anti-OX40L Monoclonal Antibody Research Articles

520 - Evaluating the expression of OX40 and OX40 ligand in lesional skin versus non-lesional skin in volunteers with atopic dermatitis and in healthy control skin

Abstract Introduction/Background Growing evidence suggests that blocking the OX40 ligand (OX40L)/OX40 costimulatory pathway could have therapeutic effects in inflammatory diseases such as atopic dermatitis (AD). Amlitelimab (SAR445229; KY1005) is a potential first-in-class, fully human, non-depleting anti-OX40L monoclonal antibody that blocks OX40L-OX40 interactions. We report OX40L and OX40 expression from skin biopsies collected in two studies: an amlitelimab Phase 2a (Ph2a) study in patients with moderate-to-severe AD, and a small-scale study of patients with mild AD and healthy volunteers. Objectives Assess OX40L and OX40 expression in AD skin (lesional and non-lesional) and healthy control skin. Methods As part of a Ph2a randomized, double-blind, placebo-controlled study (NCT03754309) of amlitelimab in moderate-to-severe AD, 174 baseline-paired lesional/non-lesional skin biopsies were collected from all patients (n=87). In a separate small-scale study, 24 skin biopsies were collected, including healthy control skin from healthy volunteers (n=6) and paired lesional/non-lesional skin from patients with mild AD (n=9). Each biopsy underwent immunohistochemical (IHC; formalin-fixed paraffin-embedded) analyses using various antibodies (OX40L: rabbit IgG antibody from Cell Signaling Technology for Ph2a biopsies, proprietary mouse IgG2a KY1005 for small-scale study biopsies; OX40: clone Ber-ACT35 from BioLegend for all biopsies). A trained pathologist assessed IHC staining in the dermis and/or epidermis using semi-quantitative scoring (0 [no stained cells] to 10 [all cells stained]) or quantitative methods (cells/mm2, Tissue Studio 4.0, and HALO link image analysis software). Wilcoxon matched-pairs signed-rank test was used to assess IHC staining of Ph2a biopsies. Ph2a biopsies also underwent transcriptomic analyses (stored in RNALater) for OX40L (TNFSF4) and OX40 (TNFRSF4) on sequenced RNA using a linear mixed model with Benjamini-Hochberg adjustment. Results In baseline Ph2a biopsies, IHC analysis revealed elevated OX40L and OX40 staining in lesional versus non-lesional AD skin (P <0.0001 for both markers). For OX40L, the median score was 3 (lesional) versus 1 (non-lesional) and the Wilcoxon matched-pairs signed-rank test found P<0.0001. For OX40+ cells, the median expression was 179.8 cells/mm2 in lesional AD skin versus 54.45 cells/mm2 in non-­lesional AD skin (P<0.0001 using the Wilcoxon matched-pairs signed-rank test). These findings were confirmed by RNA sequencing analysis, with TNFSF4 (OX40L) and TNFRSF4 (OX40) differentially expressed in lesional and non-lesional AD skin (P<0.01 and P<0.0001, respectively). A larger scale study is ongoing that may further support this finding. In the small-scale study including patients with mild AD and healthy volunteers, there was a trend for an increase of mean OX40L+ and OX40+ cells/mm2 in lesional skin versus non-lesional skin and healthy control skin in the dermis and epidermis (OX40L+ dermal lesional, non-lesional, healthy, respectively: 17.6, 11.2, 11.5; OX40L+ epidermal: 5.8, 2.1, 1.9; OX40+ dermal: 5.7, 2.1, 0.4; OX40+ epidermal: 10.9, 9.0, 1.8); the increase was significant for lesional versus healthy control skin for dermal OX40+ cells/mm2 (P<0.001). Conclusions We confirm with IHC and RNA sequencing that OX40L and OX40 expression is elevated in skin lesions of patients with moderate-to-severe AD. We also demonstrate a trend to elevated OX40L and elevated OX40 in mild AD skin versus healthy volunteers. A potential explanation for the lower levels of OX40L and OX40 in patients with mild AD in this study may be their lower disease severity versus Ph2a study patients. These findings support the OX40L/OX40 pathway as a relevant target in T cell immune dysregulation; its blockade may represent a novel treatment approach for moderate-to-severe AD.

523 - Efficacy and safety of amlitelimab (an anti-OX40 ligand antibody) in patients with moderate-to-severe atopic dermatitis: 24-week results from a phase 2b trial (STREAM-AD)

Abstract Introduction/Background Targeting and binding OX40 ligand (OX40L) expressed on antigen-presenting cells may inhibit the persistent immune response that drives atopic dermatitis (AD) pathophysiology. Amlitelimab (SAR445229; KY1005) is a potential first-in-class, fully human, non-depleting anti-OX40L monoclonal antibody that blocks OX40L-OX40 interactions and has shown efficacy and an acceptable safety profile in a Phase 2a trial in adults with moderate-to-severe AD. Here, we present 24-week efficacy and safety results (Part 1) from an ongoing dose-ranging Phase 2b trial. The study remains blinded to individual patient data (Part 2 ongoing). Objectives To evaluate the efficacy and safety of amlitelimab in adults with moderate-to-severe AD. Methods STREAM-AD (NCT05131477) is a 52-week, randomised, double-blinded, placebo-controlled Phase 2b monotherapy trial. This study is designed with 2 parts (double-blind throughout): a 24-week treatment period (Part 1, completed and presented here) and a 36-week maintenance/withdrawal period (Part 2, ongoing). Adults (18 to <75 years; n=390) with moderate-to-severe AD were randomised 1:1:1:1:1 to receive subcutaneous amlitelimab Q4W (250 mg with 500 mg loading dose [LD], n=77; 250 mg without LD, n=78; 125 mg without LD, n=77; or 62.5 mg without LD, n=79) or placebo Q4W (n=79). The primary endpoint was percentage change in Eczema Area and Severity Index (EASI) from baseline at Week 16. Key secondary endpoints included percentage change in EASI at Week 24 and percentage of patients with at least 75% reduction from baseline in EASI (EASI-75), percentage of patients with Investigator Global Assessment response of 0 (clear) or 1 (almost clear) and a reduction from baseline of ≥2 points (IGA 0/1), and proportion of patients with a weekly average reduction of Peak Pruritus Numerical Rating Scale (PP-NRS) ≥4 points from baseline. The primary efficacy analysis included all randomised patients who completed Week 24 or discontinued treatment or study prior to Week 24 visit (n=390), whereas the safety analysis included all treated patients (n=388). Results Treatment with amlitelimab resulted in statistically significant improvements in percentage change in EASI from baseline to Week 16 compared to placebo for all four doses studied. The 250 mg with LD group had the numerically highest response versus placebo at Week 16, with a least-squares mean change from baseline of –32.1% (95% CI: –43.9, –20.3; P<0.0001); the remaining groups without LD had the following responses versus placebo: 250 mg, –27.3 (95% CI: –39.1, –15.6; P<0.0001); 125 mg, –22.2 (95% CI: –34.0, –10.4; P=0.0002); and 62.5 mg, –30.2 (95% CI: –41.9, –18.5; P<0.0001). There were also clinically meaningful improvements in all key secondary efficacy outcome measures, with all amlitelimab dose groups demonstrating nominally significant (P<0.05) efficacy versus placebo for EASI-75, IGA 0/1, and PP-NRS ≥4, except 250 mg (no LD) in IGA 0/1 at Week 16 (P=0.0562). Continued improvements were generally observed through Week 24 in primary and key secondary efficacy outcomes. Amlitelimab was well tolerated across all dose groups, with no safety concerns identified. Conclusions In this dose-ranging Phase 2b trial of amlitelimab in adults with moderate-to-severe AD, amlitelimab demonstrated clinically meaningful efficacy over 24 weeks with an acceptable safety profile across all four dose groups.

345 Treatment with amlitelimab—a novel non-depleting, non-cytotoxic anti-OX40Ligand monoclonal antibody—reduces IL-22 serum levels in a phase 2a randomized, placebo-controlled trial in patients with moderate-to-severe atopic dermatitis

Abstract OX40Ligand (OX40L) upregulation on antigen-presenting cells (APCs) following antigen presentation contributes to the survival and functional activation of T helper (Th) 2 and Th1/17/22 cells, which are central to the inflammation and pathological outcomes in atopic dermatitis (AD). In a Phase 2a trial (NCT03754309), amlitelimab, a fully human, non-depleting, non-cytotoxic anti-OX40L monoclonal antibody, was effective in treating patients with moderate-to-severe AD. To assess the effects of amlitelimab on interleukin (IL)-22, an important Th22-associated disease mediator of AD. 89 patients with moderate-to-severe AD were enrolled in a Phase 2a randomized, double-blind, placebo-controlled, multicentre trial and randomized 1 : 1 : 1 to intravenous amlitelimab low dose (200 mg loading/100 mg maintenance every 4 weeks [Q4W]; n = 29), high dose (500 mg/250 mg Q4W; n = 30) or placebo (Q4W; n = 29) until week 12. The primary efficacy endpoint was the percentage change in Eczema Area and Severity Index (EASI) from baseline to week 16. Other disease severity measurements included SCORing of Atopic Dermatitis (SCORAD) and validated Investigator Global Assessment (vIGA). Serum was collected at baseline, week 4 and week 16, and further collected at week 24 and week 36 in responders, defined as patients who reached vIGA 0/1 at week 16. IL-22 levels were determined by ultrasensitive single-molecule immunoassay (Simoa). Of 88 subjects who received study treatment, 59 were evaluable at week 16. Amlitelimab was well tolerated with an unremarkable safety profile. The mean percentage change in EASI from baseline at week 16 was −80.12% for amlitelimab low dose and −69.97% for amlitelimab high dose vs. −49.37% for placebo (nominal P-values: 0.009 [amlitelimab low dose vs. placebo] and 0.072 [amlitelimab high dose vs. placebo]). Amlitelimab-treated patients who achieved vIGA 0/1 at week 16 had sustained clinical responses up to week 36. No difference in IL-22 serum levels was found between groups at baseline. IL-22 levels correlated with disease severity at baseline, as measured by EASI and SCORAD (r = 0.53, P < 0.0001; and r = 0.36, P = 0.001; respectively). A significant reduction in IL-22 levels was observed at week 16 in amlitelimab-treated patients (low dose P < 0.0001; high dose P = 0.001), but not in the placebo group (P = 0.381). The amlitelimab-induced decrease in IL-22 levels was maintained until week 36 in those defined as vIGA 0/1 responders at week 16. OX40L blockade on APCs represents a promising novel approach in the treatment of AD by effectively targeting underlying T-cell immune dysregulation. Amlitelimab monotherapy not only provided a sustained and clinically meaningful improvement in disease activity compared with placebo in patients with moderate-to-severe AD, but also significantly decreased serum levels of IL-22, a Th22 cell-associated cytokine involved in the underlying immunopathogenesis of AD.

OX40L Inhibition Suppresses KLH-driven Immune Responses in Healthy Volunteers: A Randomized Controlled Trial Demonstrating Proof-of-Pharmacology for KY1005.

The safety, tolerability, immunogenicity, and pharmacokinetic (PK) profile of an anti‐OX40L monoclonal antibody (KY1005, currently amlitelimab) were evaluated. Pharmacodynamic (PD) effects were explored using keyhole limpet hemocyanin (KLH) and tetanus toxoid (TT) immunizations. Sixty‐four healthy male subjects (26.5 ± 6.0 years) were randomized to single doses of 0.006, 0.018, or 0.05 mg/kg, or multiple doses of 0.15, 0.45, 1.35, 4, or 12 mg/kg KY1005, or placebo (6:2). Serum KY1005 concentrations were measured. Antibody responses upon KLH and TT immunizations and skin response upon intradermal KLH administration were performed. PD data were analyzed using repeated measures analysis of covariances (ANCOVAs) and post hoc exposure‐response modeling. No serious adverse events occurred and all adverse events were temporary and of mild or moderate severity. A nonlinear increase in mean serum KY1005 concentrations was observed (median time to maximum concentration (Tmax) ~ 4 hours, geometric mean terminal half‐life (t½) ~ 24 days). Cutaneous blood perfusion (estimated difference (ED) −13.4 arbitrary unit (AU), 95% confidence interval (CI) −23.0 AU to −3.8 AU) and erythema quantified as average redness (ED −0.23 AU, 95% CI −0.35 AU to −0.11 AU) decreased after KY1005 treatment at doses of 0.45 mg/kg and above. Exposure‐response analysis displayed a statistically significant treatment effect on anti‐KLH antibody titers (IgG maximum effect (Emax) −0.58 AU, 95% CI −1.10 AU to −0.06 AU) and skin response (erythema Emax −0.20 AU, 95% CI −0.29 AU to −0.11 AU). Administration of KY1005 demonstrated an acceptable safety and tolerability profile and PK analyses displayed a nonlinear profile of KY1005. Despite the observed variability, skin challenge response after KY1005 treatment indicated pharmacological activity of KY1005. Therefore, KY1005 shows potential as a novel pharmacological treatment in immune‐mediated disorders.

Anti-OX40L monoclonal antibody prolongs secondary heart allograft survival based on CD40/CD40L and LFA-1/ICAM-1 blockade

BackgroundMemory T cells (Tms) form a barrier against long-term allograft survival; however, CD4+Foxp3+ regulatory T cells (Tregs) can suppress allograft rejection. The OX40/OX40L pathway is critical to the generation of Tms and turns off Treg suppressor function. MethodsB6 mice that rejected BALB/c skin grafts after 4weeks were used as the secondary heart transplant recipients. The skin recipient mice, termed S0, S2 and S3, were treated with the isotype antibodies, anti-CD40L/LFA-1 or anti-OX40L combined with anti-CD40L/LFA-1 mAbs, respectively. The secondary heart recipients, termed H0 and H2, received anti-CD40L/LFA-1 mAbs or not, respectively (Fig. 1). ResultsFour weeks after primary skin transplantation, the Tms in the S3 group that received anti-OX40L with anti-CD40L/LFA-1 mAbs were reduced compared to those in the S2 group (CD4+ Tm: 32.61±2.20% in S2 vs. 25.36±1.16% in S3; CD8+ Tm: 27.76±1.96% in S2 vs. 20.95±1.30% in S3; P<0.01). Meanwhile, the proportions of Tregs in S3 increased compared to those in S2 (P<0.05). The anti-OX40L with anti-CD40L/LFA-1 mAbs group (S3H2) prolonged the mean survival time (MST) following secondary heart transplantation from 9.5days to 21days (P<0.001). Furthermore, allogeneic proliferation of recipient splenic T cells and graft-infiltrating lymphocytes were significantly inhibited in the S3H2 group. Additionally, a higher level of IL-10 was detected in sera and allografts. ConclusionsAnti-OX40L mAb could prolong secondary heart allograft survival based on CD40/CD40L and LFA-1/ICAM-1 blockade. The mechanism of protecting allografts using anti-OX40L mAb involved impairing the generation of Tm and up-regulating IL-10 producing Tregs, inhibiting the function of T cells.

Erratum to: Memory T Cells Mediate Cardiac Allograft Vasculopathy and are Inactivated by Anti-OX40L Monoclonal Antibody

Erratum to: Memory T Cells Mediate Cardiac Allograft Vasculopathy and are Inactivated by Anti-OX40L Monoclonal Antibody

OX40L blockade and allergen‐induced airway responses in subjects with mild asthma

BackgroundThe OX40/OX40L interaction contributes to an optimal T cell response following allergic stimuli and plays an important role in the maintenance and reactivation of memory T effector cells.ObjectiveWe tested whether treatment with an anti-OX40L monoclonal antibody (MAb) would inhibit allergen-induced responses in subjects with asthma.MethodsTwenty-eight mild, atopic asthmatic subjects were recruited for a double-blind, randomized, placebo-controlled, parallel-group trial (ClinicalTrials.gov identifier NCT00983658) to compare blockade of OX40L using a humanized anti-OX40L MAb to placebo-administered intravenously in 4 doses over 3 months. Allergen inhalation challenges were carried out 56 and 113 days after the first dose of study drug. The primary outcome variable was the late-phase asthmatic response. Other outcomes included the early-phase asthmatic response, airway hyperresponsiveness, serum IgE levels, blood and sputum eosinophils, safety and tolerability.ResultsTreatment with anti-OX40L MAb did not attenuate the early- or late-phase asthmatic responses at days 56 or 113 compared with placebo. In the anti-OX40L MAb treatment group, total IgE was reduced 17% from pre-dosing levels, and sputum eosinophils decreased 75% by day 113 (both P = 0.04). There was no effect of anti-OX40L MAb on airway hyperresponsiveness or blood eosinophils. The frequency of AEs was similar in both groups.Conclusion and Clinical RelevancePharmacological activity of anti-OX40L MAb was observed by decreases in serum total IgE and airway eosinophils at 16 weeks post-dosing, but there was no effect on allergen-induced airway responses. It is possible that the treatment duration or dose of antibody was insufficient to impact the airway responses.

Memory T Cells Mediate Cardiac Allograft Vasculopathy and are Inactivated by Anti-OX40L Monoclonal Antibody.

Cardiac allograft vasculopathy (CAV) is a major complication limiting the long-term survival of cardiac transplants. The role of memory T cells (Tmem) in the pathogenesis of CAV remains elusive. This study investigated the role of Tmem cells in the development of CAV and the therapeutic potential of targeting the OX40/OX40L pathway for heart transplant survival. Tmem cells were generated in Rag-1(-/-) C57BL/6 (B6) mice by homeostatic proliferation (HP) of CD40L null CD3(+) T cells from B6 mice. Rag-1(-/-) B6 mice (H-2(b)) harboring Tmem cells received cardiac allografts from BALB/c mice (H-2(d)), and were either untreated or treated with anti-OX40L monoclonal antibody (mAb) (0.5mg/mouse/day) for 10days. Six weeks after HP, the majority of transferred CD40L(-/-) T cells in Rag-1(-/-) B6 mice were differentiated to CD44(high) and CD62L(low) Tmem cells. BALB/c heart allografts in Rag-1(-/-) B6 recipient mice in the presence of these Tmem cells developed a typical pathological feature of CAV; intimal thickening, 100days after transplantation. However, functionally blocking the OX40/OX40L pathway with anti-OX40L mAb significantly prevented CAV development and reduced the Tmem cell population in recipient mice. Anti-OX40L mAb therapy also significantly decreased cellular infiltration and cytokine (IFN-γ, TNF-α and TGF-β) expression in heart allografts. Tmem cells mediate CAV in heart transplants. Functionally blocking the OX40/OX40L pathway using anti-OX40L mAb therapy prevents Tmem cell-mediated CAV, suggesting therapeutic potential for disrupting OX40-OX40L signaling in order to prevent CAV in heart transplant patients.

Efficacy of pretreatment of allografts with methoxypolyethylene glycol-succinimidyl-propionic acid ester in combination with an anti-OX40L monoclonal antibody in relieving graft-versus-host disease in mice

We designed to determine whether pretreatment of allografts with methoxypolyethylene glycol-succinimidyl-propionic acid ester (mPEG-SPA) and an anti-OX40L monoclonal antibody (McAb) can relieve acute graft-versus-host disease in allogeneic bone marrow transplantation recipients. Responder splenocytes from C57BL/6 donor mice were incubated with stimulator splenocytes from BALB/c recipient mice for 7days in the presence or absence of anti-OX40L McAb followed by mPEG-SPA modification. Donor BM cells plus mixed culture T cells were then transplanted into myeloablatively irradiated BALB/c mice. The signs of GVHD were less evident in mice of groups B (mPEG-SPA modification group), C (anti-OX40L McAb pretreated group) and D (dual-treated group), with average survival durations all longer than those in group A (non-treated BMT group) (P<0.05). The survival rates on day 60 post-BMT in groups B, C and D were 50, 41.7 and 66.7%, respectively. After BMT, serum IL-4 and IL-10 levels elevated in groups B, C (P<0.05) and even more significantly increased in group D (P<0.01), while serum IFN-gamma levels decreased in these three groups (P<0.01). In conclusion, the combination of mPEG-SPA and anti-OX40L McAb can block T cell-activated antigens, co-stimulatory pathways and induce the immune shift of Th cells toward Th2 cells; their effects in ameliorating GVHD are synergistic.

Regulatory T Cells Are Critical to Tolerance Induction in Presensitized Mouse Transplant Recipients Through Targeting Memory T Cells

Memory T cells are a significant barrier to induction of transplant tolerance. However, reliable means to target alloreactive memory T cells have remained elusive. In this study, presensitization of BALB/c mice with C57BL/6 skin grafts generated a large number of OX40(+)CD44(hi)effector/memory T cells and resulted in rapid rejection of donor heart allografts. Recognizing that anti-OX40L monoclonal antibody (mAb) (alpha-OX40L) monotherapy prolonged graft survival through inhibition and apoptosis of memory T cells in presensitized recipients, alpha-OX40L was added to the combined treatment protocol of LF15-0195 (LF) and anti-CD45RB (alpha-CD45RB) mAb-a protocol that induced heart allograft tolerance in non-presensitized recipients but failed to induce tolerance in presensitized recipients. Interestingly, this triple therapy restored donor-specific heart allograft tolerance in our presensitized model that was associated with induction of tolerogenic dendritic cells and CD4(+)CD25(+)Foxp3(+) T regulatory cells (Tregs). Of note, CD25(+) T cell depletion in triple therapy recipients prevented establishment of allograft tolerance. In addition, adoptive transfer of donor-primed effector/memory T cells into tolerant recipients markedly reduced levels of Tregs and broke tolerance. Our findings indicated that targeting memory T cells, by blocking OX40 costimulation in presensitized recipients was very important to expansion of Tregs, which proved critical to development of tolerance.

Expression of OX40 ligand in microglia activated by IFN-γ sustains a protective CD4 + T-cell response in vitro

T-cell-dependent immunity in the central nervous system (CNS) is beneficial for neuroprotection, neurogenesis and even behavior. As a highly specialized site, the CNS is speculated to possess the means to maintain T-cell immune responses through its own resident cells. Therefore, we investigated whether microglia, the most potent antigen-presenting cells residing in the CNS, could sustain T-cell responses in vitro. We showed that interferon-γ (IFN-γ)-activated microglia (MG IFN-γ) inducibly expressed an important immune co-stimulatory molecule, OX40 ligand (OX40L). Co-culture of activated CD4 + T cells with MG IFN-γ significantly increased T-cell proliferation and decreased apoptosis, and these effects were markedly inhibited by addition of a neutralizing anti-OX40L monoclonal antibody. In addition, ligation of OX40L in MG IFN-γ enhanced their production of insulin-like growth factor I (IGF-I). These results suggest that the expression of OX40L in microglia provides a molecular basis for the maintenance of T-cell survival, expansion of T cells and increased secretion of remedial growth factor from MG IFN-γ, which may contribute to the protective effect in the CNS.

Effects of costimulatory pathway OX40/OX40L on the pathogenesis of allergic asthma in mice

Allergic asthma is thought to be mediated by CD4+ T lymphocytes producing the Th2-associated cytokines, which play a critical role in the development of the airway hyper-responsiveness and the eosinophilic inflammatory response. The costimulatory pathway CD28/B7 has been shown to play an important role in CD4+ T cell activation in allergic asthma. This study was conducted to evaluate the role of another costimulatory pathway OX40/OX40 ligand (L) in the pathogenesis of allergic asthma in BALB/c mice. An allergic asthma model in BALB/c mice was established. Thirty-six BALB/c mice were randomly divided into three groups with 12 in each. Mice in treatment group (group B) were treated with neutralizing anti-OX40L monoclonal antibody (mAb, 300 microg per mouse) during the sensitization period. Mice in two control groups, asthma model group (group A) and IgG antibody group (group C) were treated with normal saline (NS) and control IgG respectively instead of anti-OX40L mAb. Bronchoalveolar lavage fluid (BALF) was collected from the mice of each group for counting the total number of white blood cells (including neutrophil granulocyte, lymphocyte, monocyte and eosinophil granulocyte) and the proportions of these cells. The levels of IL-4 and INF-gamma in BALF were measured by ELISA. Lungs were removed for morphological examination after HE and PAS staining, and expression of OX40 in lungs was evaluated by immunohistochemical method. (1) The count of total number of white blood cells in BALF (x10(6)/ml) was lower in group B than that of group A and group C (26.6 +/- 4.6 vs. 36.8 +/- 5.2 and 34.3 +/- 6.9, respectively), the difference between the treatment group (group B) and two control groups (groups A and C) was significant; The proportions of eosinophils and lymphocytes in the BALF (%) were lower in group B than those in group A and group C (eosinophils 15.1 +/- 2.6 vs. 20.0 +/- 4.1 and 19.9 +/- 3.9, respectively; lymphocytes 7.0 +/- 0.9 vs. 8.9 +/- 1.6 and 8.6 +/- 1.8, respectively), the difference between the treatment group and two control groups was significant. (2) The IL-4 level in BALF (pg/ml) was lower in group B than that in group A and group C (672 +/- 58 vs. 809.57 +/- 106.00 and 784 +/- 58, respectively), but the INF-gamma levels in BALF (pg/ml) were higher than those in group A and group C (0.86 +/- 0.09 vs. 0.69 +/- 0.15 and 0.67 +/- 0.13 respectively), and all the differences were statistically significant. (3) The expression of OX40 in the lungs of mice in group B were at a lower level than that of group A and group C, and the morphological changes of asthma were ameliorated in the mice of the treatment group. The signs of mice in treatment group were obviously ameliorated as compared to the two control groups. Blocking the costimulatory pathway by administering the neutralizing anti-OX40L mAb during the sensitization period of allergic asthma model could balance the Th1/Th2 responses, inhibit lung inflammation and ameliorate the signs of mice model of asthma.

Ligation of OX40 (CD134) regulates graft-versus-host disease (GVHD) and graft rejection in allogeneic bone marrow transplant recipients

OX40 (CD134) is expressed on activated T cells; its ligand, OX40 ligand (OX40L) is expressed on dendritic cells, B cells, and activated endothelial cells. To determine how OX40-OX40L interaction affects graft-versus-host disease (GVHD), we used antagonistic anti-OX40L monoclonal antibody (mAb) or OX40(-/-) donor or OX40L(-/-) recipient mice. Similar degrees of GVHD reduction were observed with each approach. Despite the fact that OX40 is up-regulated on both CD4(+) and CD8(+) T cells isolated during GVHD, the major effects of OX40 ligation were on CD4(+) and not CD8(+) T-cell-mediated alloresponses as assessed in both GVHD and engraftment model systems. GVHD inhibition by blockade of the OX40/OX40L pathway did not require CD28 signaling. Some studies have indicated OX40 is essential for inducing T-helper type 2 (Th2) responses. However, in vivo blockade of OX40-OX40L interactions reduced GVHD mortality induced by either signal transducer and activator of transcription-6(-/-) (Stat-6(-/-)) (Th2-defective) or Stat-4(-/-) (Th1-defective) major histocompatibility complex (MHC)-disparate splenocytes, indicating that the GVHD-ameliorating effects did not require Stat-4 or Stat-6 signaling. Although OX40L has been reported to be expressed on activated T cells, no effects on GVHD were observed when OX40L(-/-) versus OX40L(+/+) T cells were infused in different models. These data provide insights as to the mechanisms responsible for OX40/OX40L regulation of GVHD.

Contribution of OX40/OX40 ligand interaction to the pathogenesis of rheumatoid arthritis.

OX40 ligand (OX40L) and OX40 (CD134) are a pair of cell surface molecules belonging to the TNF/TNF receptor family. Interaction of OX40L with its receptor OX40 is thought to be important in T cell activation through T cell/antigen-presenting cell interaction. However, involvement of these molecules in the pathogenesis of rheumatoid arthritis (RA) remains unclear. To explore the contribution of OX40/OX40L interaction to the pathogenesis of RA in vivo, we evaluated the effect of a neutralizing anti-OX40L monoclonal antibody (mAb) on the development of collagen-induced arthritis (CIA) in DBA/1 mice as an animal model for RA. Administration of anti-OX40L mAb into type II collagen (CII) -immunized DBA/1 mice dramatically ameliorated the disease severity. In vivo treatment with anti-OX40L mAb did not inhibit the expansion of CII-reactive T cells, but suppressed IFN-gamma and anti-CII IgG2a production. Therefore, OX40/OX40L interaction appears to play a critical role in the development of CIA by enhancing Th1-type autoimmune response. In addition, T lymphocytes in synovial fluid and synovial tissue from RA patients expressed OX40, while OX40L was expressed on sublining cells in synovial tissue. These results indicate that OX40/OX40L interaction may play a critical role in the development of RA.

Contribution of OX40/OX40 ligand interaction to the pathogenesis of rheumatoid arthritis

OX40 ligand (OX40L) and OX40 (CD134) are a pair of cell surface molecules belonging to the TNF/TNF receptor family. Interaction of OX40L with its receptor OX40 is thought to be important in T cell activation through T cell/antigen-presenting cell interaction. However, involvement of these molecules in the pathogenesis of rheumatoid arthritis (RA) remains unclear. To explore the contribution of OX40/OX40L interaction to the pathogenesis of RA in vivo, we evaluated the effect of a neutralizing anti-OX40L monoclonal antibody (mAb) on the development of collagen-induced arthritis (CIA) in DBA/1 mice as an animal model for RA. Administration of anti-OX40L mAb into type II collagen (CII) -immunized DBA/1 mice dramatically ameliorated the disease severity. In vivo treatment with anti-OX40L mAb did not inhibit the expansion of CII-reactive T cells, but suppressed IFN-gamma and anti-CII IgG2a production. Therefore, OX40/OX40L interaction appears to play a critical role in the development of CIA by enhancing Th1-type autoimmune response. In addition, T lymphocytes in synovial fluid and synovial tissue from RA patients expressed OX40, while OX40L was expressed on sublining cells in synovial tissue. These results indicate that OX40/OX40L interaction may play a critical role in the development of RA.

Impairment of antigen-presenting cell function in mice lacking expression of OX40 ligand.

OX40 expressed on activated T cells is known to be an important costimulatory molecule on T cell activation in vitro. However, the in vivo functional significance of the interaction between OX40 and its ligand, OX40L, is still unclear. To investigate the role of OX40L during in vivo immune responses, we generated OX40L-deficient mice and a blocking anti-OX40L monoclonal antibody, MGP34. OX40L expression was demonstrated on splenic B cells after CD40 and anti-immunoglobulin (Ig)M stimulation, while only CD40 ligation was capable of inducing OX40L on dendritic cells. OX40L-deficient and MGP34-treated mice engendered apparent suppression of the recall reaction of T cells primed with both protein antigens and alloantigens and a significant reduction in keyhole limpet hemocyanin-specific IgG production. The impaired T cell priming was also accompanied by a concomitant reduction of both T helper type 1 (Th1) and Th2 cytokines. Furthermore, antigen-presenting cells (APCs) derived from the mutant mice revealed an impaired intrinsic APC function, demonstrating the importance of OX40L in both the priming and effector phases of T cell activation. Collectively, these results provide convincing evidence that OX40L, expressed on APCs, plays a critical role in antigen-specific T cell responses in vivo.