(18)F-labeled-bioorthogonal liposomes for in vivo targeting.

Liposomes are attractive vehicles for the controlled release of drugs and cytotoxins and have a long-standing history in medical research and clinical practice. In addition to established therapeutic indications, liposomes have several favorable properties for molecular imaging, including high stability and the ability to be labeled with radioisotopes, as well as paramagnetic and fluorescent contrast agents. However, long circulation times and difficulties in creating targeted liposomes have proven challenges for imaging. In this study, we have addressed these limitations using a recently developed strategy for bioorthogonal conjugation, the reaction between tetrazines and trans-cyclooctenes. By coating radiolabeled liposomes with trans-cyclooctene and pretargeting with a tetrazine coupled to a targeted peptide, we were able to selectively enhance the retention of liposomes and bind them to tumor tissue in live animals. The rapid reaction between tetrazines and trans-cyclooctenes allowed imaging to be performed with the short-lived PET tracer (18)F, yielding signal-to-background activity ratios of 7:1. The covalent, bioorthogonally driven tumor-targeting of liposomes by in vivo click chemistry is promising and should be explored for more selective and rapid delivery of radiodiagnostics and radiotherapeutics, two classes of drugs which particularly benefit from fast clearance, low nonspecific binding, and the associated reduced toxicity to kidneys and bone marrow.

Endogenous expression of Hras(G12V) induces developmental defects and neoplasms with copy number imbalances of the oncogene.

We developed mice with germline endogenous expression of oncogenic Hras to study effects on development and mechanisms of tumor initiation. They had high perinatal mortality, abnormal cranial dimensions, defective dental ameloblasts, and nasal septal deviation, consistent with some of the features of human Costello syndrome. These mice developed papillomas and angiosarcomas, which were associated with Hras(G12V) allelic imbalance and augmented Hras signaling. Endogenous expression of Hras(G12V) was also associated with a higher mutation rate in vivo. Tumor initiation by Hras(G12V) likely requires augmentation of signal output, which in papillomas and angiosarcomas is achieved via increased Hras-gene copy number, which may be favored by a higher mutation frequency in cells expressing the oncoprotein.

A modular labeling strategy for in vivo PET and near-infrared fluorescence imaging of nanoparticle tumor targeting.

UNLABELLED: Advances in preclinical molecular imaging have generated new opportunities to noninvasively visualize the biodistribution and tumor targeting of nanoparticle therapeutics. Capitalizing on recent achievements in this area, we sought to develop an (89)Zr-based labeling strategy for liposomal nanoparticles that accumulate in tumors via passive targeting mechanisms. METHODS: (89)Zr-labeled liposomes were prepared using 2 different approaches: click labeling and surface chelation. Pharmacokinetic and biodistribution studies, as well as PET/CT imaging of the radiolabeled nanoparticles, were performed on a mouse model of breast cancer. In addition, a dual PET/optical probe was prepared by incorporation of a near-infrared fluorophore and tested in vivo by PET and near-infrared fluorescence imaging. RESULTS: The surface chelation approach proved to be superior in terms of radiochemical yield and stability, as well as in vivo performance. Accumulation of these liposomes in tumor peaked at 24 h after injection and was measured to be 13.7 ± 1.8 percentage injected dose per gram. The in vivo performance of this probe was not essentially perturbed by the incorporation of a near-infrared fluorophore. CONCLUSION: We have developed a highly modular and efficient strategy for the labeling of liposomal nanoparticles with (89)Zr. In xenograft and orthotopic mouse models of breast cancer, we demonstrated that the biodistribution of these nanoparticles can be visualized by PET imaging. In combination with a near-infrared dye, these liposomal nanoparticles can serve as bimodal PET/optical imaging agents. The liposomes target malignant growth, and their bimodal features may be useful for simultaneous PET and intraoperative imaging.

Fluorine-labeled dasatinib nanoformulations as targeted molecular imaging probes in a PDGFB-driven murine glioblastoma model.

Dasatinib, a new-generation Src and platelet-derived growth factor receptor (PDGFR) inhibitor, is currently under evaluation in high-grade glioma clinical trials. To achieve optimum physicochemical and/or biologic properties, alternative drug delivery vehicles may be needed. We used a novel fluorinated dasatinib derivative (F-SKI249380), in combination with nanocarrier vehicles and metabolic imaging tools (microPET) to evaluate drug delivery and uptake in a platelet-derived growth factor B (PDGFB)-driven genetically engineered mouse model (GEMM) of high-grade glioma. We assessed dasatinib survival benefit on the basis of measured tumor volumes. Using brain tumor cells derived from PDGFB-driven gliomas, dose-dependent uptake and time-dependent inhibitory effects of F-SKI249380 on biologic activity were investigated and compared with the parent drug. PDGFR receptor status and tumor-specific targeting were non-invasively evaluated in vivo using (18)F-SKI249380 and (18)F-SKI249380-containing micellar and liposomal nanoformulations. A statistically significant survival benefit was found using dasatinib (95 mg/kg) versus saline vehicle (P < .001) in tumor volume-matched GEMM pairs. Competitive binding and treatment assays revealed comparable biologic properties for F-SKI249380 and the parent drug. In vivo, Significantly higher tumor uptake was observed for (18)F-SKI249380-containing micelle formulations [4.9 percentage of the injected dose per gram tissue (%ID/g); P = .002] compared to control values (1.6%ID/g). Saturation studies using excess cold dasatinib showed marked reduction of tumor uptake values to levels in normal brain (1.5%ID/g), consistent with in vivo binding specificity. Using (18)F-SKI249380-containing micelles as radiotracers to estimate therapeutic dosing requirements, we calculated intratumoral drug concentrations (24-60 nM) that were comparable to in vitro 50% inhibitory concentration values. (18)F-SKI249380 is a PDGFR-selective tracer, which demonstrates improved delivery to PDGFB-driven high-grade gliomas and facilitates treatment planning when coupled with nanoformulations and quantitative PET imaging approaches.

Optimizing tumor targeting of the lipophilic EGFR-binding radiotracer SKI 243 using a liposomal nanoparticle delivery system.

Positron emission tomography (PET) of epidermal growth factor receptor (EGFR) kinase-specific radiolabeled tracers could provide a means for non-invasively characterizing EGFR expression and signaling activity in patients' tumors before, during, and after therapy with EGFR inhibitors. Towards this goal, our group has developed PET tracers which irreversibly bind to EGFR. However, tumor uptake is relatively low because of both the lipophilicity of such tracers (e.g. the morpholino-[124I]-IPQA [SKI 212243]), with octanol-to-water partition coefficients of up to 4, and a short dwell time in the blood and significant hepatobiliary clearance and intestinal reuptake. Liposomal nanoparticle delivery systems may favorably alter the pharmacokinetic profile and improve tumor targeting of highly lipophilic but otherwise promising cancer imaging tracers, such as the EGFR inhibitor SKI 243. SKI 243 is therefore an interesting model molecule for incorporation into lipid-based nanoparticles, as it would not only improve their solubility but also increase the circulation time, availability and, potentially, targeting of tumors. In the current study, we compared the pharmacokinetics and tumor targeting of the bare EGFR kinase-targeting radiotracer SKI 212243 (SKI 243) with that of the same tracer embedded in liposomes. SKI 243 and liposomal SKI 243 are both taken up by tumor xenografts but liposomal SKI 243 remained in the blood longer and consequently exhibited a 3- to 6-fold increase in uptake in the tumor among several other organs.

Synthesis of a tissue-engineered periosteum with acellular dermal matrix and cultured mesenchymal stem cells.

Periosteal grafts can aid in bone repair by providing bone progenitor cells and acting as a barrier to scar tissue. Unfortunately, these grafts have many of the same disadvantages as bone grafts (donor site morbidity and limited donor sites). In this article, we describe a method of synthesizing a periosteum-like material using acellular human dermis and osteoblasts or mesenchymal stem cells (MSC). We show that osteoblasts readily attach to and proliferate on the acellular human dermis in vitro. In addition, osteoblasts retained the potential for differentiation in response to bone morphogenetic protein stimulation. Cells grown on the acellular human dermis were efficiently transfected with adenoviruses with no evidence of cellular toxicity. To assess for in vivo cell delivery and bone-forming potential, the acellular human dermis was seeded with green fluorescent protein (GFP)-positive MSCs, transfected with bone morphogenetic protein 2, wrapped around the adductor muscle in syngeneic mice, and used to treat critical-sized mandibular defects in nude rats. After 3 weeks, GFP-positive cells were still present, and bone had replaced the interface between the muscle and the constructs. After 6 weeks, critical-sized bone defects had been successfully healed. In conclusion, we show that an acellular human dermis can be used to synthesize a tissue-engineered periosteum capable of delivering cells and osteoinductive proteins.