Antiviral and Antifungal Agents Celia R. Ravelo, MD, DPPS Objectives 1.

To know the mechanism of action of different antiviral drugs 2. To know the application on what virus the drugs are utilized 3. To know the different adverse effects of the these drugs

Inside the endosome there would be uncoating and releases of viral genome The viral genome will be replicated, transcribed and translated A new virus will then be formed through the process of assembly and maturation It will then be released through the process of budding

I. Viral Life Cycle Virus-cell adsorption (binding, attachment) o The virus has to attach on the host cell o The binding depends on the receptors present on the different cells of the body Virus-cell fusion (entry, penetration) o Once they are attached they will fuse into the host cell membrane Uncoating (decapsidation) o Then eventually they will uncoat and release the viral genome GENOME REPLICATION Early transcription Early translation Replication of the viral genome o the uncoated viral genome will undergo replication, transcription and translation o different proteins necessary for the virus will be formed o the proteins will form proteases Late transcription Late translation Virus assembly o The proteins will form proteases and they will form together o This called the viral assembly Release o Once the mature virus has been formed and it has already matured, it will then be released into the circulation where it will infect other cells

Reverse Transcriptase The HIV viruses is an RNA virus But since it has a reverse transcriptase it is then reversed into a DNA virus The HIV DNA genome is then integrated into the host DNA to produce different RNA and proteins The HIV virus uses the host DNA genome to make its own RNA and proteins for its own formation and maturation Viral Genome Replication Requirements: DNA viruses Deoxyribonucleoside triphosphate RNA Viruses Ribonucleoside triphosphate Pathways for deoxyribonucleoside triphosphates synthesis Salvage pathway uses the enzyme thymidine kinase De Novo pathway - uses the enzyme thymidylate kinase Nucleoside triphosphates - are incorporated into new viral genomes by a viral or cellular polymerase. HSV phosphorylation of nucleosides via the salvage pathway by a viral thymidine kinase;

- a viral DNA polymerase then adds deoxyribonucleoside triphosphates to the growing DNA genome II. Pharmacologic Classes of Agents Anti-HIV drugs Anti-Herpes drugs o Drugs used against CMV Anti-Influenza drugs A. ANTI-HIV DRUGS

Human Immunodeficiency Virus (HIV)

Viral Life Cycle The virus will attach on the receptors Through the process of endocytosis it will enter the cell

HIV contains glycoproteins on its surface o Gp41 o Gp120 Both of them are necessary in order to attach to the T-cell Only the T-cell has the chemokine receptors (CCR5 and CXR4) and CD4 Receptors Special Thanks to: Carla Medina for the recordings ^^

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HIV Life Cycle

Because inside you are replicating viruses, and these viruses in order for them to attack other T cells they need attachment This drug will prevent the virus from infecting other T cells in the body

Enfuvirtide: Mode of Action

Gp120 of HIV will attach into the chemokine and CD4 receptors of the host cell The gp41 will then be exposed, injecting itself into the host cell membrane There will then be fusion of the viral membrane and the host membrane Penetration will follow, there would then be a release of nucleocapsid into the cytoplasm of the host cell Once it is released, the reverse transcriptase enzyme (connected to the viral genome) would now be active Reverse transcriptase enzyme will transcribe the viral RNA into viral DNA The integrase enzyme would integrate the viral DNA into the host cell DNA The RNA will then be transcribed Then there would be synthesis of different proteins in order to assemble the different viral parts The proteases would cleave each protein and through the process of assembly different proteins will come together to form an mature virus Through the process of budding the mature virus will be released from the host cell

The gp120, after binding into the CD4 or chemokine receptors, will expose the gp41 (Fig. a) It is the gp41 that will inject itself into the host cell membrane (Fig. b) Enfuvirtide insert itself into gp41 particles (fig. f), preventing gp41 from injecting itself into the host cell The viral outer layer will not fuse with outer host cell membrane

*Note: Penetration allows the nucleocapsid -- the genetic core -- of the virus to be injected directly into the cell's cytoplasm HIV The process by which HIV's RNA is converted to DNA is called reverse transcription

* Notes from Ppt The native gp41 protein is trapped in the virion in a conformation that prevents its ability to fuse membranes or to bind T-20. Binding of HIV to its cellular receptors triggers a conformational change in gp41 that exposes the fusion-active segment (fusion peptide), heptad repeat region, and a second heptad repeat region mimicked by T-20. The gp41 then refolds, so that the segments mimicked by T-20 bind to the first set of heptad repeats. If the fusion peptide has properly inserted into the host cell membrane, this refolding brings the virion envelope and the cell membrane into close proximity, allowing membrane fusion to occur (by mechanisms that remain poorly understood). When T-20 is present, however, the drug binds to the first set of heptad repeats and prevents the refolding process, thereby preventing fusion of the HIV envelope with the host cell membrane

1. a.

Inhibitors of viral entry Inhibits the attachment of the virus into the host cell Enfuvirtide Peptide First drug that acts by inhibiting viral entry to be approved by the FDA Structurally similar to a segment of gp41 o gp41 - HIV protein that mediates membrane fusion You can still take this drug even though you are already infected with HIV

Administered parenterally, typically by twice daily subcutaneous injections o Since it is a peptide it cant be given orally Special Thanks to: Carla Medina for the recordings ^^

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Adverse effects o Not yet well recognized o Common: Irritation at the site of injection

Excellent substrate for cellular thymidine kinase which phosphorylates AZT to AZT monophosphate

2.

Inhibitor of Viral Genome Replication a. Inhibitor of Reverse Transcriptase o It is important to inhibit this enzyme since it is the one responsible for transcription of viral RNA to viral DNA that will be integrated into the host cell genome o Inhibit Viral polymerases or reverse transcriptase(human herpesviruses, the retrovirus HIV, and the hepadna virus HBV) Nucleoside analogue Non-nucleoside inhibitor of DNA polymerase or reverse transcriptase All nucleoside analogues must be activated by phosphorylation, usually to the triphosphate form, in order to exert their effect Nucleoside analogues inhibit polymerases by competing with the natural triphosphate substrate; incorporated into the growing DNA chain, where they often terminate elongation. Either or both of these featuresenzyme inhibition and incorporation into DNAcan be important for antiviral Role of RT (reverse transcriptase) 2 main categories of nucleoside analogues o Antiherpesvirus agent o Anti-HIV agents o 2 anti-HIV agents (adefovir and lamivudine) and a third drug, entecavir are also approved for use agai*nst hepatitis B virus This drug will enter the cell as nucleoside and by the action of the cellular thymidyllate kinase it will be phosphorylated Thus ultimately becoming a Zidovudine triphosphate The triphosphate form will the one to attach to the reverse transcriptase The Reverse transcriptase will attach the nucleotide into the protein chain of the viral DNA Nucleotide is needed for the formation of the protein chain of the viral DNA Since the drug is a false triphosphate, the growth of DNA will not progress and at the same time Since the drug attaches to the active site of the enzyme, the said enzyme will be paralyzed Thus the RNA genome of HIV will not progress into DNA, thereby breaking the process of the viral life cycle

o o o

o o o

*Note:

Reverse transcriptase (RT) - DNA polymerase that can copy both DNA and RNA Copies the RNA retrovirus genome into doublestranded DNA after the virus enters a new cell

2 Groups of Drugs that Inhibit Reverse Transcriptase NRTI Nucleotide Reverse Transcriptase Inhibitors NNRTI Non-Nucleotide Reverse Transcriptase Inhibitors Nucleoside Reverse Transcriptase Inhibitor (NRTI) Zidovudine Lamivudine Zidovudine (AZT) Nucleoside analogue with an altered sugar moiety o A nucleoside has to be converted to a nucleotide, so this drug needs to undergo 3 phosphorylation inside the cell As with acyclovir, AZT is an obligatory chainterminator More potent inhibitor of HIV RT but it also affects the human DNA polymerases (reason for having adverse effects) Adverse effect: bone marrow suppression neutropenia anemia The limited clinical effectiveness of AZT, and problems with its toxicity and resistance, have led to the development of other anti-HIV drugs and to the use of combination chemotherapy for HIV

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Lamivudine (3TC) L-stereoisomer biologic nucleosides Contains sulfur atom in its five-membered ring Weakly inhibits mitochondrial DNA polymerase therefore less toxic Resistance to 3TC develops quickly in patients so it is used in combination with other anti-HIV drugs Non- Nucleoside Reverse Transcriptase Inhibitors (NNRTI) Efavirenz first anti-HIV drug to be taken OD Nevirapine Delavirdine o Note that since they are non-nucleoside they are in nucleotide form already o They dont need to be phosphorylated by the kinases o It will be inserted directly into the reverse transcriptase *Approved in combination with other antiretroviral drugs for the treatment of HIV-1 infection.

HIV Proteases as Target for anti-HIV drugs First, it is essential for HIV replication Second, a point mutation is sufficient to inactivate the enzyme Third, the substrates of HIV protease are conserved and somewhat unusual, suggesting both specificity and a starting point for drug design. Fourth, HIV proteaseunlike the human proteases most closely related to itis a symmetric dimer of two identical subunits, each of which contributes to the active site, again suggesting both specificity and a starting point for drug design. Fifth, the enzyme can be easily overexpressed and assayed, and its crystal structure has been solved. Common Toxicities of Antiretroviral Drugs

b.

Inhibitor of Integrase Enzyme o Integrase enzyme is the one responsible for integrating HIV pro-viral DNA to the host cell DNA to produce the viral mRNA o Drug: __________

3.

Inhibitor of Viral Maturation a. HIV Protease Inhibitors o HIV protease are the one responsible for cleaving different protein that were synthesized for the assembly of a new virus o The inhibitors will prevent this assembly, thus preventing the formation of a new virus Saquinavir Ritonavir Amprenavir Indinavir Nelfinavir Lopinavir Atazanavir Tipranavir Darunavir

B. 1.

ANTI-INFLUENZA Inhibitor of Viral Uncoating (Exclusive for Influenza A viruses) Amantadine Rimantadine

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envelope hemagglutinin (HA) protein triggers fusion of the viral membrane with the endosomal membrane. HA binding alone is not sufficient to cause viral uncoating, however. In addition, protons from the low-pH endosome must enter the virus through M2, a pHgated 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. NA, neuraminidase; ADP, adenosine diphosphate.

Adverse Effects: Amantadine GIT o CNS o o o

Nausea, vomiting, diarrhea, anorexia

Lightheadedness Headache difficulty concentrating likely due to its effects on host ion channel Note: adverse effects are more common in CNS Used also for treatment of Parkinson's disease

Amantadine and Rimantadine act on the uncoating of the viral core If the viral core will not be uncoated then the viral genome will not move out, thus replication of these genome will cease Uncoating o The uncoating happens within the endosome o There is a M2 channel on the surface of the endosome o M2 is important for the acidification of the endosome and ultimately the viral matrix o If acidity is achieved the matrix will be disrupted or endosome will be ruptured, releasing now the viral genome Action of Amantadine and Rimantadine o Both of them block the M2 channel o If M2 channels are blocked the acidification of the endosome will cease o The viral matrix and the endosome will not rupture, therefore the viral genome will not be released Amantadine and Rimantadine only differ in their Pkinetics o Rimantadine has longer half life, metabolized in the liver o Amantadine is the only drug utilized in the management of Parkinsons disease o Adverse effect is more common in amantadine

Rimantadine Analogue of amantadine Similar antiviral mechanism Lacks adverse effects as seen in amantadine especially the neurological o Less adverse effect o Reason why patients prefer this drug Prophylactic agent in settings where there is a large population at risk from influenza morbidity (e.g., nursing homes) Inhibitor of Viral Release Zanamivir o Poor oral bioavailability o Administered by inhalation Oseltamivir o Oral availability is approximately 75% o When taken prophylactically, oseltamivir reduces the number of flu cases in susceptible populations (e.g., nursing home residents) o Both oseltamivir and zanamivir reduce the duration of flu symptoms in patients who are already infected with the virus. However, this reduction is only 1 day on average, and even this modest effect requires that the drugs be taken within 2 days of the onset of symptoms. o Oseltamivir is effective in preventing human mortality due to H5N1 avian influenza (bird flu) Both of them inhibit influenza virus neuramidase, causing newly synthesized virions to attached to the host cell

2.

Notes from ppt + 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

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o o o o o

Administered intravitreally Antisense oligonucleotide Antisense oligonucleotides target specific RNAs It is utilized for the treatment of CMV The drug is approved for treatment of ophthalmic CMV disease and it is used mainly in CMV retinitis

Neuramidase o An enzyme that reduces the stickiness of the virus from the host cell o It is needed for the release of the new virus from the host cell Zanamivir and Oseltamivir inhibit the neuramidase o Thus the virus will remain attach to the host cell o The newly formed virus will not be able to infect other host cells

Ribavirin o Broad-spectrum antiviral that exhibits activity against many viruses in vitro and efficacy against several in vivo o Approved only in aerosol form (in effect, topical application to the lungs) for severe respiratory syncytial virus (RSV) infection, and only in combination with an interferon for chronic hepatitis C virus (HCV) infection. o Ribavirin is converted to a monophosphate by cellular adenosine kinase and is known to inhibit cellular inosine monophosphate dehydrogenase, thereby lowering cellular GTP pools o Inhibition of viral RNA polymerase could represent a second possible selective mechanism for ribavirin action o Interestingly, both ribavirin diphosphate and ribavirin triphosphate have inhibitory activity against the RNA polymerase from certain viruses. o A third possible mechanism also involves viral RNA polymerase. o It has been approved only in aerosol form (in effect, topical application to the lungs) for severe respiratory syncytial virus (RSV) infection It is also approved only in combination with an interferon for chronic hepatitis C virus (HCV) infection

Zanamivir and Oseltamivir o Both of them has the same mechanism of action their difference lies at the Pkinetics Zanamivir o Poor bioavailability o So it is only given via Aerosol Oseltamivir o With oral preparation o Used in H1N1 virus o It has to be taken within 2 days of the onset of symptoms o Lifesaving drug endemic regions

C.

Anti-Herpes Agents o Phosphorylation of drug by viral kinases leads to inhibition of DNA synthesis in virus-infected cells Inhibitor of Viral Replication

1.

3.

Antiviral Drug with Unknown Mechanisms of Action Fomivirsen o An antisense oligonucleotide. that target specific RNAs o If the viral RNA is an mRNA, binding of the oligonucleotide should prevent the synthesis of the protein encoded by the mRNA. o Fomivirsen is the first FDA-approved oligonucleotide drug o Designed to bind to an mRNA that encodes IE2, a gene-regulatory protein of CMV o More potent than ganciclovir against CMV, with activity at submicromolar concentrations. o Approved for treatment of ophthalmic CMV disease and is used mainly in CMV retinitis

Nucleoside and Nucleotide Analogue Acyclovir Valacyclovir Ganciclovir Valganciclovir Penciclovir Famciclovir Cidofovir a. Acyclovir Against HSV and VZV o It most common drug against HSZ and VZV because it is the cheapest drug Nucleoside analogue Has high therapeutic index A guanosine analog that acts as an antimetabolite It is the only approved drug for neonatal encephalitis It has long bioavailability Special Thanks to: Carla Medina for the recordings ^^

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Acyclovir Mode of Action

delivers DNA to the human nucleus; the active drug acyclovir triphosphate exerts its action on the viral DNA located in the nucleus.

Acyclovir once it enters the virally infected cell it will be phosphorylated It will then be doubly phosphorylated by cellular kinase o Note that it only requires the virally infected cellular kinase o This is also the reason for the resistance to Acyclovir o Some viruses can mutate and will no longer produce the kinase needed by the Acyclovir Once Acyclovir is phosphorylated and eventually become Acyclovir triphosphate the herpes DNA polymerase will now insert it into the herpes simplex genome

Notes from ppt The acyclovir triphosphate competes with 2-deoxyguanosine triphosphate (dGTP) as a substrate for viral DNA polymerase, as well as acting as a chain terminator. In actual infection, the HSV releases its naked capsid that delivers DNA to the human nucleus; the active drug acyclovir triphosphate exerts its action on the viral DNA located in the nucleus. Resistance vs Acyclovir HSV Absence of partial production of viral thymidine kinase Altered thymidine kinase substrate specificity Altered viral DNA polymerase VZV Mutation in VZV thymidine kinase Mutation in viral DNA polymerase A vaccine is produced for VZV o Since it has no subspecies or different strain, a vaccine was produced o This is also the reason why VZV can be eradicated by immunization Adverse effects ORAL: nausea, diarrhea, rash, headache, renal insufficiency, neurotoxicity TOPICAL: mucosal irritation and transient burning to genital lesions IV: renal insufficiency b. Valacyclovir A prodrug form of acyclovir Has approximately fivefold greater oral bioavailability than acyclovir Rapidly converted to acyclovir after oral administration The active metabolite acyclovir can easily go into the circulation Penciclovir and famciclovir Famciclovir is the diacetyl 6-deoxy analogue of penciclovir Penciclovir is the active form MOA same as Acyclovir Used for treatment of shingles, cold sores and HSV infections It should be given at the early phase of the illness, what it do is that it reduces the duration of the illness

If the Acyclovir triphosphate is inserted into the growing DNA of herpes virus, the next nucleotide will not be inserted, thereby terminating the growth of the viral DNA Acyclovir triphosphate will also paralyze the DNA polymerase, it can no longer carry another nucleotide to be inserted into the growing viral DNA

Notes from ppt The acyclovir molecules enter the cell and are converted to acyclovir monophosphate by the HSV enzyme thymidine kinase (TK). Enzymes in the human cell add two more phosphates to eventually form the active drug acyclovir triphosphate. The acyclovir triphosphate competes with 2-deoxyguanosine triphosphate (dGTP) as a substrate for viral DNA polymerase, as well as acting as a chain terminator. In actual infection, the HSV releases its naked capsid that

c.

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d.

Ganciclovir A nucleoside analogue Originally synthesized as a derivative of acyclovir, with the intention of developing another anti-HSV drug Much more potent than acyclovir against CMV First antiviral drug approved for use against CMV More closely resembles deoxyguanine accounting for its toxicity Used only for more serious infection Viral protein kinase called UL97 phosphorylates ganciclovir (30-fold increase in the amount of phosphorylated ganciclovir in infected versus uninfected cells) Ganciclovir triphosphate inhibits CMV DNA polymerase more potently than it does cellular DNA polymerases Selective against CMV at two steps: phosphorylation and DNA polymerization. More toxic than acyclovir Toxicity: bone marrow suppression, especially neutropenia It is a problem among pregnant individual o It can cause fetal abnormality It is also a problem among immunocompromised patient

Approved for use in the treatment of CMV retinitis in patients with HIV/AIDS Long intracellular half-life Cleared in the kidneys Most prominent toxicity: Nephrotoxicity o To prevent the toxicity it must be administered with probenecid o Probenecid inhibits a proximal tubule anion transporter and thereby decreases cidofovir excretion Two related phosphonate-containing drugs: Tenofovir and Adefovir - acyclic deoxyadenosine monophosphate analogues o Tenofovir - approved as an anti-HIV drug in 2001, can be administered just once each day an important advantage for HIV-infected individuals who must comply with complex combination chemotherapy regimens o Adefovir - approved as an anti-HBV drug in 2002 The mechanisms of action of these drugs against their respective viruses are similar to that of cidofovir against CMV

e. f.

Valganciclovir Prodrug of ganciclovir For better bioavailabilty With same mechanism of action Cidofovir Also known as hydroxyphosphonylmethoxypropylcytosine (HPMPC), Phosphonate-containing acyclic cytosine analogue o It means that this drug is already a monophosphate Can be considered a nucleotide rather than a nucleoside analogue Mimics deoxycytidine monophosphate;( thus, in effect, it is already phosphorylated) Does not require viral kinases for its phosphorylation Active against kinase-deficient viral mutants that are resistant to ganciclovir Enters cells efficiently Further phosphorylated (twice) by cellular enzymes to yield an analogue of dCTP, which inhibits herpesvirus DNA polymerases more potently than cellular DNA polymerases

Non-Nucleoside DNA polymerase Inhibitor Foscarnet a. Foscarnet Inhibits both DNA and RNA polymerases encoded by a wide variety of viruses Relatively broad spectrum of activity in vitro (including against HIV), but clinically it is used for certain serious HSV and CMV infections where therapy with acyclovir or ganciclovir has not succeeded Differs from nucleoside analogues in that it does not require activation by cellular or viral enzymes: rather, foscarnet inhibits viral DNA polymerase directly by mimicking the pyrophosphate product of DNA polymerization Selectivity results from the increased sensitivity of viral DNA polymerase relative to cellular enzymes; Selectivity is not as high as acyclovir's It inhibits cell division at concentrations not much higher than its effective antiherpesvirus concentration. Major drawbacks to foscarnet use include its lack of oral bioavailability and its poor solubility; renal impairment is its major dose-limiting toxicity DRUGS THAT MODULATE IMMUNE SYSTEM Immunization o Active o Passive Interferons Imiquimod Immunization Active and passive immunization inhibit viral infection by providing antibodies against viral envelope proteins Antibodies then block the attachment and penetration of virions into cells and increase virion clearance Some antibodies are directly virucidal, causing virions to be destroyed or inactivated before the virus can interact with its receptor(s) on target cells Special Thanks to: Carla Medina for the recordings ^^

Cidofovir Mecahnism of Action

D. a.

This drug is a phosphate It will then be phosphorylated by the host cell kinase This will then be inserted into the growing viral genome Eventually it will also inhibit the viral polymerase enzyme

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Active o o

It involves challenging our body with inactivated organism or a particle of an organism The weakened particle will boost our immune system to develop antibodies so that the next time we encounter those antigen, we are already immune to them Antibodies are given Given to new born, gainst tetanus toxin

ANTIVIRAL DRUGS SUMMARY

HIV
Zidovudine Didanosine Zalcitabine Stavudine Lamivudine Nevirapine Delavirdine Efavirenz Saquinavir Ritonavir Indinavir Nelfinavir Amprenavir Lopinavir Abacavir

CMV
Ganciclovir Foscarnet Cidofovir Fomivirsen

HSV
Acyclovir Valaciclovir Penciclovir Famciclovir Idoxuridine Trifluridine Brivudi

Influenza A
Amantadine Rimantadine

Influenza A and B
Oseltamivir Zanamivir

Passive o o

b.

Interferon Proteins that were produced in response to virus infection and that could inhibit replication of the same or other viruses o Note that it has no specificity o No specific virus is targeted Two major types of interferons Type I interferons - interferon a and interferon , which are produced by many cell types and interact with the same cell-surface receptor Type II interferons - interferon , which is typically produced by cells of the immune system, especially T cells, and interacts with a separate receptor Interaction of interferons with their receptors induces a series of signaling events that activate and/or induce the expression of proteins that combat virus infections One relatively well-understood example of such a protein is a protein kinase, called PKR, which is activated by double-stranded RNA. PKR phosphorylates a component of the host translational machinery, thereby turning off protein synthesis and thus the production of virus in infected cells Used as a therapeutic agent in the treatment of: HCV HBV condyloma acuminata (which is caused by certain HPVs Kaposi's sarcoma (which is caused by Kaposi's sarcoma-associated herpesvirus [KSHV] Imiquimod Approved for reatment of certain diseases caused by HPVs Interacts with the Toll-like receptors TLR7 and TLR8 to boost innate immunity, including the secretion of interferons. Toll-like receptors are cell surface proteins that recognize pathogen-associated molecular patterns. Activation of Toll-like receptors induces intracellular signaling events that are important for defense against pathogens It enhances our body to produce interferon

c.

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Anti-Fungal Agents Main Target Site of Anti-Fungal Drugs Cell membrane Cell wall Nucleic Acid Synthesis Sites of action of anti-fungal drugs

b.

a.

Inhibitor of Fungal Nucleic Acid Synthesis: Flucytosine Flucytosine is selectively taken up by fungal cells via cytosine-specific permeases that are expressed only in fungal membranes Lacking these transporters, mammalian cells are protected

Fungistatic under most circumstances Fungi and bacteria in the intestine can convert flucytosine into 5-fluorouracil, which can cause adverse effects in host cells o The adverse effect is not directly given by flucystosine because the adverse effects comes from the 5FU that is secreted by the fungi and bacteria inside the intestine Used in combination with amphotericin B to treat systemic mycoses Used singly, resistance emerges rapidly due to mutations in fungal cytosine permease or cytosine deaminase Can kill aspergillus when combined with amphotericin B The mechanism of this synergistic interaction appears to involve enhancement of flucytosine uptake by fungal cells due to amphotericin-induced damage to the fungal plasma membrane As a single agentactionis limited to o candidiasis, cryptococcosis, and chromomycosis o has large volume of distribution, with excellent penetration into the central nervous system (CNS), eyes, and urinary tract Dose-dependent adverse effects: bone marrow suppression leading to leukopenia and thrombocytopenia, nausea, vomiting, diarrhea, and hepatic dysfunction Contraindicated during pregnancy Inhibitor of fungal Mitosis: Griseofulvin Binds to tubulin and a microtubule-associated protein disrupting assembly of the mitotic spindle. Also inhibit fungal RNA and DNA synthesis Accumulates in keratin precursor cells and binds tightly to keratin in differentiated cell that allows new growth of skin, hair, or nail to be free of dermatophyte infection. Fungistatic under most circumstances. Therapeutic uses: topical antifungal medications Griseofulvin can be used to treat fungal infection of the skin, hair, and nail due to Trichophyton, Microsporum, and Epidermophyton. Not effective against yeast (such as Pityrosporum) and dimorphic fungi. It is important to continue treatment until the infected skin, hair, or nail is completely replaced by normal tissue Inhibitor of Ergosterol biosynthesis Squalene epoxidase Inhibitor Inhibitors of 14a- Sterol Demethylase

c.

Inside the fungal cell, the enzyme cytosine deaminase converts flucytosine to 5-fluorouracil (5-FU) Subsequent reactions convert 5-FU to 5fluorodeoxyuridylic acid (5-FdUMP), which is a potent inhibitor of thymidylate synthase Inhibition of thymidylate synthase results in inhibition of DNA synthesis and cell division

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Divided into allylamines and benzylamines based on their chemical structures: terbinafine and naftifine are allylamines, whereas butenafine is a benzylamine.

Ergosterol is synthesized in fungal cells from acetyl CoA building blocks One of the intermediates, squalene, is converted to lanosterol by the 14a-Sterol demethylase, a cytochrome P450 enzyme not expressed in mammalian cells, catalyzes the first step in the conversion of lanosterol to the unique fungal sterol ergostero Imidazoles and triazoles inhibit 14a-sterol demethylase and thereby prevent the synthesis of ergosterol, which is the principal sterol in fungal membranes. Fluconazole and voriconazole are two representative triazoles action of squalene epoxidase Allylamines and benzylamines inhibit the action of squalene epoxidase 1. Squalene epoxidase Inhibitor: Terbinafine and Naftifine

Terbinafine Available in both oral and topical formulations When taken orally, the drug is 99% protein-bound in the plasma and it undergoes first-pass metabolism in the liver Oral bioavailability of terbinafine is 40%. Elimination half-life is extremely long, approximately 300 hours, because terbinafine accumulates extensively in the skin, nails, and fat. Oral form is used in the treatment of onychomycosis, tinea corporis, tinea cruris, tinea pedis, and tinea capitis. Not recommended in patients with renal or hepatic failure and in pregnant women Very rarely, the oral form of terbinafine can lead to hepatotoxicity, Stevens-Johnson syndrome, neutropenia, and exacerbation of psoriasis or subacute cutaneous lupus erythematosus. Liver function enzymes should be monitored during the treatment course. Plasma levels of terbinafine are increased by coadministration with cimetidine (a cytochrome P450 inhibitor) and decreased by coadministration with rifampin (a cytochrome P450 inducer) Topical terbinafine is available in cream or spray form and is indicated for tinea pedis, tinea cruris, and tinea corporis. Naftifine broad-spectrum antifungal activity Available topically as a cream or gel Effective in tinea corporis, tinea cruris, and tinea pedis Butenafine A benzylamine Topical antifungal agent with a mechanism of action and spectrum of antifungal activity similar to that of the allylamine More effective than topical azole agents against common dermatophytes

2.

Inhibitors of 14a- Sterol Demethylase Imidazoles and Triazoles Results in decreased ergosterol synthesis and accumulation of 14a-methyl sterols disrupting the tightly packed acyl chains of the phospholipids in fungal membrane Destabilization of the fungal membrane leads to dysfunction of membrane-associated enzymes, including those in the electron transport chain, and may ultimately lead to cell death. Not completely selective for the fungal P450 enzyme, however, and they can also inhibit hepatic P450 enzymes

Squalene is converted to lanosterol by the action of squalene epoxidase Prevent the formation of lanosterol, which is a precursor for ergosterol Promote accumulation of the toxic metabolite squalene in the fungal cell, making them fungicidal under most circumstances

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Azoles

Antifungal Activity: B. Dermatitidis Cryptococcus neoformans H. capsulatum, Coccidioides species, P. brasiliensis, dermatophytes, and most Candida species Generally fungistatic rather than fungicidal against susceptible organisms

Amphotericin B Binding to ergosterol produces channels or pores that alter fungal membrane permeability and allow for leakage of essential cellular contents, leading ultimately to cell death Destabilize fungal membranes by generating toxic free radicals upon oxidation of the drug Affinity to ergosterol is 500 times greater than its affinity for cholesterol Concentration of membrane-associated ergosterol in a given fungal species determines whether it is fungicidal or fungistatic for that species Resistance is attributable to a decrease in the ergosterol content of the fungal membraen Highly insoluble Supplied as a buffered deoxycholate colloidal suspension o Given via IV Poorly absorbed from the gastrointestinal tract; administered intravenously In the bloodstream, more than 90% of the drug binds rapidly to tissue sites Low CSF, vitreous humor and amniotic fluid penetration Intrathecal therapy may be necessary for treatment of serious meningeal disease o To target CNS infection Notes from ppt For decades, this drug provided the only effective treatment vs systemic mycoses .Its therapeutic and toxic effects are related to its affinity for plasma membrane sterols. Adverse Effects Immediate systemic reactions Renal effects Hematologic effects Systemic Reactions Cytokine storm - elicits release of tumor necrosis factor-alpha (TNF-a) and interleukin-1 (IL-1) from cells of the host immune system fever chills hypotension within the first several hours after drug administration loss of consciousness May be minimized by decreasing the rate of drug administration or by pretreatment with antipyretic agents (e.g., acetaminophen, nonsteroidal antiinflammatory drugs [NSAIDs], or hydrocortisone) Renal Toxicity Mechanism of renal toxicity is unknown but may be related to amphotericin-mediated vasoconstriction of afferent arterioles leading to renal ischemia Often the limiting factor in determining the extent of the therapeutic response to amphotericin B Discontinue therapy temporarily if the blood urea nitrogen (BUN) exceeds 50 mg/dL or the serum creatinine exceeds 3 mg/dL. Special Thanks to: Carla Medina for the recordings ^^

2 Types of Azoles Imidazoles ketoconazole, clotrimazole, miconazole, econazole, butoconazole, oxiconazole, sertaconazole, and sulconazole Triazoles itraconazole, fluconazole, voriconazole, terconazole, and posaconazole; one additional member of this class, ravuconazole, is currently in clinical trials Fluconazole Most widely used antifungal drug Hydrophilic triazole that is available in both oral and intravenous formulations The bioavailability of oral fluconazole is nearly 100%, and, unlike ketoconazole and itraconazole, its absorption is not influenced by gastric pH. Once absorbed, fluconazole diffuses freely into CSF, sputum, urine, and saliva. Fluconazole is excreted primarily by the kidneys. low adverse-effect profile excellent CSF penetration drug of choice for systemic candidiasis and cryptococcal meningitis also the drug of choice for coccidioidal meningitis Not effective against aspergillosis

d.

Polyene antifungal agents Amphotericin B Nystatin Both are derived from Streptomyces sp.

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Renal tubular acidosis Cylindruria (the presence of renal cell casts in the urine) Hypokalemia

Summary of Anti-Fungal Drugs and their MOA

Hematologic Toxicity Anemia - secondary to decreased production of erythropoietin. Renal and hematologic toxicities of amphotericin B are cumulative and dose-related Therapeutic measures that can minimize these toxicities: o avoidance of other nephrotoxic drugs, such as aminoglycosides and cyclosporine o maintenance of euvolemia to provide adequate renal perfusion Nystatin Structural relative of amphotericin B Acts by binding ergosterol and causing pore formation in fungal cell membranes Used topically to treat candidiasis involving the skin, vaginal mucosa, and oral mucosa e. Glucan Synthesis Inhibitor: Echinocandins Inhibitor of fungal cell wall synthesis

Noncompetitively inhibit the synthesis of -(1,3)-D-glucans Disrupts cell wall integrity resulting in osmotic stress, lysis of the fungal cell, and ultimately fungal cell death caspofungin micafungin Anidulafungin all are semisynthetic lipopeptides derived from natural products (not oral) Fungicidal against Candida species, including Candida glabrata and Candida krusei Fungistatic against Aspergillus species Currently available only in parenteral form because they are insufficiently bioavailable for oral use

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Special Thanks to: Carla Medina for the recordings ^^