1. INTRODUCTION
- Background
Biotechnology-derived pharmaceuticals, or biopharmaceuticals, were first developed in the early 1980s, with initial marketing authorizations occurring later in the decade. Various regulatory agencies have issued guidelines for the safety assessment of these products, which are beneficial for developing new biopharmaceuticals. Extensive experience with application submissions has been the basis for the development of guidance aimed at establishing general principles for preclinical safety evaluation programs.
Considerable experience has now been gathered with submission of applications for biopharmaceuticals. Critical review of this experience has been the basis for development of this guidance that is intended to provide general principles for designing scientifically acceptable preclinical safety evaluation programs.
- Objectives
The primary goals of preclinical safety evaluation include identifying a safe initial dose and dose escalation for humans, determining potential target organs for toxicity and its reversibility, and establishing safety parameters for clinical monitoring. Adhering to these principles aims to enhance the quality and consistency of preclinical safety data for biopharmaceutical development.
- Scope
This guidance recommends a framework for preclinical safety evaluation of biotechnology-derived pharmaceuticals, encompassing products from various expression systems such as bacteria, yeast, and mammalian cells. It addresses active substances including proteins, peptides, and their derivatives, with uses in diagnostics, therapeutics, or prophylactics. Examples include cytokines, growth factors, and monoclonal antibodies. The principles also apply to recombinant DNA protein vaccines and oligonucleotide drugs, but exclude antibiotics, conventional vaccines, and gene therapies.
2. SPECIFICATION OF THE TEST MATERIAL
Safety concerns include impurities or contaminants, necessitating purification over preclinical testing. Proper product characterization is essential for designing safety studies. Host cell contaminants, including bacteria and mammalian cells, can pose risks such as allergic reactions, while nucleic acid contaminants may theoretically integrate into host genomes, and products from certain cells may carry viral infection risks. The test product for pharmacology and toxicology studies should match the product for clinical trials, though manufacturing changes may occur. Comparability during development must be demonstrated with biochemical and biological characterizations, and additional studies may be required, supported by scientific rationale.
3. PRECLINICAL SAFETY TESTING
3.1 General Principles
The objectives of the preclinical safety studies are to define pharmacological and toxicological effects not only prior to initiation of human studies but throughout clinical development. Both in vitro and in vivo studies can contribute to this characterisation. Biopharmaceuticals that are structurally and pharmacologically comparable to a product for which there is wide experience in clinical practice may need less extensive toxicity testing.
Preclinical safety testing should consider: 1) selection of the relevant animal species; 2) age; 3) physiological state; 4) the manner of delivery, including dose, route of administration, and treatment regimen; and 5) stability of the test material under the conditions of use.
Toxicity studies should follow Good Laboratory Practice (GLP), but some tests for biopharmaceuticals may not fully comply. It’s important to identify areas of non-compliance and evaluate their impact on safety assessments. In certain cases, these studies can still support clinical trials and marketing authorizations even with partial GLP adherence.
Conventional toxicity testing methods for pharmaceuticals are often unsuitable for biopharmaceuticals, owing to their unique structural and biological characteristics such as species specificity, immunogenicity, and unpredictable pleiotropic effects.
3.2 Biological Activity/Pharmacodynamics
In vitro assays can evaluate biological activity of biotechnology-derived pharmaceuticals, examining direct effects on cellular phenotype and proliferation. Cell lines derived from mammalian cells can predict in vivo activity and assess species sensitivity to biopharmaceuticals. These studies help select appropriate animal species for further pharmacology and toxicology studies. The combined results from in vitro and in vivo studies support the rationale for clinical studies.
Monoclonal antibodies should have their immunological properties thoroughly detailed, covering aspects like antigenic specificity, complement binding, and any unintended reactivity or cytotoxic effects on human tissues separate from the target. Cross-reactivity studies must be conducted using relevant immunohistochemical techniques on various human tissues.
3.3 Animal Species/Model Selection
The biological activity together with species and/or tissue specificity of many biotechnology-derived pharmaceuticals often preclude standard toxicity testing designs in commonly used species (e.g., rats and dogs). Safety evaluation programs should include the use of relevant species. A relevant species is one in which the test material is pharmacologically active due to the expression of the receptor or an epitope (in the case of monoclonal antibodies). A variety of techniques (e.g., immunochemical or functional tests) can be used to identify a relevant species. Knowledge of receptor/epitope distribution can provide greater understanding of potential in vivo toxicity.
Relevant animal species for testing of monoclonal antibodies are those that express the desired epitope and demonstrate a similar tissue cross-reactivity profile as for human tissues. This would optimise the ability to evaluate toxicity arising from the binding to the epitope and any unintentional tissue cross-reactivity. An animal species which does not express the desired epitope may still be of some relevance for assessing toxicity if comparable unintentional tissue cross-reactivity to humans is demonstrated.
Safety evaluation programs typically involve two relevant species, though one may be sufficient in some cases, such as when only one can be identified or the biopharmaceutical’s biological activity is well understood. Even when two species are needed for short-term toxicity studies, justification may allow for using only one species in long-term studies if the toxicity profiles are comparable.
Toxicity studies in non-relevant species are discouraged due to their potential misleading results. In the absence of relevant species, transgenic animals expressing human receptors or homologous proteins should be considered. The effectiveness of transgenic models is heightened when the interaction with the humanized receptor mimics physiological effects in humans. Although homologous proteins may provide useful data, differences in production, impurities, pharmacokinetics, and pharmacological mechanisms must be acknowledged. If neither transgenic models nor homologous proteins can be utilized, limited toxicity evaluations in a single species should be conducted, such as a repeated dose study under 14 days that assesses critical functional endpoints like cardiovascular and respiratory health.
In recent years, there has been much progress in the development of animal models that are thought to be similar to the human disease. These animal models include induced and spontaneous models of disease, gene knockout(s), and transgenic animals. These models may provide further insight, not only in determining the pharmacological action of the product, pharmacokinetics, and dosimetry, but may also be useful in the determination of safety (e.g., evaluation of undesirable promotion of disease progression). In certain cases, studies performed in animal models of disease may be used as an acceptable alternative to toxicity studies in normal animals (Note 1). The scientific justification for the use of these animal models of disease to support safety should be provided.
3.4 Number/Gender of Animals
The number of animals used per dose influences toxicity detection, as small sample sizes may miss toxic events despite their severity. In non-human primate studies, increasing monitoring frequency and duration can help compensate for these limitations. Both genders should typically be included unless specific justifications are provided for their omission.
3.5 Administration/Dose Selection
The administration route and frequency for clinical trials should closely align with proposed human use, factoring in pharmacokinetics, bioavailability, and safe volumes for test animals. Increased administration frequency in laboratory animals may be necessary due to faster clearance or low solubility of active ingredients, and exposure levels must be defined accordingly. Additionally, aspects such as volume, concentration, formulation, and administration site warrant consideration. Alternative routes may be warranted if bioavailability is limited or due to the species’ size or physiology.
Dosage levels must be chosen to elucidate a dose-response relationship, specifying toxic doses and NOAEL. For less toxic products, a defined maximum dose may not exist, requiring scientific justification for dose selection and exposure multiples. High dose selection justification should reflect pharmacological effects, available test materials, and intended clinical use. If a product exhibits lower affinity or potency in animal models compared to humans, higher doses are essential for safety margin evaluation, which varies with biotechnology-derived pharmaceuticals and their clinical uses.
3.6 Immunogenicity
Biotechnology-derived pharmaceuticals can be immunogenic in animals, necessitating antibody measurements during repeated dose toxicity studies to interpret results accurately. The characterization of antibody responses—such as titer, number of responding animals, and type (neutralising or non-neutralising)—is essential. These responses should be correlated with pharmacological and toxicological changes, considering their impact on pharmacokinetics, adverse effect severity, complement activation, and new toxic effects. Evaluating potential pathological changes from immune complex formation and deposition is also crucial.
The early termination of a preclinical safety study should not solely rely on antibody detection unless the immune response significantly neutralizes the biopharmaceutical’s effects in a large proportion of test animals. Typically, immune responses to biopharmaceuticals vary, similar to human responses. If the safety study’s data interpretation remains intact, the antibody response should not be given undue importance.
The induction of antibody formation in animals is not predictive of a potential for antibody formation in humans. Humans may develop serum antibodies against humanised proteins, and frequently the therapeutic response persists in their presence. The occurrence of severe anaphylactic responses to recombinant proteins is rare in humans. In this regard, the results of guinea pig anaphylaxis tests, which are generally positive for protein products, are not predictive for reactions in humans; therefore, such studies are considered of little value for the routine evaluation of these types of products.
The entire text of ICH S6 (R1) could not be presented in this article due to space constraints. To properly grasp, PRECLINICAL SAFETY EVALUATION OF BIOTECHNOLOGY-DERIVED PHARMACEUTICALS, it is advised that you read the entire guideline. The link is provided below:
Reference:
https://database.ich.org/sites/default/files/S6_R1_Guideline_0.pdf
– Dr Subramanian S Iyer
