(T1030-01-04) Short-Term Tacrolimus/Sirolimus Combination Therapy Induces Long-Term Immune Tolerance to a Human Monoclonal Antibody in Rats

Purpose: Administration of human therapeutic proteins (TPs) such as monoclonal antibodies (mAbs) to animals during preclinical drug development may be accompanied by an immune response. This includes development of anti-drug antibodies (ADA) which bind to mAb and may reduce the systemic exposure of the mAb through enhancing its immune complex mediated clearance. ADA formation may thus hinder the preclinical pharmacokinetic (PK) and toxicology assessments of mAbs. To mitigate immune mediated impact, we have explored the ability of immunosuppressive drugs (tacrolimus and sirolimus or methotrexate (as a monotherapy)) to induce long-term immune tolerance towards a human mAb in rats with a short-term administration of immunosuppressants and thereby maintaining mAb systemic exposure in study animals (1). The purpose of this study was to further confirm our preliminary findings (1). The aim was to investigate the ability of short-term treatment with a combination of tacrolimus and sirolimus to suppress ADA formation to a human mAb aggregates in rats, to induce immune tolerance beyond the treatment period with the immunosuppressants and to confirm its ability to maintain mAb serum exposure in treated animals throughout the study period.

Methods: Erenumab (Aimovig^®), a human mAb that does not bind to any specific target in rats is used here as a representative TP. Erenumab aggregates were prepared by stressing erenumab monomer solution (1 mg/mL) stirred at room temperature for 24 hours at 760 rpm. We performed a 12-week tolerance study in rats with 2 treatment groups (n=10 per group; male Sprague Dawley rats). The study was divided into 3 phases: induction (week 1-4), washout (week 5-8), and rechallenge (week 9-12). In Group 1, animals received 1 mg/kg erenumab aggregate solution by subcutaneous injection on the first day of week 1, 2, 3, 4, 9, 10, 11 and 12; without any immune suppression. In contrast, rats in Group 2 received the same erenumab injections but also received treatment with tacrolimus and sirolimus during the induction and washout phase according to Table 1. Blood samples were drawn from each animal at predefined time points and serum was obtained for measuring erenumab concentration and ADA. Predose samples were obtained in week 1 to 12 for PK and ADA analysis. In addition, PK samples were collected at 2, 8 and 24 hours post dose in week 1, 4, 9 &12 and 72 hours post dose in week 1 to 12. ELISA assays were utilized for PK and ADA assessments (1).

Results: Treatment with erenumab aggregates in group 1 showed 30% ADA incidence at the end of induction phase and 60% at the end of rechallenge phase. Group 2 (aggregates with immunosuppression) animals had 0% ADA incidence in both the induction and the rechallenge phase. We observed a statistically significant difference in ADA incidence between group 1 and group 2 (Fisher’s exact test; p=0.01). The ADA+ animals of group 1 showed the expected reduction in erenumab systemic exposure during the rechallenge phase (AUC_w9-12: 0.7 ± 1.1 h*mg/mL) compared to ADA - animals (AUC_w9-12: 3.6 ± 0.4 h*mg/mL). All animals in group 2 maintained erenumab exposure (AUC_wk9-12: 3.4 ± 0.2 h*mg/ mL) similar to what was seen in group 1 ADA- animals. Systemic erenumab exposure (AUC_w9-12) was also statistically different (Unpaired t-test; p= 0.003) between ADA+ animals of group 1 and ADA- animals of group 2.

Conclusion: This study confirms our previous study results (1) that a short-term administration with a combination of tacrolimus and sirolimus administered concurrently with a human mAb is a promising approach to induce immune tolerance and thus prevent ADA formation and thereby maintain drug exposure in preclinical mAb studies in rats. The lack of immune response in tacrolimus and sirolimus treated rats following rechallenge with the mAb, also suggests a long-term immune tolerance. Further confirmatory studies with other therapeutic proteins are necessary to explore full potential of this approach.

References: 1. P.Gupta, A. Srivastava, J. Ryman, M. Swanson, V. Jawa and B. Meibohm, “Short-Term Administration with Immunosuppressants Suppresses Preclinical Immunogenicity against a Human Monoclonal Antibody in Rats”, AAPS National Biotechnology Conference, May 2022, Anaheim, USA.


Table 1. Treatment Groups