Antivirals
Lecture 20
Biology W3310/4310
Virology
Spring 2016
You cant go back and you cant stand
still. If the thunder dont get you, then
the lightning will.
JERRY GARCIA
The Wheel (lyrics by Robert Hunter)
Vaccines can prevent viral disease
But they have modest or no therapeutic eect if
an individual is already infected (exception?)
Our second arm of antiviral defense is antivirals
Can stop infection once it has started
Despite 50 years of research,
our arsenal of antiviral drugs remains
dangerously small
Only about 100 antiviral drugs are available on the US market
Most against HIV, HCV, herpesviruses - Persistent infections
Why are there so few antiviral drugs?
Compounds interfering with virus growth can adversely
aect the host cell
- Side eects are common (unacceptable)
- Every step in viral life cycle engages host functions
Some medically important viruses cant be propagated,
have no animal model, or are dangerous
- HBV, HPV
- Smallpox
- Ebola, Lassa
An unappreciated third reason
may be the most important
A compound must block virus replication
completely! It must be potent
Many standard pharmaceuticals can be
eective if enzyme activity is partially blocked
Partial inhibition is not acceptable for an
antiviral drug - resistant mutants will arise
Makes drug discovery expensive
Principles of Virology, ASM Press
Another serious problem for antiviral
discovery:
Many acute infections are of short duration
By the time the patient feels ill, it is too late to impact
clinical disease
Antiviral drugs for these viruses must be given early in
infection or prophylactically to populations at risk
- Safety issues; giving drugs to healthy people not wise
No broad-spectrum antiviral agents are currently available
Lack of rapid diagnostic reagents has hampered
development of antiviral drugs
LJ1001, a broad spectrum antiviral
http://www.ncbi.nlm.nih.gov/pubmed/20133606
LJ1001, a broad spectrum antiviral
Antiviral history
The rst modest search for antiviral drugs occurred in
the early 1950s
- Chemists looked at derivatives of the sulfonamide antibiotics
- Synthesis of thiosemicarbazones active against poxviruses
- Smallpox was still a major threat after WWII
1960s and 1970s: blind screening programs to nd
chemicals with antiviral activity
- Spurred on by successes in the treatment of bacterial infections with
antibiotics
Blind screening
No attempt to focus discovery on a virus or a virus-specic
mechanism
Random chemicals and natural product mixtures tested
for ability to block replication of a variety of viruses in cell
culture systems
Hits, compounds or mixtures that block in vitro viral
replication; puried and fractions tested in various cell
culture and animal models for safety and ecacy
Promising molecules called leads were modied
systematically by medicinal chemists
- To reduce toxicity, increase solubility and bioavailability
- To improve other pharmacokinetic properties
Thousands of molecules were made and
screened before a specic antiviral was
even tested in humans
Considerable eort, very little success
One exception: Symmetrel (amantadine)
- Approved late 1960s for treatment of inuenza A virus infections
- One of three drugs now available for inuenza
Mechanism of action was often unknown or speculative
- Mechanism of action of Symmetrel deduced early 1990s
Antiviral discovery today
Recombinant DNA technology & sophisticated
chemistry make targeted discovery possible
Essential viral genes cloned, expressed in genetically
tractable organisms, puried, analyzed in atomic detail
Life cycles of most viruses known, targets for
intervention can be generalized
Modern technology allows inhibitors to be found even
for viruses that cannot be propagated in cell culture
Blind screening procedures are dead
Principles of Virology, ASM Press
The path of drug discovery
Will the compound get to the right place in
the body at the right concentration?
(bioavailability)
proof of principle Will the compound persist in the body long
enough to be eective? (pharmacokinetics)
Will the compound be safe? (toxicity and
specicity)
Principles of Virology, ASM Press
Signicant hurdles stand in the way of
nding eective antiviral drugs
It is not unusual for the cost to bring an antiviral drug to market to
exceed $100-200 million dollars! Principles of Virology, ASM Press
From drug discovery to the clinic
Principles of Virology, ASM Press
Mechanism-based screens
Principles of Virology, ASM Press
Cell-based screen
Principles of Virology, ASM Press
Antiviral screening
High-throughput: 10,000 compounds/day
Chemical libraries
Natural products
Combinatorial chemistry
Structure-based design
In silico screening
Principles of Virology, ASM Press
High throughput screening
Go to:
m.socrative.com
room number: virus
We have many antibiotics, but fewer antivirals. What
is a reason for the dierence?
1. Robotic screening is slow
2. There are few serious viral infections
3. Resistance is a problem
4. Antivirals must be potent
5. All of the above
1
Resistance to antiviral drugs
Resistance to any antiviral drug must be anticipated
- Viruses replicate eciently
- Modest to high mutation frequencies
Special concern during extended therapy for chronic
infections (HIV, HBV, HCV)
Viral mutants resistant to every antiviral drug in arsenal
have been detected
Disconcerting because antiviral arsenal is small
Dangers of drug resistance
Patient cannot be treated with same drug
If no other drug is available, infection cannot
be stopped
Genetic analysis of resistance provides insight
into antiviral mechanism
May reveal new strategies to reduce or
circumvent problem
Mechanisms of drug resistance
RNA viruses: error prone RNA polymerase, no
correction mechanism
One misincorporation in 104 - 105 nucleotides
polymerized (106 greater than host DNA
genome)
In RNA viral genome of 10 kb, this frequency
leads to one mutation in 1-10 genomes
Mechanisms of drug resistance
DNA viruses: most DNA polymerases can
excise and replace misincorporated
nucleotides
DNA viruses evolve more slowly than RNA
viruses because they have less diversity
Principles of Virology, ASM Press
Consider Acyclovir, the highly
eective,
anti-herpes simplex virus drug
A prodrug; a nucleoside analog
Many antiviral compounds are nucleoside
and nucleotide analogs
Principles of Virology, ASM Press
Acyclovir mechanism of action
Principles of Virology, ASM Press
Improving acyclovir
Valacyclovir (valatrex), an L-valyl ester derivative of
acyclovir, has markedly improved bioavailability
Ester is taken up after oral administration, acyclovir is
released when the ester is cleaved by cellular enzymes
Principles of Virology, ASM Press
Acyclovir-resistant HSV
Arise spontaneously during virus replication
Some mutants cannot phosphorylate the pro-
drug
- Mutations are in viral thymidine kinase gene
Some mutants cannot incorporate
phosphorylated drug into DNA
- Mutations are in viral DNA polymerase gene
Acyclovir-resistant HSV
TK mutants can be devastating in AIDS patients
- May cause disseminated disease
- Often resistant to other nucleoside analogs that require
viral TK (cross-resistance)
- Treat with Foscarnet, DNA polymerase inhibitor (side
eects)
DNA polymerase mutants may also be
resistant to Foscarnet: no treatment options
left
Symmetrel (amantadine)
Interacts with inuenza viral M2 protein (ion
channel)
Blocks entry of protons into virion, prevents
uncoating
Principles of Virology, ASM Press
Principles of Virology, ASM Press
Go to:
m.socrative.com
room number: virus
Resistance to which antiviral would involve amino
acid changes in a viral enzyme?
1. Acyclovir
2. Amantadine
3. LJ001
4. Penicillin
5. All of the above
2
Inuenza virus NA inhibitors
Principles of Virology, ASM Press
Inuenza virus NA inhibitors
Zanamivir Oseltamivir
(Relenza) (Tamiu)
Designed to mimic natural ligand, sialic acid
Closer inhibitor to natural compound, less
likely target can change to avoid binding drug
while maintaining viable function
How inhibitors of NA (Tamiu, Relenza) work
Circulating H1N1 and H3N2 viruses are largely resistant to
Adamantanes, not recommended for use
http://www.cdc.gov/u/weekly/index.htm
WIN compounds
Principles of Virology, ASM Press
Inhibitors of picornavirus uncoating
Pharmac. Ther.
29:287, 1985
New HCV drugs
Principles of Virology, ASM Press
miRNA target: HCV
http://www.nejm.org/doi/full/10.1056/NEJMoa1209026
HCV new drug pipeline
http://www.hcvdrugs.com/ Principles of Virology, ASM Press
Targets for intervention: HIV replication
Principles of Virology, ASM Press
The problem with AIDS therapy:
relentless viral replication for years
Azido-deoxythymidine (AZT) - rst HIV
drug
Initially discovered during screens for anti-
tumor cell compounds
Phosphorylated to active form by cellular
kinases
Chain terminator
Principles of Virology, ASM Press
AZT
Substantial side eects (unlike acyclovir)
Can be given orally, is absorbed rapidly, but
half-life is ~1 hr (degraded by liver enzymes)
Consequently patients dosed 2-3x daily
Short half-life, multiple dose regimen
problematic: resistant mutants will be selected
Resistance to AZT
Mutants resistant to AZT arose immediately after
drug was licensed
Single aa changes at one of four sites in RT
Altered RT do not bind phosphorylated AZT
New nucleoside analogs developed: Didanosine
(ddI), Zalcitabine (ddC), Stavudine (d4T), Lamivudine
(3TC)
This lead to combination therapy, use of two
antiviral drugs to combat resistance
Mutants resistant to two drugs arose <1 yr
Non-nucleoside RT inhibitors (NNRTI)
nevirapine delavirdine efavirenz
(Viramune)
11-cyclopropyl-4-methyl-5,11-dihydro-6H- dipyrido[3,2-b:2,3-e][1,4]diazepin-6-one Principles of Virology, ASM Press
Resistance to NNRTIs
Resistant mutants are selected rapidly
Amino acid substitutions in any of seven
residues that line binding sites on enzyme
confer resistance
Cannot be used alone for treatment of AIDS
Now used largely in combination therapy
Antiviral drugs that target HIV protease
HIV protease absolutely required for production of infectious virions
Principles of Virology, ASM Press
Antiviral drugs that target HIV protease
Key nding: HIV protease recognizes and cleaves small synthetic peptides
Peptidomimetic
Principles of Virology, ASM Press
IN inhibitors
Principles of Virology, ASM Press
Maraviroc: CCR5 inhibitor
gp120
Free binding site
wt gp120
receptor on CCR5
High anity
Binding site
Disrupted
MVC ( ) by MVC
Very low anity
Bound
to CCR5
Go to:
m.socrative.com
room number: virus
Which of the following targets for HIV antiviral
therapy inhibits the earliest stage of infection?
1. Nucleoside inhibitors
2. NNRTIs
3. CCR5
4. Integrase
5. Fusion
3
Combination therapy
HAART: HIV can be treated as a chronic disease
Target dierent mechanisms
One pill containing three inhibitors
Mathematics of drug resistance
Assume one mutation needed for drug
resistance
Mutation rate 1 every 104 bases polymerized
Each base is substituted in every 104 viruses
Each person makes 1010 new viruses/day
1010/104 = 106 viruses will be produced each
day with resistance to one drug
Do the math
Developing resistance to two drugs: 104 x 104 =
108
1010/108 = 100 viruses resistant to two drugs
per day
Resistance to three drugs: 104 x 104 x 104 = 1012
viruses needed
Remember replication is suppressed by drugs
Complera (Efavirenz, emtricitabine, rilpivirine) 2011, Stribild (Efavirenz, emtricitabine, cobicistat) 2012
ART saves
lives
Principles of Virology, ASM Press
There are 1016 HIV genomes on the planet
today
With this number of genomes, it is highly probable that HIV genomes exist that are
resistant to every one of the antiviral drugs that we have now,
or EVER WILL HAVE!
