Discovery of novel glycomimetic ligands for Siglec-8 : doctoral dissertation

Girardi, Benedetta (author), Mravljak, Janez (mentor), Schwardt, Oliver (co-mentor)

This thesis principally focuses on the development of glycomimetic ligands with improved affinity to Siglec-8. Siglec-8 is a member of the Siglec family, I-type lectins that contain a sialic acid-binding domain and that play different roles in cell-cell interactions and cell signaling. Siglec-8 is a CD33- related protein expressed exclusively on mast cells and eosinophils, and weakly on basophils. Crosslinking of Siglec-8 with antibodies leads to apoptosis of eosinophils and inhibition of degranulation of mast cells. Apoptosis has also been induced when eosinophils were treated with a glycopolymer (6’-sulfo-sLex-polyacrylamide polymer). Therefore, Siglec-8 represents a very interesting new pharmacological target for the treatment of eosinophil- or mast cell-associated diseases, such as asthma, chronic rhinosinusitis, chronic urticaria, hypereosinophilic syndromes, mast cell and eosinophil malignancies, and eosinophilic gastrointestinal disorders. Recently, the NMR solution structure of the Siglec-8 lectin domain was solved in complex with its preferred ligand, the tetrasaccharide 6′-sulfo sialyl Lewisx (Neu5Acα2-3[6S]Galβ1-4[Fucα1-3]GlcNAc). The main interactions involve a salt bridge between Arg109 and the carboxylate of sialic acid and a second salt bridge between Arg56 and Gln59 and the sulfate at the 6-position of the galactose moiety. Interestingly, the sulfate plays a key role in both binding and specificity to Siglec-8. In addition, hydrogen bonds exist between hydroxyl groups 7, 8 and 9 of sialic acid and Tyr7, Ser118 and Gln122. In contrast, the fucose and glucosamine subunits show only minor interactions. This led to the suggestion that the disaccharide substructure 6-sulfo-Sia-Gal might represent the minimal binding epitope for Siglec-8.

• Language: English
• Work type: Dissertation
• Typology: 2.08 - Doctoral Dissertation
• Organization: FFA - Faculty of Pharmacy
• Place of publishing: Ljubljana
• Publisher: [B. M. Girardi]
• Year: 2022
• Number of pages: 221 str.
• PID: 20.500.12556/RUL-143829
• UDC: 615.4:54:576.36(043.3)
• COBISS.SI-ID: 105504003
• Publication date in RUL: 13.01.2023

Secondary language

• Language: Slovenian

Abstract

Ta doktorska naloga se osredotoča predvsem na razvoj glikomimetičnih ligandov z izboljšano afiniteto do Siglec-8. Siglec-8 je član družine Siglec, lektinov tipa I, ki vsebujejo domeno, ki veže sialično kislino, in imajo različne vloge pri interakcijah med celicami in celični signalizaciji. Siglec- 8 je CD33 sorodni protein, izražen izključno v tkivnih bazofilcih (mastocitih) in eozinofilcih, šibko pa v bazofilcih. Povezovanje Siglec-8 s protitelesi vodi do apoptoze eozinofilcev in zaviranja degranulacije mastocitov. Apoptoza se je pojavila tudi pri tretiranju eozinofilcev z glikopolimerom (6'-sulfo-sLex-poliakrilamidnim polimerom). Zato Siglec-8 predstavlja zelo zanimivo novo farmakološko tarčo za zdravljenje bolezni, povezanih z eozinofilci ali mastociti, kot so astma, kronični rinosinuzitis, kronična urtikarija, hipereozinofilni sindromi, malignosti mastocitov in eozinofilcev ter eozinofilne gastrointestinalne motnje. Pred kratkim so razrešili NMR strukturo lektinske domene Siglec-8 v kompleksu s svojim ligandom tetrasaharidnim 6'-sulfo sialil Lewisx (Neu5Acα2-3[6S]Galβ1-4[Fucα1-3]GlcNAc). Glavne interakcije vključujejo solni most med Arg109 in karboksilatom sialične kisline ter drugi solni most med Arg56 in Gln59 ter sulfatom na mestu 6 galaktoznega dela. Zanimivo je, da ima sulfat ključno vlogo pri vezavi in specifičnosti na Siglec-8. Poleg tega obstajajo vodikove vezi med hidroksilnimi skupinami 7, 8 in 9 sialične kisline ter Tyr7, Ser118 in Gln122. Nasprotno pa podenote fukoze in glukozamina tvorijo le šibkejše interakcije. To je pripeljalo do predloga, da bi disaharidna podstruktura 6-sulfo-Sia-Gal lahko predstavljala minimalni vezavni epitop za Siglec-8.

• Keywords: ligandi, Siglec-8, farmacevtska kemija, apoptoza