Category: Amino acid/peptide conjugates

The conjugation of different amino acids/peptides to various biologically active compounds has fetched the outstanding results as are very promising drug candidates.

An intranasally delivered peptide drug ameliorates cognitive decline in Alzheimer transgenic mice

Alzheimer’s disease (AD) is the most common neurodegenerative disease. Imbalance between the production and clearance of amyloid β (Aβ) peptides is considered to be the primary mechanism of AD pathogenesis. This amyloid hypothesis is supported by the recent success of the human anti‐amyloid antibody aducanumab, in clearing plaque and slowing clinical impairment in prodromal or mild patients in a phase Ib trial. Here, a peptide combining polyarginines (polyR) (for charge repulsion) and a segment derived from the core region of Aβ amyloid (for sequence recognition) was designed. The efficacy of the designed peptide, R8‐Aβ(25–35), on amyloid reduction and the improvement of cognitive functions were evaluated using APP/PS1 double transgenic mice. Daily intranasal administration of PEI‐conjugated R8‐Aβ(25–35) peptide significantly reduced Aβ amyloid accumulation and ameliorated the memory deficits of the transgenic mice. Intranasal administration is a feasible route for peptide delivery. The modular design combining polyR and aggregate‐forming segments produced a desirable therapeutic effect and could be easily adopted to design therapeutic peptides for other proteinaceous aggregate‐associated diseases.
Alzheimer’s disease (AD) is the most common neurodegenerative disease that causes dementia across multiple cognitive domains. Its incidence increases significantly with age and doubles every 5 years among the geriatric population ≧ 65 years of age. Despite remarkable scientific advancement and the vast resources invested in drug development, no effective therapy is currently available for AD. Thus, it is listed as one of the major unmet medical needs worldwide. Peptide drugs have been used with consistent benefits for many years and have advantages over small molecules, such as higher potency and fewer off‐target side effects (Craik et al, 2013). In addition, the properties of easy customization and synthesis under a well‐controlled environment make peptides excellent candidates for AD drug development. Neurodegenerative diseases encompass a heterogeneous group of neurological diseases characterized by synoptic and neuronal losses caused by multiple factors. Misfolded proteinaceous aggregates which exist in a variety of these diseases besides AD, including Parkinson’s disease, Huntington’s disease, amyotrophic lateral sclerosis, are considered one of them, and may cause or contribute to these diseases through their prionlike property (Kim & Holtzman, 2010; de Calignon et al, 2012; Luk et al, 2012; Smethurst et al, 2016). In spite of the difference in the constituent proteins and complexity of assembly mechanism, the proteinaceous aggregates across these diseases share common structural conformations such as a β‐sheet conformation in the backbone (Funke & Willbold, 2012). This provides the basis for a rational design of therapeutic peptides for these misfolded aggregate‐associated diseases by applying a universal principle to reverse the process of formation. (EMBO Mol Med. 2017 May; 9(5): 703–715.)

A calorimetric evaluation of the interaction of amphiphilic prodrugs of idebenone with a biomembrane model

Lipoamino acids (LAA) are useful promoieties to modify physicochemical properties of drugs, namely lipophilicity and amphiphilicity. The resulting membrane-like character of drug–LAA conjugates can increase the absorption profile of drugs through cell membranes and biological barriers. To show the role of amphiphilicity with respect to lipophilicity in the interaction of drugs with biomembranes, in the present study we evaluated the mode of such an interaction of lipophilic conjugates of LAA with the antioxidant drug idebenone (IDE). DSC analysis and transfer kinetic studies were carried out using dimyristoylphosphatidylcholine (DMPC) multilamellar liposomes (MLVs) as a model. For comparison, two esters of IDE with alkanoic acids were synthesized and included in the analysis. The experimental results indicate that based on their different structure, IDE–LAA conjugates interacted at different levels with respect to pure IDE with DMPC bilayers. In particular, a progressive penetration inside the vesicles was observed upon incubation of IDE–LAA compounds with empty liposomes. The enhanced amphiphilicity of the drug due to the LAA moieties caused more complex interactions with DMPC bilayers, compared to those registered with the native drug or IDE alkanoate esters.(J Colloid Interface Sci. 2006 Jul 15;299(2):626-35.)

Novel derivatives of abietic and dehydroabietic acid

Abietic and dehydroabietic acid are interesting diterpenes with a highly diverse repertoire of associated bioactivities. They have, among others, shown antibacterial and antifungal activity, potentially valuable in the struggle against the increasing antimicrobial resistance and imminent antibiotic shortage. In this paper, we describe the synthesis of a set of 9 abietic and dehydroabietic acid derivatives containing amino acid side chains and their in vitro antimicrobial profiling against a panel of human pathogenic microbial strains. Furthermore, their in vitro cytotoxicity against mammalian cells was evaluated. The experimental results showed that the most promising compound was 10 [methyl N-(abiet-8,11,13-trien-18-yl)-d-serinate], with an MIC90 of 60 μg/mL against Staphylococcus aureus ATCC 25923, and 8 μg/mL against methicillin-resistant S. aureusStaphylococcus epidermidis and Streptococcus mitis. The IC50 value for compound 10 against Balb/c 3T3 cells was 45 μg/mL. ( 2017 Jan 1;25(1):132-137.)

Galloyl-RGD as a new cosmetic ingredient

Background

The cosmetics market has rapidly increased over the last years. For example, in 2011 it reached 242.8 billion US dollars, which was a 3.9% increase compared to 2010. There have been many recent trials aimed at finding the functional ingredients for new cosmetics. Gallic acid is a phytochemical derived from various herbs, and has anti-fungal, anti-viral, and antioxidant properties. Although phytochemicals are useful as cosmetic ingredients, they have a number of drawbacks, such as thermal stability, residence time in the skin, and permeability through the dermal layer. To overcome these problems, we considered conjugation of gallic acid with a peptide.

Results

We synthesized galloyl-RGD, which represents a conjugate of gallic acid and the peptide RGD, purified it by HPLC and characterized by MALDI-TOF with the aim of using it as a new cosmetic ingredient. Thermal stability of galloyl-RGD was tested at alternating temperatures (consecutive 4°C, 20°C, or 40°C for 8 h each) on days 2, 21, 41, and 61. Galloyl-RGD was relatively safe to HaCaT keratinocytes, as their viability after 48 h incubation with 500 ppm galloyl-RGD was 93.53%. In the group treated with 50 ppm galloyl-RGD, 85.0% of free radicals were removed, whereas 1000 ppm galloyl-RGD suppressed not only L-DOPA formation (43.8%) but also L-DOPA oxidation (54.4%).

Conclusions

Galloyl-RGD is a promising candidate for a cosmetic ingredient. (. 2014; 15: 18.)

Traversal of the Blood–Brain Barrier by Cleavable l-Lysine Conjugates of Apigenin

Apigenin, a flavone abundant in parsley and celery, is known to act on several CNS receptors, but its very poor water solubility (<0.001 mg/mL) impedes its absorption in vivo and prevents clinical use. Herein, apigenin was directly conjugated with glycine, l-phenylalanine, and l-lysine to give the corresponding carbamate derivatives, all of which were much more soluble than apigenin itself (0.017, 0.018, and 0.13 mg/mL, respectively). The Lys-apigenin carbamate 10 had a temporary sedative effect on the mice within 5 min of intraperitoneal administration (single dose of 0.4 mg/g) and could be detected in the mice brain tissues at a concentration of 0.82 μg/g of intact Lys-apigenin carbamate 10 and 0.42 ug/g of apigenin at 1.5 h. This study accomplished the delivery of apigenin across the BBB in a manner that might be applicable to other congeners, which should inform the future development of BBB-crossing flavonoids.(J. Agric. Food Chem., 2018, 66 (30), pp 8124–8131)

Synthesis of doxorubicin cell‐penetrating peptide bioconjugates

Cell‐penetrating peptides, also known as protein transduction domain (PTD), have attracted interest as carriers for intracellular drug delivery. We report the first drug conjugate with a negatively charged amphipathic cell‐penetrating peptide. Furthermore, we compare two different doxorubicin cell‐penetrating peptide conjugates, which are both unique in their properties, due to their net charge at physiological pH, namely the positively charged octaarginine and the negatively charged proline‐rich amphipathic peptide. These conjugates were prepared exploiting a novel heterobifunctional crosslinker to join the N‐terminal cysteine residue of the peptides with the aliphatic ketone of doxorubicin. This small linker contains an activated thiol as well as aminooxy functionality, capable of generating a stable oxime bond with the C‐13 carbonyl group of doxorubicin. The disulfide bond formed between the peptide and doxorubicin enables the release of the drug in the cytosol, as confirmed by drug‐release studies performed in the presence of glutathione. Additionally, the cytotoxicity as well as the cellular uptake and distribution of this tripartite drug delivery system was investigated in MCF‐7 and HT‐29 cell lines.

Heterobifunctional crosslinkers are extensively applied in drug modifications, and their utilization has proven to be very important for the attachment of diverse carriers, such as monoclonal antibodies, proteins, polymers, and peptides, to doxorubicin. Established conjugation technique is applied at the C‐13 keto group by hydrazones, due to their fast hydrolysis in acidic environment existing in biological compartments like endosomes and lysosomes. However, the insufficient stability of the doxorubicin hydrazone conjugates has been reported even at physiological pH (7.4), leading to the release of the free drug in the bloodstream. In order to overcome these difficulties, we have chosen a crosslinker capable of creating an oxime bond on doxorubicin’s ketone, due to the higher hydrolytic stability of the oxime group. Additionally, thiol‐containing carriers, like albumin proteins, have also been conjugated to anthracyclines utilizing functional groups that are highly specific for sulfhydryl groups, e.g. maleimides and pyridyl disulfides. The application of pyridyl disulfides is advantageous, because a disulfide bond between the linker and the cargo is formed, which can be reduced in the cytoplasm by glutathione to deliver the freight. Furthermore, pyridyl disulfides can serve as a protective group during synthesis to avoid undesired dimerization as well as an activating group for the thiol to facilitate disulfide formation. In contrast, maleimides react with a thiol via Michael addition; thus, a covalent bond is created that cannot be cleaved under physiological conditions. Therefore, we have selected a heterobifunctional crosslinker that contains a protected aminooxy group and pyridyl disulfide.

Antimicrobial peptide-fatty acid conjugate

The rise of resistant bacteria has prompted the search for new antimicrobial agents. Antimicrobial membrane lytic peptides have potential as future microbial agents due to their novel mode of action. Recently conjugation of a fatty acid to antimicrobial peptides has been explored as a method to modulate the activity and selectivity of the peptide. Our work further explores these phenomena by testing two peptides, YGAAKKAAKAAKKAAKAA (AKK) and LKKLLKLLKLLKL (LKK), conjugated to fatty acids of varying length for their activity, structure, solution assembly properties and the ability to bind model membranes. We found that increasing the length of fatty acids conjugated to peptide AKK, up to a 16 carbons in length, increases the antimicrobial activity. Peptide AKK appears to lose activity when the minimal active concentration is higher than the critical miscelle concentration (CMC) of the molecule. Thus, if the CMC of the peptide conjugate is too low the activity is lost. Peptide LKK has no activity when conjugated to lauric acid and appears to aggregate at very low concentrations. Conjugation of AKK with a fatty acid increases its affinity to model supported lipid membranes. It appears that the increased hydrophobic interaction imparted by the fatty acid increases the affinity of the peptide to the surface thus increasing its activity. At concentrations above the CMC, solution self-assembly inhibits binding of the peptide to cell membranes.

Cleavable Doxorubicin–Peptide Conjugate

Although recent methods for targeted drug delivery have addressed many of the existing problems of cancer therapy associated with undesirable side effects, significant challenges remain that have to be met before they find significant clinical relevance. One such area is the delicate chemical bond that is applied to connect a cytotoxic drug with targeting moieties like antibodies or peptides. Here we describe a novel platform that can be utilized for the preparation of drug–carrier conjugates in a site-specific manner, which provides excellent versatility and enables triggered release inside cancer cells. Its key feature is a cleavable doxorubicin–octreotide bioconjugate that targets overexpressed somatostatin receptors on tumor cells, where the coupling between the two components was achieved through the first cleavable disulfide-intercalating linker. The tumor targeting ability and suppression of adrenocorticotropic hormone secretion in AtT-20 cells by both octreotide and the doxorubicin hybrid were determined via a specific radioimmunoassay. Both substances reduced the hormone secretion to a similar extent, which demonstrated that the tumor homing peptide is able to interact with the relevant cell surface receptors after the attachment of the drug. Effective drug release was quickly accomplished in the presence of the physiological reducing agent glutathione. We also demonstrate the relevance of this scaffold in biological context in cytotoxicity assays with pituitary, pancreatic, and breast cancer cell lines.(Mol Pharm. 2015 Dec 7;12(12):4290-300. )