Exploring the Limits of Ligand Rigidification in Transition Metal Complexes with Mono-N-Functionalized Pyclen Derivatives.

We report the synthesis of a new family of side-bridged pyclen ligands. The incorporation of an ethylene bridge between two adjacent nitrogen atoms was reached from the pyclen-oxalate precursor described previously. Three new side-bridged pyclen macrocycles, Hsb-3-pc1a, sb-3-pc1py, and Hsb-3-pc1pa, were obtained with the aim to assess their coordination properties toward Cu2+ and Zn2+ ions. We also prepared their nonreinforced analogues H3-pc1a, 3-pc1py, and H3-pc1pa as comparative benchmarks. The two series of ligands were characterized and their coordination properties were investigated in detail. The Zn2+ and Cu2+ complexes with the nonside-bridged series H3-pc1a, 3-pc1py, and H3-pc1pa were successfully isolated and their structures were assessed by X-ray diffraction studies. In the case of the side-bridged family, the synthesis of the complexes was far more difficult and, in some cases, unsuccessful. The results of our studies demonstrate that this difficulty is related to the extreme stiffening and basicity of such side-bridged pyclens.

Synthesis and Photophysical Properties of Lanthanide Pyridinylphosphonic Tacn and Pyclen Derivatives: From Mononuclear Complexes to Supramolecular Heteronuclear Assemblies.

Synthetic methodologies were developed to achieve the preparation of ligands L1 and L2 consisting of tacn- and pyclen-based chelators decorated with pyridinylphosphonic pendant arms combined with ethylpicolinamide or acetate coordinating functions, respectively. Phosphonate functions have been selected for their high affinity toward Ln3+ ions compared to their carboxylated counterparts and for their steric hindrance that favors the formation of less-hydrated complexes. Thanks to regiospecific N-functionalization of the macrocyclic backbones, the two ligands were isolated with good yields and implicated in a comprehensive photophysical study for the complexation of Eu3+, Tb3+, and Yb3+. The coordination behavior of L1 and L2 with these cations has been first investigated by means of a combination of UV-vis absorption spectroscopy, steady-state and time-resolved luminescence spectroscopy, and 1H and 31P NMR titration experiments. Structural characterization in solution was assessed by NMR spectroscopy, corroborated by theoretical calculations. Spectroscopic characterization of the Ln3+ complexes of L1 and L2 was done in water and D2O and showed the effective sensitization of the lanthanide metal-centered emission spectra, each exhibiting typical lanthanide emission bands. The results obtained for the phosphonated ligands were compared with those reported previously for the corresponding carboxylated analogues.

Multistep synthesis of a novel copper complex with potential for Alzheimer's disease diagnosis.

Positron emission tomography (PET) imaging of Aß plaques, is recognized as a tool for the diagnosis of Alzheimer's disease. As a contribution to the development of new strategies for early diagnosis of the disease, using PET medical imaging technique, a new copper complex, the [Cu(TE1PA-ONO)]+ was synthesized in ten steps. The key step of our strategy is the coupling of a monopicolinate-N-alkylated cyclam-based ligand with a moiety capable of recognizing Aß plaques via a successful and challenging Buchwald-Hartwig coupling reaction. To our knowledge, it is the first time that such a strategy is used to functionalize polyazamacrocyclic derivatives. The thermodynamic stability constants determined in MeOH/H2O solvent indicate that the attachment of this moiety does not weaken the chelating properties of TE1PA-ONO ligand in relation to parent HTE1PA. The novel complex described here is able to recognize amyloid plaques in brain sections from Alzheimer's disease patients and shows low toxicity to human neuronal cells.

["New Modalities in Cancer Imaging and Therapy" XVth edition of the workshop organized by the network "Tumor Targeting, Imaging, Radiotherapies" of the Cancéropôle Grand-Ouest, 5-8 October 2022, France]. / « New Modalities in Cancer Imaging and Therapy ¼ XVe édition de l'atelier organisé par le réseau « Vectorisation, Imagerie, Radiothérapies ¼ du Cancéropôle Grand-Ouest, 5­8 octobre 2022, Erquy, France.

The fifteenth edition of the international workshop organized by the "Tumour Targeting and Radiotherapies network" of the Cancéropôle Grand-Ouest focused on the latest advances in internal and external radiotherapy from different disciplinary angles: chemistry, biology, physics, and medicine. The workshop covered several deliberately diverse topics: the role of artificial intelligence, new tools for imaging and external radiotherapy, theranostic aspects, molecules and contrast agents, vectors for innovative combined therapies, and the use of alpha particles in therapy.

Design of Bifunctional Pyclen-Based Lanthanide Luminescent Bioprobes for Targeted Two-Photon Imaging.

In the past, Lanthanide Luminescent Bioprobes (LLBs) based on pyclen-bearing π-extended picolinate antennas were synthesized and demonstrated well-adapted optical properties for biphotonic microscopy. The objective of this work is to develop a strategy to design bifunctional analogues of the previously studied LLBs presenting an additional reactive chemical group to allow their coupling to biological vectors to reach deep in vivo targeted two-photon bioimaging. Herein, we elaborated a synthetic scheme allowing the introduction of a primary amine on the para position of the macrocyclic pyridine unit. The photophysical and bioimaging studies demonstrate that the introduction of the reactive function does not alter the luminescent properties of the LLBs paving the way for further applications.

Tuning the lipophilic nature of pyclen-based 90Y3+ radiopharmaceuticals for ß-radiotherapy.

Pyclen-dipicolinate chelates proved to be very efficient chelators for the radiolabeling with ß--emitters such as 90Y. In this study, a pyclen-dipicolinate ligand functionalized with additional C12 alkyl chains was synthesized. The radiolabeling with 90Y proved that the addition of saturated carbon chains does not affect the efficiency of the radiolabeling, whereas a notable increase in lipophilicity of the resulting 90Y radiocomplex was observed. As a result, the compound could be extracted in Lipiodol® and encapsulated in biodegrable pegylated poly(malic acid) nanoparticles demonstrating the potential of lipophilic pyclen-dipicolinate derivatives as platforms for the design of radiopharmaceuticals for the treatment of liver or brain cancers by internal radiotherapy.

Pyclen-Based Ligands Bearing Pendant Picolinate Arms for Gadolinium Complexation.

We report the synthesis of two pyclen-based regioisomer ligands (pyclen = 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene) functionalized with picolinic acid pendant arms either at positions 3,9-pc2pa (L5) or 3,6-pc2pa (L6) of the macrocyclic fragment. The ligands were prepared by the regiospecific protection of one of the amine nitrogen atoms of the macrocycle using Boc and Alloc protecting groups, respectively. The X-ray structure of the Gd(III) complex of L5 contains trinuclear [(GdL5)3(H2O)3]3+ entities in which the monomeric units are joined by µ2-η1:η1-carboxylate groups. However, the 1H and 89Y NMR spectra of its Y(III) analogue support the formation of monomeric complexes in solution. The Tb(III) complexes are highly luminescent, with emission quantum yields of up to 28% for [TbL5]+. The luminescence lifetimes recorded in H2O and D2O solutions indicate the presence of a water molecule coordinated to the metal ion, as also evidenced by the 1H relaxivities measured for the Gd(III) analogues. The Gd(III) complexes present very different exchange rates of the coordinated water molecule (kex298 = 87.1 × 106 and 1.06 × 106 s-1 for [GdL5]+ and [GdL6]+, respectively). The very high water exchange rate of [GdL5]+ is associated with the steric hindrance originating from the coordination of the ligand around the water binding site, which favors a dissociatively activated water exchange process. The Gd(III) complexes present rather high thermodynamic stabilities (log KGdL = 20.47 and 19.77 for [GdL5]+ and [GdL6]+, respectively). Furthermore, these complexes are remarkably inert with respect to their acid-assisted dissociation, in particular the complex of L5.

Expanding the Scope of Pyclen-Picolinate Lanthanide Chelates to Potential Theranostic Applications.

A family of three picolinate pyclen-based ligands, previously investigated for the complexation of Y3+ and some lanthanide ions (Gd3+, Eu3+), was studied with 161Tb and 177Lu in view of potential radiotherapeutic applications. The set of six Tb3+ and Lu3+ complexes was synthesized and fully characterized. The coordination properties in the solid state and in solution were thoroughly studied. Potentiometric titrations, supported by density functional theory (DFT) calculations, showed the very high stability constants of the Tb3+ and Lu3+ complexes, associated with remarkable kinetic inertness for up to 30 days in 1 M HCl. A complete radiolabeling study performed with both 161Tb and 177Lu radionuclides, including experiments with regard to the stability with and without scavengers and kinetic inertness using challenging agents, proved that the ligands could reasonably compete with the DOTA analogue. To conclude, the potential of using the same ligand for both radiotherapy and optical imaging was highlighted for the first time.

Enabling Indium Channels for Mass Cytometry by Using Reinforced Cyclam-Based Chelating Polylysine.

The synthesis of a polylysine polymer functionalized with the previously reported astonishingly inert [In(cb-te2pa)]+ chelate was performed. A biotin end group allowed the conjugation to biotinylated beads by the intermediary of a fluorescein isothiocyanate/neutravidin receptor. High quality imaging mass cytometry trials, based on 115In detection were performed to highlight the behavior of the material. Anti-CD20 antibody was labeled by the so-obtained In(III)-modified polylysine using the biotin/neutravidin interaction. Ramos (CD20[+]) and HL-60 (CD20[-]) cell lines were costained with the In(III)-modified bioconjugate by finding the best staining conditions. Both immunofluorescence microscopy (IF-M) and mass cytometry analyses confirmed the specific binding of anti-CD20 onto Ramos cells. CyTOF histograms constructed on the 115In detection allowed us to define and to separate, with a good signal-to-noise ratio, two populations (Ramos and HL-60). The inertness of In(III)-MCP-NAv over a three-month storage period was proved by performing new functionality tests involving Jurkat cells (CD20[-]) and multiparametric trials involving the 115In channel. The results ensure a promising future use of the previously announced [In(cb-te2pa)]+ complex-based polymers for mass cytometry.

Pyclen-Based Ln(III) Complexes as Highly Luminescent Bioprobes for In Vitro and In Vivo One- and Two-Photon Bioimaging Applications.

In addition to the already described ligand L4a, two pyclen-based lanthanide chelators, L4b and L4c, bearing two specific picolinate two-photon antennas (tailor-made for each targeted metal) and one acetate arm arranged in a dissymmetrical manner, have been synthesized, to form a complete family of lanthanide luminescent bioprobes: [EuL4a], [SmL4a], [YbL4b], [TbL4c], and [DyL4c]. Additionally, the symmetrically arranged regioisomer L4a' was also synthesized as well as its [EuL4a'] complex to highlight the astonishing positive impact of the dissymmetrical N-distribution of the functional chelating arms. The investigation clearly shows the high performance of each bioprobe, which, depending on the complexed lanthanide, could be used in various applications. Each presents high brightness, quantum yields, and lifetimes. Staining of the complexes into living human breast cancer cells was observed. In addition, in vivo two-photon microscopy was performed for the first time on a living zebrafish model with [EuL4a]. No apparent toxicity was detected on the growth of the zebrafish, and images of high quality were obtained.

Cationic Biphotonic Lanthanide Luminescent Bioprobes Based on Functionalized Cross-Bridged Cyclam Macrocycles.

Cationic lanthanide complexes are generally able to spontaneously internalize into living cells. Following our previous works based on a diMe-cyclen framework, a second generation of cationic water-soluble lanthanide complexes based on a constrained cross-bridged cyclam macrocycle functionalized with donor-π-conjugated picolinate antennas was prepared with europium(III) and ytterbium(III). Their spectroscopic properties were thoroughly investigated in various solvents and rationalized with the help of DFT calculations. A significant improvement was observed in the case of the Eu3+ complex, while the Yb3+ analogue conserved photophysical properties in aqueous solvent. Two-photon (2P) microscopy imaging experiments on living T24 human cancer cells confirmed the spontaneous internalization of the probes and images with good signal-to-noise ratio were obtained in the classic NIR-to-visible configuration with the Eu3+ luminescent bioprobe and in the NIR-to-NIR with the Yb3+ one.

Picolinate-appended tacn complexes for bimodal imaging: Radiolabeling, relaxivity, photophysical and electrochemical studies.

Based on our previous works involving two 1,4,7-triazacyclononane (tacn)-based ligands Hno2py1pa (1-Picolinic acid-4,7-bis(pyridin-2-ylmethyl)-1,4,7-triazacyclononane) and Hno1pa (1-Picolinic acid-1,4,7-triazacyclononane), we report here the synthesis of analogues bearing picolinate-based π-conjugated ILCT (Intra-Ligand Charge Transfer) transition antenna (HL1, HL2), using regiospecific N-functionalization of the tacn skeleton and their related transition metal complexes (e.g. Cu2+, Zn2+ and Mn2+). Coordination properties as well as their photophysical and electrochemical properties were investigated in order to quantify the impact of such antenna on the luminescent or relaxometric properties of the complexes. The spectroscopic properties of the targeted ligands and metal complexes have been studied using UV-Vis absorption and fluorescence spectrocopies. While the zinc complex formed with HL1 possesses a moderate quantum yield of 5%, complexation of Cu2+ led to an extinction of the luminescence putatively attributed to a photo-induced electron transfer, as supported by spectroscopic and electrochemical evidences. The [Mn(L2)]+ complex is characterized by a fluorescence quantum yield close to 8% in CH2Cl2. The potential interest of such systems as bimodal probes has been assessed from radiolabeling experiments conducted on HL1 and 64Cu2+ as well as confocal microscopy analyses and from relaxometric studies carried out on the cationic [Mn(L2)]+ complex. These results showed that HL1 can be used for radiolabeling, with a radiochemical conversion of 40% in 15 min at 100 °C. Finally, the relaxivity values obtained for [Mn(L2)]+, r1p = 4.80 mM-1·s-1 and r2p = 8.72 mM-1·s-1, make the Mn(II) complex an ideal candidate as a probe for Magnetic Resonance Imaging.

Highly Stable and Inert Complexation of Indium(III) by Reinforced Cyclam Dipicolinate and a Bifunctional Derivative for Bead Encoding in Mass Cytometry.

H2 cb-te2pa, a cross-bridged cyclam functionalized by two picolinate arms, was used for the formation of an incredible inert InIII chelate. The inertness of the complex was evaluated by UV/Vis experiments in several competitive media and was highlighted by the comparison with [In(dota)]- and [In(dtpa)]2- (H4 dota = 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid, H5 dtpa = diethylenetriamine pentaacetic acid), which are currently used in biological applications. For the first time, a bifunctional analogue of H2 cb-te2pa was prepared by C-functionalization to keep its coordination properties intact. However, this strategy leads to the formation of two diastereoisomers as evidenced and studied by NMR experiments and DFT calculations. Kinetic studies proved nevertheless that both isomers of the complex are equally inert. They were therefore used without distinction for their covalent grafting on polystyrene beads. The so-called metal-encoded beads were tested for imaging mass cytometry. The detection of 115 In allows the generation of images with high quality, proving the great potential of the bifunctional [In(cb-te2pa)]+ derivatives for single-cell analysis by mass cytometry.

Expanding the Family of Pyclen-Based Ligands Bearing Pendant Picolinate Arms for Lanthanide Complexation.

We report a detailed characterization of lanthanide complexes with two azaligands based on the pyclen macrocycle containing two picolinate and one acetate pendant arms. The two picolinate arms are attached to either opposite (L3) or adjacent (L4) amine nitrogen atoms of the macrocyclic platform. The X-ray structures of the Yb3+ complexes show nine coordination of the ligand to the metal ion, a situation that is also observed for EuL4 in the solid state. The EuL3 complex forms centrosymmetric dimers in the solid state joined by µ2-η1:η1 carboxylate groups, which results in 10-coordinate Eu3+ ions. The emission spectra of EuL3 measured in H2O and D2O solution reveal the presence of a hydration equilibrium involving a nine-coordinate species lacking inner-sphere water molecules and a monohydrated 10-coordinate species. The Eu3+ complexes present modest emission quantum yields in buffered aqueous solution (Φ = 16 and 22% for EuL3 and EuL4, 0.1 M tris buffer, pH 7.4), while the corresponding Tb3+ complexes present very high emission quantum yields under the same conditions (∼90%). 1H NMR studies show that the complexes of L3 present a fluxional behavior in D2O solution, while those of L4 are more rigid. The analysis of the Yb3+-induced NMR shifts of YbL4 indicates that the complex presents a structure in solution similar to that observed in the solid state. The Gd3+ complexes present very high thermodynamic stability constants (log KGdL = 23.56(2) and 23.44(2) for GdL3 and GdL4, respectively). The corresponding pGd values (pGd = -log[Gd3+]free with cL = 1 × 10-5 M and cGd = 1 × 10-6 at pH 7.4) of 20.69 (GdL3) and 21.83 (GdL4) are higher than that of Gd(dota)- (pGd = 19.21). The Gd3+ complexes also show remarkable inertness with respect to their proton-assisted dissociation, with dissociation half-life times of 50 min (GdL3) and 20 h (GdL4) in 1 M HCl.

Combining a pyclen framework with conjugated antenna for the design of europium and samarium luminescent bioprobes.

The first pyclen based ligand bearing two picolinate intra-ligand charge transfer transition antennae and one acetate arm organized in a dissymmetric manner was synthesized for Eu(iii) and Sm(iii) complexation. The europium complex presents an excellent brightness and biphotonic imaging of T24-cells has been performed using Eu(iii) and the less common Sm(iii) bioprobes.

Kinetics Are Crucial When Targeting Copper Ions to Fight Alzheimer's Disease: An Illustration with Azamacrocyclic Ligands.

Targeting copper ions to either remove or redistribute them is currently viewed as a possible therapeutic strategy in the context of Alzheimer's disease (AD). Thermodynamic parameters, as for instance the copper(II) affinity of the drug candidate or the copper(II) over zinc(II) selectivity, are considered in the design of the drug candidate. In contrast, kinetic factors have been overlooked despite their probable high importance. In the present article, we use a series of azamacrocyclic ligands to demonstrate that kinetic issues must be taken into account when designing copper-targeting drug candidates in the context of AD.

Stable and Inert Yttrium(III) Complexes with Pyclen-Based Ligands Bearing Pendant Picolinate Arms: Toward New Pharmaceuticals for ß-Radiotherapy.

We report the synthesis of two azaligands based on the pyclen macrocyclic platform containing two picolinate and one acetate pendant arms. The two ligands differ in the relative positions of the pendant functions, which are arranged either in a symmetrical (L3) or nonsymmetrical (L4) fashion. The complexation properties of the ligands toward natY3+ and 90Y3+ were investigated to assess their potential as chelating units for radiopharmaceutical applications. The X-ray structures of the YL3 and YL4 complexes show nonadentate binding of the ligand to the metal ions. A multinuclear 1H, 13C, and 89Y NMR study shows that the complexes present a structure in solution similar to that observed in the solid state. The two complexes present very high thermodynamic stability constants (log KYL = 23.36(2) and 23.07(2) for YL3 and YL4, respectively). The complexes also show a remarkable inertness with respect to their proton-assisted dissociation, especially YL4. 90Y radiolabeling was proved to be efficient under mild conditions. The 90YL3 and 90YL4 radiochelates were found to be more stable than 90Y(DOTA).

A Coordination Chemistry Approach to Fine-Tune the Physicochemical Parameters of Lanthanide Complexes Relevant to Medical Applications.

The geometric features of two pyclen-based ligands possessing identical donor atoms but different site organization have a profound impact in their complexation properties toward lanthanide ions. The ligand containing two acetate groups and a picolinate arm arranged in a symmetrical fashion (L1) forms a Gd3+ complex being two orders of magnitude less stable than its dissymmetric analogue GdL2. Besides, GdL1 experiences a much faster dissociation following the acid-catalyzed mechanism than GdL2. On the contrary, GdL1 exhibits a lower exchange rate of the coordinated water molecule compared to GdL2. These very different properties are related to different strengths of the Gd-ligand bonds associated to steric effects, which hinder the coordination of a water molecule in GdL2 and the binding of acetate groups in GdL1.

The role of the capping bond effect on pyclen natY3+/90Y3+ chelates: full control of the regiospecific N-functionalization makes the difference.

Thanks to a smart regiospecific N-functionalization, a pyclen based ligand bearing one picolinate and two acetate arms organized in a dissymmetric manner was synthesized for Y3+ complexation, and compared to its symmetric analogue. The nature of the capping bonds around the metal coordination environment has a dramatic effect on the properties of the chelate, the natY3+ and 90Y3+ dissymmetric derivatives presenting enhanced thermodynamic stability and kinetic inertness.