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Abstract

  1. Top of page
  2. Abstract
  3. PATIENTS AND METHODS
  4. RESULTS
  5. DISCUSSION
  6. AUTHOR CONTRIBUTIONS
  7. ROLE OF THE STUDY SPONSORS
  8. Acknowledgements
  9. REFERENCES

Objective

CXCL10 (also known as interferon-γ–inducible 10-kd protein [IP-10]) is a chemokine that potentially plays a role in the immunopathogenesis of rheumatoid arthritis (RA). We undertook this phase II study to evaluate the efficacy and safety of MDX-1100, a fully human, anti-CXCL10 (anti–IP-10) monoclonal antibody, in RA patients whose disease responded inadequately to methotrexate (MTX).

Methods

Patients with active RA receiving stable doses of MTX (10–25 mg weekly) were randomized to receive intravenous doses of 10 mg/kg MDX-1100 (n = 35) or placebo (n = 35) every other week. The primary end point was the proportion of patients meeting the American College of Rheumatology 20% improvement criteria (achieving an ACR20 response) on day 85, and patients were followed up for safety to day 141.

Results

The ACR20 response rate was significantly higher among MDX-1100–treated patients than among placebo-treated patients (54% versus 17%; P = 0.0024). Statistically significant differences in the ACR20 response rate between treatments were observed starting on day 43 (P < 0.05). The ACR50 and ACR70 response rates on day 85 did not differ between the groups. Overall, 51.4% of MDX-1100–treated patients and 30.3% of placebo-treated patients experienced at least 1 adverse event (AE). No study drug–related serious AEs were reported.

Conclusion

MDX-1100 was well tolerated and demonstrated clinical efficacy in RA patients whose disease responded inadequately to MTX. This is the first study to demonstrate clinical efficacy of a chemokine inhibitor in RA and supports the notion of a potential role of IP-10 in the immunopathogenesis of RA.

Multiple studies in patients with rheumatoid arthritis (RA) have identified increased chemokine levels in blood, synovial fluid, and synovial membrane compared to patients with osteoarthritis or healthy controls (1–3). Interferon-γ (IFNγ)–inducible 10-kd protein (IP-10) (also known as CXCL10) is a chemokine that can potentially regulate inflammation at several levels. It induces integrin activation and generates directional migration of multiple cell types, including activated T cells, monocytes, and natural killer (NK) cells (4, 5). In addition to mediating chemotaxis, IP-10 induces apoptosis of pancreatic beta cells and inhibits the proliferation of both epithelial and endothelial cells (6–9). Other proinflammatory functions of IP-10 include induction of molecules, such as interleukin-8 (IL-8) and CXCL5, and up-regulation of costimulatory cell surface molecules, such as CD54, CD80, and CD86, on monocytes (Cardarelli PM: unpublished observations).

Multiple receptors have been implicated in these processes, including CXCR3, CXCR3b, glycosaminoglycans, Toll-like receptor 4, and an unidentified high-affinity receptor on epithelial cells (6–11). Expression of IP-10 and CXCR3 is increased in murine models of arthritis, and neutralizing anti–IP-10 or anti-CXCR3 antibodies ameliorate disease manifestations in these models (12, 13). IP-10 and CXCR3 expression is also increased in the synovial membrane of RA patients (14, 15). Taken together, these data suggest a potential role of IP-10 as a therapeutic target in RA.

MDX-1100 is a fully human anti–IP-10 (anti-CXCL10) monoclonal antibody (produced by Medarex, since acquired by Bristol-Myers Squibb) that binds to IP-10 with high affinity but not to other CXCR3 ligands, CXCL9, or CXCL11. In vitro studies demonstrated that MDX-1100 neutralizes the functional activity of IP-10, including calcium influx, leukocyte migration, and up-regulation of IP-10–responsive genes. Phase I single-dose studies of MDX-1100 in healthy volunteers or patients with ulcerative colitis demonstrated that the drug was well tolerated at all dose levels tested (0.1–10 mg/kg) and had a half-life of 10 days. The current study was a randomized, double-blind, placebo- controlled, phase II proof-of-concept study to determine the clinical efficacy and safety of MDX-1100 in RA patients whose disease responded inadequately to methotrexate (MTX).

PATIENTS AND METHODS

  1. Top of page
  2. Abstract
  3. PATIENTS AND METHODS
  4. RESULTS
  5. DISCUSSION
  6. AUTHOR CONTRIBUTIONS
  7. ROLE OF THE STUDY SPONSORS
  8. Acknowledgements
  9. REFERENCES

Patients.

We enrolled male and female adult patients (age ≥18 years) in Ukraine and Romania who were diagnosed as having RA based on the American College of Rheumatology (ACR) 1987 revised criteria (16) and whose disease responded inadequately to MTX. Inclusion criteria were at least 6 swollen joints and 6 tender joints and at least 2 of the following: a C-reactive protein (CRP) level of >10 mg/liter, an erythrocyte sedimentation rate (ESR) of ≥28 mm/hour, or morning stiffness lasting ≥45 minutes. All patients had to be seropositive for rheumatoid factor (RF) and/or anti–cyclic citrullinated peptide (anti-CCP) antibody. Patients were included if they had been taking MTX at a dosage of 10–25 mg weekly for at least 6 months and the dosage had been stable for at least 42 days prior to randomization. No change in MTX treatment was permitted during the study unless necessitated by toxicity. Concomitant treatment with low-dose corticosteroids (≤10 mg/day of prednisolone or equivalent) and nonsteroidal antiinflammatory drugs was permitted provided no change in dosage was anticipated during the study. Besides MTX, all other disease-modifying antirheumatic drugs (DMARDs) or biologic agents had to have been discontinued at least 28 days prior to randomization, with the exception of leflunomide, which had to have been discontinued at least 60 days prior to randomization. Patients were excluded if they had prior B cell–depleting therapy, were immunodeficient, had complications of RA such as pulmonary fibrosis or vasculitis, or had a history of malignancy or acute or chronic infections.

Study design and treatment administration.

Patients were randomly assigned in a 1:1 ratio to receive either 10 mg/kg of MDX-1100 or placebo every 2 weeks for a total of 12 weeks (6 infusions). MDX-1100 or placebo was administered intravenously for 60 minutes on days 1, 15, 29, 43, 57, and 71 (weeks 0, 2, 4, 6, 8, and 10). Premedication was not routinely administered.

ACR core set assessments (17) and data on the primary end point for clinical efficacy (meeting the ACR 20% improvement criteria [achieving an ACR20 response] [18]) were obtained at each visit up to day 85 (week 12), and the followup visit occurred on day 113 (week 16). Peak and trough blood samples for pharmacokinetic studies were obtained at each dosing visit, and single samples were obtained at nondosing visits up to day 113. Routine chemistry, hematology, and other safety laboratory tests were performed at each visit. Blood samples for immunogenicity testing were obtained at baseline and on days 29, 57, 85, and 113. All patients were followed up for safety until day 141. Reported efficacy and safety parameters are based on the visit days when the parameters were assessed. Written informed consent was obtained from each patient prior to screening, and the ethics review board of each participating center approved the study.

Safety assessments.

All patients were monitored for adverse events (AEs), and these were recorded at each visit and graded according to the Common Terminology Criteria for Adverse Events, version 3 (19). A serious AE was defined as an event that was fatal or life-threatening, required prolonged hospitalization, was significantly or permanently disabling or incapacitating, was associated with a congenital anomaly or birth defect, or required medical or surgical intervention to prevent one of the above-mentioned outcomes. An independent data monitoring committee reviewed unblinded safety data during the study.

Safety analyses were conducted on all subjects administered at least 1 dose or partial dose of MDX-1100 or placebo. Safety assessments, including the incidence and severity of treatment-emergent AEs, were summarized by descriptive statistics. A treatment-emergent AE was defined as any AE occurring within 70 days of the last dose of study drug or any study drug–related AE occurring at any time point in the study.

Efficacy end points.

The primary end point of the study was the proportion of patients having achieved an ACR20 response on day 85. Secondary end points included the proportion of patients achieving ACR50 and ACR70 responses (20) as well as improvements in the individual components of the ACR core set of measures (17). A 0–100-mm visual analog scale was used for the patient's assessment of pain and the patient's and physician's global assessments of disease activity. The proportion of patients having achieved disease remission or a European League Against Rheumatism (EULAR) good response (21) was determined using the Disease Activity Score in 28 joints (DAS28) (22), which assessed the number of swollen and tender joints, CRP levels, and the patient's global assessment of disease activity. Physical function was determined using the modified Health Assessment Questionnaire (23).

Pharmacokinetics assessment.

Serum MDX-1100 concentrations were determined by quantitative enzyme-linked immunosorbent assay.

Pharmacodynamics assessment.

Samples for pharmacodynamic assessments were obtained for exploratory analyses at baseline and on study days 8 and 85.

Measurement of gene expression by quantitative polymerase chain reaction (qPCR).

Total RNA was purified from peripheral blood collected in PAXgene Blood RNA tubes (Qiagen) according to the manufacturer's protocol. Complementary DNA (cDNA) was synthesized using the Invitrogen SuperScript III Reverse Transcriptase Kit and oligo(dT) primers (Invitrogen) according to the manufacturer's instructions. Gene expression was measured by qPCR using gene-specific primers and Applied Biosystems Power SYBR Green PCR Master Mix. Copy number per 5 ng cDNA was determined using a standard curve of gene-specific PCR product, and then normalized to the copy number of a housekeeping gene, UBE2D2.

Measurement of serum cytokine levels.

Cytokines were analyzed using the Luminex multiplexing system and Millipore Milliplex Human Cytokine/Chemokine Immunoassay kits. Kits were run according to the manufacturer's recommended protocol. Levels of 30 cytokines/chemokines, including IP-10 and eotaxin, were assayed, and concentrations were determined from a standard curve run with each assay. Sixteen of the 28 other cytokines/chemokines were below the detection range of the assay (granulocyte colony-stimulating factor, IFNγ, IL-1β, IL-2, IL-3, IL-4, IL-5, IL-7, IL-8, IL-10, IL-13, IL-15, IL-17, IL-12p70, tumor necrosis factor α [TNFα], and TNFβ), and 12 showed no significant differences among treatment groups (granulocyte–macrophage colony-stimulating factor, IFNα, IL-6, IL-12p40, macrophage inflammatory protein 1α [MIP-1α], MIP-1β, IL-1α, soluble shed CD25, monocyte chemotactic protein 1, vascular endothelial growth factor, macrophage-derived chemokine, and soluble CD40L).

Immunogenicity assessment.

Serum samples along with positive and negative controls were tested in an electrochemiluminescence assay using the Meso Scale Discovery Sector PR 100 Plate Reader. Briefly, biotinylated MDX-1100, ruthenylated MDX-1100, and patient serum samples were added to wells of a polypropylene microtiter plate. Upon incubation, anti–MDX-1100 antibodies bind to both the biotinylated MDX-1100 and ruthenylated MDX-1100 molecules to form an antibody complex bridge. Next, the samples were added to microwell plates coated with streptavidin (Meso Scale Discovery). After washing, a tripropylamine-containing read buffer was added, and upon voltage application, a chemiluminescent signal was triggered. Samples were screened at a dilution of 1:20. Samples with signal intensity that was less than or equal to the calculated assay cut point (as determined by a low probability for presence of any anti–MDX-100 antibodies) were reported as negative.

Statistical analysis.

A sample size of 35 patients per cohort was needed to provide 80% power to detect a difference of 40% in the ACR20 response between the MDX-1100–treated and placebo-treated cohorts at the 5% significance level (2-tailed) adjusted for a discontinuation rate of 10%. The ACR20 response rate in the placebo-treated cohort was assumed to be 30%. The intent-to-treat (ITT) population, defined as all patients who were randomized to receive MDX-1100 or placebo, was the primary population for determining efficacy. The difference in response rates was analyzed by Fisher's exact test. Subjects who discontinued prior to day 85 were imputed as nonresponders. Secondary end points and baseline demographics were summarized using descriptive statistics.

For pharmacokinetics assessments, peak and trough concentrations were summarized using descriptive statistics. For pharmacodynamics assessments, a paired t-test was used to compare FoxP3 gene expression in patient samples collected on day 1 and day 85. The Mann-Whitney U test was used to compare mean expression levels of cytokines or genes between 2 patient groups. When more than 2 groups were compared, the Kruskal-Wallis test with Dunn's correction was used. Data were analyzed using GraphPad Prism software. The reported P values for the pharmacodynamic analyses were not adjusted for multiplicity, although multiple pharmacodynamic biomarkers were analyzed.

RESULTS

  1. Top of page
  2. Abstract
  3. PATIENTS AND METHODS
  4. RESULTS
  5. DISCUSSION
  6. AUTHOR CONTRIBUTIONS
  7. ROLE OF THE STUDY SPONSORS
  8. Acknowledgements
  9. REFERENCES

Baseline characteristics.

Baseline demographic characteristics and disease activity were generally comparable between the treatment cohorts (Table 1). The majority of patients in each cohort were female, all were Caucasian, and the mean age of patients was 47.5 years and 50.7 years in the MDX-1100–treated and placebo-treated cohorts, respectively. The mean weekly MTX dose was the same in each cohort (11.8 mg), and only 1 patient had previously been treated with an anti-TNF agent. The majority of patients in each cohort were RF positive (88.6% and 94.3% in the MDX-1100–treated and placebo-treated cohorts, respectively) and/or anti-CCP positive (94.3% in both cohorts).

Table 1. Baseline characteristics in the intent-to-treat population*
 MDX-1100 (n = 35)Placebo (n = 35)
  • *

    Except where indicated otherwise, values are the mean ± SD. RA = rheumatoid arthritis; DMARDs = disease-modifying antirheumatic drugs; anti-TNF = anti–tumor necrosis factor; RF = rheumatoid factor; anti-CCP = anti–cyclic citrullinated peptide; CRP = C-reactive protein; ESR = erythrocyte sedimentation rate; VAS = visual analog scale; M-HAQ = modified Health Assessment Questionnaire.

  • Does not include methotrexate (MTX).

  • Sixty-eight joints were evaluated for tenderness and 66 joints were evaluated for swelling.

  • §

    High scores indicate more severe abnormalities.

  • Low scores indicate low disease activity and high scores indicate greater disease activity.

Age, mean (range) years47.5 (24–73)50.7 (21–74)
Weight, mean (range) kg67.5 (47–116)66.1 (40–86)
Female, %82.977.1
Caucasian, no. (%)35 (100)35 (100)
Disease duration, mean (range) years6.6 (1–23)7.6 (0–29)
Baseline medications  
 Corticosteroids, %51.454.3
 Prednisolone equivalent dose, mean (range) mg/day8.2 (3–13)7.4 (4–10)
 MTX, %100100
 MTX dose, mean (range) mg/week11.8 (10–20)11.8 (10–20)
Prior RA therapy, %  
 DMARDs34.331.4
 Anti-TNF02.9
RF positive, %88.694.3
Anti-CCP positive, %94.394.3
CRP, mean (range) mg/liter18.3 (0–109)11.6 (1–41)
ESR, mean (range) mm/hour44.0 (22–95)47.3 (18–86)
No. of tender joints22.5 ± 11.3020.5 ± 9.79
No. of swollen joints15.4 ± 6.6113.5 ± 6.16
Patient's assessment of pain, 0–100-mm VAS§63.9 ± 12.4362.2 ± 17.9
Patient's global assessment of disease activity, 0–100-mm VAS§66.6 ± 13.2167.1 ± 16.36
Physician's global assessment of disease activity, 0–100-mm VAS§61.1 ± 11.7660.3 ± 14.32
M-HAQ score, 0–31.5 ± 0.471.5 ± 0.66

Patient disposition.

Seventy patients were randomized to receive 10 mg/kg MDX-1100 plus MTX (n = 35) or placebo plus MTX (n = 35), and 68 received ≥1 doses of study drug. Two patients withdrew from the study after randomization and prior to administration of the first dose of study drug. Most patients received all 6 infusions of study drug; 32 patients (91.4%) in the MDX-1100–treated cohort and 28 patients (84.8%) in the placebo-treated cohort completed all infusions.

The majority of patients in each cohort completed the day 85 efficacy assessment (Table 2). Two patients in the MDX-1100–treated cohort withdrew early from the study; 1 patient withdrew consent for unknown reasons and 1 withdrew due to an AE. Six patients withdrew from the placebo-treated cohort prior to day 85; 5 withdrew consent for unknown reasons and 1 withdrew due to lack of efficacy.

Table 2. Patient disposition from enrollment to completion of the study*
 MDX-1100 (n = 35)Placebo (n = 35)
  • *

    Values are the number (%) of patients.

Completed through day 85  
 No2 (5.7)6 (17.1)
 Yes33 (94.3)29 (82.9)
Primary reason for discontinuation prior to day 85  
 Adverse event1 (2.9)0
 Subject withdrew consent1 (2.9)5 (14.3)
 Unsatisfactory therapeutic effect01 (2.9)
Entered extended followup to day 141  
 No2 (5.7)5 (14.3)
 Yes33 (94.3)30 (85.7)
Completed extended followup  
 No00
 Yes33 (94.3)30 (85.7)

Efficacy.

In the ITT population, 19 of 35 patients (54%) in the MDX-1100–treated cohort and 6 of 35 patients (17%) in the placebo-treated cohort had achieved an ACR20 response on day 85. The ACR20 response differed significantly (P = 0.0024) between the 2 cohorts on day 85. Analysis of response by visit demonstrated a difference in ACR20 response rates from day 29 that achieved statistical significance by day 43 (P < 0.05) and all subsequent visits (Figure 1).

thumbnail image

Figure 1. Efficacy responses to treatment with MDX-1100 or placebo in the intent-to-treat population. Top, Proportion of patients meeting the American College of Rheumatology 20% improvement criteria (achieving an ACR20 response) by week. Differences in ACR20 response rates in patients treated with MDX-1100 versus those treated with placebo were observed starting at week 4 and achieved statistical significance by week 6 and at all subsequent visits. Bottom, Mean ± SD change in the Disease Activity Score in 28 joints (DAS28) by week. Mean change from baseline in the DAS28 was similar between cohorts up to week 2, and there were nonsignificant differences from week 4 to week 12.

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While the response rates were greater in the MDX-1100–treated cohort than in the placebo-treated cohort, none of the secondary efficacy end points differed significantly. The ACR50, ACR70, and EULAR good responses in the MDX-1100–treated and placebo-treated cohorts on day 85 were 8.6% versus 2.9% (P = 0.61), 2.9% versus 0 (P = 1.00), and 5.7% versus 0 (P = 0.49), respectively. Mean change from baseline in the DAS28 was similar in the 2 cohorts up to day 15, and there were nonsignificant differences from day 29 to day 85 (Figure 1). For each of the ACR core set components with the exception of the ESR, a greater change in improvement from baseline was observed on day 85 in the MDX-1100–treated cohort (Table 3). There was no change in RF levels.

Table 3. Mean ± SD change from baseline in the ACR core components on day 85*
ACR core componentMDX-1100 (n = 35)Placebo (n = 35)
  • *

    ACR = American College of Rheumatology (see Table 1 for other definitions).

Tender joint count (66 joints assessed)−9.9 ± 15.07−5.1 ± 11.09
Swollen joint count (68 joints assessed)−10.3 ± 7.06−7.0 ± 6.81
Patient's assessment of pain, 0–100-mm VAS−16.6 ± 14.23−11.1 ± 19.69
Patient's global assessment of disease activity, 0–100-mm VAS−18.6 ± 15.94−14.1 ± 19.69
Physician's global assessment of disease activity, 0–100-mm VAS−18.1 ± 13.11−11.5 ± 12.83
HAQ disability index−0.5 ± 0.41−0.2 ± 0.47
ESR, mm/hour−5.0 ± 20.65−10.4 ± 14.17
CRP, mg/dl−3.2 ± 13.114.2 ± 22.49

AEs.

MDX-1100 had an acceptable toxicity profile. Overall, 51.4% of patients treated with MDX-1100 and 30.3% of patients treated with placebo experienced ≥1 AEs during the study (Table 4). The difference in AE frequency was primarily driven by a higher proportion of patients reporting peri-infusional events (defined as potentially infusion-related AEs occurring on the day of or the day following infusion) in the MDX-1100–treated cohort (25.7%) compared to the placebo-treated cohort (3.0%). With the exception of 1 patient with a grade 3 infusion reaction of bronchospasm and allergic rhinitis which led to early discontinuation from the study, all of the other AEs in the MDX-1100–treated group were mild to moderate in intensity and none led to discontinuation. The most common peri-infusional AEs in the MDX-1100–treated group were pyrexia and hyperthermia, which were reported by 5 of the 9 subjects. The majority of the peri-infusional AEs occurred after the first infusion only and did not recur when the patient continued to receive the study drug for the remainder of the 3-month study. The majority of AEs seen in ≥2 patients and occurring more frequently in the MDX-1100–treated cohort than in the placebo-treated cohort were infusion-related AEs (Table 4). The frequency of infections was 8.6% and 12.1% in the MDX-1100–treated and placebo-treated cohorts, respectively.

Table 4. Summary of AEs (safety population)*
 MDX-1100 plus MTX (n = 35)Placebo plus MTX (n = 33)
  • *

    Values are the number (%) of patients. AEs = adverse events; MTX = methotrexate.

Any AE18 (51.4)10 (30.3)
AE grade ≥31 (2.9)1 (3.0)
Serious AE01 (3.0)
Deaths01 (3.0)
Discontinued due to AE1 (2.9)0
Infections3 (8.6)4 (12.1)
Peri-infusional AEs (with 24 hours of infusion)9 (25.7)1 (3.0)
AEs reported in ≥2 subjects  
 Anemia2 (5.7)0
 Asthenia2 (5.7)0
 Hyperthermia2 (5.7)0
 Infusion-related reaction2 (5.7)0
 Pyrexia3 (8.6)1 (3.0)
 Alanine aminotransferase increased02 (6.1)
 Aspartate aminotransferase increased02 (6.1)
 Lymphocyte count decreased2 (5.7)0
 Neutrophil count increased2 (5.7)1 (3.0)

With few exceptions, AEs occurring during the study were grade 1 or grade 2 in severity. One patient administered placebo developed grade 4 pancytopenia and died suddenly. One patient administered MDX-1100 developed a grade 3 infusion reaction (bronchospasms) during the first infusion and withdrew from the study. In both cohorts there were no clinically significant changes from baseline during the study in relevant safety laboratory test results such as white blood cell, neutrophil, lymphocyte, monocyte, or platelet counts, or aspartate aminotransferase, alanine aminotransferase, creatinine, cholesterol, or triglyceride levels.

Pharmacodynamics findings.

Serum IP-10 protein levels were measured at baseline but could not be determined posttreatment because the anti–MDX-1100 antibody interferes with the assay (data not shown). Mean ± SD baseline IP-10 protein levels were comparable between the MDX-1100–treated group (544 ± 56 pg/ml) and the placebo-treated group (699 ± 95 pg/ml). In the MDX-1100–treated cohort, mean ± SD IP-10 protein levels at baseline were 593 ± 78 pg/ml among ACR20 responders and 480 ± 78 pg/ml among nonresponders (Figure 2A). IP-10 gene expression at baseline was significantly higher in ACR20 responders than in nonresponders (P = 0.026). The change from baseline in IP-10 messenger RNA (mRNA) levels was not statistically significant on day 85 in patients administered MDX-1100 (Figure 2B). Of 30 soluble serum factors analyzed by Luminex assay, only eotaxin demonstrated a potentially interesting trend. In the MDX-1100–treated cohort, baseline serum eotaxin protein levels were markedly lower in ACR20 responders than in nonresponders, and levels did not change significantly over the course of treatment (Figure 2C). All the other soluble factors analyzed were either below the limit of detection or demonstrated no significant change during the study. Messenger RNA analysis of FoxP3, a marker for Treg cells, did not demonstrate any change in level of expression in the placebo-treated cohort (not shown) or in nonresponders in the MDX-1100–treated cohort. In contrast, a statistically significant increase in FoxP3 mRNA was observed in ACR20 responders on day 85 compared to baseline (Figure 2D).

thumbnail image

Figure 2. A, Mean serum interferon-γ–inducible 10-kd protein (IP-10) concentrations at baseline did not differ between MDX-1100–treated patients who would meet the American College of Rheumatology 20% improvement criteria (achieve an ACR20 response) (responders [R]) and those who would not (nonresponders [NR]). B, Shown are IP-10 mRNA copy numbers in MDX-1100–treated patients at baseline and at week 12. ACR20 responders had significantly higher baseline IP-10 levels than nonresponders. C, Shown are serum eotaxin levels in MDX-1100–treated patients at baseline and at week 12. ACR20 responders had significantly lower eotaxin levels than nonresponders. D, Shown are FoxP3 mRNA copy numbers in MDX-1100–treated patients at baseline and at week 12. ACR20 responders had significantly increased FoxP3 levels after treatment, whereas FoxP3 levels remained unchanged in nonresponders. Horizontal bars represent mean values. NS = not significant.

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Pharmacokinetics findings.

Limited pharmacokinetics assessments were performed. The serum concentration of MDX-1100 attained steady state after ∼4 doses. The mean concentration at 30 minutes after the last dose on day 71 was 267.4 μg/ml, and the mean trough concentration on day 85 (2 weeks after the last dose) was 93.4 μg/ml.

Exposure-response analysis was conducted to examine the relationship between ACR20 responses and serum MDX-1100 concentrations. Although there were differences in ACR20 responses between MDX-1100–treated and placebo-treated subjects, no differences were observed when ACR20 responders were stratified into 3 groups based on day 85 trough concentrations (bottom tertile, ∼13.3–81.0 μg/ml; middle tertile, ∼86–111 μg/ml; top tertile, ∼112–202 μg/ml).

Immunogenicity findings.

Based on both screening and titration tests, the results in 5 patients (14%) were inconclusive, and the rest of the patients in the MDX-1100–treated cohort were considered negative for anti-human antibody responses on the last postdose day (day 113).

DISCUSSION

  1. Top of page
  2. Abstract
  3. PATIENTS AND METHODS
  4. RESULTS
  5. DISCUSSION
  6. AUTHOR CONTRIBUTIONS
  7. ROLE OF THE STUDY SPONSORS
  8. Acknowledgements
  9. REFERENCES

MDX-1100 administered every 2 weeks for 12 weeks in combination with weekly MTX led to a statistically significant increase in the ACR20 response rate, the primary end point of the study, compared with placebo. Combination therapy with MDX-1100 and MTX had an acceptable toxicity profile. To our knowledge, this is the first randomized, double-blind study of a chemokine inhibitor to demonstrate clinical efficacy in RA. While the ACR20 response at 3 months conclusively favored MDX-1100 compared to placebo (54% versus 17%; P = 0.0024), the ACR50 and ACR70 responses did not differ significantly between treatment groups.

Prior studies with inhibitors of chemokine pathways that are up-regulated in RA have failed to show evidence of pharmacodynamic or clinical efficacy. Clinical trials of antibodies targeting CCR2 (a chemokine receptor expressed on monocytes and T cell subsets), CCL2 (a chemokine that regulates monocyte/macrophage and T cell trafficking), and IL-8 (a chemoattractant predominantly for neutrophils) failed to demonstrate clinical efficacy (24). Blocking of CCR1 (a receptor for several ligands with potential roles in RA immunopathogenesis) or CCR5 has also failed to demonstrate clinical, magnetic resonance imaging, or histologic improvements in RA patients (25). The redundancy in circulating chemokines possibly contributes to the lack of effect observed by others upon inhibition of these mediator pathways. Positive results from the present study indicate that IP-10 is a dominant CXCR3 ligand in the immunopathogenesis of RA and possibly plays a more critical role in the inflammatory cascade. Another potential reason for lack of efficacy observed in the other studies cited above is their relatively short treatment duration, as discussed below.

The kinetics of the MDX-1100 response are of interest. A difference in ACR20 response between the MDX-1100–treated cohort and the placebo-treated cohort was observed starting at 4 weeks. By 6 weeks, the differences were statistically significant and were maintained for the duration of the study. Biologic agents that specifically target proinflammatory molecules or their receptors, such as TNFα or IL-6 receptor, typically have a quicker onset of action, usually within just a week or two of treatment initiation (26, 27). In contrast, the MDX-1100 response kinetics were similar to those of traditional DMARDs that have a slower onset of action of weeks to months. This may be due to the mechanism of action of MDX-1100 or to a suboptimal dosing schedule or regimen.

It is possible that greater activity and a faster onset of action could be obtained with a higher MDX-1100 dose or different dosing schedule. However, exposure-response analyses did not demonstrate an increase in clinical efficacy with higher exposure. The 10 mg/kg dose was based on a pharmacodynamic effect observed in a phase I study in which peripheral blood IP-10 mRNA levels were decreased following MDX-1100 administration. The 12-week period for this study was chosen based on previous studies of blockers of proinflammatory molecules that demonstrate clinically and statistically significant activity in this time frame, particularly for the ACR20 response (26, 27). Nevertheless, based on the kinetics of response discussed here, a longer dosing period with MDX-1100 may be required for optimal responses. This may also be relevant for the ACR50 and ACR70 responses, where 12–24 weeks of treatment may be needed to elicit higher responses (26, 27).

CXCR3, which is expressed on activated T cells, NK cells, plasmacytoid and myeloid dendritic cells, B cells, and mast cells and is the receptor for IP-10, can also bind 2 other ligands, CXCL9 and CXCL11 (28–30). CXCL9 and CXCL11 are both up-regulated during inflammation and regulate migration of T cells. It could be hypothesized that greater clinical efficacy in RA might be observed with an agent that neutralizes CXCL9 and/or CXCL11, in addition to IP-10. All 3 CXCR3 ligands are present in RA synovial fluid and produced by cytokine-stimulated synovial membrane fibroblasts in vitro (28, 31). Serum CXCL9 and IP-10 levels correlate with RA disease activity and response to therapy (32). MDX-1100, a highly specific IP-10–neutralizing antibody that does not inhibit CXCL9 or CXCL11 function in vitro, was chosen for clinical development because IP-10 blockade is sufficient to ameliorate inflammation in multiple murine disease models (33, 34), including an arthritis model (13), and because of theoretical concerns over potential safety issues with simultaneous blocking of all 3 CXCR3 ligands. It is currently unknown if other agents that effectively block multiple CXCR3 ligands or CXCR3 would demonstrate enhanced clinical efficacy with an acceptable safety profile.

There are multiple potential mechanisms by which neutralization of IP-10 may have activity in RA. IP-10 is known to regulate trafficking of cell types found in RA synovial membrane, such as effector T cells, B cells, dendritic cells, and mast cells (4, 5, 29, 35), and is thought to have key roles in initiating and perpetuating inflammation. CXCR3 is preferentially expressed on Th1 cells (36, 37), and IP-10 or CXCR3 blockade can skew T cell responses to secretion of Th2 cytokines, which dampen inflammation in animal models of arthritis (38). IP-10 may also play a role in modulating the balance of Treg cells at sites of inflammation. In this regard, in an atherosclerosis model in mice genetically deficient in apolipoprotein E and IP-10, an increase in lesional Treg cells was observed in association with decreased T effector cells and decreased plaque size (39). IP-10 up-regulates RANKL and TNFα production by CD4+ T cells and induces osteoclastogenesis. Moreover, in the collagen-induced arthritis model, anti–IP-10 antibody therapy inhibited synovial T cell and macrophage migration and diminished bone erosions (40). In vitro, IP-10 can induce secretion of proinflammatory cytokines (such as IL-8, CXCL5, and IL-6) and up-regulate cell surface molecules (such as CD80, CD86, and CD54) thought to have roles in the immunopathogenesis of RA (Cardarelli PM, et al: unpublished observations).

It is not known if MDX-1100 modulated the composition of synovial inflammatory cells since synovial biopsies were not performed. No effects of MDX-1100 on leukocyte subset counts or on the levels of Th1 and Th2 cytokines in serum were observed (data not shown). In vitro data demonstrate that IP-10 induces IL-8 production (Cardarelli PM, et al: unpublished observations), and in this clinical study IL-8 mRNA levels decreased following MDX-1100 administration (data not shown). It is unlikely that modulating IL-8 secretion would account for the observed clinical efficacy, since no differences in IL-8 levels between responders and nonresponders were observed after MDX-1100 treatment. Also, an anti–IL-8 monoclonal antibody had no activity in a phase II study in a similar patient population (24). Eotaxin, a chemokine mostly associated with regulating eosinophil chemotaxis, is measurable in the serum of RA patients (41), and low serum levels were associated with rapid radiographic progression in patients with early RA (42). The finding that serum eotaxin levels were lower in responders than in nonresponders in the present study is of unclear significance.

The most intriguing pharmacodynamic observation in this study is the finding of increased peripheral blood expression of mRNA for FoxP3, a transcription factor marker for Treg cells, in responders who received MDX-1100. CD4+CD25highFoxP3+ Treg cells have been identified in RA peripheral blood, synovial fluid, and synovial membrane (43). However, studies suggest that FoxP3-expressing cells may be relatively diminished in synovial membrane compared to T effector cells, and it has been hypothesized that increasing Treg cells in RA might have therapeutic potential (43–45). It is tempting to speculate that MDX-1100–mediated IP-10 neutralization modulates Treg cell–T effector cell balances, which would be consistent with the animal model findings discussed previously and would represent a novel mechanism of action in the treatment of RA.

Finally, the MDX-1100 infusion was generally well tolerated without requirement for infusion reaction prophylaxis in the majority of patients, and most AEs were low grade. The need for administration of prophylaxis for infusion reaction is a topic for future investigations.

The incidence of infection was comparable in the placebo-treated and MDX-1100–treated cohorts. However, this was only a 12-week study in a relatively small number of patients, and therefore definitive conclusions about the safety of MDX-1100 cannot yet be made. The immunogenicity result was inconclusive in 5 patients (14%) who were administered MDX-1100. However, this finding is likely to reflect suboptimization of the assay used to determine immunogenicity in a patient population with high-titer RF. MDX-1100 immunogenicity has not been observed in phase I single-dose studies or in a phase II multidose study in patients with ulcerative colitis.

In summary, MDX-1100–mediated neutralization of IP-10 in this 12-week proof-of-concept study was safe and associated with a modest but statistically significant clinical response in RA patients with active disease who were also taking MTX. This is the first study to demonstrate clinical efficacy of a chemokine inhibitor in RA and supports the notion of a potential role of IP-10 in the immunopathogenesis of RA.

AUTHOR CONTRIBUTIONS

  1. Top of page
  2. Abstract
  3. PATIENTS AND METHODS
  4. RESULTS
  5. DISCUSSION
  6. AUTHOR CONTRIBUTIONS
  7. ROLE OF THE STUDY SPONSORS
  8. Acknowledgements
  9. REFERENCES

All authors were involved in drafting the article or revising it critically for important intellectual content, and all authors approved the final version to be published. Dr. Luo had full access to all of the data in the study and takes responsibility for the integrity of the data and the accuracy of the data analysis.

Study conception and design. Yellin, Ter-Vartanian, Xu, Nichol.

Acquisition of data. Paliienko, Balanescu, Ter-Vartanian, Tseluyko, Xu, LeBlanc, Nichol, Ancuta, Chirieac.

Analysis and interpretation of data. Yellin, Ter-Vartanian, Xu, Tao, Cardarelli, LeBlanc, Nichol, Ancuta, Chirieac, Luo.

Preclinical studies to identify lead antibody. Cardarelli.

ROLE OF THE STUDY SPONSORS

  1. Top of page
  2. Abstract
  3. PATIENTS AND METHODS
  4. RESULTS
  5. DISCUSSION
  6. AUTHOR CONTRIBUTIONS
  7. ROLE OF THE STUDY SPONSORS
  8. Acknowledgements
  9. REFERENCES

Medarex (which has been acquired by Bristol-Myers Squibb) contributed to the study design, conduct of the study, data collection, and data analysis. All authors had access to the data. The manuscript was cowritten by the lead and senior authors, with additional editorial support provided by Bristol-Myers Squibb. All authors reviewed the initial manuscript and all revisions thereafter and provided approval to submit the manuscript for publication.

Acknowledgements

  1. Top of page
  2. Abstract
  3. PATIENTS AND METHODS
  4. RESULTS
  5. DISCUSSION
  6. AUTHOR CONTRIBUTIONS
  7. ROLE OF THE STUDY SPONSORS
  8. Acknowledgements
  9. REFERENCES

The authors thank Dr. Cristina Tanaseanu for substantial contributions in data acquisition. We also give special thanks to Judy Maccarone and Einav Leiberknight for assistance in conducting the study and writing the manuscript, to Jason Tian and Jenny Zhu for statistical support, to Anu Santhanagopal, PhD for writing and editorial support, and to Sharline Chen and Fei Cao for technical assistance.

REFERENCES

  1. Top of page
  2. Abstract
  3. PATIENTS AND METHODS
  4. RESULTS
  5. DISCUSSION
  6. AUTHOR CONTRIBUTIONS
  7. ROLE OF THE STUDY SPONSORS
  8. Acknowledgements
  9. REFERENCES