Nanomedicine Research Highlights

Feng Chen¹, Weibo Cai^1,2,*

¹ Departments of Radiology and Medical Physics, University of Wisconsin - Madison

² University of Wisconsin Carbone Cancer Center, Madison, WI

*Author for correspondence

WCai@uwhealth.org

Acknowledgements

The authors would like to thank the University of Wisconsin - Madison, the National Institutes of Health (NIBIB/NCI 1R01CA169365), the Department of Defense (W81XWH-11-1-0644), and the American Cancer Society (125246-RSG-13-099-01-CCE) for research support.

Evaluation of: Hu CM, Fang RH, Copp J, Luk BT, Zhang L. A biomimetic nanosponge that absorbs pore-forming toxins. Nat. Nanotechnol. 8(5), 336-340 (2013).

Biomimetic Nanosponge Paves the Way for Future Biodetoxification in the Clinic

Pore-forming toxins (PETs) are common toxins that can disrupt cells by forming pores in cellular membranes and altering their permeability. By camouflaging biocompatible poly(lactic-co-glycolic acid) nanoparticles with preformed mouse red blood cell (RBC) membrane-derived vesicles, Prof. Liangfang Zhang and co-workers at UC San Diego (USA) developed an intriguing biomimetic nanosponge, which could function as a decoy to efficiently absorb α-toxin (and other similar PFTs) both in vitro and in vivo. The capability of the nanosponge in neutralizing α-toxin was convincingly demonstrated, which led to significant reduction in RBC hemolysis when mixing α-toxin/nanosponge with purified RBCs. More importantly, markedly improved survival rate was achieved by intravenous injection of the nanosponge either immediately before or after administration of α-toxin into mice, clearly demonstrating the efficacy of such nanosponge to sequester the toxin in vivo. The membrane-toxin affinity was found to be dependent upon the type of toxin and the sources of RBC membranes. Although more effort is needed to further optimize the detoxification capability in vivo, and concerns about the long-term in vivo stability of the nanosponge will need to be addressed in future studies, such a biomimetic nanosponge indeed holds promising potential as injectable nanocarriers for future biodetoxification in the clinical setting.

Evaluation of: Wang C, Xu H, Liang C et al. Iron Oxide@Polypyrrole Nanoparticles as a Multifunctional Drug Carrier for Remotely Controlled Cancer Therapy with Synergistic Antitumor Effect. ACS Nano 7 (8), 6782–6795 (2013)

Biocompatible Theranostic Agent Exhibits Remotely Controllable Synergistic Antitumor Effect

Design and synthesis of novel multifunctional theranostic nanomedicine with both therapeutic and imaging components have attracted tremendous attention over the last several years. In their latest endeavor, Prof. Zhuang Liu and co-workers at Soochow University (China) offered a smarter and safer design by coating biocompatible iron oxide (Fe₃O₄) nanoparticles with a layer of biodegradable near-infrared (NIR) light-absorbing polypyrrole (PPy). The resulting Fe₃O₄@PPy core/shell nanostrucuture, upon loading of the anticancer drug doxorubicin and with the control of NIR light and magnetic field, could exhibit synergistic antitumor effect in the 4T1 murine breast cancer model through a combination of photothermal therapy and chemotherapy. Although marked tumor suppression could only be achieved through intratumoral injection of the Fe₃O₄@PPy core/shell nanostrucuture in this work, future investigation and optimization on how to achieve such synergistic therapeutic effect upon systemic administration will provide this novel biocompatible theranostic agent a brighter future for potential clinical translation.

 

Evaluation of: Xing H, Zheng X, Ren Q et al. Computed tomography imaging-guided radiotherapy by targeting upconversion nanocubes with significant imaging and radiosensitization enhancements. Sci. Rep. 3, 1751 (2013).

Targeted Upconversion Nanocubes Enhance Efficacy of Computed Tomography-Guided Radiotherapy

Although radiation therapy can control tumor growth in clinical oncology by damaging the DNA of exposed tissue with ionizing radiation, its clinical use is still quite limited due to the inaccurate positioning and inherent radioresistance of many tumor types. To address these issues, Prof. Jianlin Shi and co-workers at Shanghai Institute of Ceramics (Chinese Academy of Sciences) developed a novel water soluble arginine-glycine-aspartic acid (RGD)-conjugated BaYbF₅:Er³⁺ upconversion nanocubes. These intriguing nanocubes could function as optical imaging tracker, computed tomography (CT) contrast agent, and more importantly radiosensitization enhancer. Active targeting of integrin α_vβ₃ with RGD-conjugated nanocubes was found to have ~3-fold enhancement in U87MG tumor uptake (which expresses a high level of integrin α_vβ₃), when compared to that of passive targeting alone. The capability of RGD-conjugated nanocubes as radiosensitizer was further demonstrated in vivo, where significantly enhanced tumor growth inhibition was observed after receiving the same radiation dose as the multiple control groups. Although the biocompatibility and long term effect of such heavy metal-based nanocubes need to be further elucidated in the future, such targeted upconversion nanocubes indeed hold promising potential for multimodal cancer imaging and more efficacious image-guided radiation therapy.

 

Evaluation of: Wang S, Huang P, Nie L et al. Single continuous wave laser induced photodynamic/plasmonic photothermal therapy using photosensitizer-functionalized gold nanostars. Adv. Mater. 25(22), 3055-3061 (2013).

Single Continuous Wave Laser Enables Synergistic Photodynamic and Photothermal Therapy

When compared with conventional chemotherapy and radiation therapy, photodynamic therapy (PDT) is a relatively new and much safer cancer treatment strategy, which relies on the generation of singlet oxygen after laser irradiation of photosensitizers in the presence of tissue oxygen. However, many problems such as limited tissue penetration depth of visible laser, photosensitizer self-destruction upon laser irradiation, severe local hypoxia, etc. significantly hinder the broad clinical applications of PDT. By developing a novel Chlorin e6-functionalized gold nanostars (i.e. GNS-PEG-Ce6), Dr. Xiaoyuan Chen and co-workers at the National Institute of Biomedical Imaging and Bioengineering (USA) offered a new therapeutic strategy, which enabled an impressive synergistic anticancer effect of PDT and plasmonic photothermal therapy (PPTT) in tumor-bearing mice upon irradiation with a single continuous wave laser (671 nm). Adapting the localized surface plasmon resonance of GNS to fit the absorption spectrum of Ce6 to ensure both PDT and PPTT by single continuous wave laser is vital for this approach, which was successfully achieved in this work. In vitro and in vivo (with intratumoral injection) studies clearly demonstrated significantly enhanced antitumor effects with the simplified single laser irradiation process. Future demonstration of synergistic combination of PDT and PPTT after systemic administration of GNS-PEG-Ce6 will make it more desirable for translational from bench to the bedside.

 

Evaluation of: Wang Y-F, Liu G-Y, Sun L-D, Xiao J-W, Zhou J-C, Yan C-H. Nd³⁺-Sensitized Upconversion Nanophosphors: Efficient In Vivo Bioimaging Probes with Minimized Heating Effect. ACS Nano 7 (8), 7200–7206, 2013.

Nd³⁺-Doping Enables “Cool” Excitation of Upconversion Nanoparticles at 808 nm

Lanthanide ion-doped upconversion nanoparticle (UCNP) has recently emerged as an exciting nanoplatform for molecular imaging because of its unique upconversion luminescence properties (e.g. high tissue penetration depth, extremely low background signal, just to name a few). However, the typical requirement of a 980 nm laser as the excitation source can lead to overheating of tissue, due to the strong absorbance by water at this wavelength in living subjects. In this report, Prof. Chun-Hua Yan and co-workers at Peking University (China) offered a “cool” solution to this problem by taking advantage of Nd³⁺→Yb³⁺ energy transfer in their newly designed core/shell structured NaGdF₄:Er/Yb@NaGdF₄:Nd/Yb UCNP (abbreviated as Er@Nd). Systematic in vitro and in vivo investigation clearly demonstrated a high upconversion efficiency of Er@Nd upon excitation with an 808 nm laser, along with dramatically reduced tissue heating in living mice. This study extended the upconversion excitation spectrum to shorter wavelengths, which is still in the near-infrared range and highly suitable for in vivo imaging applications, and could inspire the design of future generations of more interesting UCNPs for better and safer in vivo optical imaging.

 

Highlights from the latest articles in Radiochemistry and Radiotracer Development

Shuanglong Liu & Zibo Li*

Molecular Imaging Center, Department of Radiology, University of Southern California, Los Angeles, California 90033

*Author for correspondence: ziboli@usc.edu

Development of a New Thiol Site-Specific Prosthetic Group

Evaluation of: Yue X, Kiesewetter DO et al. Development of a New Thiol Site-Specific Prosthetic Group and Its Conjugation with [Cys⁴⁰]-exendin-4 for In Vivo Targeting of Insulinomas. Bioconjug Chem. 2013, 24(7), 1191–1200.

Radiochemistry has played a critical role in PET probe developments and applications. Novel and efficient labeling methods are being developed to meet the increasing demand for nuclear medicine. In this study, Yue and coworkers developed a new prosthetic group, [¹⁸F]FPenM, for conjugation with free thiol group in proteins and peptides. Among various maleimide-functionalized prosthetic agents, [¹⁸F]FPenM has small molecular weight, which may minimally compromise the ligand binding affinity after conjugation.

As an example, the GLP-1R targeted agent [¹⁸F]FPenM-[cys⁴⁰]-exendin-4 was also successfully synthesized using [¹⁸F]FPenM. This tracer showed high tumor-to-liver and tumor-to-kidney ratios for GLP-1R-targeted insulinama imaging. Although systematic comparison is still needed, [¹⁸F]FPenM-[cys⁴⁰]-exendin-4 already demonstrated advantages over other labeling analogs such as NOTA-MAL-[cys⁴⁰]-exendin-4 and [Lys⁴⁰(natGa-DOTA)]exendin-3 in term of the higher GLP-1R binding affinity and faster kidney clearance in vivo. The efficient labeling strategy and superior tumor targeting capability make [¹⁸F]FPenM-[cys⁴⁰]-exendin-4 a promising PET agent not only for insulinaoma imaging, but also for native or transplanted pancreatic β-cell imaging.

¹⁸F-Labeled Picolinamide Probes for Malignant Melanoma Imaging

Evaluation of: Liu H, Liu S, et al. Development of ¹⁸F-Labeled Picolinamide Probes for PET Imaging of Malignant Melanoma. J Med Chem 2013, 56(3), 895–901.

In addition to novel labeling strategies, labeling process optimization represents another important aspect of radiochemistry research. In an ideal situation, the radiotracer should be synthesized in high yield with very simple purification step(s). For example, NOTA-Al-¹⁸F labeled RGD peptides has been translated into clinical trials partially due to the simplified labeling procedures. However, for imaging probes based on small organic molecules, a different approach might be preferred due to the relatively large size of NOTA-Al-¹⁸F, which may significantly alter the in vivo behaviors of the final imaging probes.

Malignant melanoma is one of the most aggressive and lethal cancers with increasing incidence in the Caucasian population. In order to detect melanoma at its earliest stages with accurate molecular imaging techniques, Ren and co-workers have developed the ¹⁸F labeled benzamide (¹⁸F-FBZA) for melanin-targeted melanoma imaging. Although promising preliminary results has been obtained in the animal studies, the multistep radiosynthesis of ¹⁸F-FBZA set a high barrier for its large scale production and potential clinical translation. In this study, Liu and co-workers designed and synthesized two novel PET probes based on the picolinamide structure for melanoma diagnosis. The simplified one-step radiolabeling method provided final PET agent in one-hour with 10–20% decay-corrected yields. In fact, this paper reported not only the improved radiolabeling method, but also the optimized in vivo imaging result. In particular, N-(2-(diethylamino)ethyl)-5-¹⁸F-fluoropicolinamide (¹⁸F-2) showed high in vivo stability and favorable pharmacokinetic properties: the fast clearance of ¹⁸F-2 from urinal system lead to almost background level of uptake for all major organs other than tumor at 2 h post injection. Taken together, ¹⁸F-2 might be ready for potential clinical evaluation of melanoma diagnosis.

¹⁸F-Labeled Glutamic Acid and Glutamine

Evaluation of: Ploessl K, Wang L, et al. Comparative Evaluation of ¹⁸F-Labeled Glutamic Acid and Glutamine as Tumor Metabolic Imaging Agents. J Nucl Med 2012, 53(10), 1616–1624.

Chiral center is widely present in bioactive ligands. Because harsh labeling condition may racemize the imaging agent, it is crucial to confirm that the chirality is maintained after labeling. Radio-labeled amino acids represent a large category of PET imaging agents for cancer research. Although some of the probes may not be incorporated into protein, their tumor uptake could reflect the elevated expression of amino acid transporters. Previously, ¹⁸F-(2S,4R)4-fluoroglutamine (¹⁸F-(2S,4R)4F-GLN) has been developed for detecting glutaminolytic tumors. ¹⁸F-labeled glutamic acid was also synthesized as a diastereomeric mixture of ¹⁸F-(2S,4S) and (2S,4R) glutamic acid (GLU) for tumor imaging. Because the tumor uptakes of ¹⁸F-labeled glutamic acid could be highly stereoselective, analysis of the obtained imaging result using these mixed agents could be complicated.

In this study, Ploessl and co-workers prepared and evaluated enantiomerically pure ¹⁸F-(2S,4R)4F-GLN and ¹⁸F-(2S,4R)4F-GLU in vitro and in vivo. The findings demonstrated that tumor cell uptake of ¹⁸F-(2S,4R)4F-GLN was higher than that of ¹⁸F-(2S,4R)4F-GLU in vitro. Without incorporation into protein, the uptake of ¹⁸F-(2S,4R)4F-GLU is predominantly controlled by the amino acid transporter. PET imaging studies in rats bearing 9L tumor xenographs demonstrated that ¹⁸F-(2S,4R)4F-GLN exhibited a higher tumor uptake and longer retention, whereas ¹⁸F-(2S,4R)4F-GLU demonstrated a slightly higher tumor to background contrast. As both PET probes are optically pure, the information obtained is conclusive.

A PET Probe for Imaging Transplanted Islets

Evaluation of: Wu Z, Liu S, et al. Development and Evaluation of ¹⁸F-TTCO-Cys⁴⁰-Exendin-4: A PET Probe for Imaging Transplanted Islets. J Nucl Med 2013, 54(2), 244–251.

For PET probe synthesis, specific activity is another important parameter to consider, especially when receptor number is limited (such as for brain imaging and transplanted islet imaging). When the bio-ligand has larger molecular weight than a few thousand Dalton, the structural changes caused by introducing a labeling tag become less significant. Therefore, complete separation of the radiolabeled probe from the unlabeled cold ligand could be challenging, and relatively low specific activity is usually expected.

In this study, Wu et al. used the ultra-efficient ¹⁸F-labeling method based on the tetrazine trans-cyclooctene (TTCO) ligation to construct an imaging agent targeting GLP-1R. The radiochemistry showed that 4 µg (0.84 nmol) tetrazine-Cys⁴⁰-exendin-4 loading was enough to obtain the PET probe in good yield with high specific activity (130 GBq/µmol). This probe showed high and specific binding to GLP-1R when evaluated in tumor models. Furthermore, it showed that ¹⁸F-TTCO-Cys⁴⁰-exendin-4 is capable of imaging intraportally transplanted islets, where the receptor number is very limited.

Acknowledgements

We would like to thank the NIBIB (R21 1 r21 eb012294-01a1), the American Cancer Society (121991-MRSG-12-034-01-CCE), SC CTSI (12-2176-3135), and USC Radiology Department for research support.

 

Highlight in Molecular Imaging Instrumentation

Quanzheng Li

Center for Advanced Radiological Sciences, Massachusetts General Hospital, Harvard Medical School, Boston, MA 02114, USA.

Correspondence: li.quanzheng@mgh.harvard.edu

The author sincerely thanks Drs. Joyita Dutta and Chuan Huang for their help on preparing the material for this summary.

Evaluation of:  Huang  C, Ackerman J,  Petibon Y, Brady T, El Fakhri G, J Ouyang. Real-time 3D motion tracking using MR micro-coils for PET imaging. J Nucl Med. 2013; 54 (Supplement 2):44

Motion induced image quality degradation is a major problem in brain positron emission tomography (PET) which could lead to imaging failures, especially for pediatric patients, the elderly, and patients with Alzheimer’s disease (AD). In the 2013 SNMMI annual meeting, researchers from Massachusetts General Hospital presented a motion correction technology based on Magnetic Resonance (MR) miniature coils to track the motion of the head during imaging in a simultaneous PET/MR scanner. The presented technology monitors the head motion using several MR coils, which are a few millimeters in diameter, attached to the head of a patient. The measured real-time 3D fields of motion are then incorporated into an iterative PET image reconstruction which produces crisp PET images without motion induced degradation even for continuous motion that could be encountered with AD patients. In their conference proceeding, the researchers presented phantom results with wired miniature MR coils, they are also developing wireless miniature coils which are said to be cheaper to manufacture and more patient friendly.

Evaluation of: Dan M, Gulani V, Seiberlich N, Liu K, Sunshine JL, Durek JL, Griswold MA. Magnetic Resonance Fingerprinting. Nature, 2013; 495: 187-192

One hurdle for the adoption of simultaneous PET/MR scans is the unmatched acquisition durations of clinical PET scans and MRI scans. MR is rich in contrast mechanisms; as a result, conventional MR protocol requires separate image acquisitions to obtain information for several important tissue properties and/or their combinations. The necessity of multiple image acquisitions leads to long acquisition time. For example, a common PET acquisition will take less than 10 minutes for one bed position; however, a single-bed cardiac MR scan could take more than 30 minutes. Magnetic Resonance Fingerprinting (MRF) provides an alternative to gather information of multiple tissue MR properties. Instead of isolating certain properties from others in each imaging acquisition, MRF utilizes randomized imaging parameter sets to create a unique signal history (thus the name fingerprinting) for each combination of tissue properties, such as proton density, T1, T2. Effectively, several properties can be simultaneously acquired with much less acquisition time. Although the clinical utility of MRF is yet to be shown, MRF is potentially a revolutionary technique, and could provide a solution to the problem of acquisition duration mismatch between PET and MRI scans.

 

This work is a proof of principle of simultaneous or multiplexed imaging of multiple high atomic number (Z) elements using X-ray ?uorescence computed tomography (XFCT). The experimental setup included a 5-mm-diameter pencil beam produced by a polychromatic X-ray source used to stimulate emission of XRF photons from gold, gadolinium, and barium embedded within a water phantom, which is translated and rotated relative to the stationary pencil beam. One important outcome of this study is the observation of a linear relationship between the XRF intensity of each tested element and their concentrations indicating the quantitative imaging capabilities of XFCT.

 

Evaluation of: Defrise, Michel, Ahmadreza Rezaei, and Johan Nuyts. "Time-of-flight PET data determine the attenuation sinogram up to a constant." Physics in Medicine and Biology 57, no. 4 (2012): 885.

Quantitatively accurate PET image reconstruction relies on the availability of a reliable attenuation map. This is a challenge for applications where a CT image of the subject is not available. The problem of estimating the attenuation sinogram directly from the PET emission data is therefore a critical one. While there have been many efforts in this directly, most of these methods lead to crosstalk artifacts in the attenuation and activity images. Defrise et al. 2012 was a major breakthrough in this area showing that when time-of-flight (TOF) information is available, the attenuation sinogram can be estimated from the emission data up to an additive constant and that its gradient can be estimated efficiently using a simple analytic algorithm. This was done by exploiting the consistency condition for the TOF Radon transform. Furthermore, this paper provided an analytical algorithm to solve this problem and performed an approximate analysis of noise propagation. This paper is awarded the Roberts Prize for the best paper published in Physics in Medicine and Biology in 2012.

 

Evaluation of: L. Cai and L. J. Meng, "Hybrid pixel-waveform CdTe/CZT detector for use in an ultrahigh resolution MRI compatible SPECT system," Nuclear Instruments & Methods in Physics Research Section a-Accelerators Spectrometers Detectors and Associated Equipment, vol. 702, pp. 101-103, Feb 21 2013. 

A research group at the University of Illinois at Urbana-Champaign (UIUC), led by Dr. Ling-Jian Meng, has developed one of the first MR-compatible SPECT (MRC-SPECT) in the world. The UIUC system could provide an unprecedented SPECT imaging resolution, while operated inside MR scanners for simultaneous ultrahigh resolution dual-modality imaging studies. The key to the system is a state-of-art room-temperature semiconductor imaging detector platform, which offers an ultrahigh spatial resolution for gamma ray imaging and is proven to be fully compatible with MR environment. The current MRC-SPECT system is constructed using 40 CdTe detector modules, assembled in a compact fully stationary ring SPECT geometry. The group has experimentally demonstrated that this system is capable of providing an imaging resolution of <500 μm, when operated inside MR scanners. This development opens up many interesting opportunities for molecular imaging, by allowing simultaneous acquisition of naturally coregistered ultrahigh resolution nuclear and MR images.

 

New Molecular Imaging Techniques

Kinetic modeling of glucose metabolism

Evaluation of: Wong K-P, Zhang X, Huang S-C. Improved Derivation of Input Function in Dynamic Mouse [¹⁸F]FDG PET Using Bladder Radioactivity Kinetics. Mol Imaging Biol. 15(4), 486–496 (2013).

An essential step in kinetic modeling of glucose metabolism is the generation of an accurate input function (IF). The tracer concentration in arterial plasma must be measured frequently by means of an invasive and time-consuming blood sampling. This procedure is challenging, especially in rodent models due to the limited blood volume available and the size of the blood vessels. In this work, the authors reported a hybrid modeling approach to estimate the plasma IF in mice using the activity in bladder along with a single blood sample at the end of the scan with [¹⁸F]FDG-PET. During the dynamic scan that was performed on 9 mice, blood samples were drawn from the femoral artery. A renal compartment model and a single exponential function were used to fit the activity in the bladder and the blood sample drawn at the end of the acquisition. The liver was also used as an alternative reference for the estimation of input function. Patlak modeling was performed both with arterial blood samples and the bladder derived Ifs showing that the area under the bladder derived IF was not significantly different from that obtained by serial blood sampling. Displayed below, the kinetic parameters for brain, myocardium, muscle, and liver obtained from both estimated and calculated IFs were in great agreement (R² > 0.983). However, the authors did not recommend the method for the liver because the IF results derived from the liver were not reliable.  

The modeling method proposed in this study is practical and novel. Of note, the bladder must always be in the field of view for the implementation of this method. If proven successfully by follow-up studies, it can eliminate the need for serial blood sampling in small animals during image acquisition. Certainly, this is of significance to preclinical imaging given the difficulties of blood sampling from mice during imaging acquisition, which makes the blood input modeling virtually impossible due to the lack of arterial blood input and unreliable image derived input functions (e.g. small blood pool and partial volume effect). However, the paper lacks some important technical details regarding the modeling package for Patlak modeling and the analytical tool for solving the differential equations. The amount of mathematics involved in the modeling method appears overwhelming, but there should be no problems to have it simplified in a code or graphical user interface.

Saleh Ramezani¹ & Xiankai Sun^*1,2

 ¹Department of Radiology and ²Advanced Imaging Research Center, University of Texas Southwestern Medical Center, Dallas, Texas 75390-8542, USA
Author for correspondence: xiankai.sun@utsouthwestern.e

New Preclinical Imaging Technique: Cryosection Labeling and Intravital Microscopy (CLIM)

Evaluation of: Ritsma L, Vrisekoop N and van Rheenen: In vivo imaging and histochemistry are combined in the cryosection labelling and intravital microscopy technique. Nat Commun, 2013, 4, 2366.

The ability to obtain histological analysis of a specific region of interest in vivo would provide exceptional information about diseases and potential therapies. One of the common techniques to obtain high resolution information in vivo is the utilization of intravital microscopy (IVM), where surgically implanted glass windows can be used to observe real-time cellular events using optical imaging. However, one major drawback is the need to have transgenic mice/rats or specific fluorescent probes. Through immunohistochemistry (IHC) and optical imaging (e.g. confocal microscopy), high resolution ex vivo tissue information can be obtained thanks to the availability of varieties of antibodies. In this work, Ritsma and coworkers created a technique to view a specific region of interest (ROI), where they can observe a physiological process at the cellular level in vivo, followed by IHC analysis of that exact same region. The technique combines cryosection labeling and intravital microscopy (CLIM) on the basis that the creation of an autofluorescent mark using a multiphoton laser, “photo-tattoo”, can later be re-identified by a fluorescent microscope after the tissue is cryosectioned and attached to glass slides. The authors were able to show that a variety of tissues could be photo-tattooed, such as tumor xenografts, liver, pancreas, and spleen. Although the autofluorescent compounds for photo-tattoo were not identified, they were believed to be present in the blood, more specifically in the serum. As such, highly vascularized tissues with stalled vessels can be more efficiently photo-tattooed than those with flowing blood, which would wash out the marked molecules. Given that the primary/secondary imaging procedures as well as the visualization of photo-tattooed tissues can be performed on the same optical imaging device,  employing CLIM is a rather straightforward. To demonstrate the procedure of CLIM stepwise, the authors used IVM to observe a mammary tumor in mice and then create a photo-tattooed square around the region of interest. The mammary tumor was removed, placed into a tissue mould, frozen, and cyrosectioned into 16 μm slices. To account for tissue shrinkage and alterations to the tumor shape, multiple slices were analyzed for the tattooed section. When all sections were accounted for, the marked region was reconstructed and compared to the IVM derived image. Utilizing this technique, they were also able to observe breast cancer tumor cell migration in a mouse model in real-time with IVM, in which IHC revealed that CD3⁺ T cells were present. The determination of T cell subset was conducted using mice depleted of either CD4⁺ or CD8⁺ T cells, whose migration was found to be minimal.

This reported technique has the potential to assist researchers at obtaining a better understanding about the microenvironment and cellular composition of diseases in animal models. As performed in this work, specific cells can be identified with relevancy to tumor metastasis, which can be targeted for therapy. The photo-tattooed areas observable in pre and post tissue removal provide the landmarks required to correlate the IHC information with IVM. Potentially, therapeutic efficacies can be determined through the treatment and observation of tumor/diseased area before and after treatment. To monitor the efficacy and distribution of peptide/small molecule therapies, IVM potentially could observe a change in tissues of target. As necessary, secondary analysis could be performed using a mass spectrometry technique, such as inkjet printed MALDI-MS to directly detect the molecules of interest in the histology samples. Despite its success, this technique is perhaps limited to the examination of small regions. With the methodology described, tattooing a micro-region of interest takes up to 5 min, while the tissue excision must be done within 10 min to minimize the rapid infiltration of immune cells. This limits the dimensions that can be interrogated.

William Silvers¹ & Xiankai Sun^*1,2

¹Department of Radiology and ²Advanced Imaging Research Center, University of Texas Southwestern Medical Center, Dallas, Texas 75390-8542, USA
Author for correspondence: xiankai.sun@utsouthwestern.edu

 

Conjugation Chemistry for Radioimmunoimaging and Radioimmunotherapy

Evaluation of: Price, E. W.; Zeglis, B. M.; Cawthray, J. F.; Ramogida, C. F.; Ramos, N.; Lewis, J. S.; Adam, M. J.; Orvig, C. “H₄octapa-trastuzumab: Versatile Acyclic Chelate System for ¹¹¹In and ¹⁷⁷Lu Imaging and Therapy” J. Am. Chem. Soc. 2013, 135, 12707-12721.

In recent years there is a trend in clinical practice to use different radioisotopes with the same targeting vector for pre-therapy imaging and dosimetry studies. As such, a tracer labeled with a specific radionuclide can be used for scouting scans before the radiotherapy or for follow up scans, while the same agent bearing a different radioisotope can be administered for radiotherapy. The use of antibodies for such applications is gaining momentum since ImmunoPET has successfully implemented in both preclinical and clinical research. Although antibodies are extremely stable in vivo, they can be easily damaged when subjected to harsh labeling conditions. Therefore, there is a need for the development of new chelating ligand system that enables efficient radiolabeling under mild conditions. In this study, Price et al, reported a p-SCN-Bn-H₄octapa bifunctional chelator that can label trastuzumab, a HER2/neu-targeted antibody, with both ¹¹¹In and ¹⁷⁷Lu under mild conditions.

For the synthesis of the p-SCN-Bn-H₄octapa bifunctional chelator, the authors developed a new synthetic route, incorporating the 2-nitrobenzenesulfonamide protecting group. The synthesis of the chelator was accomplished in 7 steps with an overall yield of 25 – 30%. Thermodynamic stability calculations using both potentiometric titration experiments and molecular modeling revealed that [Lu(octapa)]^- and [In(octapa)]^- complexes have similar logK_ML and pM values. They lack fluxional isomerization in solution and show comparable structural properties. These results suggest that the two complexes would have similar biological behavior and can be used for SPECT imaging (¹¹¹In) along with ¹⁷⁷Lu based radiotherapy. The evaluation of p-SCN-Bn-H₄octapa was performed as compared to p-SCN-Bn-DOTA. In the case of p-SCN-Bn-H₄octapa, 3.03 ± 0.1 chelators were conjugated per antibody while for p-SCN-Bn-DOTA it was 3.40 ± 0.1. Radiolabelling of the antibody conjugates of p-SCN-Bn-H₄octapa was achieved in >95% radiochemical yield and >99% radiochemical purity. The specific activity was 4.0 ± 0.3 and 3.4 ± 0.4 mCi/mg for ¹¹¹In and ¹⁷⁷Lu, respectively. These results demonstrated a significant improvement in radiochemistry as compared to the DOTA counterparts; and a higher immunoreactivity was observed for the octapa-trastuzumab conjugates. In vivo performance and pharmacokinetic studies of the above synthesized conjugates were performed using nude female mice bearing SKOV-3 ovarian cancer xenografts. The biodistribution studies indicated high tumor to background ratios for all conjugates, while the tumor uptake with the ¹¹¹In-octapa and ¹⁷⁷Lu-octapa were significantly higher than their DOTA counterparts. Impressively, the SPECT/CT images displayed high tumor contrast for all four conjugates in the SKOV-3 tumor-bearing mouse model out to five days.

In summary, the reported synthetic route not only made feasible the synthesis of p-SCN-Bn-H₄octapa bifunctional chelator system but also reduced the burden of purification. The new chelator system was proven to be able to efficiently label antibodies with radioisotopes without sacrificing the immunoreactivity desired for radioimmunoimaging and/or radioimmunotherapy.  

Marianna Dakanali¹ & Xiankai Sun^*1,2

 ¹Department of Radiology and ²Advanced Imaging Research Center, University of Texas Southwestern Medical Center, Dallas, Texas 75390-8542, USA
Author for correspondence: xiankai.sun@utsouthwestern.edu

     

Aug/27/2013