Synthesis of 68Ga-Labeled Biopolymer-based Nanoparticle Imaging Agents for Positron-emission Tomography.

AIM: The purpose of this study was to develop a folate receptor-targeted 68Ga-labeled agent for the detection of cancer cells in mouse models of ovarian cancer by dual positron-emission tomography (PET) and magnetic resonance imaging (MRI). Moreover, we aimed to develop a controlled biopolymer-based chemistry that enables linking metal-binding (here Ga-68) chelators. MATERIALS AND METHODS: The nanoparticle (NP) agent was created by self-assembling of folic acid-modified polyglutamic acid and chelator-modified chitosan followed by radiolabeling with 68Ga (III) ions (68Ga-NODAGA-FA). The structure of modified biopolymers was characterized by spectroscopy. Particle size and mobility were determined. RESULTS: Significant selective binding of NPs was established in vitro using folate receptor-positive KB and - negative MDA-MB-231 cell lines. In vivo tumor uptake of folate-targeted 68Ga3+-radiolabeled NPs was tested using subcutaneous tumor-bearing CB17 SCID mice models. PET/MR dual modalities showed high tumor uptake with 6.5 tumor-to-muscle ratio and NP localization. CONCLUSION: In vivo results supporting the preliminary in vitro tests demonstrated considerably higher 68Ga-NODAGA-FA nanoparticle accumulation in KB tumors than in MDA-MB-231 tumors, thereby confirming the folate receptor-mediated uptake of this novel potential PET imaging agent.

Folic acid-conjugated fluorescent polymer for up-regulation folate receptor expression study via targeted imaging of tumor cells.

Thoroughly investigation of folate receptor (FR) expression related to targeting drug delivery in tumor cells has been intensively pursued in recent years. Herein, a simple and versatile strategy for determination of FR expression based on targeted imaging of tumor cells with fluorescent nano-conjugates was developed. The fluorescent nano-conjugates were composed of poly 2-vinyl-4,4-dimethyl azlactone (PVDMA) as the linker, folic acid as the targeting unit and amino-Rhodamine B as the fluorescent ligand. Owing to possessing dimethyl azlactone groups in polymer framework, PVDMA could easily reacted with amines or alcohols, and form water soluble materials. Fluorescent imaging studies indicated that the prepared nano-conjugates could specifically target tumor cells and monitor the over expressing of FR. Moreover, the FR expression up-regulation in HeLa cells through medicines regulation has been further explored. This new protocol opens an effective way through synthesis and design of novel fluorescent nano-conjugates for FR expression investigation in tumor cells via targeted imaging, showing great potential in drug delivery mechanism study and cancer therapy.

A novel DAD type and folic acid conjugated fluorescent monomer as a targeting probe for imaging of folate receptor overexpressed cells.

We describe a modification and post-functionalization technique for a donor-acceptor-donor type monomer; 6-(4,7-bis(2,3-dihydrothieno[3,4-b][1,4]dioxin-5-yl)-2H-benzo[d][1,2, 3]triazol-2-yl)hexan-1-amine. Folic acid was attached to the fluorescent structure. The conjugation was confirmed via NMR and Fourier transform infrared analyses. Cytotoxicity was investigated and the comparison of association of targeted monomeric structures in tumor cells was monitored via fluorescence microscopy.

Quantitation of rare circulating tumor cells by folate receptor α ligand-targeted PCR in bladder transitional cell carcinoma and its potential diagnostic significance.

Numerous attempts for detection of circulating tumor cells (CTC) have been made to develop reliable assays for early diagnosis of cancers. In this study, we validated the application of folate receptor α (FRα) as the tumor marker to detect CTC through tumor-specific ligand PCR (LT-PCR) and assessed its utility for diagnosis of bladder transitional cell carcinoma (TCC). Immunohistochemistry for FRα was performed on ten bladder TCC tissues. Enzyme-linked immunosorbent assay (ELISA) for FRα was performed on both urine and serum specimens from bladder TCC patients (n = 64 and n = 20, respectively) and healthy volunteers (n = 20 and n = 23, respectively). Western blot analysis and qRT-PCR were performed to confirm the expression of FRα in bladder TCC cells. CTC values in 3-mL peripheral blood were measured in 57 bladder TCC patients, 48 healthy volunteers, and 15 subjects with benign urologic pathologies by the folate receptor α ligand-targeted PCR. We found that FRα protein was overexpressed in both bladder TCC cells and tissues. The levels of FRα mRNA were also much higher in bladder cancer cell lines 5637 and SW780 than those of leukocyte. Values of FRα were higher in both serum and urine specimens of bladder TCC patients than those of control. CTC values were also higher in 3-mL peripheral blood of bladder TCC patients than those of control (median 26.5 Cu/3 mL vs 14.0 Cu/3 mL). Area under the receiver operating characteristic (ROC) curve for bladder TCC detection was 0.819, 95 % CI (0.738-0.883). At the cutoff value of 15.43 Cu/3 mL, the sensitivity and the specificity for detecting bladder cancer are 82.14 and 61.9 %, respectively. We concluded that quantitation of CTCs through FRα ligand-PCR could be a promising method for noninvasive diagnosis of bladder TCC.

A folate receptor electrochemical sensor based on terminal protection and supersandwich DNAzyme amplification.

Based on the protecting effect of folate receptor (FR) toward folic acid (FA) modified DNA and the signal amplification of supersandwich DNA structure, we designed an interesting electrochemical biosensor for FR. In the present system, with the increase of FR, protecting more FA bound DNA from hydrolysis by exonuclease I (Exo I), FA bound DNA will hybridize to form more supersandwich DNA structure resulting in an increased electrochemical signal. A relationship between the concentration of the target protein, FR, and the obtained electrochemical signal can be established. The signal was obtained by the catalysis on H2O2 in the system containing Fc and hemin/DNAzyme. The detection concentration range of FR was from 1.0 to 20.0 ng/mL with an achieved detection limit of 0.3 ng/mL which approached clinically relevant concentrations of FR.

Effective peroxidase-like activity of a water-solubilized Fe-aminoclay for use in immunoassay.

In this study, we developed a colorimetric sensor for the determination of the peroxidase-like activity of Fe-aminoclay, which was used as a novel way of immunoassay for lung cancer was examined. Fe-aminoclay was synthesized by a facile sol-gel reaction under ambient conditions, with both amino sites and Fe surface exposed outside. This Fe-aminoclay, which exhibits strong peroxidase-like activity particularly over a wide pH range, was explored as a robust and rugged replacement of peroxidase enzymes. The peroxidase-like activity of Fe-aminoclay was proved by means of the production of a blue-colored product by 3,3',5,5'-tetramethylbenzidine (TMB) in the presence of hydrogen peroxide (H2O2). This unique activity, in a folic acid-conjugated form, was evaluated for its ability to specifically detect folate receptor-positive A549 cell, which was compared to a human lung normal cell line with lack of the expression of the folate receptor. The chromatic response, which was even detectable by naked eyes, displayed gradational variation, proportional to the number of cells (up to 20,000 cells).