New research that is bringing ribozyme therapy closer to clinical
trials was presented at the recent meeting of the American Association
for Cancer Research (AACR) by Kevin Scanlon, PhD, director of
Biochemical Pharmacology at the City of Hope National Medical
Center in Duarte, California, and editor of the journal Cancer
Gene Therapy. The presentation was entitled "Therapeutic
Applications of an Anti-Oncogene Ribozyme in Cancer."
The RNA enzyme is very precise and seeks out and cleaves mutated
RNA, halting its ability to launch a complex cascade of events
leading to cancer. Ribozyme therapy is one of the new gene therapy
strategies that holds promise for significantly improving treatment
of solid tumors, such as melanoma, colon, breast, lung, bladder
and pancreatic cancer, by targeting specific molecular sites within
cancer cells, resulting in more effective treatment and fewer
side effects than conventional chemotherapy and radiation therapy.
Until now, ribozyme studies have mostly been confined to tissue
cultures and animals; however, within the next year Dr. Scanlon
expects to begin clinical trials at San Diego Regional Cancer
Center, in collaboration with Robert E. Sobol, MD, and at the
University of California, San Francisco, in collaboration with
Mohammed Kashani-Sabet, MD.
Ribozymes are a class of small enzymes the normal function of
which is to cut out unwanted segments of RNA, a step in the synthesis
of protein from RNA. Research in the late 1980s led to the approach
of modifying ribozymes to cut oncogene RNA at the mutation sites,
thereby disrupting production of the oncogene protein products
but not affecting normal gene function. Researchers accomplished
this by designing ribozymes that bind and cleave only the mutated
portion of oncogene RNA. In addition, they attached a molecular
structure--"a hammerhead"--borrowed from plant viroid
ribozymes. Inside the cancer cell, the modified ribozyme binds
to the mutated RNA; immediately, the hammerhead "snaps"
and cleaves the oncogene RNA in two, preventing production of
the oncogene protein product.
Oncogenes contain mutations and contribute to cancer by expressing
mutated proteins. Like other genes, these oncogene proteins are
expressed in two steps: DNA is transcribed into mRNA, which, in
turn, controls protein synthesis. Ribozyme gene therapy differs
from other forms of gene therapy because it does not replace or
repair oncogene DNA, but instead works by inhibiting the second
step of this process--synthesis of the oncogene protein product
from mRNA. Describing the basic approach to using ribozymes in
cancer treatment, Dr. Scanlon notes, "We have to sequence
the oncogene DNA to define the mutation, and then design the ribozyme
to specifically cleave the mutation."
From the Lab to the Clinic
In his presentation at the AACR, Dr. Scanlon focused on currrent
research aimed at making ribozyme therapy more clinically relevant.
One of the tasks has been unraveling the complexity of oncogene
interactions. "Our early assumption was that if a ribozyme
targeted to a specific oncogene worked in one type of cancer that
it would work in all types of cancer, and in reality that's not
true," reports Dr. Scanlon. "Every time we treat a cancer,
we have to define which oncogene is most important--is it ras,
fos or one of the 50 or more oncogenes identified to date?
When we have identified the key oncogene(s), and prevented its
expression using ribozyme gene therapy, we know we can have an
impact on cancer cell growth."
In ribozyme gene therapy, as in other types of gene therapy, one
of the key challenges is delivering the therapeutic agent. "The
fundamental problem is delivery to all cancer cells, and expression
of the ribozyme at a high enough level for a long enough time
to be therapeutic." Currently, Dr. Scanlon's group is studying
three viral vectors--retrovirus, adenovirus, and adeno-associated
virus (AAV)--that deliver the ribozyme "package" by
infecting cancer cells. But because the viral vectors can infect
healthy as well as malignant cells, researchers are seeking ways
to ensure that the vectors selectively "target" cancer
cells in order to minimize possible damage to normal cells and
to design mechanisms for switching on ribozymes. This approach
depends on the identification of "promoters"--substances
unique to each type of cancer cell. The ribozyme package, which
includes the nucleic acid sequence of the ribozyme, is designed
so that the ribozyme is expressed only when the tissue-specific
promoter is present in the diseased tissue.