Polymeric nanotheranostics for real-time non-invasive optical imaging of breast cancer progression and drug release.

Polymeric nanocarriers conjugated with low molecular weight drugs are designed in order to improve their efficacy and toxicity profile. This approach is particularly beneficial for anticancer drugs, where the polymer-drug conjugates selectively accumulate at the tumor site, due to the enhanced permeability and retention (EPR) effect. The conjugated drug is typically inactive, and upon its pH- or enzymatically-triggered release from the carrier, it regains its therapeutic activity. These settings lack information regarding drug-release time, kinetics and location. Thereby, real-time non-invasive intravital monitoring of drug release is required for theranostics (therapy and diagnostics). We present here the design, synthesis and characterization of a theranostic nanomedicine, based on N-(2-hydroxypropyl) methacrylamide (HPMA) copolymer, owing its fluorescence-based monitoring of site-specific drug release to a self-quenched near-infrared fluorescence (NIRF) probe. We designed two HPMA copolymer-based systems that complement to a theranostic nanomedicine. The diagnostic system consists of self-quenched Cy5 (SQ-Cy5) as a reporter probe and the therapeutic system is based on the anticancer agent paclitaxel (PTX). HPMA copolymer-PTX/SQ-Cy5 systems enable site-specific release upon enzymatic degradation in cathepsin B-overexpressing breast cancer cells. The release of the drug occurs concomitantly with the activation of the fluorophore to its Turn-ON state. HPMA copolymer-SQ-Cy5 exhibits preferable body distribution and drug release compared with the free drug and probe when administered to cathepsin B-overexpressing 4T1 murine mammary adenocarcinoma-bearing mice. This approach of co-delivery of two complementary systems serves as a proof-of-concept for real-time deep tissue intravital orthotopic monitoring and may have the potential use in clinical utility as a theranostic nanomedicine.

Tumor targeting profiling of hyaluronan-coated lipid based-nanoparticles.

Hyaluronan (HA), a naturally occurring high Mw (HMw) glycosaminoglycan, has been shown to play crucial roles in cell growth, embryonic development, healing processes, inflammation, and tumor development and progression. Low Mw (LMw, <10 kDa) HA has been reported to provoke inflammatory responses, such as induction of cytokines, chemokines, reactive nitrogen species and growth factors. Herein, we prepared and characterized two types of HA coated (LMw and HMw) lipid-based targeted and stabilized nanoparticles (tsNPs) and tested their binding to tumor cells expressing the HA receptor (CD44), systemic immunotoxicity, and biodistribution in tumor bearing mice. In vitro, the Mw of the surface anchored HA had a significant influence on the affinity towards CD44 on B16F10 murine melanoma cells. LMw HA-tsNPs exhibited weak binding, while binding of tsNPs coated with HMw HA was characterized by high binding. Both types of tsNPs had no measured effect on cytokine induction in vivo following intravenous administration to healthy C57BL/6 mice suggesting no immune activation. HMw HA-tsNPs showed enhanced circulation time and tumor targeting specificity, mainly by accumulating in the tumor and its vicinity compared with LMw HA-tsNPs. Finally, we show that methotrexate (MTX), a drug commonly used in cancer chemotherapy, entrapped in HMw HA-tsNPs slowly diffused from the particles with a half-life of 13.75 days, and improved the therapeutic outcome in a murine B16F10 melanoma model compared with NPs suggesting an active cellular targeting beyond the Enhanced Permeability and Retention (EPR) effect. Taken together, these findings have major implications for the use of high molecular weight HA in nanomedicine as a selective and safe active cellular targeting moiety.

Modular theranostic prodrug based on a FRET-activated self-immolative linker.

New chemotherapeutic prodrugs that can report on the localization and activation of the drug upon internalization into select cells are being widely investigated for cancer treatment. Here, we introduce a new molecular design for a theranostic prodrug based on a self-immolative linker attached to a pair of FRET dyes that produces a fluorescent signal upon disassembly. The prodrug evaluated here was designed to release the chemotherapeutic drug camptothecin upon activation by the model enzyme penicillin-G-amidase. Similar patterns of the disassembly of the prodrug were observed in HPLC and fluorescence assays. The obtained results demonstrate that upon specific activation of the prodrug, the increase of the emitted fluorescent signal is linearly correlated with the observed drug release. Such a design could potentially be used to monitor prodrug activation in real-time and provide information regarding the location and the amount of active drug molecules.

A simple FRET-based modular design for diagnostic probes.

In recent years, there has been a massive effort to develop molecular probes with optical modes of action. Probes generally produce detectable signals based on changes in fluorescence properties. Here, we demonstrate the potential of self-immolative molecular adaptors as a platform for Turn-On probes based on the FRET technique. The probe is equipped with identical fluorophore pairs or a fluorophore/quencher FRET pair and a triggering substrate. Upon reaction of the analyte of interest with the triggering substrate, the self-immolative adaptor spontaneously releases the two dye molecules to break off the FRET effect. As a result, a new measurable fluorescent signal is generated. The fluorescence obtained can be used to quantify the analyte. The modular structure of the probe design will allow the preparation of various chemical probes based on the FRET activation technique.