Upping the ante in biochip research
William Check, PhD
Microminiaturization techniques developed in the computer
industry have been adapted in the last 10 to 15 years to making
silicon or glass chips, called biochips, on which biological reactions,
typically nucleic acid hybridization, take place. Ulysses Balis,
MD, instructor in pathology at Massachusetts General Hospital, uses
the analogy of computers to describe progress in biochips.
"Computers started out very expensive," he notes. "People weren’t
sure what they would do. But they became a mass market commodity,
with technology doubling every 18 months or so." Biochips also started
as a powerful tool for gathering thousands of pieces of information
concurrently on random bits of gene segments. "Yet initially there
was a dearth of tools for processing that data," Dr. Balis says.
Since the invention of the biochip around 1988 by Stephen Fodor,
PhD, the ability to use information from these devices has grown
exponentially. "Taking that into account, we can expect maybe in
five to 10 years a transformation in diagnostics," predicts Dr.
Balis, "not just tumor diagnostics but inborn errors of metabolism
and genetic predispositions as well."
For example, screening for the few main mutations and the hundreds
of sporadic point mutations responsible for cystic fibrosis by current
methods is laborious and impractical. But a relatively simple biochip
could theoretically perform this task quickly and accurately.
Patrick Brown, MD, PhD, associate professor of biochemistry at Stanford University
and associate investigator at the Howard Hughes Medical Institute, quantifies
the capacity of biochips. "We make microarrays where we try to represent all
genes that can be expressed from the human genome," he says. "So far we have
about 20,000. These are not all the human genes, but we are heading in that
direction. Assuming that about 100,000 human genes are expressed, there is no
fundamental reason we can’t get them all on a single array."
From a diagnostic perspective, biochips represent "an incredibly
powerful new set of tools for metabolic disorders, neoplastic predisposition,
and neoplasia," Dr. Balis says. These tools will classify neoplasias
with a prognostic accuracy not previously possible. Just as acute
lymphoblastic leukemia and acute myeloblastic leukemia looked similar
before flow cytometry and other tools distinguished them, gene technologies
are revealing differences in tumor types that look the same morphologically.
"But I don’t think gene chips are going to replace morphology,"
Dr. Balis says. "It will just be a new set of tools. Pathologists,
especially surgical pathologists, don’t need to look on this technology
with a jaundiced eye. They should embrace it as a better way to
Advocates of bio-chips believe eventually physicians will be able
to look at the spectrum of expression of 10,000 genes in a tissue
sample, compare that profile to a database containing prognostic
data associated with clinical histories and the chemotherapy used
to achieve those outcomes, and select the best antineoplastic agents
for that patient’s tissue type. "We will shift from empiric use
of neoplastic therapy to targeting disease based on the molecular
bases of oncogenesis," Dr. Balis says.
He cites a preliminary report in which 31 of 31 chronic myelogenous
leukemia patients went into remission following treatment with a
chemotherapeutic agent targeted to a key molecule in the process
that deranges the cell cycle.
Dr. Brown presents a similar view. "A pathologist can look at
tissue sections and recognize which are similar and different,"
he says. "If it is done systematically, one can recognize even subtle
features that connect with clinical behavior, features that carry
information about underlying biology." He continues, "A microarray
too has double value, since we can trace prognostic differences
in expression to particular genes that then become potential targets
So far almost all biochips are intended for research and
commercial uses rather than for clinical application. Says Tom Gingeras,
PhD, vice president of biological sciences at the biochip pioneer
Affymetrix, "All arrays that we make are for research purposes only."
Bob Burrows, director of corporate communications at Gene Logic,
which makes expression databases, sounds a similar note: "We want
to be content providers to the pharmaceutical, biotechnology, and
life science research communities." But, he adds, "the ultimate
downstream use of this information is to put it into the hands of
doctors for patient management and on the Internet for direct access
Underscoring the distance that bio-chips have to go to reach the
clinic is the experience of LabCorp’s Center for Molecular Biology
and Pathology. Says Tim Alcorn, PhD, director of infectious diseases
at the center, "Gene chips do not have much clinical utility right
now." LabCorp was using Affy-met-rix’s chip for detecting drug resistance
in HIV. But, Dr. Alcorn says, "There were limitations to what that
HIV chip could do that forced us to move away from it." One problem
is that the chip can detect only what it has been instructed to
look for. "HIV resistance testing evolved so rapidly that by the
time a chip design was set, it was obsolete," Dr. Alcorn says. "Ultimately,"
he believes, "a chip type format may replace gel-based sequencing
and be standard for HIV resistance testing, but not now."
Most people working in this field agree with the twin ideas that
bio-chips will revolutionize diagnostics but will not make pathologists
obsolete. Kieran Gallahue, chief financial officer at Nanogen, predicts,
"The real excitement will be surrounding expansion of testing—pharmacogenomics
and susceptibility and resistance testing. The objective is not
to replace 30-cent tests, but to provide a whole new level of information
that is much more specific and direct." He adds,"I don’t think anybody
believes that laboratorians are going to go away, but they may have
a higher level of information from which to interpret. Like many
other aspects of modern life, people don’t go away; tools empower
them to make better and more timely decisions."
As Fred Waldman, MD, PhD, professor of laboratory medicine at the
University of California, San Francisco, puts it, "Our goal is to
identify genetic signatures for subtypes of cancers. Gene chips
could distinguish between renal and bladder cancer, but a pathologist
can do that already with 99.9 percent accuracy. So the real goal
is to define prognosis of tumors better than histology can do alone."
Ideally, chips would not displace pathologists, but would provide
new information for them to use.
Lawrence True, MD, associate professor of pathology at the University
of Washington, envisions a scenario in which the initial workup
of a patient with suspected prostate cancer, for example, will involve
diagnosis by conventional his-to-pa-thol-ogy—"which is probably
the most specific and cost-effective and efficient way to diagnose
prostate cancer"—while expression array data will supplement some
of the traditional measures used to distinguish that cancer’s malignant
potential, such as histologic assessment of degree of differentiation.
Expression profiles could replace assays of proliferation, such
as DNA flow cytometry, and assays for over- or underexpression of
genes such as HER-2/neu in pros-tate cancer.
Dr. Gingeras sums up these ideas by saying, "We are looking
to begin moving toward more tailored and individualized treatment."
He thinks about the impact of biochips on medicine in terms
of patient management. "One can think about the issue as a three-dimensional
box," he says. These dimensions are the cost-effectiveness of treating
an individual patient; the patient’s status, asymptomatic to symptomatic;
and the likelihood of treatment success in that patient.
"Where we most often are today," he says, "is in the corner with
the lowest likelihood of a drug being successful as well as being
fairly expensive, and we use it most often on symptomatic patients.
Using genetics and profiling, you would like to move the population
to the corner where you are treating before they have serious disease,
where there is a higher drug effectiveness, and where you are getting
more from your money by virtue of treating the right people in the
right way." Genetic profiling could help match treatments to patients.
"Then you know the patient will respond," Dr. Gingeras says. "You
will not be doing an experiment on each individual."
Two assumptions underlie this approach. The first is that variations
in response to drugs are based on differences in metabolism that
reflect detectable differences in genes and gene expression. The
second is that there will be tissues available, such as blood cells,
that will serve as surrogates when access to the affected tissue
is not possible. This latter situation might arise in neurological
conditions, for instance. And if you move from treating symptomatic
to asymptomatic persons, it won’t be possible to take tissue from
an apparently healthy heart or other organ.
W hether these researchers’ visionary predictions
become reality is still an open question. But one fact is certain—an
overview of the several novel ways in which biochips are being exploited
bears striking testimony to human ingenuity. These devices are being
used to measure the relative number of genes in tissues, to quantitate
gene expression, as discovery platforms for finding potential therapeutic
targets in cancer, for screening libraries of chemicals, to generate
reference databases, as concentrating devices—each hopeful researcher
and company has a unique wrinkle.
We can start with an exception, Cepheid. "We are different
from other companies," says Curt Petersen, PhD, Cepheid’s president.
"Our intention is to attack the clinical laboratory market first
through pathogen detection with DNA probes." Cepheid uses chips
as an adjunct device—for concentrating samples.
Cepheid’s goal is automated sample preparation, including DNA extraction
from biological samples. "We are trying to make a hands-off cartridge-based
approach," Dr. Petersen says. "You take a specimen such as urine
or blood, put it into our machine and it does everything." A prototype
using a cartridge for chlamydia/gonococcus detection has been demonstrated.
"It accepts up to 5 mL of urine and gives you a DNA probe result
in 30 minutes," Dr. Petersen says. Potentially this will allow a
small clinic or operating room to take a sample and get a rapid
result for many infectious diseases or genetic disorders.
Cepheid uses chips to concentrate DNA. Its micro-fluidic chip contains
an array of single-crystal silicon etched pillars that capture DNA
as a biological fluid flows through. With their high surface area,
the pillars adsorb DNA by nonspecific interactions.
Initial applications will be in areas where an answer is needed
fast, although Dr. Petersen won’t specify which applications the
company is working on. He does note that Ce-pheid’s prototype chlamyd-ia/gono-coccus
cartridge "gave results in less than one-half hour that were comparable
to commercially available, FDA-approved methods taking many hours."
He adds that Cepheid has done a lot of work on biowarfare detection,
including anthrax, for the Department of Defense.
Dr. Petersen predicts that Ce-pheid’s products will have a strong
impact on culture-based bacterial assays. "In the next three years,"
he asserts, "we will dramatically change that industry, making identification
a lot easier, quicker, and more accurate."
Dr. Waldman of UCSF uses bio-chips in a more conventional and research-oriented
manner. In his work on genetic alterations in solid tumors, he uses
them to evaluate changes in the copy number of genes. This application
evolved from previous experiments on chromosomal copy number alterations
using a technique called comparative genomic hybridization, in which
tumor DNA and normal human DNA are labeled with fluorochromes of
different colors and hybridized to human metaphase chromosomes.
Changes in copy number show up as altered fluorescence ratios, allowing
one to identify and quantify gains or amplifications in genetic
material located to a particular chromosome region at low resolution—10
To increase resolution, Dr. Waldman performs the same type of experiment
on biochips using much smaller 100-kb clones of DNA as probes rather
than metaphase chromosomes. A "profusion" of such clones have been
isolated and provided by laboratories funded by the Human Genome
Project, Dr. Waldman notes. Each clone or DNA segment is placed
into one well of an 864-well plate. A robot spots a sample of each
clone onto a slide via pins, creating the array. Thousands of different
DNA clones can be spotted onto one array. Then labeled tumor and
normal DNA are hybridized to the arrays, revealing relative amounts
of the two DNAs binding to each spot, which defines genetic gains
or losses at high resolution.
Dr. Waldman estimates that a useful map of tumor-related alterations
in copy number can be achieved with about 3,000 cloned fragments
distributed at one-megabase intervals along the genome. For regions
of particular interest, such as a breakpoint in a translocation,
analysis can be done at higher density.
His goal is to identify candidate markers of clinical outcome either
with therapy or in the absence of therapy. Already more than 30
oncogenes are known to be amplified in one or more forms of cancer.
And many chromosomal regions are amplified or deleted but the identity
of the presumed genes in those regions is not known. "Our hope is
that identifying these genes that are altered in copy number will
lead to better diagnosis and prognosis of cancers," Dr. Waldman
One project involves renal cancer, for which other methods have
defined specific genetic alterations associated with differing histologic
subtypes. Dr. Waldman has worked with Guyla Kovacs, MD, of Germany,
who provided 40 primary human renal cancers of differing histologic
subtype—for example, clear cell, chromophobe, capillary. Dr. Waldman
and his colleagues have produced arrays containing cloned fragments
representing regions known to be associated with the different renal
cancer subtypes. Hybridizing labeled DNAs from the human cancer
samples onto arrays allowed them to classify these cancer DNAs with
high accuracy. Although they have not yet totally unblinded the
study, Dr. Waldman says,"We can see clear differences in genetic
alterations of different renal tumors. So this becomes potentially
a renal cancer diagnostic chip.
"Right now there are not good therapies or adjuvant treatment for
renal cancer," he continues. "But identification of new genetic
markers opens up the possibility of gene-based therapies in the
Dr. Waldman’s work concerns direct genomic analysis.
More common today is expression analysis of genes, using a similar
approach to ask whether some genes are over- or underexpressed in
tumors. In this approach, messenger RNA (mRNA) is extracted from
tumors, reverse transcribed into cDNA, cloned, and applied to an
Dr. True, of the University of Washington, uses expression arrays
in his research on prostate cancer. "Our goal is to use arrays of
ultimately all genes whose expression is associated with growth
and differentiation of human prostate tissue," ranging from development
to benign proliferative diseases like nodular hypertrophy to prostate
cancer, he says. "These arrays will allow us to identify patterns
of gene expression that characterize prostate cancers that will
progress to metastatic tumors and to see which features of those
patterns are most strongly associated with progression." Once the
specificity of such putative patterns of gene expression is validated,
a standardized expression array of at least 10,000 unique genetic
sequences can be used as a laboratory assay in the initial diagnostic
workup to help tailor management to the individual patient.
"We are increasingly able to detect prostate cancers based primarily
on more sensitive serum PSA assays," Dr. True points out. "But only
a minority of men have prostate cancer that will progress. The problem
is that our present tools are not sufficiently specific to distinguish
those patients whose cancers will progress from those whose cancers
will not. The promise of expression arrays is to make that distinction
with more certainty than we can presently do."
Optimism that such distinctions are possible is based on precedents.
Dr. True cites differences in molecular abnormalities of neuroblastomas
in children, where the degree of amplification of the n-myc gene
is a predictor of the probability of metastasis. In breast cancer,
overexpression of the HER-2/neu protein characterizes tumors that
progress more rapidly, although the link is weaker.
"We have identified about 20 novel partial sequences of genes that
remain to be identified, and we have reported on a couple of known
genes that seem to be differentially expressed in the prostate,"
Dr. True reports. "We have been collecting fresh tumor tissue with
information about stage and grade for the last six years. Probably
within the year we will take some of those samples and see what
expression differences we can detect. Using an array with a couple
of thousand sequences, we will see if we can correlate expression
patterns with clinical outcomes." (Dr. True’s collaborators at the
University of Washington are Lee Hood, MD, PhD, and Peter Nelson,
MD, of the Department of Molecular Biotechnology, and Paul Lange,
MD, Alvin Liu, PhD, and Robert Vessella, PhD, of the Department
Technically, handling tissue for expression arrays differs from
handling tissue for anatomic pathology, Dr. True notes. Hybridization
to an array now requires greater care and immediacy in handling
of the tissue to minimize RNA degradation. Fresh, not formalin-fixed,
tissue is required, and tissue must be put into solutions that inhibit
enzymes that degrade RNA. Second, solid tumor biopsies contain mostly
nontumor cells, so techniques to enrich for tumor cells are needed.
Dr. True is optimistic about flow cytometric sorting using cell
surface antigens, a method now used for lymphomas. A third, more
general, issue is how representative a biopsy sample is of the whole
tumor. "We don’t yet know the answer to that question," he says.
Dr. Brown of Stanford is also an expert in expression arrays. "Recently,"
he says, "it has become possible to come close to looking at the
activity of every gene in the genome in clinical samples." With
this level of detail, he notes, "It is crucially important that
our approach is systematic, making a series of detailed molecular
characterizations of clinical samples using a rather standardized
quantitative method. That allows us to look at large sets of samples
and to use a rigorous statistical method to try to recognize correlations
between variations seen at the molecular level and differences in
the clinical behavior and biology of tumors."
Dr. Brown’s method is to "print" about 140 arrays at a time with
20,000 to 30,000 genes each using a mechanical printing process.
Then, using a selective hybridization method, he quantitatively
measures the abundance of mRNA from each of the genes in a clinical
sample relative to a standard scale.
In an article published in early February in Nature, Dr.
Brown and collaborators at the National Cancer Institute and other
centers reported results of a study of variation in gene expression
in diagnostic biopsies of diffuse large-cell B cell lymphoma. Based
on gene expression patterns, they identified two subtypes of this
malignancy not previously recognized. "Not only do these subtypes
have a distinct difference in their gene expression pattern that
suggests underlying biological differences," Dr. Brown says, "but
they have a highly statistically significant difference in clinical
outcome, a difference that has never before been possible to see."
In this case the differences in gene expression suggest possible
molecular differences that could account for the differences in
outcome. These findings are now being expanded with a larger group
of collaborators. "There will be many more such findings in the
next year or two," Dr. Brown predicts.
In a variation on gene expression analysis, David Per-sing, MD, PhD, vice president
of diagnostics development at Corixa Corp. and medical director of the Infectious
Diseases Research Institute, both in Seattle, is using expression arrays as
what he calls "gene discovery" platforms. "We use expression microarrays as
screening devices to identify potential targets for T-cell vaccines for cancer,"
Dr. Persing says. Such vaccines are based on the concept that tumor-specific
proteins or nucleic acids can prime a patient’s own T cells to become cytotoxic
to a tumor or pathogen.
"For several tumor types, we aim to identify from the roughly 100,000
genes in the human genome between six and 20 candidate genes for
which expression is essentially restricted to the tumor," Dr. Persing
explains. Candidate genes identified on expression microarrays are
subjected to real-time PCR (RT-PCR), which is more sensitive than
expression analysis, to verify specificity, and to additional verification
steps. "These candidate genes can be used for diagnostics, because
their expression is consistent with the presence of tumor, and for
vaccine targets as well as therapeutic antibody targets," Dr. Persing
Immunohistochemical testing of a prostate-specific candidate protein
discovered by Corixa showed that 65/65 prostate cancers or prostate
tissues were reactive while all 4,635 nonprostate tissues were nonreactive.
(For prostate cancer, markers only need to be tissue-specific, not
cancer-specific, since vaccines will be used in men who have had
Another candidate gene is mammoglobin, which Dr. Persing calls
"the most breast tissue-specific gene that has ever been described
in terms of its expression profile." Mammoglobin and two other markers
were used to detect breast cancer cells in the blood of women who
had had cytoreductive surgery. (Tumor cells can be detected in lymph
nodes and blood of such patients even without obvious lesions.)
Markers in addition to mammoglobin were needed since one-fourth
of breast cancers either do not express it or lose the chromosomal
segment that carries the mammoglobin gene.
Dr. Persing says that, using RT-PCR to measure the expression of
these three genes, expression of one or more targets was present
in nearly 100 percent of breast cancers. Dr. Persing is "very optimistic"
about using these techniques for detection of cancers of various
types and for monitoring therapy. "We plan to use RT-PCR profiles
on whole blood to determine if we can detect circulating cancer
cells at a very early stage," he says. Cells will be captured on
magnetic particles using antiepithelial antibodies and assayed with
RT-PCR to detect particular types of cancer. "I think this will
be a huge area of opportunity for diagnostics," Dr. Persing says.
Corixa is involved in clinical trials to evaluate this hypothesis.
Affymetrix, the company that trademarked the term
"GeneChip," employs a unique approach to making microarrays. "Arrays
are usually made with premade oligonucleotides or cloned DNAs that
are bound to a substrate," Dr. Gingeras explains. In Affymetrix’s
proprietary method, oligonucleotide probes, usually 20 to 25 nucleotides
long, are synthesized on the substrate in a combinatorial fashion
using photo-catalyzed reactions. By masking sites in a preset pattern
as oligonucleotides are lengthened, each position on the surface
identifies a unique sequence.
"This approach allows us to achieve much higher densities of information
than competitors," Dr. Gingeras says. Affymetrix’s chips routinely
carry 240,000 to 280,000 unique oligonucleotides.
With these gene chips, three kinds of questions are answerable. In resequencing
applications, a consensus sequence is defined for a portion of the genome. One
can then do base-by-base interrogation of a sample testing for identity with
each of the bases in the consensus sequence. Discovery of SNPs (single-nucleotide
polymorphisms) is a direct application of this approach, Dr. Gingeras says.
Affymetrix has relationships with bioMérieux and Roche Molecular
Systems to develop chips for this purpose. Clinical applications
include bacterial identification, viral genotyping for drug resistance
mutations, and genotyping of p53 genes and some human cytochrome
P450 genes. Both companies are building proprietary platforms for
diagnosis based on GeneChip probe arrays. Affymetrix will manufacture
and provide arrays to the companies, who will handle development
A subset of resequencing applications arises when one wants to
focus not on identifying each base in a long string of bases, but
on looking at snapshots across the whole genome. "A large amount
of work is being done identifying SNPs on the human genome," Dr.
Gingeras says. "We are beginning to develop assays and arrays to
interrogate many of those polymorphisms." Affymetrix has just released
a product that interrogates 1,500 SNP sites mapped on the human
Quantitative mRNA profiling—gene expression—is a third application.
"We currently make a variety of arrays that have varying numbers
of genes for evaluating message levels in a cell," Dr. Gingeras
says. "For each gene in the expression array, the sample is interrogated
with a set of 16 to 20 oligonucleotide pairs." Each pair consists
of an oligonucleotide that is the perfect complement of the message,
which interrogates a single base positioned along the length of
the mRNA molecule, along with its companion, which has a deliberate
mismatch incorporated at its center, and which is designed to serve
as a negative control.
"This approach allows one to have multiple interrogations for each
sequence," Dr. Gingeras says, "so you are not dependent on a single
interrogation by a single probe as with a single PCR product or
a single spotted clone." With this redundancy, results are less
likely to be compromised if a sequence in the public database has
Nanogen also supplies chips for researchers, but with a difference:
Its chips can be custom designed in the investigator’s laboratory.
As CFO Gallahue puts it, "We supply open platforms for targeted
genetic analysis. Our products are unique, since their integrated
electronics allows researchers to create their own genetic array."
With a Nanogen chip, it is possible to do an analysis and get results
in a single day, an advantage in research and eventual clinical
applications. One of today’s biggest challenges, Gallahue notes,
is to translate clinical research into clinical practice. "Nanogen’s
technology," he asserts, "allows users to bridge that transition
by designing their chips to meet requirements for either clinical
research or clinical application."
More specifically, Gallahue notes, most biochips are passive systems,
whether lithographically synthesized or spotted. In either case
a genetic sequence is put down on a glass slide and heat and chemistry
are manipulated to facilitate hybridization and stringency. "You
have to apply the same characteristics to all of the chip at the
same time," he says. Nanogen’s product integrates electronics into
the analysis process. The company’s first product will be a 100-site
chip with 100 individual controllable electrodes. A user can create
an environment around each electrode without affecting the others.
"In effect you have 100 independent test sites," Gallahue says. "You can use
the electronics to turn each site positive or negative or neutral. So a user
can manipulate the electronics to attach genetic content to a blank chip at
sites of their choice, then use reporter probes to interrogate the whole chip
or only portions of it at one time."
Later this year Nanogen will sell its first product—NanoChip molecular
biology workstation—to the research market—pharmaceutical companies,
genomics firms, biotechnology firms, and clinical investigators.
Last year, Dennis O’Kane, PhD, of Mayo Clinic, one of the beta-testing
sites for the product, reported that he and colleagues had successfully
used a customized Nanogen chip for a pharmacogenomic application,
looking at SNPs in the gene for thiopurine methyl transferase (TPMT),
which metabolizes immunosuppressant thiopurine drugs used to treat
leukemia, Crohn’s disease, and other conditions. Variants in the
TPMT gene result in high and low activity forms of the enzyme, which
can affect drug toxicity in individual patients. A Nanogen chip
customized by the Mayo researchers was 100 percent accurate in identifying
SNPs in the TPMT gene in 115 patient samples, while two widely used
comparison methods had six percent and 11 percent error rates.
A similarly positive result was reported by the Bode Technology
Group, a leading private forensics laboratory, using a custom-designed
Nanogen chip to identify short tandem repeats, genetic identifiers
used by the government for populating criminal identification databases.
Caliper Technologies makes no microarray products
as such, but it uses biochips in a unique way—as total reaction
platforms. Michael Knapp, PhD, vice president of science and technology
at Caliper, notes that a conventional array has one partner of a
single biochemical reaction, often an oligonucleotide probe, immobilized
on its surface. As a result, such chips are unifunctional: They
perform one step of a complex series of reactions, such as hybridization,
with high throughput. But other steps in the process, such as amplification,
may be done in a more labor-intensive format.
Caliper uses photolithography and photoetching techniques to create
a custom network of channels on the order of 10 µm deep and 50 µm
wide in a chip. "These channels," Dr. Knapp says, "amount to an
instruction set for a series of fluid manipulations."
Such chips incorporate three innovations: They miniaturize functional
steps to volumes of 100 picoliters to a nanoliter; integrate multiple
steps on the same device; and automate the reaction sequence with
a custom-designed software program that can exert forces on fluids
inside the chip using pressure or vacuum or electrokinetic forces.
Caliper’s main product, targeted to the pharmaceutical industry,
is a high-throughput device for screening large libraries of compounds
for desired activities, such as antagonism of a cell surface receptor,
inhibition of a kinase or calcium flux, or DNA hybridization—all
multistep assays. Several such high-throughput chips are in production.
A reservoir feeding into one processing unit of a chip stores enough
reagent for a 24-hour run, perhaps 10 µL for each reaction, adequate
for about 10,000 assays. Chips can have up to 12 processing units,
thereby having a capacity of more than 100,000 assays per day.
Finally, at Gene Logic, the product is applied expression arrays
targeted to the research market. Gene Logic uses Affymetrix GeneChip
expression arrays, supplemented by a proprietary method to quantitate
low-abundance transcripts, to generate expression databases. It
then sells these databases in two formats: tailored databases sold
exclusively to pharmaceutical customers, or their large reference
database called Gene-Express by subscription. The company just signed
up its first two subscribers to GeneExpress, one biotechnology company
and one large pharmaceutical company, says director of corporate
He notes that the goal for GeneExpress is to have quantitative
expression profiles for all 100,000 genes across 30,000 human samples
across all tissue types and all disease types plus all underlying
clinical data. This will enable researchers to query on hundreds
of factors for their influence on clinical outcome. Says Burrows,
"It will be possible to say,
’Show me all genes that are expressed in livers of men aged 15 to
45 who drink a lot of alcohol and take Prozac and compare that profile
to similar individuals who don’t take Prozac.’’" So far GeneExpress
contains about 1,200 profiles, and profiles are being added at 400
samples per month; the company plans to accelerate to 800 per month
later this year. Samples are obtained from volunteers recruited
through an international network of centers.
If the scope of Gene Logic’s database seems overwhelming, consider a pharmacogenetic
application conceived at Caliper. "With massive genotyping in the developmental
phase of a drug," Dr. Knapp says, "one can identify different responses and
different toxicities that are genetically determined." This requires 100,000
different genotypes on perhaps 1,000 patients. To enable this project, Caliper
has developed an integrated chip that does amplification and genotyping as a
high-throughput screening operation. Says Dr. Knapp, "Generating such a database
requires a scale of experimentation that is unprecedented in any field."
William Check is a freelance medical writer in Wilmette, Ill.