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Review
. 2022 Oct 17;51(20):8756-8783.
doi: 10.1039/d2cs00640e.

Multivalent glycocyclopeptides: conjugation methods and biological applications

David Goyard  1 , Angela Martin-Serrano Ortiz  1 , Didier Boturyn  1 , Olivier Renaudet  1
Affiliations
Review

Multivalent glycocyclopeptides: conjugation methods and biological applications

David Goyard et al. Chem Soc Rev. .

Abstract

Click chemistry was extensively used to decorate synthetic multivalent scaffolds with glycans to mimic the cell surface glycocalyx and to develop applications in glycosciences. Conjugation methods such as oxime ligation, copper(I)-catalyzed alkyne-azide cycloaddition, thiol-ene coupling, squaramide coupling or Lansbury aspartylation proved particularly suitable to achieve this purpose. This review summarizes the synthetic strategies that can be used either in a stepwise manner or in an orthogonal one-pot approach, to conjugate multiple copies of identical or different glycans to cyclopeptide scaffolds (namely multivalent glycocyclopeptides) having different size, valency, geometry and molecular composition. The second part of this review will describe the potential of these structures to interact with various carbohydrate binding proteins or to stimulate immunity against tumor cells.

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Conflict of interest statement

There are no conflicts to declare.

Figures

Scheme 1
Scheme 1. Strategies for the synthesis of aminooxy glycans 1–11: (A) using phase transfer catalysis from glycosyl bromides; (B) using glycosylation from glycosyl fluorides. Reagents and conditions: (i) NHS, TBAHS, CH2Cl2, Na2CO3 1 M; (ii) PhthOH, BF3·Et2O, TEA, CH2Cl2. Abbreviations: NHS: N-hydroxysuccinimide; TBAHS: tetra-butyl-ammonium-hydrogen-sulfate; PhthOH: N-hydroxyphthalimide; TEA: triethylamine; Gal: galactose; Glc: glucose; GalNAc: N-acetylgalactosamine; Man: mannose, Fuc: fucose, Lac: lactose; TF: Thomsen–Friedenreich.
Scheme 2
Scheme 2. Example of methods to incorporate an aldehyde or a ketone function into a peptide sequence: (A) using an enzymatic process; (B) using the incorporation of a keto-amino acid; (C) using a periodic acid mediated serine oxidation. Reagents and conditions: (i) NaIO4, H2O. Abbreviations: FGE: formyl-generating enzyme; SPPS: solid-phase peptide synthesis.
Scheme 3
Scheme 3. Assembly of tetravalent glycoclusters 18–22 by oxime ligation. Reagents and conditions: (i) NaIO4, H2O, r.t., 30 min, 83%; (ii) AcONa (pH 4), r.t., 1–24 h, 70–80%.
Scheme 4
Scheme 4. Assembly of glycoclusters by CuAAC: (A) from the tetravalent scaffold 23; (B) from the hexavalent scaffold 30. Reagents and conditions: (i) Cu micropowder, tBuOH/AcONH4 (pH 7.4), r.t., 3 h, 70–75%; (ii) CuSO4·5H2O, THPTA, sodium ascorbate, PBS (pH 7.4), r.t., 1 h, 85%.
Scheme 5
Scheme 5. Synthesis of glycoclusters using squaramide coupling. Reagents and conditions: (i) 33, NaHCO3, H2O/CH3OH; (ii) TFA/CH2Cl2; (iii) 34, NaHCO3, H2O/CH3OH, approx. 70%.
Scheme 6
Scheme 6. On-bead synthesis of a glycosyl carbamate-cyclopeptide 38 using glycosyl p-nitrophenyl carbonate 37. Reagent and conditions: 37, DIPEA, NMP; (ii) TFA/TIS/CH2Cl2 (1 : 1:98); (iii) NaOMe, MeOH/CHCl3, 15%. Abbreviations: DIPEA: diisopropylethylamine; NMP: N-methylmorpholine.
Scheme 7
Scheme 7. Chemoenzymatic synthesis of a trivalent GM3-based cyclopeptide as blocker of influenza virus hemagglutinin. Reagents and conditions: (i) 40, Transglutaminase; (ii) α-2,3-(N)-sialyltransferase, CMP-Neu5Ac.
Scheme 8
Scheme 8. Conjugation of high-mannose glycan 43 by Lansbury aspartylation. Reagents and conditions: (i) HATU/DIPEA/DMSO then 43. Abbreviations: HATU: hexafluorophosphate azabenzotriazole tetramethyl uronium; DMSO: dimethylsulfoxide.
Scheme 9
Scheme 9. Synthesis of di-, tri- and tetravalent glycocyclopeptides 47 using a one-pot oligomerization/cyclization protocol. Reagents and conditions: (i) DPPA, TEA, DMF, 0 °C.
Scheme 10
Scheme 10. Functionalization of the second domain of a cyclopeptide scaffold 50 using successive oxime ligation. Reagents and conditions: (i) 0.1% TFA in CH3CN/H2O (1 : 1), 37 °C, o/n then NaIO4, H2O, r.t., 1 h, 95%; (ii) 0.1% TFA in CH3CN/H2O (1 : 1), 37 °C, o/n, 80%.
Scheme 11
Scheme 11. Extension of the cyclopeptide valency from 16 to 64 by iterative oxime ligation. Reagents and conditions: (i) 0.1% TFA in H2O, 37 °C, 4 h, acetone quenching; (ii) NaIO4, H2O, r.t., 30 min then 0.1% TFA in H2O, 37 °C, 4 h, 65–88%; (iii) 0.1% TFA in H2O, 37 °C, 1 h, acetone quenching then NaIO4, H2O, r.t., 30 min, 88%; (iv) 0.1% TFA in H2O, 37 °C, 4 h, 83–85%.
Scheme 12
Scheme 12. Synthesis of fucosylated and sialylated heteroglycocluster 65. Reagents and conditions: (i) CuSO4·5H2O, THPTA, sodium ascorbate, DMF/PBS (pH 7.5) (1 : 1), r.t., 1 h, 79% for 63, 85% for 64, 71% for 65; (ii) N-succinimidyl azidoacetate or N-succinimidyl pentynoate, DIPEA, DMF, r.t., 2 h, quant.; (iii) LiOH, r.t., 2 h, quant.
Scheme 13
Scheme 13. Structure of an oligomannose-based glycocluster prepared by successive CuAAC reactions. Reagents and conditions: (i) 67, CuSO4, sodium ascorbate, t-BuOH/H2O (1 : 1), approx. 90%; (ii) 69, 0.05 M NaHCO3, CH3CN/MeOH (1 : 1); (iii) 70, CuSO4, sodium ascorbate, t-BuOH/H2O (1 : 1), 70%.
Scheme 14
Scheme 14. Synthesis of 2 : 2 tetravalent heteroglycoclusters using OL and CuAAC. Reagents and conditions: (i) 0.1% TFA in H2O; (ii) Cu micropowder, t-BuOH, AcONH4 100 mM pH 7.4 (1 : 1, v/v), conv. rate 83–93% (calculated from the crude analytical HPLC profile).
Scheme 15
Scheme 15. Convergent synthesis of the hexadecavalent GalNAc glycodendrimer 78 by CuAAC/OL. Reagents and conditions: (i) CuSO4·5H2O, THPTA, sodium ascorbate, DMF/PBS buffer (pH 7.5) (1 : 1), r.t., 2 h, 70% (divergent synthesis); (ii) TFA/H2O (6 : 4), r.t., 1 h, 77% over two steps for 77; (iii) 51, 0.1% TFA in H2O, 37 °C, 45 min, 90% for 78; 85% for 79.
Scheme 16
Scheme 16. Synthesis of heterovalent glycodendrimers as epitope carriers for antitumor vaccines. Reagents and conditions: (i) 0.1% TFA in H2O/CH3CN (1 : 1), 37 °C, 30 min, 75%; (ii) NaIO4, H2O, r.t., 40 min, 70%; (iii) 0.1% TFA in H2O/CH3CN (1 : 1), 37 °C, 30 min, 85%; (iv) NaIO4, H2O, r.t., 40 min, 78%; (v) 0.1% TFA in H2O/CH3CN (1 : 1), 37 °C, 30 min; (vi) CuSO4, THPTA, sodium ascorbate, PBS (pH 7.4, 10 mM), r.t., 90 min, 55% (over two steps).
Scheme 17
Scheme 17. Synthesis of heteroglycoclusters by thiol–ene and thiol–chloroacetyl couplings. (A) Functionalization of the bottom domain as the first step; (B) functionalization of the upper domain as the first step. Reagents and conditions: (i) NaH, DMF, r.t., 1 h; (ii) DPAP, DMF/H2O, irrad. 365 nm, r.t., 45 min, 78% for 88, 77% for 89 over two steps.
Scheme 18
Scheme 18. Synthesis of heterovalent structures using CuAAC, TEC and SqC. Reagents and conditions: (i) DPA, DMF/H2O, irrad. 365 nm, r.t., 45 min; (ii) Cu micropowder, PBS (pH 7.4), r.t., 45 min, 71% over two steps; (iii) DMF, DIPEA (pH 8.5), r.t., 4 h, 87%; (iv) 91, Na2CO3 (pH 9.5), 37 °C, 24 h, 67%.
Scheme 19
Scheme 19. Sequential one-pot synthesis of the heterovalent glycocluster 95 using four conjugation strategies and assembly of the glycodendrimer 96. Reagents and conditions: (i) 0.1% TFA in H2O, r.t., 30 min; (ii) DPAP, DMF/H2O, irrad. 365 nm, r.t., 30 min; (iii) CuSO4, THPTA, sodium ascorbate, DMF/PBS (1 : 1, pH 7.4), r.t., 1 h; (iv) KI, DMF/H2O (1 : 1), DIPEA, r.t., 1 h; 47% over four steps; (v) N-succinimidyl (Boc-aminooxy)acetate, DIPEA, DMF; (vi) 50% TFA in CH2Cl2, r.t., 30 min, 93% over two steps; (vii) 51, 0.1% TFA in H2O, 37 °C, 30 min, 87%.
Fig. 1
Fig. 1. General structure of tetravalent ligands designed to target hepatocytes. Abbreviation: lys: d-lysine residue.
Fig. 2
Fig. 2. Structure of hexavalent glycoclusters displaying fucose (100) or a fucose mimetic (101).
Fig. 3
Fig. 3. Structures of tetra-, hexa- and hexadecavalent fucosylated conjugates tested with LecB.
Fig. 4
Fig. 4. Structures of hGCs 114–117 with dual affinities for LecA and LecB.
Fig. 5
Fig. 5. Structures of tetracosavalent mannosylated glycodendrimers 118–121 evaluated as BC2L-A ligands.
Fig. 6
Fig. 6. Structure of the bis-galactoside inhibitor 122 of Candida albicans adhesion and its multivalent conjugates 123–125.
Fig. 7
Fig. 7. (A) Immobilization of glycoclusters by oxime ligation (1) and incubation with a fluorescent lectin (2); (B) direct (1) and indirect (2) approach for the preparation of glycocluster microarray; (C) glycodendrimer microarray for screening and measuring binding constants.
Fig. 8
Fig. 8. (A) Structure of two and four component antitumoral fully synthetic vaccine candidates based on cyclopeptide scaffolds (R1: Tn analogue; R2: PV peptide; R3: palmitoyl-OVA peptide; R4: PADRE peptide); (B) structure of an antitumoral vaccine candidate using a Tn mimetic (R1: Tn mimetic; R2: PADRE peptide; R3: OVA peptide).
Fig. 9
Fig. 9. Homo- and heterovalent glycosylated platforms as carrier for antitumoral synthetic vaccines.
Fig. 10
Fig. 10. Structure of tetra- and hexadecavalent vaccine candidates based on α-GalNAc or the native Tn antigen.
Fig. 11
Fig. 11. Structure of the Antibody Binding Modules 136–137 and the Tumor Binding Module 138.
Fig. 12
Fig. 12. Structure of Antibody-Recruiting Glycodendrimers to stimulate immune-mediated cytotoxicity against cancer cells.
None
David Goyard
None
Angela Martin-Serrano Ortiz
None
Didier Boturyn
None
Olivier Renaudet
See this image and copyright information in PMC

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