5th EuropEan MolEcular IMagIng MEEtIng - ESMI

5th EuropEan MolEcular IMagIng MEEtIng – EMIM2010
Integrisense: a novel near-infrared fluorescent probe for α v β 3 integrin and its applications in
drug discovery
Sur C. (1) , Lin S.A. (1) , Gleason A. (1) , Kossodo S. (1) , Pickarski M. (1) , Coleman P. (1) , Rajopadhye M. (2) , Duong L. T. (1) , Yared W. (2) ,
Peterson J. (2) , Bednar B. (1) .
(1) Merck Research Laboratories, Westpoint US
(2) ViSen Medical.
cyrille_sur@merck.com
Introduction: The α v β 3 integrin belongs to the superfamilly
of dimeric transmembrane cell adhesion
receptors involved in critical biological processes
such as cell-to-cell, and cell-to-extracellular
matrix binding. As these activities are central to
pathological conditions such as inflammation and
tumor progression, the RGD-binding α v β 3 integrin
has been considred as clinically-relevant biomarkers
of these disease states. Although, RDG peptides
have been developed for PET and SPECT imaging,
the operating cost of these modalities limits their
widespread deployment in drug discovery teams. In
contrast optical molecular imaging allows the noninvasive
monitoring and quantification of fluorescent
probes in vivo with less expensive equipment
that can be installed at multiple research sites. In
order to take full advantage of the rich biology of integrins,
a partnership has been established between
Merck Research Laboratories and VisEn Medical to
develop a novel near-infrared (NIR) fluorescent integrin
probe, IntegriSense TM . The pharmacological
and biological characterization of IntegriSense TM ,
a peptidomimetic antagonist-based molecule with
improved specificity and binding affinity will be
presented together with research applications in the
oncology and atherosclerosis research fields.
Methods: The in vitro cell biology and pharmacological
profiling of IntegriSense TM were conducted
with recombinant HEK-293 cells expressing α v β 3
and included confocal microscopy for cellular localization
and flow cytometry for binding and kinetic
constant determination. In vivo pharmacodynamics
and tumor localization studies were performed
in female NU/NU mice bearing different human
tumor xenografts. IntegriSense TM NIR fluorescent
signal was acquired with a Fluorescent Molecular
Tomography imaging system (FMT2500). The potential
application of IntegriSense TM in atherosclerosis
was evaluated in apolipoprotein E-deficient
(ApoE-/-) and human cholesteryl ester transfer
protein (CETP) knockin/LDL receptor-deficient
(C57BL/6-Tg(CETP)-Ldlrtm1) mice. Transgenic
as well as control C57Bl/6 mice were fed a cholesterol
enriched diet (9% fat, 0.15% cholesterol) to
induce atherosclerosis. IntegriSense TM NIR signal
was measured in vivo 24 and 48 hours after probe
injection and its localization in inflamed aortas was
further evaluated by ex-vivo imaging and histology.
Results: Integrisense-680 has absorption/emission
spectra centered at 674nm/692nm and an extinction
coefficient of 2.02x10 5 M -1 cm -1 . Binding of IntegriSense
TM to HEK-293 stably expressing recombinant
human α v β 3 yielded a Kd of 4.2±0.6 nM and
a dissociation constant koff of 1.08x10 -4 s -1 . Plasma
pharmacokinetic analysis revealed a two compartmental
profile with t1/2 of 6 min and 210 min corresponding
to clearance of free and bound IntegriSense
TM , respectively. Biodistribution studies in
mice with xenograft tumors showed a specific uptake
of IntegriSense TM by tumor cells and supported
using IntegriSense TM to monitor tumor growth in
animal models.
In vivo measurements with FMT2500 in mice atherosclerosis
models detected a longitudinal increase in
IntegriSense TM fluorescence in the thorax area that
was three to five fold higher when compared with
control animals. This NIR signal originated from
atherosclerotic lesions in the aortic arch, carotid,
and subclavian arteries as confirmed by ex vivo imaging
of the dissected vessels and histopathology.
Conclusions: This report demonstrated that the α v β 3
integrin optical reporter, IntegriSense TM developed
by Merck and VisEn partnership can be used as a
molecular imaging biomarker for in vivo imaging
studies. For instance, IntegriSense TM can be used to
monitor tumor growth as well as to detect inflammation
in atherosclerosis plaques. The ability of
this biomarker to report on several important cellular
and physiological functions makes it a tool of
choice to evaluate preclinically the therapeutic potential
of novel anticancer and atherosclerosis drugs.
EuropEan SocIEty for MolEcular IMagIng – ESMI
day2
Plenary Session on current contribution of IMAGING TECHNOLOGIES to DRUG DEVELOPMENT