Surfactants in Biopharmaceutical Development

Surfactants in Biopharmaceutical Development addresses the progress, challenges and opportunities for surfactant research specific to pharmaceutical development, providing a broad range of important surfactant-related topics as they relate directly to the biopharmaceutical process. Chapters address fundamental topics, like mechanisms of protein stabilization by surfactants, the latest, state-of-the-art technology and methods to illustrate the practical application to biopharmaceutical development, forward-looking chapters on control strategies and novel surfactants, with a special focus on current regulatory aspects of paramount importance for biopharmaceutical companies and regulators.

It has been widely recognized that surfactants provide protection to therapeutic proteins against interfacial stresses. Despite the fact that the very mechanism of protein stabilization by surfactants has not been completely understood, surfactants are universally regarded as critical functional excipients by the industry and by regulators.

• Describes the current state of research on surfactants in the context of biopharmaceutical development, drawing upon contributions from international experts across industry, academia, and regulators
• Addresses the opportunities and challenges associated with surfactants in biologic drug development
• Provides a defining resource for practitioners in the biopharmaceutical industry, regulators and academics by summarizing the latest knowledge of surfactants in biopharmaceutical development in one comprehensive volume

• Language: English
• Publisher: Academic Press
• Release date: Aug 12, 2023
• ISBN: 9780128125694

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Surfactants in Biopharmaceutical Development

Editor

Atanas V. Koulov

Chief Scientific Officer, Clear Solutions Laboratories AG, Basel, Switzerland

Editor

Satish K. Singh

Vice President, Drug Product Development, Moderna Inc., Cambridge, MA, United States

Table of Contents

Cover image

Title page

Copyright

Dedication

Contributors

Chapter 1. Introduction: Surfactants continue to surprise

What are the different surfactants used in biopharmaceutics today and what is their in vivo fate? What is their toxicity and how well are they tolerated by the human organism?

How are the surfactants polysorbate and poloxamer made and how is that significant for their use?

How do surfactants work to stabilize biopharmaceuticals?

How do we use surfactants in biopharmaceutical manufacturing and what are the important considerations in their use during the processing of biopharmaceuticals?

How do we understand, measure, and control the quality of surfactants?

What is the stability and degradation behavior of surfactants?

What are the regulatory (and scientific) considerations for developing protein formulations containing surfactants?

What should we expect in the future?

Chapter 2. Polysorbate in biopharmaceuticals—an overview including in vivo fate and safety perspective

Introduction

Overview of the use of surfactants for biotherapeutics

Metabolism of surfactants

PK PD modulating effect of polysorbate 80

Safety of surfactants

Safety of surfactants after degradation

Conclusions

Chapter 3. Surfactants (polysorbate and poloxamer): synthesis, characterization, and degradation

Introduction

Polysorbates overview

Techniques used to characterize polysorbates

The impact of polysorbate impurities on biotherapeutic formulation stability

Alternatives to polysorbates

Poloxamers overview

Synthesis and process controls

Purification techniques

Characterization techniques

Stability of poloxamers

Other uses of surfactants in biopharmaceuticals

Chapter 4. Mechanisms of stabilization of proteins by surfactants

Introduction

Methods for characterizing surfactant–protein behavior

Modes and factors impacting protein stabilization by surfactants

Summary

Chapter 5. Behavior of surfactants during processing

Introduction

Challenges with polysorbates

Behavior of surfactants during processing

Conclusion

Chapter 6. Analytical methods for surfactant characterization in biologic drug products

Introduction

Routine quantitative analytical methods

Characterization analytical methods

Summary and conclusion

Chapter 7. Oxidative degradation of surfactants: mechanisms

Introduction

The location of oxidation-sensitive sites on polysorbate and poloxamer

Surfactant oxidation: the initiation process

Chain oxidation

Oxidation products and mechanisms

Oxidative generation of free fatty acids

Cis/trans isomerization of unsaturated fatty acids

Perspective

Chapter 8. Hydrolytic degradation of surfactants and impact on protein stabilization

Chemical hydrolysis

Enzymatic hydrolysis

Impact of hydrolytic degradation of polysorbate on protein stabilization

Conclusions

Chapter 9. Scientific and regulatory considerations for developing protein formulations containing surfactants

Surfactants and their use in biotechnology drug products

Scientific rationales for using and controlling surfactants in protein formulations

Some strategies for controlling surfactants in protein formulations

Studies that generally support the use of surfactants in therapeutic protein formulations

Chapter 10. Control strategies to address surfactant degradation: a quality perspective

Introduction

Raw material

Monitoring surfactant levels during development

Control elements on drug substance and drug product level

Chapter 11. A look to the future—closing remarks

Surfactant functionality

Safety and adverse events

New surfactants for biopharmaceuticals

Surfactant degradation and control strategies

Author index

Subject index

Copyright

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Notices

Knowledge and best practice in this field are constantly changing. As new research and experience broaden our understanding, changes in research methods, professional practices, or medical treatment may become necessary.

Practitioners and researchers must always rely on their own experience and knowledge in evaluating and using any information, methods, compounds, or experiments described herein. In using such information or methods they should be mindful of their own safety and the safety of others, including parties for whom they have a professional responsibility.

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Dedication

We dedicate this book to the most important women in our lives: Sangeeta, Nikita, Roohi, Elena, Nanna, and Matilda.

Contributors

Christian H. Bell,     Analytical Development and Quality Control, F. Hoffmann-La Roche AG, Basel, Switzerland

Vincent Corvari,     Bioproduct Research & Development, Eli Lilly & Company, Indianapolis, IN, United States

Diana C. Gomes,     Technical Research and Development, Novartis Pharma AG, Basel, Switzerland

Christoph Grapentin,     Lonza AG/Ltd, Drug Product Services, Basel, Switzerland

Marc Heitz,     Pharma Technical Development Europe, F. Hoffmann-La Roche Ltd., Basel, Switzerland

Lihua Huang,     Bioproduct Research & Development, Eli Lilly & Company, Indianapolis, IN, United States

Michael Jahn,     Lonza DPS – Forensic Chemistry, Basel, Switzerland

Tarik A. Khan,     Pharma Technical Development Europe, F. Hoffmann-La Roche Ltd., Basel, Switzerland

Atanas V. Koulov,     Clear Solutions Laboratories AG, Basel, Switzerland

Hanns-Christian Mahler,     Ten23 Health AG, Basel, Switzerland

Jeannette E. Marine,     Croda Pharma R&T, Croda Inc., Princeton, NJ, United States

Sreejit R. Menon,     Croda Pharma R&T, Croda Inc., Princeton, NJ, United States

Claudia Mueller,     Lonza AG/Ltd, Drug Product Services, Basel, Switzerland

Janina Pfaff

Analytical Development and Quality Control, F. Hoffmann-La Roche AG, Basel, Switzerland

Pharmaceutical Development, F. Hoffmann-La Roche AG, Basel, Switzerland

V. Ashutosh Rao,     Laboratory of Applied Biochemistry, Division of Biotechnology Review and Research III, Office of Biotechnology Products, Office of Pharmaceutical Quality, Center for Drug Evaluation and Research, U.S. Food and Drug Administration, Silver Spring, MD, United States

Stephen J. Rumbelow,     Croda Pharma R&T, Croda Inc., Princeton, NJ, United States

Christian Schöneich,     Department of Pharmaceutical Chemistry, The University of Kansas, Simons Laboratories, Lawrence, KS, United States

Ariane Schröter,     Ten23 Health AG, Basel, Switzerland

Satish K. Singh,     Drug Product Development, Moderna Inc., Cambridge, MA, United States

Chapter 1: Introduction

Surfactants continue to surprise

Atanas V. Koulov     Clear Solutions Laboratories AG, Basel, Switzerland

Abstract

The introductory chapter to this book reviews the main questions put forward by the authors of the individual chapters and guides the readers' attention to the most surprising elements relevant to these questions, which hopefully move forward our current understanding of the use of surfactants in biopharmaceutical development.

Keywords

Particles; Poloxamer; Polysorbate; Surfacactant control strategy; Surfactant analytics; Surfactant degradation; Surfactant synthesis; Surfactants

… the table of elements does not contain one of the most powerful elements that make up our world, and that is the element of surprise.

Lemony Snicket, The Ersatz Elevator.

The book you are holding is devoted to the use of surfactants in the development of biopharmaceutical products. Surfactants represent a very important—if not critical—class of excipients, nearly universally present in biopharmaceutical drug product (DP) formulations today. As functional excipients, surfactants provide the biologic active pharmaceutical ingredient (API) with protection from interfacial stresses, enabling appropriate processing, storage, and handling of the drug product and ensuring adequate stability, according to the corresponding quality target product profile (QTPP). Considering the fact that surfactants have been used as a stabilizing excipient for biopharmaceuticals for decades, it is fascinating to see the number of challenges that this class of compounds has presented and the surprises it continues to serve to biopharmaceutical scientists even in the present day. In reviewing various aspects of the application of surfactants in the development of biopharmaceuticals, we have focused on those surprises, nearly all comprising hot topics and impactful current trends in the development of modern biopharmaceuticals.

Here is a list of some of the questions that the authors of the book have explored in 12 chapters (in an impressive combination of original research and review of the current literature) and which have revealed surprising answers in practically every aspect of the use of surfactants in biopharmaceutical development.

What are the different surfactants used in biopharmaceutics today and what is their in vivo fate? What is their toxicity and how well are they tolerated by the human organism?

In Chapter 2 of this book, Satish Singh reviews the various surfactants in use today and their (presumed) mechanisms of action. Notably, the chapter addresses the important and frequently overlooked set of questions related to the metabolism, the in vivo fate, and the toxicity of surfactants. Even more importantly, the chapter looks at the highly relevant and little understood question of the safety aspects of the surfactant degradation products, providing the reader with a valuable and detailed review of the literature on this topic, including a collection of reports on adverse events. Last but not least, this chapter addresses one of the most hotly debated technical topics in the field of drug product development today—what is the safety profile of surfactants after degradation?

How are the surfactants polysorbate and poloxamer made and how is that significant for their use?

Perhaps as a consequence of the increasing need for specialization, practitioners in the field of pharmaceutical development often overlook the basics of surfactant chemistry and its implications. A prime example is the impact of the chemical synthesis and manufacturing processes used to produce surfactants for use as excipients in biopharmaceutical products. In Chapter 3, Stephen Rumbelow and colleagues focus on introducing the reader to the synthetic routes for manufacturing polysorbates and poloxamer.

We consider this chapter foundational to the book, not only because of its educational value, but also because one of the main take-homes of the book—the fact that many of the surprises presented to biopharmaceutical development scientists today are rooted in the exceptional structural heterogeneity of polysorbates—is a direct consequence of the synthetic procedures used for polysorbate manufacturing. For example, current manufacturing processes use complex natural raw materials (corn, tapioca, coconut oil, palm oil, tall oil) and sometimes harsh process conditions, resulting in an extraordinary heterogeneity of these groups of compounds. Whereas this heterogeneity may seem obvious to synthetic chemists and excipient manufacturers, we believe that this important attribute of polysorbates is not well recognized by practitioners in the biopharmaceutical industry. Even more importantly, the significance of this heterogeneity is not well understood and is only recently begun to be explored.

Of course, it is important to note, that the synthetic procedures used to manufacture polysorbates are designed to consistently yield the product quality described in the pharmaceutical compendia (USP, EP, etc.). However, these pharmacopeia tolerate significant variability in the composition of surfactants. In their chapter, Rumbelow and colleagues also introduce the differences between the different qualities of polysorbates, particularly when it comes to residual impurities. The authors suggest choosing wisely, when it comes to the quality of surfactants used in biopharmaceuticals.

How do surfactants work to stabilize biopharmaceuticals?

In Chapter 4, Tarik Khan and colleagues review current data on the critical question of the role that surfactants play in stabilizing the active pharmaceutical ingredient (API) in biopharmaceuticals—that is, how do they stabilize proteins? It is fascinating to recognize the fact that despite significant efforts over the last few decades to understand the mechanisms of protein stabilization by surfactants, there is still a lot that remains to be clarified—another major surprise for many. With their chapter, Khan and coauthors provide us with a very systematic review of the physicochemical properties of surfactants and proteins, the approaches and methods to study surfactant behavior, and the interaction between proteins and surfactants. More importantly, the authors also review the current state of the art and the experimental studies addressing the mechanistic details of the stabilization of proteins by surfactants. Khan and colleagues conclude that although in some cases surfactants have been demonstrated to directly interact with proteins, in the majority of cases nonionic surfactants stabilize protein pharmaceuticals primarily by competitive adsorption at interfaces.

How do we use surfactants in biopharmaceutical manufacturing and what are the important considerations in their use during the processing of biopharmaceuticals?

In Chapter 5, Hanns-Christian Mahler and colleagues review important attributes of surfactants—the surfactant quality and impurities, their degradation, and interactions with processing materials—as well as related considerations for the use of surfactants in the development of biologics. Most significantly, the authors guide our attention to the potential pitfalls that may be encountered in drug product processing unless these considerations are appropriately addressed. The chapter provides illustrative case studies highlighting the importance of attending to the problems of surfactant interaction with filters and membranes used in the processing of biopharmaceuticals (such as UF/DF membranes, for example). Again, we believe that some readers will find that these case studies hold a lot of surprises and are very informative, particularly with regard to the extent to which such interactions may influence the final product quality. Most importantly, Mahler and colleagues guide us to some of the tools at our disposal for addressing and avoiding these potential pitfalls—analytical monitoring, risk assessments, etc.

How do we understand, measure, and control the quality of surfactants?

In Chapter 6 of this book, Michael Jahn introduces us to the diverse analytical toolbox we have at our disposal to begin understanding the composition and quality of surfactants in biopharmaceuticals. The chapter presents in a very systematic fashion, the tools for routine monitoring of surfactant quality and quantity, as well as characterization methods used mostly in investigative mode. In our opinion, the two most significant contributions of this chapter (which we believe will be very useful to a broad audience) are: (1) the comprehensive overview it provides (reviewing methods for total surfactant quantitation to characterization of free fatty acid particles as byproducts of surfactant degradation), and (2) the comparison it provides between the various complementary approaches, including critical review of advantages and deficiencies of the different analytical methods. One of the most arresting surprises that some readers will find in this volume is the fact that polysorbates are a strikingly complex mixture of molecular species. The extent of this heterogeneity is well illustrated in this chapter in the multitude of PS-80 related species by LC-MS (see Fig. 1.1 in Chapter 6, also shown on the cover of this book).

The analytical toolbox for surfactant monitoring and characterization is also a subject of review in Chapter 11, where Christian Bell and Janina Pfaff also discuss the various considerations one needs to be aware of when selecting an analytical method for polysorbate quantification or characterization. More importantly, in this chapter, the authors guide us to the critical question of control strategy for surfactants in biopharmaceutical development. The chapter provides a careful review of the various pieces that build the puzzle of holistic and comprehensive surfactant control strategy—from raw material through drug substance and drug product control elements to product specifications.

What is the stability and degradation behavior of surfactants?

A major part of this book by far is devoted to the stability of polysorbates. We intentionally sought to examine this issue in such detail. From being a stabilizer, polysorbate instability can have an adverse impact on both product quality and the ability to be in compliance with current regulatory requirements. Multiple publications—journal articles, conference presentations, and health authority BLA CMC approvability recommendations—have highlighted potential issues that can arise when polysorbates turn bad. Such reports have featured prominently the formation of subvisible and visible particles, either composed of polysorbate degradation products (fatty acid and related insoluble compounds), or proteinaceous. As polysorbate-related particle formation is notoriously difficult to control, in addition to the compromised QTTP, it usually raises important safety and quality concerns.

The various aspects of the stability of polysorbates have been addressed in different chapters of this book. In Chapter 7, Christian Schoeneich (in much detail) dissects the mechanisms of oxidative degradation of surfactants. The chapter reviews the chemical moieties sensitive to oxidation, as well as the different stages of oxidative reactions. Most importantly, there is an extensive overview of the potential products of these oxidative reactions and the implications for the quality and stability of biopharmaceuticals, which we believe the reader will find very illuminating.

In Chapter 8, Vincent Corvari and Lihua Huang review the current state of our knowledge of hydrolytic surfactant degradation—chemical and enzymatic. The authors discuss the damage that copurifying enzymes can cause to polysorbates and their functionality, the product quality risk including the specific consequences of the liberation of free fatty acids into the drug product solution, as well as diagnosis of these enzymatic reactions. The chapter provides an exhaustive review of the latest literature reports on this critical topic, with much relevance to high-concentration monoclonal antibody products being brought to market.

What are the regulatory (and scientific) considerations for developing protein formulations containing surfactants?

In the penultimate chapter of the book, Ashutosh Rao from the US FDA presents his perspective on the regulatory aspects of the use of surfactants. The chapter provides a broad overview of topics ranging from the use of surfactants in the currently licensed biological products, through scientific aspects of the surfactant use (such as their mechanisms of action as a functional excipient), quality considerations (such as the control of raw materials and the excipient stability in the drug product) to the typical regulatory considerations for licensure.

What should we expect in the future?

In the final chapter of this book, we have attempted to provide a glimpse into the future and perhaps somewhat provocatively, ask questions that may be most critical to tackle next. We have attempted to provide an outlook on the field, focusing on potential solutions to the challenges highlighted in this book. Interestingly, several of these potential solutions will require entirely novel approaches in nearly all aspects reviewed—analytical, synthetic, and regulatory.

But of course, outside of all of the questions that have been asked in this volume, the most intriguing question remains:

What other surprises will surfactants bring in the future?

Chapter 2: Polysorbate in biopharmaceuticals—an overview including in vivo fate and safety perspective

Satish K. Singh     Drug Product Development, Moderna Inc., Cambridge, MA, United States

Abstract

Surfactants are commonly added to biotherapeutic product formulations (including vaccines) to stabilize the active moiety against interfacial stresses commonly encountered during manufacturing, shipping, and storage. Polysorbates are the primary surfactant used in biopharmaceuticals, usually at small concentrations compared to the larger amounts used with small molecule (parenteral) products, where the objective is the solubilization of the active agent. Excipients are intended to be inactive, but they may not be pharmacologically inert as is the case with polysorbates. There are an increasing number of reports about injection site reactions, sensitization, anaphylactoid reactions, etc. from healthcare practitioners administering biopharmaceutical products, with the polysorbate component being generally implicated. This chapter provides a perspective on the in vivo impact of polysorbates and the safety aspects of this excipient including learnings from use in small molecule parenteral products and in biotherapeutics. While biotherapeutics avoid the significant adverse events associated with their use in small molecules, the importance of quality of this excipient both as raw material and in the biopharmaceutical is reemphasized, to minimize adverse events and improve tolerability.

Keywords

Adverse events; Immunogenicity; Polysorbates; Safety; Sensitization; Tolerability

Introduction

Surfactants are commonly added to biotherapeutic product formulations (including vaccines) to stabilize the active moiety against interfacial stresses commonly encountered during manufacturing, shipping, and storage. This functionality of the surfactants is extensively examined in other chapters of this book. Surfactants in biotherapeutics are usually added in small amounts ranging between 0.001% and 0.1% w/v and the commonly used surfactants tend to be nonionic due to their favorable tolerability and toxicity profile compared to ionic surfactants in general.

Excipients in biopharmaceutical formulations are intended to stabilize the active protein from manufacturing until use by the patient and also ensure that the product is suitable for intended use, such as providing adequate pH or tonicity [1]. Excipients are designated by the FDA to be inactive, in that they are not considered to be the primary biologically active ingredient [2]. However, this does not necessarily imply that these excipients are always biologically and pharmacologically inert on administration. In regulatory filings, the use and need for each excipient must be appropriately justified.

This chapter provides a short overview of the use of surfactants and then an assessment of in vivo fate and safety aspects of polysorbates. Although this chapter and the book focus on biologics, a significant number of nonbiologic parenteral products contain surfactants, primarily with a focus on solubilization. A wider variety of surfactants are used in these cases and are considered vehicles, with concentrations of the surface active agents used at levels significantly higher than those used in biologics. Thus, learnings about surfactants (polysorbates, PS) from the nonbiologic parenteral products are used as a springboard in this chapter.

Overview of the use of surfactants for biotherapeutics

The most commonly used surfactants for biotherapeutics are poly-oxy-ethylene (POE) based polysorbates 20 and 80 as well as to a lesser extent the Pluronic surfactant Poloxamer 188. The general preference of which one to use is often dictated by past experience since in general, formulation development studies are unable to differentiate significantly between for example PS20 or PS80 (based on author's experience). The preference for one over the other based on stability differences does not confer any significant advantages.

The mechanism of stabilization by surfactants falls into two broad categories: stabilization by prevention of access to interfaces (interfacial competition) or complexation at hydrophobic patches on the protein and enhancing colloidal stability [3,4]. Biophysical studies have shown little to some direct interaction between these (polysorbate) surfactants and proteins, depending significantly on the nature of the protein (see also review in Ref. [5]). The currently prominent monoclonal antibodies show weak interactions, potentially due to their generally more hydrophilic nature, while reasonable levels of interaction have been shown for hGH, BSA, HSA, hGH-HSA fusion, LDH, etc. (summarized by Refs. [3,5]). Poloxamer 188 (P188) has a complex solution behavior, forming multiple different states of association depending on temperature and concentration [6,7]. There are fewer studies with P188 but the studies again show that protection may be interfacial competition mediated (Fc-Fusion protein [8]); or there may be some direct but weak interaction (rFVIII [9,10]). As an associated mechanism, the binding of surfactant is proposed to stabilize partially folded structures and enable refolding, similar to molecular chaperones [11].

Various authors have tried to relate the protective effect of surfactants to their CMC but in the author's experience,