Abstract
Objectives
Antibody-based immunotherapies represent a sizable portfolio of cancer therapies largely based on the immunoglobulin G (IgG) class (Tang et al, 2021). However, in recent years the therapeutic potential of IgE mediating allergic responses has emerged as an attractive modality for targeting solid tumours (McCraw et al., 2021). The clinical use of IgEs as immunotherapies may offer formulation advantages since lower formulation concentrations are required for each dosage form. However, limited case studies on IgE formulation and physical stability characteristics are available in the literature. Therefore, we aimed to provide insights into the stability of a novel IgE in response to formulation pH and environmental (freeze-thaw, thermal, agitation) stress conditions using orthogonal analytical techniques.
Materials and methods
We exposed IgE to different formulation pH (5.5, 6.5, 7.5) to determine the optimal conditions for further exposure to physical stresses (thermal, freeze-thaw, agitation) encountered during product shelf life. Changes in physical stability were quantified using orthogonal analytical techniques, including dynamic light scattering, particle tracking analysis, size exclusion chromatography (SEC), and asymmetric flow field flow fractionation (AF4) hyphenated with UV and multiangle light scattering detectors.
Results
Formulation of IgE in pH 6.5 buffer resulted in high monomeric purity and no submicron proteinaceous particulate detection, whereas formulation at pH 5.5 and 7.5 induced significant submicron particle formation (100-400 nm). IgE formulation in pH 6.5 resisted self-association following multiple freeze-thaw cycles, retaining a high percentage of monomeric purity. Exposure to thermal stress at elevated temperatures (56°C and 80°C), resulted in a loss of monomer and a significant increase in Z-average (p<0.001). Agitation stress induced submicron particle formation (100-500 nm) but maintained a high monomeric purity after dilution in running buffers used for AF4 and SEC analysis.
Conclusions
We implemented a range of bioanalytical techniques to study changes occurring in the physical stability of IgE. Overall, we show that IgE is resistant to freeze-thaw, aggregates reversibly after agitation stress, aggregates irreversibly at elevated temperatures, and is optimally formulated in pH 6.5. This work may guide the development of future formulations of IgE-based immunotherapies.
Antibody-based immunotherapies represent a sizable portfolio of cancer therapies largely based on the immunoglobulin G (IgG) class (Tang et al, 2021). However, in recent years the therapeutic potential of IgE mediating allergic responses has emerged as an attractive modality for targeting solid tumours (McCraw et al., 2021). The clinical use of IgEs as immunotherapies may offer formulation advantages since lower formulation concentrations are required for each dosage form. However, limited case studies on IgE formulation and physical stability characteristics are available in the literature. Therefore, we aimed to provide insights into the stability of a novel IgE in response to formulation pH and environmental (freeze-thaw, thermal, agitation) stress conditions using orthogonal analytical techniques.
Materials and methods
We exposed IgE to different formulation pH (5.5, 6.5, 7.5) to determine the optimal conditions for further exposure to physical stresses (thermal, freeze-thaw, agitation) encountered during product shelf life. Changes in physical stability were quantified using orthogonal analytical techniques, including dynamic light scattering, particle tracking analysis, size exclusion chromatography (SEC), and asymmetric flow field flow fractionation (AF4) hyphenated with UV and multiangle light scattering detectors.
Results
Formulation of IgE in pH 6.5 buffer resulted in high monomeric purity and no submicron proteinaceous particulate detection, whereas formulation at pH 5.5 and 7.5 induced significant submicron particle formation (100-400 nm). IgE formulation in pH 6.5 resisted self-association following multiple freeze-thaw cycles, retaining a high percentage of monomeric purity. Exposure to thermal stress at elevated temperatures (56°C and 80°C), resulted in a loss of monomer and a significant increase in Z-average (p<0.001). Agitation stress induced submicron particle formation (100-500 nm) but maintained a high monomeric purity after dilution in running buffers used for AF4 and SEC analysis.
Conclusions
We implemented a range of bioanalytical techniques to study changes occurring in the physical stability of IgE. Overall, we show that IgE is resistant to freeze-thaw, aggregates reversibly after agitation stress, aggregates irreversibly at elevated temperatures, and is optimally formulated in pH 6.5. This work may guide the development of future formulations of IgE-based immunotherapies.
| Original language | English |
|---|---|
| Publication status | Published - 12 Nov 2024 |
| Event | JPAG Pharmaceutical Analysis Research Awards and Careers Fair 2024 - London, United Kingdom Duration: 12 Nov 2024 → 12 Nov 2024 https://www.jpag.org/?p=meetings&r=174 |
Conference
| Conference | JPAG Pharmaceutical Analysis Research Awards and Careers Fair 2024 |
|---|---|
| Country/Territory | United Kingdom |
| City | London |
| Period | 12/11/24 → 12/11/24 |
| Internet address |
