Screening methods of immunomodulatory drugs
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The document summarizes the immune system and how it distinguishes self from nonself through innate and adaptive immunity. It then discusses immunomodulators which can suppress or stimulate the immune response, dividing them into immunosuppressants and immunostimulants. Several screening methods for evaluating immunomodulatory activity are also presented, including acute systemic anaphylaxis in rats, anti-anaphylactic activity, passive cutaneous anaphylaxis, Arthus type hypersensitivity, delayed type hypersensitivity, and carbon clearance tests for phagocytic response measurement. Modifications to these screening methods are also noted.
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Screening methods of immunomodulatory drugs
- 1. PRESENTED BYPRESENTED BY:-:- Prafulla C TiwariPrafulla C Tiwari M.Pharm (2M.Pharm (2ndnd sem)sem) PharmacologyPharmacology UNDER THE GUIDANCE OF:- MR. RUPESH GAUTAM (Lecturer) Department Of Pharmacology
- 2. The immune system evolved to discriminate self from nonself. Multicellular organism shows immune response by destroying infectious invaders (microbes) while leaving normal cells intact. Immunity system shows two types of responses that are- 1.Innate immunity 2.Adaptive immunity
- 3. 1.Innate, or natural immunity-It is primitive, does not require priming, and have relatively low affinity, but is broadly reactive. The major effectors of innate immunity are granulocytes, monocytes/macrophages, natural killer cells and mast cells. 2. Adaptive, or learned, immunity-It is antigen specific, depends upon antigen exposure and have very high affinity. The major effectors of adaptive immunity are B and T lymphocytes. B lymphocytes make antibodies; T lymphocytes function as helper, cytolytic cells.
- 4. Immunomodulators are the agents that modulates the immune system by suppress or stimulate the immune response.So, these are also divided into two parts that are- 1.Immunosuppressants & 2.Immunostimulants
- 5. 1.IMMUNOSUPPRESSANTS- Immunosuppressive drugs are used to dampen the immune response in organ transplantation and autoimmune disease. (a)Specific T-cell inhibitors-Cyclosporine,Tacrolimus (b)Cytotoxic drugs-Azathioprine, Cyclophosphamide, Methotrexate, Chlorambucil, Mycophenolate mofetil(MMF) (c)Glucocorticoids-Prednisolone (d)Antibodies-Muromonab CD3, Antithymocyte globulin(ATG), Rho(D) immunoglobulin
- 6. 2.IMMUNOSTIMULANT- These drugs are used to stimulate the immune response in case of immunodeficiency. (a) Levamisole (ergamisole) (b) Thalidomide (thalomid) (c) Bacillus Calmette-Guerin (BCG) (d) Interferons- gamma1b & beta1a (e) Interleukin-2
- 7. Screening method Acute systemic anaphylaxis in rats Anti-anaphylactic activity (schultz-dale reaction) Passive cutaneous anaphylaxis Arthus type immediate hypersensitivity Delayed type hypersensitivity Reversed passive arthus reaction Adjuvant arthritis in rats
- 8. Acute systemic anaphylaxis in rats Strain: 10-20 Female Sprague-Dawley rat(120g) Firstly immunized by i.m. injection of 10mg/kg highly purified ovalbumin. Simultaneously 1ml of Bordetella pertusis suspension injected intraperitoneally. After 11 days animals injected with i.v. injection of 25mg/kg highly purified ovalbumin 18hr prior to challenge; test dexamethasone 1-10mg/kg s.c. control-vehicle
- 9. Evaluation- after treatment compared treated and control group for their shock symptoms and mortality counted. Statistical calculation is performed using the chi-square test.
- 10. Modification- By Davis and Evans(1973)-This experiment have also been performed in guinea pigs and in mice. Anaphylactic bronchospasm can be measured in isolated guinea pig lungs. By Ufkes and Ottenhof(1984)- studied on Brown- Norway rats Given a suspension of trinitrophenyl haptenized ovalbumin together with AlPO4. Bronchial and cardiovascular function were studied after treatment with antiallergic agents and antigen challenge.
- 11. Elwood et al.(1992)- studied on Brown-Norway rats. They studied the effect of dexamethasone and cyclosporin A on hyperresponsiveness and inflammatory cell responses.
- 12. Anti-anaphylactic activity (Schultz-Dale reaction) Animal-guinea pig of both sex Body wt.-300-350g Sensitized with alum precipitated egg albumin. Give injection of 0.125ml of egg albumin in each foot pad and 0.5ml subcutaneously.
- 13. After 4 weeks animals are killed and the ileum is dissected out. About 2-3cm long are mounted in an organ bath containing Tyrode solution at 370 C. Contractility of the ileum strips is tested by adding standardin one organ bath and test compound in another organ bath.
- 14. Biochemical estimation- Carbon clearance test- for phagocytic response Phagocytic index=K(sample)/K(control) Hemagglutinating antibody titer
- 15. Evalution- The contractions are recorded by a polygraph. Modification- This method modified by testing histamine release in the lung after challenging with egg albumin. Koppel et al.(1981) developed a method to induce contraction of mouse trachea by antigen. Omote et al.(1994) modified method by using sensitized guinea pigs.
- 16. Passive cutaneous anaphylaxis Animal-male rats Body wt.-100g Principle- Formation of antigen-antibody complex induces the release of mediator from mast cells.This results increase in permeability of the vessel walls and leakage of plasma.
- 17. Procedure- Antiserum are injected intradermally in to the shaved dorsal skin of rats. After 24hr each animal is challenged with the intravenous administration of 0.1ml of 2.5% Evans blue dye containing 25mg/ml of egg albumin. Test compound is also administer along with the antigen. After 30 min.,animals are sacrificed. Amount of Evans blue dye that leaked at the site of reaction is extracted and determined colorimetrically at 620milimicron wavelength.
- 18. Evaluation- Amount of Evans blue that extracted from passive cutaneous anaphylactic reaction of control group is compared with the treated group. Modification- Goose and Blair(1969)- used Bordetella pertusis as antigen in passive cutaneous anaphylaxis experiments in the rat. Patterson et al.(1971)- tested passive cutaneous reactivity to antihuman IgE in rhesus monkey.
- 19. Arthus type immediate hypersensitivity Animal-rats of both sex Strain-Wistar or Sprague-Dawley Body wt.-220-280g Principle- Antigen-antibody induced reaction leading to an inflammatory factors that characterised by edeme,hamorrhage and vasculitis.
- 20. Procedure- Seven days prior to experiment rats are sensitized by i.m. administration of 0.5ml of the ovalalbumin suspension. 1st group(treated group)- 1hr prior test compound are administered and challenged with 0.1ml of ovalalbumin in left hind paw 2nd group(positive control)-sensitized animals treated with solvent alone. 3rd group(negative control)-nonsensitized animals treated with solvent.
- 21. Evalution- The change in footpad thickness is expressed as percent change from the vehicle control group. Thickness can be measured by calipers. Modification- Omote et al.(1994)-sheep red blood cell suspension used for immunization.
- 22. Delayed type hypersensitivity Principle- Delayed type hypersensitivity is a reaction of cell mediated immunity and becomes visible after 16- 24hr. Procedure- 7days prior,rats are sensitised by i.m. administration of 0.5ml ovalbumin. After 7 days again 0.1ml of 0.04% of ovalbumin injected in the left hind paw.
- 23. Footpad thickness is measured immediately and 24hr after of administration. Modification- Mizukoshi et al.(1994)-They use sheep red blood cells for immunization.
- 24. References- Vogel.H.G. , second edition, Analgesics, page no:797-801. Bigonia P. & Rana A.C. Immunomodulatory activity of EUPHORBIA NERIIFOLIA leaf alcoholic extract on rats Indian drugs, feb.2008