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Saquinavir was the first protease inhibitor to be approved for treatment of HIV infection. It is a peptidomimetic agent designed by computer modeling as a molecule that fits into the active site of the HIV protease enzyme. Such drugs inhibit the viral protease that is required at the late stage of the replicative cycle to cleave the viral gag and gag-pol polypeptide precursors to form the mature virion core and activate the reverse transcriptase that will be used in the next round of infection. Inhibition of the protease yields noninfectious virus particles. Protease inhibitors include indinavir and ritonavir and others not listed here. FUSION INHIBITOR Fuzeon is a large peptide that blocks the virus and cellular membrane fusion step involved in entry of HIV-1 into cells. OTHER TYPES OF ANTIVIRAL AGENTS A number of other types of compounds have been shown to possess some antiviral activity under certain conditions. Amantadine and Rimantadine These synthetic amines specifically inhibit influenza A viruses by blocking viral uncoating. They must be administered prophylactically to have a significant protective effect. Foscarnet (Phosphonoformic Acid, PFA) Foscarnet, an organic analog of inorganic pyrophosphate, selectively inhibits viral DNA polymerases and reverse transcriptases at the pyrophosphatebinding site. Methisazone Methisazone is of historical interest as an inhibitor of poxviruses. It was the first antiviral agent to be described and contributed to the campaign to eradicate smallpox. It blocked a late stage in viral replication, resulting in the formation of immature, noninfectious virus particles.

Interferons
Interferons (IFNs) are host-coded proteins that are members of the large cytokine family and which inhibit viral replication. They are produced very quickly (within hours) in response to viral infection or other inducers and are one of the body's first responders in the defense against viral infection. Interferon was the first cytokine to be recognized. Interferons are central to the innate antiviral immune response. They also modulate humoral and cellular immunity and have broad cell growth regulatory activities, but the focus here will be on their antiviral effects. PROPERTIES OF INTERFERONS There are multiple species of interferons that fall into three general groups, designated IFN- , IFN- , and IFN- (Table 307). Both IFN- and IFN-b are considered type I or viral IFNs, whereas IFN- is type II or immune IFN. The IFN- family is large, being coded by at least 20 genes in the human genome; the IFN- and IFN- families are coded by one gene each. The three gene families have diverged so that the coding sequences now are not closely related.

Table 307. Properties of Human Interferons.

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Type Property Current nomenclature Former designation Type designation Number of genes that code for family Principal cell source Inducing agent Stability at pH 2.0 Glycosylated Introns in genes Homology with IFNChromosomal location of genes Alpha IFNLeukocyte Type I 20 Beta IFNFibroblast Type I 1 Gamma IFNImmune interferon Type II 1

Most cell types Most cell types Lymphocytes Viruses; dsRNA Viruses; dsRNA Mitogens Stable No No 8095% 9 Stable Yes No 30% 9 166 IFNAR 21 Labile Yes Yes < 10% 12 143 IFNGR 6

Size of secreted protein (number of amino acids) 165 IFN receptor Chromosomal location of IFN receptor genes IFNAR 21

The different interferons are similar in size, but the three classes are antigenically distinct. IFN- and IFN- are resistant to low pH. IFN- and IFN- are glycosylated, but the sugars are not necessary for biologic activity, so cloned interferons produced in bacteria are biologically active. Dendritic cells are potent interferon producers; under the same virus challenge conditions, dendritic cells can secrete up to 1000x more interferon than fibroblasts. SYNTHESIS OF INTERFERONS Interferons are produced by all vertebrate species. Normal cells do not generally synthesize interferon until they are induced to do so. Infection with viruses is a potent insult leading to induction; RNA viruses are stronger inducers of interferon than DNA viruses. Interferons also can be induced by double-stranded RNA, bacterial endotoxin, and small molecules such as tilorone. IFN- is not produced in response to most viruses but is induced by mitogen stimulation. The different classes of interferon are produced by different cell types. IFN- and IFN- are synthesized by many cell types, but IFN- is produced mainly by lymphocytes, especially T cells and natural killer (NK) cells. Dendritic cells are potent interferon producers; under the same virus challenge conditions, dendritic cells can secrete up to 1000x more interferon than fibroblasts. ANTIVIRAL ACTIVITY AND OTHER BIOLOGIC EFFECTS

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Interferons were first recognized by their ability to interfere with viral infection in cultured cells. Interferons are detectable soon after viral infection in intact animals, and viral production then decreases (Figure 307). Antibody does not appear in the blood of the animal until several days after viral production has abated. This temporal relationship suggests that interferon plays a primary role in the nonspecific defense of the host against viral infections. This conclusion is also supported by observations that agammaglobulinemic individuals usually recover from primary viral infections about as well as normal people.

Figure 307.

Illustration of kinetics of interferon and antibody synthesis after respiratory viral infection. The temporal relationships suggest that interferons are involved in the host's early defense system against viral infections.

Interferon does not protect the virus-infected cell that produces it, and interferon itself is not the antiviral agent. Rather, interferon moves to other cells where it induces an antiviral state by prompting the synthesis of other proteins that actually inhibit viral replication. Interferon molecules bind to specific cell surface receptors on target cells. IFN- and IFN- have the same receptor, whereas IFN- recognizes a different receptor. Receptor binding triggers tyrosine phosphorylation and activation of transcription factors (STAT proteins) in the cytoplasm, which then translocate into the nucleus and mediate transcription of interferon-inducible genes (which occurs within minutes after interferon binding). This results in the synthesis of several enzymes believed to be instrumental in the development of the antiviral state. Several pathways appear to be involved, including the following: (1) a dsRNA-dependent

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protein kinase, PKR, which phosphorylates and inactivates cellular initiation factor eIF-2 and thus prevents formation of the initiation complex needed for viral protein synthesis; (2) an oligonucleotide synthetase, 2-5A synthetase, which activates a cellular endonuclease, RNase L, which in turn degrades mRNA; (3) a phosphodiesterase, which inhibits peptide chain elongation; and (4) nitric oxide synthetase, which is induced by IFN- in macrophages. These explanations, however, fail to reveal why the antiviral state acts selectively against viral mRNAs and not cellular mRNAs. Other steps in viral replication may also be inhibited by interferon. Interferons are almost always host species-specific in function but are not specific for a given virus. The replication of a wide variety of DNA and RNA viruses can be inhibited. When interferon is added to cells prior to infection, there is marked inhibition of viral replication but nearly normal cell function. Interferons are extremely potent, so that very small amounts are required for function. It has been estimated that fewer than 50 molecules of interferon per cell are sufficient to induce the antiviral state. VIRUS MECHANISMS TO COUNTERACT INTERFERON Viruses display different mechanisms that block the inhibitory activities of interferons on virus replication, processes necessary to surmount this line of host defense. Examples include specific viral proteins may block induction of expression of interferon (herpesvirus, papillomavirus, filovirus, hepatitis C virus, rotavirus); may block the activation of the key PKR protein kinase (adenovirus, herpesviruses); may activate a cellular inhibitor of PKR (influenza, poliovirus); may block interferon-induced signal transduction (adenovirus, herpesviruses, hepatitis B virus); or may neutralize IFN- by acting as a soluble interferon receptor (myxoma virus). CLINICAL STUDIES It was originally hoped that interferons might be the answer to prevention of many viral diseases, such as respiratory infections in which many different viruses may be involved. However, their use turns out to be impractical because for them to be effective, high doses must be given prior to virus exposure or early in infection before the appearance of clinical signs of disease. Recombinant IFN- is beneficial in controlling hepatitis B and hepatitis C viral infections of the liver (Chapter 35), though relapse after cessation of treatment is common. Topical interferon in the eye may suppress herpetic keratitis and accelerate healing. Several interferon preparations are approved for clinical use. Interferons cause many side effects, most commonly systemic and hematologic.

Viral Vaccines
The purpose of viral vaccines is to utilize the immune response of the host to prevent viral disease. Several vaccines have proved to be remarkably effective at reducing the annual incidence of viral disease (Figure 308). Vaccination is the most cost-effective method of prevention of serious viral infections.

Figure 308.

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