64Cu tumor labeling with hexadentate picolinic acid-based bispidine immunoconjugates.

Discussed are two picolinate appended bispidine ligands (3,7-diazabicyclo[3.3.1]nonane derivatives) in comparison with an earlier described bis-pyridine derivative, which are all known to strongly bind CuII. The radiopharmacological characterization of the two isomeric bispidine complexes includes quantitative labeling with 64CuII at ambient conditions with high radiochemical purities and yields (molar activities >200 MBq/nmol). Challenge experiments in presence of EDTA, cyclam, human serum and SOD demonstrate high stability and inertness of the 64Cu-bispidine complexes. Biodistribution studies performed in Wistar rats indicate a rapid renal elimination for both 64Cu-labeled chelates. The bispidine ligand with the picolinate group in N7 position was selected for further biological experiments, and its backbone was therefore substituted with a benzyl-NCS group at C9. Two tumor target modules (TMs), targeting prostate stem cell antigen (PSCA), overexpressed in prostate cancer, and the fibroblast activation protein (FAP) in fibrosarcoma, were selected for thiourea coupling with the NCS-functionalized ligand and lysine residues of TMs. Small animal PET experiments on tumor-bearing mice showed specific accumulation of the 64Cu-labeled TMs in PSCA- and FAP-overexpressing tumors (standardized uptake value (SUV) for PC3: 2.7±0.6 and HT1080: 7.2±1.25) with almost no uptake in wild type tumors.

First-Generation Bispidine Chelators for 213 BiIII Radiopharmaceutical Applications.

Hepta- and octadentate bispidines (3,7-diazabicyclo[3.3.1]nonane, diaza-adamantane) with acetate, methyl-pyridine, and methyl-picolinate pendant groups at the amine donors of the bispidine platform have been prepared and used to investigate BiIII coordination chemistry. Crystal structure and solution spectroscopic data (NMR spectroscopy and mass spectrometry) confirm that the rigid and relatively large bispidine cavity with an axially distorted geometry is well suited for BiIII and in all cases forms nine-coordinate complexes; this is supported by an established hole size and shape analysis. It follows that nonadentate bispidines probably will be more suited as bifunctional chelators for 213 BiIII -based radiopharmaceuticals. However, two isomeric picolinate-/acetate-based heptadentate ligands already show very efficient complexation kinetics with 213 BiIII at ambient temperature and kinetic stability that is comparable with the standard ligands used in this field. The experimentally determined hydrophilicities (log D7.4 values) show that the BiIII complexes reported are relatively hydrophilic and well suited for medicinal applications. We also present a very efficient and relatively accurate method to compute charge distributions and hydrophilicities, and this will help to further optimize the systems reported here.

Bispidines for radiopharmaceuticals.

Very rigid tetra-, penta-, hexa-, hepta- and octadentate bispidine ligands are well preorganized for specific coordination geometries. The broad range of coordination numbers, geometries and donor sets allows one to design and prepare bispidines for many specific metal ions and applications. Major requirements for radiopharmaceuticals are efficient labeling, inertness under physiological conditions and easy functionalization of the ligands with biological vectors. The reasons why bispidines are among the most favorable chelators for radiopharmaceutical applications, a comprehensive list of bispidine ligands and a range of examples where bispidines have been used to develop radiopharmaceuticals for future applications in diagnosis and therapy are discussed.

Optimization of Hexadentate Bispidine Ligands as Chelators for 64 CuII PET Imaging.

The coordination chemistry of the hexadentate bispidine ligand L1 with three pyridine, one pyridazine, and two tertiary amine donors with CuII and ZnII is reported. The substitution of one of the two cis-disposed pyridine donors in-plane with the two tertiary amines of the bispidine backbone by a pyridazine group was predicted to substantially reduce distortion from planarity of the tetradentate subunit and therefore was assumed to lead to higher CuII complex stability. The X-ray single-crystal structures of the ZnII and CuII complexes, which are in excellent agreement with the DFT- and MM-optimized structures, confirmed this prediction but indicated that deviation from planarity is appreciable. This also emerges from solution spectroscopy (UV/Vis/NIR and EPR of the CuII complex), which indicated that the in-plane ligand field is only slightly increased. The geometric parameters together with the lower basicity of pyridazine versus pyridine (pKa =2.33 vs. 5.23) are also in agreement with an only slightly more negative redox potential of the CuII/I couple (Eo =-780 vs. -760 mV, MeCN, vs. Fc/Fc+ ), and the potentiometrically determined CuII stability constant with L1 is slightly lower than that with the parent ligand L2 (log K=12.7 vs. 14.5). Therefore, modification of the donor groups is a more promising approach to increasing the stabilities of these complexes and yields 64 CuII chelators that outperform known hexadentate bispidine ligands.

Octadentate Picolinic Acid-Based Bispidine Ligand for Radiometal Ions.

The synthesis of the octadentate bispidine ligand bearing two picolinic acid pendant arms (H2 bispa2 ), and its coordination chemistry with radionuclides relevant for nuclear medicine, namely indium(III) (111 In), lutetium(III) (177 Lu), and lanthanum(III) (as surrogate for 225 Ac), are reported. The non-radioactive metal complexes of the N6 O2 -type bispa ligand were characterized by 1 H and 13 C NMR spectroscopy, elemental analysis, mass spectrometry and single-crystal X-ray analysis. Experimental structural data, computational analysis, complex stabilities determined by potentiometric titration, and "radiostabilities" determined by competition studies in the presence of human serum reveal complex stabilities of H2 bispa2 comparable to those of the macrocyclic "gold standard" DOTA. After an incubation time of 1 day, 86 and 87 % of [177 Lu(bispa2 )]+ and [177 Lu(DOTA)]- , respectively, remain intact. Importantly, unlike DOTA, H2 bispa2 is radiolabeled quantitatively with 111 InIII and 225 AcIII under ambient conditions, which is an essential aspect when working with heat-sensitive antibodies as targeting vectors. In the case of 111 InIII , room temperature radiolabeling of H2 bispa2 yields molar activities as high as 70 MBq nmol-1 within 10 minutes. These are promising results for radiopharmaceutical applications of H2 bispa2 .