Abstract: The intrinsic property of Cell Penetrating Peptides (CPP) is to deliver various molecules including nucleic acids, large plasmids, therapeutic drugs, imaging molecules, liposomes, nano-molecules to various cell and tissues, which indicates about the potential of CPP as therapeutic vehicle molecules. These are easy to prepare, well characterized, versatile and have ability to link with bioactive cargo through covalent and non-covalent bonds. The interaction between cargo and CPP is highly dependent on physiochemical properties of CPP including size, pH and presence of basic residues in the carrier peptide. Primary mechanism of transport of CPP is endocytosis; however, evidences of endocytosis independent mechanisms including carpet model, inverted micelle model, barrel stave pore model and toroidal pore model are also present. Though, the natural CPP are often non- selective and passive, these may be tuned to become specific and targeted by conjugating them with functional groups and chemicals. In fact several natural CPPs such as penetratin, Tat, polyarginies etc have been modified to achieve maximum penetration and desired characters. Linear CPPs may be brought into multi-branched topology to give dendrimeric structures having more cell penetrating capacity, lower toxicity and hemolysis and higher serum stability. Site-specific targeting of CPP might be helpful in achieving several goals including enhancement in protein expression, gene silencing, formation of pleuripotent cells, reduction in inflammation and apoptosis, trans-epithelial transport, neuroprotection, ischemia treatment, treating insulin disorders, delivery of nucleic acids and agricultural pest control. The present review encompasses the detailed information regarding different modes of entry of CPP inside the cells, designing and classes of such peptides, along with theirs versatile therapeutic applications.
INTRODUCTION
Cell Penetrating Peptides (CPP) are typically 3 to 30 amino acids containing peptides having positive charge, which facilitate its interaction with negatively charged glycosaminoglycans (GAGs) and sialic acids. The history of CPP commenced in 1998 with the discovery of Frankel and Pabo1, who discovered that the trans-activator of transcription (Tat) protein of HIV has ability to translocate across the plasma membrane. In 1991, Drosophila antennapedia homeodomain protein was demonstrated to be introduced inside the cells, which now-a-days referred as penetratin2. Since then, numerous CPP have been discovered and still the number is growing. The CPP can be natural, designed, or chimeras. To enhance cell penetrating ability, several strategies including changing of stereochemistry of amino acid from L to D, inclusion of unusual amino acids3, making them branched or cyclic4 and usage of β or γ-amino acids5 are in practice. The present review encompasses the detailed information regarding different modes of entry of CPP inside the cells, designing and classes of such peptides, along with theirs versatile therapeutic applications. It discusses the role of CPP in generation of pluripotent stem cells, anti-inflammatory properties, neuroprotective actions, treating insulin disorders, managing ischemia, delivery of nucleic acids and usages in agricultural pest control.
Design of CPP: The CPP can be divided into different groups based on their origin or distinguish characteristics. Previously, only natural CPP were known such as Tat and penetratin but later several synthetic CPP were also designed possessing the cell penetration properties and sometimes even more efficient than natural ones. Positively charged amino acids are abundantly present in CPP and a stretch of arginine (R) is almost ubiquitous in CPP6. Other characters like its secondary structure also play an imperative role during penetration. The CPP can be manipulated to have artificial amino acids for enhanced activity. For example, if lysine (K) residues are replaced with ornithine residue, its susceptibility for degradation is reduced. Alterations in the CPP structure may also aid in utility such as formation of dendrimer or cyclisation is often used to modify side chains7. However, while designing and altering, it is important to consider few important points including stability, toxicity, poor delivery, aggregation or poor yield upon synthesis8.
Formulation of CPP as transport vector: The CPP to cargo peptide conjugation is dependent upon two processes. The one is chemical covalent conjugation, usually achieved by chemical bonds like disulfide bonds, amine bonds or specific linkers9,10 and the second one is through expression of CPP-cargo as fusion protein in E. coli or Saccharomyces cerevisiae11,12. Physical conjugation is achieved simply by bulk mixing of cargo and CPP; which offers the ratio of cargo and CPP flexibility. The interaction of cargo and CPP is highly dependent on physiochemical properties like size, pH and presence of R residues in the carrier peptide13.
DIRECT ENTRY OF CPP
There are evidences of endocytosis independent mechanisms of entry of CPP. There are four existing models for direct entry (Fig. 1).
Carpet model: Positively charged CPP are able to cross plasma membrane easily. Hydrophobic residue of CPP faces towards plasma membrane14. Accumulation of CPP locally causes disturbance in electric charge of the membrane and micelle formation takes place. The lipid bilayer is disturbed as if it is affected by detergents15, therefore the mechanism is called detergent-like mechanism16. A transient pore is formed, through which the micelle is internalized.
Inverted micelle model: Alain Prochiantz's group presented the inverted micelle model based on NMR studies2. In this model, CPP is internalized by the process of receptor mediated endocytosis, that is an energy consuming mechanism17. The inverted micelle is a cavity in between two cell membrane bilayers, where CPP is surrounded by hydrophilic environment. This micellar phase is transient and without help of any vesicular body it passes into cytosol and there the peptide is released. Drosophila antennapedia homeodomain proteins, HIV-1 Tat protein and octa-arginine are the CPP having high internalization efficiencies via inverted micelle mechanism18.
Barrel stave pore model: Barrel stave mechanism the commonest pathway of CPP traffic. Amphipathic α-helices form transmembrane pores. The major steps involved are (a) In α-helical form, peptide bind to the membrane (b) Peptide monomers recognize each other in membrane bound state (c) α-helices further penetrate in hydrophobic core (d) Progressive addition of monomers to the barrel increase the size of barrel. The formed pores are less than 10 nm in size16. A peptide lines the pore (Fig. 1).
Toroidal pore model: In toroidal model, peptide molecules are always associated with lipid headgroup even when it is perpendicularly inserted in the membrane. In such a pore, the lipid monolayer bends continuously in a toroidal hole fashion and pore is lined by both the peptide and lipid19.
| Fig. 1: | Possible mechanisms of endocytosis-independent entry of the cell penetrating peptides (CPP) |
CLASSES OF CPP
The CPP have been studied originated from various organisms. The CPP differ in their length, sequence, hydrophobicity and polarity with variable confirmation. These can be broadly classified into cationic peptides, hydrophobic sequences, amphipathic peptides, proline-rich and chimeric or bipartite peptides etc.
Cationic CPP: These are mainly comprised of multiple R and K residues and carry net positive charge at physiological pH with the pKa value ∼12. The cationic CPP are generally derived from heparin, RNA and DNA-binding proteins. R possesses the guanidine head group, which stably binds with negatively charged phosphates and sulphates on the surface of cell membranes. K residues also are positively charged but devoid of guanidine head group. A comparison of polymers of R, histidine (H), K, or ornithine revealed that R polymers are most efficacious20. The number and the position of R or K residues also determine the CPP transporting efficacy. The R7-R9 peptides are most efficacious in translocation and the efficacy might be reduced by polysulfonated compounds or acidic pH. Tat CPP (RKKRRQRRR) derived from transcriptional activator protein of HIV-1 virus, is a classical cationic CPP and its dimer (RKKRRQRRRRKKRRQRRR) is high in ability to translocate across plant and human cells21. The truncated N-terminal Drosophila Antennapedia Transcription Factor (ATF) peptide, a 16 amino acid residue containing peptide is another example of cationic peptide (RQIKIWFQNRRMKWKK) and also referred as penetratin3. The seven R residues containing M918, a peptide derived from tumor suppressor protein p14ARF (22 residue long peptide) shares cationic nature as penetratin22.
Hydrophobic CPP: These are derived from signal peptide sequences. The example of hydrophobic CPP includes the Kaposi Fibroblast Growth Factor (FGF) signal peptide (AAVALLPAVLLALLAP) and integrin β3 signal peptide (VTVLALGALAGVGVG). Less number of Hydrophobic CPP has been reported. The SG3 (RLSGMNEVLSFRWL)23, contains only two R and one glutamic acid (E) residue and 6 hydrophobic amino acid residues24. Hydrophobic CPP contain only non-polar amino acid residues. In comparison to cationic or amphipathic CPP, hydrophobic CPP are less studied.
Amphipathic CPP: Amphipathic CPP contain amphipathic helices with hydrophilic and hydrophobic faces. These CPP insert into the lipid bilayer through hydrophobic interaction. The α-helical structure acquired by the primary amphipathic CPP molecule is responsible for its insertion into plasma membrane regardless of the strength of ionic interaction. The Model Amphipathic Peptide (MAP) is having α-helical structure with an 18 amino acids (KLALKLALKALKAALKLA). In case of secondary amphipathic CPP with random confirmation, the interaction with plasma membrane is greatly charge dependent and after binding, the CPP attain α-helical structure25. Pep-1 is a 21-amino acid long commercially available synthetic CPP, having a tryptophan (W) rich hydrophobic domain, K-rich hydrophilic domain and a spacer domain to separate and maintain the integrity of these two domains. While it prevents degradation of cargo proteins, it is being used for non-covalent transport of proteins.
Proline-rich sequences: Proline (P)-rich peptides are able to attain polyproline I (PPI) and polyproline II (PPII) confirmations. In the presence of aliphatic alcohols, the PPI form is dominating confirmation with right-handed helix with all cis peptide bonds, where in presence of aqueous media or aliphatic acids PPII form is dominating having left-handed extended helix with all trans bonds26. By functionalisation of a polyproline helix by O-alkylation of a hydroxyproline monomer, delivery vectors have been prepared. Proline-based leucine (L), K and R mimics also have been prepared. Synthetic proline-derived γ-peptides can be functionalized by acylation, alkylation or guanidylation to get hydrophobic, hydrophilic or amphipathic γ-peptides. Proline-based dendrimers are highly branched structures able to transport drugs, vaccines or DNA. However, at concentrations higher than 60 μM dendrimers are lethal, at lower concentrations these are efficient in carrying DNA even in the presence of serum. Amphipathic pro-rich CPP, appear to be most promising due to reduced toxicity and ease in synthesis27.
Chimeric or bipartite peptides: Chimeric CPP are the combination of two or more above listed peptides. A chimera containing Ala46-Tyr51 amino acid residue from β-lactamase inhibitory protein (BLIP) and LLIIL residues from cell-penetrating vascular endothelial-cadherin (pVEC) sequence, decreased the number of viable cells in presence of antibiotic. Thus, have greater importance in conquering β-lactamase-mediated ampicillin drug resistance28. Lysins are phage derived hydrolases, targeted to digest bacterial peptidoglycan cell wall in order to release the progeny. Because lysins are highly species specific and rarely are subjected to develop bacterial resistance, are now considered as potent alternative to antibiotics. The lysins have a typical structure of N-terminus catalystic domain and C-terminus cell-binding domain. A chimera of lysins could be prepared to improve the host range, activity, solubility and intracellular uptake. Such one chimera staphylolytic chimeolysin (ClyF) is able to lyse all clinical isolates of S. aureus including Methicillin-resistant Staphylococcus aureus (MRSA)29.
APPLICATIONS OF CPP
After the discovery of CPP, these fetch the attention of scientific community for targeted drug delivery across the plasma membrane. Many applications of CPP, in field of medicine have been invented and are in phase of clinical trials.
CPP for generation of pluripotent stem cells: Wilmut and his colleagues, demonstrated for the first time that adult somatic cell may be converted to undifferentiated embryonic stem cells using Somatic Cell Nuclear Transfer (SCNT) technique30. However, a less complicated technique was discovered later for reprogramming somatic cells to pluripotency by introducing transcription factors Oct4, Sox2, Klf4 and c-Myc through retroviral vectors31. However, the technique is impeded by integration of retroviral genomes in the host genomes and resulting risk of mutagenesis and genetic dysfunction. The CPP have provided a safer way to deliver these transcription proteins into human cells. If 9 R residues containing CPP is conjugated to these reprogramming proteins by fusion, it successfully trafficked these factors into fibroblasts from human origin and convert them into pluripotent stem cells5,32. In the experiment of Kaitsuka and Tomizawa33, both the mice and human iPS cells were differentiated to obtain a pancreatic lineage using transcription factors including Pdx1, NeuroD and MafA. A fusion of Tat peptide with nucleus localization signal is used to transduce four transcriptional factors (Oct4, Klf4, Sox2 and cMyc) and resulting embryoids express the most common pleuripotency markers such as SSEA1, Oct4, Sox2, Klf4 and Nanog34. Oct4 alone is also able to induce pleuripotency without the requirement of additional transcriptional factors32. The transcription factor Oct4 is having a 16 amino acid long peptide which has 68% amino acid homology with the CPP penetration and when tagged with fluorescein isothiocyanate (FITC), revealed its efficient uptake via endocytic pathway. Also, unmodified Oct4 self-penetrated in CVI-5B cells and human BJ foreskin fibroblasts in the experiment of Kim et al.32 revealed the potential of Oct4 CPP as the pluripotency reprogramming factor (Table 1).
CPP against inflammation: There are several mediators of inflammation including TNFα and IL-1β, reactive oxygen species, reactive nitrogen species and NF-κB pathway is upregulated65. A series of evidences suggest that NF-κB inhibition may block inflammation and the associated tissue damage.
| Table 1: | Cell penetrating peptides and their functions |
Anti-inflammatory peptide-6 (AIP6), interacts directly with p65 subunit and inhibit DNA-binding and transcriptional activities of NF-κB and the downstream cascade of generation of inflammatory mediators38. In the acute organ injury animal model, pathogen-derived inducers including staphylococcal enterotoxin B (SEB), lipopolysaccharide (LPS), or lectin concanavalin A (ConA) cause inflammation and apoptosis of the liver, which is dependent on the signalling pathways by tumor necrosis factor-alpha (TNF-α), IFN-γ and Fas-Fas ligand interaction. Suppressor of cytokine signalling (SOCS)1 and SOCS3 are rapidly produced to conquer proinflammatory signalling and then degraded. In case of acute inflammation SOCS are insufficient to curb proinflammatory signalling. Khandia et al .65 and Jo et al.66, reasoned the replenishment of exogenous SOCS3 as intracellular protein therapy. Recombinant cell penetrating form of SOCS3 having 12 amino acids long hydrophobic CPP derived from fibroblast growth factor-439, effectively suppressed cytokine-mediated acute inflammation and liver apoptosis40. Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is a regulator molecule in cancer progression and inflammation. Tat peptide conjugated with Nrf2 sequence has been shown to activate Nrf2 and its downstream target gene heme-oxygenase-1 (HO-1) in THP-1 monocytes. Also, the Nrf2 has anti-inflammatory role via inhibition of the TNF37.
CPP in targeting cancer: The CPP may play an important role in both the screening as well in targeting of tumor cell67. Specialized CPP named as tumor penetrating peptides are used to specifically target tumor cells68. The EPPT1 peptide linked to cationic polyacrylamide (CPAA) and FITC labelled is capable of targeting transmembrane MUC-1 protein present in colorectal cell lines69. Magainins, is a peptide derived from Xenopus laevis, which forms amphipathic α-helical structure. It causes pore formation in cancer cells and selectively kills them exempting lymphocytes or fibroblasts. Conjugation of Magainins with tumor homing peptide Bombesin, the anticancer effect is exemplified70. Defensins are small (3.5-4 kD), cysteine-rich peptides, produced by body as part of innate defence system. α-defensins (HNP1, HNP2, HNP3) are present in granules of neutrophils and their membrane permeabilization character is attributed to its ability to make pores in lipid bilayers71 and is effective against squamous cell carcinoma72 owning to its DNA damaging effects. However, the cytotoxicity of HNPs is not confined to cancers cells only and also harm normal leukocytes or epithelial cell73. CopA3 a synthetic peptide derived from defensin like molecules of insects has anticancer potential74. Bax is a pro-apoptotic peptide family, having nine α-helices. α5, α6 and α9 helices have ability to bind to cellular membrane and α5 and α6 helices of Bax, involving arginine lysine residues, are able to make toroidal like pore structure. The structure of Bax protein explains how it permeated mitochondrial membrane and cause apoptosis75. Designing a poro-peptide encompassing the pore forming region of Bax is able to induce caspase mediated apoptosis in cancer cells. Its peri-tumoral application regressed tumor in a nude mice xenograft model, thus effective ion anticancer therapy76. Bacteriorhodopsin C helix derived peptide called as pH (low)-dependent Insertion Peptide (pHLIP), is present in 3 forms. The first one is water soluble form at pH 7 or above; at neutral pH, it resides at the lipid bilayer and at acidic pH below 6, it is inserted into the lipid bilayer as α helix77. With the drop of pH folding of the peptide changes and the energy released in this confirmation change, is used to move the cargo present along with peptide across the membrane. pHLIP is a peptide which uses the inherent acidic character of cancer cells to target them78. (KLAKLAK)2 synthetic peptide, when fused with CPP or THPs, enter inside the cell and disrupt mitochondria and efficiently caused apoptosis in tumor cells79,80 (Fig. 2). More such CPP41-55 have been summarized in Table 1.
CPP in neuroprotective role: R rich CPP have been shown to be neuroprotective, with or without fusion to a neuroprotective cargo peptide and their efficacy increases with increasing length. The charge present on the CPP and number of R residue is important in the neuroprotective role of CPP, which is evident by the fact that charge neutral CPP i.e., polyglutamic acid (E9) and poly-lysine K10 peptide are only weakly protective in E induced neuronal death81. Maximum protection is achieved by R15 peptide, which is dependent on the endocytic property of peptide82. Meloni et al.83 demonstrated the neuroprotective role of Tat (GRKKRRQRRR), penetratin (RQIKIWFQNRRMKWKK) and oligoarginine-9 (R9: RRRRRRRRR) in in vitro neuronal cell stroke models and reduces excitotoxic calcium influx and its toxicity at high concentrations56. Iron is an element essential for oxygen transport, electron transfer and redox processes, transported by transferrin protein. Iron overload may result in accumulation of iron in redox active iron, toxic to cell. It causes oxidation of amino acids, proteins and DNA and thereby causes oxidative stress and cell death84. Iron overload in central nervous system leads to brain degenerative diseases including Alzheimer’s and Parkinson’s diseases85. With iron if Al, Cu and Zn metals are present, it further increases the reactive oxygen species86. Desferrioxamine-B (DFO), is an iron siderophore having significant affinity for Al and Zn too. In clinical trials DFO has shown anti-Alzheimer activity but poor bioavailability reduces its utility for use in neurodegenerative disorders87.
| Fig. 2: | Beneficial applications of cell penetrating peptides (CPP) |
To improve its bioavailability CPP may be very useful. In the experiment of Goswami et al.57, DFO has been covalently conjugated with TAT and penetratin and the redox activity of Fe was suppressed in HeLa and RBE4 cell lines.
CPP in ischemia: Blockage in the coronary artery of heart leads to ischemia. Even after restoring the revascularization by thrombolysis or angioplasty, it ends up in apoptosis of cardiac cells88. The BH4-peptide, derived from the Bcl-xL anti-apoptotic protein, conjugated with four different CPP [Tat, (RXR)4, Bpep and Pip2b] and these conjugates minimized staurosporine-induced apoptosis in primary cardiomyocytes. Intravenous administration of Tat-BH4 and Pip2b-BH4 conjugates, at a single dose of 1 mg kg‾1, before reperfusion, was able to reduce infarct size by 50% and apoptosis by 60%. Such targeted delivery of antiapoptotic peptide to myocardium may be of greater therapeutic potential in clinical conditions58.
CPP in delivery of nucleic acids: Delivery of nucleic acid into the cell is a comparatively difficult task due to negative charge on them, their comparatively higher molecular weight and propensity to degradation. Viral capsid proteins, which have inherent tendency to pack, protect and deliver the nucleic acid into its capsid may be an useful tool to deliver nucleic acid inside the cell89; however large size of capsid protein limits its utility. Hence the consideration is to search for capsid domains instead full protein, which can serve as CPP peptides. DENV C protein is large protein of 100 amino acid size and can facilitate large nucleic acid molecules translocation. pepR and pepM are the peptides, derived from DENV C protein, which have tendency to bind with RNA and membrane respectively59. pepM is highly hydrophobic, where pepR is highly cationic (LKRWGTIKKSKAINVLRGFRKEIGRMLNILNRRRR)90 (Fig. 2). pepR translocate through endocytic pathway, where pepM directly pass through lipid membranes (KLFMALVAFLRFLTIPPTAGILKRWGTI)91. All the three-dengue virus capsid protein C, pepM and pepR can translocate small interfering RNA (siRNA) and ssDNA molecule in mammalian cell59. The S413PV, Penetratin, Pep-1, Polyarginines, Transportan and Tat peptides are few CPP that are able to translocate nucleic acid61. The CPP and nucleic acids are often conjugated non-covalently, due to virtue of negatively charged nucleic acid and positively charged CPP. In vivo such conjugation offers greater reproducibility92.
CPP for agricultural pest control applications: There are some peptides which share similar characters with that of CPP. These are Membrane Acting Microbial Peptides (MAMP) having antimicrobial properties with carrying positive charge. Because some of the peptides are pathogenic to insects, these can be further exploited as biopesticides93. Small amphipathic peptide lycotoxin-1 (Lyt-1) from wolf spider (Lycosa carolinensis) forms a pore in the membrane and increase cell permeability thereby bringing the lysis of the cell by reducing ion and voltage gradient62. Amphipathic α-helical structure occurred in Lyt-1 peptide due to repetition of lysine residue at each 4th or 5th position in the peptide. This amphipathic alpha helical structure is present in other pore forming pore-forming peptides, including magainins, dermaseptins and adenoregulin94. Mutant Lyt-1 peptide has been identified through scanning mutant analysis technique, which is specific against armyworms, which causes damage to corn and other crops. The puroindoline proteins (PINA and PINB) of wheat, has a lipid binding ability and due to tryptophan rich domain it exhibits antibacterial and antifungal properties. Based on tryptophan rich domain, constructed synthetic peptide also exhibit the antibacterial and antifungal ability sparing mammalian cells. The PIN based peptides enter inside the cell by carpet method of entering63. Bipartite nuclear localization sequence virE2 protein, of Agrobacterium is able to mediate ssDNA transfer to nucleus of plant cells. In plant protoplasts, CPP like Tat, pVEC and transportan have shown concentration dependent, non-saturable and endocytosis independent internalization95.
CPP in trans-epithelial transport: Intestinal absorption of insulin has been seen to be enhanced by co administrating CPP like penetratin, however high amounts of penetratin is needed to stimulate intestinal absorption of insulin. In a study of Kamei et al.96, 26 penetratin analogues were evaluated for the absorption-enhancing efficiency based on chain length, hydrophobicity, basicity and amphipathicity. Oligoarginie significantly improved delivery of insulin from the intestine to the systemic circulation without the requirement of interlinking. Conjugation of nona-arginine (R9) to biologically active part of parathyroid hormone [PTH(1-34)], was toxic to the intestinal enterocytes. R rich CPP like R8, penetratin, pVEC and RRL helix significantly increase insulin absorption from intestine, however different CPP exhibited variation in potency while using D or L form97. Higher amount of CPP with insulin lead to formation of aggregates in solution and L-penetratin enhanced the insulin absorption even in aggregated form.
CPP in treating insulin disorders: Hyperglycemia is the feature of Diabetes. Patients who are unable to balance critical glucose level in blood Patients with diabetes are unable to effectively manage blood glucose levels, often suffer different complications like heart diseases, an increased risk of blindness and renal failure. The ailment is due to insufficient insulin. When insulin was conjugated to different cationic CPP like Tat, oligoarginine or oligolysine, the transmembrane transport to cultured rat alveolar epithelial cell monolayer was maximum for oligoarginine conjugate with 27 folds increase in comparison to native insulin98. Insulin is often taken by oral route; however enzymatic barrier rapidly degrades it and secondly mucosal barrier limits its bioavailability. CPP enhance transport ability as well as bioavailability up to 50% in comparison to subcutaneously administered insulin99. Also, low molecular weight protamine (LMWP) acts like CPP peptide, with a cell translocation efficiency equivalent to TAT100. Oligoarginine (R8), conjugated insulin transport efficacy was 5-7 times higher and co administration of oligoarginie-hydroxypropyl-β-cyclodextrin (HP-β-CD)- insulin was 8-10 times higher than normal insulin across the Caco-2 cell monolayer101.
SIGNIFICANCE STATEMENT
| • | Present review gives a brief account for the guidelines to design Cell penetrating peptides (CPP) as well as describes the mode of CPP trafficking inside the cell |
| • | Different categories of CPP including cationic, hydrophobic, amphipathic, proline-rich sequences and chimeric/bipartite peptides have been described in detail |
| • | Various applications of CPP encompass their use as anticancer peptide, anti-inflammatory, neuroprotective, anti-ischemic, trans-epithelial transporter of insulin, in treating insulin disorders, nucleic acid delivery vehicle and differentiation tool for induced pluripotent stem cells and in agricultural pest control |
CONCLUSION AND FUTURE PERSPECTIVE
The numbers of CPP are continuously expanding since day of its discovery. Several CPP have been designed to deliver various cargos not only inside the cells but also across the epithelial and endothelial barriers. The individual amino acid’s structure and any other factors are responsible for its intrinsic property to cargo therapeutic molecules and tailor made CPP might be there with improved characteristics. However, a proper mechanism of its delivery through which epithelial, endothelial and blood brain barrier are crossed and interaction with mucus is carried out, need to be elucidated. More reliable techniques are required to elucidate, those are enabled to tell the precise quantity of CPP delivered to target organ. The safer introduction of CPP into target tissue demands enhanced tissue specificity to reduce possible detrimental effects due to off-target delivery of cargo.
ACKNOWLEDGMENTS
Authors of the manuscript thank and acknowledge their respective Universities/Institutes.