Coordination Chemistry of Sulfur-Containing Bifunctional Chelators: Toward in Vivo Stabilization of 64Cu PET Imaging Agents for Alzheimer's Disease.

The broader utilization of 64Cu positron emission tomography (PET) imaging agents has been hindered by the unproductive demetalation induced by bioreductants. To advance the development of 64Cu-based PET imaging tracers for Alzheimer's Disease (AD), there is a need for novel ligand design strategies. In this study, we developed sulfur-containing dithiapyridinophane (N2S2) bifunctional chelators (BFCs) as well as all nitrogen-based diazapyridinophane (N4) BFCs to compare their abilities to chelate Cu and target Aß aggregates. Through spectrophotometric titrations and electrochemical measurements, we have demonstrated that the N2S2-based BFCs exhibit >10 orders of magnitude higher binding affinity toward Cu(I) compared to their N4-based counterparts, while both types of BFCs exhibit high stability constants toward Cu(II). Notably, solid state structures for both Cu(II) and Cu(I) complexes supported by the two ligand frameworks were obtained, providing molecular insights into their copper chelating abilities. Aß binding experiments were conducted to study the structure-affinity relationship, and fluorescence microscopy imaging studies confirmed the selective labeling of the BFCs and their copper complexes. Furthermore, we investigated the potential of these ligands for the 64Cu-based PET imaging of AD through radiolabeling and autoradiography studies. We believe our findings provide molecular insights into the design of bifunctional Cu chelators that can effectively stabilize both Cu(II) and Cu(I) and, thus, can have significant implications for the development of 64Cu PET imaging as a diagnostic tool for AD.

C-H bond activation via concerted metalation-deprotonation at a palladium(iii) center.

Herein we report the direct observation of C-H bond activation at an isolated mononuclear Pd(iii) center. The oxidation of the Pd(ii) complex (MeN4)PdII(neophyl)Cl (neophyl = -CH2C(CH3)2Ph; MeN4 = N,N'-dimethyl-2,11-diaza[3.3](2,6)pyridinophane) using the mild oxidant ferrocenium hexafluorophosphate (FcPF6) yields the stable Pd(iii) complex [(MeN4)PdIII(neophyl)Cl]PF6. Upon the addition of an acetate source, [(MeN4)PdIII(neophyl)Cl]PF6 undergoes Csp2-H bond activation to yield the cyclometalated product [(MeN4)PdIII(cycloneophyl)]PF6. This metalacycle can be independently prepared, allowing for a complete characterization of both the starting and final Pd(iii) complexes. The C-H activation step can be monitored directly by EPR and UV-Vis spectroscopies, and kinetic isotope effect (KIE) studies suggest that either a pre-association step such as an agostic interaction may be rate limiting, or that the C-H activation is partially rate-limiting in conjunction with ligand rearrangement. Density functional theory calculations support that the reaction proceeds through a κ3 ligand coordination and that the flexible ligand structure is important for this transformation. Overall, this study represents the first example of discrete C-H bond activation occurring at a Pd(iii) center through a concerted metalation-deprotonation mechanism, akin to that observed for Pd(ii) and Pd(iv) centers.

Isolation and Characterization of Heteroleptic Mononuclear Palladium(I) Complexes.

Catalytic transformations involving Pd(0)/Pd(II) catalytic cycles are very well known, and processes involving high-valent Pd(III) and Pd(IV) and low-valent Pd(I) intermediates have also gained interest in recent years. Although low-valent Pd(I) intermediates are proposed in these catalytic cycles, isolated and characterized mononuclear Pd(I) species are very rare. Herein, we report the isolation of two heteroleptic mononuclear Pd(I) complexes stabilized by dithiapyridinophane ligands that were fully characterized by single-crystal X-ray diffraction; EPR, IR, UV-vis spectroscopies; and computational studies. Excitingly, one of these Pd(I) complexes shows Kumada Csp3-Csp2 cross-coupling competency, and initial studies of the other shows direct evidence for Csp3-H bond activation proposed to occur at the Pd(I) center.

Visible Light Mediated Bidirectional Control over Carbonic Anhydrase Activity in Cells and in Vivo Using Azobenzenesulfonamides.

Two azobenzenesulfonamide molecules with thermally stable cis configurations resulting from fluorination of positions ortho to the azo group are reported that can differentially regulate the activity of carbonic anhydrase in the trans and cis configurations. These fluorinated probes each use two distinct visible wavelengths (520 and 410 or 460 nm) for isomerization with high photoconversion efficiency. Correspondingly, the cis isomer of these systems is highly stable and persistent (as evidenced by structural studies in solid and solution state), permitting regulation of metalloenzyme activity without continuous irradiation. Herein, we use these probes to demonstrate the visible light mediated bidirectional control over the activity of zinc-dependent carbonic anhydrase in solution as an isolated protein, in intact live cells and in vivo in zebrafish during embryo development.

Responsive fluorinated nanoemulsions for 19F magnetic resonance detection of cellular hypoxia.

We report two highly fluorinated Cu-based imaging agents, CuL1 and CuL2, for detecting cellular hypoxia as nanoemulsion formulations. Both complexes retained their initial quenched 19F MR signals due to paramagnetic Cu2+; however, both complexes displayed a large signal increase when the complex was reduced. DLS studies showed that the CuL1 nanoemulsion (NECuL1) had a hydrodiameter of approximately 100 nm and that it was stable for four weeks post-preparation. Hypoxic cells incubated with NECuL1 showed that 40% of the Cu2+ taken up was reduced in low oxygen environments.

Highly fluorinated metal complexes as dual 19F and PARACEST imaging agents.

We reported a set of water-soluble transition metal complexes that can serve as both 19F and PARACEST magnetic resonance imaging agents. The high number of equivalent fluorine atoms and the paramagnetic effect of metals offer these complexes high 19F sensitivity as demonstrated by in vitro19F MRI experiments. The complexes contain carboxamide groups appended onto a cyclen macrocycle, which provide 1H CEST peaks well differentiated from bulk water. The Co(ii) agent displays two CEST peaks that can be utilized for ratiometric pH determination and the concept of combining 19F MR and PARACEST as complementary imaging techniques was demonstrated with the Fe(ii) complex.

Copper(ii) complexes for cysteine detection using 19F magnetic resonance.

Cysteine plays an essential role in maintaining cellular redox homeostasis and perturbations in cysteine concentration are associated with cardiovascular disease, liver disease, and cancer. 19F MRI is a promising modality for detecting cysteine in biology due to its high tissue penetration and negligible biological background signal. Herein we report fluorinated macrocyclic copper complexes that display a 19F NMR/MRI turn-on response following reduction of the Cu(ii) complexes by cysteine. The reactivity with cysteine was studied by monitoring the appearance of a robust diamagnetic 19F signal following addition of cysteine in conjunction with UV-vis and EPR spectroscopies. Importantly, complexes with -CH2CF3 tags display good water solubility. Studies with this complex in HeLa cells demonstrate the applicability of these probes to detect cysteine in complex biological environments.

19F PARASHIFT Probes for Magnetic Resonance Detection of H2O2 and Peroxidase Activity.

Elevated levels of reactive oxygen species and peroxidase expression are often associated with inflammation and inflammatory diseases. We developed two novel Co(II) complexes that can be used to detect oxidative activity associated with inflammation using 19F magnetic resonance imaging (MRI). These agents display a large change in 19F chemical shift upon oxidation from Co(II) to Co(III), facilitating selective visualization of both species using chemical shift selective pulse sequences. This large chemical shift change is attributed to a large magnetic anisotropy in the high spin Co(II) complexes. Importantly, the differing reactivity of the two agents allows for detection of either H2O2 production and/or the activity of peroxidase enzymes, providing two useful platforms for 19F MR hot spot imaging of oxidative events associated with biological inflammation.