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Article Released Thu-3rd-October-2013 06:10 GMT
Contact: gro-pr Institution: RIKEN
 Advanced technology for gene expression analysis can facilitate drug development

Researchers from the RIKEN Center for Life Science Technologies report a new method to monitor and quantify the activity of gene promoters during the response to a drug, using the advanced gene expression analysis method CAGE followed by single-molecule sequencing.


When developing new drugs, monitoring cellular responses to candidate
compounds is essential for assessing their efficacy and safety.
Researchers from the RIKEN Center for Life Science Technologies report a
new method to monitor and quantify the activity of gene promoters during
the response to a drug, using the advanced gene expression analysis
method CAGE followed by single-molecule sequencing. This research paves
the way to a more precise analysis of cellular responses to drugs, at
the level of individual promoters.

The study is published this week in the journal CPT: Pharmacometrics &
Systems Pharmacology.

Microarray-based technologies are widely used to monitor cellular
changes in response to drug administration at the level of genes.
However, microarrays have several limitations due to the fact that they
rely on pre-designed oligonucleotide probes and detection based on
hybridization.

In order to circumvent the limitations imposed by the use of
microarray-based technology for the development of new drugs, Dr
Harukazu Suzuki and his team at CLST developed a new technique combining
Cap Analysis of Gene Expression (CAGE) with 3rd generation,
single-molecule sequencing.

CAGE is a method developed at RIKEN to comprehensively map human
transcription start sites and their promoters, and quantify the set of
mRNAs in a cell, also called the transcriptome.

During CAGE the 5’-end of mRNAs is sequenced in order to produce a
series of 20-30 nucleotide sequences that can then be mapped onto the
genome and provide information about the level of expression of genes.

Dr Suzuki and his team used CAGE, combined with a single-molecule
sequencer, to monitor the effect of three drugs, U0126, wortmannin and
gefitinib on human breast cancer cells.

U0126 and wortmannin are known to inhibit the Ras-ERK and
phosphatidylinositol-3-kinase (PI3K)-Akt signalling pathways within
cells. Gefinitib is a potent inhibitor of the epidermal growth factor
receptor kinase (EGFR kinase) and mainly inhibits the Ras-ERK and
PI3K-Akt pathways downstream of EGFR.

The researchers identified a distinct set of promoters that were
affected by low doses of the drugs, and therefore showed sensitivity to
a weak inhibition of the Ras-ERK and PI3K-Akt signal-transduction
pathways. This level of precision would would have been very difficult
to achieve using microarray-based profiling.

Furthermore, a quantitative analysis showed that the inhibitory profiles
of both U0126 and wortmannin are constitutive components of the
transcriptome profile obtained by inhibition of the EGFR kinase. Using a
regression model, the researchers were able to quantitatively predict
the promoter activity profile of gefitinib, based on the U0126 and
wortmannin profiles.

These results demonstrate the potential utility of highly quantitative
promoter activity profiling in drug research.

“Quantitative transcriptome analysis is potentially widely applicable to
determine the target proteins and action mechanisms of uncharacterized
compounds,” concludes Dr Suzuki. “Our study paves the wayfor
quantitative analysis of drug responses at the promoter level, and
moreover, is potentially applicable for the evaluation of combinatorial
or serial drug treatment in a clinical setting,” he adds.

# # #

This press release is available online at:
http://www.riken.jp/en/pr/press/2013/20131003_3/

Dr Suzuki is available for interviews on the phone at +81 45-503-9222
or by email at harukazu@gsc.riken.jp

Alternatively please contact:
Juliette Savin
RIKEN
Tel: +81-(0)48-462-1225
Email: pr@riken.jp

Reference:
Kazuhiro Kajiyama et al.
“Capturing drug responses by quantitative promoter activity profiling”
CPT: Pharmacometrics & Systems Pharmacology, 2013 DOI: 10.1038/psp.2013.53


The paper is available at
http://www.nature.com/psp/journal/v2/n9/full/psp201353a.html


About RIKEN
RIKEN is Japan's flagship research institute for basic and applied
research. Over 2500 papers by RIKEN researchers are published every year
in reputable scientific and technical journals, covering topics ranging
across a broad spectrum of disciplines including physics, chemistry,
biology, medical science and engineering. RIKEN's advanced research
environment and strong emphasis on interdisciplinary collaboration has
earned an unparalleled reputation for scientific excellence in Japan and
around the world.

About the Center for Life Science technologies
The RIKEN Center for Life Science Technologies aims to develop key
technologies for breakthroughs in the medical and pharmaceutical
applications of life science as well as conduct ground-breaking research
and development for the next-generation life sciences.

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