[123I]CC1: A PARP-Targeting, Auger Electron-Emitting Radiopharmaceutical for Radionuclide Therapy of Cancer.

Poly(adenosine diphosphate ribose) polymerase (PARP) has emerged as an effective therapeutic strategy against cancer that targets the DNA damage repair enzyme. PARP-targeting compounds radiolabeled with an Auger electron-emitting radionuclide can be trapped close to damaged DNA in tumor tissue, where high ionizing potential and short range lead Auger electrons to kill cancer cells through the creation of complex DNA damage, with minimal damage to surrounding normal tissue. Here, we report on [123I]CC1, an 123I-labeled PARP inhibitor for radioligand therapy of cancer. Methods: Copper-mediated 123I iododeboronation of a boronic pinacol ester precursor afforded [123I]CC1. The level and specificity of cell uptake and the therapeutic efficacy of [123I]CC1 were determined in human breast carcinoma, pancreatic adenocarcinoma, and glioblastoma cells. Tumor uptake and tumor growth inhibition of [123I]CC1 were assessed in mice bearing human cancer xenografts (MDA-MB-231, PSN1, and U87MG). Results: In vitro and in vivo studies showed selective uptake of [123I]CC1 in all models. Significantly reduced clonogenicity, a proxy for tumor growth inhibition by ionizing radiation in vivo, was observed in vitro after treatment with as little as 10 Bq [123I]CC1. Biodistribution at 1 h after intravenous administration showed PSN1 tumor xenograft uptake of 0.9 ± 0.06 percentage injected dose per gram of tissue. Intravenous administration of a relatively low amount of [123I]CC1 (3 MBq) was able to significantly inhibit PSN1 xenograft tumor growth but was less effective in xenografts that expressed less PARP. [123I]CC1 did not cause significant toxicity to normal tissues. Conclusion: Taken together, these results show the potential of [123I]CC1 as a radioligand therapy for PARP-expressing cancers.

A radioiodinated rucaparib analogue as an Auger electron emitter for cancer therapy.

INTRODUCTION: Radioligand therapy (RLT) is an expanding field that has shown great potential in the fight against cancer. Radionuclides that can be carried by selective ligands such as antibodies, peptides, and small molecules targeting cancerous cells have demonstrated a clear improvement in the move towards precision medicine. Poly (ADP-ribose) polymerase (PARP) is a family of enzymes involved in DNA damage repair signalling pathway, with PARP inhibitors olaparib, talazoparib, niraparib, veliparib, and rucaparib having FDA approval for cancer therapy in routine clinical use. Based on our previous work with the radiolabelled PARP inhibitor [18F]rucaparib, we replaced the fluorine-18 moiety, used for PET imaging, with iodine-123, a radionuclide used for SPECT imaging and Auger electron therapy, resulting in 8-[123I]iodo-5-(4-((methylamino)methyl)phenyl)-2,3,4,6-tetrahydro-1H-azepino[5,4,3-cd]indol-1-one, ([123I]GD1), as a potential radiopharmaceutical for RLT. METHODS: [123I]GD1 was synthesized via copper-mediated radioiodination from a selected boronic esters precursor. In vitro uptake, retention, blocking, and effects on clonogenic survival with [123I]GD1 treatment were tested in a panel of cancer cell lines. Enzymatic inhibition of PARP by GD1 was also tested in a cell-free system. The biodistribution of [123I]GD1 was investigated by SPECT/CT in mice following intravenous administration. RESULTS: Cell-free enzymatic inhibition and in vitro blocking experiments confirmed a modest ability of GD1 to inhibit PARP-1, IC50 = 239 nM. In vitro uptake of [123I]GD1 in different cell lines was dose dependent, and radiolabelled compound was retained in cells for >2 h. Significantly reduced clonogenic survival was observed in vitro after exposure of cells for 1 h with as low as 50 kBq of [123I]GD1. The biodistribution of [123I]GD1 was further characterized in vivo showing both renal and hepatobiliary clearance pathways with a biphasic blood clearance. CONCLUSION: We present the development of a new theragnostic agent based on the rucaparib scaffold and its evaluation in in vitro and in vivo models. The data reported show that [123I]GD1 may have potential to be used as a theragnostic agent.

Radiofluorination of a highly potent ATM inhibitor as a potential PET imaging agent.

PURPOSE: Ataxia telangiectasia mutated (ATM) is a key mediator of the DNA damage response, and several ATM inhibitors (ATMi) are currently undergoing early phase clinical trials for the treatment of cancer. A radiolabelled ATMi to determine drug pharmacokinetics could assist patient selection in a move towards more personalised medicine. The aim of this study was to synthesise and investigate the first 18F-labelled ATM inhibitor [18F]1 for non-invasive imaging of ATM protein and ATMi pharmacokinetics. METHODS: Radiofluorination of a confirmed selective ATM inhibitor (1) was achieved through substitution of a nitro-precursor with [18F]fluoride. Uptake of [18F]1 was assessed in vitro in H1299 lung cancer cells stably transfected with shRNA to reduce expression of ATM. Blocking studies using several non-radioactive ATM inhibitors assessed binding specificity to ATM. In vivo biodistribution studies were performed in wild-type and ATM-knockout C57BL/6 mice using PET/CT and ex vivo analysis. Uptake of [18F]1 in H1299 tumour xenografts was assessed in BALB/c nu/nu mice. RESULTS: Nitro-precursor 2 was synthesised with an overall yield of 12%. Radiofluorination of 2 achieved radiochemically pure [18F]1 in 80 ± 13 min with a radiochemical yield of 20 ± 13% (decay-corrected) and molar activities up to 79.5 GBq/µmol (n = 11). In vitro, cell-associated activity of [18F]1 increased over 1 h, and retention of [18F]1 dropped to 50% over 2 h. [18F]1 uptake did not correlate with ATM expression, but could be reduced significantly with an excess of known ATM inhibitors, demonstrating specific binding of [18F]1 to ATM. In vivo, fast hepatobiliary clearance was observed with tumour uptake ranging 0.13-0.90%ID/g after 1 h. CONCLUSION: Here, we report the first radiofluorination of an ATM inhibitor and its in vitro and in vivo biological evaluations, revealing the benefits but also some limitations of 18F-labelled ATM inhibitors.

Imaging PARP with [18F]rucaparib in pancreatic cancer models.

PURPOSE: Rucaparib, an FDA-approved PARP inhibitor, is used as a single agent in maintenance therapy to provide promising treatment efficacy with an acceptable safety profile in various types of BRCA-mutated cancers. However, not all patients receive the same benefit from rucaparib-maintenance therapy. A predictive biomarker to help with patient selection for rucaparib treatment and predict clinical benefit is therefore warranted. With this aim, we developed [18F]rucaparib, an 18F-labelled isotopologue of rucaparib, and employed it as a PARP-targeting agent for cancer imaging with PET. Here, we report the in vitro and in vivo evaluation of [18F]rucaparib in human pancreatic cancer models. METHOD: We incorporated the positron-emitting 18F isotope into rucaparib, enabling its use as a PET imaging agent. [18F]rucaparib binds to the DNA damage repair enzyme, PARP, allowing direct visualisation and measurement of PARP in cancerous models before and after PARP inhibition or other genotoxic cancer therapies, providing critical information for cancer diagnosis and therapy. Proof-of-concept evaluations were determined in pancreatic cancer models. RESULTS: Uptake of [18F]rucaparib was found to be mainly dependent on PARP1 expression. Induction of DNA damage increased PARP expression, thereby increasing uptake of [18F]rucaparib. In vivo studies revealed relatively fast blood clearance of [18F]rucaparib in PSN1 tumour-bearing mice, with a tumour uptake of 5.5 ± 0.5%ID/g (1 h after i.v. administration). In vitro and in vivo studies showed significant reduction of [18F]rucaparib uptake by addition of different PARP inhibitors, indicating PARP-selective binding. CONCLUSION: Taken together, we demonstrate the potential of [18F]rucaparib as a non-invasive PARP-targeting imaging agent for pancreatic cancers.

multi-patient dose synthesis of [18F]Flumazenil via a copper-mediated 18F-fluorination.

BACKGROUND: Flumazenil (FMZ) is a functionally silent imidazobenzodiazepine which binds to the benzodiazepine binding site of approximately 75% of the brain γ-aminobutyric acid-A receptors (GABAARs). Positron Emission Tomography (PET) imaging of the GABAARs with [11C]FMZ has been used to evidence alterations in neuronal density, to assess target engagement of novel pharmacological agents, and to study disorders such as epilepsy and Huntington's disease. Despite the potential of FMZ PET imaging the short half-life (t1/2) of carbon-11 (20 min) has limited the more widespread clinical use of [11C]FMZ. The fluorine-18 (18F) isotopologue with a longer t1/2 (110 min) is ideally suited to address this drawback. However, the majority of current radiochemical methods for the synthesis of [18F]FMZ are non-trivial and low yielding. We report a robust, automated protocol that is good manufacturing practice (GMP) compatible, and yields multi-patient doses of [18F]FMZ. RESULTS: The fully automated synthesis was developed on the Trasis AllinOne (AIO) platform using a single-use cassette. [18F]FMZ was synthesized in a one-step procedure from [18F]fluoride, via a copper-mediated 18F-fluorination of a boronate ester precursor. Purification was performed by semi-preparative radio-HPLC and the collected fraction formulated directly into the final product vial. The overall process from start of synthesis to delivery of product is approximately 55 min. Starting with an initial activity of 23.6 ± 5.8 GBq (n = 3) activity yields of [18F]FMZ were 8.0 ± 1 GBq (n = 3). The synthesis was successfully reproduced at two independent sites, where the product passed quality control release criteria in line with the European Pharmacopoeia standards and ICH Q3D(R1) guidelines to be suitable for human use. CONCLUSION: Reported is a fully automated cassette-based synthesis of [18F]FMZ that is Good Manufacturing Practice (GMP) compatible and produces multi-patient doses of [18F]FMZ.

Closing the gap between 19F and 18F chemistry.

Positron emission tomography (PET) has become an invaluable tool for drug discovery and diagnosis. The positron-emitting radionuclide fluorine-18 is frequently used in PET radiopharmaceuticals due to its advantageous characteristics; hence, methods streamlining access to 18F-labelled radiotracers can make a direct impact in medicine. For many years, access to 18F-labelled radiotracers was limited by the paucity of methodologies available, and the poor diversity of precursors amenable to 18F-incorporation. During the last two decades, 18F-radiochemistry has progressed at a fast pace with the appearance of numerous methodologies for late-stage 18F-incorporation onto complex molecules from a range of readily available precursors including those that do not require pre-functionalisation. Key to these advances is the inclusion of new activation modes to facilitate 18F-incorporation. Specifically, new advances in late-stage 19F-fluorination under transition metal catalysis, photoredox catalysis, and organocatalysis combined with the availability of novel 18F-labelled fluorination reagents have enabled the invention of novel processes for 18F-incorporation onto complex (bio)molecules. This review describes these major breakthroughs with a focus on methodologies for C-18F bond formation. This reinvigorated interest in 18F-radiochemistry that we have witnessed in recent years has made a direct impact on 19F-chemistry with many laboratories refocusing their efforts on the development of methods using nucleophilic fluoride instead of fluorination reagents derived from molecular fluorine gas.

Copper-Mediated Radiosynthesis of [18F]Rucaparib.

The poly(ADP-ribose) polymerase (PARP) inhibitor rucaparib is used in the clinic to treat BRCA-mutated cancers. Herein, we report two strategies to access the 18F-isotopologue of rucaparib by applying a copper-mediated nucleophilic 18F-fluorodeboronation. The most successful approach features an aldehydic boronic ester precursor that is subjected to reductive amination post-18F-labeling and affords [18F]rucaparib with an activity yield of 11% ± 3% (n = 3) and a molar activity (Am) up to 30 GBq/µmol. Preliminary in vitro studies are presented.

Direct Carbon Isotope Exchange of Pharmaceuticals via Reversible Decyanation.

The incorporation of carbon-14 allows tracking of organic molecules and provides vital knowledge on their fate. This information is critical in pharmaceutical development, crop science, and human food safety evaluation. Herein, a transition-metal-catalyzed procedure enabling carbon isotope exchange on aromatic nitriles is described. By utilizing the radiolabeled precursor Zn([14C]CN)2, this protocol allows the insertion of the desired carbon tag without the need for structural modifications, in a single step. By reducing synthetic costs and limiting the generation of radioactive waste, this procedure will facilitate the labeling of nitrile containing drugs and accelerate 14C-based ADME studies supporting drug development.

Carbon isotope labeling of carbamates by late-stage [11C], [13C] and [14C]carbon dioxide incorporation.

A general procedure for the late-stage [11C], [13C] and [14C]carbon isotope labeling of cyclic carbamates is reported. This protocol allows the incorporation of carbon dioxide, the primary source of carbon-14 and carbon-11 radioisotopes, in a direct, cost-effective and sustainable manner. A disconnection/reconnection strategy, involving ring opening/isotopic closure, was also implemented.

Transition-Metal-Free Carbon Isotope Exchange of Phenyl Acetic Acids.

A transition-metal-free carbon isotope exchange procedure on phenyl acetic acids is described. Utilizing the universal precursor CO2 , this protocol allows the carbon isotope to be inserted into the carboxylic acid position, with no need of precursor synthesis. This procedure enabled the labeling of 15 pharmaceuticals and was compatible with carbon isotopes [14 C] and [13 C]. A proof of concept with [11 C] was also obtained with low molar activity valuable for distribution studies.

Dynamic Carbon Isotope Exchange of Pharmaceuticals with Labeled CO2.

A copper-catalyzed procedure enabling dynamic carbon isotope exchange is described. Utilizing the universal precursor [14C]CO2, this protocol allows to insert, in one single step, the desired carbon tag into carboxylic acids with no need of structural modifications. Reducing synthetic costs and limiting the generation of radioactive waste, this procedure will facilitate the access to carboxylic acids containing drugs and accelerate early 14C-based ADME studies supporting drug development.

Recent developments in heterocycle labeling with carbon isotopes.

Heterocycles play an essential role in modern pharmaceutical and agrochemical developments, representing a very common structural unit in marketed drugs. Over the 46 new drugs approved in 2017 by the FDA, 25 contain in their structure a heterocyclic core. The development of novel and straightforward labeling strategies for the effective insertion of carbon isotopes into heterocylic scaffolds is an inspiring and vibrant field of research. The use of carbon-11, carbon-13, and carbon-14 isotopes is well established in life science and particularly in pharmaceutical and agrochemical industry. Their introduction into small organic molecules represents a crucial step for the radiochemists. Because the labeling should occur in metabolically stable positions and in the shortest synthetic route, their incorporation into the heterocycles represents a viable solution. This review summarizes recent contributions to this area of research through the analysis of different industrial and academic cases.