GMHC Treatment Issues 1993 Oct 1; 7(9): 13
Gene therapy is a new branch of medicine with revolutionary
potential. One type of gene therapy, called "gene transfer,"
implants new, functional genes into cells to alter their
function or confer resistance to infection. (See Treatment
Issues May, 1993) Gene transfer experiments are on-going in
several congenital diseases such as cystic fibrosis and ADA-
deficiency, as well as cancer and AIDS. Yet other gene therapy
strategies do not insert new genes into cells, but rather
attempt to inhibit or repress specific genes already in the
cell. Antisense, the first example of these strategies to enter
clinical studies, attacks the RNA, a crucial messenger in the
life cycle of HIV.
Background and Overview
All viral infections, including AIDS, can be described as
acquired genetic diseases. Viruses are packages of genetic
material that insert themselves into DNA, the double-stranded
chain of genes inside the nucleus of every cell, and transform
the cell into a factory for producing new copies of the virus.
Each sequence on the DNA provides the blueprint for the
production of a specific protein. Proteins are complex
structures which form the building blocks of all life. Human
DNA contains all the necessary information to produce the
proteins that compose our bodies. However, once infected by the
virus, the DNA also programs the cell to manufacture the
component proteins of HIV.
In order to produce proteins, the DNA must transmit its
information to a messenger RNA (mRNA) molecule, also known as
the sense strand. The mRNA uses this information to organize
the building and assembly of proteins into the finished product
-- usually essential cellular components but, in the case of
infected cells, new copies of HIV. An anti-sense drug is the
exact opposite (i.e. the mirror image) of a specific "sense"
strand. Antisense drugs attach to mRNA, and thereby block the
production of particular proteins at the genetic level.
Traditional drugs attack proteins, which, in the case of
anti-HIV treatments, include reverse transcriptase, protease,
and other familiar targets. However, proteins are large,
complex structures that are produced in massive quantities by
infected cells. In order to be successful, traditional drugs
must disable every copy of the protein. Traditional drugs often
attack healthy, normal proteins as well, and cause toxicities.
Antisense is an attractive option because each mRNA molecule
produces large numbers of each protein. Anti- sense technology,
by attacking a single mRNA strand which is responsible for
producing large quantities of single protein, may be a more
efficient way of eliminating large quantities of unwanted
proteins at once. Since an antisense drug only binds with its
exact opposite, it should be extremely specific, producing
Whereas proteins are large and complex, RNA is composed of
various combinations of just four well-known amino acids --
adenosine, cytosine, thymidine, and guanosine.
Antisense proponents have discovered that the real world does
not follow theory as quickly nor as easily as they would like.
As simple and elegant as it seems, antisense has not yet
translated into clinical reality. The technological challenges
are numerous. Since antisense drugs will be administered in
traditional ways -- intravenously or subcutaneously --
scientists face significant hurdles. The compounds must be
large enough to be extremely specific to the right piece of
RNA, but stable enough to avoid destruction by the body, and
small enough to reach and penetrate target cells. Chemists have
been modifying antisense compounds for years in order to solve
these problems, with as yet unknown success.
Even with an optimal compound, the therapy, in the case of HIV
at least, would have to be taken chronically, and probably in
high doses. Viral resistance, as with regular antivirals, could
theoretically develop. Furthermore, the expense could be
staggering because large-scale production of these compounds is
still costly and difficult.
For all these reasons, the first efforts with antisense drugs
have been for topical, rather than systemic, uses. The first
antisense drug to enter clinical trials was a therapy aimed at
genital warts caused by Human Papilloma Virus (HPV). Isis
Pharmaceuticals, a Seattle-based biotechnology company, began
this first antisense trial in 1992. Thus far, the company has
no evidence of efficacy, but reports about absorption and
safety are promising. Again, this is only topical use, and its
lessons for systemic HIV therapy are limited.
The First AIDS Trial
Hybridon, a Worcester, MA-biotechnology company, seems poised
to put the first systemic antisense drug into HIV-positive
individuals. GEM-91, its lead compound, blocks HIV's gag gene.
The gag gene of HIV is common to all retroviruses and encodes
for the critical core proteins of HIV, such as p9, p17, and p25
(the nucleiod shell). If gag, production is blocked, the
Hybridon investigators theorize HIV replication could be
dramatically slowed. Phase I trials are currently planned to
begin in United States at the University of Alabama at
Birmingham and through the ANRS, the French national AIDS
research network. The French study could enroll its first
patient in October.
Hybridon believes GEM-91 may have two unique features.
Laboratory studies indicate mRNA may not be the only target.
HIV, like all other retroviruses, enters the cell as a piece of
viral RNA (vRNA). GEM-91 may also inhibit vRNA before its
integration into DNA. This suggests that GEM-91 might interfere
with both early and late stages in HIV's replication cycle.
Hybridon also claims another advantage to its compound over the
traditional antisense model. Instead of merely disabling a
single RNA strand, its drug might be able to destroy many RNA
strands. According to Hybridon, GEM-91 first binds to the
target RNA strand, then activates a cellular enzyme (RNAseH)
which destroys the strand, leaving the drug free to attack more
Even if GEM-91 fails in humans, Hybridon is positioned to be a
leading player in antisense research. Hybridon has already
synthesized GEM-92, its second generation product. Also, the
company claims broad patent rights over all antisense
approaches to AIDS, although this has not been tested in the
In addition, Hybridon believes it has resolved manufacturing
issues. The company was recently awarded a patent that covers
new, more efficient methods of antisense production and has
started construction of a large manufacturing facility. The
company believes that it will have sufficient supply of the
compound to meet the needs of clinical research.
Since the genes of HIV have been extensively studied by
molecular biologists, researchers can design antisense
compounds aimed at specific mRNA molecules that produce
proteins essential to HIV's survival. Although very few
clinical trials have actually begun, and clinical efficacy is a
long way from certain, the pharmaceutical industry has devoted
significant resources to this burgeoning field. Its proponents
believe antisense will radically transform medicine and open up
the possibility for new treatments for AIDS and other viral
Other Gene Modulators in Development
Other gene therapy approaches similar to antisense are in more
preliminary stages of development.
Triple Helix DNA
An innovative twist on the antisense idea is the "Triple Helix"
technology being developed by several companies. Whereas
naturally-occurring DNA consists of two interlocking strands in
the famous double helix structure, scientist have developed the
means to attach a third strand of DNA to the double helix,
effectively blocking transcription. Now they are working to
synthesize small strands of DNA which are targeted against
specific sequences of the HIV genome. In the same way that
RNA-targeted therapeutics are more efficient than drugs which
bind with proteins, Triple Helix promises to be more efficient
than conventional antisense. The compound would bind to the DNA
itself, rather that to the thousands of mRNA transcripts. Thus
less drug would be needed, greater efficacy would be achieved,
and transcription of the unwanted gene would cease completely.
However, this technology is at an earlier stage of development,
and many biochemical obstacles remain.
Another promising variation on antisense are a class of
compounds called Ribozymes. These are naturally-occurring RNA
molecules which function as catalytic molecular scissors,
chopping up RNA strands at selected sites. Ribozymes can be
synthesized, and targeted against specific RNA sequences, just
like antisense compounds. However since ribozymes can
effectively destroy many targets, according to its proponents,
lower levels of drug might be needed, and therapy could be more
However, chemists have not yet figured out how ribozymes can
reach and penetrate cells in the body. Recently the RAC
approved the first clinical study of ribozymes (see page 6).
The investigator is Flossie Wong-Staal at the University of
California at San Diego. In the test tube, her so-called
"hairpin" ribozyme completely blocked HIV infection. In order
to overcome the pharmacological difficulties, Wong-Staal is
attempting a gene therapy double whammy: she will plant a gene
in the CD4 cells of the subjects that will produce the
One hope is the creation of a ribozyme which can cleave DNA,
rather than RNA, and thus could simply cut the viral genes out
of the host cell DNA. If such a compound were in existence, it
would be an extremely promising modality for HIV treatment.
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