Dendritic polymer nanoparticles (NPs) are promising vehicles for drug delivery. Most dendrimer polymer NPs, however, exhibit positive surface charge which make them, in many instances, cytotoxic. We constructed noncationic, amphiphilic dendrimer NPs embedding curcumin and resveratrol, natural polyphenols exhibiting anticancer properties. The curcumin/resveratrol/dendrimer NPs both effectively shielded the embedded polyphenols and facilitated their slow release and, notably, targeted cancer cells. The experimental data trace the cancer cell toxicity of the curcumin/resveratrol/dendrimer NPs to impairment of mitochondrial functions, specifically giving rise to enhanced intracellular calcium release, inhibition of cytochrome c oxidase enzyme activity, decreased mitochondrial membrane potential, and mitochondrial membrane perturbation. Importantly, synergism between the dendrimer-NP-embedded curcumin and resveratrol was observed, as more pronounced cancer cell death and mitochondrial disruption were induced by the curcumin/resveratrol/dendrimer NPs as compared to either the freely dissolved polyphenols or amphiphilic dendrimer NPs incorporating curcumin or resveratrol separately. This work suggests that amphiphilic dendrimer NPs encapsulating curcumin and resveratrol may constitute a promising anticancer therapeutic platform.

Although myriad protein–protein interactions in nature use polyvalent binding, in which multiple ligands on one entity bind to multiple receptors on another, to date an affinity advantage of polyvalent binding has been demonstrated experimentally only in cases where the target receptor molecules are clustered prior to complex formation. Here, we demonstrate cooperativity in binding affinity (i.e., avidity) for a protein complex in which an engineered dimer of the amyloid precursor protein inhibitor (APPI), possessing two fully functional inhibitory loops, interacts with mesotrypsin, a soluble monomeric protein that does not self-associate or cluster spontaneously. We found that each inhibitory loop of the purified APPI homodimer was over three-fold more potent than the corresponding loop in the monovalent APPI inhibitor. This observation is consistent with a suggested mechanism whereby the two APPI loops in the homodimer simultaneously and reversibly bind two corresponding mesotrypsin monomers to mediate mesotrypsin dimerization. We propose a simple model for such dimerization that quantitatively explains the observed cooperativity in binding affinity. Binding cooperativity in this system reveals that the valency of ligands may affect avidity in protein–protein interactions including those of targets that are not surface-anchored and do not self-associate spontaneously. In this scenario, avidity may be explained by the enhanced concentration of ligand binding sites in proximity to the monomeric target, which may favor rebinding of the multiple ligand binding sites with the receptor molecules upon dissociation of the protein complex.

It is currently believed that molecular agents that specifically bind to and neutralize the toxic proteins/peptides, amyloid β (Aβ42), tau, and the tau-derived peptide PHF6, hold the key to attenuating the progression of Alzheimer's disease (AD). We thus tested our previously developed nonaggregating Aβ42 double mutant (Aβ42 DM ) as a multispecific binder for three ADassociated molecules, wild-type Aβ42, the tau K174Q mutant, and a synthetic PHF6 peptide. Aβ42 DM acted as a functional inhibitor of these molecules in in vitro assays and in neuronal cell-based models of AD. The double mutant bound both cytotoxic tau K174Q and synthetic PHF6 and protected neuronal cells from the accumulation of tau in cell lysates and mitochondria. Aβ42 DM also reduced toxic intracellular levels of calcium and the overall cell toxicity induced by overexpressed tau, synthetic PHF6, Aβ42, or a combination of PHF6and Aβ42. Aβ42 DM inhibited PHF6-induced overall mitochondrial dysfunction: In particular, Aβ42 DM inhibited PHF6-induced damage to submitochondrial particles (SMPs) and suppressed PHF6-induced elevation of the ζ-potential of inverted SMPs (proxy for the inner mitochondrial membrane, IMM). PHF6 reduced the lipid fluidity of cardiolipin/DOPC vesicles (that mimic the IMM) but not DOPC (which mimics the outer mitochondrial membrane), and this effect was inhibited by Aβ42 DM . This inhibition may be explained by the conformational changes in PHF6 induced by Aβ42 DM in solution and in membrane mimetics. On this basis, the paper presents a mechanistic explanation for the inhibitory activity of Aβ42 DM against Aβ42-and tauinduced membrane permeability and cell toxicity and provides confirmatory evidence for its protective function in neuronal cells.

Although myriad protein–protein interactions in nature use polyvalent binding, in which multiple ligands on one entity bind to multiple receptors on another, to date an affinity advantage of polyvalent binding has been demonstrated experimentally only in cases where the target receptor molecules are clustered prior to complex formation. Here, we demonstrate cooperativity in binding affinity ( i.e ., avidity) for a protein complex in which an engineered dimer of the amyloid precursor protein inhibitor (APPI), possessing two fully functional inhibitory loops, interacts with mesotrypsin, a soluble monomeric protein that does not self-associate or cluster spontaneously. We found that each inhibitory loop of the purified APPI homodimer was over three-fold more potent than the corresponding loop in the monovalent APPI inhibitor. This observation is consistent with a suggested mechanism whereby the two APPI loops in the homodimer simultaneously and reversibly bind two corresponding mesotrypsin monomers to mediate mesotrypsin dimerization. We propose a simple model for such dimerization that quantitatively explains the observed cooperativity in binding affinity. Binding cooperativity in this system reveals that the valency of ligands may affect avidity in protein–protein interactions including those of targets that are not surface-anchored and do not self-associate spontaneously. In this scenario, avidity may be explained by the enhanced concentration of ligand binding sites in proximity to the monomeric target, which may favor rebinding of the multiple ligand binding sites with the receptor molecules upon dissociation of the protein complex.