Stimulating Macropinocytosis for Intracellular Nucleic Acid and Protein Delivery: A Combined Strategy with Membrane-Lytic Peptides To Facilitate Endosomal Escape

Abstract

Delivery of biomacromolecules via endocytic pathways requires the efficient accumulation of cargo molecules into endosomes, followed by their release to the cytosol. We propose a unique intracellular delivery strategy for bioactive molecules using a new potent macropinocytosisinducing peptide derived from stromal-derived factor (SDF)-1α (SN21). This peptide allowed extracellular materials to enter cells through the activation of macropinocytosis. To provide the ability to release internalized cargoes from endosomes, we conjugated SN21 with membranelytic peptides. The combination of a macropinocytosis-inducing peptide and a membrane lytic peptide successfully delivered functional siRNA and proteins, which include antibodies, Cre recombinase, and an artificial transcription regulator protein having a transcription activator-like effector (TALE) motif. This study shows the feasibility of combining physiological stimulation of macropinocytosis with physicochemical disruption of endosomes as a strategy for intracellular delivery.

The Dual Role of Macropinocytosis in Cancers: Promoting Growth and Inducing Methuosis to Participate in Anticancer Therapies as Targets

“…These two phenomena have major contributions to the extreme vacuolation of cancer cells. Some scholars believe that the emergence of extreme vacuolation led to the death of cancer cells ( 128 , 140 ). However, more detailed, and specific evidence linking macropinocytosis directly to cell death is lacking.…”

“…The key players in this process are Arf and Rab GTPases ( 94 , 126 ). Over time, continuous high expressing of RAS (G12V) with subsequent slow stimulating of Rac1 causes extreme vacuolation of tumor cells, eventually leading to methuosis ( 128 ). One step that takes effect in methuosis is that the activity of macropinosomes recycled to the plasma membrane surface is reduced during the over-activated macropinocytosis process.…”

“…Heptapeptide R7, a novel peptide, was synthesized to promote internalization of chlorin e6(a photosensitizer) into HepG2 cells by activation of endocytosis and/or macropinocytosis in comparison with initial NPs without peptide R7 ( 168 ). SN21, a peptide based on stromal-derived factor 1α, can induce macropinocytosis and deliver functional siRNA and proteins such as antibodies and Cre recombinases ( 128 ).…”

“…SN21 was combined with membrane cleavage peptides by Arafiles et al. to realize the internalization through macropinocytosis and release of endosomes of extracellular materials ( 128 ). This strategy has been shown to be effective in promoting cellular uptake of anticancer drugs.…”

Membrane Trafficking and Subcellular Drug Targeting Pathways

“…Another strategy describes conjugation of stromal-derived factor 1a (SN 21 ) with membrane lytic peptides which induces micropinocytosis or membrane lysis and aids in the successful delivery of functional siRNAs and proteins. It also gives the loaded drug cargo the ability to be released from endosomes or endosomal escape (Arafiles et al, 2020). Fraire et al, describe the escape from endosomes in terms of endosomal rupture through the photothermal properties of gold nanoparticles by vapor bubble formation and heat induction.…”