Direct entry of cell-penetrating peptide can be controlled by maneuvering the membrane curvature

AbstractA biomembrane's role is to be a barrier for interior cytosol from an exterior environment to execute the cell's normal biological functions. However, a water-soluble peptide called cell-penetrating peptide (CPP) has been known for its ability to directly penetrate through the biomembranes into cells (cytolysis) without perturbating cell viability and expected to be a promising drug delivery vector. Examples of CPP include peptides with multiple arginine units with strong cationic properties, which is the key to cytolysis. Here we show the conclusive evidence to support the mechanism of CPP’s cytolysis and way to control it. The mechanism we proposed is attributed to biomembrane’s physicochemical nature as lamellar liquid crystal (Lα). Cytolysis occurs as the temporal and local dynamic phase transitions from Lα to an undulated lamellar with pores called Mesh1. We have shown this phase transfer of Lα composed of dioleoyl-phosphatidylcholine (DOPC) with water by adding oligo-arginine (Rx) as CPP at the equilibrium. Using giant unilamellar vesicle composed of DOPC as a single cell model, we could control the level of cytolysis of CPP (FITC-R8) by changing the curvature of the membrane through osmotic pressure modulation. The cytolysis of CPP utilizes biomembrane's inherent topological and functional flexibility corresponding to the stimuli.

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Supramolecular intracellular delivery of an anionic porphyrin by octaarginine-conjugated per-O-methyl-β-cyclodextrin

Chemical Communications ◽

10.1039/c4cc09042j ◽

2015 ◽

Vol 51 (12) ◽

pp. 2421-2424 ◽

Cited By ~ 24

Author(s):

Hiroaki Kitagishi ◽

Fumihiko Chai ◽

Shigeru Negi ◽

Yukio Sugiura ◽

Koji Kano

Keyword(s):

Efficient Method ◽

Intracellular Delivery ◽

Water Soluble ◽

Cell Penetrating Peptide ◽

Supramolecular Interaction ◽

Cell Penetrating ◽

A Cell

A convenient and efficient method for intracellular delivery of a water-soluble anionic porphyrin has been developed by utilizing its supramolecular interaction with per-O-methyl-β-cyclodextrin bearing an octaarginine chain as a cell-penetrating peptide.

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Cell Penetrating Peptide as a High Safety Anti-Inflammation Ingredient for Cosmetic Applications

Biomolecules ◽

10.3390/biom10010101 ◽

2020 ◽

Vol 10 (1) ◽

pp. 101 ◽

Cited By ~ 1

Author(s):

Tse-Kai Fu ◽

Ping-Hsueh Kuo ◽

Yen-Chang Lu ◽

Hsing-Ni Lin ◽

Lily Hui-Ching Wang ◽

...

Keyword(s):

Cell Model ◽

Skin Barrier ◽

Skin Condition ◽

Cell Penetrating Peptide ◽

Market Demand ◽

Model Following ◽

Cell Penetrating ◽

Oil Formulation ◽

A Cell ◽

Anti Inflammation

Cosmeceutical peptides have become an important topic in recent decades in both academic and industrial fields. Many natural or synthetic peptides with different biological functions including anti-ageing, anti-oxidation, anti-infection and anti-pigmentation have been developed and commercialized. Current cosmeceutical peptides have already satisfied most market demand, remaining: “cargos carrying skin penetrating peptide with high safety” still an un-met need. To this aim, a cell-penetrating peptide, CPPAIF, which efficiently transported cargos into epithelial cells was exanimated. CPPAIF was evaluated with cell model and 3D skin model following OECD guidelines without using animal models. As a highly stable peptide, CPPAIF neither irritated nor sensitized skin, also did not disrupt skin barrier. In addition, such high safety peptide had anti-inflammation activity without allergic effect. Moreover, cargo carrying activity of CPPAIF was assayed using HaCaT cell model and rapid CPPAIF penetration was observed within 30 min. Finally, CPPAIF possessed transepidermal activity in water in oil formulation without disruption of skin barrier. All evidences indicated that CPPAIF was an ideal choice for skin penetrating and its anti-inflammatory activity could improve skin condition, which made CPPAIF suitable and attractive for novel cosmeceutical product development.

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Effect of Vesicle Size on the Cytolysis of Cell-Penetrating Peptides (CPPs)

International Journal of Molecular Sciences ◽

10.3390/ijms21197405 ◽

2020 ◽

Vol 21 (19) ◽

pp. 7405

Author(s):

Kazutami Sakamoto ◽

Takeshi Kitano ◽

Haruka Kuwahara ◽

Megumi Tedani ◽

Kenichi Aburai ◽

...

Keyword(s):

Phase Transfer ◽

Membrane Lipids ◽

Positive Curvature ◽

Membrane Lipid ◽

Cell Penetrating Peptides ◽

Membrane Curvature ◽

Local Phase ◽

Vesicle Size ◽

Cell Penetrating ◽

A Cell

A specific series of peptides, called a cell-penetrating peptide (CPP), is known to be free to directly permeate through cell membranes into the cytosol (cytolysis); hence, this CPP would be a potent carrier for a drug delivery system (DDS). Previously, we proposed the mechanism of cytolysis as a temporal and local phase transfer of membrane lipid caused by positive membrane curvature generation. Moreover, we showed how to control the CPP cytolysis. Here, we investigate the phospholipid vesicle’s size effect on CPP cytolysis because this is the most straightforward way to control membrane curvature. Contrary to our expectation, we found that the smaller the vesicle diameter (meaning a higher membrane curvature), the more cytolysis was suppressed. Such controversial findings led us to seek the reason for the unexpected results, and we ended up finding out that the mobility of membrane lipids as a liquid crystal is the key to cytolysis. As a result, we could explain the cause of cytolysis suppression by reducing the vesicle size (because of the restriction of lipid mobility); osmotic pressure reduction to enhance positive curvature generation works as long as the membrane is mobile enough to modulate the local structure. Taking all the revealed vital factors and their effects as a tool, we will further explore how to control CPP cytolysis for developing a DDS system combined with appropriate cargo selection to be tagged with CPPs.

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Key Process and Factors Controlling the Direct Translocation of Cell-Penetrating Peptide through Bio-Membrane

International Journal of Molecular Sciences ◽

10.3390/ijms21155466 ◽

2020 ◽

Vol 21 (15) ◽

pp. 5466 ◽

Cited By ~ 2

Author(s):

Kazutami Sakamoto ◽

Taku Morishita ◽

Kenichi Aburai ◽

Kenichi Sakai ◽

Masahiko Abe ◽

...

Keyword(s):

Time Course ◽

Phase Transfer ◽

Membrane Lipid ◽

Cell Penetrating Peptide ◽

Giant Unilamellar Vesicle ◽

Dynamic Membrane ◽

Chemical Mediator ◽

Lipid Molecule ◽

Cell Penetrating ◽

Lipid Adsorption

Cell-penetrating peptide (CPP) can directly penetrate the cytosol (cytolysis) and is expected to be a potent vector for a drug delivery system (DDS). Although there is general agreement that CPP cytolysis is related to dynamic membrane deformation, a distinctive process has yet to be established. Here, we report the key process and factors controlling CPP cytolysis. To elucidate the task, we have introduced trypsin digestion of adsorbed CPP onto giant unilamellar vesicle (GUV) to quantify the adsorption and internalization (cytolysis) separately. Also, the time-course analysis was introduced for the geometric calculation of adsorption and internalization amount per lipid molecule consisting of GUV. As a result, we found that adsorption and internalization assumed to occur successively by CPP molecule come into contact with membrane lipid. Adsorption is quick to saturate within 10 min, while cytolysis of each CPP on the membrane follows successively. After adsorption is saturated, cytolysis proceeds further linearly by time with a different rate constant that is dependent on the osmotic pressure. We also found that temperature and lipid composition influence cytolysis by modulating lipid mobility. The electrolyte in the outer media is also affected as a chemical mediator to control CPP cytolysis by following the Hoffmeister effect for membrane hydration. These results confirmed the mechanism of cytolysis as temporal and local phase transfer of membrane lipid from Lα to Mesh1, which has punctured bilayer morphologies.

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Towards customized studies of G-quadruplex DNA structures in live cells

10.1101/2021.04.07.438780 ◽

2021 ◽

Author(s):

Bagineni Prasad ◽

Mara Doimo ◽

Mans Andreasson ◽

Valentin L Hote ◽

Erik Chorell ◽

...

Keyword(s):

Cell Model ◽

Model Systems ◽

Live Cells ◽

Cell Penetrating Peptide ◽

Dna Structures ◽

Cellular Effects ◽

G4 Dna ◽

G Quadruplex ◽

Cell Penetrating

G-quadruplex (G4) DNA structures are implicated in central biological processes and are considered promising therapeutic targets because of their links to human diseases such as cancer. However, functional details of how, when, and why G4 DNA structures form in vivo are largely missing leaving a knowledge gap that requires tailored chemical biology studies in relevant live-cell model systems. Towards this end, we developed a synthetic platform centered around one of the most effective and selective G4 stabilizing compounds, Phen-DC3. We used a structure-based design to equip Phen-DC3 with an amine in a position that does not interfere with G4 interactions. To evaluate the power of the approach, we next used this reactive handle to conjugate a BODIPY fluorophore and a cell-penetrating peptide to Phen-DC3. The BODIPY conjugation generated a fluorescent derivative with retained G4 selectivity, G4 stabilization, and cellular effects that revealed the localization and function of Phen-DC3 in human cells. On the other hand, the cell-penetrating peptide conjugation, while retaining G4 selectivity and stabilization, increased nuclear localization and cellular effects, showcasing the potential of this approach to modulate and direct cellular uptake e.g. as delivery vehicles. The developed platform can thus generate tailored biochemical tools for the studies of G4 biology to uncover molecular details and therapeutic approaches.

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