Selektiv funktionalisierbare Glycopeptide und Glycodendrimere in biologischen Testsystemen

Glycopeptides and dendrimer-functionalized peptides were synthesized. These derivatives were labeled with ruthenium bipyridyl complexes and coupled with HIV-haptenes for utilisation in luminescence immunoassays. Solid phase peptide synthesis was carried out applying the Fmoc-solid phase strategy. In addition the synthesis of selectively functionalized AnX-type dendrimers is described. The coupling of peptides with dendritic residues as well as biotinylation of these compounds was performed in solution. The AnX-type dendrimers were coupled with mannose and fucose derivatives as biologically relevant carbohydrate epitopes and thus were converted into glycodendrimers. By this approach multivalent mannose and fucose clusters could be obtained to serve as oligosaccharide mimics. Fucose clusters were functionalized using alkyl and oligo ethylene glycol thioacetates. These compounds could be immobilized on gold surfaces in self assembled monolayers. N-glycosides were synthesized from anomeric azides by a modified Staudinger reaction. In doing so building blocks were obtained, which can be used for the synthesis of glycopeptido mimetics

CD38-Specific Biparatopic Heavy Chain Antibodies Display Potent Complement-Dependent Cytotoxicity Against Multiple Myeloma Cells

CD38 is overexpressed by multiple myeloma cells and has emerged as a target for therapeutic antibodies. Nanobodies are soluble single domain antibody fragments derived from the VHH variable domain of heavy chain antibodies naturally occurring in camelids. We previously identified distinct llama nanobodies that recognize three non-overlapping epitopes of the extracellular domain of CD38. Here, we fused these VHH domains to the hinge, CH2, and CH3 domains of human IgG1, yielding highly soluble chimeric llama/human heavy chain antibodies (hcAbs). We analyzed the capacity of these hcAbs to mediate complement-dependent cytotoxicity (CDC) to CD38-expressing human multiple myeloma and Burkitt lymphoma cell lines. Combinations of two hcAbs that recognize distinct, non-overlapping epitopes of CD38 mediated potent CDC, in contrast to the hcAb monotherapy with only weak CDC capacity. Similarly, combining daratumumab with a hcAb that recognizes a non-overlapping epitope resulted in dramatically enhanced CDC. Further, introducing the E345R HexaBody mutation into the CH3 domain strongly enhanced the CDC potency of hcAbs to CD38-expressing cells. Exploiting their high solubility, we genetically fused two distinct nanobodies into heteromeric dimers via a flexible peptide linker and then fused these nanobody dimers to the hinge, CH2 and CH3 domains of human IgG1, yielding highly soluble, biparatopic hcAbs. These biparatopic hcAbs elicited CDC toward CD38-expressing myeloma cells more effectively than daratumumab. Our results underscore the advantage of nanobodies vs. pairs of VH and VL domains for constructing bispecific antibodies. Moreover, the CD38-specific biparatopic heavy chain antibodies described here represent potential new powerful therapeutics for treatment of multiple myeloma

A glycopeptide dendrimer inhibitor of the galactose specific lectin LecA & of Pseudomonas aeruginosa biofilms

Biofilm inhibition is achieved with a phenylgalactosyl peptide dendrimer (see picture) that binds to the galactose-specific lectin LecA of P. aeruginosa. The multivalency of the ligands is critical for biofilm inhibition, although the nature of the linker between the peptide dendrimer and the galactose can provide additional contacts to the lectin and also has an effect on the interaction

Dendritic glycopolymers based on dendritic polyamine scaffolds: view on their synthetic approaches, characteristics and potential for biomedical applications

In this review we highlight the potential for biomedical applications of dendritic glycopolymers based on polyamine scaffolds. The complex interplay of the molecular characteristics of the dendritic architectures and their specific interactions with various (bio)molecules are elucidated with various examples. A special role of the individual sugar units attached to the dendritic scaffolds and their density is identified, which govern ionic and H-bond interactions, and biological targeting, but to a large extent are also responsible for the significantly reduced toxicity of the dendritic glycopolymers compared to their polyamine scaffolds. Thus, the application of dendritic glycopolymers in drug delivery systems for gene transfection but also as therapeutics in neurodegenerative diseases has great promisePublikacja w ramach programu Royal Society of Chemistry "Gold for Gold" 2014 finansowanego przez Uniwersytet Łódzk

Design of Membrane Active Peptides Considering Multi-Objective Optimization for Biomedical Application

A multitude of membrane active peptides exists that divides into subclasses, such as cell penetrating peptides (CPPs) capable to enter eukaryotic cells or antimicrobial peptides (AMPs) able to interact with prokaryotic cell envelops. Peptide membrane interactions arise from unique sequence motifs of the peptides that account for particular physicochemical properties. Membrane active peptides are mainly cationic, often primary or secondary amphipathic, and they interact with membranes depending on the composition of the bilayer lipids. Sequences of these peptides consist of short 5–30 amino acid sections derived from natural proteins or synthetic sources. Membrane active peptides can be designed using computational methods or can be identified in screenings of combinatorial libraries. This review focuses on strategies that were successfully applied to the design and optimization of membrane active peptides with respect to the fact that diverse features of successful peptide candidates are prerequisites for biomedical application. Not only membrane activity but also degradation stability in biological environments, propensity to induce resistances, and advantageous toxicological properties are crucial parameters that have to be considered in attempts to design useful membrane active peptides. Reliable assay systems to access the different biological characteristics of numerous membrane active peptides are essential tools for multi-objective peptide optimization

Molecular evolution of peptide ligands with custom-tailored characteristics for targeting of glycostructures.

As an advanced approach to identify suitable targeting molecules required for various diagnostic and therapeutic interventions, we developed a procedure to devise peptides with customizable features by an iterative computer-assisted optimization strategy. An evolutionary algorithm was utilized to breed peptides in silico and the "fitness" of peptides was determined in an appropriate laboratory in vitro assay. The influence of different evolutional parameters and mechanisms such as mutation rate, crossover probability, gaussian variation and fitness value scaling on the course of this artificial evolutional process was investigated. As a proof of concept peptidic ligands for a model target molecule, the cell surface glycolipid ganglioside G(M1), were identified. Consensus sequences describing local fitness optima were reached from diverse sets of L- and proteolytically stable D lead peptides. Ten rounds of evolutional optimization encompassing a total of just 4400 peptides lead to an increase in affinity of the peptides towards fluorescently labeled ganglioside G(M1) by a factor of 100 for L- and 400 for D-peptides