Fmoc Strategy Research Articles

Novel Gd-DTPA-peptide for targeted breast tumor magnetic resonance imaging

The prevalence of breast cancer underscores the imperative for early diagnosis in guiding treatment decisions. This study introduces a novel contrast agent, Gd-DTPA-VGB3, derived from the peptide VGB3 targeting vascular endothelial growth factor receptor-1 (VEGFR1) and VEGFR2, to enhance the contrast of conventional drug Magnevist in breast tumor MRI. The MRI contrast agent was synthesized on rink amide resin via Fmoc strategy, incorporating amino acids, and coupling to diethylenetriaminepentaacetic acid (DTPA). Gadolinium (Gd)-DTPA-VGB3 displayed specific binding to VEGFR1/2 in a displacement binding assay. Gd-DTPA-VGB3 exhibited minimal cytotoxicity to normal MCF-10 cells while inhibiting 4T1 mammary carcinoma cell proliferation. Compared to Magnevist, Gd-DTPA-VGB3 demonstrated a 2.8-fold increase in contrast-to-noise ratio (CNR) (355 vs. 125). Gd-DTPA-VGB3 exhibited enhanced accumulation in 4T1 tumor-bearing mice, resulting in significant signal intensity improvement. The findings highlight Gd-DTPA-VGB3's specific binding to VEGFRs, substantiating its potential as a candidate for enhancing MRI contrast in breast cancer diagnostics.

Synthesis and biological evaluation of cyanobacterial-inspired peptides

Cyanobacteria are known producers of structurally diverse and potent natural products; the majority are peptides with unique modifications. Yet, there remains a huge underexplored chemodiversity from cyanobacteria. Here, we designed a linear octapeptide as a product of combinatorial peptide design inspired by the natural products from the filamentous cyanobacteria Hapalosiphon welwitschii and Leptolyngbya sp. The target peptide was synthesized via solid-phase peptide synthesis (SPPS) using fluorenylmethyloxycarbonyl-protecting group (Fmoc) strategy. Structural diversity was expanded by the substitution of unnatural amino acids to yield five analogues. The structure and sequence of the synthesized peptides were confirmed using nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry (MS). Biological activity evaluation was done; with none of the peptides showing antimicrobial or cytotoxic activities against microbial pathogens and mammalian cells, respectively. To our knowledge, this study is the first to report a combinatorial peptide design inspired by a natural product and a predicted biosynthetic product. This strategy of peptide design expands the chemistry of a known bioactive natural product with the aid of unexplored cyanobacterial biosynthetic gene clusters.

Assessment of bioactive peptides derived from laminin-111 as prospective breast cancer-targeting agents

Breast cancer remains a pressing public health issue primarily affecting women. Recent research has spotlighted bioactive peptides derived from laminin-111, implicated in breast tumor development. Remarkably, the sequences IKVAV, YIGSR, and KAFDITYVRLKF from the α1, β1, and γ1 chains, respectively, have garnered significant attention. This study aims to assess the potential of these radiolabeled peptides as targeting agents for breast cancer. The three peptides were synthesized using the Fmoc strategy, purified via reversed-phase high-performance liquid chromatography (RP-HPLC), and characterized through mass spectrometry. Iodine-131 (131I) radiolabeling was performed using the chloramine T method, exhibiting high radiochemical yield and stability for [131I]I-YIKVAV and [131I]I-YIGSR. Conversely, [131I]I-KAFDITYVRLKF demonstrated low radiochemical yield and stability and was excluded from the biological studies. The lipophilicity of the compounds ranged from − 2.12 to − 1.10. Serum protein binding assay for [131I]I-YIKVAV and [131I]I-YIGSR reached ≅ 48% and ≅ 25%, respectively. Affinity for breast cancer cells was evaluated using MDA-MB-231 and MCF-7 tumor cell lines, indicating the affinity of the radiopeptides with these tumor cells. Ex vivo biodistribution profiles of the radiopeptides were assessed in the MDA-MB-231 breast tumor animal model, revealing tumor tissue accumulation, supported by a high tumor-to-contralateral muscle ratio and autoradiography. These results signify the effective penetration of YIKVAV and YIGSR into tumor tissue. Therefore, the synthesized α1 and β1 peptide fragments exhibit favorable characteristics as potential breast cancer-targeting agents, promising future exploration as radiopharmaceuticals for breast cancer.

PEGylation of the antimicrobial peptide LyeTx I-b maintains structure-related biological properties and improves selectivity.

The biological activity of antimicrobial peptides and proteins is closely related to their structural aspects and is sensitive to certain post-translational modifications such as glycosylation, lipidation and PEGylation. However, PEGylation of protein and peptide drugs has expanded in recent years due to the reduction of their toxicity. Due to their size, the PEGylation process can either preserve or compromise the overall structure of these biopolymers and their biological properties. The antimicrobial peptide LyeTx I-bcys was synthesized by Fmoc strategy and coupled to polyethylene glycol 2.0 kDa. The conjugates were purified by HPLC and characterized by MALDI-ToF-MS analysis. Microbiological assays with LyeTx I-bcys and LyeTx I-bPEG were performed against Staphylococcus aureus (ATCC 33591) and Escherichia coli (ATCC 25922) in liquid medium. MIC values of 2.0 and 1.0 µM for LyeTx I-bcys and 8.0 and 4.0 µM for LyeTx I-bPEG were observed against S. aureus and E. coli, respectively. PEGylation of LyeTx I-bcys (LyeTx I-bPEG) decreased the cytotoxicity determined by MTT method for VERO cells compared to the non-PEGylated peptide. In addition, structural and biophysical studies were performed to evaluate the effects of PEGylation on the nature of peptide-membrane interactions. Surface Plasmon Resonance experiments showed that LyeTx I-b binds to anionic membranes with an association constant twice higher than the PEGylated form. The three-dimensional NMR structures of LyeTx I-bcys and LyeTx I-bPEG were determined and compared with the LyeTx I-b structure, and the hydrodynamic diameter and zeta potential of POPC:POPG vesicles were similar upon the addition of both peptides. The mPEG-MAL conjugation of LyeTx I-bcys gave epimers, and it, together with LyeTx I-bPEG, showed clear α-helical profiles. While LyeTx I-bcys showed no significant change in amphipathicity compared to LyeTx I-b, LyeTx I-bPEG was found to have a slightly less clear separation between hydrophilic and hydrophobic faces. However, the similar conformational freedom of LyeTx I-b and LyeTx I-bPEG suggests that PEGylation does not cause significant structural changes. Overall, our structural and biophysical studies indicate that the PEGylation does not alter the mode of peptide interaction and maintains antimicrobial activity while minimizing tissue toxicity, which confirmed previous results obtained in vivo. Interestingly, significantly improved proteolytic resistance to trypsin and proteinase K was observed after PEGylation.

Optimization of the Linker Length in the Dimer Model of E22P-Aβ40 Tethered at Position 38.

Since amyloid β (Aβ) oligomers are more cytotoxic than fibrils, various dimer models have been synthesized. We focused on the C-terminal region that could form a hydrophobic core in the aggregation process and identified a toxic conformer-restricted dimer model (E22P,G38DAP-Aβ40 dimer) with an l,l-2,6-diaminopimelic acid linker (n = 3) at position 38, which exhibited moderate cytotoxicity. We synthesized four additional linkers (n = 2, 4, 5, 7) to determine the most appropriate distance between the two Aβ40 monomers for a toxic dimer model. Each di-Fmoc-protected two-valent amino acid was synthesized from a corresponding dialdehyde or cycloalkene followed by ozonolysis, using a Horner-Wadsworth-Emmons reaction and asymmetric hydrogenation. Then, the corresponding Aβ40 dimer models with these linkers at position 38 were synthesized using the solid-phase Fmoc strategy. Their cytotoxicity toward SH-SY5Y cells suggested that the shorter the linker length, the stronger the cytotoxicity. Particularly, the E22P,G38DAA-Aβ40 dimer (n = 2) formed protofibrillar aggregates and exhibited the highest cytotoxicity, equivalent to E22P-Aβ42, the most cytotoxic analogue of Aβ42. Ion mobility-mass spectrometry (IM-MS) measurement indicated that all dimer models except the E22P,G38DAA-Aβ40 dimer existed as stable oligomers (12-24-mer). NativePAGE analysis supported the IM-MS data, but larger oligomers (30-150-mer) were also detected after a 24 h incubation. Moreover, E22P,G38DAA-Aβ40, E22P,G38DAP-Aβ40, and E22P,G38DAZ-Aβ40 (n = 5) dimers suppressed long-term potentiation (LTP). Overall, the ability to form fibrils with cross β-sheet structures was key to achieving cytotoxicity, and forming stable oligomers less than 150-mer did not correlate with cytotoxicity and LTP suppression.

Fmoc solid-phase synthesis of RF9 optimization with mass spectrometry verification

Abstract The RF9 compound, which is an antagonist of the FF neuropeptide receptors is used as a therapeutic substance to improve the effectiveness of opioids in the chronic treatment of pain. The purpose of this study was to find the most efficient method of RF9 synthesis. The optimization experiment involved solid-phase peptide synthesis. The Fmoc strategy is based on the usage of the 9-fluorenylmethoxycarbonyl group to block reactive amino groups. Commonly applied RF9 synthesis is based on DIC/HOBt activation of 1-adamantanecarboxylic acid prior to its substitution. The experiments carried out in this research were based on the routinely applied DIC/HOBt carboxylic group activation and this scheme was compared with the COMU/DIPEA and DIC approach. The obtained results showed that COMU/DIPEA was the most efficient and effective method of RF9 synthesis. Using this strategy, pure compound was obtained, without any by-products, and at a highest yield. The use of COMU/DIPEA can be an excellent alternative to the routinely used RF9 synthesis.

Chimeric Agonist of Galanin Receptor GALR2 Reduces Heart Damage in Rats with Streptozotocin-Induced Diabetes.

Neuropeptide galanin and its N-terminal fragments reduce the generation of reactive oxygen species and normalize metabolic and antioxidant states of myocardium in experimental cardiomyopathy and ischemia/reperfusion injury. The aim of this study was to elucidate the effect of WTLNSAGYLLGPβAH-OH (peptide G), a pharmacological agonist of the galanin receptor GalR2, on the cardiac injury induced by administration of streptozotocin (STZ) in rats. Peptide G was prepared by solid phase peptide synthesis using the Fmoc strategy and purified by preparative HPLC; its structure was confirmed by 1H-NMR spectroscopy and MALDI-TOF mass spectrometry. Experimental animals were randomly distributed into five groups: C, control; S, STZ-treated; SG10, STZ + peptide G (10 nmol/kg/day); SG50, STZ + peptide G (50 nmol/kg/day); G, peptide G (50 nmol/kg/day). Administration of peptide G prevented hyperglycemia in SG50 rats. By the end of the experiment, the ATP content, total pool of adenine nucleotides, phosphocreatine (PCr) content, and PCr/ATP ratio in the myocardium of animals of the SG50 group were significantly higher than in rats of the S group. In the SG50 and SG10 groups, the content of lactate and lactate/pyruvate ratio in the myocardium were reduced, while the glucose content was increased vs. the S group. Both doses of peptide G reduced the activation of creatine kinase-MB and lactate dehydrogenase, as well as the concentration of thiobarbituric acid reactive products in the blood plasma of STZ-treated rats to the control values. Taken together, these results suggest that peptide G has cardioprotective properties in type 1 diabetes mellitus. Possible mechanisms of peptide G action in the STZ-induced diabetes are discussed.

Synthesis and evaluation of chloro SPIRO and amino SPIRO supports: Novel hydrophilic cross-linked polystyrene polymers for solid phase peptide synthesis

Synthesis of a novel, beaded, hydrophobic/hydrophilic balanced and chemically robust chloro SPIRO polymer matrix consisting of styrene and 4-Chloromethyl styrene (VBC) as backbone and 3,9-Divinyl-2,4,8,10-tetraoxaspiro[5.5]undecane as cross-linking units is demonstrated here. The suspension polymerisation technique was preferred to achieve complete monomer to polymer conversion as well as to attain spherical beaded form. Two different cross-linking densities (2 and 4 mol.%) of polymer supports were synthesized and the composition of the functional bearing site VBC was fixed as 3 mol.% for both supports. The integrity of chloro SPIRO polymer formed was verified by Fourier transform infrared spectroscopy and scanning electron microscopy analysis. Chloro SPIRO resin was subjected to Gabriel Phthalimide reaction to form amino SPIRO resin. Chemical integrity of chloro SPIRO resin was tested in different standard solid phase peptide synthetic conditions and swelling capacity was studied using amino SPIRO resin in different polar solvent conditions. The synthetic capacity of novel amino SPIRO polymer matrix was evaluated by synthesising classically challenging acyl carrier protein peptide by Fmoc strategy. The purity of peptide was checked by high performance liquid chromatography and mass by matrix assisted laser desorption.

Total synthesis of xylapeptide B [Cyclo‐(L‐Leu‐L‐Pro‐N‐Me‐Phe‐L‐Val‐D‐Ala)

AbstractXylapeptide B is a cyclopentapeptide isolated from Xylaria sp. derived from the Chinese medicinal plant Sophora tonkinensis. Xylapeptide B was successfully synthesized by a combination of solid‐ and solution‐phase, using the Fmoc strategy, and 2‐chlorotrityl chloride resin. The coupling reagent used is a combination of HBTU/HOBt to assist in the formation of amide bonds. D‐Ala was chosen as the C‐terminal because it has a small residue and can facilitate the cyclization process. Linear peptide was cleaved from the resin using a dilute acid concentration of 20% TFA in DCM because the peptide has no protecting group at the side chain. Crude linear peptide was purified by semi‐preparative RP‐HPLC using 0%–100% ACN eluent for 35 min and obtained a pure mass of 22.4 mg (18.83%). Cyclization was carried out in solution phase using HBTU (3 eq.) and DIPEA (1% v/v) in diluted concentration (1.25 mM) for 72 h at room temperature. The cyclization stage was monitored by thin‐layer chromatography (TLC). Crude xylapeptide B was purified by semi‐preparative RP‐HPLC using 30%–80% ACN eluent for 40 min, to result in 6 mg (8.91%) of the desired xylapeptide B. All synthesized products were characterized by HR‐TOFMS, 1H‐, and 13C‐NMR.

Scope and Limitations of Barbituric and Thiobarbituric Amino Acid Derivatives as Protecting Groups for Solid‐Phase Peptide Synthesis: Towards a Green Protecting Group

AbstractDMB (Dimethylbarbituric) and DETB (Diethylthiobarbituric) are both barbituric and thiobarbituric acid derivatives respectively, that forms enamines with the Nα amine of amino acids. These compounds were found to be stable crystalline solids and show stability in the standard acidic and basic conditions used for solid‐phase peptide synthesis (SPPS) strategies. These protecting groups are cleaved by a mild solution of 2 % hydrazine hydrate in DMF and 2 % hydroxylamine in DMF, both at short reaction times. Their use in SPPS showed that DMB‐protected amino acids allow the preparation of peptides and therefore could be an alternative to the Fmoc strategy currently used. A further advantage of these protecting groups is that their preparation does not involve the concourse of phosgene derivatives and therefore they could be considered greener protecting groups than the carbamate‐based one.

Skipjack (Katsuwonus pelamis) elastin hydrolysate‐derived peptides attenuate UVA irradiation‐induced cell damage in human HaCaT keratinocytes

AbstractWe aimed to identify skipjack elastin hydrolysate‐derived peptides, and explore its mechanism of photoprotection against ultraviolet A (UVA) induced skin damage. Two peptides, TGVLTVM (peptide A) and NHIINGW (peptide B), were successfully identified using Sephadex G‐15, analyzed and detected using LC‐MS/MS fitted with electrospray ionization (ESI‐MS/MS), and further synthesized using the Fmoc strategy. A UVA radiation‐induced damage cell model of Human keratinocytes cell line (HaCaT) was established and applied. We found that both peptides significantly reduced reactive oxygen species (ROS) generation, reversed mitochondrial membrane potential (MMP) by 12.40%–13.13% and 10.80%–12.90% and reduced cell apoptosis by 3.43%–6.44%, and 4.56%–6.03% for peptide A and B, respectively, compared to the UVA model group. In conclusion, peptides TGVLTVM and NHIINGW protected against UVA‐induced photoaging through the attenuation of oxidative stress, thus both peptides might be useful as promising ingredients in sunscreens or antiaging cosmetic products.

Total synthesis of cyclohexadepsipeptides exumolides A and B

Abstract Exumolides A and B are two cyclodepsipeptides that was previously isolated from marine fungus. Both peptides were synthesized using a combination of solid- and solution-phase method. The linear precursor was synthesized through solid-phase method on 2-chlorotrityl resin with standard Fmoc strategy. The hydroxy acid, (S)-2-hydroxy-4-methylpentanoic acid (Hmp), was prepared from its precursor l -leucine, and attached on the resin with a double-coupling protocol. The depside bond formation that was carried out at the end of the coupling process, was beneficial, particularly, in the blockage of the diketopiperazine formation during Fmoc deprotection. The cyclic product was obtained through HATU-based cyclisation. Exumolides A and B were successfully synthesized with overall yields of 4.12% and 6.39%, respectively.

Cationic peptidomimetics derived from CAP-18 cathelicidin shown activity in vitro against mycobacterium tuberculosis H37RA and synergy combined with isoniazid

Aims & objectives: Tuberculosis (TB) is the infectious disease that results in the largest number of deaths around the world caused by a bacterial infectious agent. According to the World Health Organization (WHO), there were 10 million new cases and 1.2 million deaths caused by TB in 2018. The problem increases due to drug resistance and HIV/AIDS coinfection, which is why it is necessary to develop new anti-TB compounds with different mechanisms of action. Antimicrobial peptides are molecules gener42ated in the innate immune response of the host with an effect on pathogens. This is the case of the cathelicidins present in mammals involved in the formation of channels in the bacterial cell membrane that leads to cell lysis. Methods: In this study, the antimycobacterial activity and the possible mechanism of action of peptides derived from CAP-18 with unnatural amino acids were evaluated. Using bioinformatic tools, the epitope of the native sequence with the best characteristics of helical structure, amphipathicity and antibacterial value was determined. The corresponding peptides, CAP18-B and CAP18-C, had changes from L-Lys to D-Lys and from L-Gly to N-methyl-Gly in the sequence, generating the peptides CAP18-BD, CAP18-CD and the peptoid CAP18-BP. Peptides and peptoids were synthesized using the Fmoc strategy, purified by RP-HPLC and characterized by ESI-MS, circular dichroism and H1 -NMR. The in vitro assesament of the designed sequences was carried out in Mycobacterium smegmatis mc2 155 and Results: Mycobacterium tuberculosis H37Ra, obtaining a minimum inhibitory concentration increase from 2 to 6 times in the ranges from 1200 to 60 μg/mL, less than 10% hemolytic activity and no cytotoxic activity on U937 monocytes with LC50 values at least 10 times higher than the MIC and synergy combined with isoniazid. Confocal microscopy showed the insertion of peptides and peptoids in Mycobacterium smegmatis mc2 155 and transmission electron microscopy showed alterations in the cell membrane of Mycobacterium smegmatis mc2 155 evidenced in the formation of blisters, microspheres, channels and cell lysis. Conclusions: The results suggest that analogous sequences derived from CAP-18 can be considered as potential antimycobacterial compounds.

Shortened derivatives from native antimicrobial peptide LyeTx I: In vitro and in vivo biological activity assessment.

In the continuing search for novel antibiotics, antimicrobial peptides are promising molecules, due to different mechanisms of action compared to classic antibiotics and to their selectivity for interaction with microorganism cells rather than with mammalian cells. Previously, our research group has isolated the antimicrobial peptide LyeTx I from the venom of the spider Lycosa erythrognatha. Here, we proposed to synthesize three novel shortened derivatives from LyeTx I (LyeTx I mn; LyeTx I mnΔK; LyeTx I mnΔKAc) and to evaluate their toxicity and biological activity as potential antimicrobial agents. Peptides were synthetized by Fmoc strategy and circular dichroism analysis was performed, showing that the three novel shortened derivatives may present membranolytic activity, like the original LyeTx I, once they folded as an alpha helix in 2.2.2-trifluorethanol and sodium dodecyl sulfate. In vitro assays revealed that the shortened derivative LyeTx I mnΔK presents the best score between antimicrobial (↓ MIC) and hemolytic (↑ EC50) activities among the synthetized shortened derivatives, and LUHMES cell-based NeuriTox test showed that it is less neurotoxic than the original LyeTx I (EC50 [LyeTx I mnΔK] ⋙ EC50 [LyeTx I]). In vivo data, obtained in a mouse model of septic arthritis induced by Staphylococcus aureus, showed that LyeTx I mnΔK is able to reduce infection, as demonstrated by bacterial recovery assay (∼10-fold reduction) and scintigraphic imaging (less technetium-99m labeled-Ceftizoxime uptake by infectious site). Infection reduction led to inflammatory process and pain decreases, as shown by immune cells recruitment reduction and threshold nociception increment, when compared to positive control group. Therefore, among the three shortened peptide derivatives, LyeTx I mnΔK is the best candidate as antimicrobial agent, due to its smaller amino acid sequence and toxicity, and its greater biological activity.

Inhibition properties of free and conjugated leupeptin analogues.

Leupeptin is a naturally occurring inhibitor of various proteases, in particular serine proteases. Following its discovery, the inhibitory properties of several other peptidyl argininals have been studied. The specificity of leupeptin is most likely due to the Leu–Leu–Argininal sequence, and its C‐terminal aldehyde group has been suggested to enhance the binding efficiency and to be essential for function. The terminal aldehyde group makes the structure less vulnerable to carboxypeptidases. Here, we investigated whether the inhibitory function of leupeptin toward serine proteases is retained after oxidation or reduction of the aldehyde group. The oxidized form, which corresponds to the natural precursor, was shown to be superior to the reduced form in terms of inhibitory properties. However, the original leupeptin possessed enhanced inhibitory properties as compared with the oxidized form. Based on these results, new synthetic leupeptin analogues, 6‐aminohexanoic acid (Ahx)–Phe–Leu–Arg–COOH and Ahx–Leu–Leu–Arg–COOH, were prepared by solid‐phase peptide synthesis using the Fmoc strategy. In these analogues, the N‐terminal capping acetyl group was replaced with a 6‐aminohexanoyl group to allow conjugation. The structures of the modified leupeptin and the synthetic peptides were confirmed by mass spectrometry. Determination of the inhibitory properties against trypsin (IEC 3.4.21.4, Chymotrypsin IEC 3.4.21.1) revealed that these further modified tripeptides were tight binding inhibitors to their target enzyme, similar to the naturally occurring leupeptin, with Ki values generally in the micromolar range. The Ahx–Phe–Leu–Arg–COOH analogue was selected for conjugation to inorganic oxide nanoparticles and agarose gel beads. All conjugates exhibited inhibitory activity in the same range as for the free peptides.

Sintesis Tetrapeptida Linear (DPAP) Menggunakan Metode Sintesis Peptida Fasa Padat (SPPS) Dan Aktivitas Insektisidanya Terhadap Ulat Krop Kubis

Crop cabbage worm (Crocidolomia pavonana) is one of the pests that has a bad effect on the cabbage production in Indonesia. One of the efforts that has been applied to control this pest is by using bayrusil. However, the bayrusil can cause negative effects such as poisoning and environmentally polluting effects. To prevent the undesired effects, it has been developed the use of environmentally friendly insecticide and one of them is the use of insecticidal peptide. Tripsin-modulating oostatic factor (TMOF) and ist analogues is one of the peptides that has an insecticidal activity. The aim of this research is to synthesise a linear tetrapeptide (DPAP) an analogues of TMOF and to test insecticidal activities against crop cabbage worm. A DPAP has been synthesised by Solid Phase Peptide Synthesis on 2-chlorotrytil chloride resin. Fmoc strategy was applied on the synthesis and a combination DIC/Oxime was employed in the coupling reaction.The tetrapeptidyl resin was cleaved by TFA:water:EDT (90:5:5). The peptide was purified by reverse-phase column chromatography and characterized by TOF ES-MS spectroscopy. A linear peptide was biologically tested towards the C. pavonana, showing percentage of mortality values at 1000 ppm.

A Strategy To Prepare Peptide Heterodimers in the Solid Phase with an Acid-Labile Linker.

The currently synthetic methods of peptide heterodimer involve tedious synthesis and purification steps. An acid-labile traceless linker was prepared, which is highly compatible with the Fmoc strategy and could be used to prepare peptide heterodimer on resin. The linker could be cleaved concomitantly with peptide cleavage, and two model heterodimers were synthesized. The proposed synthesis procedure is simple, straightforward, and provides great convenience for preparing disulfide-linked peptide heterodimers.

Synthesis of Antioxidant Peptide SCAP1 (Leu-Ala-Asn-Ala-Lys)

SCAP1 is a pentapeptide isolated from the hydrolysate of oyster (Saccostrea cucullata), which has antioxidant and anticancer properties. This study aims to find a novel strategy to obtain antioxidant peptide SCAP1 (Leu-Ala-Asn-Ala-Lys) through chemical synthesis. This study reports the first synthesis of SCAP1 through the solid phase peptide synthesis (SPPS) method with Fmoc strategy on 2-chlorotrityl chloride resin. A HATU-HOAt coupling reagent was employed in all amide bond formations and 95% TFA in water was employed to release the desired peptide from the solid support. The crude peptide was purified using successive preparative-RP-MPLC and RP-HPLC to obtain the pure peptide. The synthesized peptide was confirmed by mass spectrometry, showing a correct molecular ion peak at m/z 516.3143 (M+H)+, referring to the desired SCAP1. 1H-NMR spectra of the peptide revealed protons signals consistent to the structure of SCAP1. During the synthesis, the coupling between Ala and Asn(Trt) was not straightforward, which led to a low yield of the final product. The use of Trt-based Asn was not a suitable option, as it could cause aggregation during the synthesis, eventually leading to a poor yield of the final product. SCAP1 was obtained as a white solid with a percent yield of 8.28%.

Synthesis, in vitro biological activity and docking of new analogs of BIM-23052 containing unnatural amino acids.

Somatostatin (SST) is an endogenous cyclic tetradecapeptide hormone that exerts multiple biological activities via a family of five receptors. BIM-23052 (DC-23-99) D-Phe-Phe-Phe-D-Trp-Lys-Thr-Phe-Thr-NH2 is a linear SST analog with established in vitro GH-inhibitory activity and high affinity to sstr5, sstr3 and sstr2. The different SSTR subtypes are expressed in different tissues and in some tumor cells. Based on this finding, a series of new analogs of BIM-23052 with expected antitumor activity have been synthesized. The Thr at position 6 in BIM-23052 was replaced by the conformationally hindered Tle, Aib, Ac5c and Ac6c of the new analogs. The peptides were synthesized by standard solid-phase peptide chemistry methods, Fmoc strategy. The cytotoxic effects of the compounds were tested in vitro against a panel of tumor cell lines: HT-29, MDA-MB-23, Hep-G2, HeLa and the normal human diploid cell line Lep-3. All five somatostatin receptor subtypes were modeled and docking was performed to determine the binding affinity of the analogs. The new peptides exhibited different concentration-dependent antiproliferative effect on the tumor cell lines after 24h of treatment. The compound 3B (Aib6) demonstrated the most pronounced antiproliferative effects on HepG-2 cells with the IC50 = 0.01349nM. Docking confirmed that all compounds bind well to SST receptors with preference to sstr3 and sstr5, which is most probably the reason for the observed biological effects.

A faster solid phase peptide synthesis method using ultrasonic agitation

Solid phase-supported synthesis is a widely used strategy in peptide chemistry. A factor which limits the product purity is the individual stages yields. Here, we reported that the use of ultrasonic agitation allows to reduce tenfold the time of synthesis in the Fmoc strategy and improve the purity of the final product. Our method is a promising alternative to traditional synthetic methods and microwave synthesizers.