Biosimilars: Proteins, PDF

PROPRIETARY. DO NOT SHARE. Transcript: Biosimilars Section 1: Proteins Welcome Welcome to Biotech Primer’s online course titled Biosimilars. In this course, we will discuss the science, manufacturing, safety, and regulatory considerations associated with Biosimilars. The course is divided into six...

PROPRIETARY. DO NOT SHARE. Transcript: Biosimilars Section 1: Proteins Welcome Welcome to Biotech Primer’s online course titled Biosimilars. In this course, we will discuss the science, manufacturing, safety, and regulatory considerations associated with Biosimilars. The course is divided into six sections --- Proteins, Biologics, Biosimilars, Manufacturing, Safety, and Regulation, and we will end the course by looking at various Biosimilar Case Studies. Since biosimilars are protein therapeutics, let’s begin our course with a review of proteins. Section 1: Proteins Objective In section one we will take a closer look at Proteins. Why proteins? Biosimilars are proteins. Biosimilars are generic versions of brand-name biologics. Understanding the complexity of protein structure and function is critical to understanding how both brand name biologics and biosimilars function. By the end of section one, you should be able to: • Describe three protein functions. • Arrange the steps of protein synthesis in the correct order. • Explain why proteins fold. • Deduce what happens when a protein folds properly and improperly. • List the purposes for post-translational modifications. Cells Make Proteins All living things are made up of cells. Within the cells are organelles. Organelles called ribosomes make proteins. Proteins are made by cells. What Do Proteins Do? Proteins are large molecules responsible for cellular structure and function. Proteins are the workhorse of the cell. 1 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. Proteins: Critical to Cellular Function: Enzymes Almost all vital physiological functions are carried out by proteins. One example of a type of cellular protein is enzymes. Enzymes are proteins that enable important chemical reactions to occur in the body. This figure depicts an enzyme catalyzing the conversion of a complex sugar into a simple sugar. Proteins: Critical to Cellular Function: Antibodies Antibodies are proteins produced naturally by the immune system in response to foreign invaders - viruses, bacteria, etc. Antibodies recognize their target based on shape - the antibody’s shape is complementary to the target. Antibodies defend the body against foreign invaders and are highly specific for their target. Proteins: Critical to Cellular Function: Receptors Receptors are proteins on the surface of cells that receive messages (also called signals) from the rest of the body. These messages are often carried in the form of other proteins, such as the hormone insulin. Receptors enable cellular communication. A specific messenger recognizes and binds to a particular receptor protein based on their complementary shapes. Once the messenger protein has attached, it causes the receptor protein to undergo a subtle conformational change that transmits the message to the inside of the cell, ultimately to the nucleus, where the expression of a specific gene is activated. How Do Cells Make Proteins? The nucleus, found in the center of the cell, houses the DNA. Sections of DNA, called genes, store the information the cell uses to make proteins. Each gene is a recipe for one or more proteins. Protein Synthesis Protein synthesis occurs inside cells. It is a two-step process. DNA is first transcribed or copied into a chemically related molecule, RNA, which is then translated into the actual protein. 2 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. Amino Acids Make Up Proteins There are 20 amino acids in the human body. Each amino acid has unique chemical and physical properties. To learn more about the specifics of the amino acids click on the button to learn more, otherwise, click next to continue. Amino Acids Make Up Proteins Let’s look at the close-up of an amino acid. Each amino acid molecule has a common structure an amino group, an acidic carboxyl group, and an “R” group. The “R” group is what differs between each amino acid and makes each one unique. For example, some such as Arginine are positively charged; Others, such as Aspartic Acid, have a negative charge. Amino Acids Interact The order, or sequence, of the amino acids in the chain, determines the final structure of the protein molecule. The string of beads takes on a specific shape because some R groups attract one another while others tend to repel one another. This attraction/repulsion is shown here-the positive charge of Arginine is attracted to the negative charge of Aspartic Acid, resulting in a fold in the amino acid chain. Amino Acids Fold into Complex, Functional Proteins To recap, each amino acid has unique chemical and physical properties. How these amino acids interact with each other up and down the length of the amino acid chain, as well as how they interact with their environment, determines how the protein folds into its final complex shape and structure, which in turn determines its function. This slide shows two alternative ways of representing complex protein structures: the space-filling model and the ribbon model. Protein Shape Determines Function The folding pattern dictates the protein’s function. Therefore, proteins that are folded correctly function as intended. Proteins that do not fold properly, do not function properly. In the case of the human body, misfolded proteins may cause disease. In the case of therapeutic proteins misfolded proteins may cause unintended or adverse reactions. 3 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. Post-Translational Modification Post-translational modification simply means any modification or change that is made to a protein after it is produced by the cell. Chemical modifications may range from the addition of a single chemical group to multiple small modifications, to the addition of sugar chains, or even the chemical attachment of small proteins - all with the potential to impact key activities of the protein. Modifications Are Critical For example, cellular enzymes might add a specific chemical group to the protein or might cut the protein in a specific place. Modifications are critical to protein function and may serve to activate or deactivate a protein, change its function by changing its cellular location or interaction with other cellular proteins, or alter the stability of the protein. Phosphorylation One common type of post-translational modification is phosphorylation or the addition of a negatively charged phosphate group to a protein by a kinase enzyme. Glycosylation Another very common type of post-translational modification is glycosylation, or the addition of a sugar group to the protein by glycosylase enzymes. Glycosylation: A Closer Look Glycosylation patterns - how a protein is glycosylated, or not - are associated with key aspects of protein function, including conformation, stability, receptor binding, protein function, and immune response to protein. This slide illustrates the same protein with two different glycosylation patterns and therefore two different functions. Different Modifications Using different production cell lines to produce the same protein may result in slightly altered glycosylation patterns on that protein. 4 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. Section 1: Proteins Summary Understanding the complexity of protein function and structure is critical to understanding biosimilars. Let’s summarize the main points made in section one titled Proteins. ● Three important proteins and their functions are: 1. Enzymes. Enzymes enable chemical reactions to occur in the body. 2. Receptors. Receptors communicate with other cells by receiving messages. 3. Antibodies. Antibodies fight disease. ● The two steps of protein synthesis are transcription and translation. ● Proteins fold because each amino acid has unique chemical and physical properties. These properties interact with each other up and down the length of the protein. This interaction determines how the protein folds into its final complex shape and structure. A protein’s structure determines its function. ● When a protein folds properly it functions properly. When a protein folds improperly it does not function as it should. ● 5 Post-translational modifications help the protein function properly. Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. Section 2: Biologics Welcome Now that you have a solid understanding of proteins let’s learn more about protein therapeutics in this section titled Biologics. Section 2: Biologics Objectives After the completion of this section, you should be able to: ● Define biologics, ● List the functions of therapeutic proteins, and ● Compare and contrast small molecule drugs with biologics. Biologics Definition A biologic is a substance derived from living material, such as cells or tissues, used to treat, prevent or cure disease. The WHO, EMA, FDA, and other regulatory bodies recognize that there is a wide range of biologics, many of which require different regulatory considerations. One of those classes is protein therapeutics, which will be the focus of this course. Biologic Uses in Healthcare Because proteins play such a critical role in cellular function and human health, it’s not surprising that they have been adapted for use as therapeutics. In the next few slides, we will look at a few common clinical applications of therapeutic proteins. The ways in which biologic drugs treat disease include replacing endogenous proteins, modulating the immune system, and modulating protein activity. Biologics Function: Replacing Missing Proteins Some diseases are caused when the patient’s body doesn’t produce enough of the required protein that functions properly. For example, Type 1 diabetes is the result of the pancreatic beta cells not producing the protein insulin. The job of insulin is to reduce blood sugar after a meal. Why is reducing blood sugar so important? High blood sugar results in damaged organs. To replace the insulin not produced in the body, diabetics inject man-made insulin. Insulin is a biologic. 6 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. Biologics Function: Modulating Protein Activity Modulating activity can mean either boosting or suppressing activity. In the context of monoclonal antibodies, “modulating” usually means inhibiting or blocking the activity of a disease-causing protein, such as an overactive growth factor. Let’s look at the drug, Herceptin as a classic example of a monoclonal antibody modulating protein activity. In a percentage of breast cancer patients, the breast cancer cells overexpress a gene known as HER2. The overexpression of HER2 leads to the creation of a large number of HER2 receptors on the surface of the breast cancer cell. Because of this large number of receptors, the cell receives too many signals. Signals known as growth factors bind to this abundance of HER2 receptors, causing uncontrolled cell growth and cell division leading to cancer. Herceptin is a monoclonal antibody that works by binding to the HER2 receptors on the outside of these breast cells. Once bound to the HER2 receptor, Herceptin—the monoclonal antibody— blocks the receptor so that the growth factor signal cannot bind to the HER2 receptor. Herceptin also flags the breast cancer cell as foreign. When a cancer cell has most of its HER2 receptors bound by monoclonal antibodies, the immune system is alerted and immune cells like the macrophage are deployed to the site to destroy the flagged cancer cells. Biologics Function: Modulating the Immune System Monoclonal antibodies have been used to treat autoimmune disorders such as rheumatoid arthritis and Crohn’s disease either by helping to destroy cells involved in the immune response, or by blocking immune cells from targeting the patient’s tissues. Interferon-beta is an antiinflammatory signaling protein - it helps to “calm down” an overactive immune response. Small Molecule Drugs vs. Biologic It is important to stress the key differences between small molecule drugs and biologics, in terms of complexity, characterization, and manufacturing. Let’s start with small-molecule drugs. Small molecule drugs are those that come in pill or capsule form. They have a simple chemical structure. Because of this simple structure, they are easy to characterize, meaning it is simple to determine their structure. They are easy to manufacture 7 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. because they are chemically synthesized in the lab. They are stable, meaning they don’t break apart. Biologics are protein therapeutics taken by injection; if taken orally, the intestine and stomach would digest the protein and it would not be available to fight disease. Biologics have complex structures, so complex it is difficult to determine their structure. Biologics are difficult and expensive to manufacture because they are made in living cells. Biologics are unstable and must be kept refrigerated so they maintain their structure and therefore their function. Manufacturing Implications What are the manufacturing implications? Two small molecule drugs made in different labs can be chemically analyzed and shown to be identical due to the simplicity of the chemical structure. Two biologics made in different labs cannot be chemically analyzed and shown to be identical due to the complexity of the protein structure. Distinctly Different All biologic drugs are larger and more complex than small molecule drugs. Here, we are comparing an aspirin, a small molecule drug, with two biologic Interferons and a monoclonal antibody. Interferon alpha, used to rev up the immune system to help fight cancer, is about 500 times larger than the typical small molecule drug aspirin, while the typical monoclonal antibody is about 750 times larger than aspirin. But even within the category of biologics, there is a range of size and complexity. Section 2: Biologics Summary In summary, section 2 explained: ● A biologic is a substance derived from living material, such as cells or tissues, used to treat, prevent or cure disease. An example of a biologic is a monoclonal antibody. ● Common clinical applications of therapeutic proteins include replacing missing or damaged proteins, modulating the immune system, and increasing or decreasing protein activity. 8 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. ● As compared to small molecule drugs, biologics are made in cells, have a more complex structure, are more difficult and more expensive to manufacture, and are more unstable so most must be refrigerated. 9 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. Section 3: Biosimilars Welcome Now that we have completed our review of proteins and biologics, we can take a closer look at biosimilars. Other terms you may have heard used to describe generic versions of biologics are subsequent-entry biologics or follow-on biologics. Section 3: Biosimilars Objectives After this section, you should be able to: ● Define biosimilars. ● Contrast small molecule drug generics with biosimilars. ● Compare the EMA and FDA definitions of biosimilars. Biosimilar Defined A biosimilar is a biologic drug that is produced using a different cell line, master cell bank, and or a different process than the one that originally produced the brand-name product. We will explain terms such as cell line and master cell bank in greater detail later in this course. Making a brand name small molecule drug into a generic is relatively straightforward. It is easy matter to prove the generic is identical to the small molecule innovator drug using analytical tests. However, it is difficult to prove that a biosimilar is identical to the brand name biologic, especially when the biologic is a large protein. Product is the Process The product is the process. We’ll begin by contrasting biosimilars with small molecule generics. Because the structure of a small molecule drug is relatively simple, scientists use standard analytic techniques to determine its exact structure - down to the level of where individual atoms are concerning each other. Therefore, it’s generally straightforward for generic manufacturers to demonstrate that their generic drug is structurally identical to the brand name small molecule drug. In contrast, biologics have a highly complex structure. We do not currently have the technology to precisely determine the exact molecular structure of a biologic. So, if a version of the biologic, the biosimilar, is produced using a different cell line or process, it is not possible to demonstrate 10 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. the brand name biologic and the biosimilar are structurally identical. This idea is sometimes summarized by saying “The product is the process” EMA Biosimilar Definition The European Medicines Agency or EMA, the regulatory agency for the European Union, uses the following criteria to determine if the therapeutic protein is a biosimilar medicine. Specifically, they state the biosimilar is expected to have certain similarities including a similarity to the active ingredient of the reference drug and a similarity to clinical results in patients. However, the EMA stipulates the name, appearance, and packaging of a biosimilar medicine must differ from reference biological medicinal products to distinguish between the two. FDA Biosimilar Definition The Food and Drug Administration or FDA, the regulatory agency for the United States of America, states that a biosimilar is a biological product highly similar to the reference product notwithstanding minor differences in clinically inactive components. And that it has the same route of administration, dosage form, and strength. Therefore, the FDA and the EMA both define biosimilarity with respect to a reference product - the innovator biologic being copied. Both agencies concluded if a biosimilar is shown to be highly similar to the reference product, it is likely to have the same clinical result. We will discuss how companies can demonstrate their product is highly similar to a reference product in more detail later in this course. Section 3: Biosimilars Summary Let’s review the main points detailed in section three, Biosimilars. ● A biosimilar is a biologic drug that is produced using a different cell line, master cell bank, and/or a different process than the one that originally produced the brand name product. ● As compared to small molecule drugs, biologics have a highly complex structure that cannot be determined exactly, and they are expensive and difficult to manufacture. ● The FDA and the EMA both define biosimilarity concerning a reference product— the brand name biologic being copied. Both agencies concluded if a biosimilar is shown to be highly similar to the reference product, it is likely to have the same clinical result. 11 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. Section 4: Manufacturing Welcome In this section, we will demonstrate how biologics, and therefore biosimilars, are manufactured. This section will explain how cell banks ensure access to the original cell line to enable identical protein processing for decades. We will also learn how biologics are made, and what impact that has on the production of biosimilars. As stated earlier- the product is the process. Section 4: Manufacturing Objectives After this section, you should be able to: ● Define biomanufacturing. ● Contrast the manufacturing process for small molecule drugs and biologics. ● List the steps used in biomanufacturing in their correct order. ● Distinguish between the master cell bank and the working cell bank. ● Explain how different master cell lines, with the same gene, can produce different proteins. ● Identify where variations can occur between two different production campaigns. Making a Small Molecule Drug vs. Making a Biologic The process of making a biologic drug is completely different from the process of making a small molecule drug. Small molecule drug manufacture starts with chemicals that are synthesized together in specialized glass or metal containers. In contrast, biologics manufacture starts by transferring a specific gene sequence (DNA) into a cell, plant, or animal. The initial product of chemical synthesis is a relatively pure chemical compound, whereas the initial product of biologic manufacturing is the broken-apart cells or the growth medium into which the protein product was secreted. The components of a small molecule synthesis are clearly defined, while the components of biologics manufacture are not. Biomanufacturing Overview In the next few slides, we will review key concepts related to biomanufacturing, and then discuss their potential impact on biosimilars. Biomanufacturing refers to the use of living cells to make biological products. The four essential steps are establishing cell banks, cell culture seeding and scale up; harvest and purification; and formulation, fill, and finish. 12 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. Therapeutic Proteins Are Made in Cells Therapeutic proteins are made in cells. To get a cell line to produce a particular protein, scientists use genetic engineering techniques to create a plasmid that contains the gene of interest. A circular piece of DNA that contains the “gene of interest”, which codes for the therapeutic protein is inserted into the plasmid. The plasmid is then transferred into the production cell line such as the Chinese hamster ovary or CHO cells. Establishing Production Cell Line Since there are variations in every cell, each must be screened to identify the best clone to move forward with into production. This involves multiplying each cell and measuring each prospective cell line’s production capacity - which means calculating how many grams of protein can be produced per liter of cells and the level of impurities produced. Once the best individual cell or cell clone is identified, it is moved forward into production. Cell Banks The term Cell Banking refers to the process of storing cells, of a specific genome, for future use in the production of medicinal products. Cell banks are critical to ensuring the continuity of product identity and quality. When cells are stored at extremely low temperatures - 210 degrees Celsius - all cell growth is stopped. Essentially capturing a moment in time. Decades after a production cell line is created, a company can access the original cell line by going back to the stored cells. Why is the ability to go back to the original cell line important? All cells grow and change with time so companies must have access to the original cell line. Two Types of Cell Banks There are two types of cell banks- master cell banks (MCBs) and working cell banks (WCBs). MCB Production A master cell bank is created by expanding the original production cell line for a few generations and creating a few hundred vials of cells, each vial containing about one million cells. Because of its critical importance, the master cell bank will usually be divided into two or three and stored in distinct geographic locations. That way, if storage conditions are not able to be maintained due to natural disasters or other unforeseen circumstances in one location, the company will not lose its entire master cell bank. Importantly, no two master cell banks are alike. 13 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. WCB Production A working cell bank is created by taking one vial from the master cell bank, expanding it for a few generations, and again freezing down hundreds of vials that each contain a few million cells. Working cell banks are the cell banks from which cells are taken to start each new manufacturing campaign. Importance of Cell Lines Why is accessing the original cell line important? Differences in protein structure may exist even between two proteins made from the same gene but produce by different cell lines. Changes to Cell Line or Growth Conditions: Same Gene, Different Protein Because cells are living and growing things, they change with time. This means that two different cells, even if they are both CHO cells, may have differences, and produce slightly different proteins. For example, one protein may have different post-translational modifications than the other. In Summary In summary, even if derived from the same gene - changes in the cell and/or changes in the growth conditions may result in different therapeutic proteins. Cell Culture Seeding & Scale-Up The biomanufacturing process starts by taking a vial from a working cell bank and using that to start a small-scale culture of approximately 1 liter. The goal is to grow thousands of liters of cells; however, cells need to grow at a certain density to survive. So, to maintain that required cell density, the cell culture is transferred into successively larger volumes until the final production volume is reached. At each step along the way of the scale-up process, there must be continuous, real-time quality control monitoring to ensure cell viability and concentration, product concentration and activity, and optimal growth conditions. Variations in the Scale-Up Process The process of cell culture scale-up has many possible variations between manufacturers. Any of these variations could potentially impact the structure and function of the final product. Variations include differences in vessels used, components of the growth media including 14 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. nutrients, growth factors, type of carbohydrates, and physical conditions of the growth culture such as temperature, oxygen and carbon dioxide concentration, pH, and nutrient circulation. All of these are possible variations. Production Options Both bacteria and mammalian cells are used in the production of biologic drugs. Bacteria cells are easier and less expensive to grow than mammalian cells. And because bacteria grow quickly, a production campaign can be completed in days instead of weeks, as required with the slowergrowing mammalian cells. However, bacteria cells can only be used to produce relatively simple biologics such as insulin. For more complex biologics, such as monoclonal antibodies, mammalian cells are required. Only mammalian cells are capable of producing intricately folded proteins with the correct posttranslational modifications. Harvesting & Purification Process The final protein product - which will be the active ingredient in the biologic drug - must be purified away from all other cellular components to be therapeutically useful. Some proteins, such as monoclonal antibodies and growth factors, are secreted by the cell and will be present in the cell culture fluid. Other proteins, such as enzymes, will be retained inside the cell. Chromatography In either case, all the proteins are extracted and loaded onto a column - a tool that can separate proteins based on charge, shape, and size. This process is called chromatography. Harvesting & Purification Process Overview The exact steps taken to extract and separate proteins have a significant impact on the purity profile of the final biologic. Any change to the purification process may alter the purity profile of the product and may affect its clinical efficacy and safety. Harvesting and purification steps include separating cells from the cell culture fluid to obtain a cell-free supernatant; extracting product from the supernatant, and product purification. 15 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. Biologic Formulation Defined Once purified, the protein product must be formulated, or mixed with other ingredients to provide an adequate volume and consistency for patient administration. The formulation is defined as the process in which different chemical substances, including the active drug, are combined to produce a final medicinal product. Biologic Formulation: Critical to Stability Key elements of the formulation protect the protein product from degradation. Correct buffer choice enables the final formulation to resist changes in pH and maintain a consistent pH. pH is a key parameter that has a major impact on stability. Protein stability is critical for correct protein structure and function. Surfactants are added to protect the protein product from unfolding at air/water or glass/water interface, and a range of different stabilizers may be added to help maintain the product’s shelf life. Because formulation influences product attributes such as bioavailability and stability, it is critical to the safety and efficacy of the final biologic. Biosimilar Formulation In many cases, the formulation of a biosimilar is likely to vary from the formulation of the innovator biologic due to patent protection on the original formulation which expires later than patent protection on the product itself. Stability Studies: Testing Protein Quality Over Time For every new formulation or change to an existing product formulation, product stability studies must be conducted. Stability studies involve storing the product for 6 to 12 months at room temperature or refrigerated conditions, and different humidity levels. Throughout that period, product samples are monitored for the following attributes: appearance, pH, protein concentration, particulate matter, protein aggregates, product fragmentation, and product activity. Section 4: Manufacturing Summary In this section, we learned how biologics are made, and what impact that has on the production of biosimilars. A common saying in biopharma is “The product is the process.” To summarize the main points of section 4: 16 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. ● Biomanufacturing is the use of living cells to make biological products. ● The steps of a biomanufacturing campaign include establishing cell banks, cell culture seeding, and scale-up, harvest and purification, formulation, fill and finish. ● A master cell bank is produced from the original therapeutic-producing cell line. A working cell bank is derived from the master cell bank and used to establish the cell line to initiate the manufacturing process. ● Different master cell lines with the same gene can produce different proteins because changes in the cell and/or changes in the growth conditions may result in different biologics. ● Variations could potentially impact the structure and function of the final product. Variations include differences in vessels used, components of the growth media including nutrients, growth factors, type of carbohydrates, and physical conditions of the growth culture such as temperature, oxygen and carbon dioxide concentration, pH, and nutrient circulation. 17 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. Section 5: Biosimilars Safety & Regulation Welcome In this section, we will describe the possible safety concerns regarding biosimilar products and learn the framework of how regulatory agencies are approving biosimilars for sale in the marketplace. Section 5: Biosimilars Safety & Regulation Objectives After this section, you should be able to: ● Define immunogenicity. ● Summarize possible outcomes of an immune response to a therapeutic protein. ● Define data exclusivity. ● Explain how companies can gain approval for a biosimilar. Protein Complexity As discussed, proteins are highly complex, and their therapeutic effect is dependent on having the correct structure. Even a slight change to the protein sequence, shape, or post-translational modifications may alter a biologic drug’s safety and efficacy. Immunogenicity Defined Biologic drugs typically have a very good safety profile. However, one possible concern for any biologic drug is immunogenicity. Immunogenicity is the ability of a foreign substance, such as a biologic, to provoke an immune response in the body. The consequences of this are unpredictable, and in some cases may be severe. Biologics can trigger an unwanted immune response with varying, unpredictable consequences. The immune response includes the body producing antibodies to attack the biologic. Proving these adverse reactions do not occur as part of the biologics drug development process before getting regulatory approval. Biologics Safety: Immunogenicity Immunogenicity is the ability of a foreign substance, such as a biologic, to provoke an immune response in the body. Immunogenicity is a rare but serious autoimmune response that may be 18 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. life-threatening. Immunogenicity is always a potential safety issue for biologic drugs. It is caused by protein aggregates, clumps of proteins, in the biologic. Since the biosimilar may not be identical to the brand name biologic, unanticipated immunogenic responses may arise from its use. Therefore, biosimilars must be thoroughly evaluated for immunogenicity both pre- and post-market approval. Clinical Impact Let’s take a closer look at why the binding of a patient’s antibodies to a biologic drug has a potential clinical impact. Antibody binding may neutralize - render ineffective - a product’s activity. This is called a neutralizing antibody. A non-neutralizing antibody may influence how the patient clears the drug, by either extending or reducing the half-life in the blood or may influence where the biologic goes in the body, impacting biodistribution. Extending/Reducing Drug Half-Life Extending the drug’s half-life - the amount of time that it stays in circulation - may enhance its efficacy, but it may also enhance any potential toxicity. Reducing the amount of time the drug is in circulation, is likely to reduce its efficacy. Antibody-binding may also impact where the biologic drug ends up within a patient’s body - its biodistribution. This may be critically important for the product’s efficacy - if it is not in the right place in the body, it will not work as expected. Similar Does Not Mean Same Making a brand-name chemical compound drug into a generic is pretty straightforward. It is technically possible to prove the generic is identical to the chemical compound brand drug using standard analytic techniques. The more complex structure of a protein therapeutic, or biologic drug, means that we can only ascertain the similarity between 2 proteins. Current technical capabilities make it extremely difficult to analyze the exact structure of a protein to demonstrate that it is identical to the reference biologic. Data Exclusivity For a new drug or biologic to gain marketing approval, it must undergo years of rigorous clinical testing to demonstrate that it is both safe and effective. Since a small molecule generic drug is an exact copy of the innovator small molecule, the generic manufacturer may use the clinical safety and efficacy data generated by the innovator company to support its case for approval. However, 19 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. a generics manufacturer will not be able to use this data for 5 years after the innovator product is approved. This is called the data exclusivity period - the period during which the innovator has exclusive use of clinical trial data to make a case for the safety and efficacy of the product. The number of years of data exclusivity varies between different regions of the world, as well as between small molecule and biologic drugs. In the U.S. and Europe, biologic drugs are given a longer data exclusivity period than small molecule drugs. Makers of biologics enjoy a longer period of data exclusivity, in recognition of the fact that the much higher production costs for these types of products mean that companies require more time to recover their development costs. Biosimilars Need to Show Similarity In the next few slides, we will discuss how a biosimilar product may benefit from an innovator biologic’s safety and efficacy data by first showing that it is highly similar, or comparable, to the innovator’s product. How to Gain Approval for Biosimilars The EMA and FDA have released guidance documents outlining the regulatory pathway for the approval of biosimilars. Both organizations require the biosimilar manufacturer to demonstrate comparability to a reference product - the already approved biologic that the biosimilar is meant to imitate. The first step for companies to receive biosimilar approval is extensive analytical characterization, in an effort to show comparability to the reference product. If biosimilarity is established, the biosimilar may qualify for abbreviated preclinical and clinical testing and use some of the innovator’s safety and efficacy data, once the exclusivity period is expired. Biosimilarity: Cell-Based Assays Cell-based, preclinical, and clinical testing programs for biosimilars are developed on a case-bycase basis, using product knowledge gained from preceding innovator biologic studies. Cellbased assays can be undertaken prior to preclinical and clinical studies to establish biosimilar comparability in reactivity. An example of a cell-based assay would be to demonstrate that the biosimilar binds to a given receptor in the same manner as the innovator biologic or reference product. 20 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. Biosimilarity: Preclinical Data A biosimilar manufacturer should also be able to demonstrate that their biosimilar behaves comparably to the innovator or reference product when tested in animals. Therefore, animal studies should be undertaken before embarking on clinical trials. Biosimilarity: Clinical Trials Most biosimilar products will require at least some clinical data for approval, as non-clinical and preclinical data will not be sufficient to identify all of the potential differences from the reference product. In most cases, comparative clinical trials will be necessary to demonstrate biosimilarity between biosimilar products and reference product. Clinical trials of proposed biosimilar products are necessary to establish equivalent clinical safety and efficacy. Data from an innovator or reference product clinical studies normally are insufficient to identify all potential differences in the biosimilar. Section 5: Biosimilars Safety & Regulation Summary In section 5 we took an in-depth look at biosimilar safety and regulation and ended with a checklist of what is needed for a biosimilar to gain approval in both the United States and the European Union. The main points made include: ● Immunogenicity is the ability of a foreign substance, such as a biologic, to provoke an immune response in the body. ● Possible outcomes of an immune response to a biologic includes the body producing antibodies to attack the therapeutic protein. For innovator biologics and biosimilars proving these adverse reactions do not occur is part of the drug development process before getting regulatory approval to market that innovator biologic. ● Data exclusivity period is the time during which the innovator has exclusive use of clinical trial data to make a case for the safety and efficacy of the product. ● The first step for companies to receive biosimilar approval is extensive analytical characterization to show comparability to the innovator's product. If biosimilarity is established, the biosimilar may qualify for abbreviated preclinical and clinical 21 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. testing and use some of the innovator’s safety and efficacy data once the exclusivity period is expired. 22 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. Section 6: Biosimilars Case Studies Welcome Over the next few slides, we will present a few case studies of biosimilars. The case studies demonstrate that differences in proteins can be created by small changes in the protein sequence, changes to the process of purification of the protein, or even changes to the packaging of the protein. These changes can then lead to changes in the safety or efficacy of the biologic. It is important to consider these case studies because they demonstrate that care must be taken when attempting to demonstrate biosimilarity between a reference product and a biosimilar because it is difficult to determine how even small changes in the creation of the product might lead to different clinical outcomes. Section 6: Biosimilars Case Studies Case Study 1: Changes in Amino Acid Sequence Affect Properties of Biologics A slow-acting version of insulin was created by substituting three amino acids in the native sequence. This substitution made insulin less soluble at physiologic pH, resulting in one injection lasting for 24 hours. Case Study 2: Differences in Impurity Profile May Result in Differences in Immunogenicity Slight differences in a purification protocol may result in differences in the final product that could invoke an unexpected immune response. An example of this can be found in the development of Omnitrope, a biosimilar of Genotropin, or human growth hormone. Initial clinical studies of Omnitrope detected antibodies against the biosimilar in the patient's blood. This immune response was traced to differences in the purity profile of the biosimilar relative to the reference product. Additional purification steps succeeded in reducing the immune response in Phase 3 clinical testing. The EMA was approved after the revised purification protocol was established. Case Study 3: Careful Analysis of Proposed Biosimilar Product May Detect Significant Differences Before Clinical Trials Differences in impurity profiles may lead to decreased efficacy and a higher rate of adverse events due to the medication. An example of this is Alpheon, a biosimilar to Roferon-A or interferonalpha. Alpheon did not show the same rates of safety and efficacy in clinical trials as the reference product, likely due to a different impurity profile. Developers were not able to fix this and the drug was not approved. 23 Copyright 2024 Biotech Primer, Inc. PROPRIETARY. DO NOT SHARE. Case Study 4: Packaging Changes May Have Serious Safety Consequences Changes to the formulation and packaging of a biologic may lead to unanticipated immunogenic reactions. An example of this is the serious safety issues that arose from a change in packaging for an erythropoietin biosimilar, Eprex. The formulation of Eprex differed from the reference product in one component. Polysorbate was used as a stabilizer instead of human serum albumin. Polysorbate absorbed organic compounds from the rubber stoppers in the medicine vials, which triggered an immune response in patients. The immune response caused these patients’ immune cells to attack their red blood cells or red blood cell aplasia. The problem was solved by replacing the rubber stoppers with Teflon-coated stoppers. Case Study 5: Need for Clinical Trials to Investigate Therapeutic Outcomes in Humans Clinical outcomes are not always predictable based on preclinical results. An example of this is found in Silapo, a biosimilar to the reference product Eprex. In preclinical animal studies, researchers detected a 5-fold increase in antibody production against Silapo as compared to Eprex. However, in human clinical studies, this was not seen, allowing the drug to be approved. 24 Copyright 2024 Biotech Primer, Inc.