EJNMMI Radiopharmacy and Chemistry最新文献

Background

Hsp90 is a molecular chaperone that is often overexpressed across multiple cancer types and has a potential value as a prognostic marker as well as a therapeutic target. Given the high interest in Hsp90 therapies, positron emission tomography or PET imaging of Hsp90 can be a valuable tool for patient selection. The limitations of the previously developed Hsp90 tracers prompted us to evaluate the recently developed brain-permeable [¹¹C]HSP990 PET probe to advance the development of Hsp90-targeted therapeutics. Given the brain accumulation of [¹¹C]HSP990 probe, application for glioblastoma imaging of this tracer is of particular interest.

Results

In vitro [¹¹C]HSP990 binding was assessed in breast cancer and glioma cell lines including patient-derived cells using Hsp90 inhibitors and RNA interference knockdown of Hsp90 isoforms. Saturation binding studies were conducted on these cells and tumour tissue homogenates, and autoradiography was performed on tissue sections. Ex vivo biodistribution and in vivo dynamic µPET/CT studies were performed in healthy mice and tumour-bearing mice, including immunocompromised subcutaneous human U87 and MDA-MB-231models and immunocompetent intracranial murine NS/CT-2A models at baseline and following a pre-treatment with Hsp90 inhibitors. High Hsp90-specific tracer uptake was observed in breast cancer and glioma cells, with Hsp90β inhibition resulting in the most substantial reduction in uptake. In vivo uptake was high in U87 tumours but low in MDA-MB-231, presumably due to the differences in Hsp90 expression in tumour tissue versus cultured cells. Differences in maximum binding capacity or B_max across cell and tissue types support this hypothesis, especially given that the affinity measured as dissociation constant K_d remained similar across all tissue types. Despite high NS/CT-2A tumour uptake in vitro, no contrast between the healthy brain tissue and the NS/CT-2A glioma was observed in vivo due to the high uptake by the healthy brain.

Conclusion

[¹¹C]HSP990 is a promising tracer for identifying Hsp90-overexpressing tumours and may hold potential for patient stratification, prognosis, and therapy monitoring of novel Hsp90 therapeutics. High healthy brain uptake of this tracer precluded the differentiation of the tumour in the intracranial NS/CT-2A tumour model, therefore [¹¹C]HSP990 might not be a suitable tracer for the glioblastoma imaging. Tracer with a longer half-life might be needed to compare the washout of the tracer from the brain and the tumour tissue over several hours to identify a suitable imaging window.

Background

Prostate-specific membrane antigen (PSMA) is an established target for the imaging and treatment of prostate cancer. This study focused on the preclinical evaluation of three novel PSMA inhibitors—P15, P16, and P19—which were structurally modified compared to the clinically used PSMA-617. Two main strategies were pursued: a chemical approach following the so-called reversed synthetic strategy, and the replacement of the naphthyl-based linker moiety with an analogous diphenyl-based moiety. The aim was to assess the impact of these modifications on physicochemical properties, in vitro behaviour, and in vivo pharmacokinetics following radiolabelling with ⁶⁸Ga.

Results

Radiolabelling of all three novel compounds with ⁶⁸Ga resulted in high radiochemical purity above 98% under physiological pH conditions and above 97% during stability testing in human plasma. All compounds exhibited hydrophilic characteristics based on partition coefficient measurements. Notable differences were observed in plasma protein binding, with P15 and P16 showing significantly lower binding compared to PSMA-617 and P19. In vitro assays using LNCaP prostate cancer cells demonstrated similar cellular uptake and internalization across all tested compounds. In vivo evaluation using Positron Emission Tomography/Computed Tomography (PET/CT) imaging in LNCaP tumour-bearing mice confirmed the tumour-targeting ability of all three inhibitors. These findings were further supported by biodistribution studies, which highlighted efficient and specific accumulation in tumour tissue. Among the tested compounds, P19 demonstrated the most promising overall profile in terms of stability, binding characteristics, and tumour uptake.

Conclusions

The stereochemical modifications in the linker region significantly influenced the in vitro and in vivo behaviour of the PSMA inhibitors. Despite similar cellular uptake, differences in plasma protein binding and pharmacokinetics were evident. Among the three novel compounds, P19 emerged as a particularly promising candidate for further investigation, also indicating that the diphenyl moiety might serve as a favourable linker building block in analogy to the naphthyl moiety. Our observations suggest potential not only for diagnostic imaging with ⁶⁸Ga, but also for therapeutic applications using ¹⁷⁷Lu, which offers a longer half-life suitable for delayed imaging and treatment intervals in prostate cancer management.

Background

The March 2022 approval of [¹⁷⁷Lu]Lu-PSMA-617—the first targeted radioligand therapy (TRT) for metastatic castration-resistant prostate cancer (mCRPC)—addressed a critical need in therapeutic diagnostics. However, approximately 30% of patients fail to respond to PSMA-targeted therapies, with suboptimal pharmacokinetics implicated as a key factor. To address this challenge, fatty acids, known for their role in binding albumin and enhancing drug pharmacokinetics, were explored. Although commonly used in small molecules and peptides, their application in radiopharmaceuticals remains limited. In response, a novel fatty acid-modified PSMA-targeting agent was developed to optimize pharmacokinetics by enhancing circulation half-life, tumor uptake, radioligand selectivity, and safety while reducing off-target accumulation. This innovative approach aims to maximize therapeutic efficacy, improve patient outcomes in mCRPC care, and reduce the overall treatment burden.

Results

In vitro studies confirmed the strong binding affinity and high specificity of [¹⁷⁷Lu]Lu-BT-117016 for PSMA. Biodistribution studies in LNCaP clone FGC tumor-bearing mice demonstrated significantly enhanced tumor uptake and retention of [¹⁷⁷Lu]Lu-BT-117016 compared to [¹⁷⁷Lu]Lu-PSMA-617. SPECT/CT imaging and therapy studies highlighted the superior profile of [¹⁷⁷Lu]Lu-BT-117016. It achieved significant tumor growth suppression with favorable tolerability at a minimal dose of 3 MBq, demonstrating comparable efficacy to a nearly 6-fold higher dose (20 MBq) of [177Lu]Lu-PSMA-617. The improved safety profile of [¹⁷⁷Lu]Lu-BT-117016 over [¹⁷⁷Lu]Lu-PSMA-617 was confirmed through radiation dosimetry.

Conclusion

Preclinical studies demonstrate that [¹⁷⁷Lu]Lu-BT-117016, a novel fatty acid-modified PSMA-targeting agent, optimizes albumin binding and harmonizes ligand-isotope half-lives. This results in advantageous biodistribution—characterized by high tumor uptake, rapid non-target clearance, and extended circulation—leading to complete tumor remission in LNCaP models with reduced toxicity. Crucially, [¹⁷⁷Lu]Lu-BT-117016 achieved equivalent efficacy to [¹⁷⁷Lu]Lu-PSMA-617 at 3–6 times lower doses. These findings indicate [¹⁷⁷Lu]Lu-BT-117016’s potential as a safer, more potent CRPC therapy enabling reduced dosages and less frequent administration. Further clinical trials are warranted to confirm these benefits.

Background

Alzheimer’s disease (AD) is increasingly recognized as a multifactorial disorder with vascular contributions, including a pro-coagulant state marked by fibrin deposition in the brain. Fibrin accumulation may exacerbate cerebral hypoperfusion and neuroinflammation, leading to neurodegeneration. Identifying patients with this pathology could enable targeted anticoagulant therapy. However, current imaging tools lack the specificity and sensitivity to detect fibrin in the brain non-invasively. This study aimed to develop and evaluate brain-penetrating peptide- and antibody-based PET radioligands targeting fibrin to enable individualized treatment strategies in AD.

Results

A fibrin-binding peptide (FBP) was conjugated to the antibody fragment scFv8D3, which targets the transferrin receptor (TfR), to facilitate transcytosis across the blood-brain barrier. FBP-scFv8D3 bound TfR and with modest affinity to fibrin. In vivo studies in Tg-ArcSwe mice, that exhibit fibrin along with brain amyloid-β pathology, and wild-type mice showed that [¹²⁵I]FBP-scFv8D3 retained brain-penetrating properties but did not demonstrate significant fibrin-specific retention. In contrast, the monoclonal antibody 1101 and its bispecific, brain penetrant variant 1101-scFv8D3 exhibited higher fibrin selectivity and TfR binding. Both antibodies showed a trend towards higher brain retention in Tg-ArcSwe mice and [¹²⁵I]1101-scFv8D3 showed a higher brain-to-blood ratio compared to [¹²⁴I]1101. PET imaging with [¹²⁴I]1101 and [¹²⁴I]1101-scFv8D3 revealed low global brain uptake. However, ex vivo autoradiography and regional PET quantification (ROI-to-cerebellum ratios) indicated significant cortical and caudate retention of [¹²⁴I]1101-scFv8D3 in Tg-ArcSwe mice, supporting region-specific target engagement.

Conclusion

This proof-of-concept study demonstrates the feasibility of using bispecific antibody-based PET radioligands to target fibrin in the AD brain. While the FBP-scFv8D3 conjugate showed limited specificity, the bispecific antibody 1101-scFv8D3 exhibited higher brain penetration and fibrin selectivity. These findings support further development of antibody-based imaging tools toward the goal to stratify AD patients who may benefit from anticoagulant therapy.

Background

Copper-mediated radiofluorination (CMRF) is a breakthrough in ¹⁸F-radiochemistry, enabling ¹⁸F incorporation into molecules even at electron-rich aromatic positions. In recent years, several improved protocols have been reported to advance the application of CMRF. These advancements primarily focus on improving radiochemical conversion, expanding substrate scope, and enabling scalability for remote-controlled radiotracer production. Despite these improvements, one major challenge remains: the protodemetallation. Protodemetallation is a common side reaction in transition metal-mediated cross-couplings that takes place by a mechanism that is not yet fully elucidated. In ¹⁸F-chemistry, the formation of the hydrogenated side product (HSP) can interfere with the chromatographic purification of the desired radiotracer, resulting in complex radiotracer production.

Results

The present work investigates the factors influencing the rate of the hydrogenation reaction as well as the source of hydrogen in the CMRF by use of model precursors bearing -B(OH)₂, -Bpin, -BEpin and -SnBu₃ as leaving groups. While the CMRF reactions are usually carried out under anhydrous conditions, the formation rate of the HSP was evaluated by controlling the chemical constituents (type and molarity of reagents) as well as the physical parameters (time and temperature). Moreover, experiments with deuterated reagents complemented by high-resolution mass spectrometry (HRMS) analysis were carried out to identify the source of hydrogen for the reductive elimination step.

Conclusion

This study identifies reaction parameters that influence hydrogenation side reactions in CMRF, enabling high RCC with minimal HSP formation. The optimal reaction conditions include low temperature, short reaction time, and minimal amount of precursor, copper, and ideally no base and alcohols as solvents. Among the evaluated precursors, –BEpin afforded the lowest HSP formation, while –B(OH)₂ afforded the highest. Overall, this study showed that the selection of proper reaction reagents and the fine-tuning of reaction parameters can substantially reduce the HSP formation while maintaining optimal radiochemical conversion.

Graphical abstract

Background

 Bladder cancer remains a significant global health challenge, with approximately 75% of cases presenting as non-muscle-invasive bladder cancer. Despite standard treatment with transurethral resection and intravesical Bacillus Calmette-Guérin immunotherapy, up to 40% of patients develop resistance or progress to muscle-invasive disease. Targeted alpha-emitting radionuclide therapy offers promising therapeutic potential through the selective delivery of high linear energy transfer radiation to tumor cells while minimizing damage to healthy tissues. PTK7 is overexpressed in various malignancies, including bladder cancer, and is therefore a viable therapeutic target. This study evaluated the preclinical efficacy of [²¹²Pb]Pb-TCMC-chOI-1, a ²¹²Pb-labeled antibody targeting PTK7, for targeted alpha-emitting radionuclide therapy in bladder cancer using 2D adherent cultures (clonogenic assay) and 3D multicellular spheroid models (spheroid growth inhibition).

Results

 PTK7 expression analysis revealed varying antigen densities across five bladder cancer cell lines, ranging from approximately 10,000 to 70,000 sites per cell. The chimeric anti-PTK7 antibody demonstrated apparent equilibrium dissociation constants of 10–44 nM with moderate binding affinity suitable for therapeutic applications. [²¹²Pb]Pb-TCMC-chOI-1 treatment resulted in activity- and time-dependent cytotoxicity, with enhanced sensitivity observed in cell lines with higher PTK7 levels. In clonogenic assays, the activity concentration required for 50% growth reduction was 48–74 kBq/mL, corresponding to 22–51 bound and 9–16 internalized ²¹²Pb atoms per cell. In 3D models, similar therapeutic effects were observed despite significantly lower activities (values of approximately 1 and 30 kBq/mL for KU-19–19 and 647-V cells, respectively), suggesting a more pronounced cross-fire effect. Flow cytometry demonstrated treatment-induced DNA damage, cell cycle perturbations and cell death, with response patterns correlating with overall treatment sensitivity. RT-112 and KU-19–19 cells showed superior responses compared to 647-V and T-24 cells, consistent with their higher PTK7 expression.

Conclusions

 These findings support PTK7 as a therapeutic target for bladder cancer and demonstrate the potential of [²¹²Pb]Pb-TCMC-chOI-1 for targeted alpha-emitting radionuclide therapy. The results provide a rationale for further preclinical optimization of this therapeutic approach.

Trial registration number (TRN): Not applicable.

Background

Recent EMA and FDA approvals of Lu-DOTATATE and Lu-PSMA-617 have led to increased demand for radiotherapeutic (^{177})Lu, due to its promising potential to treat castration-resistant neuroendocrine cancers. Conventional reactor production methods pose challenges related to cost, waste management, and local availability. In comparison, accelerators produce less waste, have lower maintenance costs, and can be directly integrated into hospital settings. In this study, we evaluate the production of radiotherapeutic (^{177})Lu using a 10 mA, 18 MeV (D^+) compact linear accelerator design. The design consists of a single radio-frequency quadrupole (RFQ) and seven drift tube linacs (DTLs) that achieve a beam efficiency of 98.5% over a total length of (12,text {m}). Deuteron activations on a 99% enriched [(^{176})Yb](hbox {Yb}_2hbox {O}_3) target are estimated using experimental and simulated excitation functions.

Results

A circular target with a radius of 1 cm and 0.36 mm thickness is selected to optimize the yield of (^{177})Lu while minimizing the production of undesirable radioisotopes, including (^{174g+m})Lu and (^{177m})Lu. Model calculations indicate that the accelerator design can produce 11.3 μg of (^{177})Lu per hour. A 5-day irradiation is expected to yield approximately 1.07 mg of (^{177})Lu (4.4 TBq), while a 12-day irradiation can produce up to 1.9 mg (7.8 TBq). Following a 2-day processing period, the specific activity of the 5-day irradiated sample is projected to approach 0.6 TBq/mg, with a radiopurity of approximately 99.8%. The minimal burn-up of the (hbox {Yb}_2hbox {O}_3) target suggests it may be recycled and reused over multiple irradiations.

Conclusions

The study confirms the feasibility of accelerator-based (^{177})Lu production as an alternative to existing reactor-based methods. The 10 mA, 18 MeV (D^+) RFQ-DTL design achieves an exceptionally high (^{177})Lu radiopurity and a competitive overall yield, which can meet the dose requirements of thousands of patients.

Background

Fibroblast activation protein (FAP)-targeting radioligands have gained attention for the ability to image multiple tumor types. Current FAP-targeting radioligands are labeled with ⁶⁸Ga and ¹⁸F, but their short half-lives limit distribution range after production and later time-point imaging. This study describes the development Kalios, a novel class of NODAGA-conjugated FAP-targeting radioligands labeled with the cyclotron-produced Copper-61 (t_1/2 = 3.33 h), for greater temporal range for FAP-targeted imaging.

Results

Four Kalios ligands were synthesized and radiolabeled with [⁶¹Cu]CuCl₂ in high yield and radiochemical purity within 5 min at room temperature. All radioligands demonstrated high hydrophilicity and strong affinity for FAP, and were primarily internalized after incubation with FAP-positive cells. PET/CT images obtained at 0–1 h and 4 h post-injection (p.i.) illustrated accumulation of all radioligands in FAP-positive tumors. Biodistribution studies of [⁶¹Cu]Cu-Kalios-02 demonstrated stable tumor uptake between 1 and 4 h p.i., with washout from normal tissues at 4 h, resulting in improved tumor-to-background ratios.

Conclusions

Kalios ligands represent a new class of FAP-targeting ⁶¹Cu-labeled radioligands. The half-life of ⁶¹Cu allowed delayed 4-h imaging with improved tumor-to-background ratios. The improved delayed imaging and greater distribution range of these ⁶¹Cu-labeled FAP-targeting radioligands demonstrates their clear potential for clinical translation, while combination with the therapeutic twin ⁶⁷Cu allows for truly paired Kalios theranostics.

Background

Mesothelin is a glycoprotein overexpressed in various cancers, with limited expression in healthy tissues. The single-domain antibody (sdAb, or nanobody) A1-His has previously successfully been validated in mice for the SPECT imaging of mesothelin positive tumors following radiolabeling with ^99mTc. Our objective was to radiolabel this sdAb with ⁶⁸Ga for PET imaging, exhibiting superior sensitivity and resolution than SPECT in clinical practice. To this aim, it was conjugated to NOTA chelator that is commonly employed for ⁶⁸Ga labeling of antibody-derived tracers. In addition, the high affinity and specificity of A1-His sdAb position it as a promising candidate for theranostic applications. In anticipation of future radiolabeling with beta-emitting radionuclides, DOTA-conjugated A1-His was also evaluated. Given the high thermal stability of sdAbs, this DOTA-conjugated sdAb could potentially be implemented in future studies as a theranostic agent with beta-emitting radionuclides.

Results

A1-His was successfully conjugated to p-SCN-Bn-DOTA and p-SCN-Bn-NOTA under optimized conditions, achieving chelator-to-sdAb ratios of 1.8 and 1.3, respectively. NOTA-A1-His allowed rapid radiolabeling with ⁶⁸Ga at room temperature, achieving high radiochemical purity (> 98%) within 5 min. Using DOTA, similar purity was obtained at 60 °C for 15 min. Both radiotracers demonstrated stability over 4 h in the radiolabeling medium and 2 h in human blood. However, some instability was observed in murine blood. Biodistribution and imaging studies in mice bearing mesothelin-expressing tumors showed specific tumor targeting for both tracers. Notably, [68Ga]Ga-DOTA-A1-His exhibited twofold lower kidney uptake compared to [68Ga]Ga-NOTA-A1-His, potentially enhancing imaging contrast and reducing renal radiation exposure. His-tag removal, further improves the biodistribution profile of the 2 tracers.

Conclusions

Both p-SCN-Bn-DOTA and p-SCN-Bn-NOTA chelators can be effectively conjugated to the A1 sdAb and radiolabeled with ⁶⁸Ga, producing stable radiotracers with specific tumor-targeting capabilities. NOTA chelator offers advantages in rapid, room-temperature radiolabeling. However, DOTA would offer the advantage to be employed for theranostic approaches using β⁻ emitters such as ¹⁷⁷Lu or ¹⁶¹Tb. The lower kidney retention of DOTA-A1 also suggests that its dosimetry, a key factor in theranostic, would be more favorable.