Principles of Pharmacology The Pathophysiologic Basis of Drug Therapy

Pharmacology of Viral Infections Chapter 37 Pharmacology of Viral Infections

Viral Life Cycle and Pharmacologic Intervention Figure 37-1 The viral life cycle can be divided into a sequence of individual stages, each of which is a potential site for pharmacologic intervention. Shown is a generic replication cycle of viruses in cells, alongside which are listed the names of drug classes and examples of individual agents that block each stage. Many of the currently approved antiviral agents are nucleoside analogues that target genome replication, typically by inhibiting viral DNA polymerase or reverse transcriptase. Several other drug classes target other stages in the viral life cycle, including attachment and entry, uncoating, assembly and maturation, and egress and release. It should be noted that the details of viral replication differ for each type of virus, often presenting unique targets for pharmacologic intervention and drug development.

Life Cycle of HIV Figure 37-2 HIV is a retrovirus that infects CD4+ cells. 1. Virus attachment is dependent on binding interactions between viral gp41 and gp120 proteins and host cell CD4 and certain chemokine receptors. 2. Fusion of the viral membrane (envelope) with the host cell plasma membrane allows the HIV genome complexed with certain virion proteins to enter the host cell. 3. Uncoating permits the single-stranded RNA (ssRNA) HIV genome to be copied by reverse transcriptase into double-stranded DNA. 4. The HIV DNA is integrated into the host cell genome, in a reaction that depends on HIV-encoded integrase. 5. Gene transcription and posttranscriptional processing by host cell enzymes produce genomic HIV RNA and viral mRNA. 6. The viral mRNA is translated into proteins on host cell ribosomes. 7. The proteins assemble into immature virions that bud from the host cell membrane. 8. The virions undergo proteolytic cleavage, maturing into fully infective virions. Currently approved anti-HIV drugs target viral attachment and fusion, reverse transcription, integration, and maturation. The development of drug resistance can be significantly retarded by using combinations of drugs that target a single stage (e.g., two or more inhibitors of reverse transcription) or more than one stage in the HIV life cycle (e.g., reverse transcriptase inhibitors and protease inhibitors).

Model for HIV gp41-Mediated Fusion and Maraviroc and Enfuvirtide (T-20) Action Figure 37-3 A. HIV glycoproteins exist in trimeric form in the viral membrane (envelope). Each gp120 molecule is depicted as a ball attached noncovalently to gp41. B. The binding of gp120 to CD4 and certain chemokine receptors in the host cell plasma membrane causes a conformational change in gp41 that exposes the fusion peptide, heptad-repeat region 1 (HR1) and heptad-repeat region 2 (HR2). The fusion peptide inserts into the host cell plasma membrane. C. gp41 undergoes further conformational changes, characterized mainly by unfolding and refolding of the HR2 repeats. D. Completed refolding of the HR regions creates a hemifusion stalk, in which the outer leaflets of the viral and host cell membranes are fused. E. Formation of a complete fusion pore allows viral entry into the host cell. F. Enfuvirtide (T-20) is a synthetic peptide drug that mimics HR2, binds to HR1, and prevents the HR2–HR1 interaction (dashed arrow). Therefore, the drug traps the virus–host cell interaction at the attachment stage, preventing membrane fusion and viral entry. G. Maraviroc is a small-molecule antagonist of the CCR5 chemokine receptor; the drug blocks cellular infection of HIV strains that use CCR5 for attachment and entry (dashed arrow). The structure of maraviroc is shown.

Uncoating of Influenza Virus and Effect of Amantadine and Rimantadine Figure 37-4 The structures of the adamantanes, amantadine and rimantadine, are shown. Influenza virus enters host cells by receptor-mediated endocytosis (not shown) and is contained within an early endosome. The early endosome contains an H+-ATPase that acidifies the endosome by pumping protons from the cytosol into the endosome. A low pH-dependent conformational change in the viral envelope hemagglutinin (HA) protein triggers fusion of the viral membrane with the endosomal membrane. Fusion alone is not sufficient to cause viral uncoating, however. In addition, protons from the low-pH endosome must enter the virus through M2, a pH-gated proton channel in the viral envelope that opens in response to acidification. The entry of protons through the viral envelope causes dissociation of matrix protein from the influenza virus ribonucleoprotein (RNP), releasing RNP and thus the genetic material of the virus into the host cell cytosol. Amantadine and rimantadine block M2 ion channel function and thereby inhibit acidification of the interior of the virion, dissociation of matrix protein, and uncoating. Note that the drug is shown as “plugging” the channel (lower right panel, upper channel graphic); however, there is also evidence that the drug may bind to the outside of the channel instead (lower right panel, lower channel graphic). NA, neuraminidase.

Antiviral Nucleoside and Nucleotide Analogues Figure 37-5 A. The nucleosides used as precursors for DNA synthesis are depicted in their anti conformations. Each nucleoside consists of a purine (adenine and guanine) or pyrimidine (cytosine and thymidine) base attached to a deoxyribose sugar. These deoxynucleosides are phosphorylated in stepwise fashion to the triphosphate forms (not shown) for use in nucleic acid synthesis. B. Except for cidofovir, the antiherpesvirus nucleoside and nucleotide analogues are structural mimics of deoxyguanosine. For example, acyclovir consists of a guanine base attached to an acyclic sugar. Cidofovir, which mimics the deoxynucleotide deoxycytidine monophosphate, uses a phosphonate (C–P) bond to mimic the physiologic P–O bond of the native nucleotide. Valacyclovir, famciclovir, and valganciclovir are more orally bioavailable prodrugs of acyclovir, penciclovir, and ganciclovir, respectively. C. Anti-HIV nucleoside and nucleotide analogues mimic a variety of endogenous nucleosides and nucleotides and contain variations not only in the sugar but also in base moieties. For example, AZT is a deoxythymidine mimic that has a 3′-azido group in place of the native 3′-OH. Stavudine, zalcitabine, and lamivudine also contain modified sugar moieties linked to natural base moieties. Tenofovir, which is shown as its prodrug tenofovir disoproxil, is a phosphonate analogue of deoxyadenosine monophosphate. Of the analogues that contain modified base moieties, didanosine mimics deoxyinosine and is converted to dideoxyadenosine, while emtricitabine contains a fluoro-modified cytosine and abacavir contains a cyclopropyl-modified guanine. D. Adefovir is a phosphonate analogue of the endogenous nucleotide deoxyadenosine monophosphate, while entecavir is a deoxyguanosine analogue with an unusual moiety substituting for deoxyribose. These two compounds and lamivudine (see panel C) are approved for use in the treatment of HBV infection. E. Ribavirin, which contains a purine mimic attached to ribose, is approved for use against the RNA viruses HCV and RSV.

Mechanism of Action of Acyclovir Figure 37-6 A. Acyclovir is a nucleoside analogue that is selectively phosphorylated by HSV or VZV thymidine kinase to generate acyclovir monophosphate. Host cellular enzymes then sequentially phosphorylate acyclovir monophosphate to its diphosphate and triphosphate (pppACV) forms. B. Acyclovir triphosphate has a three-step mechanism of inhibition of herpesvirus DNA polymerase in vitro: (1) it acts as a competitive inhibitor of dGTP (pppdG) binding; (2) it acts as a substrate and is base-paired with dC in the template strand to become incorporated into the growing DNA chain, causing chain termination; and (3) it traps the polymerase on the ACV-terminated DNA chain when the next deoxyribonucleoside triphosphate (shown here as dCTP, or pppdC) binds.

Nonnucleoside DNA Polymerase and Reverse Transcriptase Inhibitors Figure 37-7 Foscarnet is a pyrophosphate analogue that inhibits viral DNA and RNA polymerases. Foscarnet is approved for the treatment of HSV and CMV infections that are resistant to antiherpesvirus nucleoside analogues. The nonnucleoside reverse transcriptase inhibitors (NNRTIs) efavirenz, nevirapine, delavirdine, and etravirine inhibit HIV-1 reverse transcriptase. The NNRTIs are approved in combination with other antiretroviral drugs for the treatment of HIV-1 infection. Note that the structures of the NNRTIs are significantly different from those of the anti-HIV nucleoside and nucleotide analogues (compare with Fig. 37-5).

Integration of HIV DNA into Cellular DNA and Effect of Anti-HIV Integrase Inhibitor Figure 37-8 A. Schematic rendering of the action of HIV integrase. Double-stranded HIV DNA is generated by reverse transcription as a blunt-ended, linear molecule with repeated sequences known as long terminal repeats (LTR) at both ends. The 5′ LTR includes the promoter/enhancer for HIV transcription, and the 3′ LTR includes the polyadenylation signal. At the termini of both LTRs are identical sequences of four base pairs. In the first step of integration (3′ end processing), HIV integrase removes the two terminal nucleotides from the 3′ strands from both ends of the viral DNA, resulting in two-base (AC), 5′ overhangs. In the second step (strand transfer), integrase creates a staggered cleavage of host DNA, and then catalyzes the attack of the 3′ OH ends of the viral DNA on phosphodiester bonds in the host DNA, resulting in the formation of new phosphodiester bonds linking host and viral DNA at both ends of the viral genome. The AC overhang of viral DNA is not joined, and the process also results in single stranded gaps in the host DNA on each side of the viral genome. This leads to the third step (repair/ligation), in which the AC overhangs are removed and the gaps in host DNA filled in, creating a short duplication of host sequences on either side of the integrated viral DNA. Raltegravir inhibits the strand transfer reaction. B. Domain structure of an HIV integrase monomer. Raltegravir binds at the active site in the catalytic core domain and inhibits the strand transfer reaction. The catalytic triad Asp-64, Asp-116, and Glu-152 is shown as D-D-E in the core domain. C. Structure of raltegravir.

Anti-HIV Protease Inhibitors Figure 37-9 Shown are the structures of the anti-HIV protease inhibitors amprenavir, saquinavir, lopinavir, indinavir, ritonavir, nelfinavir, atazanavir, and tipranavir. These compounds mimic peptides (peptidomimetics), and all but tipranavir contain peptide bonds. Two additional anti-HIV protease inhibitors, darunavir and fosamprenavir (a prodrug form of amprenavir), are not shown.

Steps in the Evolution of Ritonavir Figure 37-10 A. The HIV pol gene product has a phenylalanine (Phe)-proline (Pro) sequence that is unusual as a cleavage site for human proteases. HIV protease cleaves this Phe-Pro bond. The transition state of the protease reaction includes a rotational axis of symmetry. B. Structure-based development of a selective HIV protease inhibitor began with a compound (A-74702) that contained two phenylalanine analogues and a CHOH moiety between them. This compound, which had weak inhibitory activity, was then modified to maximize antiprotease activity while also maximizing antiviral activity, aqueous solubility, and oral bioavailability. The maximization of antiprotease activity was measured as a progressive reduction in IC50, the drug concentration required to cause 50% inhibition of the enzyme. See Box 37-3 for details.

Structure-Based Design of Neuraminidase Inhibitors Figure 37-11 A. Shown is a model of sialic acid (space-filling structure) bound to the influenza A virus neuraminidase, with the amino acids that bind sialic acid depicted in stick form. This structure was used to design transition state analogues that bind more tightly to neuraminidase than sialic acid does, resulting in potent inhibitors of the enzyme. B. Structures of sialic acid and the neuraminidase inhibitors zanamivir and oseltamivir. C. Schematic diagram of the active site of influenza virus neuraminidase, depicting the binding of sialic acid, zanamivir, and GS4071 to several different features of the active site. (Oseltamivir is the ethyl ester prodrug of GS4071.)