Clinical qualification of [18F] JNJ-64413739, a novel candidate PET ligand for the P2X7 receptor

Introduction: The P2X7 purinergic receptor is a membrane bound, ATP-gated cation channel embedded in the cell membrane. In the CNS, P2X7 receptors have been identified on several cell types with the most abundant expression on microglia. Activation of the P2X7 promotes the NLRP3 inflammasome assembly and release of neuroactive cytokines such as IL1β. Over-activation of this pathway is hypothesized to play a role in the pathophysiology of mood disorders. Selective, brain penetrant inhibitors of P2X7 have been developed as candidate therapeutics. A site-specific PET ligand for the P2X7 receptor could provide evidence of target engagement and assist in clinical dose selection. [18F] JNJ-64413739, a selective P2X7 inhibitor, has been extensively evaluated in preclinical models as a candidate PET ligand. We now report the clinical qualification of the ligand in healthy adult males including dosimetry, brain distribution, test-retest (TRT) variability, and blocking by pharmacological doses of the P2X7 inhibitor JNJ-54175446. Methods: For the radiation dosimetry 3 healthy male subjects were given a dose of ~185 MBq [18F] JNJ-64413739 and underwent 10 consecutive whole body PET/CT scans (Siemens Biograph) to assess the tracer biodistribution for approximately 5 hours post injection. Delineation of source organs allowed estimation of the normalized cumulated activity and to calculate the corresponding organ doses and effective dose (ED) with OLINDA. To evaluate the tracer distribution in the brain, 5 healthy male subjects were given 115-196 MBq [18F] JNJ-64413739 and completed a 120-minute dynamic PET scan (PETMR GE Signa) combined with arterial blood sampling and tracer metabolite analysis. 1 and 2 tissue compartment models (1-2TCM) and Logan graphical analysis (LGA) were evaluated to quantify tracer kinetics and calculate regional volumes of distribution (VT) (PMOD v.3.7). Retest scans were acquired for 3 of the 5 subjects (interscan interval 26-44 days). As part of a PET dose occupancy study, a subject underwent a baseline and post dose [18F] JNJ-64413739 PETMR scan. The latter was performed after oral dosing of JNJ-54175446 predicted to saturate binding based on preclinical PET. Occupancy was estimated by analyzing baseline and post dose VT values using a Lassen plot. Results: Average ED was 22 ± 1 µSv/MBq with the highest organ absorbed doses for gall bladder, urinary bladder, small and large intestine. 2TCM provide to be the most appropriate kinetic model to estimate regional VT with very similar estimates for LGA. Regional VT values were similar across all brain volumes with slightly higher values in thalamus, striatum, and brain stem. Inter subject variability of VT was relatively high with a composite cortical region of interest VT showing an approximate 3-fold range across subjects. Average regional TRT was 12.7 ± 8.4% by 2TCM and 13.5 ± 8.0 % by LGA while the acquisition time could be reduced to 90 minutes with only limited increase in bias and TRT variability. A single oral dose of JNJ-54175446 resulted in 78% occupancy across all brain regions and no reference region could be identified. Tracer metabolism was not altered by pretreatment with JNJ-54175446. Conclusions: [18F] JNJ-64413739 appears to be a suitable PET ligand for quantitation of P2X7 receptors in human brain. As such it can be used for testing target engagement by P2X7 inhibitors, assist in guiding dose selection of JNJ-54175446 for Phase 2 clinical trials, and provide insight into the expression of P2X7 receptors in health and disease.status: submitte

F-18-JNJ-64413739, a Novel PET Ligand for the P2X7 Ion Channel: Radiation Dosimetry, Kinetic Modeling, Test-Retest Variability, and Occupancy of the P2X7 Antagonist JNJ-54175446

The P2X7 receptor (P2X7R) is an adenosine triphosphate-gated ion channel that is predominantly expressed on microglial cells in the central nervous system. We report the clinical qualification of P2X7-specific PET ligand 18F-JNJ-64413739 in healthy volunteers, including dosimetry, kinetic modeling, test-retest variability, and blocking by the P2X7 antagonist JNJ-54175446. Methods: Whole-body dosimetry was performed in 3 healthy male subjects by consecutive whole-body PET/CT scanning, estimation of the normalized cumulated activity, and calculation of the effective dose using OLINDA (v1.1). Next, 5 healthy male subjects underwent a 120-min dynamic 18F-JNJ-64413739 PET/MRI scan with arterial blood sampling to determine the appropriate kinetic model. For this purpose, 1- and 2-tissue compartment models and Logan graphic analysis (LGA) were evaluated for estimating regional volumes of distribution (VT). PET/MRI scanning was repeated in 4 of these subjects to evaluate medium-term test-retest variability (interscan interval, 26-97 d). For the single-dose occupancy study, 8 healthy male subjects underwent baseline and postdose 18F-JNJ-64413739 PET/MRI scans 4-6 h after the administration of a single oral dose of JNJ-54175446 (dose range, 5-300 mg). P2X7 occupancies were estimated using a Lassen plot and regional baseline and postdose VTResults: The average (mean ± SD) effective dose was 22.0 ± 1.0 μSv/MBq. The 2-tissue compartment model was the most appropriate kinetic model, with LGA showing very similar results. Regional 2-tissue compartment model VT values were about 3 and were rather homogeneous across all brain regions, with slightly higher estimates for the thalamus, striatum, and brain stem. Between-subject VT variability was relatively high, with cortical VT showing an approximate 3-fold range across subjects. As for time stability, the acquisition time could be reduced to 90 min. The average regional test-retest variability values were 10.7% ± 2.2% for 2-tissue compartment model VT and 11.9% ± 2.2% for LGA VT P2X7 occupancy approached saturation for single doses of JNJ-54175446 higher than 50 mg, and no reference region could be identified. Conclusion:18F-JNJ-64413739 is a suitable PET ligand for the quantification of P2X7R expression in the human brain. It can be used to provide insight into P2X7R expression in health and disease, to evaluate target engagement by P2X7 antagonists, and to guide dose selection.status: publishe

Zoledronic acid treatment of 5T2MM-bearing mice inhibits the development of myeloma bone disease: evidence for decreased osteolysis, tumor burden and angiogenesis, and increased survival

Multiple myeloma is characterized by the growth of plasma cells in the bone marrow and the development of osteolytic bone disease. Myeloma cells are found closely associated with bone, and targeting this environment may therefore affect both the bone disease and the growth of myeloma cells. We have investigated the effect of the potent bisphosphonate, zoledronic acid, on the development of bone disease, tumor burden, and disease-free survival in the 5T2MM model of myeloma. 5T2MM murine myeloma cells were injected intravenously into C57BL/KaLwRij mice. After 8 weeks, all animals had a paraprotein. Animals were treated with zoledronic acid (120 microg/kg, subcutaneously, twice weekly) or vehicle, from the time of tumor cell injection or from paraprotein detection for 12 or 4 weeks, respectively. All animals injected with tumor cells developed osteolytic lesions, a decrease in cancellous bone volume, an increase in osteoclast perimeter, and a decrease in bone mineral density. Zoledronic acid prevented the formation of lesions, prevented cancellous bone loss and loss of bone mineral density, and reduced osteoclast perimeter. Zoledronic acid also decreased paraprotein concentration, decreased tumor burden, and reduced angiogenesis. In separate experiments, Kaplan-Meier analysis demonstrated a significant increase in survival after treatment with zoledronic acid when compared with control (47 vs. 35 days). A single dose of zoledronic acid was also shown to be effective in preventing the development of osteolytic bone disease. These data show that zoledronic acid is able to prevent the development of osteolytic bone disease, decrease tumor burden in bone, and increase survival in a model of established myeloma