Technical guidelines for quality control of human monoclonal antibodies!
Distribution, half-life, etc.) can be obtained from animal models; when there is no suitable animal model, animal pharmacokinetics may not be considered, and pharmacokinetic monitoring during clinical trials should be strengthened.
(2) The following aspects can guide the selection of animal species for pharmacokinetic and pharmacodynamic tests:
① It is best to choose animal models that have cross-reactivity with humans or have the same target antigen for testing. For unconjugated monoclonal antibodies that are only against human antigens or foreign antigens (bacteria, viruses, etc.) that are not expressed in animal models, there is no need to use animals lacking the target antigen for testing, etc.
② When antigen binding data indicate that primates are the most relevant species, it is appropriate to use non-human primates for testing of unconjugated monoclonal antibodies.
③The possibility of using normal rodent and murine xenograft models to accurately predict the pharmacokinetic behavior of monoclonal antibodies in humans should be rigorously evaluated. Xenograft models are more useful for evaluating the ability of monoclonal antibodies to bind to tumors in humans.
(3) Mouse-derived antibodies are non-immunogenic substances in mice, but are immunogenic in humans. This makes it difficult to extrapolate the repeated dose results obtained in mice to the repeated doses intended for use in humans. Problems arise with the use of fully human, chimeric or "humanized" monoclonal antibodies, in which case repeated dose studies in rodents are of little interest.
4. Clinical in vivo studies using immunoconjugates
(1) The stability of immunoconjugates should be tested in vivo
① The pharmacokinetics and tissue distribution of each component of the immunoconjugate in animals should be determined and compared with the distribution of unconjugated antibodies.
② The target tissues of different components and the potential toxicity they can cause should be confirmed.
(2) Since the immunoconjugate may be degraded or the activity of the action site is not the result of the binding of the monoclonal antibody to the target antigen, animal toxicity experiments should be conducted on immunoconjugates containing radionuclides, toxins or drugs, although the animals are not Target antigen is present. Depending on the nature of the immunoconjugate components and the stability of their conjugation, it may be reasonable to test the components separately. The toxicity profile, occurrence and severity of side effects of each component should be fully described. The results obtained should correlate closely with the conjugate stability testing. If possible, immunoconjugate testing should be performed in animals with relevant target antigens or disease models, or in rodents if no target antigen-positive animals exist. Toxicity tests for free toxins or nuclides can be conducted in different types of animals.
(3) For immunoconjugates of radionuclides:
① Animal biodistribution data can be used to evaluate initial human doses.
② When possible, animal models expressing the target antigen are more likely to find tissues with "reduced" antigen or with tissues that exhibit unexpected antigens in terms of biodistribution and/or toxicity.
③ The allogeneic transplantation model can solve the problem of tissue localization and antigen non-specific radioimmunoconjugate distribution, but it is not helpful in determining the scope of general tissue cross-reaction.
④ An appropriate number of animals should be studied to evaluate the radioactive dose with an acceptable coefficient of variation (usually less than 20%).
⑤ Appropriate values for metabolism using total radioactivity and determination of time points from early to late clearance periods should be fully calculated.
⑥ The in vitro stability of radioimmunoconjugates should be tested by incubation in serum or plasma. Methods should be developed to assess the percentage of radioactivity present in each component of free epitopes, conjugated monoclonal antibodies, and labeled non-monoclonal antibodies.
Note: The requirements for topical use and preparation of monoclonal antibodies for in vitro diagnostic kits are implemented in accordance with the above-mentioned relevant sections, but the requirements for production conditions, mouse cleanliness levels, antibody purity and safety tests can be reduced or reduced according to actual needs.
Appendix: Detection of mouse-derived viruses
1. Detected virus
Mouse-derived monoclonal antibody products for human use may contain potentially contaminating viruses, as shown in the table below
(1) Hemorrhagic fever virus, affected animals are rats and mice
(2) Lymphocyte choroids from meningitis virus (LCMV), the affected animals are rats
(3) Reovirus type 3 (REO), affected animals are rats and mice
(4) Rat rotavirus, the affected animals are rats
(5) Sendai virus, affected animals are rats and mice
(6) Podiatry virus, affected animals are mice
(7) KITHAM's rat virus (KRV), the affected animals are rats
(8) Mouse adenovirus (MAV), the affected animals are mice
(9) Pneumonia mouse virus (PVM), the affected animals are mice
(10) Retrovirus, affected animals are rats and mice
(11) TOOLAN virus (HI), the affected animals are rats
The first five viruses belong to Group I, which are viruses that can infect humans and primates, and the last six belong to Group II, which have no signs of infecting humans yet, but are potentially dangerous to humans and can be cultured in vitro. Replicating in cells derived from apes and monkeys, these viruses should be detected as a priority.
2. Sample
Including cell lines, ascites, animal serum, monoclonal antibody semi-finished products and finished products, etc., should be pretreated with appropriate methods.
3. Test method
Including cell tests, animal antibody production tests, chicken embryo infection tests, etc.
(1) Cell experiment
The cells and culture supernatant were inoculated into human 2BS and Vero cells respectively, cultured, and then passaged for two generations before making slices and using indirect immunofluorescence method to detect viral antigens.
(2) Animal antibody production test
Samples were inoculated into 10 suckling mice within 24 hours after birth (i.m); 20 adult mice between 15 and 20 g (i.mti.p: 10 samples were inoculated and 10 were used as controls); 20 mice (i.c : 10 animals were inoculated with samples and 10 animals were used as controls). The 4-week survival rate should be above 80%, and ELISA or other appropriate methods should be used to detect viral antibodies in the serum.
(3) Chicken embryo infection test
Chicken embryo inoculation is used for examination. Chicken embryos aged 9 to 11 days can be used to inject samples into the chorioallantoic membrane, allantoic cavity and yolk sac of 10 chicken embryos. Chicken embryos cannot be examined until at least 5 days after vaccination. And use red blood cells from guinea pigs, chickens or other poultry to check whether the allantoic fluid contains hemagglutinin.
4. Detection range
(1) Cell bank - inspection of each batch
(2) Cell bank - inspection of each batch
(3) Cell bank-inspection of each batch
(4) Rat library - regular spot checks
(5) Ascites - Check every batch
(6) Cell culture preparation-batch inspection
(7) Supernatant of monoclonal antibody
