Signals

CH I.RON

Milestones in Editors' Note-Viral Load in Review
Viral Load Testing

i V

1989
First studies in the New England
Journal of Medicine link high levels
of infectious HIV in the plasma of
AIDS patients and poor patient outcomes.'2
1990-1993
Studies confirm inverse correlation
between viral load and CD4+ cell
counts but show this relationship
does not apply in all patients.3,4
" Antivirals reduce viral load.34
~ Branched DNA (bDNA) signal
amplification used to quantify HIV
levels.5
HIV-1 RNA quantification, using
bDNA technology, correlates with
clinical progression.6
Pharmaceutical companies emphasize viral load in clinical trials.
1994
ACTG monitors viral load in clinical
trials.
Viral burden is highly predictive of
AZT resistance and death in
patients on long-term zidovudine
therapy.'
The Chiron Reference Testing Laboratory opens in August and makes
the bDNA testing service available
to clinicians.
Dr. David Ho uses bDNA technology
to demonstrate that 9 out of 10
long-term non-progressors had low
levels of virus-below 10 KEq/mL.*n
1995
~ In two independent studies, Drs.
David Ho, Michael Saag and
George Shaw use bDNA technology, n ccuncticr,,with a potent protease inhibitor, to measure virus
levels. Both studies demonstrate
that the latent phase of HIV infection is actually a dynamic period in
which viral replication is continuous
and highly productive, driving the
rapid turnover in CD4
lymphocytes.,910
* ACTG studies demonstrate that
changes in viral load from baseline
predict antiviral effect and clinical
progression11
* Dr. John Mellors shows that bDNA
levels post-seroconversion predict

which patients will progress to
AIDS most rapidly. Patients with
bDNA levels >1 0 KEq/mL progressed to AIDS 10-11 times
faster than patients with viral burdens <10 KEq/mL.12
1996
A consensus panel recommends
using viral load in all patients to
guide the initiation of antiviral therapy, to identify the development of
drug resistance, and to guide
changes in therapy.13 (See article.)
Prospective studies demonstrate
that both baseline viral load and
changes in viral load in response to
therapy are accurate predictors of
clinical progression.14
HIV protease inhibitors, in combination with nucleoside analogs,
reduce viral load to below
detectable levels.14
~ Dr. John Mellors recommends
staging HIV disease using bDNA.14
Dr. Paul Volberding describes
bDNA as a relatively easy method
of viral load testing for labs to set
up and perform with less variability
from lab to lab than RT-PCR. He
also states that bDNA quantifies
certain HIV strains that go undetected with RT-PCR.15
References on page 5

Welcome to a special issue
of Signals.
Several drug companies recently announced that combination
therapy with new protease inhibitors can significantly reduce virus
levels in the blood-in some cases
to below detectable levels. The
emergence of these and other new
drugs, once again, highlight the
importance of monitoring viral
load.
Since we created Signals in
November, 1994, our goal has been
to keep AIDS-treating physicians
updated on new developments in

using viral load to individualize
antiviral therapy. In this issue, we
take a step back to look at what
has been learned about the clinical
utility of measuring viral load by
reprinting some articles from previous editions.
With this issue, we're also
expanding our distribution of Signals to pathologists. Pathologists
play an important role in treatment
decisions and evaluating new
methods for quantifying viral load.
What physicians want to know
In 1994, most AIDS-treating
physicians were just beginning to

hear about a new technology from
Chiron called bDNA signal amplification that can accurately and
reproducibly measure HIV levels in
a patient's blood.
The most common questions
we heard from physicians were
*What is the clinical utility of
measuring viral load?
How can viral load measurements help determine when to
initiate antiviral therapy?
How can measuring viral load
be used to guide decisions to
change antiviral therapy?
Continued on page 5

7 Things Pathologists Should Know

pith the growing importance of viral load testing,
many AIDS-treating
physicians are looking to pathologists to help them determine the
best method for monitoring their
HIV-positive patients. The pathologist is faced with a daunting task.
To date, no viral load assays have
been approved by the FDA, yet
there is a proliferation of reference
laboratories offering "HIV-1 RNA
Quantification."
For those of you who are just
now becoming familiar with these
new developments, here are seven
things every pathologist should
know about viral load assays.
1. Viral load is the most important laboratory test to monitor the progression of
HIV disease.
A recent study by Dr. John Mellors demonstrated "that direct

Hybridize
HIV-I RNA

HIV-I RNA
Target
Probes

bDNA
Molecule
Hybridize
bDNA
Molecule

Hybridize
Signalgenerating
Enzyme

Capture
HIV-I
RNA
Surface of Microwell
Capture HIV-1 RNA

Amplify Signal

Quantify HIV-1 RNA

Figure 1. By amplifying the signal rather than replicating the target sequence, bONA technology is
not subject to the errors inherent in target amplification methods like PCR.

quantification of HIV RNA by
branched DNA (bDNA) predicts
HIV progression earlier and more
accurately than CD4+ cell counts...
regardless of an individual's disease
stage at measurement.'
2. bDNA and PCR are the two most common methods of measuring viral load.
Branched DNA was designed
specifically to quantify nucleic ecid
levels in blood and has been used
in clinical trials and HIV research
since 1992. In 1994, the Chiron

bDNA-Important Tool in

New research strongly suggests that measuring HIV
RNA levels is a far better
predictor of disease progression
than monitoring immune cell
decline. These findings were presented at the Third Conference on
Retroviruses and Opportunistic
Infections, held in Washington D.C.
on January 28th-February 1st.
Stage HIV/infection with bDNA
Dr. John Mellors, Associate
Professor of Medicine at the University of Pittsburgh, recommends: "All treatment strategies
should now be based on an accurate staging of HIV disease." Mellors urged clinicians to stage HIV
disease as is done when treating
cancer, and stated, "The bDNA
technology provides us with a tool
to accurately stage HIV disease
progression."
Mellors' recommendations are
based on a study that looked at
samples collected from 181 HIVinfected individuals over ten years.
Results indicate that viral load can
help predict disease progression as
far as 10 years into the future.
These findings were confirmed in
a similar study using bDNA by
Cladd Stevens, MD of the New
York Blood Center.
According to Mellors, "You can
actually see a doubling of (viral) levels at least two years before you see
a 100-cell decline in CD4 levels."
Dr. Anthony Fauci, head of the
NIAID, commented that the

results are "impressive" and prove
that such viral measurements are
important.
Initiate combination therapy
immediately after infection
Dr. David Ho, director of the
Aaron Diamond AIDS Research
Center, presented astounding new
data on HIV replication. Pioneering the use of bDNA to analyze
the pathogenesis of HIV, Dr. Ho
discovered that infected cells
become factories for viral production. They produce 10 billion
viruses every 24 hours and, in the
process, CD4+ cells die but are
rapidly replaced.

Reference Testing Laboratory first
made the bDNA viral load testing
service available to physicians.
Now, physicians can submit specimens to the bDNA testing service
through SmithKline Beecham Clinical Laboratories and Corning
Clinical Laboratories.
PCR has been used extensively
in research and also in clinical triaV_ The technology is used by
hundreds of laboratories, and a
large number of these labs have
Continued on page 3
Staging HIV
According to Dr. Ho, the magnitude of viral production suggests
that "any one drug may be
doomed to fail," because HIV
mutates with each round of reproduction. This increases the chance
of drug resistance.
Dr. Ho believes that the key to
AIDS treatment is to initiate combination therapy, using protease
inhibitors, immediately after infection, and to monitor viral load to
gauge effectiveness of therapy.
New AIDS drugs reduce viral load to
undetectable levels
New drug studies, using several
Continued on page 3

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Update on using branched
DNA (bDNA) direct
quantification to measure
viral load

MARCH 1996

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Panel Recommends
Viral Load Monitoring
panel of leading AIDS-treating physicians has recommended the use
of new assays to measure HIV RNA levels in the blood of infected
patients as a way of optimizing antiviral therapy.
The Consensus Conference on Laboratory Markers in HIV Infection,
comprised of F Aluti, C. Boucher, F Dianzanl, E.M. Fenyo, D. Ho, C. Lane,
J. Lange, J. Montaner, G. Pantalec, L. Perrin, S. Romagnani, M. Saag, R.
Schooley, and S. Vella, assembled in Rome on January 16th to discuss the
clinical use of laboratory markers in the management of HIV infection. The
physicians focused on the utility of clinical markers in the documentation
and prediction of disease progression in HIV-infected individuals.
Panel conclusions
The pathogenesis of HIV infection is characterized by continuous viral
replication and rapid virus turnover, which ultimately leads to a significant decline in CD4+ cells and immunodeficiency.
Continued on page 4

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*HIV-1 RNA kilo equivalents/milliliter of plasma as measured by branched DNA (bDNA) direct quantification. A kilo equivalent - 1,000 molecules of HIV-1 RNA.