Can someone conclude to me how we measure immune responses in Animal models in pre-clinical studies?
I know there are many ways and many essays. I want an opening to know how to start thinking in this part of immunology.
Thank you
Hi (copy from your question into discussion loop ...)
If your aim is study immunogenicity of human drugs in animal models then best way is analysis of guidances by FDA/EMA :) and total ADA (anti drug antibodies), fraction of Nab's (neutralising - high affinity antibodies) and seroconversion time. Such data could be colected after sigle dose of human protein. Sometimes pilot study is useful. Sometimes best way to analyse immunogenicity of human drug in animal model is simply analysis of drug pharmacokinetics. Then you just see what the difference is with Nab's because this fraction of antibodies increases the clearance of the drug that is administered. You can measure the most important cytokines and cells. The meaning of such work increases when your model is relevant in pharmacokinetics and pharmacodynamic meaning. Please remember that using the animal models for comparision test versus reference drug immunogenicity could be valuable (see guidelines below) but translation into humans impossible and off value. Please remember that different doses of the duman drug can induce different immune responce. For example higher dose of the drug may give lower immune responce if drug has immunosupresive action. Moreover in case of some drugs multiple dosing may decrease immune responce for example ustekinumab or infliximab ....
Some explanations from old FDA guidance could be usefull in your work (document is not available now)
„Animal immunogenicity assessments generally do not predict potential immunogenic responses to protein products in humans. However, when differences in manufacturing (e.g., impurities or excipients) between the proposed product and the reference product may result in differences in immunogenicity,
measurement of anti-protein antibody responses in animals may provide useful information relevant to patient safety. Additionally, significant differences in the immune response profile in inbred strains of mice, for example, may indicate that the proposed product and the reference product differ in one or more product attributes not captured by other analytical methods.”
GUIDANCE FOR INDUSTRY SCIENTIFIC CONSIDERATIONS IN DEMONSTRATING BIOSIMILARITY TO A REFERENCE PRODUCT FDA 2012
" A risk-based evaluation, focused on the mechanism of action of the therapeutic protein product as well as results of animal and in vitro evaluations should be performed to determine the need for collection of preand post-dose cytokine levels in the early phase of clinical development. In case of a clinical adverse event, such an evaluation may provide evidence to support the clinical diagnosis of cytokine release syndrome and help distinguish this entity from other acute drug reactions "
Guidance for Industry Immunogenicity Assessment for Therapeutic Protein Products
https://www.fda.gov/media/85017/download
Immunogenicity in animal models is not predictive of immunogenicity in humans.
Assessment of immunogenicity in animals may be useful to interpret nonclinical toxicology and pharmacology data.
Immunogenicity in animal models may reveal potential antibody related toxicities that could be monitored in clinical trials.
May reveal immunogenicity differences between biosimilar and reference product
The immunogenicity of therapeutic proteins- what you don’t know can hurt YOU and the patient
https://www.fda.gov/files/drugs/published/The-immunogenicity-of-therapeutic-proteins--what-you-don%E2%80%99t-know-can-hurt-YOU-and-the-patient--Fall-2014.pdf
" Therapeutic proteins show species differences in most cases. Thus, human proteins will be recognised as foreign proteins by animals. For this reason, the predictivity of non-clinical studies for evaluation of immunogenicity is considered low."
but .......
" the comparison of the antibody response to the reference product in an animal model may be part of the comparability exercise both for similar biological medicinal products "
https://www.ema.europa.eu/en/documents/scientific-guideline/guideline-immunogenicity-assessment-biotechnology-derived-therapeutic-proteins-first-version_en.pdf
" Qualitative or quantitative difference(s) of product-related variants (e.g. glycosylation patterns, charge variants) may affect biological functions of the biotechnology-derived protein and are expected to be evaluated by appropriate in vitro assays. These differences and impurities may have an effect on immunogenic potential and the potential to cause hypersensitivity. It is acknowledged that these effects are difficult to predict from animal studies and should be further assessed in clinical studies. "
Guideline on similar biological medicinal products containing biotechnology-derived proteins as active substance: non-clinical and clinical issues
https://www.ema.europa.eu/en/documents/scientific-guideline/guideline-similar-biological-medicinal-products-containing-biotechnology-derived-proteins-active_en-2.pdf
Some nice papers about could be useful
Article Immunogenicity of Therapeutic Proteins: The Use of Animal Models
Article Animal models as indicators of immunogenicity of therapeutic...
Article Preclinical Models Used for Immunogenicity Prediction of The...
Article Mouse Models for Assessing Protein Immunogenicity: Lessons a...
Article Development of a Human Antibody Tolerant Mouse Model to Asse...
Article Development of a Transgenic Mouse Model to Study the Immunog...
Article Preclinical Immunogenicity Testing for Recombinant Therapeut...
Hope it's helpful
Best regards
Tomasz
it depends on what kind of immunity you want to evaluate. your question is too general
Hi (copy from your question into discussion loop ...)
If your aim is study immunogenicity of human drugs in animal models then best way is analysis of guidances by FDA/EMA :) and total ADA (anti drug antibodies), fraction of Nab's (neutralising - high affinity antibodies) and seroconversion time. Such data could be colected after sigle dose of human protein. Sometimes pilot study is useful. Sometimes best way to analyse immunogenicity of human drug in animal model is simply analysis of drug pharmacokinetics. Then you just see what the difference is with Nab's because this fraction of antibodies increases the clearance of the drug that is administered. You can measure the most important cytokines and cells. The meaning of such work increases when your model is relevant in pharmacokinetics and pharmacodynamic meaning. Please remember that using the animal models for comparision test versus reference drug immunogenicity could be valuable (see guidelines below) but translation into humans impossible and off value. Please remember that different doses of the duman drug can induce different immune responce. For example higher dose of the drug may give lower immune responce if drug has immunosupresive action. Moreover in case of some drugs multiple dosing may decrease immune responce for example ustekinumab or infliximab ....
Some explanations from old FDA guidance could be usefull in your work (document is not available now)
„Animal immunogenicity assessments generally do not predict potential immunogenic responses to protein products in humans. However, when differences in manufacturing (e.g., impurities or excipients) between the proposed product and the reference product may result in differences in immunogenicity,
measurement of anti-protein antibody responses in animals may provide useful information relevant to patient safety. Additionally, significant differences in the immune response profile in inbred strains of mice, for example, may indicate that the proposed product and the reference product differ in one or more product attributes not captured by other analytical methods.”
GUIDANCE FOR INDUSTRY SCIENTIFIC CONSIDERATIONS IN DEMONSTRATING BIOSIMILARITY TO A REFERENCE PRODUCT FDA 2012
" A risk-based evaluation, focused on the mechanism of action of the therapeutic protein product as well as results of animal and in vitro evaluations should be performed to determine the need for collection of preand post-dose cytokine levels in the early phase of clinical development. In case of a clinical adverse event, such an evaluation may provide evidence to support the clinical diagnosis of cytokine release syndrome and help distinguish this entity from other acute drug reactions "
Guidance for Industry Immunogenicity Assessment for Therapeutic Protein Products
https://www.fda.gov/media/85017/download
Immunogenicity in animal models is not predictive of immunogenicity in humans.
Assessment of immunogenicity in animals may be useful to interpret nonclinical toxicology and pharmacology data.
Immunogenicity in animal models may reveal potential antibody related toxicities that could be monitored in clinical trials.
May reveal immunogenicity differences between biosimilar and reference product
The immunogenicity of therapeutic proteins- what you don’t know can hurt YOU and the patient
https://www.fda.gov/files/drugs/published/The-immunogenicity-of-therapeutic-proteins--what-you-don%E2%80%99t-know-can-hurt-YOU-and-the-patient--Fall-2014.pdf
" Therapeutic proteins show species differences in most cases. Thus, human proteins will be recognised as foreign proteins by animals. For this reason, the predictivity of non-clinical studies for evaluation of immunogenicity is considered low."
but .......
" the comparison of the antibody response to the reference product in an animal model may be part of the comparability exercise both for similar biological medicinal products "
https://www.ema.europa.eu/en/documents/scientific-guideline/guideline-immunogenicity-assessment-biotechnology-derived-therapeutic-proteins-first-version_en.pdf
" Qualitative or quantitative difference(s) of product-related variants (e.g. glycosylation patterns, charge variants) may affect biological functions of the biotechnology-derived protein and are expected to be evaluated by appropriate in vitro assays. These differences and impurities may have an effect on immunogenic potential and the potential to cause hypersensitivity. It is acknowledged that these effects are difficult to predict from animal studies and should be further assessed in clinical studies. "
Guideline on similar biological medicinal products containing biotechnology-derived proteins as active substance: non-clinical and clinical issues
https://www.ema.europa.eu/en/documents/scientific-guideline/guideline-similar-biological-medicinal-products-containing-biotechnology-derived-proteins-active_en-2.pdf
Some nice papers about could be useful
Article Immunogenicity of Therapeutic Proteins: The Use of Animal Models
Article Animal models as indicators of immunogenicity of therapeutic...
Article Preclinical Models Used for Immunogenicity Prediction of The...
Article Mouse Models for Assessing Protein Immunogenicity: Lessons a...
Article Development of a Human Antibody Tolerant Mouse Model to Asse...
Article Development of a Transgenic Mouse Model to Study the Immunog...
Article Preclinical Immunogenicity Testing for Recombinant Therapeut...
Hope it's helpful
Best regards
Tomasz
Teaching yourself immunology is difficult. I'd find an immunology lab and ask them to teach you. Immunology is a profession with a lot of lore.
Tomasz Grabowski Thank you so much. Very helpful.
Do you know any regulations for immunogenicity measurement against vaccines in animal models?
Thank you again.
I don't think there are any regulations; even the FDA guidance is just that-guidance as to standard of proof the FDA requires to support a definitive statement. But I suppose that this is not the issue here rather the issues is convincing yourself, I think. The ultimate proof is protection from disease, otherwise antibodies, first by ELISA, then by neutralization activity; and cellular immunity measured by a number of possible methods of in vitro manipulation of T cells. The main issue here is the relevant target cells.
Hi,
Some statements in regulatory documents are available:
NOTE FOR GUIDANCE ON PRECLINICAL PHARMACOLOGICAL AND TOXICOLOGICAL TESTING OF VACCINES (1997 now
Withdrawn)
"It is recognised that suitable animal models are not always available and responses in such models are not always predictive of human responses. Therefore, selection of animal species should be made on a case by case basis."
"The availability of animal models to address these issues should be considered in the development of a new vaccine."
"Single dose toxicity data from at least one animal species should be performed with a dose providing an adequate safety margin in relation to the human dose. However, if toxic findings are seen in this study, the dose response relationship should be further characterised. These data may be part of animal immunogenicity studies"
"A study on repeated dose toxicity in one animal species is normally requested for vaccines that will require multiple doses in the clinical setting. Even in cases where only single doses will be administered in clinical use, a repeated dose toxicity study may nevertheless be appropriate. The selection of an appropriate animal species should be carefully evaluated on a case by case basis."
"Valuable information can be obtained by expanding multiple-dose immunogenicity studies to include measurements normally conducted in preclinical toxicity studies (i.e. body weight, food consumption, clinical pathology, gross necropsy and histopathology). The value of the protocol would normally be enhanced by concurrent measurement of the antibody response to all important components of the vaccine (immunogenicity studies). Incorporation of safety pharmacology endpoints in the design of these studies should be considered"
"- Where appropriate, specific consideration should be given to immunological aspects of toxicity, such as production of complexes with host immunoglobulins (e.g. antibodydependent enhancement of disease) or release of immunofunctional molecules, (e.g. cytokines) affecting functions of the immune system.
- Hypersensitivity reactions, induced by the antigen itself, by antigens (toxins) modified in new ways (new detoxification procedure, by antigen-carrier complex or presence of minute amounts of impurities) or by additives adjuvants/excipients/preservatives) may be increased (especially for vaccines proposed to be injected more than once).
- In some rare cases, antigenic substances can induce antibodies that can cross-react with human tissue resulting in possible adverse effects and the availability of an animal model to address these issues should be considered."
"Primary pharmacodynamic studies with respect to the ‘antigen-protective response’ should be carried out in a relevant species. The endpoint in these kind of studies should preferably be the protection against a challenge from the pathogenic organism where there is an animal model reflecting the infections in humans."
"It is preferable to study new combined vaccines in comparison with the individual
antigens in animals to determine if any augmentation or diminution of response occurs."
"It is recommended that these preclinical studies are also designed to evaluate the adjuvant effects on the immune response when a relevant animal model is available."
https://www.ema.europa.eu/en/documents/scientific-guideline/note-guidance-preclinical-pharmacological-toxicological-testing-vaccines_en.pdf
and current effective version of the document
Questions and answers on the withdrawal of the CPMP Note for guidance on preclinical pharmacological and toxicological testing of vaccines (CPMP/SWP/465)
" Due to the complex mode of action of vaccines, animal studies are normally the only the possibility to address the pharmacological and toxicological activity of a vaccine. Since vaccines in most cases are given to large numbers of healthy individuals, there is a need for a solid nonclinical safety evaluation "
" animal experiments are considered pivotal for the development of vaccines. Nevertheless, all efforts should be taken to limit the number of animals and the impact on well-being in animals. "
https://www.ema.europa.eu/en/documents/other/questions-answers-withdrawal-cpmp-note-guidance-preclinical-pharmacological-toxicological-testing/swp/465_en.pdf
Please check other documents by FDA
Considerations for Developmental Toxicity Studies for Preventive and Therapeutic Vaccines for Infectious Disease Indications
https://www.fda.gov/files/vaccines,%20blood%20%26%20biologics/published/Guidance-for-Industry--Considerations-for-Developmental-Toxicity-Studies-for-Preventive-and-Therapeutic-Vaccines-for-Infectious-Disease-Indications.pdf
GUIDANCE FOR INDUSTRY FOR THE EVALUATION OF COMBINATION VACCINES FOR PREVENTABLE DISEASES:
" B. Animal Immunogenicity It is recommended that new combinations be studied for the appropriate immunogenicity parameters in an animal model, if available, before the initiation of studies of human clinical trials. The response to each of the antigens in the vaccine should be assessed as well as the quality of the response. This evaluation may include a characterization of antibody class, avidity, affinity, half-life, or function, e.g. examining the ability to neutralize the target agent or toxin. It is preferable to study the new combination in comparison to the individual antigens in animals to determine if any augmentation or diminution of response occurs. Interference between live vaccine strains also may be studied in animal immunogenicity studies.
C. Animal Challenge Studies Protection studies are recommended using an animal model, wherever one is available, for new vaccines or combination vaccines with a new antigen that has not been previously studied in humans. Protection should be demonstrated upon challenge with a virulent strain(s) of each organism against which the vaccine is intended to protect. The study should be conducted with statistically and scientifically valid procedures for verifying the results and these should be described. It is advisable to perform such studies early in the product development cycle. "
https://www.fda.gov/media/77191/download
Guidance for Industry General Principles for the Development of Vaccines to Protect Against Global Infectious Diseases
https://www.fda.gov/files/vaccines,%20blood%20&%20biologics/published/Guidance-for-Industry--General-Principles-for-the-Development-of-Vaccines-to-Protect-Against-Global-Infectious-Diseases.pdf
Its useful set of the document but please remember that "case by case" philosophy is always present and its important aspect of interaction with agency.
Moreover models should be "apropriate", "relevant", "well controled" .... and finaly
" The use of the word should in Agency guidances means that something is suggested or recommended, but not required. " :-) (https://www.fda.gov/media/78034/download)
In short, all responsibility lies solely with you. The Authority can only advise you.
Best regards :-)
Tomasz
Your question is too general. The measurement of the immune response depend on the type of response you want to evaluate, the disorder you are going to search.
There are many ways to detection and meaure of immune response in labs animals such as Rabbit ,mice, Rats , Guinea pig, Hamester etc. Theses methods are :
1- evaluate of humoral immune response such as determination immunoglobulins in experimental animal after injection microbials agents . 2- measuring cellular immune response by using Skin test and Cytokines as well as you can use other way by going to search to investigate of experimental models to do your ideas.
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