Mechanism based heparanase inhibitors reduce cancer metastasis in vivo

Heparan sulfate proteoglycans (HSPGs) mediate essential interactions throughout the extracellular matrix (ECM), providing signals that regulate cellular growth and development. Altered HSPG composition during tumorigenesis strongly aids cancer progression. Heparanase (HPSE) is the principal enzyme responsible for extracellular heparan sulfate catabolism and is markedly up-regulated in aggressive cancers. HPSE overactivity degrades HSPGs within the ECM, facilitating metastatic dissemination and releasing mitogens that drive cellular proliferation. Reducing extracellular HPSE activity reduces cancer growth, but few effective inhibitors are known, and none are clinically approved. Inspired by the natural glycosidase inhibitor cyclophellitol, we developed nanomolar mechanism-based, irreversible HPSE inhibitors that are effective within physiological environments. Application of cyclophellitol-derived HPSE inhibitors reduces cancer aggression in cellulo and significantly ameliorates murine metastasis. Mechanism-based irreversible HPSE inhibition is an unexplored anticancer strategy. We demonstrate the feasibility of such compounds to control pathological HPSE-driven malignancies.NWO"Endoglycoprobe”714.018.002Bio-organic Synthesi

Effective Melanoma Immunotherapy in Mice by the Skin-Depigmenting Agent Monobenzone and the Adjuvants Imiquimod and CpG

Background: Presently melanoma still lacks adequate treatment options for metastatic disease. While melanoma is exceptionally challenging to standard regimens, it is suited for treatment with immunotherapy based on its immunogenicity. Since treatment-related skin depigmentation is considered a favourable prognostic sign during melanoma intervention, we here aimed at the reverse approach of directly inducing vitiligo as a shortcut to effective anti-melanoma immunity. Methodology and Principal Findings: We developed an effective and simple to use form of immunotherapy by combining the topical skin-bleaching agent monobenzone with immune-stimulatory imiquimod cream and cytosine-guanine oligodeoxynucleotides (CpG) injections (MIC therapy). This powerful new approach promptly induced a melanoma antigen-specific immune response, which abolished subcutaneous B16. F10 melanoma growth in up to 85% of C57BL/6 mice. Importantly, this regimen induced over 100 days of tumor-free survival in up to 60% of the mice, and forcefully suppressed tumor growth upon re-challenge either 65- or 165 days after MIC treatment cessation. Conclusions: MIC therapy is effective in eradicating melanoma, by vigilantly incorporating NK-, B-and T cells in its therapeutic effect. Based on these results, the MIC regimen presents a high-yield, low-cost and simple therapy, readily applicable in the clini

Functionalization of Mono- and Oligonucleotides with Phosphane Ligands by Amide Bond Formation

Seven phosphane-functionalized deoxyuridines have been prepared by amide bond formation between aminodeoxyuridines and phosphanylcarboxylic acids. X-ray crystal structures for two of these new modified nucleosides have been obtained. The same coupling method has been extended to oligonucleotides. The phosphane containing strands have been purified and characterized by MALDI-TOF and, for the first time, P-31 NMR spectrometry. Coordination of a phosphane-modified 15-mer to a [PdCl(eta(3)-allyl)] moiety has been confirmed by P-31 NMR spectroscopy.</p

Phosphine containing oligonucleotides for the development of metallodeoxyribozymes

Novel transition metal catalysts based on oligonucleotides can be easily obtained by functionalization of 5-iodouridine with phosphine ligands, resulting in good asymmetric induction in palladium catalyzed allylic nucleophilic substitution.</p

C-mannosyl lysine for solid phase assembly of mannosylated peptide conjugate cancer vaccines

Dendritic cells (DCs) are armed with a multitude of Pattern Recognition Receptors (PRRs) to recognize pathogens and initiate pathogen-tailored T cell responses. In these responses, the maturation of DCs is key, as well as the production of cytokines that help to accomplish T cell responses. DC-SIGN is a frequently exploited PRR that can effectively be targeted with mannosylated antigens to enhance the induction of antigen-specific T cells. The natural O-mannosidic linkage is susceptible to enzymatic degradation, and its chemical sensitivity complicates the synthesis of mannosylated antigens. For this reason, (oligo)mannosides are generally introduced in a late stage of the antigen synthesis, requiring orthogonal conjugation handles for their attachment. To increase the stability of the mannosides and streamline the synthesis of mannosylated peptide antigens, we here describe the development of an acid-stable C-mannosyl lysine, which allows for the inline introduction of mannosides during solid-phase peptide synthesis (SPPS). The developed amino acid has been successfully used for the assembly of both small ligands and peptide antigen conjugates comprising an epitope of the gp100 melanoma-associated antigen and a TLR7 agonist for DC activation. The ligands showed similar internalization capacities and binding affinities as the O-mannosyl analogs. Moreover, the antigen conjugates were capable of inducing maturation, stimulating the secretion of pro-inflammatory cytokines, and providing enhanced gp100 presentation to CD8 + and CD4 + T cells, similar to their O-mannosyl counterparts. Our results demonstrate that the C-mannose lysine is a valuable building block for the generation of anticancer peptide-conjugate vaccine modalities

The Optimization of Bioorthogonal Epitope Ligation within MHC‑I Complexes

Antigen recognition followed by the activation of cytotoxic T-cells (CTLs) is a key step in adaptive immunity, resulting in clearance of viruses and cancers. The repertoire of peptides that have the ability to bind to the major histocompatibility type-I (MHC-I) is enormous, but the approaches available for studying the diversity of the peptide repertoire on a cell are limited. Here, we explore the use of bioorthogonal chemistry to quantify specific peptide–MHC-I complexes (pMHC-I) on cells. We show that modifying epitope peptides with bioorthogonal groups in surface accessible positions allows wild-type-like MHC-I binding and bioorthogonal ligation using fluorogenic chromophores in combination with a Cu­(I)-catalyzed Huisgen cycloaddition reaction. We expect that this approach will make a powerful addition to the antigen presentation toolkit as for the first time it allows quantification of antigenic peptides for which no detection tools exist

Self-Adjuvanting Cancer Vaccines from Conjugation-Ready Lipid A Analogues and Synthetic Long Peptides

Self-adjuvanting vaccines, wherein an antigenic peptide is covalently bound to an immunostimulating agent, have been shown to be promising tools for immunotherapy. Synthetic Toll-like receptor (TLR) ligands are ideal adjuvants for covalent linking to peptides or proteins. We here introduce a conjugation-ready TLR4 ligand, CRX-527, a potent powerful lipid A analogue, in the generation of novel conjugate-vaccine modalities. Effective chemistry has been developed for the synthesis of the conjugation-ready ligand as well as the connection of it to the peptide antigen. Different linker systems and connection modes to a model peptide were explored, and in vitro evaluation of the conjugates showed them to be powerful immune-activating agents, significantly more effective than the separate components. Mounting the CRX-527 ligand at the N-terminus of the model peptide antigen delivered a vaccine modality that proved to be potent in activation of dendritic cells, in facilitating antigen presentation, and in initiating specific CD8+ T-cell-mediated killing of antigen-loaded target cells in vivo. Synthetic TLR4 ligands thus show great promise in potentiating the conjugate vaccine platform for application in cancer vaccination.Bio-organic Synthesi