Targeted, Molecular Europium(III) Probes Enable Luminescence-Guided Surgery and 1 Photon Post-Surgical Luminescence Microscopy of Solid Tumors.

Discrete luminescent lanthanide complexes represent a potential alternative to organic chromophores due to their tunability of optical properties, insensitivity to photobleaching, and large pseudo-Stokes shifts. Previously, we demonstrated that the lack of depth penetration of UV excitation required to sensitize discrete terbium and europium complexes can be overcome using Cherenkov radiation emitted by clinically employed radioisotopes in situ. Here, we show that the second-generation europium complexes [Eu(III)(pcta-PEPA2)] and [Eu(III)(tacn-pic-PEPA2)] (Φ = 57% and 76%, respectively) lower the limit of detection (LoD) to 1 nmol in the presence of 10 µCi of Cherenkov emitting isotopes, 18F and 68Ga. Bifunctionalization provides access to cysteine-linked peptide conjugates with comparable brightness and LoD. The conjugate, [Eu(tacn-(pic-PSMA)-PEPA2)], displays high binding affinity to prostate-specific membrane antigen (PSMA)-expressing PC-3 prostate cancer cells in vitro and can be visualized in the membrane-bound state using confocal microscopy. Biodistribution studies with the [86Y][Y(III)(tacn-(pic-PSMA)-PEPA2)] analogue in a mouse xenograft model were employed to study pharmacokinetics. Systemic administration of the targeted Cherenkov emitter, [68Ga][Ga(III)(PSMA-617)], followed by intratumoral injection or topical application of 20 or 10 nmol [Eu(III)(tacn-(pic-PSMA)-PEPA2)], respectively, in live mice resulted in statistically significant signal enhancement using conventional small animal imaging (620 nm bandpass filter). Optical imaging informed successful tumor resection. Ex vivo imaging of the fixed tumor tissue with 1 and 2 photon excitation further reveals the accumulation of the administered Eu(III) complex in target tissues. This work represents a significant step toward the application of luminescent lanthanide complexes for optical imaging in a clinical setting.

Correction: Radiolabeling and in vivo evaluation of lanmodulin with biomedically relevant lanthanide isotopes.

[This corrects the article DOI: 10.1039/D3CB00020F.].

Radiolabeling and in vivo evaluation of lanmodulin with biomedically relevant lanthanide isotopes.

Short-lived, radioactive lanthanides comprise an emerging class of radioisotopes attractive for biomedical imaging and therapy applications. To deliver such isotopes to target tissues, they must be appended to entities that target antigens overexpressed on the target cell's surface. However, the thermally sensitive nature of biomolecule-derived targeting vectors requires the incorporation of these isotopes without the use of denaturing temperatures or extreme pH conditions; chelating systems that can capture large radioisotopes under mild conditions are therefore highly desirable. Herein, we demonstrate the successful radiolabeling of the lanthanide-binding protein, lanmodulin (LanM), with medicinally relevant radioisotopes: 177Lu, 132/135La and 89Zr. Radiolabeling of the endogenous metal-binding sites of LanM, as well exogenous labeling of a protein-appended chelator, was successfully conducted at 25 °C and pH 7 with radiochemical yields ranging from 20-82%. The corresponding radiolabeled constructs possess good formulation stability in pH 7 MOPS buffer over 24 hours (>98%) in the presence of 2 equivalents of natLa carrier. In vivo experiments with [177Lu]-LanM, [132/135La]-LanM, and a prostate cancer targeting-vector linked conjugate, [132/135La]-LanM-PSMA, reveal that endogenously labeled constructs produce bone uptake in vivo. Exogenous, chelator-tag mediated radiolabeling to produce [89Zr]-DFO-LanM enables further study of the protein's in vivo behavior, demonstrating low bone and liver uptake, and renal clearance of the protein itself. While these results indicate that additional stabilization of LanM is required, this study establishes precedence for the radiochemical labeling of LanM with medically relevant lanthanide radioisotopes.

N,N-Alkylation Clarifies the Role of N- and O-Protonated Intermediates in Cyclen-Based 64Cu Radiopharmaceuticals.

Radioisotopes of Cu, such as 64Cu and 67Cu, are alluring targets for imaging (e.g., positron emission tomography, PET) and radiotherapeutic applications. Cyclen-based macrocyclic polyaminocarboxylates are one of the most frequently examined bifunctional chelators in vitro and in vivo, including the FDA-approved 64Cu radiopharmaceutical, Cu(DOTATATE) (Detectnet); however, connections between the structure of plausible reactive intermediates and their stability under physiologically relevant conditions remain to be established. In this study, we share the synthesis of a cyclen-based, N,N-alkylated spirocyclic chelate, H2DO3AC4H8, which serves as a model for N-protonation. Our combined experimental (in vitro and in vivo) and computational studies unravel complex pH-dependent speciation and enable side-by-side comparison of N- and O-protonated species of relevant 64Cu radiopharmaceuticals. Our studies suggest that N-protonated species are not inherently unstable species under physiological conditions and demonstrate the potential of N,N-alkylation as a tool for the rational design of future radiopharmaceuticals.

Construction of the Bioconjugate Py-Macrodipa-PSMA and Its In Vivo Investigations with Large 132/135La3+ and Small 47Sc3+ Radiometal Ions.

To harness radiometals in clinical settings, a chelator forming a stable complex with the metal of interest and targets the desired pathological site is needed. Toward this goal, we previously reported a unique set of chelators that can stably bind to both large and small metal ions, via a conformational switch. Within this chelator class, py-macrodipa is particularly promising based on its ability to stably bind several medicinally valuable radiometals including large 132/135La3+, 213Bi3+, and small 44Sc3+. Here, we report a 10-step organic synthesis of its bifunctional analogue py-macrodipa-NCS, which contains an amine-reactive -NCS group that is amenable for bioconjugation reactions to targeting vectors. The hydrolytic stability of py-macordipa-NCS was assessed, revealing a half-life of 6.0 d in pH 9.0 aqueous buffer. This bifunctional chelator was then conjugated to a prostate-specific membrane antigen (PSMA)-binding moiety, yielding the bioconjugate py-macrodipa-PSMA, which was subsequently radiolabeled with large 132/135La3+ and small 47Sc3+, revealing efficient and quantitative complex formation. The resulting radiocomplexes were injected into mice bearing both PSMA-expressing and PSMA-non-expressing tumor xenografts to determine their biodistribution patterns, revealing delivery of both 132/135La3+ and 47Sc3+ to PSMA+ tumor sites. However, partial radiometal dissociation was observed, suggesting that py-macrodipa-PSMA needs further structural optimization.

The expanding role of primary care providers in care of individuals with kidney disease.

An estimated 37 million Americans have chronic kidney disease (CKD). Primary care providers (PCPs) have long played a critical role in detecting CKD and preventing disease progression, particularly in the early stages of the disease. With recent studies demonstrating substantial improvements in kidney outcomes with use of sodium glucose cotransporter 2 (SGLT2) inhibitors, PCPs have an even greater opportunity to improve care of individuals with CKD. Health disparities in nephrology have recently come to the forefront - again, PCPs will play a key role in efforts to reduce such disparities and ensure all patients receive high quality care. This review summarizes the latest guidelines for treatment of CKD and its complications, explores health disparities affecting patients with CKD, and highlights the role of the PCP in caring for this population.

Multiplex and In Vivo Optical Imaging of Discrete Luminescent Lanthanide Complexes Enabled by In Situ Cherenkov Radiation Mediated Energy Transfer.

Recently, we pioneered the application of Cherenkov radiation (CR) of radionuclides for the in situ excitation of discrete Eu(III) and Tb(III) complexes. CR is produced by isotopes decaying under emission of charged particles in dielectric media and exhibits a maximum intensity below 400 nm. We have demonstrated that luminescent lanthanide antenna complexes are ideal acceptors for Cherenkov radiation-mediated energy transfer (CRET). Here, we develop and assess peptide-functionalized Tb(III) and Eu(III) complexes in conjunction with CRET excitation by the positron emissive radioisotope 18F for simultaneous, multiplexed imaging and in vivo optical imaging. This work demonstrates, for the first time, that the detection of the luminescence emission of a discrete Eu(III) complex in vivo is feasible. Our results open possibilities for discrete luminescent lanthanide complexes to be used as diagnostic, optical tools for the intrasurgical guidance of tumor resection.

Establishing Radiolanthanum Chemistry for Targeted Nuclear Medicine Applications.

We report the first targeted nuclear medicine application of the lanthanum radionuclides 132/135 La. These isotopes represent a matched pair for diagnosis via the positron emissions of 132 La and therapy mediated by the Auger electron emissions of 135 La. We identify two effective chelators, known as DO3Apic and macropa, for these radionuclides. The 18-membered macrocycle, macropa, bound 132/135 La with better molar activity than DO3Apic under similar conditions. These chelators were conjugated to the prostate-specific membrane antigen (PSMA)-targeting agent DUPA to assess the use of radiolanthanum for in vivo imaging. The 132/135 La-labeled targeted constructs showed high uptake in tumor xenografts expressing PSMA. This study validates the use of these radioactive lanthanum isotopes for imaging applications and motivates future work to assess the therapeutic effects of the Auger electron emissions of 135 La.