(T0930-02-09) Short-Term Administration with Immunosuppressants Suppresses Preclinical Immunogenicity against a Human Monoclonal Antibody in Rats

Purpose: Immunogenicity can often alter the concentration of therapeutic proteins in pharmacokinetic and toxicology studies during preclinical drug development. Human therapeutic proteins such as monoclonal antibodies (mAb) are often immunogenic in animal studies leading to the formation of anti-drug antibodies (ADA). ADA formation may reduce the systemic exposure of the mAb by forming immune complexes and triggering their elimination. The purpose of this preliminary study was to evaluate if long-term immune tolerance towards a human mAb can be induced in rats with short-term administration of immunosuppressants.

Methods: We performed a 12-week tolerance study in rats with 4 parallel treatment groups (n=8 per group) that received erenumab (Aimovig ®), a human monoclonal antibody that does not bind to any specific target in rats and is used here as a stereotypical human mAb. The study consisted of an induction phase (week 1-4), a wash-out phase (week 5-8), and a re-challenge phase (week 9-12). In all four groups, animals (male, Sprague-Dawley) received 10 mg/kg erenumab by subcutaneous injection on the first day of week 1, 2, 3, 4, 9, 10, 11 and 12. While rats in Group 1 did not receive any immunosuppression, animals in groups 2, 3, and 4 received treatments with methotrexate or a sirolimus/tacrolimus combination during the induction and washout phase as summarized in Table 1. Assessments for erenumab PK and ADA were made at predefined time points. Predose and 72 hours post dose samples were obtained in week 1 to 12 for PK and ADA analysis. In addition, PK samples were drawn at 2, 8 and 24 hours post dose in week 1, 4, 9 and 12. Terminal blood was drawn in week 14 for PK and ADA assessment. Blood samples (300 µL) were drawn via tail vein. Serum was separated by centrifugation (2000×g for 15 min at 4°C) and stored at -80°C until analysis. Erenumab concentrations were measured using streptavidin coated plates coated with biotin-tagged mouse anti-human IgG against the heavy chain constant region of erenumab. A sulfo-tagged goat anti-human IgG against the human lambda light chain constant region of erenumab was used to detect the captured erenumab. The ECL signal was measured using SECTOR® Imager 600 plate reader. The range of this assay was 0.078 - 10 µg/ml. ADA assessments were performed using a bridging immunogenicity assay. Biotin-tagged erenumab was used to capture the ADA. Serum samples were treated with acetic acid and neutralized with Tris-HCL and sulfo-tagged erenumab was used for detection. The response was reported as ratio of sample ECL to negative control ECL (rat serum) as a signal to noise ratio.

Results: In this pilot study, the four treatment groups exhibited different incidences in ADA formation. In Group 1, the group without immunosuppressive therapy, no animals had ADA in the induction phase but 25% of animals exhibited ADA formation against erenumab at the end of the re-challenge phase. Group 2 (MTX 5 mg/kg) had 25% ADA incidence in the induction phase and 50% at the end of the re-challenge phase. Group 3 (MTX 3 mg/kg) had no ADA incidence in the induction phase and 12.5% ADA incidence at the end of the re-challenge phase. In contrast, group 4, the combination of tacrolimus and sirolimus, had 0% ADA incidence in both the induction and the re-challenge phase. The occurrence of ADA formation in the re-challenge phase of group 1 resulted in the expected reduction of erenumab systemic exposure (AUC_w9-12: ADA- 176 ± 12.2 h*mg/mL vs. ADA+ 44.3 ± 58.5 h*mg/mL), while all animals in group 4 maintained full erenumab exposure (AUC_wk9-12: 145 ± 18.7 h*mg/ mL).

Conclusion: Our preliminary results suggest that a short-term induction treatment of a combination with tacrolimus and sirolimus administered concurrently with a human mAb may be a promising approach to induce immune tolerance and thus suppress ADA formation in preclinical studies in rats. Confirmatory studies with adequate statistical power are currently ongoing to further investigate these preliminary findings.


Table 1