Mingwei Wang

A rapid pathway toward a superb gene delivery system: programming structural and functional diversity into a supramolecular nanoparticle library.

Nanoparticles are regarded as promising transfection reagents for effective and safe delivery of nucleic acids into a specific type of cells or tissues providing an alternative manipulation/therapy strategy to viral gene delivery. However, the current process of searching novel delivery materials is limited due to conventional low-throughput and time-consuming multistep synthetic approaches. Additionally, conventional approaches are frequently accompanied with unpredictability and continual optimization refinements, impeding flexible generation of material diversity creating a major obstacle to achieving high transfection performance. Here we have demonstrated a rapid developmental pathway toward highly efficient gene delivery systems by leveraging the powers of a supramolecular synthetic approach and a custom-designed digital microreactor. Using the digital microreactor, broad structural/functional diversity can be programmed into a library of DNA-encapsulated supramolecular nanoparticles (DNA⊂SNPs) by systematically altering the mixing ratios of molecular building blocks and a DNA plasmid. In vitro transfection studies with DNA⊂SNPs library identified the DNA⊂SNPs with the highest gene transfection efficiency, which can be attributed to cooperative effects of structures and surface chemistry of DNA⊂SNPs. We envision such a rapid developmental pathway can be adopted for generating nanoparticle-based vectors for delivery of a variety of loads.

Covalent Functionalization of Graphene Oxide with Biocompatible Poly(ethylene glycol) for Delivery of Paclitaxel

Graphene oxide (GO), a novel 2D nanomaterial prepared by the oxidation of natural graphite, has been paid much attention in the area of drug delivery due to good biocompatibility and low toxicity. In the present work, 6-armed poly(ethylene glycol) was covalently introduced into the surface of GO sheets via a facile amidation process under mild conditions, making the modified GO, GO-PEG (PEG: 65 wt %, size: 50-200 nm), stable and biocompatible in physiological solution. This nanosized GO-PEG was found to be nontoxic to human lung cancer A549 and human breast cancer MCF-7 cells via cell viability assay. Furthermore, paclitaxel (PTX), a widely used cancer chemotherapy drug, was conjugated onto GO-PEG via π-π stacking and hydrophobic interactions to afford a nanocomplex of GO-PEG/PTX with a relatively high loading capacity for PTX (11.2 wt %). This complex could quickly enter into A549 and MCF-7 cells evidenced by inverted fluorescence microscopy using Fluorescein isothiocyanate as a probe, and it also showed remarkably high cytotoxicity to A549 and MCF-7 cells in a broad range of concentration of PTX and time compared to free PTX. This kind of nanoscale drug delivery system on the basis of PEGylated GO may find potential application in biomedicine.

Delivery of Paclitaxel Using PEGylated Graphene Oxide as a Nanocarrier

Paclitaxel (PTX) is an extensively used potent chemotherapy drug; however, low water solubility, poor bioavailability, and emergence of drug resistance in patients limited its biological application. In this report, we proposed a new drug delivery system for cancer therapy based on graphene oxide (GO), a novel 2D nanomaterial obtained from the oxidation of natural graphite, to improve the utilization rate of PTX. PTX was first connected to biocompatible 6-armed poly(ethylene glycol), followed by covalent introduction into the surface of GO sheets via a facile amidation process under mild conditions, affording the drug delivery system, GO-PEG-PTX (size 50-200 nm). GO-PEG nanosized carrier could quickly enter into human lung cancer A549 and human breast cancer MCF-7 cells verified by inverted fluorescence microscope using fluorescein isothiocyanate as probe. This nanocarrier was nontoxic to A549 and MCF-7 cells without linking with PTX. Nevertheless, GO-PEG-PTX showed remarkably high cytotoxicity to A549 and MCF-7 cells in a broad range of concentration of PTX and time compared to free PTX. This kind of nanoscale drug delivery system based on PEGylated GO may find widespread application in biomedicine.

Microfluidic-based 18F-labeling of biomolecules for immuno-positron emission tomography.

Methods for tagging biomolecules with fluorine 18 as immuno–positron emission tomography (immunoPET) tracers require tedious optimization of radiolabeling conditions and can consume large amounts of scarce biomolecules. We describe an improved method using a digital microfluidic droplet generation (DMDG) chip, which provides computer-controlled metering and mixing of 18F tag, biomolecule, and buffer in defined ratios, allowing rapid scouting of reaction conditions in nanoliter volumes. The identified optimized conditions were then translated to bench-scale 18F labeling of a cancer-specific engineered antibody fragments, enabling microPET imaging of tumors in xenografted mice at 0.5 to 4 hours postinjection.

Microfluidics for Positron Emission Tomography Probe Development

Owing to increased needs for positron emission tomography (PET), high demands for a wide variety of radiolabeled compounds will have to be met by exploiting novel radiochemistry and engineering technologies to improve the production and development of PET probes. The application of microfluidic reactors to perform radiosyntheses is currently attracting a great deal of interest because of their potential to deliver many advantages over conventional labeling systems. Microfluidics-based radiochemistry can lead to the use of smaller quantities of precursors, accelerated reaction rates, and easier purification processes with greater yield and higher specific activity of desired probes. Several proof-of-principle examples along with the basics of device architecture and operation and the potential limitations of each design are discussed. Along with the concept of radioisotope distribution from centralized cyclotron facilities to individual imaging centers and laboratories (“decentralized model”), an easy-to-use, stand-alone, flexible, fully automated, radiochemical microfluidic platform can provide simpler and more cost-effective procedures for molecular imaging using PET.

Can Fluorine-18 Fluoroestradiol Positron Emission Tomography–Computed Tomography Demonstrate the Heterogeneity of Breast Cancer In Vivo?

AIM Our study was to investigate the heterogeneity of estrogen receptor (ER) expression among tumor sites by using fluorine-18 ((18)F) fluoroestradiol (FES) positron-emission tomography-computed tomography (PET-CT) imaging. METHODS Thirty-two breast cancer patients underwent both (18)F-FES and (18)F fluorodeoxyglucose (FDG) PET-CTs from June 2010 to December 2011 in our center (mean age, 53 years; range, 27-77 years). We used the maximum standardized uptake value to quantify ER expression and a cutoff value of 1.5 to dichotomize results into ER(+) and ER(-). The difference of heterogeneity between the initial patients and patients with recurrent or metastatic disease after treatments was assessed by using the χ(2) test. Also, the (18)F-FES uptake was compared with the (18)F-FDG uptake by use of Spearman correlation coefficients. RESULTS A total number of 237 lesions in 32 patients were detected. Among them, most lesions (64.1% [152/237]) were bone metastasis. A striking 33.4-fold difference in (18)F-FES uptake was observed among different patients (maximum standardized uptake value range, 0.5 to approximately 16.7), and a 8.2-fold difference was observed among lesions within the same individual (1.0 to approximately 8.2). As for (18)F-FDG uptake, the difference was 11.6-fold (1.3 to approximately 15.1) and 9.9-fold (1.4 to approximately 13.8), respectively. In 28.1% (9/32) of the patients, both (18)F-FES(+) and (18)F-FES(-) metastases were present, which suggests partial discordant ER expression. After treatments, 37.5% (9/24) patients with recurrent or metastatic breast cancer showed heterogeneity, whereas no untreated patient was detected to exist discordant ER expression (χ(2), 4.174; P < .05). In addition, the (18)F-FES uptake showed a weak correlation with the (18)F-FDG uptake (ρ = 0.248; P < .05). CONCLUSION (18)F-FES and (18)F-FDG uptake varied greatly both within and among patients. (18)F-FES PET-CT demonstrated a conspicuous number of patients with the heterogeneity of ER expression.

Prevalence and risk of cancer of thyroid incidentaloma identified by fluorine-18 fluorodeoxyglucose positron emission tomography/computed tomography.

OBJECTIVE To investigate the prevalence and risk of thyroid incidentaloma identified by positron emission tomography/computed tomography (PET/CT). STUDY DESIGN Historical cohort study. SETTING Fudan University Shanghai Cancer Center. METHODS A total of 15 948 non-thyroid disease patients who underwent fluorine-18 fluorodeoxyglucose (18F-FDG) PET/CT from November 2006 to September 2010 were included. They were divided into two groups: 12 080 patients for metastatic evaluation and 3868 patients for cancer screening. When thyroid incidentaloma was found, further diagnostic examination was conducted. MAIN OUTCOME MEASURES Prevalence and risk of thyroid incidentaloma. RESULTS The prevalence of incidental thyroid 18F-FDG uptake was approximately 2.5% (395 of 15 948). The prevalence of incidentaloma in healthy subjects (118 of 3868; 3.1%) was statistically higher than that in patients with suspected or known cancer (277 of 12 080; 2.3%) (p < .05). Among 395 incidentalomas, 146 patients had further examinations (53 patients with histologic confirmations, 93 patients with clinical monitoring). Finally, 43 lesions were confirmed to be malignancies. Therefore, the cancer risk was 29.5% (43 of 146), and it was higher in cancer screening patients (24 of 59; 40.7%) than in alleged cancer patients (19 of 87; 21.8%) (p < .05). As for FDG uptake pattern, the prevalence of thyroid cancer was 11.6% (5 of 43) and 36.9% (38 of 103) in the group of patients with diffuse and focal uptake, respectively (p < .05). After logistic regression analysis, age, sex, maximal standardized uptake value, and calcification were the potent predictors of differentiation. CONCLUSION The presence of focal uptake with high SUVmax and calcification detected on CT images correlates with a high likelihood of thyroid malignancy. When a focal thyroid incidentaloma is detected, further examination should be performed.

MR/SPECT Imaging Guided Photothermal Therapy of Tumor-Targeting Fe@Fe3O4 Nanoparticles in Vivo with Low Mononuclear Phagocyte Uptake

The (125)I-c(RGDyK) peptide PEGylated Fe@Fe3O4 nanoparticles ((125)I-RGD-PEG-MNPs) with the average hydrodynamic diameter of ∼40 nm as a novel multifunctional platform were developed for tumor-targeting MR/SPECT imaging guided photothermal therapy in vivo. On the αvβ3-positive U87MG glioblastoma xenograft model, the signals of tumor from T2-weighted MR and SPECT imaging were much higher than those in the blocking group at 6 h post injection (p.i.) of RGD-PEG-MNPs and (125)I-RGD-PEG-MNPs intravenously, respectively. The pharmacokinetics and biodistribution were analyzed quantitatively by gamma counter ex vivo. The fact suggested that RGD-PEG-MNPs exhibited excellent targeting property and low mononuclear phagocyte uptake. At 6 h p.i. for (125)I-RGD-PEG-MNPs, the maximum uptake of 6.75 ± 1.24% of the percentage injected dose per gram (ID/g) was accumulated in the tumor. At 48 h p.i., only 1.11 ± 0.21% and 0.16 ± 0.09% ID/g were accumulated in the liver and spleen, respectively. With the guidance of MR/SPECT imaging, the multifunctional nanoparticles achieved a good photothermal therapeutic efficacy in vivo.

The Preliminary Study of 16α-[18F]fluoroestradiol PET/CT in Assisting the Individualized Treatment Decisions of Breast Cancer Patients

Objective To evaluate the clinical value of 16α-[18F]fluoroestradiol (18F-FES) PET/CT in assisting the individualized treatment decisions of breast cancer patients. Methods Thirty-three breast cancer patients, who underwent both 18F-FES and 18F-FDG PET/CT from July 2010 to March 2013 in our center, were enrolled in this preliminary study. All the patients used 18F-FES PET/CT as a diagnostic tool with a clinical dilemma. We used the maximum Standardized Uptake Value (SUVmax) to quantify ER expression and a cutoff value of 1.5 to dichotomize results into ER positive and negative lesions. All patients were clinically followed up at least 6 months. Results In evaluating equivocal lesions on conventional work-up group (n = 4), three lung lesions and another iliac lesion were enrolled. As for three lung lesions, 18F-FES PET/CT showed one lesion with high uptake, which suggested it was an ER positive metastasis. The other two lesions were 18F-FES negative, which meant an ER negative metastasis or secondary primary tumor. Additionally, one iliac lesion was detected by MRI. 18F-FDG uptake was high at the suspected lesion, whereas 18F-FES uptake was absent; In predicting origin of metastasis group (n = 2), two breast cancer patients had secondary primary tumors were collected. They were 18F-FES negative, which showed low possibility of metastasis from breast cancer and they were all confirmed by biopsy. In detecting ER status in metastasis group (n = 27), 18F-FES PET/CT showed increased 18F-FES uptake in all metastatic lesions in 11 patients; absent in all lesions in 13 patients; and the remaining 3 patients had both 18F-FES positive and negative lesions. Totally, on the basis of the 18F-FES PET/CT results, we found changes in the treatment plans in 16 patients (48.5%, 16/33). Conclusions 18F-FES PET/CT could assess the entire tumor volume receptor status; therefore, it may be used to assist the individualized treatment decisions of breast cancer patients.

Microwave-assisted One-pot Synthesis of N-succinimidyl-4-[18F]fluorobenzoate ([18F]SFB)

Biomolecules, including peptides¹⁻⁹, proteins¹⁰⁻¹¹, and antibodies and their engineered fragments¹²⁻¹⁴, are gaining importance as both potential therapeutics and molecular imaging agents. Notably, when labeled with positron-emitting radioisotopes (e.g., Cu-64, Ga-68, or F-18), they can be used as probes for targeted imaging of many physiological and pathological processes.¹⁵⁻¹⁸ Therefore, significant effort has devoted to the synthesis and exploration of ¹⁸F-labeled biomolecules. Although there are elegant examples of the direct ¹⁸F-labeling of peptides,¹⁹⁻²² the harsh reaction conditions (i.e., organic solvent, extreme pH, high temperature) associated with direct radiofluorination are usually incompatible with fragile protein samples. To date, therefore, the incorporation of radiolabeled prosthetic groups into biomolecules remains the method of choice.²³(,)²⁴ N-Succinimidyl-4-[¹⁸F]fluorobenzoate ([¹⁸F]SFB),²⁵⁻³⁷ a Bolton-Hunter type reagent that reacts with the primary amino groups of biomolecules, is a very versatile prosthetic group for the ¹⁸F-labeling of a wide spectrum of biological entities, in terms of its evident in vivo stability and high radiolabeling yield. After labeling with [¹⁸F]SFB, the resulting [F]fluorobenzoylated biomolecules could be explored as potential PET tracers for in vivo imaging studies.¹ Most [¹⁸F]SFB radiosyntheses described in the current literatures require two or even three reactors and multiple purifications by using either solid phase extraction (SPE) or high-performance liquid chromatography (HPLC). Such lengthy processes hamper its routine production and widespread applications in the radiolabeling of biomolecules. Although several module-assisted [¹⁸F]SFB syntheses have been reported²⁹⁻³²,⁴¹⁻⁴² they are mainly based on complicated and lengthy procedures using costly commercially-available radiochemistry boxes (Table 1). Therefore, further simplification of the radiosynthesis of [¹⁸F]SFB using a low-cost setup would be very beneficial for its adaption to an automated process. Herein, we report a concise preparation of [¹⁸F]SFB, based on a simplified one-pot microwave-assisted synthesis (Figure 1). Our approach does not require purification between steps or any aqueous reagents. In addition, microwave irradiation, which has been used in the syntheses of several PET tracers,³⁸⁻⁴¹ can gives higher RCYs and better selectivity than the corresponding thermal reactions or they provide similar yields in shorter reaction times.³⁸Most importantly, when labeling biomolecules, the time saved could be diverted to subsequent bioconjugation or PET imaging step. ²⁸(,)⁴³The novelty of our improved [¹⁸F]SFB synthesis is two-fold: (1) the anhydrous deprotection strategy requires no purification of intermediate(s) between each step and (2) the microwave-assisted radiochemical transformations enable the rapid, reliable production of [¹⁸F]SFB.