Size-Dependent Cellular Uptake of RGD Peptides

Kemker I, Feiner R, Müller K, Sewald N. Size-Dependent Cellular Uptake of RGD Peptides. ChemBioChem. 2020;21(4):496-499.Monomeric RGD peptides show unspecific fluid-phase uptake in cells, whereas multimeric RGD peptides are thought to be internalized by integrin-mediated endocytosis. However, a potential correlation between uptake mechanism and molecular mass has been neglected so far. A dual derivatization of peptide c(RGDw(7Br)K) was performed to investigate this. A fluorescent probe was installed by chemoselective Suzuki-Miyaura cross-coupling of the 7-bromotryptophan and a poly(ethylene glycol) (PEG) linker was attached to the lysine residue. Flow cytometry and live cell imaging confirmed unspecific uptake of the small, non-PEGylated peptide, whereas the PEG(5000) peptide conjugate unveiled a selective internalization by M21 cells overexpressing alpha(v)beta(3) and no uptake in alpha(v)-deficient M21L cells

Synthesis of substituted cycloalkene-1,1-dicarboxylates via olefin metathesis in water

Tenbrink K, Kemker I, Schatz J, Gröger H. Synthesis of substituted cycloalkene-1,1-dicarboxylates via olefin metathesis in water. ARKIVOC. 2015;2015(2):10-19.A range of substituted cycloalkene-1,1-dicarboxylates was synthesized through olefin metathesis starting from readily available acylic malonate precursors in an efficient fashion. As a metathesis catalyst, a Grubbs II-type catalyst was used in these experiments, which were run in water and gave the cyclic malonate products with high conversions of 94-100%. The catalytic amount was in the range of 0.5-5 mol% dependent on the structure of the starting material. The generality of this metathesis reaction in water was demonstrated as well as its suitability for the preparation of five and six-membered and alkyl as well as aryl-substituted prochiral cycloalkene-1,1-dicarboxylates

Self-Assembly of the Bio-Surfactant Aescin in Solution. A Small-Angle X-ray Scattering and Fluorescence Study

Dargel C, Geisler R, Hannappel Y, Kemker I, Sewald N, Hellweg T. Self-Assembly of the Bio-Surfactant Aescin in Solution. A Small-Angle X-ray Scattering and Fluorescence Study. Colloids and Interfaces. 2019;3(2): 47.This work investigates the temperature-dependent micelle formation as well as the micellar structure of the saponin aescin. The critical micelle concentration ( cmc ) of aescin is determined from the concentration-dependent autofluorescence (AF) of aescin. Values between cmcaescin,AF (10 ∘ C) = 0.38 ± 0.09 mM and cmcaescin,AF (50 ∘ C) = 0.32 ± 0.13 mM were obtained. The significance of this method is verified by tensiometry measurements. The value determined from this method is within the experimental error identical with values obtained from autofluorescence ( cmcaescin,T(WP) (23 ∘ C) = 0.33 ± 0.02 mM). The structure of the aescin micelles was investigated by small-angle X-ray scattering (SAXS) at 10 and 40 ∘ C. At low temperature, the aescin micelles are rod-like, whereas at high temperature the structure is ellipsoidal. The radii of gyration were determined to ≈31 Å (rods) and ≈21 Å (ellipsoid). The rod-like shape of the aescin micelles at low temperature was confirmed by transmission electron microscopy (TEM). All investigations were performed at a constant pH of 7.4, because the acidic aescin has the ability to lower the pH value in aqueous solution

Synthesis and Biological Evaluation of RGD–Cryptophycin Conjugates for Targeted Drug Delivery

Borbély AN, Figueras Agustí E, Martins A, et al. Synthesis and Biological Evaluation of RGD–Cryptophycin Conjugates for Targeted Drug Delivery. Pharmaceutics. 2019;11(4): 151.Cryptophycins are potent tubulin polymerization inhibitors with picomolar antiproliferative potency in vitro and activity against multidrug-resistant (MDR) cancer cells. Because of neurotoxic side effects and limited efficacy in vivo, cryptophycin-52 failed as a clinical candidate in cancer treatment. However, this class of compounds has emerged as attractive payloads for tumor-targeting applications. In this study, cryptophycin was conjugated to the cyclopeptide c(RGDfK), targeting integrin αvβ3, across the protease-cleavable Val-Cit linker and two different self-immolative spacers. Plasma metabolic stability studies in vitro showed that our selected payload displays an improved stability compared to the parent compound, while the stability of the conjugates is strongly influenced by the self-immolative moiety. Cathepsin B cleavage assays revealed that modifications in the linker lead to different drug release profiles. Antiproliferative effects of Arg-Gly-Asp (RGD)–cryptophycin conjugates were evaluated on M21 and M21-L human melanoma cell lines. The low nanomolar in vitro activity of the novel conjugates was associated with inferior selectivity for cell lines with different integrin αvβ3 expression levels. To elucidate the drug delivery process, cryptophycin was replaced by an infrared dye and the obtained conjugates were studied by confocal microscop

Halotryptophan-containing RGD peptides: biological acitivity and late-stage diversification

Kemker I. Halotryptophan-containing RGD peptides: biological acitivity and late-stage diversification. Bielefeld: Universität Bielefeld; 2019

One-Pot Synthesis of D-Halotryptophans by Dynamic Stereoinversion Using a Specific L-Amino Acid Oxidase

Schnepel C, Kemker I, Sewald N. One-Pot Synthesis of D-Halotryptophans by Dynamic Stereoinversion Using a Specific L-Amino Acid Oxidase. ACS Catalysis. 2019;9(2):1149-1158.Tryptophan (Trp) derivatives constitute important building blocks found in many natural products, peptides, and drugs. Accordingly, there is a high demand for suitable biocatalytic pathways providing selective derivatization of Trp like C-H functionalization or C-alpha-oxidation. The specific l-amino acid oxidase RebO from the actinomycete L. aerocolonigenes was harnessed for chemoenzymatic oxidation of substituted Trp derivatives. An array of potential substrates was tested, and several Trp derivatives are being accepted with reasonable turnover. The highest selectivity was observed for 7-halotryptophan being converted about 35-fold faster than nonsubstituted Trp. This selectivity is also useful for establishing a colorimetric halogenase high-throughput assay. RebO was also employed in dynamic stereoinversion in the presence of an ammonia-borane complex to provide access to non-native D-configured Trp analogues. Optimized reaction conditions yielded similar to 70% D-amino acid with >98% ee for halotryptophans on an analytical scale. Dynamic stereoinversion preceded by enzymatic halogenation in a sequential one-pot reaction cascade provided D-configured 5- or 7-bromotryptophan with improved conversion of approximately 90%, >92% ee, and good isolated yields

Bioactive fluorescent cyclopeptides through on-resin Suzuki cross-coupling reaction

Ghabraie E, Tonali NM, Kemker I, Sewald N. Bioactive fluorescent cyclopeptides through on-resin Suzuki cross-coupling reaction. In: JOURNAL OF PEPTIDE SCIENCE. Vol 24. Hoboken: Wiley; 2018: S89

Cyclization of RGD Peptides by Suzuki-Miyaura Cross-Coupling

Kemker I, Schnepel C, Schröder DC, Marion A, Sewald N. Cyclization of RGD Peptides by Suzuki-Miyaura Cross-Coupling. Journal of Medicinal Chemistry. 2019;62(16):7417-7430.Halogenated L- or D-tryptophan obtained by biocatalytic halogenation was incorporated into RGD peptides together with a variety of alkyl or aryl boronic acids. Suzuki-Miyaura cross-coupling either in solution or on-resin results in side chain-to-tail-cyclized RGD peptides, for example, with biaryl moieties, providing a new dimension of structure-activity relationships. An array of RGD peptides differing in macrocycle size, the presence of D-amino acid, N-methylation, or connectivity between the indole moiety and the boronic acid showed that, in particular, connectivity exhibits a major impact on affinities toward integrins, for example, a(v)beta(3). Structure-activity relationship studies yielded peptides with affinities toward a(v)beta(3) in the low nanomolar range, good selectivity, and high plasma stability. Structural characteristics of representative molecules have been investigated by molecular dynamics simulations, which allowed understanding the observed activity differences

Cyclization of RGD Peptides by Suzuki-Miyaura Cross-Coupling

Halogenated L- or D-tryptophan obtained by biocatalytic halogenation was incorporated into RGD peptides together with a variety of alkyl or aryl boronic acids. Suzuki-Miyaura cross-coupling either in solution or on-resin results in side chain-to-tail-cyclized RGD peptides, for example, with biaryl moieties, providing a new dimension of structure-activity relationships. An array of RGD peptides differing in macrocycle size, the presence of D-amino acid, N-methylation, or connectivity between the indole moiety and the boronic acid showed that, in particular, connectivity exhibits a major impact on affinities toward integrins, for example, a(v)beta(3). Structure-activity relationship studies yielded peptides with affinities toward a(v)beta(3) in the low nanomolar range, good selectivity, and high plasma stability. Structural characteristics of representative molecules have been investigated by molecular dynamics simulations, which allowed understanding the observed activity differences

Self-Assembly of the Bio-Surfactant Aescin in Solution: A Small-Angle X-ray Scattering and Fluorescence Study

This work investigates the temperature-dependent micelle formation as well as the micellar structure of the saponin aescin. The critical micelle concentration ( c m c ) of aescin is determined from the concentration-dependent autofluorescence (AF) of aescin. Values between c m c aescin , AF (10 ∘ C) = 0.38 ± 0.09 mM and c m c aescin , AF (50 ∘ C) = 0.32 ± 0.13 mM were obtained. The significance of this method is verified by tensiometry measurements. The value determined from this method is within the experimental error identical with values obtained from autofluorescence ( c m c aescin , T ( WP ) (23 ∘ C) = 0.33 ± 0.02 mM). The structure of the aescin micelles was investigated by small-angle X-ray scattering (SAXS) at 10 and 40 ∘ C. At low temperature, the aescin micelles are rod-like, whereas at high temperature the structure is ellipsoidal. The radii of gyration were determined to ≈31 Å (rods) and ≈21 Å (ellipsoid). The rod-like shape of the aescin micelles at low temperature was confirmed by transmission electron microscopy (TEM). All investigations were performed at a constant pH of 7.4, because the acidic aescin has the ability to lower the pH value in aqueous solution