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Background: Elevated glutamate production and release from glial cells is a common feature of many CNS disorders. Inhibitors of glutaminase (GLS), the enzyme responsible for converting glutamine to glutamate have been developed to target glutamate overproduction. However, many GLS inhibitors have poor aqueous solubility, are unable to cross the blood brain barrier, or demonstrate significant toxicity when given systemically, precluding translation. Enhanced aqueous solubility and systemic therapy targeted to activated glia may address this challenge. Here we examine the impact of microglial-targeted GLS inhibition in a mouse model of Rett syndrome (RTT), a developmental disorder with no viable therapies, manifesting profound central nervous system effects, in which elevated glutamatergic tone, upregulation of microglial GLS, oxidative stress and neuroimmune dysregulation are key features. Methods: To enable this, we conjugated a potent glutaminase inhibitor, N-(5-{2-[2-(5-amino-[1,3,4]thiadiazol-2-yl)-ethylsulfanyl]-ethyl}-[1,3,4]thiadiazol-2-yl)-2-phenyl-acetamide (JHU29) to a generation 4 hydroxyl PAMAM dendrimer (D-JHU29). We then examined the effect of D-JHU29 in organotypic slice culture on glutamate release. We also examined GLS activity in microglial and non-microglial cells, and neurobehavioral phenotype after systemic administration of D-JHU29 in a mouse model of RTT. Results: We report successful conjugation of JHU29 to dendrimer resulting in enhanced water solubility compared to free JHU29. D-JHU29 reduced the excessive glutamate release observed in tissue culture slices in a clinically relevant Mecp2-knockout (KO) RTT mouse. Microglia isolated from Mecp2-KO mice demonstrated upregulation of GLS activity that normalized to wild-type levels following systemic treatment with D-JHU29. Neurobehavioral assessments in D-JHU29 treated Mecp2-KO mice revealed selective improvements in mobility. Conclusion: These findings demonstrate that glutaminase inhibitors conjugated to dendrimers are a viable mechanism to selectively inhibit microglial GLS to reduce glutamate production and improve mobility in a mouse model of RTT, with broader implications for selectively targeting this pathway in other neurodegenerative disorders.

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A combination of donor-acceptor and electrostatic interactions in a three-component supramolecular system has been shown to form the basis for selective and sensitive detection of peptides. Different substituents in the polymer and the detection matrix were compared to demonstrate that the favorable donor-acceptor interactions explain the observed signal enhancement. The ternary supramolecular interactions discovered in this work are enabled by the self-packing behavior of amphiphilic homopolymers and their ability to mediate interactions between the detection matrix and peptide that facilitate sensitive detection of peptides.

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Zwitterionic amphiphilic homopolymers can be conveniently prepared in one-pot using activated ester-based polymer precursors. We show that these zwitterionic polymers can (i) spontaneously self-assemble to form micelle-like and inverse micelle-like assemblies depending on the solvent environment; (ii) act as hydrophilic and hydrophobic nanocontainers in apolar and polar solvents respectively; (iii) undergo pH-responsive surface charge and size variations; (iv) exhibit least cytotoxicity compared to structurally analogous amphiphilic homopolymers.

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Supramolecular nano-assemblies that reduce nonspecific interactions with biological macromolecules, such as proteins, are of great importance for various biological applications. Recently, zwitterionic materials have been shown to reduce nonspecific interactions with biomolecules, owing both to their charge neutrality and their ability to form a strong hydration layer around zwitterions via electrostatic interactions. Here, new triazole-based zwitterionic moieties are presented that are incorporated as the hydrophilic functionalities in facially amphiphilic dendrons. The amphiphilic zwitterionic dendrons spontaneously self-assemble in aqueous solutions forming micelle-type aggregates, which were confirmed by DLS, TEM, and fluorescence techniques. The structural and functional characteristics of the zwitterionic dendrons are also compared with the corresponding charge-neutral PEG-based dendrons and anionic carboxylate-based dendrons. Surface-charge measurements, temperature sensitivity and evaluation of interactions of these assemblies with proteins form the bases for these comparisons.

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Supramolecular assemblies formed by amphiphilic homopolymers with negatively charged groups in the hydrophilic segment have been designed to enable high labeling selectivity toward reactive side chain functional groups in peptides. The negatively charged interiors of the supramolecular assemblies are found to block the reactivity of protonated amines that would otherwise be reactive in aqueous solution, while maintaining the reactivity of nonprotonated amines. Simple changes to the pH of the assemblies' interiors allow control over the reactivity of different functional groups in a manner that is dependent on the pKa of a given peptide functional group. The labeling studies carried out in positively charged supramolecular assemblies and free buffer solution show that, even when the amine is protonated, labeling selectivity exists only when complementary electrostatic interactions are present, thereby demonstrating the electrostatically controlled nature of these reactions.

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Oligo(ethylene glycol)-decorated supramolecular assemblies have been of great interest due to their charge-neutral character and thus their propensity to avoid nonspecific interactions. These systems are known to exhibit a macroscopic temperature-sensitive transition, where the assembly phase-separates from the aqueous phase at higher temperatures. While this so-called lower critical solution temperature (LCST) behavior has been well-studied, there have been no studies on the fate of these supramolecular assemblies below this transition temperature. The work here brings to light the presence of a second, sub-LCST transition, observed well below the LCST of oligo(ethylene glycol) (OEG)-based dendrons, where the host-guest properties of the assembly are significantly altered. This sub-LCST transition is accompanied by changes in the guest encapsulation stability and dynamics of host exchange.

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