Biology Reference
In-Depth Information
the order in which NER factors
24
are added at DNA
damage sites. Similarly, no one is sure about the order
of the steps involved in NHEJ (Chapter 8). More
complete knowledge of such processes is necessary to
accurately identify biomarkers and determine which
repair proteins to inhibit for the greatest therapeutic
benefits.
Characterizing these pathways also must include
widespread efforts to collect prospective patient
samples. Large-scale screening of tissue samples from
populations of both healthy people and people with
cancer need to be tested to accurately characterize
DNA repair pathways, their deficiencies in various
cancers at different stages of transformation, and the
potential therapeutic response that could be gained
from targeted inhibition.
2
This molecular characteriza-
tion could also include identifying other deficiencies
related to cell growth and survival.
that may be clinically significant, additional
capabilities are needed, such as denaturing high-
performance liquid chromatography (DHPLC)
e
and the surveyor nuclease assay, which can capture
SNPs not easily detected by DHPLC. Both are being
used with the Gene-PARE project.
14
Other existing
technologies or ones in development may prove to
be more accurate or comprehensive.
4. Correlate Biomarker Findings to Specific
DNA Repair Activities
A variety of rare human diseases supports the
hypothesis that decreased DNA repair capacity causes
increased mutations, genetic instabilities, and eventual
cancers. Recent technological advances have enabled
researchers to start testing to determine whether this
hypothesis applies to the population at large. However,
to date, analyses of gene expression and polymorphisms
have presented contradictory results, muddying our
ability to determine any phenotypical connections to
cancers. In addition, many of those studies do not
measure DNA repair capacity
per se
. Thus, further
genetic and biochemical analyses are needed to deter-
mine the true clinical importance of individual SNPs
and certain groups of SNPs.
24
2. Rethink Testing Methods for Gene
Expression
Gene expression profiling for DNA repair activity is
still in its infancy. While investigators can test for
a single gene that is known to be altered in a certain
cancer, that gene by itself may not accurately represent
DNA repair activity. Testing also needs to take into
account compensatory mechanisms and other alter-
ations that may not yet be characterized. Thus, testing
for gene combinations may not paint a complete picture
of DNA repair ability. Similarly, protein expression
levels may not equate to DNA repair ability, or lack
thereof. Researchers are far from identifying the entire
genome and epigenome of normal versus tumorigenic
DNA repair. But it is equally important, if not more
important, to understand the functional interactions of
those pathways and their regulatory components.
Thus,
functional
repair assays, such as measurements
of the ability of proteins to localize into foci, may
provide a better measure of repair activity without
needing to know the status of all of every repair compo-
nent (see Chapter 7).
5. Develop Readily Available Tests for
Biomarkers
The specialized equipment needed for current
testing, such as the Comet assay, is not practical in a clin-
ical setting. Issues such as protein extraction procedures,
antibody selection, and detection limits need to be
addressed to create and optimize useful clinical assays.
This includes exploring and expanding upon the use
of new technologies, such as microfluidics and xMAP
technology.
25
New laboratory tests that are specific,
accurate, and fast are needed to identify defects in
sensing and/or repairing DNA damage
particularly
e
that of DSBs
to pinpoint the proper pathway-specific
therapeutic strategy for patients.
7
e
3. Identify Biomarkers for Cancer
This encompasses two realms:
6. Develop Selective Inhibitors Against Those
Biomarkers
While many inhibitors are in preclinical or clinical
development and more potential inhibitors are being
identified, researchers need to zero in on which step
and which protein in a particular DNA repair pathway
will produce the greatest therapeutic results when
blocked. This relates to the first action item of more fully
characterizing each pathway.
Determining the best way to assess biomarkers as
prognostic or predictive indicators (testing, scoring,
defining/quantifying cutoff points).
Employing the right [best] technologies to detect
existing variants with altered expression, as well as
to find new combinations of alleles. It is not enough
to perform genotyping, as that can identify only
known SNPs. To discover new sequence variants
