Evaluation of the effects of locally applied rosuvastatin on bone formation in a three-dimensional reconstruction rabbit xenograft model

Background/aim: Guided bone regeneration (GBR) is commonly performed to repair bone defects, and rigid occlusive titanium barriers play a vital role in bone formation in regions with no prior bone tissue. The statin, rosuvastatin (RSV), strongly affects bone apposition when applied locally. Here, we aimed to evaluate the anabolic effects of locally applied RSV with a xenograft placed on rabbit calvaria. Materials and methods: Two rigid occlusive titanium caps were used in 16 rabbits after decorticating the calvarial bone. In the control group, the area under the cap was filled with a xenograft, while in the RSV group, a xenograft in combination with RSV (1 mg) was used. In both groups, at 6 and 12 weeks, new bone, residual graft, soft tissue areas, and histological and radiological bone volume were evaluated. Results: At 12 weeks, histologically, the RSV group exhibited superior new bone proportion values, and radiologically, new bone and total bone volume in the RSV group were significantly higher than in the control group (p < 0.05); there were no significant differences at 6 weeks (p > 0.05). Conclusion: According to our results, RSV applied locally under a titanium barrier on an area to be repaired with bone grafts increases new bone and total bone volume.

3D DESI-MS lipid imaging in a xenograft model of glioblastoma: a proof of principle.

Desorption electrospray ionisation mass spectrometry (DESI-MS) can image hundreds of molecules in a 2D tissue section, making it an ideal tool for mapping tumour heterogeneity. Tumour lipid metabolism has gained increasing attention over the past decade; and here, lipid heterogeneity has been visualised in a glioblastoma xenograft tumour using 3D DESI-MS imaging. The use of an automatic slide loader automates 3D imaging for high sample-throughput. Glioblastomas are highly aggressive primary brain tumours, which display heterogeneous characteristics and are resistant to chemotherapy and radiotherapy. It is therefore important to understand biochemical contributions to their heterogeneity, which may be contributing to treatment resistance. Adjacent sections to those used for DESI-MS imaging were used for H&E staining and immunofluorescence to identify different histological regions, and areas of hypoxia. Comparing DESI-MS imaging with biological staining allowed association of different lipid species with hypoxic and viable tissue within the tumour, and hence mapping of molecularly different tumour regions in 3D space. This work highlights that lipids are playing an important role in the heterogeneity of this xenograft tumour model, and DESI-MS imaging can be used for lipid 3D imaging in an automated fashion to reveal heterogeneity, which is not apparent in H&E stains alone.

Long-Term Monitoring of Donor Xenogeneic Testis Tissue Grafts and Cell Implants in Recipient Mice Using Ultrasound Biomicroscopy.

Testis tissue xenografting and testis cell aggregate implantation from various donor species into recipient mice are novel models for the study and manipulation of testis formation and function in target species. Thus far, the analysis of such studies has been limited to surgical or post-mortem retrieval of samples. Here we used ultrasound biomicroscopy (UBM) to monitor the development of neonatal porcine testis grafts and implants in host mice for 24 wk, and to correlate UBM and (immuno)histologic changes. This led to long-term visualization of gradual changes in volume, dimension and structure of grafts and implants; detection of a 4 wk developmental gap between grafts and implants; and revelation of differences in implant development depending on the craniocaudal site of implantation on the back of host mice. Our data support the reliability and precision of UBM for longitudinal study of transplants, which eliminates the need for frequent surgical sampling.

Validation of the use of a fluorescent PARP1 inhibitor for the detection of oral, oropharyngeal and oesophageal epithelial cancers.

For oral, oropharyngeal and oesophageal cancer, the early detection of tumours and of residual tumour after surgery are prognostic factors of recurrence rates and patient survival. Here, we report the validation, in animal models and a human, of the use of a previously described fluorescently labelled small-molecule inhibitor of the DNA repair enzyme poly(ADP-ribose) polymerase 1 (PARP1) for the detection of cancers of the oral cavity, pharynx and oesophagus. We show that the fluorescent contrast agent can be used to quantify the expression levels of PARP1 and to detect oral, oropharyngeal and oesophageal tumours in mice, pigs and fresh human biospecimens when delivered topically or intravenously. The fluorescent PARP1 inhibitor can also detect oral carcinoma in a patient when applied as a mouthwash, and discriminate between fresh biopsied samples of the oral tumour and the surgical resection margin with more than 95% sensitivity and specificity. The PARP1 inhibitor could serve as the basis of a rapid and sensitive assay for the early detection and for the surgical-margin assessment of epithelial cancers of the upper intestinal tract.

Circadian rhythm impacts preclinical FDG-PET quantification in the brain, but not in xenograft tumors.

The inner clock of biological organisms plays a pivotal role and has strong effects on metabolic processes such as glucose consumption. Since the commonly used positron emission tomography (PET) tracer 18F-flourodeoxygucose (FDG) is a glucose analogue, it is not surprising that the FDG distribution in mice and humans has been shown to succumb to daily rhythms. In preclinical studies, the circadian rhythm of animals is often not considered, and studies are performed at different times of day. Only a few studies have analyzed the effect of the circadian rhythm on FDG uptake in mice, and none of these studies included human tumor xenografts. Therefore, it is not known how strongly a preclinical tumor study is influenced by the time of day. In this work, the effect of the circadian rhythm on FDG uptake in human tumor xenografts and other organs was analyzed. CD1 nu/nu mice were kept for three weeks under a 12 h light/12 h dark rhythm and then injected s.c. with PC3 or A431 tumor cells. When the tumors had reached an appropriate volume, FDG-PET scans were performed on different animal groups (n = 4-5) every 4 h over a time period from 8 A.M. to 8 P.M. Tracer uptake in the tumors and in other organs was determined based on the PET scans and biodistribution studies. The standardized uptake value and %injected dose/cc of the tumors remained constant over the whole observed time period, and no statistically significant differences were determined according to the PET analysis. In the brain, we found a small but statistically significant increase from noon to 4 P.M., which led to a decrease in the tumor-to-brain ratio. No evidence for an effect of the circadian rhythm on FDG uptake could be found in subcutaneous tumors, however, in brain studies the circadian rhythm needs to be considered.

Development of a high affinity Anticalin® directed against human CD98hc for theranostic applications.

Enhanced amino acid supply and dysregulated integrin signaling constitute two hallmarks of cancer and are pivotal for metastatic transformation of cells. In line with its function at the crossroads of both processes, overexpression of CD98hc is clinically observed in various cancer malignancies, thus rendering it a promising tumor target. Methods: We describe the development of Anticalin proteins based on the lipocalin 2 (Lcn2) scaffold against the human CD98hc ectodomain (hCD98hcED) using directed evolution and protein design. X-ray structural analysis was performed to identify the epitope recognized by the lead Anticalin candidate. The Anticalin - with a tuned plasma half-life using PASylation® technology - was labeled with 89Zr and investigated by positron emission tomography (PET) of CD98-positive tumor xenograft mice. Results: The Anticalin P3D11 binds CD98hc with picomolar affinity and recognizes a protruding loop structure surrounded by several glycosylation sites within the solvent exposed membrane-distal part of the hCD98hcED. In vitro studies revealed specific binding activity of the Anticalin towards various CD98hc-expressing human tumor cell lines, suggesting broader applicability in cancer research. PET/CT imaging of mice bearing human prostate carcinoma xenografts using the optimized and 89Zr-labeled Anticalin demonstrated strong and specific tracer accumulation (8.6 ± 1.1 %ID/g) as well as a favorable tumor-to-blood ratio of 11.8. Conclusion: Our findings provide a first proof of concept to exploit CD98hc for non-invasive biomedical imaging. The novel Anticalin-based αhCD98hc radiopharmaceutical constitutes a promising tool for preclinical and, potentially, clinical applications in oncology.

Mean Scatterer Spacing Estimation Using Cepstrum-Based Continuous Wavelet Transform.

The goal of this study was to develop an ultrasound (US) scatterer spacing estimation method using an enhanced cepstral analysis based on continuous wavelet transforms (CWTs). Simulations of backscattering media containing periodic and quasi-periodic scatterers were carried out to test the developed algorithm. Experimental data from HT-29 pellets and in vivo PC3 tumors were then used to estimate the mean scatterer spacing. For simulated media containing quasi-periodic scatterers at 1-mm and 100- [Formula: see text] spacing with 5% positional variation, the developed algorithm yielded a spacing estimation error of ~1% for 25- and 55-MHz US pulses. The mean scatterer spacing of HT-29 cell pellets (31.97 [Formula: see text]) was within 3% of the spacing obtained from histology and agreed with the predicted spacing from simulations based on the same pellets for both frequencies. The agreement extended to in vivo PC3 tumors estimation of the spacing with a variance of 1.68% between the spacing derived from the tumor histology and the application of the CWT to the experimental results. The developed technique outperformed the traditional cepstral methods as it can detect nonprominent peaks from quasi-random scatterer configurations. This work can be potentially used to detect morphological tissue changes during normal development or disease treatment.

Two-Photon Autofluorescence Imaging of Fixed Tissues: Feasibility and Potential Values for Biomedical Applications.

The value of optical redox imaging (ORI) of cells/tissues based on the intrinsic fluorescences of NADH (nicotinamide adenine dinucleotide) and oxidized flavoproteins (containing flavin adenine dinucleotide, i.e., FAD) has been demonstrated for potential biomedical applications including diagnosis, prognosis, and determining treatment response. However, the Chance redox scanner (a 3D cryogenic tissue imager) is limited by spatial resolution (~50 µm), and tissue ORI using fluorescence microscopy (single or multi-photon) is limited by the light penetration depth. Furthermore, viable or snap-frozen tissues are usually required. In this project, we aimed to study whether ORI may be achieved for unstained fixed tissue using a state-of-the-art modern Serial Two-Photon (STP) Tomography scanner that can rapidly acquire multi-plane images at micron resolution. Tissue specimens of mouse muscle, liver, and tumor xenografts were harvested and fixed in 4% paraformaldehyde (PFA) for 24 h. Tissue blocks were scanned by STP Tomography under room temperature to acquire the autofluorescence signals (NADH channel: excitation 750 nm, blue emission filter; FAD channel: excitation 860 nm, green emission filter). We observed remarkable signals with significant intra-tissue heterogeneity in images of NADH, FAD and redox ratio (FAD/(NADH+FAD)), which are worthy of further investigation for extracting biological information.

Imaging CAR T Cell Trafficking with eDHFR as a PET Reporter Gene.

Cell-based therapeutics have considerable promise across diverse medical specialties; however, reliable human imaging of the distribution and trafficking of genetically engineered cells remains a challenge. We developed positron emission tomography (PET) probes based on the small-molecule antibiotic trimethoprim (TMP) that can be used to image the expression of the Escherichia coli dihydrofolate reductase enzyme (eDHFR) and tested the ability of [18F]-TMP, a fluorine-18 probe, to image primary human chimeric antigen receptor (CAR) T cells expressing the PET reporter gene eDHFR, yellow fluorescent protein (YFP), and Renilla luciferase (rLuc). Engineered T cells showed an approximately 50-fold increased bioluminescent imaging signal and 10-fold increased [18F]-TMP uptake compared to controls in vitro. eDHFR-expressing anti-GD2 CAR T cells were then injected into mice bearing control GD2- and GD2+ tumors. PET/computed tomography (CT) images acquired on days 7 and 13 demonstrated early residency of CAR T cells in the spleen followed by on-target redistribution to the GD2+ tumors. This was corroborated by autoradiography and anti-human CD8 immunohistochemistry. We found a high sensitivity of detection for identifying tumor-infiltrating CD8 CAR T cells, ∼11,000 cells per mm3. These data suggest that the [18F]-TMP/eDHFR PET pair offers important advantages that could better allow investigators to monitor immune cell trafficking to tumors in patients.

Multilayered Activatable Nanoprobe for Ultra-Bright Tumor Imaging.

The development of tumor targeted probes with strong signal and high contrast is always challenging in cancer imaging. Here, a unique multilayered activatable nanoprobe (MAN) is prepared to fulfill this long-standing goal. MAN adopts a versatile layer-by-layer fabrication technique that sequentially assembles multifunctional polyelectrolytes onto nanoparticles via charge-charge interaction. Unlike the common one-probe-one-fluorochrome construct, MAN offers a dramatic fluorescence enhancement by transporting a large quantity of quenched fluorochromes for maximal signal and contrast. Excellent signal amplification and retention with negligible cytotoxicity is observed in cell study. Upon systemic injection into mice, MAN quickly accumulates in tumor and its fluorescent signal is turned on by proteases overexpressed in tumors, resulting in >700% tumor-to-normal-tissue contrast. This multilayered fabrication provides a simple and powerful universal platform to design sensitive tumor imaging probes.

Test-Retest Performance of a 1-Hour Multiparametric MR Image Acquisition Pipeline With Orthotopic Triple-Negative Breast Cancer Patient-Derived Tumor Xenografts.

Preclinical imaging is critical in the development of translational strategies to detect diseases and monitor response to therapy. The National Cancer Institute Co-Clinical Imaging Resource Program was launched, in part, to develop best practices in preclinical imaging. In this context, the objective of this work was to develop a 1-hour, multiparametric magnetic resonance image-acquisition pipeline with triple-negative breast cancer patient-derived xenografts (PDXs). The 1-hour, image-acquisition pipeline includes T1- and T2-weighted scans, quantitative T1, T2, and apparent diffusion coefficient (ADC) parameter maps, and dynamic contrast-enhanced (DCE) time-course images. Quality-control measures used phantoms. The triple-negative breast cancer PDXs used for this study averaged 174 ± 73 µL in volume, with region of interest-averaged T1, T2, and ADC values of 1.9 ± 0.2 seconds, 62 ± 3 milliseconds, and 0.71 ± 0.06 µm2/ms (mean ± SD), respectively. Specific focus was on assessing the within-subject test-retest coefficient-of-variation (CVWS) for each of the magnetic resonance imaging metrics. Determination of PDX volume via manually drawn regions of interest is highly robust, with ∼1% CVWS. Determination of T2 is also robust with a ∼3% CVWS. Measurements of T1 and ADC are less robust with CVWS values in the 6%-11% range. Preliminary DCE test-retest time-course determinations, as quantified by area under the curve and Ktrans from 2-compartment exchange (extended Tofts) modeling, suggest that DCE is the least robust protocol, with ∼30%-40% CVWS.

Microfluidic Assembly of siRNA-Loaded Micelleplexes for Tumor Targeting in an Orthotopic Model of Ovarian Cancer.

The use of cationic polymers to interact with negatively charged siRNA via charge complexation to form polyelectrolyte complexes has been widely studied ever since the 1998 report on RNA interference. These polyelectrolyte complex formulations aim to overcome the many pitfalls associated with the use of RNA interference as a potential cancer therapy. The triblock copolymer polyethylenimine-polycaprolactone-polyethylene glycol (PEI-PCL-PEG) contains the cation PEI and has been shown to be an efficient carrier capable of complexing with nucleic acids for gene delivery. This copolymer system also allows for targeting moieties to be linked to the micelleplex, thereby exploiting overexpressed receptors (such as the folate receptor) located within tumors. Additionally, we demonstrated recently that microfluidic mixing of PEI-PCL-PEG nanoparticles allows for the rapid, scaled-up production of micelleplexes while maintaining small and uniform particle distributions. The preparation of small and reproducible particles is imperative for clinical translation of nanomedicine and for tumor targeting via systemic administration. Furthermore, to enable tracing of its deposition in vivo after its administration, micelleplexes can be radiolabeled. To assess tumor targeting over time, the noninvasive imaging technique single-photon emission computed tomography (SPECT) offers the ability to examine the same subject at multiple time points and generate biodistribution profiles. Since the biodistribution and tumor targeting of the therapeutic load of micelleplexes is of foremost interest, we recently described an approach to modify siRNA with a DTPA (diethylenetriaminepentaacetic acid) chelator. Herein, we explain the details of encapsulating indium-labeled siRNA via microfluidic mixing in PEI-PCL-PEG nanoparticles with a folic acid targeting ligand for assessment of their in vivo tumor targeting in an orthotopic ovarian cancer model.

In vivo imaging of insulin-secreting human pancreatic ductal cells using MRI reporter gene technique: A feasibility study.

PURPOSE: The purpose of this study was to investigate the feasibility of in vivo imaging of human pancreatic ductal cells by OATP1B3 reporter gene under MRI. METHODS: A human cell line (PANC-1) derived from the pancreatic ductal epithelium was used in this study. After transduction of OATP1B3, the cellular physiological functions and the ability of intracellular uptake of the MRI contrast medium (Gd-EOB-DTPA) were examined. Induced differentiation of the PANC-1 cells into hormone-secreting cells were performed to simulate pancreatic ß-like cells. The hormone-secreting cells were implanted into rats and in vivo MRI was evaluated. RESULTS: The mRNA and proteins of OATP1B3 were highly expressed. No significant change of cellular physiological functions was found after the expression. After induced differentiation, the hormone secretion capacities of the OATP1B3-expressing PANC-1 cells were confirmed. Intra-cellular uptake of Gd-EOB-DTPA was determined in vitro by inductively coupled plasma mass spectrometry and MRI. In vivo MRI of the OATP1B3-expressing xenograft revealed an increased signal intensity after contrast enhancement. CONCLUSION: OATP1B3 can be used as a safe and feasible in vivo MRI gene reporter for human pancreatic ductal cells.

Near infrared imaging of epidermal growth factor receptor positive xenografts in mice with domain I/II specific antibody fragments.

Epidermal growth factor receptor (EGFR) is a transmembrane cell surface receptor that is frequently overexpressed and/or mutated in many cancers. Therapies targeting EGFR have poor outcomes due to the lack of reliable diagnostic tests to monitor EGFR. Current in vitro EGFR diagnostic methods are invasive, requiring biopsies, which limits tumor sampling and availability. EGFR molecular imaging provides non-invasive whole-body images capable of detecting primary tumors and metastases, which can be used to diagnose and monitor response to therapy. Methods: We evaluated properties of two anti-EGFR fragments, 8708 and 8709, as molecular-targeted imaging probes. 8708 and 8709 are anti-EGFR antigen binding fragments (Fabs) that recognize domain I/II of EGFR, which is distinct from epitopes recognized by current anti-EGFR therapeutic antibodies. We used complementarity determining region sequences from 8708 and 8709 Fabs to generate an anti-EGFR IgG and (scFv)2 and scFv-Fc antibody fragments. We expressed, purified, and labeled the IgG and fragments with IRDye800CW and used them to image EGFR-positive and -negative xenografts in CD-1 nude mice. 8709 scFv-Fc was also tested for competitive binding with the therapeutic anti-EGFR antibody nimotuzumab and for quantifying ratios of EGFR and EGFRvIII deletion mutant. Results: IRDye800CW-labeled 8708 (scFv)2 and 8709 scFv-Fc imaging probes showed high levels of accumulation and good retention in EGFR-positive xenografts, with peak accumulation occurring at 24 and 48 hours post injection, respectively. IRDye680RD-labeled 8709 scFv-Fc did not compete with IRDye800CW-labeled nimotuzumab for EGFR binding as assayed by flow cytometry using an EGFR-positive cell line. IRDye680RD-labeled 8709 scFv-Fc and IRDye800CW-labeled nimotuzumab used in combination were able to determine the ratio of cells expressing EGFR and a deletion mutant EGFRvIII. Conclusion: IRDye800CW-labeled 8708 (scFv)2 and 8709 scFv-Fc had desirable binding affinities, clearance times, and tumor accumulation to be used for imaging in combination with current EGFR targeted therapies. This study highlights the potential for using 8708 (scFv)2 and 8709 scFv-Fc as EGFR diagnostic and therapy monitoring tools.

Intravenous Administration-Oriented Pharmacokinetic Model for Dynamic Bioluminescence Imaging.

OBJECTIVE: In vivo bioluminescence imaging (BLI) is a promising tool for monitoring the growth and metastasis of tumors. However, quantitative BLI research based on intravenous (IV) injection is limited, which greatly restricts its further application. To address this problem, we designed a pharmacokinetic (PK) model which is suitable for applying on IV administration of small amounts of D-Luciferin. METHODS: After three weeks of postimplantation, mkn28-luc xenografted mice were subjected to 40-min dynamic BLI immediately following D-Luciferin intravenous injection on days 1, 3, 5, 7, and 9. Furthermore, the PK model was applied on dynamic BLI data to obtain the sum of kinetic rate constants (SKRC). RESULTS: Results showed that the SKRC values decreased rapidly with the growth of the tumor. There was a statistical difference between the SKRC values measured at different time points, while the time point of luminous intensity peak was unaffected by the growth of the tumor. CONCLUSION: In short, our results imply that dynamic BLI combined with our PK model can predict tumor growth earlier and with higher sensitivity compared to the conventional method, which is crucial for improving drug evaluation efficacy. In addition, the dynamic BLI may provide a valuable reference for the noninvasive acquiring arterial input function, which may also provide a new application prospect for hybrid PET-optical imaging.

Optical Redox Imaging of Lonidamine Treatment Response of Melanoma Cells and Xenografts.

PURPOSE: Fluorescence of co-enzyme reduced nicotinamide adenine dinucleotide (NADH) and oxidized flavoproteins (Fp) provides a sensitive measure of the mitochondrial redox state and cellular metabolism. By imaging NADH and Fp, we investigated the utility of optical redox imaging (ORI) to monitor cellular metabolism and detect early metabolic response to cancer drugs. PROCEDURES: We performed ORI of human melanoma DB-1 cells in culture and DB-1 mouse xenografts to detect the redox response to lonidamine (LND) treatment. RESULTS: For cultured cells, LND treatment for 45 min significantly lowered NADH levels with no significant change in Fp, resulting in a significant increase in the Fp redox ratio (Fp/(NADH+Fp)); 3-h prolonged treatment led to a decrease in NADH and an increase in Fp and a more oxidized redox state compared to control. Significant decrease in the mitochondrial redox capacity of LND-treated cells was observed for the first time. For xenografts, 45-min LND treatment resulted in a significant reduction of NADH content, no significant changes in Fp content, and a trend of increase in the Fp redox ratio. Intratumor redox heterogeneity was observed in both control and LND-treated groups. CONCLUSION: Our results support the utility of ORI for evaluating cellular metabolism and monitoring early metabolic response to cancer drugs.

Evaluation of antibody fragment properties for near-infrared fluorescence imaging of HER3-positive cancer xenografts.

In vivo imaging is influenced by the half-life, tissue penetration, biodistribution, and affinity of the imaging probe. Immunoglobulin G (IgG) is composed of discrete domains with known functions, providing a template for engineering antibody fragments with desired imaging properties. Here, we engineered antibody-based imaging probes, consisting of different combinations of antibody domains, labeled them with the near-infrared fluorescent dye IRDye800CW, and evaluated their in vivo imaging properties. Antibody-based imaging probes were based on an anti-HER3 antigen binding fragment (Fab) isolated using phage display. Methods: We constructed six anti-HER3 antibody-based imaging probes: a single chain variable fragment (scFv), Fab, diabody, scFv-CH3, scFv-Fc, and IgG. IRDye800CW-labeled, antibody-based probes were injected into nude mice bearing FaDu xenografts and their distribution to the xenograft, liver, and kidneys was evaluated. Results: These imaging probes bound to recombinant HER3 and to the HER3-positive cell line, FaDu. Small antibody fragments with molecular weight <60 kDa (scFv, diabody, and Fab) accumulated rapidly in the xenograft (maximum accumulation between 2-4 h post injection (hpi)) and cleared primarily through the kidneys. scFv-CH3 (80 kDa) had fast clearance and peaked in the xenograft between 2-3 hpi and cleared from xenograft in a rate comparable to Fab and diabody. IgG and scFv-Fc persisted in the xenografts for up to 72 hpi and distributed mainly to the xenograft and liver. The highest xenograft fluorescence signals were observed with IgG and scFv-Fc imaging probes and persisted for 2-3 days. Conclusion: These results highlight the utility of using antibody fragments to optimize clearance, tumor labeling, and biodistribution properties for developing anti-HER3 probes for image-guided surgery or PET imaging.

The Distribution and Imaging of 99mTc-nGO-PEG-FA in Human Patu8988 Tumor-Bearing Nude Mice.

Background: To study the distribution and imaging of 99mTc-nGO-PEG-FA in human pancreatic cancer Patu8988 tumor-bearing nude mice, and to explore its usefulness as an imaging reagent for pancreatic cancer. Materials and Methods: Natural graphite powder was used as raw material to prepare the nanosized graphene oxide (nGO) by using the modified Hummers method, and then was covalently modified by polyethylene glycol (PEG) on the surface of nGO. The nGO was further optimized by in vitro cell experiment, and then conjugated with the targeting molecule folic acid (FA) to form nGO-PEG-FA system. The nGO-PEG-FA was finally labeled by radioactive nuclide 99mTc by direct labeling method to form the 99mTc-nGO-PEG-FA molecular imaging probe. Nude mice bearing patu8988 pancreatic cancer xenografts were intravenous injection (I.V.) injected with 99mTc-nGO-PEG-FA, and the distribution of 99mTc-nGO-PEG-FA in nude mice at different time course was investigated by determination of tissue uptake of radioactivity (%ID/g), as well as the single photon emission computed tomography (SPECT) imaging at different time course. Results: The labeling rate of nGO-PEG-FA with 99mTc was (90.08 ± 2.34)%, and the highest binding rate of 99mTc-nGO-PEG-FA with Patu8988 cells was (3.15 ± 0.31)%. The radioactive uptake in tumor reached (5.11 ± 1.23)%ID/g at 6 h after I.V. injection of 99mTc-nGO-PEG-FA in nude mice. Meanwhile, the radioactive uptake in liver, spleen, and lung was also high and reached (10.33 ± 1.22)%ID/g, (5.86 ± 0.59)%ID/g, and (3.55 ± 0.93)%ID/g, respectively, whereas less radioactivity uptake was observed in the heart (1.12 ± 0.33)%ID/g and blood (2.76 ± 0.39)%ID/g, respectively. The tumors can be clearly imaged at 4.0-6.0 h after 99mTc-nGO-PEG-FA injection. Conclusions: 99mTc-nGO-PEG-FA can efficiently target pancreatic cancer, which may be developed as an imaging agent for pancreatic cancer.

Tomographic late evaluation of xenogeneic bone grafts in sockets of impacted third molars.

OBJECTIVE: It is necessary to preserve height and thickness of the alveolar bone to facilitate rehabilitation with osteointegratable implants or simply to maintain bone integrity after extraction. Biomaterials associated with resorbable or non-resorbable membranes, when placed in the region of the socket, may contribute to avoid this unwanted reabsorption. OBJECTIVE: The objective of this study was to evaluate the distance of the crest of alveolar ridge to the cementoenamel junction (CEJ) of the lower second molars and the bone density of the third molar socket filled with Gen-Tech®, 5 years after an exodontia using cone beam computed tomography (CBCT) to visualize the central region of the sockets, without overlapping of the buccal and lingual cortical bones. MATERIAL AND METHODS: A total of 12 individuals from an initial group of 39 patients submitted to extraction of the unruptured lower third molars and grafting of an association of inorganic bovine bone matrix, organic bovine bone matrix, collagen and bone morphogenetic proteins (BMP) (Gen-Tech®) on one side and the contralateral sockets filled only by clot, returned to control after 5 years, and were submitted to CBCT. The distance from the crest of alveolar bone to the CEJ and the bone density (BD) were measured using the i-CAT Vision Software. RESULTS: The results showed that the distance from the crest of alveolar bone to the CEJ in the control group was similar to that observed before the exodontia; in the experimental group, this distance was smaller. Considering the BD measurement, a significantly higher density was observed in the experimental group (p<0.05). CONCLUSION: Part of the biomaterial was not absorbed and allowed the stability of the evaluated parameters after 5 years, being able to be used as a bone substitute in the socket.

Preparation of 68Ga-PSMA-11 with a Synthesis Module for Micro PET-CT Imaging of PSMA Expression during Prostate Cancer Progression.

Objective: To synthesize 68Ga-Glu-urea-Lys(Ahx)-HBED-CC (68Ga-PSMA-11) with a synthesis module and investigate PET-CT imaging to monitor PSMA expression during prostate cancer (PCa) progression and tumor growth in mice bearing subcutaneous PCa xenografts. Method: The radiochemical purity and stability of 68Ga-PSMA-11 were determined via radio-HPLC. The PCa cell lines of different PSMA expression levels (PC3, VCAP±, CWR22RV1+, and LNCaP++) were selected to mimic the PCa progression. 68Ga-PSMA-11 biodistribution was studied by dissection method and in vivo imaging with micro PET-CT. The expression levels of PSMA in tumor cells and tissues were analyzed by immunofluorescence, flow cytometry, and western blot. The correlation between PSMA expression and radio-uptake was also evaluated. 2-PMPA preadministration served as a block group. Results: The radiochemical purity of 68Ga-PSMA-11 was 99.6 ± 0.1% and stable in vitro for 2 h. The equilibrium binding constant (Kd) of 68Ga-PSMA-11 to LNCaP, CWR22Rv1, PC-3, and VCAP cells was 4.3 ± 0.8 nM, 16.4 ± 1.3 nM, 225.3 ± 20.8 nM, and 125.6 ± 13.1 nM, respectively. Results of tumor uptake (% ID and % ID/g or % ID/cm3) of 68Ga-PSMA-11 in biodistribution and micro PET imaging were LNCaP > CWR22RV1 > PC-3 and VCAP due to different PSMA expression levels. It was confirmed by flow cytometry, western blot, and immunofluorescence. Tumor uptake (% ID/cm3) of 68Ga-PSMA-11 increased with the tumor anatomical volume in quadratic polynomial fashion and reached the peak (when tumor volume was 0.5 cm3) earlier than tumor uptake (% ID). Tumor uptake (% ID/cm3) of 68Ga-PSMA-11 based on functional volume correlated well with the PSMA expression in a linear manner (y = 9.35x + 2.59, R2 = 0.8924, and p < 0.0001); however, low dose 2-PMPA causes rapid renal clearance of increased tumor/kidney uptake of 68Ga-PSMA-11. Conclusions: The 68Ga-PSMA-11 PET-CT imaging could invasively evaluate PSMA expression during PCa progression and tumor growth with % ID/cm3 (based on functional volume) as an important index. Low dose 2-PMPA preadministration might be a choice to decrease kidney uptake of 68Ga-PSMA-11.