ID 20: Pharmacology of Anti-fungal Agents

Dermatophytoses?

"dermatophytes infections" - Dermatophytes are the type of fungal organism that can only feed off of dead tissue (nails, top part of skin). - These fungi cannot pass the immune system, only feed on dead tissue. - Use keratinized tissue as energy source.

Topical azoles?

- Include all imidazole such as clotrimazole, econazole, miconazole, oxiconazole - Widespread use as topical antifungals in: 1) Mucocutaneous candidiasis (vaginal candidiasis, oropharyngeal thrush) 2) Dermatophyte infections of the skin (e.g. ringworm)

CYP-P450 interactions with triazoles?

- She does not expect us to memorize all the the cytochrome P450 interactions. We just need to know their relative importance. (which azole should be prescribed. - The number of major drug interactions is lowest for fluconazole. Then in order of increasing adverse reactions; fluconazole, voriconazole, posaconazile, itraconazole.

Another way for fungal infections to be classified?

Fungal infections can also be classified by the type of species you are infected with. 1) Primary pathogens 2) Opportunistic pathogens.

MOA of topical allylamines?

MOA: inhibit squalene epoxidase • Fungicidal against dermatophytes, fungistatic against Candida • Used as topical cream in the treatment of tinea infections

Fungal infections are known as?

Mycoses

Topical Polyenes (nystatin)?

binds to ergosterol

Ciclopirox 8% nail lacquer (Penlac®)?

• For topical treatment of mild to moderate fungal nail infections • although clinical trials showed poor evidence that this treatment option works against onchyomycosis

Voriconazole (Vfend®)?

• Is a triazole • Newer azole, well absorbed orally (Highly prescribed orally available agent) • May result in hepatotoxicity, transient visual disturbances, skin rash • Likely teratogenic, contra-indicated in pregnant women (category D) • smaller number of drug interactions, but includes decreased metabolism of phenytoin, warfarin, omeprazole • Broad spectrum of antifungal activity, including invasive Aspergillis (more effective and better tolerated than AmB), and as salvage therapy for infections from Scedosporium and Fusarium species

Spectrum of Echinocandins?

• Limited to Candida and Aspergillus species

Amphotericin B (Fungizone®): Antifungal spectrum and Therapeutic uses?

• The most broad spectrum intravenous antifungal available • Active against almost all common invasive fungi (including many Candida species and some Aspergillis species) as well as endemic fungi • One of the mainstay for treatment of serious systemic fungal infections: - wide spectrum of antifungal activity - low cost - however, lack of oral bioavailability is a major roadblock • Development of resistance during therapy is uncommon • Development of toxicity is a major limitation to use

Flucytosine (Ancobon®)?

**not available in Canada MOA: anti-metabolite, fluorinated pyrimidine analog • metabolized in fungal cells to 5-FU and interfere with DNA and RNA metabolism - Takes the place of uracil monophosphate. If you inhibit the synthesis of the building blocks, cannot make DNA • fungicidal Adverse effects: bone marrow, liver and renal toxicities which could be life-threatening. This is why its not available in Canada. Therapeutic Uses: only use is in the systemic treatment of severe candidiasis or cryptococcal meningitis, in combination with AmB (US FDA protocol)

How is the fungal cell wall different from a human cell wall?

- A human cell stops at the phospholipid bilayer. The cell wall structure is all fungal specific. - There is steroid in the phospholipid bilayer than is known as ergosterol, this is different compared to the human membrane in which we have the same steroid but in the form of cholesterol. - The substitution of cholesterol to ergosterol, allows us to target ergosterol in antifungal medication.

How can broad spectrum antibiotics cause fungal infections?

- Broad spectrum antibiotic treatment destroys the balance that used to exist between your normal flora and fungal species. - A side effect of antibiotics is fungal infection.

Dimporhic fungi?

- Can stay in two morphological forms: single cell organism like a bacteria or multicell form (mold) = dimorphic fungi. - In body usually exist in the unicellular form.

Candidiasis: Candida infections?

- Candida is the most common opportunistic fungal species causing systemic and non-systemic mycoses - includes about 20 species of yeast - common cause of fungal infections: both systemic and non-systemic - many species (e.g. C. albicans) are a normal part of flora of mouth, colon, vagina - most Candidiasis are opportunistic infections: resulting from antibiotic treatment, immunologic depression, stress among other factors

Additional topical antifungal drugs for cutaneous infections?

- Ciclopirox olamine (Loprox®) - Tolnaftate (Tinactin®, others) - Undecylenic acid (Desenex® in various forms) - Tavaborole (Kerydin®, others)

Amphotericin B (Fungizone®) and Nephrotoxicity?

- Nephrotoxicity is a therapy limiting toxicity, has to do with the fact that amp B also binds to human cholesterol and because it is mainly eliminated by kidney filtration the site of damage is in the kidney (nephron). - Acute toxicity: reversible renal insufficiency associated with electrolyte disturbances occurs in up to 80% of patients within 2 weeks of initiation of therapy, may be reduced by co-administration of normal saline -Irreversible toxicity: however, irreversible renal dysfunction may occur on prolonged administration (total dose > 3-4g) - reduction of nephrotoxicity is the major rationale for the development of AmB lipid formulations

(B-(1,3)-glucan synthase?

- On top of the chitin there is a cross linking structure that gives the rigidity and the structure to the cell wall. - The cross linking entity is formed by the beta-(1,6)-glucan and Beta-(1.3)-glucan structure, kind of like peptidoglycan. beta-(1,6)-glucan and Beta-(1.3)-glucan are being manufactured by an enzyme in the cell membrane ,(B-(1,3)-glucan synthase, is a drug targeted.

PAMPs?

- On top of the crosslinking part is the mannoproteins - which is a PAMP, pathogen associated molecular pattern. - PAMPs are recognized by the innate immune system (not just phagocytes) , when the immune system encounters PAMPs they know it is foreign and respond accordingly. A lot of these fungal pathogens learn to coexist with us, our immune systems learns to tolerate them - there is a phenomenon where the immune system is able to maintain them at a certain level. They do so by recognizing the PAMPs, that it is foreign cell and needs to be accounted for. And that mannoprotein is highly immunogenic .

Class I: Polyenes Amphotericin B (Fungizone®)?

- Polyene macrolide antibiotic produced by Streptomyces nodosus. There are multiple other polyenes isolated from this bacteria. - In clinical use since 1960. Polyene macrolide is the oldest anti-fungal agent that is still in clinical use. - This one is still used in a number of systemic antifungal therapies. - There is two phases to this molecule, there is a double bond heavy side and then a hydroxyl group heavy side. - The structure tells you that the double bond heavy side is hydrophobic, whereas the other side is hydrophilic.

Which Allylamines are we going to learn about?

- Terbinafine (Lamasil®)

Echinocandins MOA?

- The MOA is the inhibition of the B-(1,3)-glucan synthase that is situated in the cell membrane. - It is responsible for the biosynthesis of the B-(1,3)-glucan which is important for the structural integrity of the cell wall. - When you have the inhibition of (1,3) synthesis, the (1,6) cannot be cross linked and it causes the cell wall to collapse. - Disrupts fungal cell wall leading to osmotic stress and cell death: Fungicidal!!!! - Selective, as β(1-3) glucan is unique to fungal cells Inhibition of β(1,3) glucan synthesis leading to cell wall stress and fungal cell death

Azoles : MOA?

- The MOA of azoles is to inhibit ergosterol synthesis. But instead of binding to ergosterol like amp B it inhibits the biosynthesis of the molecule - Azoles inhibit the biosynthesis of ergosterol by inhibiting Erg11 also known as Cyp51. - In some fungal species there is two isoforms of this enzyme (Cyp51a/b), and in some fungal species there is only 1. -Depending on the ability of the azole to inhibit the isoforms, it limits the spectrum of activity of the agent. - There are many steps in the synthesis of ergosterol, when you inhibit Erg11 the actually biosynthesis can shunt to a different pathway that allows Erg3 to take over. - It allows the sterol precursor to be manufactured into a different type of steroid precursor, this type of sterol is toxic when inserted into the fungal membrane. - The synthesis of these toxic sterols that get inserted into the membrane are what causes the fungal cell to die. - Inhibition of ergosterol synthesis leading to the production and membrane poisoning by toxic sterol

New England Compounding Center meningitis outbreak: 2012-13?

- This happened in the US, almost 6-7 years ago. All people getting compounded medication made in this centre were exposed to this outbreak. - There was contamination of epidural steroid injection doses by: 1) Aspergillus 2) Setosphaeria - 14,400 patients exposed from May to September, 2012 - as of March 2013, 48 patients had died and 720 being treated from persistent fungal infection - CDC had to trace to the origin, all using injectable steroid from this compounding company. - Pharmacists Glenn Chin and Barry Cadden were indicted in December 2014 - Were responsible for forging sterilization and quality control documents. - Both Glenn Chin and Barry Cadden were given prison sentences for 8 and 9 years in 2018 and 2017, respectively.

Therapeutic concerns with azoles: Pharmacokinetic Drug Interactions?

- We are not expected to know all the cytochrome /azole interactions. She wants us to know the relative importance of each of the azoles. So count how many times ketoconazole is showing up (we will have a summary table, don't have to count). - Ketoconazole (being an imidazole) is actually the worse in terms of drug interactions because it can bind to almost all of the mammalian cytochrome p450s in the liver. That means it interacts with almost everything that is metabolized by the p450 pathway. - Imidazole is generally and rapidly becoming a less prescribed systemic agent because of its drug interactions. - Other triazoles, depending on which one they are have varying degrees of ability to bind to the mammalian p450, and depending on how many they bind to , the number of drug interactions will go up if they bind to more.

Mucosal fungal infections?

- affect mucocutaneous tissues and mucous membranes - mucosal membranes are the barrier of the major organ to the outside world. -he urogenital area is covered with mucosal membrane as well as oral cavity. These are hard to cure sites of infection

Systemic (deep) fungal infections ?

- affect the whole body or an organ - Goes through the circulation and can infect any part of your body.

Polyenes (AmB)?

- binds to ergosterol and disrupts cell membrane

What echinocandins are we going to learn about?

- caspofungin - micafungin - anidulafungin

Opportunistic Pathogens?

- cause systemic or nonsystemic infections in compromised hosts 1) Aspergillus: Most of the hospital intensive care fungal infections is caused by Aspergillus. 2) Cryptococcus: Cryptococcus is responsible for the major form of fungal meningitis. 3) Rhizopus

Echinocandin (Caspofungin)?

- disrupts cell wall by inhibition of β(1-3) glucan synthesis

Allylamides?

- ergosterol synthesis inhibitors. - Disrupt cell membrane

Class IV: Allylamine?

- highly lipophilic synthetic molecule - in clinics since 1990s - Another orally available agent but the spectrum of activity is only against dermatophytes., nothing against systemic fungal.

Azoles?

- interrupts sterol synthesis in cell and mitochondial membranes

Ketoconazole (Nizoral®)?

- is an early Azoles: imidazole - represents older imidazole antifungals - first oral antifungal introduced into clinical use - wide spectrum of adverse effects and drug interactions (due to inhibition of mammalian cytochrome P450s), higher than all of the triazoles (makes it undesirable to prescribe). - largely replaced by newer triazoles for systemic use - only imidazole still used systematically in a limited clinical scope - also used topically for superficial fungal infections

Tavaborole (Kerydin®, others) ?

- not yet approved in Canada - MOA: Inhibits fungal tRNA(leu) synthetase (and therefore inhibits protein synthesis • Effective in the treatment of onychomycoses (fungicidal), due to the agent's ability to penetrate nail bed • data from clinical studies showed effective fungicidal activity within 5 days of topical applications

Where do dermatophytes infections occur?

- produce localized infections of keratinized, non-viable tissues (stratum corneum, nails, hair)

Class II: Azoles?

- synthetic compounds all containing a 5- membered azole ring - in clinics since 1990s - R represents side chains on the molecule. - Side chains change the PK and distribution parameters of the patient. - The only structure in the drug molecule that is important for its mechanism of action is the five membered azole ring . - When there are two nitrogen on this five membered ring = imidazole. - But when there are 3 nitrogens in the structure is considered a triazole.

Other systemic antifungal agents:?

1) Flucytosine (Ancobon®) 2) Griseofulvin (Grifulvin®)

Any chemotherapy or mechanical therapy against fungal organisms can have two outcomes?

1) Static reaction 2) Cidal reaction

How are fungal infections classified?

1) Systemic (deep) fungal infections 2) Non-systemic fungal infections a) Mucosal fungal infections b) Superficial fungal infections

Common dermatophytes infections?

1) Tinea pedis (athlete's foot), 2) Tinea cruris (jock itch), and 3) Onychomycosis (fungal nail infections)

Allylamine 1: Terbinafine (Lamasil®)?

ADME • Orally available • biotransformation mediated by multiple CYP-P450 enzymes • accumulates in skin and nails Adverse effects • Well-tolerated, GI upset and headache occur rarely • Rarely causes idiopathic liver toxicity: Patients at risk of hepatotoxicity should have liver enzymes monitored

Echinocandin 1: caspofungin?

ADME: Very poorly absorbed after oral administration, only available for IV administration Extensive hepatic and non-enzymatic biotransformation Limited distribution to the CNS Adverse effects: (low rate) Generally well-tolerated Minor GI upset and flushing Therapeutic Uses: • Active against mucocutaneous and systemic candidiasis • Also approved for use in patients with invasive Aspergillis infection who have failed to respond to Amphotericin B • not useful in meningitis or endophthalmitis

Non-systemic (Superficial) fungal infections?

Can be classified as either: a) Mucosal fungal infections: b) Superficial fungal infections: Superficial infections do not usually have the same severity to the person as systemic infection.

Amphotericin B (Fungizone®)?

MOA: binds to ergosterol and forms pores in the membrane - Binds to ergosterol in fungal cell membrane. The hydrophobic (double bond side) of Amp B binds to the ergosterol. - Ergosterol is not static in the cell membrane, it is like a sea of proteins. - So Amphotericin B can move the ergosterol around to find another hydrophobic structure, which is another Amphotericin B hydroxyl side. -All the Amphotericin B with the ergosterol molecules gets together and forms a pore (a central channel like structure) on the cell membrane of the fungal organism - Forms pores in membrane, altering cell permeability -Results in leakage of ions and macromolecules, changing osmolarity and eventually resulting in cell death - Selectivity results from higher affinity for ergosterol than cholesterol. Binds to ergosterol with much higher affinity than cholesterol.

Adverse effects of azoles?

Relatively nontoxic as a group • most common adverse effect: mild GI irritation • may cause elevation of liver enzymes, but rarely hepatitis BUT, drug interactions are common (due to inhibition of mammalian CYP-P450s). Some congeners are likely teratogenic (can cause harm on pregnant women).

FKS?

The B(1-3) glucan synthase has a component called FKS, FKS is inhibited when there is no B(1,3( glucan being made.

What is another potenital target for antifungal medications?

The inner most layer of the cell wall is a keratinized layer known as chitin, the enzyme involved in chitin metabolism is being targeted as a new drug therapy. Right now it is in clinical trials.

Summary of the uses of topical antifungals?

These are all the available topical agents that we have talked about so far.

Simplified pathway of ergosterol synthesis + inhibition points of antifungals?

This is a very simplified version of what the ergosterol synthesis pathway looks like. We are not expected to have to memorize the full synthesis pathway. Here are the most important steps because they are considered drug targets.

Almost all dermatophytes belonged to what 3 genera?

Trichophyton, Microsporum, Epidermophyton

What are adverse effects of Amphotericin B (Fungizone®) due to?

due to interaction of AmB with cholesterol in mammalian cell membranes

The term for a fungal cell dying from mechanical stress ?

fungicidal.

Superficial fungal infections?

infections of the body surface keratinized tissues (skin, nails, hair)

Ciclopirox olanine?

inhibits cellular transport

Tolnaftate?

inhibits ergosterol synthesis

Topical allylamines (terbinafine, naftifine)?

inhibits ergosterol synthesis

Topical azoles (clotrimazole, econazole)?

inhibits ergosterol synthesis

Selenium disuphide?

inhibits mitosis

Tavaborole?

inhibits tRNA(leu) synthesis

Mannoprotein is important in?

mannoprotein is important in immune recognition.

Griseofulvin (Grifulvin®)?

oral fungistatic compound MOA: inhibits mitotic spindle assembly - Binds to keratin in newly forming skin, protecting from infection - Must be administered for 2-6 weeks (skin and hair) or up to 18 months (nails) Adverse effects: headache, hepatitis, increases metabolism of warfarin and phenobarbital Therapeutic Uses: only use is in the systemic treatment of mycosis of nail and keratinized tissues Largely replaced by new antifungals such as terbinafine

Topical allylamines?

terbinafine, naftifine, butenafine

Undecylenic acid?

unknown MoA

Therapeutic uses of Terbinafine (Lamasil®)?

• Available in oral formulation for use in the treatment of dermatophytoses, particularly onychomycosis •Treatment for 6 weeks (fingernails) to 12 weeks (toenails) results in up to 80% cure rate • Also available as cream, for topical use in skin infections (e.g. ringworm, jock itch)

Topical Echinocandin: CD101?

• Echinocandin use is limited by low chemical solubility and unstable chemical structures. (limitation to developing topical echinocandin) - So finding a formulation that can deliver this echinocandin topically is a challenge in the drug delivery field. • newer congeners were designed to overcome such physical limitations • CD101 is currently under fast-track status by the FDA, but not yet available in Canada. Only in clinical trials and pre-clinical development.

Examples of primary pathogens?

• Examples of systemic pathogens: a) Histoplasma capsulatum - dimorphic fungi - NA prevalence concentrate in Ohio and Mississippi b) Coccidioides immitis - dimorphic fungi, - NA prevalence in southwestern USA, - Valley Fever: Causes this, when in hot desert environment stays dormant, but when it rains they are woken up and start an endemic "valley fever" These are two examples of primary pathogens, both are geographically restricted. In most circumstances they are laying dormant in the soil and when there are changes in the environment, like development, storm, fire. These fungi become living or active. Once they become active in division they will become airborne and people can breathe them in or get into food that people consume.

Echinocandin 2: micafungin?

• IV administration, adverse effects are rare but may include immunosuppression • Extensive hepatic enzymes dependent biotransformation. - Because extensive hepatic metabolism may interfere with other drug metabolism. • Example of interfence: Increase serum levels of nifedipine (Calcium channel blocker, use against hypertension) • Approved for use in Canada against Candidiasis

Echinocandin 3: anidulafungin?

• IV, minimal adverse effects • Metabolism is largely non-enzymatic (spontaneous degradation to open-ring peptide). So not a lot of drug interactions. • no significant drug interactions • Approved for use in Canada against Candidiasis

Allylamine MOA?

• Inhibits squalene epoxidase, leads to accumulation of squalene (toxic) and disruption of ergosterol synthesis • Fungicidal, and fungistatic in some candida species that are able to tolerate high levels of squalene

Itraconazole (Sporanax®)?

• Is a triazole • Limited oral bioavailability, absorption increased by food and low gastric pH • Penetrates poorly into CNS • Large number of drug interactions (e.g. decreasing metabolism of warfarin, omeprazole, digoxin) • Broader spectrum than fluconazole, used in systemic infections by histoplasma and blastomyce. • may also be used in dermatophytoses, including onychomycosis

Posaconazole (Spriafil®)?

• Is a triazole • Well absorbed orally • May result in hepatotoxicity and increase risk of arrhythmia • Also has a number of drug interactions with CYP3A4 and other P450 enzyme substrates (drug interactions profile is more than fluconazole, but less than itraconazole or voriconazole) • approved for prophylaxis against Candida and Aspergillus in immuno-compromised (hematological stem cell transplant recipients with GVHD and patients of hematological malignancy) individuals at risk of systemic infections • broadest spectrum triazole, breakthrough drug in 2007, also in clinical trials against Chagas disease Same adverse affects as voriconazole but broader spectrum amongst triazole, why its commonly used.

Fluconazole (Diflucan®)?

• Is a triazole • higher oral bioavailability than other azoles, administered IV, excellent CNS penetration • lowest incidence of adverse effects among azoles (Interacts with the least number of cytochrome p450s): - least effect on hepatic microsomal enzymes - better GI tolerance - long-term/high dose usage may be teratogenic • narrowest antifungal spectrum of the azoles, but is effective against many Candida species and in cryptococcal meningitis.

Drug resistance of Echinocandins?

• Low incidences but increasingly observed mutations of drug target FKS (β(1-3) glucan synthase)

Undecylenic acid (Desenex® in various forms) ?

• MOA is unknown • present in human sweats • Fungistatic • Active against a variety of fungi • Used in the treatment of various dermatophytoses (e.g. athlete's foot)

Tolnaftate (Tinactin®, others)?

• MOA: Inhibits squalene epoxidase (and therefore ergosterol synthesis) targeting the same enzyme as allylamines • Effective in the treatment of most cutaneous mycoses (fungicidal), but not those caused by Candida (fungistatic due to tolerance against squalene accumulations)

Ciclopirox olamine (Loprox®)?

• MOA: not clear, may inhibit fungal intracellular transport • Has broad spectrum fungicidal activity • Exclusive topical agent that is not formulated from a systemic agent. • Also has antibacterial and anti-inflammatory activity • Available as cream or lotion for tinea infections, and cutaneous Candida

ADME of Amphotericin B (Fungizone®)?

• Nearly insoluble in water, not absorbed on oral administration. So has to be IV administration. • Formulated for IV administration by complexing with deoxycholate (bile acid) • Widely distributed except to cerebrospinal fluid. IV inhection not useful in CNS infection • Binds to plasma proteins, accumulates in tissues • Slow renal elimination

Topical Polyenes - Nystatin (Mycostatin®)?

• Polyene macrolide antifungal agent, produced by Streptomyces noursei. • too toxic for parenteral administration • Well-tolerated on topical use • Available in creams and ointments, for use in mucocutaneous Candida infections (particularly oropharyngeal thrush, vaginal candidiasis)

When are topical antifungal agents for skin and mucous membrane fungal infections used?

• Topical agents are preferred treatment for cutaneous fungal infections because of the low risk for adverse side-effects • Not usually useful in mycoses affecting nails (onychomycosis) and hair (tinea capitis). Hard to apply drug to these areas. • Most topical agents are very poorly absorbed • Adverse effects are minor and localized at the site of application. Formulated to be applied to the exact site where the fungal infection occurs, instead of having to use the systemic circulation bring it where it needs to be. Because these drugs never get into the systemic circulation you can use much higher concentrations. Toxicity to the human cell stops being a concern.

Primary pathogens?

• highly pathogenic • capable of establishing an infection in all exposed individuals Those if us who come into contact with these pathogens, there is a high chance we will become infected. Luckily most of these primary pathogens are geographically restricted.

Therapeutic use of azoles?

• important alternative to AmB in the treatment of systemic fungal infections because its orally available. • Spectrum of antifungal activities is congener dependent.

Azoles and drug resistance mechanisms?

• increase drug efflux • mutations of the drug target, Erg 11 • increase expression of Erg11 • replacement of ergosterol with 14α-methylfecosterol (crossresistance with polyene)

Azoles - ADME?

• major family of orally available antifungal agent • bioavailability depends on individual formulation (refer to drug table) • CNS availability also differs for individual agent (implications on the treatment of CNS infections) • most azoles underwent extensive hepatic metabolism at by hepatic enzymes before renal or GI elimination

Acute and chronic dermatophytes infections?

• mostly minor illness and are self-limiting if cells eliminated quickly • can become chronic in tissues that turnover slowly (e.g. toenails, soles of the feet)

Amphotericin B (Fungizone®) and Infusion reactions?

• occur in most patients • major symptoms: fever and chills • end spontaneously in several hours • various approaches to reducing severity: - slow infusion rate, reduce dose - pre-medication with acetaminophen, antihistamines and/or corticosteroids

Class III: Echinocandins?

• semi-synthetic cyclic peptide linked to long chain fatty acid (lipopeptides) - Produced in fermentation reactions from a fungal species. • newest class of antifungal agents, in clinical use since 2000s