PET/CT Imaging of 89Zr-N-sucDf-Pembrolizumab in Healthy Cynomolgus Monkeys.

PURPOSE: Programmed cell death-1 receptor (PD-1) and its ligand (PD-L1) are the targets for immunotherapy in many cancer types. Although PD-1 blockade has therapeutic effects, the efficacy differs between patients. Factors contributing to this variability are PD-L1 expression levels and immune cells present in tumors. However, it is not well understood how PD-1 expression in the tumor microenvironment impacts immunotherapy response. Thus, imaging of PD-1-expressing immune cells is of interest. This study aims to evaluate the biodistribution of Zirconium-89 (89Zr)-labeled pembrolizumab, a humanized IgG4 kappa monoclonal antibody targeting PD-1, in healthy cynomolgus monkeys as a translational model of tracking PD-1-positive immune cells. PROCEDURES: Pembrolizumab was conjugated with the tetrafluorophenol-N-succinyl desferal-Fe(III) ester (TFP-N-sucDf) and subsequently radiolabeled with 89Zr. Four cynomolgus monkeys with no previous exposure to humanized monoclonal antibodies received tracer only or tracer co-injected with pembrolizumab intravenously over 5 min. Thereafter, a static whole-body positron emission tomography (PET) scan was acquired with 10 min per bed position on days 0, 2, 5, and 7. Image-derived standardized uptake values (SUVmean) were quantified by region of interest (ROI) analysis. RESULTS: 89Zr-N-sucDf-pembrolizumab was synthesized with high radiochemical purity (> 99 %) and acceptable molar activity (> 7 MBq/nmol). In animals dosed with tracer only, 89Zr-N-sucDf-pembrolizumab distribution in lymphoid tissues such as mesenteric lymph nodes, spleen, and tonsils increased over time. Except for the liver, low radiotracer distribution was observed in all non-lymphoid tissue including the lung, muscle, brain, heart, and kidney. When a large excess of pembrolizumab was co-administered with a radiotracer, accumulation in the lymph nodes, spleen, and tonsils was reduced, suggestive of target-mediated accumulation. CONCLUSIONS: 89Zr-N-sucDf-pembrolizumab shows preferential uptake in the lymphoid tissues including the lymph nodes, spleen, and tonsils. 89Zr-N-sucDf-pembrolizumab may be useful in tracking the distribution of a subset of immune cells in non-human primates and humans. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02760225.

Preclinical evaluation of [11 C]L-235 as a radioligand for Positron Emission Tomography cathepsin K imaging in bone.

The cathepsin K (CatK) enzyme is abundantly expressed in osteoclasts, and CatK inhibitors have been developed for the treatment of osteoporosis. In our effort to support discovery and clinical evaluations of a CatK inhibitor, we sought to discover a radioligand to determine target engagement of the enzyme by therapeutic candidates using positron emission tomography (PET). L-235, a potent and selective CatK inhibitor, was labeled with carbon-11. PET imaging studies recording baseline distribution of [11 C]L-235, and chase and blocking studies using the selective CatK inhibitor MK-0674 were performed in juvenile and adult nonhuman primates (NHP) and ovariectomized rabbits. Retention of the PET tracer in regions expected to be osteoclast-rich compared with osteoclast-poor regions was examined. Increased retention of the radioligand was observed in osteoclast-rich regions of juvenile rabbits and NHP but not in the adult monkey or adult ovariectomized rabbit. Target engagement of CatK was observed in blocking studies with MK-0674, and the radioligand retention was shown to be sensitive to the level of MK-0674 exposure. [11 C]L-235 can assess target engagement of CatK in bone only in juvenile animals. [11 C]L-235 may be a useful tool for guiding the discovery of CatK inhibitors.

In Vivo Evaluation and Dosimetry Estimate for a High Affinity Affibody PET Tracer Targeting PD-L1.

PURPOSE: In vivo imaging of programmed death ligand 1 (PD-L1) during immunotherapy could potentially monitor changing PD-L1 expression and PD-L1 expression heterogeneity within and across tumors. Some protein constructs can be used for same-day positron emission tomography (PET) imaging. Previously, we evaluated the PD-L1-targeting Affibody molecule [18F]AlF-NOTA-ZPD-L1_1 as a PET tracer in a mouse tumor model of human PD-L1 expression. In this study, we evaluated the affinity-matured Affibody molecule ZPD-L1_4, to determine if improved affinity for PD-L1 resulted in increased in vivo targeting of PD-L1. PROCEDURES: ZPD-L1_4 was conjugated with NOTA and radiolabeled with either [18F]AlF or 68Ga. [18F]AlF-NOTA-ZPD-L1_4 and [68Ga]NOTA-ZPD-L1_4 were evaluated in immunocompromised mice with LOX (PD-L1+) and SUDHL6 (PD-L1-) tumors with PET and ex vivo biodistribution measurements. In addition, whole-body PET studies were performed in rhesus monkeys to predict human biodistribution in a model with tracer binding to endogenous PD-L1, and to calculate absorbed radiation doses. RESULTS: Ex vivo biodistribution measurements showed that both tracers had > 25 fold higher accumulation in LOX tumors than SUDHL6 ([18F]AlF-NOTA-ZPD-L1_4: LOX: 8.7 ± 0.7 %ID/g (N = 4) SUDHL6: 0.2 ± 0.01 %ID/g (N = 6), [68Ga]NOTA-ZPD-L1_4: LOX: 15.8 ± 1.0 %ID/g (N = 6) SUDHL6: 0.6 ± 0.1 %ID/g (N = 6)), considerably higher than ZPD-L1_1. In rhesus monkeys, both PET tracers showed fast clearance through kidneys and low background signal in the liver ([18F]AlF-NOTA-ZPD-L1_4: 1.26 ± 0.13 SUV, [68Ga]NOTA-ZPD-L1_4: 1.11 ± 0.06 SUV). PD-L1-expressing lymph nodes were visible in PET images, indicating in vivo PD-L1 targeting. Dosimetry estimates suggest that both PET tracers can be used for repeated clinical studies, although high kidney accumulation may limit allowable radioactive doses. CONCLUSIONS: [18F]AlF-NOTA-ZPD-L1_4 and [68Ga]NOTA-ZPD-L1_4 are promising candidates for same-day clinical PD-L1 PET imaging, warranting clinical evaluation. The ability to use either [18F] or [68Ga] may expand access to clinical sites.

Systemic pan-AMPK activator MK-8722 improves glucose homeostasis but induces cardiac hypertrophy.

5'-Adenosine monophosphate-activated protein kinase (AMPK) is a master regulator of energy homeostasis in eukaryotes. Despite three decades of investigation, the biological roles of AMPK and its potential as a drug target remain incompletely understood, largely because of a lack of optimized pharmacological tools. We developed MK-8722, a potent, direct, allosteric activator of all 12 mammalian AMPK complexes. In rodents and rhesus monkeys, MK-8722-mediated AMPK activation in skeletal muscle induced robust, durable, insulin-independent glucose uptake and glycogen synthesis, with resultant improvements in glycemia and no evidence of hypoglycemia. These effects translated across species, including diabetic rhesus monkeys, but manifested with concomitant cardiac hypertrophy and increased cardiac glycogen without apparent functional sequelae.

Identification and in Vivo Evaluation of Liver X Receptor ß-Selective Agonists for the Potential Treatment of Alzheimer's Disease.

Herein, we describe the development of a functionally selective liver X receptor ß (LXRß) agonist series optimized for Emax selectivity, solubility, and physical properties to allow efficacy and safety studies in vivo. Compound 9 showed central pharmacodynamic effects in rodent models, evidenced by statistically significant increases in apolipoprotein E (apoE) and ATP-binding cassette transporter levels in the brain, along with a greatly improved peripheral lipid safety profile when compared to those of full dual agonists. These findings were replicated by subchronic dosing studies in non-human primates, where cerebrospinal fluid levels of apoE and amyloid-ß peptides were increased concomitantly with an improved peripheral lipid profile relative to that of nonselective compounds. These results suggest that optimization of LXR agonists for Emax selectivity may have the potential to circumvent the adverse lipid-related effects of hepatic LXR activity.