Atomic force microscopy probing of receptor-nanoparticle interactions for riboflavin receptor targeted gold-dendrimer nanocomposites.

Riboflavin receptors are overexpressed in malignant cells from certain human breast and prostate cancers, and they constitute a group of potential surface markers important for cancer targeted delivery of therapeutic agents and imaging molecules. Here we report on the fabrication and atomic force microscopy (AFM) characterization of a core-shell nanocomposite consisting of a gold nanoparticle (AuNP) coated with riboflavin receptor-targeting poly(amido amine) dendrimer. We designed this nanocomposite for potential applications such as a cancer targeted imaging material based on its surface plasmon resonance properties conferred by AuNP. We employed AFM as a technique for probing the binding interaction between the nanocomposite and riboflavin binding protein (RfBP) in solution. AFM enabled precise measurement of the AuNP height distribution before (13.5 nm) and after chemisorption of riboflavin-conjugated dendrimer (AuNP-dendrimer; 20.5 nm). Binding of RfBP to the AuNP-dendrimer caused a height increase to 26.7 nm, which decreased to 22.8 nm when coincubated with riboflavin as a competitive ligand, supporting interaction of AuNP-dendrimer and its target protein. In summary, physical determination of size distribution by AFM imaging can serve as a quantitative approach to monitor and characterize the nanoscale interaction between a dendrimer-covered AuNP and target protein molecules in vitro.

Design and mechanistic investigation of oxime-conjugated PAMAM dendrimers as the catalytic scavenger of reactive organophosphate.

Pyridinium aldoxime (PAM) molecules constitute a group of small molecule antidotes essential for the treatment of reactive organophosphate (OP) poisoning. Their therapeutic efficacy stems from a combination of two activities: (i) reactivation of acetylcholine esterase inhibited by OP; (ii) scavenging of free OP. Here, we report the design, synthesis and in vitro functional characterization of fifth generation poly(amidoamine)dendrimer conjugates, each tethered with PAM or hydroxamate, as OP scavenging macromolecules. We chose paraoxon (POX) as the model OP, and performed extensive reaction kinetic studies in aqueous media to determine the activities and mechanisms of these dendrimer conjugates in POX hydrolysis with 1H NMR, UV-vis and LCMS/MS methods. Quantitative kinetic analysis suggests that the conjugate hydrolyzed POX through a catalytic mechanism as effective as that of the unconjugated molecules. In summary, we report the first class of dendrimer conjugates which are stable in the plasma and functioning as OP-responsive catalytic scavengers.

Design and in vitro validation of multivalent dendrimer methotrexates as a folate-targeting anticancer therapeutic.

Design of cancer-targeting nanotherapeutics relies on a pair of two functionally orthogonal molecules, one serving as a cancer cell-specific targeting ligand, and the other as a therapeutic cytotoxic agent. The present study investigates the validity of an alternative simplified strategy where a dual-acting molecule which bears both targeting and cytotoxic activity is conjugated to the nanoparticle as cancer-targeting nanotherapeutics. Herein, we demonstrate that methotrexate is applicable for this dual-acting strategy due to its reasonable affinity to folic acid receptor (FAR) as a tumor biomarker, and cytotoxic inhibitory activity of cytosolic dihydrofolate reductase. This article describes design of new methotrexate-conjugated poly(amidoamine) (PAMAM) dendrimers, each carrying multiple copies of methotrexate attached through a stable amide linker. We evaluated their dual biological activities by performing surface plasmon resonance spectroscopy, a cell-free enzyme assay and cell-based experiments in FAR-overexpressing cells. This study identifies the combination of an optimal linker framework and multivalency as the two key design elements that contribute to achieving potent dual activity.

Synthesis of a photocaged tamoxifen for light-dependent activation of Cre-ER recombinase-driven gene modification.

We report the design of a water-soluble, quaternized tamoxifen photoprobe and demonstrate its application in light-controlled induction of green fluorescent protein expression via a Cre-ER recombinase system.