Exact detection and complete removal of cancer is a key point to minimize cancer recurrence. However, it is
currently very difficult to detect small tumors inside human body and continuously monitor tumors using a non-invasive
imaging modality. Presently, positron emission tomography (PET) can provide the most sensitive cancer images in the
human body. However, PET imaging has very limited imaging time because they typically use isotopes with short halflives.
PET imaging cannot also visualize anatomical information. Magnetic resonance imaging (MRI) can provide highresolution
images inside the body but it has a low sensitivity, so MRI contrast agents are necessary to enhance the
contrast of tumor. Near infrared fluorescent (NIRF) imaging has a good sensitivity to visualize tumor using optical
probes, but it has a very limited tissue penetration depth. Therefore, we developed multi-modality nanoparticles for MRI
based diagnosis and NIRF imaging based surgery of cancer. We utilized glycol chitosan of 350 nm as a vehicle for MRI
contrast agents and NIRF probes. The glycol chitosan nanoparticles were conjugated with NIRF dye, Cy5.5 and bladder
cancer targeting peptides to increase the internalization of cancer. For MR contrast effects, iron oxide based 22 nm nanocubes
were physically loaded into the glycol chitosan nanoparticles. The nanoparticles were characterized and evaluated
in bladder tumor bearing mice. Our study suggests the potential of our nanoparticles by both MRI and NIRF imaging for
tumor diagnosis and real-time NIRF image-guided tumor surgery.
Enhanced permeability and retention (EPR) effects for tumor treatment have been utilized as a representative strategy to
accumulate untargeted nanoparticles in the blood vessels around tumors. However, the EPR effect itself was not
sufficient for the nanoparticles to penetrate into cancer cells. For the improvement of diagnosis and treatment of cancer
using nanoparticles, many more nanoparticles need to specifically enter cancer cells. Otherwise, can leave the tumor
area and not contribute to treatment. In order to enhance the internalization process, specific ligands on nanoparticles
can help their specific internalization in cancer cells by receptor-mediated endocytosis. We previously developed glycol
chitosan based nanoparticles that suggested a promising possibility for in vivo tumor imaging using the EPR effect. The
glycol chitosan nanoparticles showed a long circulation time beyond 1 day and they were accumulated predominantly in
tumor. In this study, we evaluated two peptides for specific targeting and better internalization into urinary bladder
cancer cells. We conjugated the peptides on to the glycol chitosan nanoparticles; the peptide-conjugated nanoparticles
were also labeling with near infrared fluorescent (NIRF) dye, Cy5.5, to visualize them by optical imaging in vivo.
Importantly real-time NIRF imaging can also be used for fluorescence (NIRF)-guided surgery of tumors beyond normal
optical penetration depths. The peptide conjugated glycol chitosan nanoparticles were characterized with respect to size,
stability and zeta-potential and compared with previous nanoparticles without ligands in terms of their internalization
into bladder cancer cells. This study demonstrated the possibility of our nanoparticles for tumor imaging and
emphasized the importance of specific targeting peptides.
Magnetic resonance imaging (MRI) is one of the best imaging modalities for noninvasive cancer detection but MRI does
not have enough sensitivity to delineate tumor margins during surgery. Moreover, since most surgical tools contain
metal substances, image-guided surgery is hard to perform with a MR machine using magnets. Also, MR imaging is too
slow for real-time guided-surgery. On the other hand, near infrared fluorescence (NIRF) imaging has recently received
great interest for in vivo imaging due to its high signal-to-noise ratios and short image-acquisition times. NIRF imaging
can be used to delineate tumor margins during surgery, but current NIRF imaging cannot provide the penetration depth
to detect early-stage cancer inside body. Thus, we have developed dual-modality in vivo imaging for MRI detection of
tumors and NIRF-guided surgery using multi-component nanoparticles. NIRF dye (cyanine 5.5, Cy5.5), conjugated
glycol chitosan nanoparticles (HGC) exhibited excellent tumor targeting ability with NIRF imaging. Superparamagnetic
iron oxide (SPIO) nanoparticles as a MR contrast agent were loaded into the nanoparticles, resulting in SPIO-HGC-Cy5.5
nanoparticles. SPIO-HGC-Cy5.5 nanoparticles were characterized and evaluated in mice by both NIRF and MR
imaging. Our results indicate SPIO-HGC-Cy5.5 nanoparticles have the potential for dual-modality in vivo imaging with
MRI detection of tumors and NIRF-guided surgery.
Emily Haglund, Mary-Margaret Seale-Goldsmith, Deepika Dhawan, Jane Stewart, Jose Ramos-Vara, Christy Cooper, Lisa Reece, Timothy Husk, Donald Bergstrom, Deborah Knapp, James Leary
Nanomedical approaches to diseases such as cancer provide great promise with respect to diagnostic and therapeutic
applications. The impact of nanomedicine versus conventional therapies will be realized with regard to their specific cell
targeting capabilities. Semiconductor nanoparticles have distinct advantages due to their chemical conjugation and
detection characteristics. The attachment of a peptide sequence, LTVSPWY, was completed. These nanoparticles
successfully targeted in vitro and in vivo systems. This technology can be utilized as a base mechanism for the
construction of a multifunctional nanomedical system. Nanomedicine has great potential for impacting the treatment of
specific diseases and healthcare delivery methods.
Approximately 12,000 people are diagnosed with invasive transitional cell carcinoma of the urinary bladder (InvTCC) each year in the United States. Surgical removal of the bladder (cystectomy) and regional lymph node dissection are considered frontline therapy. Cystectomy causes extensive acute morbidity, and 50% of patients with InvTCC have occult metastases at the time of diagnosis. Better staging procedures for InvTCC are greatly needed. This study was performed to evaluate an intra-operative near infrared fluorescence imaging (NIRF) system (Frangioni laboratory) for identifying sentinel lymph nodes draining InvTCC. NIRF imaging was used to map lymph node drainage from specific quadrants of the urinary bladder in normal dogs and pigs, and to map lymph node drainage from naturally-occurring InvTCC in pet dogs where the disease closely mimics the human condition. Briefly, during surgery NIR fluorophores (human serum albumen-fluorophore complex, or quantum dots) were injected directly into the bladder wall, and fluorescence observed in lymphatics and regional nodes. Conditions studied to optimize the procedure including: type of fluorophore, depth of injection, volume of fluorophore injected, and degree of bladder distention at the time of injection. Optimal imaging occurred with very superficial injection of the fluorophore in the serosal surface of the moderately distended bladder. Considerable variability was noted from dog to dog in the pattern of lymph node drainage. NIR fluorescence was noted in lymph nodes with metastases in dogs with InvTCC. In conclusion, intra-operative NIRF imaging is a promising approach to improve sentinel lymph node mapping in invasive urinary bladder cancer.
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