October 12, 2001
Revealing New Clues About the Spread of Colon Cancer
Researchers have discovered a gene that appears to allow colon
tumors to spread to other parts of the body, a process called
metastasis. The gene codes for an enzyme that may be central to the
metastatic process, suggesting the possibility that the enzyme could be
targeted by drugs to block the spread of colon cancers. Metastasis is
the primary cause of death from colon cancer.
In an article published online in the October 12, 2001, edition of
Science, Howard Hughes Medical Institute investigator Bert
Vogelstein and colleagues at the Johns Hopkins University Oncology
Center reported identifying the gene, called PRL-3.
“This gene was expressed at very low or undetectable levels in normal colon epithelium. It was expressed at low levels in the early stages of colorectal neoplasia. And its expression was clearly much higher in metastatic lesions from the liver.”
Bert Vogelstein
The researchers identified PRL-3 after developing a profile
of gene expression in cells they microdissected from cancers that had
metastasized to the liver. The genetic profile was developed using SAGE
(serial analysis of gene expression), a technique invented by the
researchers to determine the level of expression of genes. In SAGE, the
enzyme reverse transcriptase is used to produce complementary DNA from
the messenger RNA (mRNA) derived from cells under study. The DNA is
then snipped at a defined position, creating a unique identifier "tag"
that corresponds to a single gene. The researchers can then analyze the
number of unique tags present in their sample and deduce how much mRNA
exists for each gene — a measure of gene activity.
Vogelstein said that the group's initial analyses of gene expression
in the metastatic tissue yielded confusing results. "When we initially
took metastatic lesions from patients, purified the RNA, and then
looked at their gene expression profiles, we found many of the [gene]
transcripts were clearly derived from non-neoplastic cells," he said.
"The problem is that tumors are composed of multiple cell types; not
just the neoplastic cells. Liver metastases from colon cancers contain
a supporting stroma tissue, as well as inflammatory cells and normal
liver cells," he said.
The researchers devised a purification procedure that enabled them
to isolate only the metastatic cells. This technique involved
separating cells based on their attraction to an antibody specific for
colorectal epithelial cells in liver tissue, which constitute the
metastatic cancer cells. "Once we did this separation," said
Vogelstein, "the gene expression patterns became much clearer and more
reliable."
SAGE analysis of these isolated cells revealed 38 gene transcripts
that appeared to be enriched in the metastatic cells, indicating that
they were switched on during metastasis. Although all the genes were
elevated in some of the metastatic lesions compared to normal
colorectal epithelium and non-metastatic cancers, only one, called
PRL-3, was consistently overexpressed.
"We were surprised and pleased that several of the transcripts were
overexpressed in many metastases," said Vogelstein. "And it was
particularly striking that this one gene, PRL-3, appeared to be
quite consistently overexpressed."
Vogelstein and his colleagues were especially interested in
PRL-3 because it codes for an enzyme. "Most genes implicated in
solid tumors are tumor suppressor genes whose activities are switched
off during tumor formation. The proteins encoded by these genes are not
good targets for potential drug therapies because drugs generally
inhibit enzymatic activities. You cannot inhibit an activity that is
not present in the tumor cell because it has been inactivated. On the
other hand, there aren’t many enzymes that have been shown to be
overexpressed in cancers in a pathogenic way and related to
metastasis," said Vogelstein. PRL-3 codes for a tyrosine
phosphatase, an enzyme that likely controls the activity of other
proteins by removing a phosphate from them.
When the scientists quantified the level of PRL-3 expression in
normal, non-metastatic and metastatic tissues, they found clear
differences. "This gene was expressed at very low or undetectable
levels in normal colon epithelium," said Vogelstein. "It was expressed
at low levels in the early stages of colorectal neoplasia. And its
expression was clearly much higher in metastatic lesions from the
liver."
Importantly, the scientists were also able to compare PRL-3
expression in normal epithelium, primary cancers and metastatic lesions
from the same patients. Such comparison eliminates the potentially
misleading differences when tissues from different patients are
compared. "In each of the six cases studied, PRL-3 expression
was quite a bit higher in the metastases than in the primary tumor,"
said Vogelstein.
The scientists also discovered that the higher levels of
PRL-3 expression they measured were associated in a few cases
with a process known as gene amplification — in which overexpression
of a gene is caused by a large increase in the number of copies of the
gene. Gene amplification is a characteristic mechanism by which the
overexpression of growth-regulating genes occurs in human cancers.
"The discovery that this gene was not only overexpressed but also
amplified provided very strong evidence for causality of this gene in
the metastatic process," said Vogelstein.
According to Vogelstein, the scientists will now search for the
biochemical and physiologic roles of PRL-3 in colon cancer
metastasis. One of the goals of future studies will be to launch a
search for molecules that inhibit the phosphatase function of PRL-3 and
to discover whether this inhibition thwarts metastasis.
|
