE patients were injected intraperitoneally (i.p.) into 8–10-week-old recipient SCID mice at a concentration of 30 × 106 cells in 0·5 ml phosphate buffered saline (PBS). PBL of each donor were transferred into six to eight mice that were divided equally into two groups. One group was treated once a week, starting at the day of cell transfer with 50 µg hCDR1 given subcutaneously (s.c.) in PBS, whereas a control group was injected with the scrambled peptide. Mice were bled periodically and the sera were evaluated for the presence of human IgG, human anti-dsDNA antibodies and anti-tetanus toxoid (TT) antibodies.

Determination of human IgG

Levels of human IgG were measured by enzyme-linked imunosorbent assay (ELISA) [18], using a goat F(ab)2 purified antihuman IgG (Jackson ImmunoResearch Laboratories, West Grove, PA, USA) as the capture antibody and peroxidase-conjugated goat antihuman Ig (Jackson) as the detection antibody. Human IgG at a known concentration was used as a standard in all assays.

ELISA for human antibodies

For determination of human anti-dsDNA antibodies in the recipient mouse sera maxisorb 96-well microtitre plates were coated with poly l-lysine (5 µg/ml, Sigma, St. Louis, MO, USA), followed by coating with lambda phage dsDNA (5 µg/well, Boehringer Mannheim, Germany). Plates were then blocked with 10% fetal calf serum (FCS) in PBS, and the sera (diluted 1 : 5–1 : 40) were incubated for 2 h. Plates were washed and incubated with a goat-antihuman IgG antibody conjugated to horseradish peroxidase (Jackson) for 90 min. Following washing, plates were incubated with the substrate ABTS (2·2′azino-bis-3-ethylbenzthiazoline-6-sulphonic acid; Sigma) and read at 405 nm using an ELISA reader (Tecan Spectra Classic, Austria).

For the detection of human tetanus, toxoid-specific antibodies purified TT (5 µg/ml, kindly provided by RAFA Laboratories Ltd, Israel) was used for coating of microtitre plates (Nunc Roskilde, Denmark). Plates were then washed, blocked, incubated with serum samples (diluted 1 : 10–1 : 1280) and developed with a goat-antihuman IgG Fc-specific antibody conjugated to horseradish peroxidase. Plates were developed with ABTS. Results of anti-dsDNA and anti-TT are presented as OD per 1 mg of human IgG [18]. Positive and negative murine sera for dsDNA reactivity were further tested by Hep 2 (ANA) and by Crithidia luciliae (dsDNA) assays [18].

Proteinuria

Proteinuria was measured semiquantitively by using Albustix dipsticks (Ames Division, Bayer Diagnostics, Newbury, UK) on the following scale: 0 = undetectable, 1 = 0·3 g/l, 2 = 1 g/l, 3 = 3 g/l, 4 = ≥20 g/l.

Immunohistology of kidneys

Kidneys of sacrificed mice were frozen immediately in vials containing 0·5 ml isopentan (2-methylbutan) in liquid nitrogen. Kidney sections (6 micron) were air-dried and fixed in acetone. Staining with FITC-conjugated goat-antihuman IgG- (Jackson) and FITC-conjugated goat-antimouse complement C3 antibodies (Cappel, ICN Pharmaceuticals Inc., Aurora, OH, USA) were performed. Sections were visualized and graded, using a fluorescence microscope (Zeiss Axioskop 2, Germany).

Cytokine secretion and detection

Spleen cells (5 × 106/ml) of the tested mice were incubated with enriched medium and supernatants were removed after 48 h (for IFN-γ and IL-10) and 72 h (for TGF-β). Cytokine levels were measured by ELISA as described previously [13].

Statistical analysis

To test statistical significance for anti-dsDNA antibody production, proteinuria and kidney stainings between the treatment groups, the Mann–Whitney test with a two-tailed P-value and Fisher's exact test with two-sided P-value were performed.

RESULTS

Engraftment of PBL of SLE patients

Three separate experiments were performed in which SCID mice were injected i.p. with 30 × 106 PBL/mouse obtained from seven SLE patients. The recipients of PBL of each SLE patient were divided equally into two treatment groups that received weekly s.c. injections of either 50 µg hCDR1 or the control scrambled peptide (50 µg). Altogether, 58 recipient mice were injected with PBL of SLE patients. Engraftment of the human PBL in mice was determined based on the levels of human IgG in the mouse sera. Levels equal or above 100 µg/ml of human IgG indicated successful engraftment. Of the 58 mice transplanted with human PBL, 38 (66%) were engrafted successfully. Figure 1 demonstrates that the mean levels of human IgG produced by the engrafted PBL were about 500 µg/ml. It is also shown in Fig. 1 that the levels of human IgG were similar in the sera of mice that were treated with hCDR1 or scrambled peptide.

Fig. 1.

Fig. 1

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Levels of total human IgG in sera of successfully engrafted SCID mice. Mean (± s.e.) levels of human IgG in sera of SCID mice 4–5 weeks following administration of human PBL.

dsDNA-specific autoantibodies

During treatment of SCID mice, the titre of human anti-dsDNA antibodies was determined periodically. Figure 2 shows the kinetic of anti-dsDNA antibody production for two representative mice engrafted with PBL of the same lupus patient, one treated with hCDR1 and the other with the control peptide. It can be seen that the dsDNA-specific antibody levels peaked between days 30 and 40 following cell transplantation and then started to decline. Figure 2 also shows that treatment with hCDR1 down-regulated significantly the production of the human anti-dsDNA antibodies. Figure 3a shows the mean anti-dsDNA antibody levels (OD/1 mg human IgG) in the sera of all mice in the two groups at the time that antibody levels peaked. As can be seen, anti-dsDNA antibody titres were significantly lower (P = 0·0053) in the sera of the hCDR1-treated mice compared to sera of the control, scrambled peptide-treated mice. It is noteworthy that sera of 10 of the 21 (48%) SCID mice that were treated with the control peptide had levels of anti-dsDNA antibodies that were higher than 1 OD/1 mg of human IgG. In contrast, an OD of 1·03/1 mg IgG was observed in serum of only one of the 17 (6%) successfully engrafted mice that were treated with hCDR1. It should be noted that the results for dsDNA reactivity measured by ELISA correlated with the clinical assays for ANA and dsDNA reactivity measured by Hep 2 and C. luciliae assays. Thus, 90% of the sera determined to be positive for dsDNA reactivity by ELISA were determined to be positive by the clinical assays for ANA and dsDNA (≥ + +). Furthermore, as shown by ELISA, serum of only one mouse treated by hCDR1 was positive for ANA by the Hep 2 assay.

Fig. 2.

Fig. 2

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Kinetics of anti-dsDNA specific antibody production. Kinetics of human anti-dsDNA antibody production, shown for two representative mice transplanted with PBL of the same lupus patient that were treated either with hCDR1 or with the control peptide. Mice were bled every 10 days and human anti-dsDNA antibody levels were determined by ELISA. The results are presented as OD/1 mg of human IgG determined for the same sera samples.

Fig. 3.

Fig. 3

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Human anti-dsDNA and anti-tetanus toxoid antibodies in sera of SCID mice. Titres of human anti-dsDNA (a) and anti-tetanus toxoid (b) antibodies were determined by ELISA on the same sera samples (∼ 40 days following cell transplantation). Shown are values of the mean OD/1 mg human IgG (± s.e.) for the two groups of mice, treated with either hCDR1 or the control peptide. *P = 0·0053.

Anti-tetanus toxoid antibodies

In order to determine whether hCDR1 treatment is specific to SLE-associated antibodies and does not affect other immune responses, we studied its effect on anti-TT antibody levels. Figure 3b shows levels of the anti-TT antibodies in the sera of mice from both treatment groups. The results indicate that there was no significant decrease in the antibody titres specific to TT, when sera samples of hCDR1-treated SCID mice were compared to those of control peptide-treated mice. Thus, the effect of hCDR1 is restricted to SLE-associated responses.

Cytokine secretion

Supernatants of splenocytes obtained from mice of the two treatment groups were tested for the secretion of IFN-γ, IL-10 and TGF-β. The secreted levels of IFN-γ and IL-10 were low (below the detectable sensitivity of the assay, 15 pg/ml for both IFN-γ and IL-10).

However, the secretion of TGF-β was higher (1102 ± 249 pg/ml) in supernatants of splenocytes of hCDR1-treated mice than in supernatants obtained from cells of scrambled peptide-treated mice (751 ± 62 pg/ml). Although the differences between groups did not reach significance, an increased secretion of TGF-β following hCDR treatment was demonstrated clearly.

SLE-associated clinical manifestations

The levels of proteinuria were determined periodically in the SCID mice. Although the measured levels were relatively low (up to 1 g/l) in both treatment groups, Fig. 4 shows a significant (P = 0·0002) reduction in the mean proteinuria levels (Fig. 4a) and in the number of mice with detectable proteinuria (Fig. 4b) following treatment with hCDR1 (P = 0·0017).

Fig. 4.

Fig. 4

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Proteinuria in SCID mice engrafted with PBL of SLE patients. Levels of proteinuria were measured before sacrifice (∼ 60 days following cell transfer). Results are expressed as (a). Mean (± s.e.) proteinuria (g/l) for all mice within a treatment group *P = 0·0002. (b) Proteinuria score for individual mice in both treatment groups. **P = 0·0017

Kidney sections of all successfully engrafted mice were analysed for human IgG and mouse complement C3 depositions. The results summarized in Table 1 indicate that immune complex deposits were determined in 14 of 21 (66%) mice treated with the control peptide. In contrast, in kidney sections of only one of 17 (6%) mice treated with hCDR1 were human Ig immune complex deposits detected (P = 0·0001). The human IgG deposits in the glomeruli were associated with inflammatory glomerulonephritis, as demonstrated by the presence of murine complement C3 deposits. As also shown in Table 1 kidney sections of 11 of 21 (52%) control peptide-treated mice compared to only one of 17 (6%) hCDR1-treated mice had murine complement C3 deposits in their kidneys (P = 0·0023). It is noteworthy that the only kidney in the hCDR1-treated group that was positively stained for both human Ig and mouse complement C3 was that of the only mouse determined to have high dsDNA-specific antibody titre (1·03OD/1 mg of human IgG). The representative kidney sections which are shown in Fig. 5 demonstrate the absence of human Ig (Fig. 5d,e) and murine complement C3 (Fig. 5f) deposits in a kidney of a mouse treated with hCDR1 compared to the positive staining of the kidney of a control peptide-treated mouse (human Ig deposits Fig. 5a,b; murine complement C3 deposits, Fig. 5c).

Table 1.

Human Ig and murine complement C3 deposits in kidney sections of SCID mice engrafted with PBL of SLE patients and treated with hCDR1

Human IgG deposits Murine C3 deposits
Scrambled peptide hCDR1 Scrambled peptide hCDR1
66% (14/21) 5·8% (1/17)P = 0·0001 52% (11/21) 5·8% (1/17)P = 0·0023
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Frozen cryostat sections (6 micron) of kidneys were stained with FITC-labelled goat antihuman IgG (γ -chain specific) or with FITC-goat-antimouse complement C3.

Fig. 5.

Fig. 5

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Deposits of human IgG and mouse complement C3 in kidney sections of SCID mice engrafted with PBL of SLE patients. Frozen cryostat sections (6 micron) of kidneys (taken ∼ 60 days following cell transfer) were air-dried, fixed with acetone and stained with either (a, b,d, e) FITC-labelled goat antihuman IgG (γ-chain specific) or (c, f) FITC-labelled goat antimouse complement C3 antibodies. Shown are representative kidney sections of two mice, engrafted with PBL of the same SLE patient. (a, b, c) Kidney sections of a mouse treated with the control scrambled peptide; (d, e, f) kidney sections of a mouse treated with hCDR1 (a, d ×100 and b, e, c, f ×400).

DISCUSSION

The main findings of the present report are that a peptide based on the complementarity determining region (CDR)1 of a human monoclonal anti-dsDNA antibody that bears the 16/6Id is capable of ameliorating disease manifestations in SCID mice engrafted with PBL of SLE patients. Thus the peptide, hCDR1, has beneficial effects on a disease model that is the closest possible to human SLE.

We have reported previously successful attempts to establish an SLE model in SCID mice [18], showing then that engraftment with the PBL of SLE patients was successful in 71% of the recipient SCID mice. Similar results were obtained in the present study, where engraftment was successful in 66% of the recipients. The percentage of recipients with successful engraftment as well as the levels of human IgG measured in the engrafted mice was the same for mice that were treated either with the hCDR1 or with the control scrambled peptide (Fig. 1).

Production of dsDNA-specific antibodies is one of the hallmarks of SLE. Weekly injections of as little as 50 µg/mouse of the hCDR1 down-regulated significantly the levels of dsDNA-specific antibodies in the treated mice compared to mice engrafted with PBL of the same lupus donors that were treated with the control scrambled peptide (Figs 2 and 3a). The down-regulating effect of the hCDR1 on the human autoantibody levels was demonstrated further by using the standard clinical assays for ANA (Hep 2) and dsDNA (C. luciliae)-specific antibodies.

The optimal treatment for SLE, as well as for other autoimmune diseases, should down-regulate specifically SLE-associated responses without affecting other unrelated arms of the immune system. To this end, in the present study we tested the levels of the acquired antibody activity in the sera of recipient SCID mice to tetanus toxoid. The production of antibodies specific to tetanus toxoid by SCID mice engrafted with human PBL has been shown previously [18,22]. We showed here that treatment with hCDR1 did not affect the antibody levels to tetanus toxoid. Thus a comparable binding activity was determined in sera of recipients of PBL of the same donors that were treated with either hCDR1 or with the control scrambled peptide (Fig. 3b), suggesting that the down-regulating effect by the hCDR1 is indeed specific.

One of the major manifestations of lupus is kidney involvement. In the present study, 14 of the 21 (66%) successfully engrafted recipient mice that were treated with the control peptide developed immune complexes of human IgG in their kidneys. In the group that was treated once a week with 50 µgof hCDR1, only one of the 17 recipient mice developed immune complex deposits. Thus, treatment with hCDR1 abolished almost completely the kidney involvement in SCID mice with the lupus-like disease. No correlation could be determined between kidney disease in the lupus donor patients, in the past or at the time of the study, and the immune complex deposits in the recipient mice. These results are in agreement with our previous publication [18]. The human IgG deposits in the glomeruli were associated with inflammatory glomerulonephritis as suggested by the presence of proteinuria (Fig. 4a,b) which correlated with the glomerular IgG deposits. In addition, deposits of murine complement C3 were detected in the kidneys of 11 of the 14 SCID mice that were treated with the scrambled control peptide and had human IgG immune complexes (Table 1, Fig. 5). The ability of human Ig deposits to cause inflammatory glomerulonephritis by the interaction with mouse complement C3 in kidneys of SCID mouse/human model of lupus was reported previously by Duchosal et al. [15] as well as by us [18].

Previous studies utilizing the human/SCID mouse model dealt mainly with the production of anti-DNA antibodies and a limited number of other clinical symptoms that are characteristic for SLE [15–17]. As for treating SCID mice with lupus manifestations, it has been reported previously that anti-IL-10 monoclonal antibodies inhibited the production of anti-DNA antibodies in SCID mice engrafted with PBL of SLE patients [23]. Similarly, Kalechman et al. reported the reduction in autoantibody levels in a lupus model in SCID mice, following treatment with AS101 that down-regulates IL-10 [24]. The two latter studies utilized non-specific means that indeed down-regulated total immunoglobulin production, whereas the treatment with hCDR1 used in the present study inhibited specifically SLE-related responses without affecting antibody reactivity to tetanus toxoid and total IgG. More recently, Suzuki et al. reported the therapeutic effect of the chemically modified ribozyme (RZ-1) to target the V3-7 gene. In the reported model, PBL of patients with active lupus nephritis were treated in vitro with RZ-1 prior to their transfer to SCID mice. Beneficial effects were observed on the serological and clinical manifestations [25]. Unlike the in vitro manipulation performed in the latter report, treatment with hCDR1 was given in vivo by s.c. injections resembling possible treatment protocols of patients.

The peptides that are based on the CDR of the murine anti-DNA 16/6Id autoantibody as well as the hCDR1 were shown by us to be capable of ameliorating experimental SLE in induced and spontaneous animal models [8,11–13 and Luger et al., unpublished data]. Other laboratories have also reported the beneficial effects of peptides based on variable regions of autoantibodies or of a consensus peptide based on amino acid sequences of murine anti-DNA monoclonal antibodies on experimental models of SLE [26–30]. The latter reports as well as ours support the crucial role of peptides derived from heavy chain variable regions in the immunomodulation of murine lupus. We have shown previously that peptides based on the murine or human anti-DNA monoclonal autoantibodies down-regulated the in vitro anti-DNA-specific proliferative responses of PBL of SLE patients [14]. The present study shows that the hCDR1 is also efficient in the in vivo amelioration of disease symptoms manifested by PBL of SLE patients.

The mechanism by which the hCDR1 ameliorates SLE characteristic manifestations is currently studied. We have shown previously that treatment of SLE-afflicted mice with the CDR-based peptides down-regulates IFN-γ, IL-10 and proinflammatory cytokines (TNF-α, IL-1β). In contrast, the treatment leads to an up-regulated production of the immunosuppressive cytokine TGF-β [11,13]. We have shown further that the inhibition of proliferative responses of PBL of SLE patients is associated with an increase in the secretion of TGF-β [18]. In agreement, in the present study treatment with hCDR1 also up-regulated the secretion of TGF-β, suggesting that this immunosuppressive cytokine plays an important role in the mechanism by which the hCDR1 ameliorates SLE. Our recent studies indicate that hCDR1 up-regulates a population of CD4+CD25+ immunoregulatory cells that play a key role in the inhibitory activity of the hCDR1 (Mauermann et al., unpublished data). These cells may either secrete TGF-β by themselves or they may trigger other regulatory T cell populations to secrete this suppressive cytokine. Nevertheless, as shown here treatment with hCDR1 is beneficial in an in vivo model of human lupus. The hCDR1 is therefore a novel potential candidate for the specific treatment of SLE patients.

Acknowledgments

This work was supported by Teva Pharmaceutical Industries, Ltd, Israel.

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