Improved detection of ovarian cancer metastases by intraoperative quantitative fluorescence protease imaging in a pre-clinical model

doi:10.1016/j.ygyno.2008.11.018

Gynecologic Oncology

Volume 112, Issue 3, March 2009, Pages 616-622

https://doi.org/10.1016/j.ygyno.2008.11.018 Get rights and content

Abstract

Objectives

Cytoreductive surgery is a cornerstone of therapy in metastatic ovarian cancer. While conventional white light (WL) inspection detects many obvious tumor foci, careful histologic comparison has shown considerable miss rates for smaller foci. The goal of this study was to compare tumor detection using WL versus near infrared (NIR) imaging with a protease activatable probe, as well as to evaluate the ability to quantify NIR fluorescence using a novel quantitative optical imaging system.

Methods

A murine model for peritoneal carcinomatosis was generated and metastatic foci were imaged using WL and NIR imaging following the i.v. administration of the protease activatable probe ProSense750. The presence of tumor was confirmed by histology. Additionally, the ability to account for variations in fluorescence signal intensity due to changes in distance between the catheter and target lesion during laparoscopic procedures was evaluated.

Results

NIR imaging with a ProSense750 significantly improved upon the target-to-background ratios (TBRs) of tumor foci in comparison to WL imaging (minimum improvement was approximately 3.5 fold). Based on 52 histologically validated samples, the sensitivity for WL imaging was 69%, while the sensitivity for NIR imaging was 100%. The effects of intraoperative distance changes upon fluorescence intensity were corrected in realtime, resulting in a decrease from 89% to 5% in signal variance during fluorescence laparoscopy.

Conclusions

With its molecular specificity, low background autofluorescence, high TBRs, and quantitative signal, optical imaging with NIR protease activatable probes greatly improves upon the intraoperative detection of ovarian cancer metastases.

Introduction

Cytoreductive surgery in metastatic ovarian cancer has been shown to have a significant beneficial effect on patient outcome and represents an important cornerstone in the treatment of this disease [1], [2], [3]. Current guidelines define an “optimal” debulking procedure as one in which the surgeon leaves the abdomen with less than 1 cubic cm of residual tumor; additional evidence suggests that greater improvements in clinical outcome can be achieved by setting even lower thresholds for residual tumor [4]. There is an unmet clinical need for an intraoperative imaging modality to assist the surgeon in identifying tumor foci. Improved intraoperative tumor detection has the potential to yield both more accurate staging as well as decreased residual metastatic disease.

A variety of imaging modalities have been applied towards more sensitive detection of diffuse peritoneal carcinomatosis, with near infrared (NIR) optical imaging showing particular promise. The advent of fluorescent molecular probes with a propensity for differentially illuminating tumor foci against normal background tissue has allowed for the application of optical imaging towards a number of cancer models [5], [6], including ovarian cancer [7], [8], [9]. Our laboratory has developed a novel class of “smart” probes that are administered in an optically silent state, but in the presence of specific proteases, are cleaved and thus activated, resulting in up to a hundred-fold signal amplification [10], [11]. Protease, particularly cathepsin B, up-regulation has been shown to occur in a variety of disease states, including ovarian cancer[12], and thus these probes represent highly sensitive molecular beacons that may be applied for the detection of metastatic disease [13].

One commonly cited drawback to intravital optical imaging is the inability to quantify fluorescent signal. The collected signal varies dramatically as the distance between the tissue under investigation and the imaging apparatus changes during an operative procedure; this variability precludes quantitative analysis of the molecular probe's signal intensity, thus severely limiting the applicability of optical imaging to reliable minimally invasive tumor detection. To account for this distance dependence, we have recently developed methods that allow for the intravital and real-time quantification of NIR fluorescence [6], [14]. In this study, we demonstrate the ability of NIR “smart” probes to detect ovarian cancer metastases with high TBRs, as well as to improve upon the sensitivity for the visualization of cancer foci adherent to and within abdominal structures, versus standard white light (WL) evaluation. Finally, we apply a distance dependence correction algorithm to allow fluorescence imaging in a quantitative manner during in vivo laparoscopic procedures.

Section snippets

NIR activatable probe

The imaging probe used for this study is a commercially available protease-activatable NIR fluorescent probe, Prosense750 (VisEn Medical, Woburn, MA). The probe's structure consists of a synthetic graft polymer composed of poly-l-lysine that is sterically protected by multiple methoxypolyethylene glycol side chains. Conjugated onto this lysine backbone are multiple NIR fluorochromes whose fluorescence is quenched by their proximity to one another in a phenomenon similar to fluorescence

Ex vivo imaging of metastatic ovarian cancer with conventional WL and ProSense750

To evaluate the ability of ProSense750 to detect ovarian cancer metastases, explanted abdominal and pelvic organs from a peritoneal carcinomatosis animal model were imaged under both conventional WL and NIR fluorescence; the results of this study are summarized in Fig. 1 by images of representative tissue samples with tumor foci. Fig. 1A demonstrates a section of omentum with a large, fixed, and irregular mass visible in WL that fluoresces vividly under NIR interrogation; histologic analysis

Discussion

Optical imaging with molecularly targeted probes has numerous advantages compared to other detection methods for evaluation of potential residual disease. The approach focally highlights disease, lacks ionizing radiation, and has a low barrier to incorporation in the operating room. Near infrared protease imaging in particular offers a number of distinct advantages in the detection of ovarian cancer peritoneal metastases, by targeting an enzyme class widely present across most ovarian cancers

Conflict of interest statement

RW is a shareholder and member of the Board of Directors for VisEn Medical. All other authors have no conflicts of interest to declare.

Acknowledgments

This research was supported in part by NIH grant R01-EB001872 and U24-CA92782.

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Improved detection of ovarian cancer metastases by intraoperative quantitative fluorescence protease imaging in a pre-clinical model

Abstract

Objectives

Methods

Results

Conclusions

Introduction

Section snippets

NIR activatable probe

Ex vivo imaging of metastatic ovarian cancer with conventional WL and ProSense750

Discussion

Conflict of interest statement

Acknowledgments

Obstet. Gynecol.

Obstet. Gynecol.

Gynecol. Oncol.

Gynecol. Oncol.

Cancer statistics

CA: Cancer J. Clinic.

Near infrared thoracoscopy of tumoral protease activity for improved detection of peripheral lung cancer

Int. J Cancer

Miniaturized multichannel near infrared endoscope for mouse imaging

Mol. Imaging

Catheter-based in vivo imaging of enzyme activity and gene expression: feasibility study in mice

Radiology