Labeling Of Peptides Research Articles

Enhancing and verification of dose in external radiation therapy using Gd nanoparticles as a theranostic agent: A Monte Carlo simulation study

Introduction: Theranostics, in particular, the use of radionuclides with the capability of simultaneous imaging and treatment has opened new horizons in personalized treatment planning of targeted radiation therapy. In this approach, positive beta or gamma emitters are required for imaging and alpha, beta and Auger electrons for treatment purpose. On the other hand, studies have shown that due to high biological effectiveness the combination of external radiation modalities such as neutrons and protons with internal radiation modalities, can have a potential impact on cancer cell killing. Therefore, given the advances made in nanoparticles and radiopharmaceuticals, the necessity of designing combined therapy modalities and radiopharmaceuticals with a diagnostic-therapeutic capability is of particular importance, and Monte Carlo computational methods are a powerful tool in this regard. Materials and Methods: In this study, by using the Geant4 and MCNPX Monte Carlo Codes, the absorbed dose of the Auger electrons, KERMA and the flux of the prompt gamma photons was calculated. the distribution of gadolinium nanoparticles with different concentrations and also without them, in two geometries of uniform PMMA environments surrounded by water and the cell model, was subjected to neutron irradiation and the dose was measured in macroscopic and microscopic conditions. Regarding the higher energy range compared to the diagnostic items in nuclear medicine, a gamma camera was optimized for counting the prompt gamma photons by using the F4 and F2 flux calculations as well as count calculation F1 MCNP tallies Results: Dose enhancement was observed in micro and macro dosimetry conditions. For example, the amount of dose in the cellular model assigned to the nucleus was 5.1 E-5 Gy, 3.01E-03 Gy, and 4.05E-7 Gy for the distribution of Gd in the core, total nucleus, and cytoplasm for particle history, respectively. There was also a direct correlation between the number of prompt gamma rays and the nanoparticle concentration Conclusion: Labeling of antibodies or peptides with gadolinium nanoparticles in the presence of external radiation beam can be used as a marker for drug distribution, as well as targeted radiation therapy theranostic agent because of enhancement of doses due to auger electrons and prompt gamma photons with imaging capability that showing the synergy impact of diagnostic imaging and therapeutics.

Docking, synthesis, in-vitro evaluation, and optimization of reaction conditions for direct radiolabeling of CGPRPPC with technetium-99m through the GAGG sequence.

With respect to the reported promising results of cyclic peptide CGPRPPC in early detection of thrombotic lesions, we developed a practical approach for technetium-99m labeling of this peptide using the Glycine-Alanine-Glycine-Glycine (GAGG) sequence as a chelating moiety. The peptide conjugated to GAGG was prepared using the solid-phase method. The optimization of radiolabeling conditions was performed on the basis of such variables as incubation time, reaction temperature, pH, and concentration of peptide and stannous chloride. Moreover, the stability and fibrin-binding affinity of the radiolabeled peptide were measured. The peptide-fibrin interactions were analyzed by docking studies using HEX and Auto dock 4.2. Softwares. The amounts of synthesized peptide and stannous chloride required for optimal radiolabeling through GAGG were 10 µmol/l and 5 µg, respectively. The best radiochemical purity% (>93%) was achieved at pH 7-8 within 15 min and a reaction temperature of 37°C. On the basis of in silico and in-vitro results, the GAGG-conjugated CGPRPPC peptide showed better binding affinity versus the HYNIC-conjugated one. We could radiolabel the fibrin-targeting peptide with high radiochemical purity% and stability during a short incubation period without a boiling step. Compared with the HYNIC-conjugated peptide, a higher binding affinity was found. Therefore, the GAGG chelating moiety possesses a considerable potentiality in Technetium 99m labeling of peptides while CGPRPPC maintains its binding properties to thrombotic lesions.

Pyrrole-Mediated Peptide Cyclization Identified through Genetically Reprogrammed Peptide Synthesis.

Flexible in vitro translation (FIT) was used as a screening method to uncover a new methodology for peptide constraining based on the attack of a nucleophilic side-chain functionality onto an oxidized furylalanine side chain. A set of template peptides, each containing furylalanine as furan-modified amino acid and a nucleophilic residue (Cys, His, Lys, Arg, Ser, or Tyr), was produced through FIT. The translation mixtures were treated with N-bromosuccinimide (NBS) to achieve selective furan oxidation and subsequent MALDI analysis demonstrated Lys and Ser as promising residues for cyclisation. Solid-phase peptide synthesis (SPPS) was used to synthesize suitable amounts of material for further in-depth analysis and characterisation. It was found that in the case of the peptide containing lysine next to a furylalanine residue, a one-pot oxidation and reduction reaction leads to the generation of a cyclic peptide featuring a pyrrole moiety as cyclisation motif, resulting from the attack of the lysine side chain onto the oxidized furylalanine side chain. Structural evidence was provided via NMR and the generality of the methodology was explored. We hereby expand the scope of our previously developed furan-based peptide labeling and crosslinking strategy.

A Ubiquitous but Overlooked Side Reaction in Dimethyl Labeling of Peptides.

Reductive dimethylation using formaldehyde and NaBH3CN to label peptides or proteins on their N-termini and lysine residues is one of the most widely used labeling methods in the quantitative proteomics field. In this study, we characterized a ubiquitous side reaction in dimethylation labeling, causing mass increments of 26 Da on the N-termini of peptides. It can occur extensively on most peptides, which significantly compromises data quality in terms of sensitivity, dynamic range, and peptide- and protein-identification rates. Nevertheless, this side reaction was so-far overlooked, largely because the current database search algorithms limited the detection of unknown modifications. In order to illustrate the chemical nature of this side reaction, 1D and 2D nuclear magnetic resonance (NMR) was performed to elucidate the exact structure of the modification formed through this side reaction, revealing that the side reaction produced an N-methyl-4-imidazolidinone moiety between the first two residues of the undesirably labeled peptides. On the basis of the mechanism proposed for the side reaction, we optimized the reaction conditions for dimethyl-labeling. Compared with the current typical labeling method, our approach can dramatically suppress the side reactions at both the standard protein and proteome levels. As a result, with our optimal labeling method, peptide- and protein-identification rates were significantly increased compared with those from the traditional labeling method.

IEDDA Conjugation Reaction in Radiometal Labeling of Peptides with 68Ga and 64Cu: Unexpected Findings.

The inverse electron demand Diels–Alder conjugation reaction has gained increasing importance over the past few years for efficient in vivo and ex vivo radiometal labeling of antibodies. However, the application of this very fast reaction type has not been studied for radiolabeling of peptides so far. We show here the synthesis of 3-benzyl-1,2,4,5-tetrazine-comprising ((1,4,7,10-tetraazacyclododecane-4,7,10-triyl)triacetic acid-1-glutaric acid) (DOTA–GA) and ((1,4,7-triazacyclononane-4,7-diyl)diacetic acid-1-glutaric acid) (NODA–GA) chelators and their radiometal labeling with 68Ga3+ and 64Cu2+. The secondary labeling precursors 68Ga–DOTA–GA–Tz, 68Ga–NODA–GA–Tz, and 64Cu–DOTA–GA–Tz were obtained in high radiochemical yields (RCYs) and purities as well as molar activities for further labeling of trans-cyclooctene (TCO)-modified peptides. However, the following reactions of the radiometal-labeled tetrazines with different TCO-comprising model peptide analogs unexpectedly resulted in the formation of a considerable amount of side products (20–55%) which limits the overall achievable RCYs and purities as well as molar activities of the target radiopeptides. Under otherwise identical, nonradioactive reaction conditions, this effect could however not be observed. In contrast, the corresponding one-step radiolabeling protocols provided the target 68Ga-labeled radiopeptides in exceptionally high RCYs and purities of ≥99% and molar activities of 68–72 GBq/μmol starting from activities of 340–358 MBq of 68Ga. Thus, the usefulness of the two-step labeling of TCO-modified peptides with radiometal-labeled chelator-tetrazines seems to be limited.

A16523 Mapping of the angiotensin AT2 receptor-coupled signalling network by time-resolved quantitative phosphoproteomics in human aortic endothelial cells identified HDAC-1 and p53 to be involved in AT2 receptor-mediated anti-proliferation

Objectives: The angiotensin AT2-receptor (AT2R) is a main component of the protective arm of the renin-angiotensin system. In order to get new and deeper insights into AT2R mediated signalling, this study mapped the AT2R-coupled phosphoproteome in human aortic endothelial cells (HAEC) by quantitative phosphoproteomics. Methods: HAEC were stimulated with the AT2R agonist C21 (1 μM) for 1, 3, 5 or 20 minutes. Following protein digestion and TMT peptide labelling, samples were subjected to TiO2−SIMAC phospho-peptide enrichment and LC−MS/MS. Data were analysed using Proteome Discoverer and the human sequence libraries SwissProt and UniProt. For confirmation of phosphoproteomics results, expression of p53 and phosphorylation of HDAC-1 were determined by Western Blot. Cellular translocation of HDAC-1 and p53 was assessed by immunofluorescence and activity of HDAC-1 by deacetylation assay. HAEC proliferation was determined by resazurin assay and apoptosis by caspase 3/7 assay. Results: AT2R stimulation of HAEC changed the phosphorylation status of 265 proteins at 471 sites (281 phosphorylations and 190 dephosphorylations). Gene Ontology and STRING analysis revealed the predominant phospho-modification of proteins involved in anti-proliferation, cell differentiation and apoptosis including HDAC-1, p53, BCL2 or HSP90. AT2R-mediated inhibition of HDAC-1 and activation of p53 were confirmed by additional experiments demonstrating changes in cellular localisation of HDAC-1 and p53, in total expression of p53, and in enzymatic activity and phosphorylation of HDAC1. Conclusion: This study mapped the AT2R coupled phospho-signalling network. It identified dephosphorylation and deactivation of HDAC-1 as well as nuclear importation and increased expression of p53 as novel, potential mediators of AT2R-coupled anti-proliferation.

Development of Chemical Proteomics for the Folateome and Analysis of the Kinetoplastid Folateome.

The folate pathway has been extensively studied in a number of organisms, with its essentiality exploited by a number of drugs. However, there has been little success in developing drugs that target folate metabolism in the kinetoplastids. Despite compounds being identified which show significant inhibition of the parasite enzymes, this activity does not translate well into cellular and animal models of disease. Understanding to which enzymes antifolates bind under physiological conditions and how this corresponds to the phenotypic response could provide insight on how to target the folate pathway in these organisms. To facilitate this, we have adopted a chemical proteomics approach to study binding of compounds to enzymes of folate metabolism. Clinical and literature antifolate compounds were immobilized onto resins to allow for “pull down” of the proteins in the “folateome”. Using competition studies, proteins, which bind the beads specifically and nonspecifically, were identified in parasite lysate (Trypanosoma brucei and Leishmania major) for each antifolate compound. Proteins were identified through tryptic digest, tandem mass tag (TMT) labeling of peptides followed by LC-MS/MS. This approach was further exploited by creating a combined folate resin (folate beads). The resin could pull down up to 9 proteins from the folateome. This information could be exploited in gaining a better understanding of folate metabolism in kinetoplastids and other organisms.

Reagents for Isobaric Labeling Peptides in Quantitative Proteomics.

Currently, the commercial reagents for isobaric peptides labeling (TMT and iTRAQ) have some drawbacks, such as high cost in experiments, especially in quantitation for the modified peptides, and inconvenient handling for variable sizes of samples. Herein, we developed a set of 10-plex isobaric tags (IBT) with high stability and low cost. The labeled peptides were sensitively detected on Orbitrap Q Exactive MS with an MS2 resolution of 35 000 at 30% NCE, while the peptides were efficiently labeled over 97% by IBT at a ratio of 10:1 of reagent/peptide (w/w) in 200 mM TEAB buffer for 2 h. The IBT labeling was demonstrated with a wide dynamic range of 50-fold without obvious matrix effects on quantification. Importantly, there was little quantification bias found among the individual IBT tags, indicating that the peptides labeled by different tags were quantitatively comparable. The IBT 10-plex reagents were applied for dynamically monitoring the quantitative responses of phosphoproteome stimulated by EGF treatment in HeLa cells. In total, 5 361 unique phosphopeptides were identified, which reached a similar conclusion as others reported. The IBT reagents were therefore experimentally proven as a new type of reagents for isobaric peptides labeling and useful in a large quantity peptides of quantitative proteomics.

Synthesis and characterization of a new MeCAT reagent containing a photocleavable linker for labeling of proteins and peptides in mass spectrometric analyses

The quantification of proteins and peptides becomes more important besides mere identification in modern life sciences. Therefore, we have developed a new reagent that adds to the known metall-coded affinity tagging strategy employed in molecular and elemental mass spectrometry containing a photocleavable linker. A synthesis route was developed that provides the new reagent in good yields. The stability of the synthesized reagents was assessed under different temperature and illumination conditions. Labeling reactions were carried out at peptide and protein level, while also the fragmentation behavior of labeled peptides was assessed. In additional experiments, the photocleavability of the new reagent was examined. Upon irradiation with ultraviolet light, the photoproducts were liberated and could be used for quantification of labeled peptides.

Abstract P240: Identifying the Angiotensin AT2-Receptor Coupled Phosphoproteome in Human Aortic Endothelial Cells by Time-Resolved, Quantitative Phosphoproteomics

Objectives: The angiotensin AT2-receptor (AT2R) is a main component of the protective arm of the renin-angiotensin system. In order to get new and deeper insights into AT2R mediated signalling, this study mapped the AT2R-coupled phosphoproteome in human aortic endothelial cells (HAEC) by quantitative phosphoproteomics. Methods: HAEC were stimulated with the AT2R agonist C21 (1 μM) for 1, 3, 5 or 20 minutes. Following protein digestion and TMT peptide labelling, samples were subjected to TiO2–SIMAC phospho-peptide enrichment and LC–MS/MS. Data were analysed using Proteome Discoverer and the human sequence libraries SwissProt and UniProt. For confirmation of phosphoproteomics results, expression of p53 and phosphorylation of HDAC-1 were determined by Western Blot. Cellular translocation of HDAC-1 and p53 was assessed by immunofluorescence and activity of HDAC-1 by deacetylation assay. HAEC proliferation was determined by resazurin assay and apoptosis by caspase 3/7 assay. Results: AT2R stimulation of HAEC changed the phosphorylation status of 265 proteins at 471 sites (281 phosphorylations and 190 dephosphorylations). Gene Ontology and STRING analysis revealed the predominant phospho-modification of proteins involved in anti-proliferation, cell differentiation and apoptosis including HDAC-1, p53, BCL2 or HSP90. AT2R-mediated inhibition of HDAC-1 and activation of p53 were confirmed by additional experiments demonstrating changes in cellular localisation of HDAC-1 and p53, in total expression of p53, and in enzymatic activity and phosphorylation of HDAC1. Conclusion: This study mapped the AT2R coupled phospho-signalling network. It identified dephosphorylation and deactivation of HDAC-1 as well as nuclear importation and increased expression of p53 as novel, potential mediators of AT2R-coupled anti-proliferation.

Comparative Proteome Analysis Reveals Lipid Metabolism-Related Protein Networks in Response to Rump Fat Mobilization

Altay is a typical fat-tailed sheep breed displaying the unique ability to rapidly mobilize fat, which is vital for maintaining a normal metabolism that facilitates its survival in lengthy winter conditions. However, the physiological, biochemical, and molecular mechanisms underlying fat mobilization remain to be elucidated. In this study, the monitoring of rump fat adipocyte sizes disclosed a positive correlation between cell size and fat deposition ability. In addition, we subjected sheep to persistent starvation to imitate the conditions that trigger rump fat mobilization and screened 112 differentially expressed proteins using the isobaric peptide labeling approach. Notably, increased secretion of leptin and adiponectin activated the key fat mobilization signaling pathways under persistent starvation conditions. Furthermore, the upregulation of resistin (RETN), heat-shock protein 72 (HSP72), and complement factor D (CFD) promoted lipolysis, whereas the downregulation of cell death-inducing DFFA-like effector C (CIDEC) inhibited lipid droplet fusion, and the increase in HSP72 and apolipoprotein AI (Apo-AI) levels activated the body’s stress mechanisms. The synergistic actions of the above hormones, genes, and signaling pathways form a molecular network that functions in improving the adaptability of Altay sheep to extreme environments. Our findings provide a reference for elucidating the complex molecular mechanisms underlying rump fat mobilization.

Abstract 2708: Imputation-free analysis of high throughput TMT proteomics of 116 lung squamous samples

Abstract Introduction: Chemical labeling of peptides using tandem mass tags (TMT) is a “barcoding” strategy, enabling relative protein quantification across a single panel of samples (as opposed to each run separately). Each multiplex assay is, effectively, its own "batch" of samples, and thus direct comparison of intensities between TMT multiplexes is problematic. Additionally, although there is relatively little missing data within a single plex, there can be large differences in missingness across plexes, with the two types of missingness exhibiting different behavior (infrequent and biased towards low abundances within-plex; more frequent and more stochastic between-plex). We have addressed these issues by developing new pipelines for data normalization, protein-level rollup, and downstream clustering, which seek to minimize the negative impact of missingness. This method development was driven by, and applied to, a set of 116 human lung squamous (SQLC) tumors, with the aim of improving the strength of down-stream biological signal and interpretation. Experiment: TMT analysis was performed on 116 SQLC samples. Each 6-plex contained 4 tumors and 2 pool replicates. The shared pool of 116 tumors was assayed on every multiplex to allow for controlling for variability between plexes, with one pool in ch-126 and the other varying channel between plexes. IDPicker was used for spectral quantification. Spectra abundances were normalized within-plex, and ratios calculated for each channel against the ch-126 pool. Spectra-level ratios were rolled up into protein-level ratios using the geometric mean of ratios within each protein group. Geometric mean protein-level abundance rollup was performed on abundances for each ch-126 pool, normalized across pools, and the geometric mean calculated for each protein group across pools. These mean protein-level abundances were then used to scale the ratios back into final normalized abundances. Average linkage hierarchical clustering was performed on abundance z-scores using a novel distance metric, calculated as the root mean squared deviation (RMSD) of points present in both vectors, divided by a binary presence/absence similarity coefficient such as Ochiai similarity. Results: After normalization, principal component analysis showed no batch effect due to differences between plexes. Heat maps generated using the novel distance metric exhibited improved biological signal over RMSD alone. Tumors cluster into 3 major groupings: high immune + low transcriptional/translational activity, low immune + high transcriptional/translational activity, and samples with medium levels of both. Conclusion: Missingness-aware methods of shared-pool TMT normalization and clustering minimize the negative impact of missingness and yield strong biological signal. Preliminary results suggest that immune response is a major source of differences between lung squamous tumors. Citation Format: Eric A. Welsh, Paul A. Stewart, Matthew C. Chambers, Guolin Zhang, Bin Fang, Steven A. Eschrich, John M. Koomen, Eric B. Haura. Imputation-free analysis of high throughput TMT proteomics of 116 lung squamous samples [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2708.

Age-dependent alpha-synuclein accumulation and aggregation in the colon of a transgenic mouse model of Parkinson\u2019s disease

BackgroundParkinson’s disease (PD) is one of the most common neurodegenerative diseases, neuropathologically characterized by misfolded protein aggregation, called Lewy bodies and Lewy neurites. PD is a slow-progressive disease with colonic dysfunction appearing in the prodromal stage and lasting throughout the course of the disease.MethodsIn order to study PD pathology in the colon, we examined the age-dependent morphological and pathological changes in the colon of a PD mouse model expressing human wildtype α-synuclein (α-syn) fused with the green fluorescent protein (GFP), under the endogenous mouse α-syn promoter.ResultsWe observed an age-dependent progressive expression and accumulation of α-syn-GFP in the enteric neurons of Meissner’s (submucosal) and Auerbach’s (myenteric) plexuses of the colon. Additionally, the phosphorylation of α-syn at serine 129 also increased with age and the aggregation of α-syn-GFP coincided with the appearance of motor deficits at 9 months of age. Furthermore, α-syn (-GFP) distinctly co-localized with different subtypes of neurons, as identified by immunohistochemical labeling of vasoactive intestinal peptide (VIP), neuronal nitric oxide synthase (nNOS), and calretinin.ConclusionsOur results show the development of α-syn pathology in the enteric neurons of the colon in a PD mouse model, which coincide with the appearance of motor deficits. Our mouse model possesses the potential and uniqueness for studying PD gastrointestinal dysfunction.

Fluorophore labeling of a cell-penetrating peptide significantly alters the mode and degree of biomembrane interaction

The demand for highly efficient macromolecular drugs, used in the treatment of many severe diseases, is continuously increasing. However, the hydrophilic character and large molecular size of these drugs significantly limit their ability to permeate across cellular membranes and thus impede the drugs in reaching their target sites in the body. Cell-penetrating peptides (CPP) have gained attention as promising drug excipients, since they can facilitate drug permeation across cell membranes constituting a major biological barrier. Fluorophores are frequently covalently conjugated to CPPs to improve detection, however, the ensuing change in physico-chemical properties of the CPPs may alter their biological properties. With complementary biophysical techniques, we show that the mode of biomembrane interaction may change considerably upon labeling of the CPP penetratin (PEN) with a fluorophore. Fluorophore-PEN conjugates display altered modes of membrane interaction with increased insertion into the core of model cell membranes thereby exerting membrane-thinning effects. This is in contrast to PEN, which localizes along the head groups of the lipid bilayer, without affecting the thickness of the lipid tails. Particularly high membrane disturbance is observed for the two most hydrophobic PEN conjugates; rhodamine B or 1-pyrene butyric acid, as compared to the four other tested fluorophore-PEN conjugates.

Increasing the Efficiency of Amino Acids Detection By Electrochemical Methods on Amorphous Carbon Nitride a-CNx Electrodes

The naphthalene-2,3-dicarboxyaldehyde (NDA) label is widely used for the derivatization of amino acids (AAs) to form N-2-substituted-1-cyanobenz-[f]-isoindole derivatives (CBI) which are both fluorescent and electroactive. [1],[2] Optimal conditions for labeling were previously obtained when [NDA]tot/[CN-] ratio =1 and [NDA]tot /[amino acid or peptide]=40 with a buffer pH of 10. [3],[4] The detection step was achieved by using a carbon reference material (glassy carbon electrode ) exhibiting an obvious oxidation peak for CBI-AA observed at 0.5V/SCE [1]. These experiments confirmed that NDA can by used as a derivative agent for non electroactive AA, allowing for simultaneous electrochemical and fluorescence detections. In this work, amorphous carbon nitride a-CN0.26 thin films were elaborated on transparent and conductive glass/indium-tin oxide (ITO) wafers to improve the electroanalytical detection of AAs. The results obtained on as-grown a-CN0.26 were compared to the glassy carbon (GC). It was shown that a soft anodic pre-treatment protocol on glass/ITO/a-CN0.26 electrode in KCl solution drastically improved the CBI-AA oxidation peak performances. The oxidation peak potential for all CBI derivatives varied in the same range that those on GC and pre-treated glass/ITO/a-CN0.26, while no discrimination could be obtained for as-grown glass/ITO/a-CN0.26. For almost all the tested CBI derivatives, the electrochemical properties determined by electrochemical impedance spectroscopy (R ct, charge transfert resistance) and by differential pulse voltammetry (peak areas, full-widths at peak mid-height, peak current density and their SD values) were improved on pre-treated a-CN0.26 in comparison with GC. [1] M. Faure, S. Korchane, I. Le Potier, A. Pallandre, C. Deslouis, A.-M. Haghiri-Gosnet, J. Gamby, Investigating of labeling and detection of transthyretin synthetic peptide derivatized with naphthalene-2,3-dicarboxaldehyde, Talanta, 116 (2013) 8-13. [2] M. Faure, A. Pallandre, S. Chebil, I. Le Potier, M. Taverna, B. Tribollet, C. Deslouis, A.-M. Haghiri-Gosnet, J. Gamby, Improved electrochemical detection of a transthyretin synthetic peptide in the nanomolar range with a two-electrode system integrated in a glass/PDMS microchip, Lab Chip, 14 (2014) 2800-2805. [3] S. Korchane, A. Pallandre, C. Przybylski, C. Poas, F. Gonnet, M. Taverna, R. Daniel, I. Le Potier, Derivatization strategies for CE-LIF analysis of biomarkers: Toward a clinical diagnostic of familial transthyretin amyloidosis, Electrophoresis, 35 (2014) 1050-1059. [4] M. Faure, I. Le Potier, A. Pallandre, S. Chebil, A.-M. Haghiri-Gosnet, C. Deslouis, E. Maisonhaute, J. Gamby, Determination of the isomeric forms proportion of fluorogenic naphthalene-2,3-dicarboxaldehyde in a binary mixture of water:methanol using electrochemical methods, Talanta, 148 (2016) 494-501.

One-step labelling of a novel small-molecule peptide with astatine-211: preliminary evaluation in vitro and in vivo

In this paper, VP2, a novel small molecule fusion peptide, was labelled with 211At through a one-step method (coupled with bifunctional intermediate SPC first and then labelled) with a radiochemical yield of about 45%. The radiochemical purity was still > 95% after 24 h at room temperature. Specificity studies in vitro indicated that 211At-SPC-VP2 has a high affinity for several tumour cells. Additionally, biodistribution in KM mice showed that 211At-SPC-VP2 has sufficient stability in vivo. This research suggested that 211At–SPC-VP2 produced by the convenient method has the potential to be a novel targeted drug for cancer radiotherapy.

Nucleophilic 18F-Fluorination of Anilines via N-Arylsydnone Intermediates

Preparation of [18F]fluoroarenes with nucleophilic [18F]fluoride for positron emission tomography (PET) molecular imaging ­research is a challenging chemical endeavor. Advances in radiofluorination have soared in the last decade, broadening the availability of potential [18F]fluoroarenes. In this Synpacts article, we highlight the ­recent development from our laboratory of a practical radiofluorination of anilines via N-arylsydnone intermediates to afford [18F]fluoroarenes. Further, we emphasize the utility of this methodology towards peptide labeling applications by preparing an 18F-labeled neuropeptide.1 Introduction2 Nucleophilic Radiofluorination of N-Arylsydnones3 Substrate Scope and Applications4 Conclusion

18F-AlF Labeled Peptide and Protein Conjugates as Positron Emission Tomography Imaging Pharmaceuticals

The clinical applications of positron emission tomography (PET) imaging pharmaceuticals have increased tremendously over the past several years since the approval of 18fluorine-fluorodeoxyglucose (18F-FDG) by the Food and Drug Administration (FDA). Numerous 18F-labeled target-specific potential imaging pharmaceuticals, based on small and large molecules, have been evaluated in preclinical and clinical settings. 18F-labeling of organic moieties involves the introduction of the radioisotope by C-18F bond formation via a nucleophilic or an electrophilic substitution reaction. However, biomolecules, such as peptides, proteins, and oligonucleotides, cannot be radiolabeled via a C-18F bond formation as these reactions involve harsh conditions, including organic solvents, high temperature, and nonphysiological conditions. Several approaches, including 18F-labeled prosthetic groups, silicon, boron, and aluminum fluoride acceptor chemistry, and click chemistry have been developed, in the past, for 18F labeling of biomolecules. Linear and macrocyclic polyaminocarboxylates and their analogs and derivatives form thermodynamically stable and kinetically inert aluminum chelates. Hence, macrocyclic polyaminocarboxylates have been used for conjugation with biomolecules, such as folate, peptides, affibodies, and protein fragments, followed by 18F-AlF chelation, and evaluation of their targeting abilities in preclinical and clinical environments. The goal of this report is to provide an overview of the 18F radiochemistry and 18F-labeling methodologies for small molecules and target-specific biomolecules, a comprehensive review of coordination chemistry of Al3+, 18F-AlF labeling of peptide and protein conjugates, and evaluation of 18F-labeled biomolecule conjugates as potential imaging pharmaceuticals.

A Sulfoxide-Based Isobaric Labelling Reagent for Accurate Quantitative Mass Spectrometry.

Modern proteomics requires reagents for exact quantification of peptides in complex mixtures. Peptide labelling is most typically achieved with isobaric tags that consist of a balancer and a reporter part that separate in the gas phase. An ingenious distribution of stable isotopes provides multiple reagents with identical molecular weight but a different mass of the reporter groups, allowing relative quantification of multiple samples in one measurement. Here we report a new isobaric labelling reagent, where the balancer and the reporter are linked by a sulfoxide group, which, based on the sulfoxide pyrolysis, leads to easy and asymmetric cleavage at low fragmentation energy. The fragmentation of our new design is significantly improved, yielding more intense complementary ion signals, allowing complementary ion cluster analysis as well.

Highly Efficient and Stable Strain-Release Radioiodination for Thiol Chemoselective Bioconjugation.

We report a novel thiol selective radioiodination method based on strain-release reaction. A new heterobifunctional radioiodination agent which has very good thiol selectivity and excellent stability with high radiolabeling yield was synthesized, characterized, and applied successfully for thiol-contained peptide labeling.