Host Defense: Host Defense: Vaccine, Vaccine,

Interferon, Interferon, Antiviral DrugsAntiviral Drugs

Host Defense: Host Defense: Vaccine, Vaccine,

Interferon, Interferon, Antiviral DrugsAntiviral Drugs

Combating Infectious Diseases

• Prevention by Public Health:– Epidemiology– Control– Education– Vaccines

• Treatment by Medical Science:– Supportive– Corrective– Drugs

• Which is preferred?• More cost effective?

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)

Killed-Virus Vaccine• Virulent wild type virus• Made noninfectious by chemical

treatment (formalin, betapropiolactone)

• Salk inactivated poliovirus (IPV)

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

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

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

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

Experimental Recombinant Vaccine

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

Experimental DNA Vaccine

• Direct intramuscular injection of viral DNA gene

• Gene expression and viral protein synthesis

Experimental Plant Vaccine

• Insert viral gene into plants (potato, tomato, banana)

• Oral vaccine ingested, elicit immune response

Experimental HIV Vaccine: Neutralizing (Blocking) Ab

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)

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

Classes of Interferon (IFN)

• IFN-α made by leukocytes• IFN-β made by fibroblasts• IFN-γ made by T lymphocytes

Inducers of Interferon

• RNA viruses• dsRNA (synthetic

poly I-C)• Intracellular MO’s

(protozoa, rickettsia)

• Bacterial endotoxin

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 three proenzymes termed antiviral effector molecules (AVEM)

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

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

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

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

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)

Interferon: Toxic Side-Effects

• Fever• Chills• Headache• Nausea & vomiting• Anorexia• Rash, dry skin• CNS – depression, dizziness, confusion

Current Use of Interferon

• Combination with other antivirals• Lipid formulation

– Not as readily degraded– Better delivery to target cell– Less toxicity

Antiviral Drugs• Virus & host cell replication closely

related• Antiviral requires

– Low toxicity for host cell– Selective toxicity for virus

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

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)

Antivirals: Nucleoside Analogues

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

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

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

Neuraminidase Inhibitors

• Oseltamvir (Tamiflu) – oral, pill• Zanamivir (Relenza) – nasal,

inhaler• Need to be given within 48 hr.

infection

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

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.

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 #)

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

Reading & Questions

• Chapter 8: Strategies to Protect Against and Combat Viral Infections (omit Question 1)

QUESTIONS???

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?