1. Introduction
Ligelizumab (also known as QGE031) is a highly potent, humanized IgG1 isotype monoclonal antibody that binds to human immunoglobulin E (IgE) [
1]. Upon binding to specific epitopes in the C3 region of IgE, ligelizumab blocks the interaction of free IgE with both the high- and low-affinity IgE receptors (FcεRI and FcεRII/CD23). Ligelizumab treatment results in rapid binding of free IgE, resulting in dose- and time-dependent neutralization of free IgE by rendering it inaccessible to IgE receptors on mast cells and basophils [
2,
3].
Ligelizumab has been explored in selected atopic conditions and autoimmune disorders for which there is evidence of an involvement of the IgE pathway, such as asthma [
2,
3,
4]. Currently, ligelizumab is being studied for the treatment of food allergies (NCT04984876). It has also been studied in chronic spontaneous urticaria (CSU), a common disease of the skin characterized by the development of pruritic hives that can be associated with angioedema, and its efficacy and safety have been demonstrated [
5,
6,
7,
8].
During the development of ligelizumab, several drug substance manufacturing changes were introduced. Ligelizumab is formulated as a 120 mg/1 mL solution for subcutaneous (s.c.) injection in glass vials as a liquid-in-vial formulation (LIVI), and as a solution for s.c. injection in a prefilled syringe (PFS) in needle safety devices with 1 mL glass syringes. The changes between LIVI used in the CSU Phase 3 studies and PFS used in the CSU Phase 3 extension study, and intended for commercial use, were solely at the level of the drug product and included changes related to the drug product manufacturing line, the manufacturing process, the primary packaging, and the device. There was no change in the drug product formulation or in the drug substance, as compared to the material used in the CSU Phase 3 studies.
To optimize the s.c. delivery system of ligelizumab, including the injection volume and duration, the formulation (LIVI vs. PFS) and administration method (patient self-administration vs. healthcare provider (HCP) administration), the pharmacokinetics (PK), pharmacodynamics (PD), and safety data were evaluated in two Phase 1 studies in healthy volunteers (HVs) and four Phase 2 and 3 studies in patients with chronic spontaneous urticaria (CSU). Within- and between-study comparisons in both populations informed the optimization of the s.c. delivery system of ligelizumab and supported the selection of the formulation and administration method in its commercial use.
2. Materials and Methods
Data from two Phase 1 studies in HVs (CQGE031B2101 (B2101) and CQGE031C2101 (C2101)), a phase 2 study (CQGE031C2202 (C2202)) [
7], and three Phase 3 studies (pivotal studies CQGE031C2302 (C2302) and CQGE031C2303 (C2303) [
8], and an extension study CQGE031C2302E1 (C2302E1)) in patients with CSU for ligelizumab were included in the assessment and optimization of its s.c delivery system (
Figure S1). The formulations of ligelizumab used in all the studies are summarized in
Table 1. All the clinical studies were conducted in accordance with the principles of the International Conference on Harmonization requirements for Good Clinical Practice, the Declaration of Helsinki, and with the approval of a National Health Service Ethics Review Committee.
2.1. Phase 1 Studies in Healthy Volunteers
2.1.1. Study B2101
Study B2101 was a Phase 1 double-blind study to investigate the safety, tolerability, PK, and PD of single doses of ligelizumab administered via s.c. injection or infusion (via syringe pumps) using the LIVI formulation in HVs. In particular, this study investigated whether increased volumes of ligelizumab administered via different delivery systems showed an acceptable local tolerability profile. The differences in the delivery systems are the site of injection (thigh or abdomen) and the mode of administration (s.c. infusion or s.c. injection). Subjects in Part 1 (32 subjects) were randomly assigned to 1 of 6 treatments (treatment 1, 2, 3, 4, 5, or 6) in a planned ratio of 8:2:8:2:8:2. The samples for PK and total/free IgE were collected at baseline, Day 1 (at 2 h, 4 h, and 12 h), Days 2, 3, and 5, and Weeks 2, 3, 5, 7, 9, 11, 13, 15, and 18, and the samples for the antidrug antibody (ADA) were collected at baseline and Days 29, 57, 85, and 113.
Subjects in Part 2 (11 subjects) were randomly assigned to 1 of 2 treatments (treatment 7 or 8) in a planned ratio of 8:2.
Part 1 treatments
Treatment 1: Two s.c. bolus injections of ligelizumab 120 mg/1.0 mL in two different areas (right and left thigh), administered manually in a sequential fashion.
Treatment 2: Two s.c. bolus injections of placebo 0 mg/1.0 mL in two different areas (right and left thigh), administered manually in a sequential fashion.
Treatment 3: Single s.c. bolus injection of ligelizumab 240 mg/2.0 mL within 15 s, administered manually in the thigh (the non-dominant thigh was used unless clinically not appropriate).
Treatment 4: Single s.c. bolus injection of placebo 0 mg/2.0 mL within 15 s, administered manually in the thigh (the non-dominant thigh was used unless clinically not appropriate).
Treatment 5: Single s.c. infusion of ligelizumab 240 mg/2.0 mL over approximately 5 min, administered via a syringe pump in the abdomen.
Treatment 6: Single s.c. infusion of placebo 0 mg/2.0 mL over approximately 5 min, administered via a syringe pump in the abdomen.
Part 2 treatments
Treatment 7: Single s.c. infusion of ligelizumab 420 mg/3.5 mL over approximately 9 min, administered via a syringe pump in the abdomen.
Treatment 8: Single s.c. infusion of placebo 0 mg/3.5 mL over approximately 9 min, administered via a syringe pump in the abdomen.
2.1.2. Study C2101
Study C2101 was an open-label, randomized, parallel-group, comparative PK study of ligelizumab administered as a LIVI or via a PFS in HVs to compare the PK of ligelizumab following a single s.c. administration of 120 mg ligelizumab, as the LIVI formulation (reference), against the PFS presentation (test). A total of 132 subjects were enrolled in the study and randomized in a 1:1 ratio, with 67 subjects in the LIVI treatment group and 65 in the PFS treatment group. PK samples were collected at baseline, Day 1 (at 2 h, 4 h, and 12 h), Days 2, 3, and 5, and Weeks 2, 3, 5, 7, 9, 11, 13, 15, and 18. Total IgE samples were collected at baseline and Days 3, 29, 57, 85, and 125, and ADA samples were collected at baseline and Days 29, 57, 85, and 125.
2.2. Phase 2 and 3 Studies in CSU Patients
2.2.1. Study C2202
Study C2202 (NCT03437278) was a multicenter, randomized, double-blind, placebo-controlled Phase IIb dose-finding study to investigate the efficacy and safety of ligelizumab (QGE031) in adolescent patients with CSU [
7]. Ligelizumab was administered s.c. to patients at 24 mg or 120 mg once every 4 weeks (Q4W) as a LIVI formulation. Pre-dose PK and total IgE samples were collected on Days 1, 8, 29, 57, 85, 113, 141, 169, 225, and 281.
2.2.2. Studies C2302 and C2303
Studies C2302 (NCT03580369) and C2303 (NCT03580356) were multicenter, randomized, double-blind, active, and placebo-controlled studies conducted to investigate the efficacy and safety of ligelizumab in the treatment of CSU in adolescents and adults inadequately controlled with H1-antihistamines [
8]. Ligelizumab was administered s.c. to patients at 72 mg or 120 mg Q4W as a LIVI formulation. Pre-dose PK and total IgE samples were collected on Days 1, 29, 85, 141, 169, 197, 253, 309, 337, 365, and 449.
2.2.3. Study C2302E1
Study C2302E1 (NCT04210843) was a multicenter, double-blind, and open-label extension study conducted to evaluate the efficacy and safety of ligelizumab as a retreatment, self-administered therapy, and monotherapy in CSU patients who completed core studies, including C2202, C2302, and C2303. In this extension study, patients from the preceding core studies were administered with ligelizumab 120 mg/1 mL as a LIVI Q4W but switched to ligelizumab 120 mg/1 mL as a PFS Q4W from Week 12 onwards. At this time, subjects were offered an opportunity to administer ligelizumab outside the clinic. Subjects who expressed interest in self-administration outside the clinical setting and fulfilled the suitability criteria performed their first outside-the-clinic self-administration at Week 24. Subjects who did not self-administer at Week 24 were evaluated for self-administration at Week 52. In the data analysis, the self-administration subpopulation was defined as the subjects who had at least one injection with PFS administered by the participant themself or a caregiver, either in the clinic or outside the clinic. The in-clinic staff-administration was defined as the subjects who had all injections with the PFS administered in the clinic and by the site staff.
2.3. Sample Analysis
Total ligelizumab serum concentrations were determined by a validated enzyme-linked immunosorbent assay (ELISA), which was established and validated at several sites over time. The lower limit of quantification (LLOQ) was 0.5 µg/mL in studies B2101 and C2202, 0.2 µg/mL in study C2101, and 1.0 µg/mL in studies C2302, C2303, and C2302E1. The PK assay format remained constant but was refined over time. Thereby, sensitivity varied from 0.2 to 1.0 µg/mL, but complied with all validation acceptance criteria and the needs of the program. The measured LLOQ and ULOQ (upper limit of quantification) are required to be within a bias of ±25% from the nominal concentrations, while for all other QCs the acceptable bias is within ±20%.
Free IgE serum concentrations in the B2101 study were determined by a validated ELISA, with an LLOQ of 2.50 ng/mL and a dynamic range up to 250 ng/mL.
Total IgE (free IgE plus IgE bound to ligelizumab) serum concentrations were determined on the commercial ImmunoCAP platform with the ImmunoCAP total IgE assay. The LLOQ was 2 IU/mL, corresponding to an IgE concentration of 4.8 ng/mL, and the assay dynamic range was up to 5000 IU/mL, corresponding to 12,000 ng/mL IgE.
Anti-ligelizumab antibodies (anti-drug antibodies (ADAs)) were detected using a validated homogeneous bridging assay based on biotin- and ruthenium-labeled ligelizumab, with read-out on the Meso Scale Discovery (MSD) platform. Assay interferences caused by the target and drug required a dedicated sample treatment: IgEs were removed via magnetic beads coated with anti-IgE antibodies and drug interference was reduced via an acid treatment step.
2.4. Data Analysis
Data analyses were conducted on the PK analysis sets that included all subjects who had PK data available and received at least one dose of the study drug. Where applicable, PK parameters of ligelizumab, such as the area under the curve from time zero to the last measurable concentration (AUClast) and from time zero to infinity (AUCinf), maximum serum concentration after dose administration (Cmax), time to reach Cmax (Tmax), elimination half-life (T1/2), and apparent clearance (CL/F), were derived by non-compartmental analysis (NCA). Concentrations below the LLOQ (BLOQ) were treated as missing for the calculation of the geometric means and geometric CV%, and as zero in concentration summary statistics and for PK parameter calculations.
In study C2101, a statistical analysis was performed on the PK parameters of PFS (Test) and LIVI (Reference) in order to assess their comparability. Natural log-transformed ligelizumab PK parameters (Cmax, AUClast, and AUCinf) were analyzed using a linear model, which included treatment as a fixed factor. The estimated mean and 90% confidence interval (CI) of the treatment difference between ligelizumab PFS and LIVI were back-transformed (from natural log-transformed to linear) to obtain the geometric mean ratio and the 90% CI of the ratio.
For a cross-study comparison, the values of the trough serum concentration (Ctrough) of ligelizumab at steady state were averaged by subject and compared between the core studies C2202, C2303, and C2302 and the extension study C2302E1 (this corresponded to data from Week 12 onwards in studies C2302, C2303, and C2202 and from Weeks 24 and 52 in study C2302E1).
The data presented from studies B2101, C2101, and C2202 are based on the final database lock. The data presented for C2302, C2303, and C2302E1 are based on interim analyses that were conducted when all adult subjects had completed the Week 52 visit in the studies C2302 and C2303.
4. Discussion
PK and PD of ligelizumab have been characterized in atopic subjects following single-dose intravenous (i.v.) administration and multiple-dose s.c. administration [
3]. Following both i.v. and s.c administration, ligelizumab exhibited PK properties expected of an IgG antibody, characterized by a bi-exponential decline with a rapid initial and slower terminal disposition phase. Dose-dependent PK as a result of target-mediated drug disposition was observed over the i.v. dose range of 0.1–10 mg/kg and the s.c. dose range of 0.2–4 mg/kg. The half-life was dose-dependent and was 17–23 days at the i.v. doses of 3 and 10 mg/kg [
3]. Target engagement was demonstrated by dose- and time-dependent suppression of free IgE, basophil FcεRI, and basophil surface IgE, as well as the accumulation of total IgE (free and bound IgE) [
3]. Ligelizumab has been investigated in different patient populations, including patients with asthma and CSU. Based on the efficacy and safety data of a Phase 2 study in CSU patients [
5], ligelizumab was studied in two pivotal Phase 3 trials in CSU patients at the s.c. doses of 72 and 120 mg Q4W [
8].
Various formulations have been developed and used in the clinical trials of ligelizumab during its course of clinical development. Given the LIVI formulation was selected for Phase 2 and 3 clinical trials in patients, a Phase 1 study (B2101) was conducted in HVs to optimize the injection volume and duration of LIVI. The results of study B2101 informed the delivery system of the LIVI formulation used in Phase 2 patient trials and the pivotal Phase 3 trials in CSU patients. The PFS formulation was introduced in the Phase 3 extension study in CSU patients and was intended for commercial use.
Data in the Phase 1 study B2101 showed that ligelizumab AUC and Cmax were comparable across dose groups receiving the 240 mg dose of the LIVI formulation, regardless of the method of administration (injection volume and duration). Exposure to ligelizumab after administration of the 420 mg dose, when normalized to the dose, was also consistent with exposures at the 240 mg dose, suggesting linear PK between 240 and 420 mg. T
max and T
1/2 values were comparable across all dose groups regardless of the dose level or the method of administration, and were consistent with previously reported data [
3]. As ligelizumab inhibits binding of IgE to both its high- and low-affinity receptors, FcεRI and CD23, respectively, after administration of ligelizumab across studies, free IgE levels were suppressed and total IgE concentrations were increased, reflecting ligelizumab target binding. In this study, free IgE was almost completely suppressed and accumulation of total IgE during exposure to ligelizumab was observed, regardless of injection volumes or durations or the method of administration (injection or infusion). These free and total IgE data were in line with a previous report [
3]. Based on these results, the LIVI formulation with ligelizumab formulated as a 120 mg/1 mL concentrate as a solution for infusion/injection in glass vials was used in Phase 3 pivotal trials in CSU patients.
To bridge the LIVI formulation used in Phase 2 and 3 clinical trials and the PFS formulation intended for commercial use, the Phase 1 PK comparability study C2101 in HVs was conducted to compare the LIVI and PFS. The geometric mean ratios for Cmax, AUClast, and AUCinf were 1.07, 1.13, and 1.14, respectively. The 90% CI for Cmax was within the conventional bioequivalence range (0.8, 1.25), and the upper bounds of the 90% CI for AUClast (1.01, 1.26) and AUCinf (1.02, 1.27) were only slightly above (0.01 and 0.02, respectively) the upper limit of the bioequivalence range. Therefore, ligelizumab PK exposures were considered comparable between the LIVI and PFS groups. Besides HV data, patient data in were also compared between the LIVI and PFS based on clinical trials in patients with CSU (one Phase 2, two pivotal Phase 3, and one Phase 3 extension trial). Both intra-subject and inter-study comparisons indicated no apparent difference in the steady-state Ctrough between LIVI and PFS, confirming the results in HVs. Total IgE increased similarly for the two formulations in both HVs and patients. PFS administration was better tolerated than LIVI, with lower incidence rates of injection site reactions; however, no formal comparison was performed to draw a definitive conclusion. Overall, the data in both HVs and CSU patients demonstrated that the small changes (LIVI vs. PFS) during the formulation development process of ligelizumab did not have a clinically significant impact on the in vivo performance of the drug product, supporting the switch from the LIVI to the PFS in clinical practice without the need for dose adjustments.
PFS, the formulation intended for commercial use, could be administered by patients (self-administration) or their caregivers/HCPs. Self-administration will bring benefits to both patients and HCPs, including increased treatment adherence, reduced frequency of clinic visits, convenience, and economic benefits for the patient and the healthcare system [
9,
10]; therefore, it was evaluated during the clinical development of ligelizumab. Our data showed that PK was comparable for the PFS formulation between the self-administration and in-clinic staff-administration subpopulations, and the PD effect (change of the total IgE level) and tolerability were also comparable. These data support the self-administration of ligelizumab using PFS.
ADA data were consistent across delivery systems and the two different formulations (LIVI and PFS). Tolerability data also showed that ligelizumab was well tolerated and comparable across different delivery systems, formulations, or administrators, consistent with the conclusion of PK and PD data.
In formulation development, clinical pharmacology or PK studies in HVs are often conducted to compare the PK of different formulations to evaluate the in vivo impact of the formulation drug treatment effect. In our work, the comparability of the LIVI and the PFS of ligelizumab was established in a dedicated PK study in HVs. Moreover, the comparability was also demonstrated in Phase 2 and 3 clinical trials. Clinical pharmacology studies are conducted under restricted and well-controlled conditions, offering a gold standard to assess the clinical effect of a variable of interest (e.g., formulation). Phase 2 and 3 trials, on the other hand, represent the clinical setting in patient populations, administered with multiple doses in long-term treatment. The consistency of the patient data with the HV data strengthened the conclusion that the LIVI and the PFS of ligelizumab were comparable. Furthermore, besides PK and tolerability, the PD marker, total IgE level, was also assessed in both HVs and patients. The similarity of the total IgE data between the LIVI and PFS formulations also strengthened the PK comparability, further confirming that there was no clinically relevant difference between the two formulations.