Colon cancer markers still in limbo
Microsatellite instability as a maker
William Check, PhD
Investors in Internet stocks are beginning to learn the same
hard lesson that pathologists and clinicians who work with prognostic
and predictive markers in colon cancer have taken to heart in the
last few years: Faith and promises will take you only so far. When
the Nasdaq lost one-fourth of its value during one week in mid-April,
Newsweek Wall Street editor Allan Sloan wrote that stocks of many
companies in the so-called New Economy "trade largely on the basis
of hopes, dreams, hype and momentum because there is little or nothing
in the way of profits or assets for investors to weigh." For many
years, molecular markers for colorectal cancer were in a similar
In 1994 the CAP sponsored a conference on prognostic factors in
colorectal cancer (along with breast and prostate cancers) at which
"many factors looked as though they were going to be important,"
says Carolyn Compton, MD, PhD, professor of pathology at Harvard
Medical School and director of gastrointestinal pathology at Massachusetts
General Hospital. "It seemed at that time that several tissue-based
prognostic factors were poised on the verge of being validated,"
Dr. Compton told CAP TODAY. But evidence reviewed at a five-year
followup conference sponsored by the CAP and held in June 1999 revealed
that, "Not a single prognostic factor [for colorectal cancer] had
been validated in that time period," says Dr. Compton, chair of
the group that wrote the conference summary on prognostic factors
in colorectal cancer, which will be published next month in the
Archives of Pathology & Laboratory Medicine.
As with dot-com stocks, there was a dearth of necessary information.
"What prevented us from accumulating a critical mass of data that
would either validate or eliminate these factors from further consideration,"
says Dr. Compton, "was that all studies were too small to achieve
the statistical power needed and they varied in methodology and
statistical analysis so you couldn’t amalgamate the data from different
Another conference participant, Clive Taylor, MD, PhD, chair of
the Department of Pathology at the University of Southern California
School of Medicine, cites the same obstacle. "The problem with prognostic
markers," Dr. Taylor says, "is that when they first appear in the
literature, you get two or three positive reports and people get
enthusiastic. But many years later you still have only inconclusive
datasets," which won’t support a definitive evaluation, much less
a positive recommendation.
Two other conferences have reached similar conclusions in the last
two years. Mark Welton, MD, associate professor of surgery at the
University of California-San Francisco School of Medicine, took
part in the 1999 CAP conference and a 1998 conference on the same
topic sponsored by the American Joint Commission on Cancer, or AJCC.
(The summary report from that conference was published in the April
1, 2000 issue of Cancer.) The outcomes and recommendations of the
two conferences were similar, Dr. Welton says. For prostate cancer,
PSA is indicated. "But there is not an equivalent molecular marker
for colon cancer," he says. "That is the exact place where we came
down from both the CAP meeting and the AJCC meeting."
Gershon Locker, MD, chief of hematology/oncology at Evanston (Ill.)
Northwestern Healthcare, sits on the American Society of Clinical
Oncology’s tumor marker panel, which last year reviewed evidence
for the clinical utility of molecular markers in colorectal cancer.
"We only advocated the use of carcinoembryonic antigen [CEA] and
only for monitoring in limited circumstances, and even that was
somewhat controversial," Dr. Locker reports. "That coincides with
my own practice. It is my view that until a marker is prospectively
shown to change a patient’s prognosis and our clinical practice
or treatment, that marker shouldn’t be advocated. Retrospective
analysis is not the right way to choose what markers should be used
clinically," he adds.
What has changed fundamentally during the past few years in the
tumor marker field is what may now be occurring for Internet stocks-a
demand that promises be redeemed with proof of performance. "Just
as molecular biology and the discovery of molecular markers have
been a major scientific advance," Dr. Locker says, "there has been
a major advance in how we look at evidence. We have moved toward
evidence-based medicine. It is no longer sufficient to adopt in
clinical practice based on retrospective studies what looks like
a hopeful marker." Today long-term prospective clinical outcomes
are the standard of adoption. And in colorectal cancer no molecular
marker has met that standard.
"Molecular markers are a big field, but when it comes down to it
very few are used in management decisions," agrees Mark Redston,
MD, staff pathologist at Mt. Sinai Hospital, Toronto. "To justify
clinical adoption, we need to show that a marker predicts response
to treatment, rather than just predicting behavior."
"I don’t think at the present time any study or series has led
to a change in clinical management and treatment on the basis of
molecular markers," states Chris Willett, MD, professor of radiation
oncology at Massachusetts General Hospital, who took part in the
CAP conference. "I have been taking care of patients with bowel
cancer for 20 years," Dr. Willett continues. "There have been a
lot of promises based on markers, but the accepted methods for guiding
therapy remain essentially the same."
Currently accepted methods for guiding therapy made up the
second major topic at the CAP conference on prognostic factors in
colorectal cancer. Were this part of the conference made into a
movie, it could star Michael J. Fox and be titled "Forward Into
the Past." For at the outset of the 21st century, the best prognostic
factors are related to the traditional, decades-old TNM staging
system-local extent of tumor (T), regional lymph node status (N),
and presence or absence of distant metastases (M). "Evaluation for
those three factors remains the single most powerful indicator for
any solid tumor," Dr. Compton states.
"At the present time, all clinical decisions are based on the TNM
staging system or classical surgical pathology staging," Dr. Willett
agrees. "That is still the fundamental basis of treatment."
This system is hardly static, however. Conference participants
made many substantive recommendations for improving the TNM system.
In fact, most of the conference turned out to be "an intense discussion
enumerating the reasons on a very specific point-by-point basis
why each of the identified tissue-based factors had not advanced
useful but unproven histopathologic factors include, among others,
vascular invasion, tumor border configuration, and host immunologic
response. "The problem was dissected for each factor as to why we
weren’t closer to knowing whether this factor was important or not,"
says Dr. Compton.
As M. Elizabeth Hammond, MD, chair of pathology at LDS Hospital,
Salt Lake City, succinctly summarizes it, "The main conclusion of
the conference was that significant work needs to be done in improving
the way we stage colon cancer. In all three of these cancers we
made significant recommendations about how to do grading so we would
get better information. In colon cancer, for instance, we recommended
changing to a two-grade rather than a three- or four-grade system."
Standardization was also emphasized. "What has come to the fore,"
adds conference participant Stanley R. Hamilton, MD, head of pathology
and laboratory medicine at the University of Texas M. D. Anderson
Cancer Center, "is the recognition that there are sets of criteria
that have to be met as to how evaluation of specimens should be
undertaken for results to be reproducible and useful in patient
Standardization of basic surgical pathology is essential to the
clinician as well. "It is hard to make innovations if the fundamentals
of surgical pathology are not done well," Dr. Willett says. "Proximal
and distal margins plus radial or soft tissue resection margins
are very relevant in colorectal cancer," he emphasizes, "as are
adequate number of lymph nodes and the presence or absence of lymphatic
or venous invasion."
If widely adopted, the suggested improvements will have a further
advantage: They will make it easier to evaluate molecular markers.
Major obstacles to interpreting the data in this field are that
investigators follow their own methods and there are no standards
for interpreting or reporting data or for statistical analysis.
"Some good ideas came out of the conference about what needs to
be done to quality control testing of molecular markers and to accumulate
data for their clinical validity," Dr. Hamilton says. "Molecular
markers have tended to be in the purview of molecular biologists
and laboratory researchers, who don’t have the skill set to be able
to evaluate them or to apply them in clinical settings. We as pathologists
need to get involved in that process."
In the remainder of this article we will discuss proposed enhancements
to traditional histopathologic staging, followed by a look at molecular
markers-why they remain in clinical limbo, and a few of the most
promising factors under investigation.
"Histopathology has been in a state of evolution," Dr. Hamilton
points out, "with the addition of important pathologic features
over and above the standard ones such as depth of invasion and lymph
node status that have been the mainstay of the TNM classification
for nondisseminated disease. We have now begun to recognize that
some newer features are useful for identifying subsets of patients
at higher risk. Tumor differentiation, extramural venous invasion,
and depth of invasion beyond the muscularis propria have now begun
to move into the classification with synoptic reports, so that they
are emphasized and provide maximal information for clinicians who
need to make decisions."
Another notable change, Dr. Hammond points out, is in thinking
about how to stage a colon cancer patient who has chemotherapy and
then surgery. "A number of [colon cancer] patients have chemotherapy
prior to surgery," she notes. "A tumor that hasn’t been treated
before surgery can’t be compared directly with one treated with
chemotherapy prior to surgery. It is in a different stage category."
Dr. Welton cites several ways in which the conference’s recommendations
have already helped him manage patients.
"Our pathologists’ reports became significantly more useful soon
after the conference," he says. (Whether these changes were due
to his input or some other route is not important, he says.) "Perineural
and vascular involvement, large vessel involvement, and inflammatory
response-all issues we discussed at the CAP conference-are now itemized
on my final pathology report and available on my computer," Dr.
"One of my major goals," he adds, "was to dig in my heels and ask
CAP to set an adequate number of lymph nodes to be reported in the
surgical specimen. This is an ongoing argument between pathologists
and clinicians and is really important for us surgeons and radiation
and medical oncologists." He acknowledges that this is "a very sticky
issue," due to constraints on how much time a pathologist can spend
on each specimen. "In the end, we got a minimum of 12 lymph nodes,"
Dr. Welton reports. If fewer nodes are seen, the pathologist might
go back and use visual enhancement techniques such as fat clearing.
"Clinically that has already been important for me," Dr. Welton
says. "It gives me as a surgeon making treatment decisions something
to stand on when I come up short of lymph nodes. And it takes some
of the acrimony out of the discussion."
Guidelines on how many lymph nodes or tissue sections are acceptable
are important to pathologists as well, Dr. Hamilton points out.
These numbers represent a "target," rather than a new standard of
care. "Especially in this era of such severe constraints on cost,
pathologists are under pressure to hold down costs," Dr. Hamilton
notes. "But we don’t want to compromise patient care. Having a document
of this sort that provides guidelines means that now it is not just
one individual pathologist doing battle with an administrator about
the number of blocks being processed for a given colon cancer case."
Dr. Welton notes that the conference participants also spent time
clarifying T3 and T4 lesions. A lesion that comes up to the serosa
but does not penetrate it is a T3 lesion; if it involves the serosa,
that is a T4 lesion. "There was a lot of discussion on this," Dr.
Welton reports, "even a drawing by [Dr. Compton] illustrating that
you can have sloughing of serosa on the colon because of inflammation.
Is that T3 or T4? And what is T4 with residual disease-when you
know you have left residual tumor behind?" These questions are clarified
in the conference report.
But even with improved histopathologic staging, there will
still be clinical roles for molecular markers. Dr. Compton cites
one possible application. For patients with stage 2 colorectal cancer,
who have no regional lymph node metastases (N0), the current standard
of care is surgery alone. Cure is achieved by complete excision
of the tumor in about 75 percent of such patients, but 25 percent
have recurrent tumor and die of their disease. "We can’t predict
which 25 percent of stage 2 patients those are," Dr. Compton points
out. A molecular marker that reliably identifies stage 2 patients
at high risk of recurrence could guide the use of adjuvant chemotherapy.
In a reverse situation, Dr. Willett notes the tendency to overtreat
stages 2 and 3 (N0 or N+) rectal cancer after surgical removal.
For these patients, postoperative chemotherapy and radiation are
now recommended. Some, however, are cured by surgery alone. "It
would be valuable to have markers to elucidate which patients are
more likely to have a good outcome and so can be spared further
treatment," he says.
Dr. Hamilton agrees that the "most exciting and encouraging" aspect
of molecular markers is their potential ability to individualize
therapy. For instance, in stage 3 colon cancer, where adjuvant chemotherapy
(5-fluorouracil plus leucovorin) has become standard, it would be
useful to be able to identify which tumors will respond and which
will be resistant. "Some patients appear not to get a survival benefit
from that regimen," Dr. Hamilton says. "It would be good to be able
to pick those patients in whom newer, more toxic regimens should
be tested as first-line adjuvant therapy." One such regimen is 5-fluorouracil
plus leucovorin with CPT-11 (irinotecan), which looks advantageous
in clinical trials. "Doing that will require input from molecular
techniques," Dr. Hamilton believes. "It is essential that pathologists
be involved in that process."
Given the perceived value of adding molecular markers to the staging
repertoire, why are the needed studies still lacking?
"One reason," says Dr. Hammond, "is that we have not been able
to be precise enough in how we collect routine information about
tumors-how far the tumor has spread through the wall, what constitutes
metastasis in a lymph node, and being precise about the histologic
grade of the tumor. There is so much variation in the way we examine
specimens, the way we interpret results, and how we report those
results that it has confounded our ability to get adequate amounts
of data to tell us whether one marker or another is important. If
we don’t learn to standardize the way we examine, report, and interpret
things," she predicts, "we will never get enough information to
make conclusions about molecular markers."
Dr. Compton describes another obstacle: the need for large multicenter
studies. "We already have very strong prognostic factors related
to TNM classification," she notes. "To know whether a newer factor
is important, you have to validate it by determining whether it
has independent prognostic significance over and above staging.
You have to stratify patients according to stage, then look at the
factor in a defined patient population in a randomized prospective
fashion. You can’t do that unless you have many institutions working
But only about three percent of cancer patients go on to cooperative
studies. "Even funded studies are having trouble accruing adequate
numbers of patients," Dr. Compton says. Cancer trials are labor-intensive
and grants pay for ancillary support but not for physicians who
enter patients. "So," she points out, "you are asking physicians
who are already beleaguered in this time of managed care to do more
work for no additional pay."
Pharmaceutical companies tend not to fund such studies because
most markers are not patentable; government grants tend to emphasize
treatment. "Prognostic factors are not high on the fundability list,"
Dr. Compton notes wryly.
She sees a possible change in attitude. "People are starting to
recognize now that we are not going to get to the goal as efficiently,
we are not going to improve the care of the cancer patient fast
enough, unless the emphasis comes off the RO1 grant-the independent
investigator, my-favorite-marker approach-to the cooperative group
mechanism," she thinks. "That is the only way to get data conclusively
At the CAP conference, data were examined for a large number
of markers, among them p53, K-ras, c-myc, bcl-2, thymidylate synthase,
mitotic index by flow cytometry, loss of heterozygosity (LOH) at
chromosome 18q (also called deleted in colon cancer, DCC), and microsatellite
instability. A few newer markers are considered worthy of further
investigation. Dr. Hammond says, "Microsatellite instability and
loss of heterozygosity at 18q are promising, but there are not enough
clinical data to recommend them for routine clinical use" at this
time. (See "Microsatellite instability as a marker," page 19.)
At M. D. Anderson, Dr. Hamilton and colleagues are looking at microsatellite
instability [by DNA analysis and immunohistochemistry for the products
of genes responsible for nucleotide mismatch repair] and 18q allelic
loss, as well as p53 gene status. "This is a field that is still
very much in the development stage," Dr. Hamilton says. "We are
trying to move from the research laboratory into the clinical setting."
As for the two methods to measure microsatellite instability, he
says, "Obviously IHC is particularly appropriate, because that is
a standard technique available in the vast majority of pathology
Dr. Taylor’s clinical research at USC concerns proliferation markers,
which, he believes, "have a promising future, although they are
not proven yet." In his view, "Topoisomerase II will be of greater
value in the long run than mitotic counts." Tumor proliferation
is assessed now by morphologic factors, such as tumor grade and
counting mitotic figures. Flow cytometry is thought to be more accurate,
but that method is not always available and is relatively expensive.
Thus the interest in immunostaining for molecular markers of proliferation.
One of the first proliferation markers described was PCNA, which
did not pan out. Next was Ki 67, which was better but still had
a fairly long half-life, so it was not specific for the proliferative
phase of the cell cycle. "Now we have topoisomerase II, which is
most closely restricted to S phase, though not totally so," Dr.
Taylor says. "It provides a pretty good index of proliferating cells.
One remaining challenge is to find a scoring system that is reproducible
across institutions as well as defining the best antibodies." Work
on proliferation markers is spurred by the increasing number of
papers appearing across a number of tumor types showing that rate
of division relates to tumor aggressiveness and eventual outcomes.
Of course, even these newest and most promising markers will still
be question marks in five years if the correct studies are not done.
But progress is being made. Most trials groups are now requiring
tissue blocks so that various molecular markers can be assayed,
according to Dr. Willett. "Cooperative groups are running tests
on batteries of markers to determine which are valuable in a consistent
way," he says. "So far results vary among studies and are very heterogeneous."
If the improvements in methodology recommended by the CAP prognostic
markers conference report are adopted, perhaps more convincing data
William Check is a freelance medical writer in Wilmette, Ill.