Drs. You and Pan contributed equally to this work.
Effects of a novel tylophorine analog on collagen-induced arthritis through inhibition of the innate immune response
Version of Record online: 28 FEB 2006
Copyright © 2006 by the American College of Rheumatology
Arthritis & Rheumatism
Volume 54, Issue 3, pages 877–886, March 2006
How to Cite
You, X., Pan, M., Gao, W., Shiah, H.-S., Tao, J., Zhang, D., Koumpouras, F., Wang, S., Zhao, H., Madri, J. A., Baker, D., Cheng, Y.-C. and Yin, Z. (2006), Effects of a novel tylophorine analog on collagen-induced arthritis through inhibition of the innate immune response. Arthritis & Rheumatism, 54: 877–886. doi: 10.1002/art.21640
- Issue online: 28 FEB 2006
- Version of Record online: 28 FEB 2006
- Manuscript Accepted: 10 NOV 2005
- Manuscript Received: 3 AUG 2005
- Arthritis Foundation Investigator award
- NIH. Grant Numbers: K01-AR-02188, 1-R01-AI-056219
To test the effects of a novel tylophorine analog, DCB 3503, on the prevention and treatment of collagen-induced arthritis (CIA) and to elucidate its underlying mechanisms.
DBA/1J mice were immunized with type II collagen, and in some cases, lipopolysaccharide (LPS) was used to boost the development of arthritis. DCB 3503 was injected intraperitoneally before or after the onset of CIA. Mice were monitored to assess the effects of DCB 3503 on the clinical severity of the disease, and pathologic changes in the joints were examined histologically. Levels of tumor necrosis factor α (TNFα) and interleukin-1β (IL-1β) in serum and joint tissues were measured by enzyme-linked immunosorbent assay and by cytometric bead array analysis. The effect of DCB 3503 on LPS-induced proinflammatory cytokines from bone marrow–derived dendritic cells was determined by flow cytometry.
DCB 3503 significantly suppressed the development and progression of CIA. Moreover, DCB 3503 completely blocked the LPS-triggered acceleration of joint inflammation and destruction. Consistent with its effects in vivo, DCB 3503 significantly suppressed the synthesis of proinflammatory cytokines in inflamed joints as well as cytokine synthesis by macrophages examined ex vivo. Treatment also reduced the levels of inflammatory cytokines (IL-6, IL-12, TNFα, and monocyte chemotactic protein 1) produced by bone marrow–derived dendritic cells in vitro. However, DCB 3503 showed no direct effects on T cell proliferation and B cell antibody response.
Because of its ability to specifically suppress innate immune responses, DCB 3503 may be a novel therapeutic agent for inflammatory arthritis in humans.
Rheumatoid arthritis (RA) is a common systemic autoimmune disease that leads to joint inflammation and progressive cartilage and bone erosion (1–3). Although current treatments that specifically block a single cytokine, such as anti–tumor necrosis factor α (anti-TNFα) or anti–interleukin-1 (anti–IL-1), have proved clinically effective, such treatments are often insufficient to produce complete disease remission in some patients (4, 5). Consequently, there has been a great demand for new antirheumatic agents that are able to act on multiple cytokines or mediators of inflammation, but have fewer toxic effects. There are effective treatments for rheumatism available from practitioners of traditional Chinese medicine, including the medicinal plant Tylophora atrofolliculata, from the family Asclepiadaceae (6). Thus, studies exploring the herbs of traditional Chinese medicine may ultimately provide additional therapeutic agents for treating this chronic inflammatory disease.
Collagen-induced arthritis (CIA) is a well-studied animal model of RA (7). Although the precise mechanisms whereby immunization with type II collagen (CII) leads to the development of chronic arthritis are not known, there is considerable evidence implicating CII-specific CD4+ T cells as primary mediators of disease induction, with the production of complement-fixing anti-CII antibody being the major immune mechanism for the initiation of joint inflammation (8–10). In the joints, activated T cells and Th1-like cytokines induce macrophages to produce TNFα, which in turn, leads to infiltration by other inflammatory cells. These cells then release other cytokines (IL-1 and IL-6) and mediators of inflammation (inducible nitric oxide synthase and cyclooxygenase 2), resulting in persistent inflammation and joint destruction (3, 8, 11). The CIA model has proved useful in the development of new therapies for RA, such as anti-TNFα and anti–IL-1 (12).
Several alkaloid extracts have recently been isolated and synthesized from Tylophora species. One synthesized tylophorine analog, DCB 3503 (NSC-716802), has been shown to selectively inhibit TNFα-induced NF-κB activity and to suppress the growth of human tumor xenografts in nude mice (13). Since NF-κB is pivotal in joint inflammation, we hypothesized that DCB 3503 might also be beneficial in the treatment of inflammatory arthritis. To test our hypothesis, we used the DBA/1J (H-2q) mouse model of arthritis (14) and administered DCB 3503 at different time points postimmunization, either before or after the onset of CIA. We show herein that DCB 3503 significantly suppressed joint inflammation and destruction, especially in the case of lipopolysaccharide (LPS)–triggered acceleration of CIA. Furthermore, DCB 3503 inhibited the production of multiple mediators of inflammation ex vivo as well as by cultured dendritic cells in vitro, with little effect on T cells and B cells.
MATERIALS AND METHODS
Male DBA/1J mice were purchased from The Jackson Laboratory (Bar Harbor, ME) and maintained under specific pathogen–free conditions at the Yale University School of Medicine. All animal procedures were performed with the approval of the Institutional Animal Care and Use Committee of the Yale University School of Medicine.
Chemicals, antibodies, and cytokines.
Tylophorine analog DCB 3503 (NSC-716802) was synthesized in the laboratory of one of us (DB) (13). Antibodies for flow cytometry and enzyme-linked immunosorbent assay (ELISA) were purchased from BD PharMingen (San Diego, CA), except where indicated otherwise. Recombinant cytokines used for T cell activation and dendritic cell differentiation were purchased from R&D Systems (Minneapolis, MN), except where indicated otherwise.
CIA model and DCB 3503 treatment.
The CIA model was adapted as described previously (14). On day 0, male DBA/1J mice ages 8–10 weeks were injected at the base of the tail with bovine CII (Chondrex, Redmond, WA) emulsified in equal volumes of Freund's complete adjuvant (CFA) (containing 2 mg/ml of inactivated Mycobacterium tuberculosis; Chondrex). On day 21, a booster injection with an emulsion of 100 μg of bovine CII dissolved in Freund's incomplete adjuvant (IFA; Chondrex) was given. Thereafter, mice were monitored daily for signs of arthritis. Assessors had no knowledge of the group to which the mice belonged. Arthritis severity was graded as described previously (15). Briefly, each paw was scored individually on a scale of 0–3, where 0 = normal; 1 = mild/moderate, but definitely visible, swelling of 1 or more digits; 2 = severe erythema and swelling affecting an entire paw or joint; 3 = deformed paw or joint, with ankylosis and rigidity (significantly reduced hock joint motion on flexion/extension). Scores for all 4 paws were summed for each mouse (maximum score 12).
For the LPS-enhanced CIA model, LPS was injected intraperitoneally (50 μg/mouse) on day 28 postimmunization, as previously described (16).
To test the effects of DCB 3503 on CIA, mice were treated with either DCB 3503, or vehicle control at different time points postimmunization. Intraperitoneal injections of DCB 3503 or vehicle were administered at a dosage of 6 mg/kg of body weight twice daily every 3 days. This injection schedule was determined from a schedule-optimizing animal study using human pancreatic cancer PANC-1 xenografts in nude mice. Our results showed that treatment with DCB 3503 twice daily on every third day resulted in minimal toxicities in terms of loss of body weight and decrease in blood cell counts (red blood cells, white blood cells, and platelets).
Based upon the kinetics of onset of CIA in this model, the mice were treated either before the onset of inflammation (early treatment) or after the onset of inflammation (late treatment). In the early treatment cohort, mice were immunized with CII/CFA and on day 21 postimmunization, were randomly divided into 2 groups and administered DCB 3503 or vehicle just prior to their booster immunization (CII/IFA). Treatment continued until day 30 (twice daily every 3 days). In the late treatment cohort, mice were immunized with CII/CFA, and on day 29 postimmunization, those with arthritis (score of 2–7) were divided into 2 equal cohorts and administered DCB 3503 or vehicle for a period of 13 days (twice daily every 3 days).
To define the effects of DCB 3503 on the LPS-enhanced CIA model, mice with arthritis scores of ≤1 were injected intraperitoneally with LPS (50 μg/mouse). Mice were then treated either 24 hours before (early) or 48 hours after (late) LPS challenge. The severity of arthritis was monitored daily and scored as described above.
Cell preparation and culture.
Bone marrow–derived dendritic cells were generated using granulocyte–macrophage colony-stimulating factor for 7 days, as described previously (17, 18). Bone marrow–derived dendritic cells were activated with different concentrations of LPS (10 ng/ml to 1 μg/ml) or poly(I-C) (1–10 μg/ml) in the presence or absence of DCB 3503 for 6 or 24 hours, with the addition of brefeldin A for the last 3 hours, and cells were used for intracellular cytokine (IL-12 and TNFα) staining. Supernatants were also collected for analysis of cytokine levels.
Cytokine levels were determined by the following 3 methods. For cytokine determinations in cultured cells or macrophages ex vivo, intracellular cytokine staining was used as described in our previous studies (19). For cytokine determinations in culture supernatants or serum, cytometric bead array kits (BD Biosciences, San Diego, CA) were used as described previously (20). For cytokine determinations in joints, ankle joints were removed, frozen in liquid nitrogen, smashed, mixed with 200 μl of phosphate buffered saline (PBS) at 4°C, centrifuged at 10,000 revolutions per minute, and the supernatants were assayed by ELISA (for IL-1β; R&D Systems) and by cytometric bead array (for IL-12, TNFα, interferon-γ [IFNγ], IL-6, IL-10 and monocyte chemotactic protein 1 [MCP-1]).
T cell proliferation assay.
Single-cell suspensions derived from the spleen were prepared and cultured as described previously (19). CD4+ T cells were negatively selected from splenocytes of mice that had been immunized and treated with either DCB 3503 or vehicle, as described previously (14). The purity of CD4+ T cells was >95%, and similar percentages of activated CD4+ T cells (CD62Llow,CD44high) were detected in these 2 groups of mice (data not shown). T cell–depleted splenocytes from DBA/1J mice were treated with mitomycin C (Sigma-Aldrich, St. Louis, MO) for 45 minutes (0.5 μg/ml) and used as antigen-presenting cells. CD4+ T cells (1 × 106/ml) were incubated together with antigen-presenting cells (5 × 106/ml) in the presence of different concentration of T cell–grade bovine CII for 72 hours, and pulsed with 3H-thymidine (20 μCi/well) for 18 hours before harvesting. Uptake of radiolabeled thymidine (in counts per minute) was measured in a liquid scintillation counter as described elsewhere (14).
ELISA for serum anti-CII antibodies and their isotypes.
Serum anti-CII antibodies and their isotypes were measured by ELISA, as described previously (14). Briefly, flat-bottomed 96-well ELISA plates were coated overnight at 4°C with bovine CII (0.5 μg/well). After blocking the plates with 5% fetal bovine serum–PBS, mouse sera were diluted 1:800, added to duplicate wells, and incubated for 2 hours at room temperature. Plates were washed, and biotin-conjugated goat anti-mouse IgG, IgG1, and IgG2a were added at a dilution of 1:2,000 for 1 hour, followed by the addition of streptavidin–horseradish peroxidase (Sigma-Aldrich). Plates were then developed and antibody levels measured as described previously (14).
Mice were killed and joints were removed, immediately fixed in 10% buffered formalin, and decalcified in a decalcifying solution for 3–5 days. The tissue was then processed and embedded in paraffin. Five-micrometer tissue sections were prepared and stained with hematoxylin and eosin using standard methods. All sections were stained by the Yale Pathology Laboratory and read blindly by a pathologist (JAM).
Analyses of arthritis severity data were performed using SAS version 8.1 software (SAS Institute, Cary, NC). Statistical testing was 2-sided at a 5% level of significance. Exploratory data analysis was performed by assessing descriptive statistics (mean and SD) for each variable using SAS PROC Univariate and PROC Freq programs. The study variable was arthritis scores over time. Outcomes were assessed for normality prior to analysis, using normal probability plots and Kolmogorov-Smirnov test statistics. Arthritis scores were analyzed using the PROC Mixed program and mixed effects models, with treatment group, time, and treatment group by time interaction as the fixed effects and a structured variance–covariance pattern matrix. The best-fitting variance–covariance matrix according to the Bayesian information criterion was selected. Cytokine levels were compared using Student's unpaired 2-tailed t-test or nonparametric analysis (Wilcoxon's or Mann-Whitney test) if the SDs were significantly different between the 2 groups being compared, using InStat 2.03 software (for Macintosh; GraphPad, San Diego, CA).
Significant blocking of the development and progression of CIA by treatment with DCB 3503.
To define the therapeutic effects of DCB 3503 on CIA in DBA/1J mice, we used 2 different treatment protocols, early and late treatment (before and after the onset of CIA). In the early treatment cohort, DCB 3503 significantly decreased the severity and delayed the onset of arthritis (Figure 1A). On day 30, only 1 of the 8 mice in the active treatment group (12.5%) developed a mild arthritis, whereas 9 mice of the 10 mice in the vehicle treatment group (90%) developed arthritis (mean score >3.0) (P < 0.0001). It should be noted that upon the termination of therapy, the incidence and severity of arthritis increased in the treated group; however, 10 days after treatment termination, the mean ± SD arthritis scores were still significantly lower than those in the control group (2.75 ± 2.8 versus 4.6 ± 2.2). Thus, DCB 3503 significantly blocked the development of arthritis and delayed its onset.
In the late treatment cohort, mice treated with vehicle continued to develop severe arthritis, whereas mice treated with DCB 3503 showed no progression or a reduced severity of arthritis (P < 0.0001) (Figure 1B). Thus, our results suggest that DCB 3503 can block the development and progression of inflammatory arthritis.
Significant suppression of LPS-triggered joint inflammation by treatment with DCB 3503.
To amplify and equalize the degree of arthritis in mice with CIA, animals were injected with LPS, as previously described (16) and then treated with DCB 3503 or vehicle according to 2 difference protocols, early and late treatment (1 day before or 2 days after LPS injection). In the early treatment cohort, all 5 mice treated with vehicle developed severe arthritis, with marked swelling and erythema of the hind paws and fore paws (Figure 2A). Inflammation in these mice affected the ankle joints and extended distally through the limb and the digits. The maximum arthritis score was 12, with mean score of 10. In contrast, the group treated with DCB 3503 was still free of arthritis after LPS challenge; only 2 mice had mild arthritis, with scores of <2 (P < 0.001) (Figure 3A).
In the late-treatment group, upon LPS injection, mice with a mean arthritis score of 5 were divided into 2 groups and treated with either DCB 3503 or vehicle from day 30 to day 42 postimmunization. Mice in the vehicle control group developed severe arthritis, which peaked on day 35 with a mean arthritis score of 11. Mice in the active treatment group showed no signs of progression and maintained similar arthritis scores as for the prior treatment (P < 0.0001) (Figure 3B). Our results therefore demonstrate that LPS-triggered inflammatory responses can be blocked by administering DCB 3503.
Findings of histopathologic analysis.
Further evidence to support the inhibitory effects of DCB 3503 on LPS-accelerated arthritis was obtained by histopathology analysis. On day 35, control mice exhibited robust pannus formation and significant articular cartilage erosion, with pannus eroding into and replacing the articular cartilage overlying the bone (Figure 2A). In contrast, DCB 3503–treated mice exhibited well-preserved joint spaces and articular cartilage surfaces, with minimal pannus formation (Figure 2B). On day 50, control mice exhibited more extensive pannus formation and destruction of bone and cartilage (Figure 2C) as compared with control joints on day 35. Figure 2C shows an articular cartilage remnant surrounded by the invading pannus. In contrast, DCB 3503–treated mice showed a normal joint space and well-preserved articular cartilage (Figure 2D), indicating that the administration of DCB 3503 directly correlated with a reduction in disease severity. Similar effects on joint pathology were seen in DCB 3503–treated mice with CIA (data not shown).
Suppression of the production of proinflammatory cytokines ex vivo by treatment with DCB 3503.
To define the underlying mechanisms of the effect of DCB 3503, we examined the systemic and local levels of these cytokines ex vivo. Treatment with DCB 3503 before the onset of arthritis significantly reduced the level of TNFα in serum (Figure 4A). The level of IFNγ in the same groups of serum samples was also reduced, whereas the level of IL-4 was increased, although the difference was not statistically significant (data not shown).
To test the effect of DCB 3503 on LPS-triggered TNFα production by macrophages ex vivo, splenocytes were obtained from mice with LPS-triggered CIA on day 35 postimmunization and were directly stained with intracellular TNFα. Mice treated with DCB 3503 produced significantly less TNFα than those treated with vehicle (Figure 4B).
Local levels of IL-1β were measured in joint tissues from mice with LPS-triggered CIA on day 35 postimmunization. Tissues were homogenized and washed with 200 μl of PBS per joint, and cytokine levels in the supernatant were determining by ELISA and cytometric bead array. DCB 3503 essentially blocked the production of IL-1β (Figure 4C), as well as several other inflammatory cytokines (TNFα, IL-6, IL-12, and MCP-1) (Table 1), at sites of local inflammation. Taken together, these data strongly suggest that DCB 3503 suppresses the development of inflammatory arthritis and joint destruction through blocking the production of inflammatory cytokines such as TNFα, IL-1β, and others.
|TNFα, mean ± SD pg/ml||IL-6, mean ± SD pg/ml||IL-12, mean ± SD pg/ml||MCP-1, mean ± SD pg/ml|
|Vehicle||760 ± 47†||476 ± 36†||190 ± 26‡||261 ± 18†|
|DCB 3503||68 ± 28||68 ± 29||48 ± 35||51 ± 15|
|Vehicle||4,767 ± 509†||4,659 ± 386†||950 ± 40†||651 ± 33†|
|DCB 3503||868 ± 124||544 ± 119||230 ± 30||59 ± 9|
Suppression of the production of mediators of inflammation by bone marrow–derived dendritic cells by treatment with DCB 3503.
Based upon the effects of DCB 3503 on LPS-triggered joint inflammation and destruction, we next tested the effects of this compound on the production of proinflammatory cytokines and mediators of inflammation by bone marrow–derived dendritic cells. Bone marrow–derived dendritic cells were pretreated with either vehicle or different concentrations of DCB 3503 for 1 hour, and cells were then activated with LPS (1 μg/ml) or poly(I-C) (5 μg/ml) for 24 hours, with the addition of brefeldin A for the last 3 hours. These cells were then used for intracellular cytokine staining.
DCB 3503 significantly suppressed both the LPS-triggered and the poly(I-C)–triggered production of IL-12 and TNFα, especially IL-12+,TNFα+ double-positive cells (from 12.4% to 1.73% for LPS, and from 50% to 5% for poly[I-C]) in a concentration-dependent manner (Figure 4D). Similar results were obtained with bone marrow–derived macrophages or peritoneal macrophages (data not shown). To confirm the results from flow cytometry, supernatants from parallel cultures activated with LPS (1 μg/ml) for 24 hours were analyzed by cytometric bead array techniques (6 cytokine array kits, including IL-12 p75, TNFα, IFNγ, IL-6, IL-10, and MCP-1). DCB 3503 significantly reduced proinflammatory cytokine production by 4–11-fold (Table 1), with no effect on the secretion of IL-10 and IFNγ analyzed in the same array (data not shown).
Absence of direct effects of DCB 3503 on antigen-specific T and B cell functions.
Both T cells and B cells have been demonstrated to be essential for the initiation of inflammatory arthritis (1, 8). To test the effects of DCB 3503 on the CII-specific T cell proliferation response, DBA/1J mice were immunized with CII/CFA and treated either with DCB 3503 or vehicle on the day of booster immunization (day 21). On day 28 postimmunization, the CD4+ T cell population was enriched, and the proliferative response of this T cell subset to CII was measured by 3H-thymidine uptake assay. DCB 3503 treatment showed little effect on the CII-specific T cell proliferative response (Figure 5A).
To test the effects of DCB 3503 on the function of B cells, we examined by ELISA the levels of CII-specific antibodies and their isotypes in serum samples collected at different time points postimmunization from mice with CIA or with LPS-triggered CIA. Figure 5B shows the results obtained in serum samples collected from LPS-triggered arthritic mice. On day 14, the anti-CII antibody (IgG1 and IgG2a) was readily detectable in all immunized mice. The levels of anti-CII antibodies peaked on day 28 postimmunization. DCB 3503 treatment had no direct effect on the antibody response to CII at all time points analyzed (Figure 5B). Similar results were obtained with serum samples from mice with CIA (data not shown). These results indicate that DCB 3503 has no direct effect on either T cell or B cell immune responses.
RA is a common autoimmune disease that results in progressive joint inflammation and destruction (1). Although therapy with anti-TNFα and anti–IL-1 provides an alternative or supplemental approach to treatment with corticosteroids or methotrexate, the efficacy of these antagonists can be limited (12, 21). Therefore, a search for novel therapeutic agents is greatly needed not only to improve outcomes, but also to reduce the financial burden of current antirheumatic regimens. Chinese herbal medicines, such as the Tylophora plant, have been used for the treatment of rheumatism for a thousand years (6), and it is possible that such herbs could be used as an alternative approach to the treatment of RA, provided that the effective components are identified and their toxicities are understood. A newly synthesized tylophorine analog derived from Tylophora species, DCB 3503, has recently been shown to have potent effects against tumor growth in vitro as well as in vivo by is effects on cAMP, activator protein 1 (AP-1), or on NF-κB–mediated transcription (13). This study is the first to demonstrate that DCB 3503 effectively inhibits the development and progression of CIA in DBA/1J mice by suppressing the production of proinflammatory cytokines and mediators of inflammation by innate immune cells.
We demonstrated that early treatment with DCB 3503 significantly suppressed the development of arthritis, delayed its onset, and reduced its severity (Figure 1A). More impressively, treatment with DCB 3503 completely blocked the accelerated arthritis triggered by LPS (Figure 3A), with a reduction in inflammatory cell infiltration, prevention of pannus formation and cartilage erosion, and preservation of the joint architecture (Figure 2). Late treatment with DCB 3503 (after the onset of CIA) suppressed the progression of arthritis (Figures 1B and 3B) and reduced the degree of joint destruction (data not shown). Our results suggest that this tylophorine analog may potentially be used as a therapeutic agent for inflammatory arthritis in humans.
How does DCB 3503 block joint inflammation and destruction? The striking feature of DCB 3503 is that it suppressed the production of multiple proinflammatory cytokines and mediators of inflammation. More specifically, it reduced both systemic and local levels of TNFα and IL-1β (Figure 4) and suppressed the ability of innate immune cells to produce multiple inflammatory cytokines (TNFα, IL-12, IL-6) and chemokine MCP-1 (Figure 4 and Table 1), as well as other mediators of inflammation (iNOS and COX-2) (data not shown). The roles of these molecules in the pathogenesis of joint inflammation and destruction have been extensively studied (3, 8, 21, 22). All of these molecules have been shown to play a divergent role in the different processes of joint inflammation and destruction. Moreover, it has been demonstrated that TNFα plays a major role in the infiltration of inflammatory cells, whereas IL-1 has been shown to be involved in the destruction of the cartilage matrix (23). These 2 cytokines can also trigger the activation of NF-κB, which leads to the production of additional inflammatory cytokines that will activate a cascade of events that cause joint inflammation and damage (24, 25). Both iNOS and COX-2 are important mediators of IL-1–induced bone erosion and edema in the CIA model (26, 27).
Our results indicate a role of DCB 3503 in inhibiting both the inflammation and the destruction caused by inflammatory arthritis, possibly through multiple mechanisms. DCB 3503 has potent inhibitory effects on AP-1 and NF-κB (13), transcription factors that play a critical role in the pathogenesis of RA in humans and CIA in mice (24, 28, 29). It is therefore reasonable to speculate that DCB 3503 could suppress the development and progression of inflammatory arthritis by acting on these essential signaling pathways. Further studies are needed to define the underlying molecular mechanisms that mediate the effects of DCB 3503 on these signaling pathways.
In contrast to its effects on innate immune cells, DCB 3503 had no direct effect on the function of T cells or B cells, as evidenced by the fact that both the CII-specific T cell proliferative response and anti-CII antibody production were not altered (Figure 5). Although DCB 3503 significantly suppressed the production of proinflammatory cytokines, it is very likely that DCB 3503 does not affect the ability of dendritic cells to present antigen and prime the T cell response. Indeed, in our preliminary studies, DCB 3503 had little effect on LPS-triggered dendritic cell maturation (class II major histocompatibility complex and B71/B7-2 expression) in vitro (data not shown). The fact that DCB 3503 attenuated joint inflammation and destructive arthritis without affecting the adaptive immune response implies that DCB 3503 has little or no effect on the initiation of inflammatory joint disease, but rather, exerts its effects on downstream events that are important for the propagation of inflammatory joint disease.
It should be mentioned that in these studies, the administration of DCB 3503 did not lead to a reduction in the body weight of the mice and did not change the peripheral blood cell counts (data not shown), and no obvious toxicity was observed. Similarly, no liver toxicity was observed even after prolonged periods of administration of DCB 3503 in the MRL/lpr mouse model of lupus (Choi J-Y, Craft J: personal communication). Thus, DCB 3503 has the potential to be a safe agent in the treatment of humans with RA.
In conclusion, we have discovered a new compound that selectively targets the production of proinflammatory cytokines by innate immune cells. This resulted in striking attenuation of joint inflammation and joint destruction associated with arthritis in the CIA mouse model. This novel compound may have potential in the treatment of humans with inflammatory arthritis.
We thank Dr. Yunfei Gao and Dr. Insoo Kang for helpful suggestions. We thank Dr. Bohdan Harvev for critical review of the manuscript.
- 6The elevated ratio of interferon γ/interleukin-4-positive T cells found in synovial fluid and synovial membrane of rheumatoid arthritis patients can be changed by interleukin-4 but not by interleukin-10 or transforming growth factor β. Rheumatology (Oxford) 1999; 38: 1058–67., , , , , , et al.