Research underscores need to combine genomics and basic biology in cancer gene hunt
Lynda Chin, MD
Dana-Farber Cancer Institute scientists' discovery of a
cancer-causing gene — the first in its family to be linked to cancer —
demonstrates how the panoramic view of genomics and the close-up
perspective of molecular biology are needed to determine which genes are
involved in cancer and which are mere bystanders.
The findings are reported in the June 25 issue of the journal Nature.
"In the coming years, we can expect genomic studies [which chart the
activity of thousands of cell genes] to generate hundreds or thousands
of genetic elements of interest in cancer research," says the study's
senior author, Lynda Chin, MD, of Dana-Farber.
"To narrow that group to the genes that actually drive cancer growth
and metastasis, it's necessary to do functional studies, which focus on
what individual genes do to turn a cell cancerous, and mechanistic
studies, which examine how they turn cells cancerous and in what
setting. It is a long and intensive effort that will leverage knowledge
from different fields and different model systems."
In the study, Chin, lead author Kenneth Scott, PhD, of Dana-Farber,
and their colleagues worked their way through a series of experiments —
in yeast cells, multiple types of human cancer cells, laboratory cell
cultures, and mouse models — to demonstrate that a surplus of a gene
known as GOLPH3 can spur cancer cell growth in a variety of tissues.
It is the first gene associated with the Golgi complex, a tiny
packaging plant that prepares proteins for their journey within and
outside the cell, which has been found to play a role in cancer.
Chin's team also found that the protein made from GOLPH3 may
serve as a biomarker for tumors that can be effectively treated with
the chemotherapy drug rapamycin: tumors with a high level of the protein
are more apt to shrink in response to the drug than those with low
The study began with an observation made years ago that a section of
chromosome 5p13 is often duplicated, or amplified, in cancers of the
lung, ovary, breast, and prostate gland, as well as melanoma. The
presence of this abnormality in so many different types of cancer led
Chin and her associates to take a closer look at that stretch of
chromosome to see what genes reside there.
Using a method called genomic qPCR that can pick out specific
sequences of DNA, they found four genes in the amplified region, two of
which, GOLPH3 and SUB1, were expressed at high levels,
due to the increase in gene copy. To determine whether both, or
either, of these genes are involved in cancer, they conducted "loss of
function" tests, in which they lowered each gene's activity in a set of
lab-grown tumor cells.
"When we 'knocked down' GOLPH3 expression [or activity] by
95 percent, it significantly inhibited the ability of these cell lines
to grow in a semi-solid condition, a cancerous quality that normal cells
do not typically share," Chin says. "Knocking down SUB1 to a comparable level had only a minimal effect."
Intriguing as this finding was, it was hardly enough to prove that GOLPH3 is an oncogene — a contributor to cancer when overexpressed within a cell.
Demonstrating that would require several experiments to ensure that GOLPH3
itself, and not a nearby "shadow" gene, is responsible for the effects.
Next came gain-of-function studies to see whether revving up GOLPH3 activity can turn a non-cancerous cell cancerous. It did in both mouse and human cells.
"All these results enabled us to build a case that GOLPH3 is
an oncogene," Chin states. But there was a problem. "This information
wasn't very helpful for achieving our ultimate goal, which is the
translation of our findings into a form that is clinically useful for
Despite their discovery that GOLPH3 can promote cancer,
researchers didn't know what the gene's role is in normal cells. "There
was literally no information on what it does," Chin remarks. The only
hint was that the protein it encodes — designated GOLPH3 — is found in
the Golgi network.
The team's first attempt to uncover GOLPH3's role — using gene
expression profiling to see how protein levels track with various cell
functions — was fruitless. So the researchers ran experiments with yeast
cells to see which proteins share GOLPH3's cell neighborhood and which
proteins it interacts with.
One such partner was found to be VPS35, a component of a structure
called the retromer complex. The complex's job is to recycle the
antenna-like receptors that dot the cell surface.
From the many genetic screening tests that have been done in yeast,
researchers knew that flaws in the retromer complex can cause cells to
be vulnerable to rapamycin, just as excess GOLPH3 can. Rapamycin is
known to interfere with a protein called TOR, whose job is to control
yeast cell size. This suggested that the retromer complex in yeast is
important for chemical signals sent to and from TOR.
Chin's team theorized that mammalian GOLPH3 also works with the
retromer complex to control the activity of TOR in mammal cells (where
it's known at mTOR). To test this idea, the investigators found that
knocking down GOLPH3 reduced cell size just as rapamycin did. They
followed those experiments with biochemical studies to explore how
GOLPH3 affects cell size.
The team next sought to answer whether high GOLPH3 levels cause tumor
cells to be more susceptible to rapamycin in animal studies.
They took two sets of human melanoma skin cancer cells — one of which
had excess GOLPH3 and the other had normal levels — implanted them in
animals, allowed them to grow into tumors, then treated them with
"In the animals where GOLPH3 was overexpressed, the cancer cells grew
much faster, but the tumors were much more responsive to rapamycin,"
Chin notes, "suggesting the tumor-promoting effect of GOLPH3 is
dependent on mTOR signaling."
Lastly, the team considered whether the same mechanism might be at
work in human cancer cells. An experiment analyzing human tumor tissue
for specific proteins suggested yes. The researchers found that
non-small cell lung cancer cells with too many copies of the GOLPH3 gene also had abnormally high levels of mTOR activity.
"The mechanistic relationship we'd identified in the mouse system is
also at work in human tumors," says Chin, who is also an associate
professor at Harvard Medical School.
In addition to identifying GOLPH3 as a bona fide oncogene
and an indicator of whether rapamycin is likely to be effective against
specific tumors, the study points to the need to follow genomic studies
with a rigorous examination of the biological purpose and operation of
potential cancer genes, Chin concludes.
"Only then can we turn our intriguing discoveries in the cancer genome into something that is useful to cancer patients."
The study was supported by a grant from the National Institutes of Health.
Co-authors include Omar Kabbarah, Mei-Chih Liang, PhD, Joyce Wu,
Sabin Dhakal, Min Wu, PhD, Shujuan Chen, Tamar Feinberg, Joseph Huang,
Hans Widlund, PhD, and Kwok-Kin Wong, MD, PhD, Dana-Farber; Elena
Ivanova, PhD, Yonghong Xiao, PhD, and Alexei Protopopov, PhD,
Dana-Farber and the Broad Institute of Advanced Cancer Science; David E.
Fisher, MD, PhD, Massachusetts General Hospital; Valsamo Anagnostou,
and David Rimm, MD, PhD,Yale University School of Medicine; Abdel Saci,
PhD, Harvard Medical School.