Supplementary Materials1_si_001. various cancers, especially ovarian, breast, and non-small cell lung cancers for the past decade.21, 22 As a mitotic inhibitor, Paclitaxel exerts its cytotoxic effect by arresting mitosis through microtubule stabilization and subsequently interfering with the normal breakdown of microtubules during cell division, resulting in cellular apoptosis.22, 23 Although Paclitaxel has had a significant impact in cancer treatment, the effectiveness of traditional Paclitaxel-based treatment regimens has been limited due to the drugs low aqueous solubility, eventual acquired chemoresistance of cells, and certain side effects.24 Its commercial formulation, Taxol? (Bristol-Myers Squibb), is formulated in a high concentration in Cremophor EL, a solvent and excipient material that is necessary to solubilize Paclitaxel but can cause severe side effects, including hypersensitivity reactions, nephrotoxicity, myelosuppression, and peripheral neuropathy.25, 26 The emerging macroparticle or nanoparticle-based delivery approaches have provided interesting avenues for improving cancer treatments by the controlled delivery and release of chemotherapeutic agents.27C37 Among them, several strategies for increasing Paclitaxel solubility and efficacy have been studied, such as the use of emulsification, micellization, liposomes, non-liposomal carriers (microspheres, nanoparticles), cyclodextrins, and local drug delivery devices.25 Most of these delivery strategies are focused upon encapsulation methods,31, 38 which have been shown to yield particles with high loading and relatively large size (100 nm C micron size).39C41 Interestingly, drug-nanoparticle hybrid conjugates have recently garnered significant attention.27, 42 The smaller structures ( 100nm) made this method are more stable, can evade reticuloendothelial system (RES) capture and exhibit the ability to accumulate in a broader range of tumors creating a necessity for nano-sized ACY-1215 cost delivery vehicles.42C45 Most nanoparticle-drug conjugates have used oligoethylene glycol (OEG)-based linkers, which do not offer the versatile chemistry afforded by ACY-1215 cost synthetically and mechanically programmable oligonucleotide linkers. As such, facile methods for labeling OEG-drug conjugates for visualization and tracking within cells do not yet exist. Finally, it is important to note that OEG-functionalized particles exhibit a three orders of magnitude lower cellular uptake than antisense DNA-AuNPs.8 In the study reported herein, paclitaxel molecules are covalently attached to gold nanoparticles fluorescent antisense oligonucleotide linkers, resulting in a multimodal drug delivery system with simultaneous capabilities in imaging and enhanced therapy. The potency of the resulting conjugates was also tested within a chemoresistant cell line. The data suggest that these conjugates may solve three common problems ACY-1215 cost associated with Paclitaxel as an effective chemotherapeutic agent. First, they enhance the solubility of the drug in aqueous systems such as buffers containing high concentrations of salts and serum-containing cell culture media. Second, they increase drug efficacy in Paclitaxel-resistant cell lines. Third, they provide a useful way of tracking the movement of the conjugates and delivery of the drug. Indeed, these nanoparticle conjugates form the basis for a new strategy for generally increasing drug solubility and efficacy while simultaneously tracking intracellular uptake. RESULTS AND DISCUSSION Nanoparticle conjugates were prepared by reacting citrate-stabilized gold nanoparticles with propylthiol-capped 20-mer poly-dT oligonucleotides containing a terminal Paclitaxel group (Scheme 1). First, DNA oligomers were synthesized with a terminal amine group for covalent attachment to Paclitaxel. Paclitaxel was modified by reaction with succinic anhydride to create a Paclitaxel carboxilic acid derivative, compound 1. The amine-terminated DNA and the Paclitaxel carboxilic acid derivative 1 were covalently attached to each other EDC/Sulfo-NHS coupling chemistry to yield compound 2 (Scheme 1). After purification by RP-HPLC, the Mouse monoclonal to FCER2 Paclitaxel-DNA (PTX-DNA), compound 2 was characterized by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS), which confirmed its formation (Figure 1). Compound 2 was then immobilized on citrate-stabilized AuNPs using literature procedures for making analogous DNA-AuNPs,46 ultimately yielding the Paclitaxel-DNA gold nanoparticle conjugates (PTX-DNA@AuNPs) 3. Excess PTX-DNA was removed through repeated centrifugation and resuspension of PTX-DNA@AuNPs until no PTX-DNA was detected by MALDI-MS in the supernatant. Fluorescein-labeled PTX-DNA conjugates were synthesized as described in Scheme 1 in order to.