Department of Radiology and Bio-X Program Canary Cancer at Stanford for Cancer Early Detection, Stanford University
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Xu, Y., Liu, H., Chang, E., Jiang, H., Cheng, Z. Cerenkov Luminescence Imaging (CLI) for Cancer Therapy Monitoring. J. Vis. Exp. (69), e4341, doi:10.3791/4341 (2012).
In molecular imaging, positron emission tomography (PET) and optical imaging (OI) are two of the most important and thus most widely used modalities1-3. PET is characterized by its excellent sensitivity and quantification ability while OI is notable for non-radiation, relative low cost, short scanning time, high throughput, and wide availability to basic researchers. However, both modalities have their shortcomings as well. PET suffers from poor spatial resolution and high cost, while OI is mostly limited to preclinical applications because of its limited tissue penetration along with prominent scattering optical signals through the thickness of living tissues.
Recently a bridge between PET and OI has emerged with the discovery of Cerenkov Luminescence Imaging (CLI)4-6. CLI is a new imaging modality that harnesses Cerenkov Radiation (CR) to image radionuclides with OI instruments. Russian Nobel laureate Alekseyevich Cerenkov and his colleagues originally discovered CR in 1934. It is a form of electromagnetic radiation emitted when a charged particle travels at a superluminal speed in a dielectric medium7,8. The charged particle, whether positron or electron, perturbs the electromagnetic field of the medium by displacing the electrons in its atoms. After passing of the disruption photons are emitted as the displaced electrons return to the ground state. For instance, one 18F decay was estimated to produce an average of 3 photons in water5.
Since its emergence, CLI has been investigated for its use in a variety of preclinical applications including in vivo tumor imaging, reporter gene imaging, radiotracer development, multimodality imaging, among others4,5,9,10,11. The most important reason why CLI has enjoyed much success so far is that this new technology takes advantage of the low cost and wide availability of OI to image radionuclides, which used to be imaged only by more expensive and less available nuclear imaging modalities such as PET.
Here, we present the method of using CLI to monitor cancer drug therapy. Our group has recently investigated this new application and validated its feasibility by a proof-of-concept study12. We demonstrated that CLI and PET exhibited excellent correlations across different tumor xenografts and imaging probes. This is consistent with the overarching principle of CR that CLI essentially visualizes the same radionuclides as PET. We selected Bevacizumab (Avastin; Genentech/Roche) as our therapeutic agent because it is a well-known angiogenesis inhibitor13,14. Maturation of this technology in the near future can be envisioned to have a significant impact on preclinical drug development, screening, as well as therapy monitoring of patients receiving treatments.
1. Tumor Model
4. Representative Results
Visual comparison between CLI and PET images can be easily carried out. After unifying the scale bar across images from the same modality and place CLI and PET images side by side one can see in this representative panel (Figure 2A) that both CLI and PET revealed significantly decreased signals from H460 xenografts in treated mice from pre-treatment to day 3, suggesting significant therapeutic effect. As a comparison, moderately increased to unchanged signals were observed in untreated mice during the same time period (data not shown). By visual inspection alone one can observe that there is a good consistence between tumor contrasts that are visualized from CLI and PET. In fact, this visual correlation has sufficient resolution to show central necrosis of the tumor secondary to the anticancer treatment regimen (please compare the CLI and PET images from Day 3). To validate the imaging findings quantifications and correlation analysis can be carried out.
Quantifications of CLI and PET images and a simple fitting via linear regression showed that the two modalities indeed had an excellent correlation (Figure 2B, R2=0.9309 for 18F-FLT probed treatment group). Notably, in all of our CLI and PET imaging studies with different tumor models and different anticancer drugs the slopes of the fits are also remarkably close, thus suggesting an excellent fit of linear regression even of all data are conglomerated (data not shown). Both representative images are adapted from our previous publication12.
Figure 1. Schematic of experimental design of PET and CLI studies. Tumors were implanted bilaterally in shoulder region and allowed to grow to 150-200 mm3, and tumor-bearing mice were subjected to in vivo imaging via PET and CLI at day -1, 1, and 3. Bevacizumab treatment was performed by 2 injections of 20 mg/kg at days 0 and 2.
Figure 2. (A) In vivo CLI and PET images of mice bearing H460 xenografts treated with Bevacizumab before treatment (pre-scan) and after treatment (day 3). (B) Corresponding quantitative analysis of CLI and PET results (n=3) and their correlations. Images adapted from (6). Click here to view larger figure.
CLI is emerging as a promising molecular imaging technique that has found potentials in many basic science research applications and even clinical use4,5,15,16,17. The major advantages of CLI over traditional nuclear imaging modalities such as PET stem from its use of OI instruments, which are easier to use, characterized by short acquisition time and high throughput, significantly less expensive, and more widely available to researchers. Additionally, what sets CLI apart from OI in general is its use of β-emitting labeled molecules as imaging probes, many of which have been approved by the Food and Drug Administration (FDA), unlike traditional OI agents. With these unique and desirable qualities, CLI has quickly garnered attention from the field of molecular imaging. Yet its potentials in preclinical and clinical applications are yet to be fully investigated.
Cancer therapy monitoring is one of the areas where CLI can have some significant utility. It is a very important area that is key to probe development, drug screening, and even tailoring cancer therapy for patients. Currently, preclinical cancer therapy monitoring is carried out almost exclusively via nuclear imaging modalities such as PET. Therefore CLI provides a very attractive alternative to PET, particularly given that there is an excellent correlation between CLI and PET images. Yet, another advantage of CLI for therapy monitoring lies in the fact that CLI can image not only β+-emitters, but also β-emitters such as 32P, 90Y, and 131I, all of which are clinically relevant.
However, CLI is not without flaws. The reliance on OI instruments dictates that CLI suffers from some shortcomings that are intrinsic to optical imaging such as signal attenuation and scattering in living tissues. Moreover, the particular spectrum of CR also results in limited signal intensity and subsequently, the deeper the signal from body surface, the lower the sensitivity, and the poorer the quantification capability6. However, while the shortcomings can be viewed to be significant, one can largely bypass these obstacles in preclinical research by employing small animals such as mice. More importantly, there are at least a couple of clinical areas that can potentially benefit from CLI cancer therapy monitoring. Monitoring superficial disease entities such as dermatological inflammatory conditions and cancers can serve as one good example. Moreover, disease entities that are deep yet accessible by charge-coupled device or fiber optic based techniques can use the excellent sensitivity and quantification capability of CLI as well. Yet another exciting possibility lies in using CLI to help surgeons obtain anatomic and functional information about tumors in the operating room. Two recent proof-of-concept studies have demonstrated detection and resection of tumors in mice with intraoperative image guidance thanks to CLI18,19.
No conflicts of interest declared.
We acknowledge support from the National Cancer Institute (NCI) R01 CA128908 and Stanford Medical Scholar Research Fellowship. No other potential conflict of interest relevant to this article was reported.
|H460 Cell Line||American Type Culture Collection||ATCC Number: HTB-177|
|RPMI 1640 Medium||Invitrogen Life Technologies||12633-012|
|Fetal Bovine Serum||Invitrogen Life Technologies||10091-148|
|Penicillin/Streptomycin||Invitrogen Life Technologies||15640-055|
|Phosphate-Buffered Saline||Invitrogen Life Technologies||10010-023|
|Female Athymic Nude Mice||Charles River Laboratories, Inc.||Strain Code: 088|
|MicroPET Rodent R4||Siemens Medical Solutions USA, Inc.||N/A|
|Isoflurane (Aerrane)||Baxter||Baxter Number: AHN3637|
|IVIS Spectrum||Caliper Life Sciences||N/A|