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Biologic Therapies:
Optimizing Therapies Online Monograph

Bioengineering and Biosimilars

Jonathan Kay, MD

Summary of the Bioengineering and Biosimilars session, presented by Gregg Silverman, MD; Vibeke Strand, MD; and Jonathan Kay, MD. This article is transcribed from a talk, not a primary written piece.

How Are Immune Therapeutics Bioengineered? Monoclonals, Bi-Specific Antibodies, Nanobodies, and More

Gregg J. Silverman, MD

This presentation reviewed the structures of naturally occurring antibodies, strategies to develop therapeutic monoclonal antibodies, and novel therapeutic agents based on antibodies that are in development.

Naturally occurring IgG antibodies consist of two light chains and two heavy chains, each of which contains one variable and one to three constant regions. Six hypervariable regions, three on the variable region of the light chain and three on variable region of the heavy chain, contribute to the antigen recognition specificity of the antibody. The constant regions of the heavy chains engage Fc receptors. The connected constant regions of the heavy chains engage Fc receptors on the membranes of a range of host cells and contain sites that can activate the complement cascade.

Therapeutic monoclonal antibodies include chimeric antibodies, such as infliximab and rituximab, that contain mouse immunoglobulin sequences in the variable regions. Humanized and fully human antibodies, such as adalimumab and golimumab, use constant regions and variable regions that closely resemble those of human antibodies and are engineered to be less immunogenic.

Engineering antibodies by placing antibody genes into a filamentous phage expression vector and expressing light and heavy chains in bacterial cells has yielded monoclonal antibodies that bind both antigen and Fc receptors with greater affinity than monoclonal antibodies generated using hybridoma technology. An alternative approach, in which mice were genetically modified to remove their inherited immunoglobulin loci with the subsequent insertion of human immunoglobulin genes, has allowed generation of human antibodies of virtually any specificity by immunizing these mice to the desired antigens.

Chemical conjugation of polyethylene glycol (PEG) molecules (PEGylation) can decrease immunogenicity and increase the circulating half-life of an antibody. Certolizumab pegol is an example of a PEGylated Fab fragment of an anti-TNF antibody, which has been approved for treatment of rheumatoid arthritis and other inflammatory diseases.

Another novel approach has joined portions of different antibodies to create bifunctional, trifunctional, and tetrafunctional antibodies. These multifunctional antibodies can bind several different antigens, allowing for previously unachievable therapeutic specificity. For example, by binding both a soluble factor and the surface of a specific cell type, a bifunctional antibody could bring the soluble protein and the cell together to cause a specific therapeutic action.

Comparative studies of antibodies from different species of animals led to the discovery of the smallest antigen-binding protein: the nanobody. With molecular weight of about 8 kDa (nearly 1/20th that of an IgG molecule), nanobodies are postulated to better penetrate tissue than antibodies and may allow previously unattainable therapeutic and diagnostic applications.

Immunogenicity and Serum Levels Across Immune-Mediated Inflammatory Diseases

Vibeke Strand, MD

This presentation reviewed the various factors that contribute to the immunogenicity of biologic agents, assays for anti-drug antibodies (ADAs), and consequences of antidrug antibodies. Development of ADAs is influenced by various factors, including the following:

  • Degree to which the therapeutic protein differs from naturally occurring human proteins,
  • Route and schedule of drug administration,
  • Coadministration of other medications,
  • Underlying disease that is being treated.

Chimeric monoclonal antibodies, such as infliximab, are more immunogenic than fully human monoclonal antibodies, because a chimeric antibody contains murine or other nonhuman epitopes. When administered subcutaneously, a biopharmaceutical is more immunogenic than when it is administered intravenously. Also, intermittent treatment with a biologic agent, such as when infliximab is used to treat Crohn's disease, is more likely to induce ADAs than continuous maintenance treatment.

Concomitant treatment with methotrexate alone or with other DMARDs reduces ADA formation. Methotrexate and other antiproliferative drugs decrease B cell production of ADA and prolong the half-life of the biopharmaceutical, which also reduces its immunogenicity. Interestingly, in clinical trials of CT-P13, an infliximab biosimilar, the proportion of patients who developed ADAs when treated with either the biosimilar or infliximab was greater among those with rheumatoid arthritis who received infliximab (3 mg/kg IV) plus methotrexate than among patients with ankylosing spondylitis who received a higher dose of infliximab (5 mg/kg IV) as monotherapy. This suggests that ADA development may have been influenced by either the underlying disease or the induction of immunologic tolerance by higher drug concentrations.

The presence of ADAs correlates with lower serum trough concentrations of drug, loss of clinical response, and greater risk of infusion or injection site reactions. Neutralizing ADAs, especially if directed against the epitope binding region of a monoclonal antibody, reduces the therapeutic efficacy of a drug.

Various assays have been used to measure ADAs (Figure 1). Initially, ADAs were measured using bridging ELISA or a radioimmunoassay, in which the circulating drug interferes with ADA detection. More recently, an electro-chemiluminescent immunoassay has been used to measure ADA levels. Using the pH-shift anti-idiotype antigen-binding (PIA) method, in which drug is dissociated from ADAs, allows better detection of ADA in the presence of circulating drug.

Because many ADAs are transient, the clinical utility of ADA measurement is limited. Thus, among patients with most immune-mediated inflammatory diseases, assessment of clinical disease activity is the best way to follow response to drug therapy over time. Only in the treatment of inflammatory bowel diseases is measurement of ADA and trough drug concentrations used regularly in clinical practice.

Biosimilars

Part 1: History and Biology

Jonathan Kay, MD

Part 2: Regulatory and Current Status

Vibeke Strand, MD

These two presentations provided an overview of the emerging biosimilars market, reviewing the definition of biosimilars, both those currently available and those in development, and the pathways that have been established for their review and approval by regulatory agencies.

A biosimilar is a version of an already approved biopharmaceutical that has been developed to be highly similar to its reference product in its physicochemical characteristics, biologic activity, pharmacokinetics, efficacy, safety, and immunogenicity. Regulatory agencies require a comprehensive, stepwise comparability exercise to demonstrate similarity of the biosimilar to its reference product within a prespecified equivalence margin. Because biosimilars must be equipotent to their reference products, they must have the same dose, dosing regimen, and formulation.

Biosimilars are not biomimics, which are replicas of marketed biopharmaceuticals that have not undergone full review or approval by a regulatory agency according to a pathway for evaluation of biosimilars. Biosimilars also are not second-generation biopharmaceuticals, which are intended to improve upon the reference product. These have the same mechanism of action as the original biopharmaceutical but differ in their primary amino acid sequences. Furthermore, biosimilars are not generic medications, which are exact copies of small molecule drugs. Because proteins produced under different culture conditions and in different cell lines have different posttranslational modifications, a biosimilar never can be an exact copy of its reference product.

Guidance for the regulatory review and approval of biosimilars has been developed by the World Health Organization, the European Medicines Agency, Health Canada, the United States Food and Drug Administration (FDA), and several other national regulatory agencies. In contrast to that of an originator biopharmaceutical, for which phase II and large III phase clinical studies must be conducted, the development of a biosimilar is based on extensive physicochemical and functional characterization comparing it to its reference product (see Figure 2). Animal toxicology studies, pharmacokinetic and pharmacodynamic studies, and at least one clinical trial comparing the biosimilar to its reference product are then conducted. However, phase II clinical trials are not needed because the biosimilar must be administered at the same dose as its reference product.

Each of these regulatory agencies has included a process that provides for the possibility of extrapolation of indications, whereby a biosimilar can be approved for some or all of the indications for which the reference product is approved after the biosimilar has been compared to the reference product in only that disease which is most sensitive to detecting potential differences between the two biopharmaceuticals. On the other hand, interchangeability is a designation that has been proposed only in the United States. If a biosimilar has been shown to have equivalent efficacy and safety to its reference product after switching between the two biopharmaceuticals, the FDA may allow someone other than the prescribing health professional to substitute that biosimilar for the reference product. However, specifics regarding the evidence required to obtain this designation have not yet been disseminated and no biosimilar yet has been deemed interchangeable.

Globally, currently marketed biosimilars include six different erythropoiesis-stimulating agents, eight different granulocyte colony-stimulating factors, one human growth hormone, one insulin glargine, and one follicle stimulating hormone. Additionally, CT-P13, an infliximab biosimilar developed in South Korea, has been approved by regulatory agencies in more than 70 countries including Canada, the European Union, and Japan. It has been approved for the treatment of ankylosing spondylitis, psoriatic arthritis, plaque psoriasis, rheumatoid arthritis, and adult and pediatric Crohn's disease and ulcerative colitis. However, extrapolation of indications to inflammatory bowel diseases was not granted by Health Canada. Davictrel, an etanercept biosimilar, was approved in South Korea in November 2014, and Exemptia, an adalimumab biosimilar, was approved in India in December 2014.

CT-P13 was studied as monotherapy in PLANETAS, a phase I study in patients with ankylosing spondylitis, and in combination with methotrexate in PLANETRA, a phase III study in patients with rheumatoid arthritis. In each study, equivalence of CT-P13 and reference infliximab (Remicade) within a prespecified equivalence margin was demonstrated. Pharmacokinetic parameters for the two versions of infliximab were shown to be equivalent in the PLANETAS study. Safety, efficacy, and immunogenicity were shown to be equivalent in both studies. Interestingly, although similar proportions of patients treated with either biosimilar or reference infliximab developed antidrug antibodies (ADAs), only about 25% of ankylosing spondylitis patients treated with infliximab 5 mg/kg as monotherapy developed ADAs whereas nearly 50% of rheumatoid arthritis patients treated with infliximab 3 mg/kg in combination with methotrexate developed ADAs. Thus, the higher infliximab dose, even in the absence of methotrexate cotherapy, might have induced immunologic tolerance resulting in a lower proportion of patients with ADAs.

Many biosimilars are currently in development to treat inflammatory diseases. These include biosimilars of adalimumab, etanercept, infliximab, rituximab, and tocilizumab. When the patents expire for the originator products, some of these biosimilars will be poised to enter the market as lower-priced alternatives to the effective biological therapies.

Commentary
Bioengineering and Biosimilars - Is this ready for the clinic?

Leonard Calabrese, DO

Leonard Calabrese, DO

First I will prognosticate and easily say that biosimilars are coming our way in the US as they are already in Europe (biosimilar infliximab in UK and Europe as of 2015). The major hurdles in the US have been legal, with fierce defense positions taken up by many companies who, ironically, are developing their own biosimilar programs.

However, in terms of immunogenicity, the issue is real and with us at this very moment. Dr. Strand did a wonderful job explain this complex area. Whenever a clinician, whether it be a rheumatologist, gastroenterologist, or dermatologist sees a patient who has failed an anti-TNF, a series of questions are posed and attempted to answer. These include:

  1. Did the drug work at all (i.e. was this a primary failure)?
  2. Did the drug fail from a side effect?
  3. Did the drug work initially and then lose its punch (i.e. secondary failure)?
  4. If the drug failed secondarily, could it have been to immunogenicity?
  5. If there is reason to suspect or prove immunogenicity, is it due to a problem with the disease, comorbidity, concomitant therapy or absence thereof or another issue entirely?

In my opinion, gastroenterologists are far ahead of the rest of us. Dr. Brian Feagan gave a strong argument for an algorithmic approach to using immunogenicity assays in the clinic (Figure 3).

Perhaps a reason why gastroenterologists are more invested in this issue is that they have fewer agents to use and thus, much stretch their exiting armamentarium. Rheumatologists and now dermatologists have many more drugs available and are able to more easily switch to another mechanism of action. I predict in time that more commercial (and better) assays will become available and we too will be incorporating such an approach.

Finally, what about bioengineering? I think the future of novel constructs is bright for our field but these are several years away from practice. Stay tuned…

Annotated Bibliography

Bendtzen K, Ainsworth M, Steenholdt C, et al. Individual medicine in inflammatory bowel disease: Monitoring bioavailability, pharmacokinetics and immunogenicity of anti-tumour necrosis factor-alpha antibodies. Scand J Gastroenterol. 2009;44:774-81.

This review discusses the pharmacokinetics of anti-TNF antibodies in patients with inflammatory bowel diseases, especially as it relates to immunogenicity. It provides a clear discussion of the mechanisms by which antidrug antibodies are induced against therapeutic anti-TNF antibodies.

Castañeda-Hernández G, Gonzalez-Ramirez R, Kay J, Scheinberg M. Biosimilars in rheumatology: What clinicians should know. RMD Open. 2015;1:e000010.

This review discusses the current status of biosimilars in rheumatology and provides a clear explanation of methods to assess therapeutic equivalence in clinical trials.

De Meyer T, Muyldermans S, Depicker A. Nanobody-based products as research and diagnostic tools. Trends Biotechnol. 2014;32(5):263-70.

A review of the opportunities provided by very small therapeutic and diagnostic antibodies.

Grandjenette C, Dicato M, Diederich M. Bispecific antibodies: an innovative arsenal to hunt, grab and destroy cancer cells. Curr Pharm Biotechnol. 2015;16(8):670-83.

An opinion article of proposed approaches to the development of bifunctional biologic agents, which now are used primarily for the treatment of cancer.

Lonberg N. Fully human antibodies from transgenic mice and phage display. Curr Opin Immunol. 2008;20(4):450-9

A narrative on the development of novel antibody discovery approaches using phage-display technology and mice transgenic for human immunoglobulin genes.

Mouser JF, Hyams JS. Infliximab: a novel chimeric monoclonal antibody for the treatment of Crohn's disease. Clin Ther. 1999;21(6):932-42

An account of the development and application of the first therapeutic antibody designed to block TNF during inflammatory disease.

Scheinberg MA, Kay J. The advent of biosimilar therapies in rheumatology - "O Brave New World." Nat Rev Rheumatol. 2012;8:430-6

This article provides a comprehensive review of biosimilars to treat rheumatologic diseases, including an overview of regulatory pathways worldwide and of biosimilars in development.

van Schouwenburg PA, Rispens T, Wolbink GJ. Immunogenicity of anti-TNF biologic therapies for rheumatoid arthritis. Nat Rev Rheumatol. 2013;9(3):164-72

This article provides a comprehensive review of the issues surrounding immunogenicity of therapeutic proteins, focusing on TNF inhibitors. It provides a clear discussion of the different methods available to detect antidrug antibodies.

Yoo DH, Hrycaj P, Miranda P, et al. A randomised, double-blind, parallel-group study to demonstrate equivalence in efficacy and safety of CT P13 compared with innovator infliximab when coadministered with methotrexate in patients with active rheumatoid arthritis: the PLANETRA study. Ann Rheum Dis. 2013;72:1613-20

These landmark articles report the results of the phase I and phase III clinical trials of CT-P13, the first biosimilar that was approved to treat inflammatory diseases.