title: Modifications of the 7-Hydroxyl Group of the Transthyretin Ligand Luteolin Provide Mechanistic Insights into Its Binding Properties and High Plasma Specificity.
abstract: Amyloid formation of the plasma protein transthyretin (TTR) has been linked to familial amyloid polyneuropathy and senile systemic amyloidosis. Binding of ligands within its natural hormone binding site can stabilize the tetrameric structure and impair amyloid formation. We have recently shown that the flavonoid luteolin stabilizes TTR in human plasma with a very high selectivity. Luteolin, however, is inactivated in vivo via glucuronidation for which the preferred site is the hydroxy group at position 7 on its aromatic A-ring. We have evaluated the properties of two luteolin variants in which the 7-hydroxy group has been exchanged for a chlorine (7-Cl-Lut) or a methoxy group (7-MeO-Lut). Using an in vitro model, based on human liver microsomes, we verified that these modifications increase the persistence of the drug. Crystal structure determinations show that 7-Cl-Lut binds similarly to luteolin. The larger MeO substituent cannot be accommodated within the same space as the chlorine or hydroxy group and as a result 7-MeO-Lut binds in the opposite direction with the methoxy group in position 7 facing the solvent. Both 7-Cl-Lut and 7-MeO-Lut qualify as high-affinity binders, but in contrast to luteolin, they display a highly non-specific binding to other plasma components. The binding of the two conformations and the key-interactions to TTR are discussed in detail. Taken together, these results show a proof-of-concept that the persistence of luteolin towards enzymatic modification can be increased. We reveal two alternative high-affinity binding modes of luteolin to TTR and that modification in position 7 is restricted only to small substituents if the original orientation of luteolin should be preserved. In addition, the present work provides a general and convenient method to evaluate the efficacy of TTR-stabilizing drugs under conditions similar to an in vivo environment.
authors.affiliation: Department of Medical Biochemistry and Biophysics, Umeå University, SE-901 87, Umeå, Sweden.|Department of Chemistry, Umeå University, SE-901 87, Umeå, Sweden.
authors.forename: Lina
authors.initials: L
authors.lastname: Nilsson
authors.affiliation: Department of Chemistry, Umeå University, SE-901 87, Umeå, Sweden.|Swedish Defence Research Agency, CBRN Defence and Security, SE-906 21, Umeå, Sweden.
authors.forename: Andreas
authors.initials: A
authors.lastname: Larsson
authors.affiliation: Department of Chemistry, Umeå University, SE-901 87, Umeå, Sweden.
authors.forename: Afshan
authors.initials: A
authors.lastname: Begum
authors.affiliation: Department of Medical Biochemistry and Biophysics, Umeå University, SE-901 87, Umeå, Sweden.
authors.forename: Irina
authors.initials: I
authors.lastname: Iakovleva
authors.affiliation: Department of Chemistry, Umeå University, SE-901 87, Umeå, Sweden.
authors.forename: Marcus
authors.initials: M
authors.lastname: Carlsson
authors.affiliation: Department of Medical Biochemistry and Biophysics, Umeå University, SE-901 87, Umeå, Sweden.
authors.forename: Kristoffer
authors.initials: K
authors.lastname: Brännström
authors.affiliation: Department of Chemistry, Umeå University, SE-901 87, Umeå, Sweden.
authors.forename: A Elisabeth
authors.initials: AE
authors.lastname: Sauer-Eriksson
authors.affiliation: Department of Medical Biochemistry and Biophysics, Umeå University, SE-901 87, Umeå, Sweden.
authors.forename: Anders
authors.initials: A
authors.lastname: Olofsson
mesh_terms: D001798:Blood Proteins / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D008024:Ligands; D047311:Luteolin / Q000097:blood / Q000378:metabolism*; D008862:Microsomes, Liver / Q000378:metabolism; D011228:Prealbumin / Q000378:metabolism*; D011485:Protein Binding
chemical_list: D001798:Blood Proteins; D008024:Ligands; D011228:Prealbumin; D047311:Luteolin