Estrogen induced proliferation of mutant cells is widely understood to be the one of major risk
determining factor in the development of breast cancer. Hence determination of the Estrogen
Receptor[ER] status is of paramount importance if cancer pathogenesis is to be detected and rectified
at an early stage. Near Infrared Fluorescence [NIRf] Molecular Optical Imaging is emerging as a
powerful tool to monitor bio-molecular changes in living subjects. We discuss pre-clinical results in
our efforts to develop an optical imaging diagnostic modality for the early detection of breast cancer.
We have successfully carried out the synthesis and characterization of a novel target-specific NIRf
dye conjugate aimed at measuring Estrogen Receptor[ER] status. The conjugate was synthesized by
ester formation between 17-β estradiol and a hydrophilic derivative of Indocyanine Green (ICG)
cyanine dye, bis-1,1-(4-sulfobutyl) indotricarbocyanine-5-carboxylic acid, sodium salt. In-vitro
studies regarding specific binding and endocytocis of the dye performed on ER+ve [MCF-7] and
control [MDA-MB-231] adenocarcinoma breast cancer cell lines clearly indicated nuclear
localization of the dye for MCF-7 as compared to plasma level staining for MDA-MB-231.
Furthermore, MCF-7 cells showed ~4.5-fold increase in fluorescence signal intensity compared to
MDA-MB-231. A 3-D mesh model mimicking the human breast placed in a parallel-plate DOT
Scanner is created to examine the in-vivo efficacy of the dye before proceeding with clinical trials.
Photon migration and florescence flux intensity is modeled using the finite-element method with the
coefficients (quantum yield, molar extinction co-efficient etc.) pertaining to the dye as obtained from
photo-physical and in-vitro studies. We conclude by stating that this lipophilic dye can be potentially
used as a target specific exogenous contrast agent in molecular optical imaging for early detection of
breast cancer.
Molecular imaging provides the in vivo characterization of cellular molecular events involved in normal and pathologic
processes. With the advent of optical molecular imaging, specific molecules, proteins and genes may be tagged with a
luminescent reporter and visualized in small animals. This powerful new tool has pushed in vivo optical imaging to the
forefront as it allows for direct determination of drug
bio-distribution and uptake kinetics as well as an indicator of
biochemical activity and drug efficacy. Although optical imaging encompasses diverse techniques and makes use of
various wavelengths of light, a great deal of excitement in molecular research lies in the use of tomographic and
fluorescence techniques to image living tissues with near-infrared (NIR) light. Nonionizing, noninvasive near-infrared
optical imaging has great potential to become promising alternative for breast cancer detection. Fluorescence
spectroscopy studies of human tissue suggest that a variety of lesions show distinct fluorescence spectra compared to
those of normal tissue. It has also been shown that exogenous dyes exhibit selective uptake in neoplastic lesions and may
offer the best contrast for optical imaging. Use of exogenous agents would provide fluorescent markers, which could
serve to detect embedded tumors in the breast. In particular, the ability to monitor the fluorescent yield and lifetime may
also enable biochemical specificity if the fluorophore is sensitive to a specific metabolite, such as oxygen.
As a first step, we have synthesized and characterized one such NIR fluorescent dye conjugate, which could potentially
be used to detect estrogen receptors (ER)[2] . The conjugate was synthesized by ester formation between 17-β estradiol
and a hydrophilic derivative of indocyanine green (ICG) cyanine dye, bis-1, 1-(4-sulfobutyl) indotricarbocyanine-5-
carboxylic acid, sodium salt. The ester formed was found to have an extra binding ability with the receptor cites as
compared to ICG, which was established by the partition coefficient studies. The replacement of the sodium ion in the
ester by a larger glucosammonium ion was found to enhance the hydrophilicity and reduce the toxic effect on the cell
lines. The excitation and emission peaks for the conjugate were recorded in the NIR region as 750nm and 788nm
respectively. The ester was found nontoxic on adenocarcinoma breast cancer cell lines MCF-7/MDA-MB-231. Specific
binding and endocytosis of the estrogen-labeled conjugate was studied on the MCF-7 (ER positive) and MDA-MB-231
(ER negative). Conjugate staining of MCF-7 cells showed ~ 4-fold increase in signal intensity compared to MDA-MB-
231. Further, estrogen molecules were found to be specifically localized to the nuclear region of MCF-7 cells, whereas
MDA-MB-231 showed plasma membrane staining.
This technique offers the potential of noninvasive detection of hormone receptor status in breast cancer cells and would
help in decreasing the load of unnecessary biopsies. Here, we have reported the progress made in the development of a
novel NIR external contrast agent and the work is in progress to use this conjugate for the molecular based, diagnostic
imaging of breast cancer.
The use of near-infrared (NIR) spectroscopy to interrogate deeper tissue volume has shown enormous potential for molecular-based non-invasive imaging when coupled with appropriate excitable dyes. As most of the breast cancers are hormone dependent hence determination of the hormonal receptor status gains paramount importance when deciding the treatment regime for the patient. Since proliferations of the breast cancer cells are often driven by estrogen, we focus on to developing a technique to detect estrogen receptor status. As a first step, the objective of this work was to synthesize and characterize one such novel NIR fluorescent (NIRF) conjugate, which could potentially be used to detect estrogen receptors. The conjugate was synthesized by ester formation between 17-b estradiol and a cyanine dye namely: bis-1, 1-(4-sulfobutyl) indotricarbocyanine-5-carboxylic acid, sodium salt. The cyanine dye is a hydrophilic derivative of indocyanine green (ICG). The ester formed was found to have an extra binding ability with the receptor cites as compared to ICG, which was established by the partition coefficient studies. This cyanine dye has a partition coefficient less than 0.005 as compared to that of ICG (>200)[1]. In addition the ester showed enhanced fluorescent quantum yield than ICG. The replacement of the sodium ion in the ester by a larger glucosammonium ion was found to enhance the hydrophilicity and reduce the toxic effect on the cell lines. The excitation and emission peaks for the conjugate were recorded in the NIR region as 750nm and 788nm respectively. The ester developed was tested on the breast cancer cell lines MCF-7 and found non-toxic. The tagging characteristics were pivotal determinants underlying the ability of the fluorescent conjugate in binding the estrogen receptor of the breast cancer cells. This technique offers the potential of non-invasive detection of hormone receptor status in vivo and may help in decreasing the load of unnecessary biopsies. Here, we have reported the progress made in the development of a novel NIR external contrast agent and the work is in progress to use this conjugate for the molecular-based, diagnostic imaging of breast cancer.
Access to the requested content is limited to institutions that have purchased or subscribe to SPIE eBooks.
You are receiving this notice because your organization may not have SPIE eBooks access.*
*Shibboleth/Open Athens users─please
sign in
to access your institution's subscriptions.
To obtain this item, you may purchase the complete book in print or electronic format on
SPIE.org.
INSTITUTIONAL Select your institution to access the SPIE Digital Library.
PERSONAL Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.