The ICONA
Project
The identification of pretreatment
tumor characteristics that are associated with sensitivity
or resistance to anticancer drugs may minimize the proportion
of patients who would not benefit from treatment and
maximize the proportion of those who would.
High-throughput technologies such as cDNA- or protein-microarrays,
may pave the way for a systematic identification of
factors linked to sensitivity to therapy. Both approaches,
though, still present major pitfalls. Microarrays
that monitor cDNA expression levels require extensive
methodological standardization to obtain reproducible
results, and the interpretation of the large body
of information can be a difficult task without the
support of advanced bioinformatics. A further limitation
is that mRNA levels closely reflect protein levels
in only 20% of the cases, and sometimes this discrepancy
can be by 20-fold or larger. On the other hand, current
proteomic approaches are hampered by the labor-intensive
and poorly reproducible procedures that are required
for the quantification of proteins of interest. With
the aid of leader experts of member Universities,
CINBO plans to develop a protein chip based on a radically
different and innovative approach.
ICONA proteomic chip
This will be based on fluorescent
dye-labelled recombinant proteins, where quantification
of protein expression in tumor tissues will be performed
by competition with known amounts of the correspondent
fluorescent proteins. Key advantages of this approach
over existing methods are that:
a. tumor lysates are tested as unmodified, native
samples;
b. accurate quantification is obtained;
c. the assays is based on entirely defined systems
(antibodies and recombinant proteins).
These characteristics have important implications
for the validation of reagents for clinical use and
for ease/robustness of performance of the measuring
assay. A proteome-wide extension of this approach
is not beyond reach, given current technologies for
parallel peptide synthesis or recombinant protein
production in E. coli or yeast.
As labeling reactions are stochastic in nature (none,
single or multiple labels per protein molecule follow
typical Poisson distributions), mixtures of heterogeneously-labeled
proteins are bound to be produced. This will affect
(a) the accuracy of quantitative measurements (b)
the reproducibility of the labeling reaction, and
(c) will potentially interfere with binding to the
antibody on the revealing chip. Thus, a second-generation
ICONA assay will be developed, using proprietary technologies.
ICONA proof
of concept will be obtained in metastatic colorectal
cancer and in primary rectal cancer. |
