Dataset Information

Click Nucleic Acid Mediated Loading of Prodrug Activating Enzymes in PEG-PLGA Nanoparticles for Combination Chemotherapy.

ABSTRACT: The simultaneous delivery of multiple therapeutics to a single site has shown promise for cancer targeting and treatment. However, because of the inherent differences in charge and size between drugs and biomolecules, new approaches are required for colocalization of unlike components in one delivery vehicle. In this work, we demonstrate that triblock copolymers containing click nucleic acids (CNAs) can be used to simultaneously load a prodrug enzyme (cytosine deaminase, CodA) and a chemotherapy drug (doxorubicin, DOX) in a single polymer nanoparticle. CNAs are synthetic analogs of DNA comprised of a thiolene backbone and nucleotide bases that can hybridize to complementary strands of DNA. In this study, CodA was appended with complementary DNA sequences and fluorescent dyes to allow its encapsulation in PEG-CNA-PLGA nanoparticles. The DNA-modified CodA was found to retain its enzyme activity for converting prodrug 5-fluorocytosine (5-FC) to active 5-fluorouracil (5-FU) using a modified fluorescent assay. The DNA-conjugated CodA was then loaded into the PEG-CNA-PLGA nanoparticles and tested for cell cytotoxicity in the presence of the 5-FC prodrug. To study the effect of coloading DOX and CodA within a single nanoparticle, cell toxicity assays were run to compare dually loaded nanoparticles with nanoparticles loaded only with either DOX or CodA. We show that the highest level of cell death occurred when both DOX and CodA were simultaneously entrapped and delivered to cells in the presence of 5-FC.

SUBMITTER: Harguindey A

PROVIDER: S-EPMC6697549 | biostudies-literature | 2019 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:Understanding the mechanistic basis of prodrug delivery and activation is critical for establishing species-specific prodrug sensitivities necessary for evaluating preclinical animal models and potential drug-drug interactions. Despite significant adoption of prodrug methodologies for enhanced pharmacokinetics, functional annotation of prodrug activating enzymes is laborious and often unaddressed. Activity-based protein profiling (ABPP) describes an emerging chemoproteomic approach to assay active site occupancy within a mechanistically similar enzyme class in native proteomes. The serine hydrolase enzyme family is broadly reactive with reporter-linked fluorophosphonates, which have shown to provide a mechanism-based covalent labeling strategy to assay the activation state and active site occupancy of cellular serine amidases, esterases, and thioesterases. Here we describe a modified ABPP approach using direct substrate competition to identify activating enzymes for an ethyl ester prodrug, the influenza neuraminidase inhibitor oseltamivir. Substrate-competitive ABPP analysis identified carboxylesterase 1 (CES1) as an oseltamivir-activating enzyme in intestinal cell homogenates. Saturating concentrations of oseltamivir lead to a four-fold reduction in the observed rate constant for CES1 inactivation by fluorophosphonates. WWL50, a reported carbamate inhibitor of mouse CES1, blocked oseltamivir hydrolysis activity in human cell homogenates, confirming CES1 is the primary prodrug activating enzyme for oseltamivir in human liver and intestinal cell lines. The related carbamate inhibitor WWL79 inhibited mouse but not human CES1, providing a series of probes for analyzing prodrug activation mechanisms in different preclinical models. Overall, we present a substrate-competitive activity-based profiling approach for broadly surveying candidate prodrug hydrolyzing enzymes and outline the kinetic parameters for activating enzyme discovery, ester prodrug design, and preclinical development of ester prodrugs.

Dataset Information

Click Nucleic Acid Mediated Loading of Prodrug Activating Enzymes in PEG-PLGA Nanoparticles for Combination Chemotherapy.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets