(674c) Mitochondrial Targeting with Triphenylphosphonium (TPP)-Conjugated and TPP-Peg-Conjugated PAMAM Dendrimer Nanocarriers

Authors: 
Bielski, E., Wayne State University
Zhong, Q., Wayne State University
Yang, L., Wayne State University
Brown, M., Wayne State University
Cabral, V., State University of Maringá
da Rocha, S. R. P., Wayne State University

Mitochondria play vital roles in energy production within cells, are key to eukaryotic cell survival, in the generation of reactive oxygen species, in the regulation of calcium homeostasis, metabolism, and of programmed cell death (intrinsic apoptosis).  Mitochondrial dysfunction is related to a range of diseases including heart disease, diabetes, cancer, neurodegenerative diseases, neuromuscular disorders, and aging.  Their crucial role in apoptosis, metabolism, and energy production make these organelles a crucial target for many mitochondria-targeted drug therapies that can modulate the defects associated with the mitochondria and mitochondrial signaling pathways or induce apoptosis for cancer treatment strategies.

Among the many polymeric nanocarriers, Poly(amido amine) (PAMAM) dendrimers  are a promising nanocarriers for the delivery of therapeutics including in the treatment of mitochondrial-related diseases.  PAMAM dendrimers are polymeric hyperbranched tree-like structures of nanometer sizes comprising of a central core, repeated branches increasing with each generation, with a large number of surface functional groups that can be used to conjugate therapeutics for controlled release and other functionalities to enhance tissue/cellular/intracellular targeting.  Polyethylene glycol (PEG) has also been extensively used to modulate the properties of dendrimers including improving hydrophobicity, decreasing toxicity, protecting therapeutic cargo, improving circulation time of dendrimers, and becoming a flexible linker to targeting moieties, such as mitochondrial targeting groups.  Delocalized lipophilic cations (DLCs), such as triphenylphosphonium cations (TPP), are the widely used mitochondrial targeting agents due to their delocalized positive charge, which allows them to pass through plasma and mitochondrial membranes in a potential driven process, showing 100-1000 fold selective accumulation into the cell cytosol and mitochondrial compartments.  Therefore, the use of DLCs such as TPP may offer unique strategies to deliver numerous therapeutic agents preferentially to mitochondria within cells, and can thus be very effective in maximizing drug effects while reducing side effects.  The combination of DLCs and PEG with drug nanocarriers, such as PAMAM dendrimers, may thus afford unique opportunities for the treatment of mitochondrial-related diseases.

In this work, we report i) the synthesis and characterization of TPP and TPP-PEG conjugated to Generation 4 PAMAM amine terminated dendrimers (G4NH2), which are potential nanocarriers of therapeutic agents, ii) the cellular internalization and colocalization of such nanocarriers in A549 alveolar epithelial adenocarcinomal cell line using flow cytometry and confocal microscopy, respectively, and iii) surface properties of TPP and TPP-PEG conjugated PAMAM dendrimers (G4NH2-TPP and G4NH2-PEG-TPP) and their microstructure.  We systematically investigate the rate of internalization and colocalization of the conjugates as a function of TPP density with or without PEG.

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