Resistance to anti-HIV Medications
by Bob Munk, Ph.D.
How Do HIV Meds
Work in the Body?
When HIV multiplies inside
an infected cell, it relies on proteins called enzymes. Whenever
you see a word that ends with “-ase” you’re probably looking
at an enzyme. First, HIV uses reverse transcriptase to read
its own genetic code and copy it. This code is a set of instructions
for building a new virus. Next, HIV uses integrase to insert
a copy of its code into the infected cell’s own “code book”
in the cell nucleus. Now the virus can use the cell’s own
machinery to make copies of itself. New viral proteins get
manufactured based on the genetic code. Then HIV uses protease
to assemble these proteins into a new working copy of itself.
Our current anti-HIV drugs
don’t kill HIV. In fact, scientists haven’t figured out how
to kill any virus yet. Instead, they design drugs that make
it harder for the virus to multiply. So far, the drugs we
have to fight HIV block, or inhibit, a specific enzyme. Two
types of drug—the nucleoside analog reverse transcriptase
inhibitors (also called nukes), and the non-nucleoside reverse
transcriptase inhibitors (non-nukes or NNRTIs)—block the reverse
transcriptase enzyme. A third type of drug, protease inhibitors,
blocks the final step of viral assembly that depends on the
protease enzyme.
The HIV genetic code is like
a string of beads called nucleosides. There are just four
different nucleosides, like four colors of beads, that get
put together in different combinations. The viral nucleosides
are adenosine (A), guanosine (G), thymidine (T) and cytidine
(C). It takes three nucleosides to define a specific amino
acid. Amino acids are the basic building blocks of all life.
When HIV—or a human cell—multiplies, the genetic code gets
“read.” Amino acids are created according to the genetic code
and assembled into proteins to make a new cell, or a new virus.
Nucleoside analog reverse
transcriptase inhibitors (NRTIs)
When reverse transcriptase
(RT) reads HIV’s genetic code, it goes one nucleoside or “bead”
at a time. It identifies which nucleoside it is (which color)
and it looks around inside the cell for that nucleoside’s
“mate.” Each nucleoside pairs off with just one other type
of nucleoside. “A” pairs up with “T,” and “G” pairs off with
“C.” When the RT enzyme reads an “A,” it looks for a “T” and
vice versa. When it reads a “G,” it looks for a “C” and vice
versa.
The nucleoside analog RT
inhibitors are fake nucleosides (“analog” means “something
similar”). The enzyme can’t tell the difference and picks
up a drug molecule instead of a real nucleoside. The fake
nucleosides stop the process of reverse transcription. It’s
kind of like a zipper, where reverse transcriptase is the
“pull” that combines the two sides. One side is HIV’s genetic
code, and the other side is made up of the nucleosides that
reverse transcriptase finds inside the cell. A “nuke” drug
molecule is like a bent tooth on the zipper, and the process
can’t continue.
Non-nucleoside reverse
transcriptase inhibitors (NNRTIs)
Researchers tell us that
the RT enzyme is shaped kind of like a catcher’s mitt. The
strand of HIV genetic code slides between the “thumb” and
the “fingers” of the enzyme, along the palm of the hand. The
NNRTI drugs block the RT enzyme by filling up the pocket in
the catcher’s mitt. When there’s a baseball in the mitt, nothing
can slide along the “palm.” The enzyme can’t read the genetic
code.
Protease inhibitors (PIs)
HIV uses the protease enzyme
late in its life cycle. After all the new viral proteins have
been built, they get assembled into a new virus that takes
shape and pushes out of the infected cell. The insides of
this new virus aren’t fully formed yet, and protease plays
a key role. It’s like a pair of chemical scissors that cut
long strands of protein into the correct pieces needed to
assemble the core of the virus. The protease inhibitor drugs
block the enzyme by locking in between the two blades of the
scissors so they can’t cut anything.
What is Resistance?
If reverse transcriptase
or protease got blocked 100%, HIV couldn’t make any new copies
of itself and your viral load would eventually decline to
zero. Unfortunately, anti-HIV drugs aren’t 100% effective
and HIV continues to multiply. Sometimes, HIV develops resistance
to a drug, or in other words, it keeps on multiplying just
like the drug wasn’t even there in the first place.
Resistance is a happy accident
from the virus’ point of view. When new viral copies are made,
they almost always contain errors: slight differences in the
genetic code that result in slightly different versions of
HIV. Most of these errors, or mutations, are fatal to the
virus and they die without multiplying. But sometimes, mutant
versions of HIV can not only multiply, but can ignore some
antiviral drugs.
For the nukes, we talked
about a zipper getting stuck when it hit a bent tooth. With
some particular mutations, HIV acts like a self-repairing
zipper. Reverse transcriptase becomes able to read “around”
the bent tooth—the nucleoside analog drug molecule—and continue
creating the code for a new virus.
For the non-nukes, if the
reverse transcriptase enzyme changes shape just a little bit,
it might “drop the ball” that we put in the catcher’s mitt.
The drug molecule might not be able to stay inside the enzyme
to block it, and RT can go ahead and read the viral genetic
code.
For the protease inhibitors,
if protease changes shape just the right way, the drug molecule
might not be able to stick in between the blades of the scissors.
Protease can go ahead and do its job of assembling a new virus.
Not every mutation can give
HIV resistance to drugs. It’s a random process. The more new
copies HIV makes, the more mutations there are, and the more
likely it is that some of them will give the virus resistance
to antiviral drugs.
Why Does Resistance Matter?
If your virus develops resistance
to a medication, it will keep multiplying even though you
take the drug. Resistance can cut down your possible treatment
options very quickly. For example, the latest version of the
treatment guidelines lists 78 different antiviral combinations
as “strongly recommended” or “recommended alternatives.” But
if your virus develops resistance to AZT (Retrovir), for example,
and you can’t use it any more, then there are only 39 remaining
combinations you could choose from.
AZT and d4T (Zerit), both
from the NRTI class, are somewhat cross-resistant. If your
virus becomes highly resistant to AZT, it’s probably at least
partly resistant to d4T. If you can’t use either AZT or d4T
in your regimens, then there are only 13 combinations to choose
from.
The current guidelines don’t
include tenofovir (Viread), which was recently approved. That
will add several more options for people who can’t use AZT
or d4T. The important point is, however, you don’t want to
run out of options and have to wait for a new medication to
get approved.
What Can I Do About Resistance?
When HIV multiplies, it mutates.
That’s just a fact of viral life. Once in a while, one of
those mutations will help the virus resist medications. The
more HIV multiplies, the more it mutates, and the higher the
risk of new resistance mutations showing up. The best way
to avoid resistance is to keep it from developing in the first
place. And the best way to do that is to keep HIV under control
so it has fewer chances to multiply.
To keep HIV from multiplying,
you should take anti-HIV medications according to their instructions.
When you do this, you should have enough of each drug in your
bloodstream to keep HIV under control. The manufacturers work
hard to figure out how much drug is needed to control the
virus without causing too many side effects.
If you miss doses or if you
don’t take them with (or without) food, according to the instructions,
there might not be enough drug in your bloodstream. When drug
levels drop, HIV can multiply more quickly, at least for a
while. More multiplication of HIV means more mutations and
a higher risk of developing resistance.
There have been some studies
showing that resistance develops even at undetectable levels.
Others show it can occur even with 100% adherence. Researchers
are searching for the mechanisms that allow this to happen.
In the meantime, what you can control is your adherence (as
long as you can tolerate your regimen). Research shows that
in the era of HAART (highly active antiretroviral therapy),
your first shot is your best shot. If you have a regimen that’s
working, be sure to stick to it!
Testing For Resistance
There are two main ways to
test HIV for resistance to drugs: genotypic and phenotypic
testing. Both types of test use a blood sample. In most cases,
the patient should have a viral load of at least 1,000 for
the tests to work properly.
Genotypic testing examines
the genetic code of the virus and looks for mutations. That
is, it looks for changes from the normal (or “wild type”)
sequence of nucleosides in the genes that contain the instructions
for the reverse transcriptase and the protease enzymes. Over
the years, researchers have studied strains of HIV that are
resistant to each of the anti-HIV drugs. They analyzed the
genetic sequences of the resistant virus and defined the specific
mutations that seemed to always show up in resistant virus.
Genotypic testing looks for these mutations.
Researchers developed a code
to identify specific mutations. Since it takes three nucleosides
to define a specific amino acid, they counted them in groups
of three (called “codons”) along the gene for either reverse
transcriptase or protease. For example, a particular mutation
at codon number 184 of the reverse transcriptase gene can
give HIV resistance to the drug 3TC (Epivir). This mutation
replaces the wild type (normal) amino acid, which is Methionine,
with a different amino acid: Valine. In research shorthand,
this mutation is the “M184V” mutation: instead of Methionine
at codon 184, there is Valine. With this one mutation, HIV
has a high level of resistance to Epivir.
A single mutation can also
give HIV resistance to all of the NNRTIs: the K103N mutation.
Just like the previous example, “K” and “N” are codes for
amino acids. Instead of the genetic code for “K” at position
103 of reverse transcriptase, we find the code for amino acid
“N”. When the same mutation or group of mutations makes HIV
resistant to more than one drug, those drugs are called “cross-resistant.”
For example, the PIs Crixivan (indinavir) and Norvir (ritonavir)
are cross-resistant. All of the NNRTIs are cross-resistant.
It’s not always this clear
whether HIV has resistance to a certain drug or not. For most
of the protease inhibitor drugs, HIV has to get several mutations,
one after another, before it develops resistance.
The resistance test report
for genotypic testing is a list of mutations found in the
sample of HIV. Those mutations that are believed to cause
resistance to specific drugs are highlighted, and the report
usually indicates whether the virus is believed to be resistant
or sensitive to each anti-HIV drug. Unfortunately, when the
virus needs multiple mutations to develop resistance, it’s
not always clear whether it’s resistant or not. Genotypic
testing cannot tell you “how resistant” the virus is to any
particular drug, but resistance is not an all-or-nothing thing.
HIV can be sensitive to a drug (no resistance), slightly resistant
(the drug still works, but not as well as against wild type
virus), or highly resistant (the drug doesn’t slow HIV down
at all). In some cases, mutations can make the virus hypersusceptible:
a drug might work even better than against the wild type virus.
Phenotypic Testing is the
second main type of resistance testing. Instead of the genetic
code of the virus, it looks at how fast the virus actually
multiplies when each anti-HIV drug is present. A range of
doses of each drug is added to individual test tubes containing
cultures of either the sample virus or of “wild type” virus.
After a certain amount of time, the amount of the sample virus
in each test tube is measured and compared to the amount of
“wild type” virus. If there are more copies of the sample
virus than the wild type virus, it has resistance to the drug.
Resistance is reported as
“fold change” in a phenotypic test report. This tells you
how many times more copies there are of the sample virus compared
to the wild type virus. For example, if there are 500,000
copies of the wild type virus, and 2 million copies of the
sample virus, then it has “4-fold” resistance to the drug
being studied. Although this is easy to understand, it’s not
clear what it means. For some drugs, 4-fold resistance means
that the drug won’t work at all. For other drugs, 10-fold
resistance means that the virus is still sensitive to the
drug. There are “cut-off” levels for fold resistance used
in phenotypic resistance reports. They are different for each
drug, and are somewhat different for each company.
Which test is better?
Genotypic testing is indirect.
It analyzes the genetic code of the virus, and reports on
mutations that researchers have found to be related to resistance
to particular drugs. It can be difficult to decide whether
a certain collection of mutations means the virus is resistant
or not. Genotypic testing has to be interpreted using a set
of rules, and each company doing resistance testing might
use a slightly different set of rules for its reports. Also,
the rules keep changing as researchers learn more about exactly
which combinations of mutations are the most relevant to resistance
for each drug. Genotypic testing is faster (about one week)
and less expensive than phenotypic testing.
Phenotypic testing is a direct
measure of how the virus behaves in the presence of anti-HIV
drugs. The report is easy to understand: it tells you how
much “fold resistance” the sample virus has to each drug.
However, it can be hard to know what level of fold resistance
really matters. Phenotypic testing usually takes about two
weeks and is more expensive than genotypic testing.
New types of resistance
tests
The company Tibotec-Virco
provides the “Virtual Phenotype” resistance test. It’s priced
between genotypic and phenotypic tests and the results come
back faster than phenotypic testing. First the virtual phenotype
does a genotypic test. Then, instead of using a set of rules
to interpret the list of mutations found in the sample, the
results are compared to a large database of paired genotypic
and phenotypic test results. The test report tells you the
phenotypic test results of samples in the database with similar
mutation patterns.
ViroLogic provides the “PhenoSense
GT” test. It’s not really a different type of test. However,
for doctors who prefer to see both genotypic and phenotypic
test results, it uses a single blood sample to run both tests
and provides the test results in a side-by-side format.
Weaknesses of resistance
testing
There are other problems
with resistance testing besides the difficulties in interpreting
results.
- Neither test can detect “minority”
strains that make up less than about 20% of a patient’s
population of viruses.
- They cannot detect resistance
that might be “hiding” in resting T-cells or other viral
reservoirs, and some researchers believe that this “archived”
resistance can re-emerge quickly if it will help the virus
survive particular drugs.
- When someone stops taking antiviral
medications, the drug-resistant virus has no survival advantage,
and the wild type virus is likely to re-emerge and become
the most common strain. The tests may not detect any resistance
if the patient has been off medications for more than a
couple of weeks.
- The rules for interpreting genotypic
tests, and the fold change results for phenotypic tests
are all based on just one drug at a time and may not accurately
reflect what’s going to happen with a combination of anti-HIV
meds.
- Some doctors use both genotypic
and phenotypic tests to get a more complete picture of what’s
going on, but often the tests give conflicting results that
can be very confusing to interpret.
Does resistance testing
help?
Several clinical trials studied
whether doctors who had the results of resistance tests made
better treatment decisions for their patients. The doctors
just made their treatment decisions the normal way (without
resistance test results), or they got genotypic and/or phenotypic
test results. Sometimes they got expert advice on how to interpret
the resistance test results.
In most of the trials, resistance
test results led to viral loads about .5 log lower than those
without. That’s a significant difference. Unfortunately, not
every trial showed the same results, and in some cases it
was difficult to tell whether it was resistance test results
or expert advice that made the most difference. Although there
are still some challenges in using resistance test results,
most AIDS physicians believe that they help make better treatment
decisions.
Who should get a resistance
test?
Treatment guidelines recommend
resistance testing in the following situations:
- When antiviral treatment stops
working: if viral load rises rapidly or CD4 cell count drops.
- When antiviral treatment isn’t
working well enough: viral load doesn’t become undetectable
within a month or two.
- Pregnant HIV-positive women.
Guidelines suggest that resistance
testing be “considered” for newly-infected people. Several
research studies have documented an increasing rate of new
infections with strains of HIV that are already resistant
to one or more antiviral drugs. Resistance testing might help
doctors avoid prescribing drugs that won’t control a patient’s
virus. Using the wrong drugs could allow HIV to multiply and
develop additional resistance.
Probably the most controversial
in terms of resistance testing are people not taking antiviral
medications who have been infected for several months or more.
As mentioned above, if you’re not taking antiviral medications,
the wild type virus will usually multiply the fastest and
become the dominant strain in your body. However, some researchers
believe that certain mutations can persist and be detected
for several months or maybe even longer. There hasn’t been
a lot of research on this question yet.
The take home message is
to prevent resistance in the first place. The best way to
avoid developing resistance is to take your anti-HIV medicines
on schedule and as according to instructions. If you have
a regimen that is working for you, then stick to it!
For additional discussion
on drug resistance also see Medicine
Chest: Once Again, One a Day in this issue.
Bob Munk is the Coordinator
of the New Mexico AIDS InfoNet at www.aidsinfonet.org,
and is a frequent writer on AIDS treatment topics. He tested
HIV positive in 1987.
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