Host Defense: Vaccine, Interferon, Antiviral Drugs

1. Host Defense: Vaccine, Interferon, Antiviral Drugs

2. Combating Infectious Diseases • Prevention by Public Health: • Epidemiology • Control • Education • Vaccines • Treatment by Medical Science: • Supportive • Corrective • Drugs • Which is preferred? • More cost effective?

3. Vaccination and Immunity • The process of using inactivated or attenuated pathogen (or portion) to induce immune response in individual prior to exposure of pathogen • Humoral Immunity (antibody, complement) • Cell-Mediated Immunity (macrophage, T lymphocytes)

4. Killed-Virus Vaccine • Virulent wild type virus • Made noninfectious by chemical treatment (formalin, betapropiolactone) • Salk inactivated poliovirus (IPV)

5. Salk Inactivated Poliovirus Vaccine (IPV) • Efficacy – High • Route – Intramuscular injection • Dose / Cost – High • Antibody – IgG • CMI – Poor • Heat labile – No • Interference by other viruses – No • Revert to virulence - No • Use in immune compromised patient - Yes

6. Live-Virus Vaccine • Avirulent – mild or no disease • Vaccinia virus (cowpox?) • Attenuated – mutated wild type virus no longer pathogenic • Sabin oral poliovirus (OPV), not able to get into CNS

7. Live - Sabin Oral Poliovirus Vaccine (OPV) • Efficacy – High • Route – Oral (natural transmission, infection) • Dose / Cost – Low • Antibody – IgG, IgA • CMI – Good • Heat labile – Yes • Interference by other viruses – Yes (other Enterovirus infections) • Revert to virulence – Yes (rare, 1/300,000 doses) • Use in immune compromised patient - No

8. Subunit Vaccine • A viral protein or glycoprotein that elicits protective immunity • May be isolated from virion • Influenza vaccine (HA, NA), grow virus in chick embryo, isolate viral proteins • May be made in recombinant expression vector • HBV vaccine (HBsAg glycoprotein), grow vector in yeast cells

9. Experimental Recombinant Vaccine • Recombinant Virus – introduce viral gene into another virus by gene recombination i.e., HIV gp120 gene into Vaccinia virus

10. Experimental DNA Vaccine • Direct intramuscular injection of viral DNA gene • Gene expression and viral protein synthesis

11. Experimental Plant Vaccine • Insert viral gene into plants (potato, tomato, banana) • Oral vaccine ingested, elicit immune response

12. Experimental HIV Vaccine: Neutralizing (Blocking) Ab

15. Interferon: Cell Antiviral Defense • 1957 – Isaacs & Lindenmann studying influenza virus in chick embryo • Identify soluble substance in allantoic fluid that blocks influenza virus infection in chick embryo • Inhibitor termed “interferon” • Now known to be a cell cytokine (signal transducer)

17. Properties of Interferon • Species specific • Induced soon (hours) following virus infection • Inhibits various viruses • Acts indirectly – induces antiviral effector molecules (AVEM) • Inhibits cell division • Activates immune system • Induces fever

18. Classes of Interferon (IFN) • IFN-α made by leukocytes • IFN-β made by fibroblasts • IFN-γ made by T lymphocytes

19. Inducers of Interferon • RNA viruses • dsRNA (synthetic poly I-C) • Intracellular MO’s (protozoa, rickettsia) • Bacterial endotoxin

20. Activity of Interferon • IFN made and secreted from virus infected cell • Binds to specific cell receptor that triggers a signal transduction cascade • Prepares the cell to combat virus infection by expressing threeproenzymes termed antiviral effector molecules (AVEM)

21. Antiviral State In Cell (AVEM): Three Proenzymes • 1) 2’,5’-oligoA synthetase (OAS) • 2) dsRNA-dependent protein kinase (PKR) • 3) RNAse-L • Proenzymes, when activated, inhibit viral replication

22. OAS-Activated Cell • Viral infection (dsRNA) activates the proenzyme OAS • 2’,5’-oligoA is maded • 2’,5’-oligoA activates proenzyme RNAse-L, which cleaves mRNA • No mRNA, translation STOPPED

23. PKR-Activated Cell • Viral infection (dsRNA) activates the proenzyme PKR • Ribosome translation initiation factor (eIF2) is phosphorylated by activated PKR • No ribosome initiation complex • No ribosome formation, translation & protein synthesis STOPPED

25. Clinical Use of Interferon: Antiviral • Available in adequate amount since 1980s by recombinant DNA technology • Life threatening infections – rabies, hemorrhagic fevers, encephalitis due to arboviruses • Persistent chronic infections - hepatitis B hepatitis C, papilloma, herpesvirus, HIV

26. Clinical Use of Interferon: Anti-Cancer • Leukemia / lymphoma • Multiple myeloma • GI tumors • Kidney, Bladder cancer • Mesothelioma (lung cancer) • AIDS-related Kaposi’s sarcoma (Human Herpes Virus-8)

27. Interferon: Toxic Side-Effects • Fever • Chills • Headache • Nausea & vomiting • Anorexia • Rash, dry skin • CNS – depression, dizziness, confusion

28. Current Use of Interferon • Combination with other antivirals • Lipid formulation • Not as readily degraded • Better delivery to target cell • Less toxicity

29. Antiviral Drugs • Virus & host cell replication closely related • Antiviral requires • Low toxicity for host cell • Selective toxicity for virus

30. Testing For Safety of Antiviral • Laboratory: Cell culture & animal testing • Clinical Trials in Humans: • Phase 1 test for safety • Phase 2 test for efficacy • Phase 3 test for efficacy in a large population

31. Therapeutic Index (T.I.) for Antiviral • Measure tolerated dose (not toxic/lethal) • Measure curative dose (decrease disease or death) • Therapeutic Index = tolerated dose / curative dose • T. I. = 1/1 = ≤ 1 (toxic) • T. I. = 1/<1 = >1 (safe)

32. Antivirals: Nucleoside Analogues

33. Acyclovir & Herpesvirus • Nucleoside analogue • Target viral enzymes in DNA synthesis • Herpesvirus thymidine kinase activates acyclovir • Viral DNA polymerase incorporates acyclovir into growing DNA chain • DNA synthesis STOPPED due to chain termination

34. Rimantadine (Flumadine) - Influenza A Virus • Synthetic cyclic amine compound • Stops virus Uncoating/Maturation • Prevents acid pH required for membrane fusion, by blocking viral M2 (ion channel) protein • CNS side effects: dizziness, ataxia, insomnia, convulsions

35. Neuraminidase Inhibitor (Zanamvir) - Influenza Virus A, B • Drug blocks neuramindase (NA), virus release STOPPED • During budding, virus hemagglutinin (HA) readily binds to neuraminic acid (sialic acid) • NA cleaves sialic acid to insure release of virus from cell • NA inhibitors are sialic acid analogues • Competitive inhibitor of virus NA enzyme

36. Neuraminidase Inhibitors • Oseltamvir (Tamiflu) – oral, pill • Zanamivir (Relenza) – nasal, inhaler • Need to be given within 48 hr. infection

37. Antivirals Against HIV • Reverse transcriptase (RT) nucleoside inhibitors • Azidothymidine • RT non-nucleoside inhibitors • ddC, ddI • Protease inhibitors • Saquinavir • Entry inhibitors • Enfuvirtide • DNA Integrase inhibitor • Raltegravir

38. Protease Inhibitor • This picture shows the HIV protease (purple and green) complexed with the inhibitor (spacefill). This prevents the substrate from reacting with the protease and thus, the polypeptides are not cleaved.

39. Current Treatment for HIV • “Cocktail” – combination of four or more drugs • Decease virus load (# progeny virus) in blood • Clinical “latency” • Prolong life • High cost • HIV reservoir in lymphoid tissue (replicate in low #)

40. Experimental Antiviral: Antisense Oligonucleotide • ssRNA or ssDNA complimentary to viral mRNA made • Will combine with viral mRNA and prevent translation • Essential viral protein STOPPED and virus replication stopped

41. Reading & Questions • Chapter 8: Strategies to Protect Against and Combat Viral Infections (omit Question 1)

42. QUESTIONS???

43. Class Discussion – Lecture 4 • 1. How is virus replication stopped in interferon-treated cells? • 2. Is a cell treated with interferon able to make interferon if it is infected by a virus? • 3. At what steps are current antivirals effective in stopping virus replication? • 4. Why is the antiviral drug, acyclovir, more selective for herpesvirus? How does it stop herpesvirus replication?