Impact of Excipients Blending on Sugar-Stabilized Therapeutic Proteins
Abstract
Sugars have long been used as stabilizing excipients for protein-based therapeutics during the development and production process. Incorporating stabilizing agents, such as sucrose and trehalose into dry formulations is a common strategy approved by the US FDA. However, single-sugar formulations may not be sufficient for diverse medicinal products, necessitating the blending of excipients to improve stability, dissolution, and patient acceptability. Our goal is to comprehensively understand the impact of excipients, focusing on the impact of various sugar types on formulation stability, and to explore the effectiveness of incorporating additional excipients such as amino acids, surfactants, and polyols, to further enhance protein stability. The review begins with an overview of sugars’ used in proteins stabilization and the blending of excipients. It discusses the limitation of single-sugar formulations and explores alternatives such as oligo-or polysaccharides. The mechanism underlying sugar and excipient combinations is elucidated, emphasizing the benefits of blending multiple excipients for stabilizing therapeutic protein. Our findings demonstrate that excipients blending presents a promising strategy for improving the stability of sugar-stabilized therapeutic proteins. Combining sugars with other excipients such as amino acids and polyols effectively enhances formulation stability. Optimization of excipients ratios and quantities is crucial for achieving the desired stability profile for each specific protein. Considering the impact of excipients on stabilization is essential in therapeutic proteins development. Employing blends of various excipients in different ratios and quantities ensures appropriate stability and functionality of the protein formulations.
Introduction
Over the past decade, therapeutic proteins have emerged as lifesaving treatments, with their high efficacy and specificity leading to a substantial increase in their usage worldwide [1]. As evidenced by projections, the global protein therapeutics market is anticipated to undergo substantial growth, with its size estimated at USD 516.79 billion in 2024 and expected to reach USD 761.80 billion by 2029, reflecting a robust compound annual growth rate (CAGR) of 8.07% during the forecast period (2024-2029) [2]. This projection underscores the anticipation of the global protein therapeutics market doubling in size over the next decade, driven by the increasing demand; there is an urgent need for these therapeutic agents [3]. However, ensuring the potency and stability of therapeutic proteins poses a significant challenge due to the complexity and instability of their physical and chemical structure. Degradation of these proteins not only compromises their efficacy, but can also lead to severe side effects [1]. Consequently precise formulation methods are required, necessitating a deeper understanding of the physicochemical properties of stabilizing agents, particularly sugars, which are commonly employed in protein formulations [4].
Addressing this challenge requires a thorough examination of the physicochemical properties of sugars, which play a crucial role in maintaining protein stability. Primarily, non-reducing sugars with high glass transition temperatures (Tg) are essential to keep the protein formulation in an amorphous state, thereby preventing denaturation or aggregation [5]. Next, maintaining low water activity is necessary to further prevent protein denaturation or aggregation. In addition, ensuring high solubility is essential to ensure uniform distribution across the formulation [6]. Finally, sugars with low reactivity are important to minimize the chemical degradation of the protein, while biocompatibility and non-toxicity are crucial to avoid any adverse effects on the biological activity of the protein [7]. Extensive development efforts have led to the registration by the United States Food and Drug Administration (US FDA) of thermally stable protein formulations. Although most therapeutic protein products are available in liquid formulation [8], transitioning proteins to a dry state is a promising approach to render them stable [7].
Storing proteins in a dry state may significantly reduce breakdown rates due to restricted molecular mobility, thereby enhancing their stability and shelf life [4]. However, proteins are subjected to multiple stresses during drying and subsequent storage, which may compromise the integrity of the protein. For example, in spray drying process, protein may encounter stress due to high inlet or outlet temperatures [9]. Furthermore, in freeze-drying processes, lyophilization involves two stress factors that cause protein denaturation:freezing and drying. Therefore, certain conditions are necessary to ensure that the stresses are not harmful to the protein [10]. One strategy to protect proteins against deterioration involves incorporating stabilizing agents such as sugars and surfactants into dry formulations [10]. Sucrose and trehalose are the most commonly used sugars for stabilizing solid protein formulations. Dry sugar-stabilized therapeutic protein and other stabilizer-therapeutic protein products that have been approved and registered by the US FDA are presented in Table 1 [11, 12].
Table 1: U.S. FDA-approved dried therapeutic protein in 2022-2023
| Protein | Trade Name | Type of Protein | Dosage Forms | Excipient used for stabilization | Route of Administration |
|---|---|---|---|---|---|
| Elotuzumab | Empliciti | Monoclonal antibody | For injection (lyophilized powder) | Sucrose,trehalose | Intravenous |
| Mepolizumab | Nucala | Monoclonal antibody | For injection (lyophilized powder) | Sucrose | Subcutaneous |
| Parathyroid hormone | Natpara | Hormone | For injection (lyophilized powder) | Mannitol, glycine, histidine | Subcutaneous |
| Secukinumab | Cosentyx | Monoclonal antibody | For injection (lyophilized powder) | Sucrose (prefilled syringe), polysorbate 80, L-histidine | Subcutaneous |
| Infliximab-dyyb | Inflectra | Monoclonal antibody | For injection (lyophilized powder) | Sucrose, polysorbate 80, L-histidine, sodium chloride | Intravenous infusion |
| Canakinumab | Ilaris | Monoclonal antibody | For injection (lyophilized powder) | Sodium chloride, polysorbate 80, and L-histidine | Subcutaneous |
| Certolizumab pegol | Cimzia | Monoclonal antibody | For injection (lyophilized powder) | arginine, sucrose, polysorbate 80 | Subcutaneous |
| Efalizumab | Raptiva | Monoclonal antibody | For injection (lyophilized powder) | sodium chloride, glycine, histidine | Subcutaneous |
| Omalizumab | Xolair | Monoclonal antibody | For injection (lyophilized powder) | L-histidine, arginine, polysorbate 20, sodium chloride | Subcutaneous |
| Velmanase alfa-tycv | Lamzede | Enzyme | For intravenous infusion (lyophilized powder) | Disodium phosphate dihydrate, sodium dihydrogen phosphate dihydrate, mannitol (E 421), glycine | Intravenous infusion |
| Cipaglucosidase alfa-atga | Pombiliti | Enzyme | For intravenous infusion (lyophilized powder) | Sodium citrate dihydrate, citric acid monohydrate, mannitol (E 421) | Intravenous infusion |
For several reasons, incorporation of a protein with a single sugar may not be sufficient for producing various medicinal products that can be administered through different delivery systems and routes. Initially, protein stability can be affected by factors such as pH, temperature, and light exposure during storage and administration, which can cause degradation or aggregation. Next, proteins may interact with other components in the formulation, leading to stability issues. Furthermore, the formulation may impact protein dissolution ability, resulting in difficulty in administering the product. To overcome these challenges, extra excipients, such as stabilizers, protectants, and solubilizing agents may be added to the formulation to stability, and usability [13]. Excipients play a crucial role in improving safety, efficacy, stability, and patient acceptability, underscoring the importance of evaluating their impact on sugar-stabilized therapeutic proteins in powder formulation. This review seeks to comprehensively evaluate the influence of different sugar types on protein stability, and analyze several excipients capableof further enhancing the stability of sugar-stabilized therapeutic proteins.
Through this review, valuable insights into the development and optimization of protein formulations will be gained, ultimately improving the safety, efficacy, and usability of therapeutic proteins. A systematic electronic search was conducted across various databases, including PubMed, Springer Link, Science Direct, ACS Publications, and Google Scholar, to investigate the influence of excipient blending on sugar stabilized therapeutic proteins. The search utilized a range of keywords such as Therapeutic proteins, Excipient blending for therapeutic protein, Sugar for protein stabilization, Amino acid for protein stabilization, Polyol for protein stabilization, and Sugar-stabilized therapeutic proteins. Studies meeting the inclusion criteria were included in the analysis, which required (1) publication between 1999 and 2024, with English text, (2) availability as complete articles, and (3) provision of relevant data pertaining to therapeutic proteins and excipients for protein stabilization. However, studies published solely as proceedings were excluded from consideration.
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Cynthia Marisca Muntu, Christina Avanti, Hayun Hayun, Silvia Surini, Impact of Excipients Blending on Sugar-Stabilized Therapeutic Proteins, Journal of Medicinal and Chemical Sciences, June 2024, DOI:10.26655/JMCHEMSCI.2024.6.2
Read also our introduction article on Mannitol here:
