Med chem of antivirals Flashcards
enveloped viruses
surrounded by lipids
HIV, influenza, herpes
non-enveloped viruses
picornavirus, adenovirus
generic virus life cycle
viral attachment and entry penetration uncoating early protein synthesis nucleic acid synthesis late protein synthesis and processing packaging and assembly viral release
human immuodeficiency virus
Enveloped retrovirus
-Genus lentivirus
-Targets CD4+ T cells, macrophages, and dendritic cells
-Directly and indirectly destroys CD4+ T cells
-Causes immunosuppression
-Pathology associated with opportunistic infections
and cancer
-HIV-1 is more virulent and more easily transmitted - Responsible for most infections
-HIV-2 - primarily found in West Africa
key steps in HIV life cycle
initial binding
uncoating of the viral RNA
reverse transcription (viral RNA - proviral DNA)
integration of proviral DNA into the host genome
transcription of proviral DNA to form mRNA
translation of viral mRNA to form viral polyprotein
cleavage of the viral polyprotein to form the mature proteins of the virus
assembly and budding
HIV fusion/entry mechanism
HIV gp120 binds to CD4 on target cell
Conformational changes in gp120 exposes region
Exposed region binds to cytokine receptor (CCR5 or CXCR4 depending on the
strain of HIV)
Normal chemokine receptor ligands can interfere with HIV binding and fusion
maraviroc
HIV entry inhibitor
Selective CCR5 antagonist
Binds to CCR5 and causes a conformation change that prevents gp120 binding
No effect on cell surface levels of CCR5 or on CCR5 signaling
Can only be used in patients with HIV strains that utilize CCR5*
-Potential problem: May select for viral mutants that bind to CXCR4, Almost all transmitted HIV isolates bind CCR5, Rest either bind to CXCR4 only or to both co receptors, In ~60 % of patients, strains that bind CXCR4 emerge late during infection - Emergence is correlated with rapid disease
progression
fusion inhibitors
Enfuvirtide (T20)
36 amino acid peptide that inhibits fusion of virus membrane with cell membrane
Active only against HIV-1
Must be injected subcutaneously
resistance: Acquired resistance is due to mutations within a 10 amino acid motif in gp41: Forms part of the binding site of enfuvirtide, critical for viral fusion
-Easily acquired
reverse transcriptase inhibitors
Reverse transcriptase (RT) has three activities
-RNA dependent DNA polymerase
-Ribonuclease H
-DNA-dependent DNA polymerase
RT does the following
-Copies plus-strand RNA to produce minu sstrand DNA
-Degrades RNA template from RNA-DNA hybrid
-Synthesizes plus-strand DNA from minus-strand DNA template
NRTIs interfere with 1st and 2nd strand DNA synthesis
RT DNA polymerase activity
incorporation of dCTP into growing strand
- each nucleoside is phosphorylated 3 times
- different enxymes are used for each phosphorylation and for each nucleoside
- RT catalyzes formation of phosphodiester bond between 3’ OH of last nucleotide added and the 5’ phosphate of the next nucleotide. Pyrophosphate is released (provides energy needed for reaction)
Nucleoside RT inhibitors mechanism
NRTIs are nucleoside analogs that lack the 3’ OH
-Sometimes called false nucleotides
Two effects:
-Competitive inhibitor
of reverse transcriptase
-DNA chain terminator - inhibit elongation
Active against HIV-1 and HIV-2
NRTIs
7 NRTIs currently available for clinical use
Form backbone of initial therapy
Used in combination - 2 NRTIs plus NNRTI or PI or integrase inhibitor
Some combinations of NRTIs work better than others
-Tenofovir and emtracitabine
-Abacavir** and lamivudine
All must be activated by cellular kinases to triphosphate form (or equivalent)
Each NRTI is phosphorylated to triphosphate
Use cellular enzymes
Compete with normal nucleosides and NRTIs that are analogs of the same nucleoside
Tenofovir disoproxilfumarate
Prodrug - converted to tenofovir (TFV)
Acyclic* nucleoside phosphonate* analog of adenosine
Requires 2 phosphorylation steps
Phosphonate cannot be cleaved by cellular esterases - catabolically stable
advantages:
-Long intracellular half life
-Different resistance profile – different mutations in RT confer resistance
-Retains some activity against mutant RT
-Highly selective for HIV RT over human cellular and mitochondrial DNA polymerases
problems:
-Plasma esterases can activate TDF to TFV
-TFV is eliminated by kidney
-Relative to other NRTIs, TDF treatment is associated with: greater loss of kidney function and a higher risk of acute renal failure, larger reduction in bone mineral density
tenofovir alafenamide (TAF)
Alternative tenofovir prodrug Activated by different pathway than TDF -Lower plasma concentrations of TFV -Increased accumulation in lymphocytes -Expected to have fewer side effects May be even better at targeting HIV -Better accumulation in lymph nodes -Higher intracellular concentrations Approved for use in Nov. 2015 as a component of Genvoya (elvitegravir, cobicistat, emtricitabine, and tenofovir alafenamide) **Different activation pathway of TAF allows for 10-fold lower dose compared to TDF**
Cathepsin A (CatA)
Carboxypeptidase present in lysosomes
Expressed all cells with high levels in lymphoid cells
removes the ester from TAF (converts TAF to TVF)
activation of NRTIs - summary
Nucleosides and NRTIs must be activated to their triphosphate forms by cellular kinases
Different kinases are responsible for adding the first, second and third phosphate to the same nucleoside
Different nucleosides require different kinases to activate them
A given nucleoside and its corresponding NRTI are activated by the same enzymes
NRTI analogues of the same nucleoside can compete with each other for activation
Activated NRTIs compete with dATP, dCTP, dGTP, and dTTP to be incorporated into the growing DNA chain by RT
Ratio of activated NRTI to dNTPs is important
NRTIs have a higher affinity for HIV RT than for cellular DNA polymerases
Some NRTIs inhibit mitochondrial DNA polymerase
TDF and TAF must be processed to TFV by cellular enzymes before phosphorylation
Tenofovir requires 2 phosphorylation steps
resistance to anit-HIV drugs
Why do resistant mutations arise so quickly?
-HIV polymerase is error prone
-RT inhibitors are unable to suppress viral replication > 90%
-Large amounts of virus present: 10^7 to 10^8 infected cells, Half-life of infected cells is 1 - 2 days, HIV-1 genome is 9,181 nt, HIV RT has an error rate of ~1/2000
Rate at which mutations appear is inversely related to the serum drug concentration
-Need to maintain drug levels above MIC
genetic barriers to antiviral drug resistance
Genetic barrier = ease with which drug
resistance develops
Low = easy for virus to become resistant
High = hard for virus to become resistant
Evolution of a viral population that overcomes the activity of a drug or drug regimen
Depends on:
-Number of mutations required
-Impact of each mutation on drug activity
-Impact of each mutation of virus replication (fitness)
-Number of pre-existing mutations in the viral population
Low genetic barrier – resistance develops easily
-single mutation with low cost to viral fitness
High genetic barrier – more difficult to develop resistance
Two common scenarios:
-A primary mutation that has a high fitness cost – i.e., makes the virus less able to replicate - one or more additional mutations needed to allow the virus to grow better
-Multiple mutations needed for complete resistance
Drugs with low genetic barriers can be used in combination to develop an effective therapy
resistance to NRTIs
Discriminatory mutations: Mutations that selectively impair the ability of reverse transcriptase to incorporate analogues into DNA
Excision mutations:
-ATP molecule mediates the removal (excision) of a nucleoside analogue after it has been incorporated
-selected by thymidine analogs AZT and d4T - TAMs (Thymidine Analog Mutations)
NRTI resistance - key points
Mutations are mainly near the RT active site, but can occur at more distant locations
Mutations can either help the RT to distinguish between normal dNTPs and NRTIs or can promote removal of NRTIs after they’ve been incorporated into the growing chain
Individual NRTIs have a low genetic barrier
Some mutations confer resistance to a subset of NRTIs, but make RT more susceptible to inhibition by others
-NRTIs in preferred combination for initial therapy take advantage of this phenomenon
AEs of NRTIs - mitochondrial toxicity
Mitochondrial toxicity:
NRTIs are selective for HIV RT over human DNA polymerase-a and –b
Some NRTIs inhibit human DNA polymerase-g
Leads to anemia, granulocytopenia, myopathy, peripheral neuropathy, and pancreatitis
Lactic acidosis and hepatic steatosis (buildup of fat droplets in liver cells)
-More common with d4T, ZDV & ddI
Lipoatrophy – loss of body fat
-Higher incidence with stavudine
AEs of NRTIs - hypersensitivity
Abacavir Hypersenstivity Reaction : Black Box Warning
Occurs in 4 - 8 % of patients - can be fatal
Symptoms: Malaise, Dizziness, Headache, GI disturbances
Discontinue immediately is symptoms develop - Prompt recovery
Highly associated with the HLA-B5701 allele
Testing for HLAB5701
is recommended before initiating treatment with abacavir
recommended combinations of NRTIs
Tenofovir and emtracitabine**
-TFV has long intracellular half-life
-Once daily dosing
-Equivalent efficacy to other NRTI combinations
-Less fat maldistribution
-Different resistant mutation profiles
Abacavir and lamivudine (or emtracitabine)
-If HLA-B*5701 negative
**Lamivudine and emtricitabine are interchangeable
-Have negligible side effects
-select for the M184V mutation that can confer improved susceptibility to zidovudine or tenofovir
antiretroviral medications - should not be offered at any time
Regimens not recommended:
Monotherapy** (except possibly zidovudine used to prevent perinatal HIV transmission)
Dual NRTI therapy (with no other antiretroviral)
3-NRTI regimen (except abacavir/lamivudine/ zidovudine and possibly lamivudine/zidovudine + tenofovir)
nonnucleoside RT inhibitors (NNRTIs)
Bind directly to site on RT
-Hydrophobic pocket near catalytic site
-Near, but distinct from that of NRTIs
-Binding affects flexibility of enzyme
-NNRTIs do NOT compete with nucleotides for binding - noncompetitive inhibitors
NNRTIs do not have to be phosphorylated
Block RNA- and DNA-dependent DNA polymerase activities
NNRTI MOA
In the presence of NNRTI, nucleoside triphosphate and template bind tightly
to RT, but nonproductively
Blocks polymerization
Single mutation in binding site can promote resistance
2nd gen NNRTIs
diaryl-pyrimidine–based molecule
Designed to be inherently flexible**
Can bind in multiple orientations
-Binds to mutants that are resistant to other NNRTIs
NNRTIs AEs
All NNRTIs: Rash, Drug-drug interactions
Nevirapine – hepatotoxicity (may be severe and life threatening), rash including Stevens-Johnson syndrome
Efavirenz - neuropsychiatric, teratogenic in nonhuman primates (FDA Pregnancy Category D)
NNRTI metabolism
All are metabolized by CYP3A**
-Efavirenz, nevirapine, and etravirine are moderate inducers
-Efavirenz and delavirdine are inhibitors of CYP3A4
-Etravirine inhibits CYP2C9 and CYP2C19
Potential for interactions with other drugs that are metabolized by CYP3A
-CYP3A4 inducers (rifampin) can reduce levels
-CYP3A4 inhibitors can increase levels
NNRTI summary
NNRTIs do not require activation, do not compete with dNTPs, and are not incorporated into DNA
NNRTIs do not bind to cellular DNA polymerases
Resistance to NNRTIs can be acquired through a single mutation
Mutations that confer resistance to NNRTIs do not cause resistance to NRTIs
integrase inhibitors
Raltegravir (Mk-0518) approved in 2007
Inhibits insertion of HIV DNA into the human genome
Most commonly reported treatment-related adverse effects were diarrhea, nausea, fatigue, headache, and itching
Orally available
Compatible with other antiretroviral drugs
***Does not interact with CYP450s
integrase function
Inserts HIV DNA into host cell DNA 2 steps: -3’-processing -Strand transfer Raltegravir inhibits the strand transfer step
raltegravir mechanism
Integrase uses divalent metal ions to catalyze insertion - Coordinated by 3 amino acids in active site
Raltegravir chelates both metal ions and stabilizes enzyme-DNA complex
integrase inhibitor examples
raltegravir, elvitegravir, dolutegravir
Integrase inhibitor (INI) resistance
Caused by primary mutations that reduce INI susceptibility
secondary mutations further decrease virus susceptibility and/or compensate for the decreased fitness
Low genetic barrier
-Dolutegravir higher
extensive but incomplete cross resistance
-Dolutegravir less affected
Elvitegravir
currently available only in coformulation with cobicistat (COBI), FTC, and TDF or TAF
cobicistat is not active against HIV
Role is to boost elvitegravir concentrations by inhibiting metabolism by CYP3A4
Dolutegravir
Long plasma half life (14h) - once daily dosing
No boosting
No interactions with CYP3A4
Higher barrier for resistance
HIV protease inhibitors
HIV protease is an aspartic protease - aspartic acid in active site
Dimer - active site formed at interface
Inhibitors are transition state mimetics**
-Peptidomimetic
-Nonpeptide compounds
HIV protease
Viral proteins aggregate at inner surface of cell membrane
One protein molecule draws the two viral RNA strands into a forming virus particle
Protease cuts itself free
Protease cuts the other enzymes from the long precursors and then cuts the precursors into four pieces
p17 remains attached to the membrane
p24, p9, and p7 form the
bullet-shaped inner core of the virion
HIV protease mechanism
Peptide bond cleavage is a hydrolysis** reaction
Protease catalyzes addition of water to the amide
Forms intermediate
Breaks down to the two products, a carboxylic acid and an amine
HIV protease inhibitors
Amide bond is replaced by non-cleavable linkages
All PIs except tipranavir** are considered peptidomimetics
Inhibitor binding causes a conformation change in protease - “flaps” close
metabolism of protease inhibitors
All are substrates and some are inhibitors of
CYP3A4
-High potential for drug interactions
-Can cause increases in levels of other CYP3A4 metabolized drugs
-Levels of PI can be influenced by other CYP3A4 inhibitors
-Interactions are complicated - Delavirdine increases indinavir and saquinavir levels; Efavirenz reduces indinavir and saquinavir levels
Saquanivir
First protease inhibitor approved by FDA
Contains hydroxyethylamine moiety in place of peptide bond
Low bioavailability as hard gel capsule (4%)
Soft gel capsule increases bioavailability 3-
fold
Ritonavir
Higher bioavailability - 75% - Increased with food
Most potent PI inhibitor of CYP450s
Sub-therapeutic doses of ritonavir can be combined with other PIs - PI boosting
Currently recommended initial treatment regimes all use ritonavir combinations
PI boosting
Low doses of ritonavir inhibit CYP3A4** -Block metabolism of other PIs -Increases serum concentrations - Lopinavir and tipranvir are not indicated for use except in combination with ritonavir -Increases trough levels -Reduces emergence of resistant viruses -Improves compliance - reduced dosing, fewer pills, eliminates food restrictions Disadvantages -Drug-drug interactions with ritonavir -Increased risk of hyperlipidemia
Atazanavir
Once-daily dosing and minimal lipid and glycemic effects
most potent protease inhibitor in vitro prior to darunavir
Different resistant mutation profile
Efavirenz and tenofovir reduce ATV concentrations
