Development of Novel Chelate-Based Probes for Applications in Bioanalysis and Molecular Imaging

Chelate-based probes are complexes which require multidentate ligands or chelators in order to demonstrate their optimal properties. This thesis is about the development of three chelate-based probes: Luminescent Lanthanide Complexes (LLCs) (Chapter 2), a Gd-based MRI contrast agent (Chapter 3), and 89Zr based PET agents (Chapter 4). In order to be used effectively in bioanalysis and molecular imaging applications, these probes must show robust signals (e.g., brightness,relaxivity, and radioactivity). Furthermore, signals must be remained unchanged in a highly competitive biological environments, which are reflected in the stability of the probes. Chapter 1 describes the general and specific requirements to achieve the highest signals followed by critical factors which should be considered in rational design of stable chelate-based probes. Chapter 2 describes the synthesis, photophysical properties and kinetic stability of a series of water-soluble, highly emissive Tb(III) and Eu(III) complexes featuring triethylenetetraamine hexaacetic acid (TTHA) and cyclohexyl triethylenetetraamine hexaacetic acid (cyTTHA) chelator scaffolds and carbostyril sensitizers. The unique and modular design of the chelators gives rise to striking quantum yields of emission in aqueous solutions (up to 54%) as well as the characteristic lanthanides’ photophysical properties (long excited state lifetimes, large effective Stokes shifts, and narrow emission peaks). Furthermore, the pre-organized chelators bind metal within minutes at ambient temperature yet exhibit substantial resistance to transchelation in the presence of a challenge solution (EDTA, 1 mM). Moreover, the Eu(III) complex (L4) remains stably luminescent in HeLa cells over hours, demonstrating the suitability of these compounds for live cell imaging applications. Representative chelators suitable for derivatization and protein bioconjugation were also prepared that were functionalized with clickable azide and alkyne moieties, biotin and trimethoprim (TMP). The development of a novel Gd based contract agent, [Gd(CyMe2TA)(H2O)]-, with high relaxivity of 4.3 mM-1s-1 (serum at 60 MHz and 37 °C) is described in chapter 3. The introduction of a pre-organized cyclohexyl group in the chelator scaffold obviates the adverse effect of the ternary complex formation with important biological anions frequently observed with linear or cyclic polyaminocarboxylate chelators. Additionally, it gives rise to a high complex stability of (pM = 19.9) and 240-fold increase in HSA affinity to 12 mM-1 relative to the clinically used Magnevist, [Gd(DTPA)(H2O)]2- (0.05 mM-1). Chapter 4 discloses efficient and modular syntheses of a library of seven pre-organized polyaminopolycarboxylate chelators from commercially available starting materials. The synthetic flexibility is further assessed by facile preparation of mono- and di-reactive, tert-butyl-protected TTHA/Cy-TTHA to selectively functionalize central chelators’ carboxylic acids. The preliminary investigation of our chelators’ library with 89Zr reveals that Cy-DTPA is more kinetically inert than the current “gold standard” zirconium chelator, DFO, with the tranchelation percentage of only ~ 4% under the experimental conditions.