Dual Function MITO-Porter, a Nano Carrier Integrating Both Efficient Cytoplasmic Delivery and Mitochondrial Macromolecule Delivery

Abstract

Mitochondrial dysfunction is associated with a variety of human diseases including inherited mitochondrial diseases, neurodegenerative disorders, diabetes mellitus, and cancer. Effective medical therapies for mitochondrial diseases will ultimately require an optimal drug delivery system, which will likely be achieved through innovations in the nanotechnology of intracellular trafficking. To achieve efficient mitochondrial drug delivery, two independent processes, i.e., "cytoplasmic delivery through the cell membrane" and "mitochondrial delivery through the mitochondrial membrane" are required. In previous studies, we developed an octaarginine (R8) modified nano carrier for efficient cytoplasmic delivery, showing that R8-modified liposomes were internalized into cells efficiently. On the other hand, we also constructed MITO-Porter for the mitochondrial delivery of macromolecules, a liposome-based carrier that delivers cargos to mitochondria via membrane fusion. Here, we report the development of a dual function MITO-Porter (DF-MITO-Porter), based on the concept of integrating both R8-modified liposomes and MITO-Porter. We show that the DF-MITO-Porter effectively delivers exogenous macro-biomolecules into the mitochondrial matrix, and provide a demonstration of its potential use in therapies aimed at mitochondrial DNA.

Enhancement in selective mitochondrial association by direct modification of a mitochondrial targeting signal peptide on a liposomal based nanocarrier

“…Intracellular observations showed that the DF-MITO-Porter effectively delivered exogenous bioactive macromolecules into the mitochondria in living cells (Yamada et al, 2011;. Future study will include the packaging of MTS-modified MITO-Porter via endosome-fusogenic envelopes to achieve mitochondrial selective targeting in living cells using DF-MTS-modified MITO-Porter.…”

“…In previous studies, we developed a Dual Function (DF)-MITO-Porter, which possesses mitochondriafusogenic inner envelopes and endosome-fusogenic outer envelopes (Yamada et al, 2011;. Intracellular observations showed that the DF-MITO-Porter effectively delivered exogenous bioactive macromolecules into the mitochondria in living cells (Yamada et al, 2011;.…”

Delivery of bioactive molecules to the mitochondrial genome using a membrane-fusing, liposome-based carrier, DF-MITO-Porter

“…The DF-MITO-Porter encapsulating DNase I was constructed as described in our previous reports (see Figure S1 in supplementary data for details) [21]. Complexed DNase I particles were first prepared with STR-R8, and were then coated with a mitochondria-fusogenic envelope.…”

“…The DF-MITO-Porter, which possesses mitochondria-fusogenic inner and endosome-fusogenic outer envelopes, has the ability to pass through endosomal and mitochondrial membranes via step-wise membrane fusion ( Figure 1) [21]. Octaarginine (R8) functions as a cell-uptake device [22,23] in the outer envelope and as a mitochondrial targeting device [12,24,25] in the inner envelope.…”

“…The DNase I-encapsulated Dual Function (DF)-MITO-Porter was constructed by the multi-layering method, as previously reported (see Figure S1 in supplementary data for details) [21]. DNase I protein was gently mixed with the STR-R8, as complex-inducer, in 100 μL of 10 mM HEPES buffer (pH 7. was added to the suspension of T-SNP to modify the outer envelope with R8.…”