Therapeutic drug delivery across the blood-brain barrier (BBB) is not only inefficient, but also nonspecific to brain stroma. These are major limitations in the effective treatment of brain cancer. Transferrin peptide (Tfpep) targeted gold nanoparticles (Tfpep-Au NPs) loaded with the photodynamic pro-drug, Pc 4, have been designed and compared with untargeted Au NPs for delivery of the photosensitizer to brain cancer cell lines. In vitro studies of human glioma cancer lines (LN229 and U87) overexpressing the transferrin receptor (TfR) show a significant increase in cellular uptake for targeted conjugates as compared to un-targeted particles. Pc 4 delivered from Tfpep-Au NPs clusters within vesicles after targeting with the Tfpep. Pc 4 continues to accumulate over a 4 hour period. Our work suggests that TfR-targeted Au NPs may have important therapeutic implications for delivering brain tumor therapies and/or providing a platform for noninvasive imaging.

Targeting gold nanoparticles (AuNPs) with two or more receptor binding peptides has been proposed to address intratumoral heterogeneity of glioblastomas that overexpress multiple cell surface receptors to ultimately improve therapeutic efficacy. AuNPs conjugated with peptides against both the epidermal growth factor and transferrin receptors and loaded with the photosensitizer phthalocyanine 4 (Pc 4) have been designed and compared with monotargeted AuNPs for in vitro and in vivo studies. The (EGFpep+Tfpep)-AuNPs-Pc 4 with a particle size of ~41 nm improved both specificity and worked synergistically to decrease time of maximal accumulation in human glioma cells that overexpressed two cell surface receptors as compared to cells that overexpressed only one. Enhanced cellular association and increased cytotoxicity were achieved. In vivo studies show notable accumulation of these agents in the brain tumor regions.

In this study, we developed a stable, nontoxic novel micelle nanoparticle to attenuate responses of endothelial cell (EC) inflammation when subjected to oxidative stress, such as observed in organ transplantation. Targeted Rapamycin Micelles (TRaM) were synthesized using PEG-PE-amine and N-palmitoyl homocysteine (PHC) with further tailoring of the micelle using targeting peptides (cRGD) and labeling with far-red fluorescent dye for tracking during cellular uptake studies. Our results revealed that the TRaM was approximately 10 nm in diameter and underwent successful internalization in Human Umbilical Vein EC (HUVEC) lines. Uptake efficiency of TRaM nanoparticles was improved with the addition of a targeting moiety. In addition, our TRaM therapy was able to downregulate both mouse cardiac endothelial cell (MCEC) and HUVEC production and release of the pro-inflammatory cytokines, IL-6 and IL-8 in normal oxygen tension and hypoxic conditions. We were also able to demonstrate a dose-dependent uptake of TRaM therapy into biologic tissues ex vivo. Taken together, these data demonstrate the feasibility of targeted drug delivery in transplantation, which has the potential for conferring local immunosuppressive effects without systemic consequences while also dampening endothelial cell injury responses.

(a) Rapamycin nanotherapeutic pre-treatment improves tracheal allograft outcome after transplantation. (b) Nanotherapy reduces aortic allograft vasculopathy. (c) Dose dependency of the nanotherapy in aortic interposition allografts.

PDGF-micelles containing TMZ have specific uptake and increased killing in glial cells compared with untargeted micelles. In vivo studies demonstrated selective accumulation of PDGF-micelles containing TMZ in orthotopic gliomas implanted in mice. Targeted micelle-based drug carrier systems hold potential for delivery of a wide variety of hydrophobic drugs thereby reducing its systemic toxicity.

Bioluminescence is a useful tool for imaging of cancer in in vivo animal models that endogenously express luciferase, an enzyme that requires a substrate for visual readout. Current bioluminescence imaging, using commonly available luciferin substrates, only lasts a short time (15–20 min). To avoid repeated administration of luciferase substrate during cancer detection and surgery, a long lasting bioluminescence imaging substrate or system is needed. A novel water-soluble biotinylated luciferase probe, B-YL (1), was synthesized. A receptor-targeted complex of B-YL with streptavidin (SA) together with a biotinylated epidermal growth factor short peptide (B-EGF) (SA/B-YL/B-EGF = 1:3:1, molar ratio) was then prepared to demonstrate selective targeting. The complex was incubated with brain cancer cell lines overexpressing the EGF receptor (EGFR) and transfected with the luciferase gene. Results show that the complex specifically detects cancer cells by bioluminescence. The complex was further used to image xenograft brain tumors transfected with a luciferase gene in mice. The complex detects the tumor immediately, and bioluminescence lasts for 5 days. Thus, the complex generates a long lasting bioluminescence for cancer detection in mice. The complex with selective targeting may be used in noninvasive cancer diagnosis and accurate surgery in cancer treatment in clinics in the future.

No abstract

The development of selectively targeted nanoparticles that can act as drug delivery vehicles is critical for improving the treatment and monitoring of glioblastoma, a life threatening disease. Rapamycin (Sirolimus, rapa), a large, lipophilic carboxylic lactone-lactam macrolide antibiotic, is recognized for its potent anti-proliferative and immunosuppressive effects in vitro and in vivo. These properties make rapa a potential chemotherapeutic agent against several tumors. Despite its promising properties, clinical applications of rapa have been limited due to its hydrophobicity, limiting its utility as an intravenously administered drug. Presently, the commercially available formulations of rapa include tablet or oral forms. Nevertheless, the low oral bioavailability of rapa limits the effectiveness of both of these forms. In addition, the lipophilicity makes the drug susceptible to attachment to the lipid membranes of normal as well as cancer cells. A selectively targeted carrier for rapa will enhance its delivery to malignant cells, avoid non-specific interactions, and reduce non-tumor toxicity. In order to design an efficient and effective drug carrier, we created a multifunctional nanocarrier that contains a tailored surface on the carrier to attach biomolecules for targeted drug delivery; a biocompatible coating which can efficiently encapsulate the hydrophobic drug thereby reducing cytotoxicity; and the capability for stimuli-induced (pH) disruption of the carrier agent for slow and controlled drug release to the desired environment, Micelles are the preferred choice of carrier as they fulfill these requirements based on their composition. Micelles containing rapamycin drug are synthesized using PEG-PE-Amine and N-palmitoyl homocysteine (PHC, pH sensitive lipid breaks in endosome pH 5.5). Specific targeting of the micelles to glioblastoma cells is achieved by PDGF (platelet derived growth factor) or EGF (epidermal growth factor) coupled to the amine moeity of the DSPE-PEG. In addition these micelles have been labeled with a NIR fluorphore to track them for cellular uptake. These micelles have an advantage of small size (<50 nm, to cross blood brain barrier) and reduced toxicity due to robust packaging of rapa drug inside the core. Leaching of the drug out of the micelle is prevented and reduces offsite cytotoxicity. Preliminary cellular uptake studies via fluorescence imaging of glioblastoma cells treated with targeted and untargeted labeled-micellar particles demonstrate receptor-mediated endocytosis. Uptake occurs rapidly within 1 to 4 hours after treatment. Once released from the micelle, the rapa kills the cells. Future experiments involve performing cytotoxicity assays, pharmacokinetic and tissue distribution studies in in vivo animal models. Note: This abstract was not presented at the meeting. Citation Format: Ann-Marie Broome, Suraj K. Dixit, Kayla Miller, Alfred Moore, Amy-Lee Bredlau. Treating brain tumors with targeted-micelles containing rapamycin. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4467. doi:10.1158/1538-7445.AM2014-4467