ABSTRACT
Breast cancer resistance protein (BCRP, ABCG2) is an efflux transporter that plays a crucial role in multidrug resistance to antineoplastic drugs. Ko143, an analogue of the natural product fumitremorgin C, is a potent inhibitor of ABCG2 but is rapidly hydrolyzed to an inactive metabolite in vivo. To identify ABCG2 inhibitors with improved metabolic stability, we have assessed a series of Ko143 analogues for their ability to inhibit ABCG2-mediated transport in ABCG2-transduced MDCK II cells and determined the stability of the most potent compounds in liver microsomes. The most promising analogues were evaluated in vivo by positron emission tomography. In vitro, three of the tested analogues were potent ABCG2 inhibitors and stable in microsomes. In vivo, they increased the distribution of the ABCG2/ABCB1 substrate [11C]tariquidar to the brain both in wild-type (with Abcb1a/b transport blocked by tariquidar) and Abcb1a/b(-/-) mice. One analogue was more potent than Ko143 in both animal models.
Subject(s)
ATP-Binding Cassette Transporters , Antineoplastic Agents , Mice , Animals , ATP-Binding Cassette Transporters/metabolism , ATP Binding Cassette Transporter, Subfamily G, Member 2/metabolism , Neoplasm Proteins/metabolism , Brain/metabolism , Antineoplastic Agents/pharmacology , Antineoplastic Agents/metabolismABSTRACT
Monoacylglycerol lipase (MAGL) is a gatekeeper in regulating endocannabinoid signaling and has gained substantial attention as a therapeutic target for neurological disorders. We recently discovered a morpholin-3-one derivative as a novel scaffold for imaging MAGL via positron emission tomography (PET). However, its slow kinetics in vivo hampered the application. In this study, structural optimization was conducted and eleven novel MAGL inhibitors were designed and synthesized. Based on the results from MAGL inhibitory potency, in vitro metabolic stability and surface plasmon resonance assays, we identified compound 7 as a potential MAGL PET tracer candidate. [11C]7 was synthesized via direct 11CO2 fixation method and successfully mapped MAGL distribution patterns on rodent brains in in vitro autoradiography. PET studies in mice using [11C]7 demonstrated its improved kinetic profile compared to the lead structure. Its high specificity in vivo was proved by using MAGL KO mice. Although further studies confirmed that [11C]7 is a P-glycoprotein (P-gp) substrate in mice, its low P-gp efflux ratio on cells transfected with human protein suggests that it should not be an issue for the clinical translation of [11C]7 as a novel reversible MAGL PET tracer in human subjects. Overall, [11C]7 ([11C]RO7284390) showed promising results warranting further clinical evaluation.
Subject(s)
Monoacylglycerol Lipases , Tomography, X-Ray Computed , Animals , Mice , Humans , Monoacylglycerol Lipases/metabolism , Positron-Emission Tomography/methods , Brain/metabolism , Kinetics , Enzyme Inhibitors/chemistryABSTRACT
PURPOSE: The co-stimulatory molecules CD80 and CD86 are upregulated on activated antigen-presenting cells (APC). We investigated whether local APC activation, induced by subcutaneous (s.c.) inoculation of lipopolysaccharides (LPS), can be imaged by positron emission tomography (PET) with CD80/CD86-targeting 64Cu-labelled abatacept. PROCEDURES: Mice were inoculated s.c. with extracellular-matrix gel containing either LPS or vehicle (PBS). Immune cell populations were analysed by flow cytometry and marker expression by RT-qPCR. 64Cu-NODAGA-abatacept distribution was analysed using PET/CT and ex vivo biodistribution. RESULTS: The number of CD80+ and CD86+ immune cells at the LPS inoculation site significantly increased a few days after inoculation. CD68 and CD86 expression were higher at the LPS than the PBS inoculation site, and CD80 was only increased at the LPS inoculation site. CTLA-4 was highest 10 days after LPS inoculation, when CD80/CD86 decreased again. A few days after inoculation, 64Cu-NODAGA-abatacept distribution to the inoculation site was significantly higher for LPS than PBS (4.2-fold). Co-administration of unlabelled abatacept or human immunoglobulin reduced tracer uptake. The latter reduced the number of CD86+ immune cells at the LPS inoculation site. CONCLUSIONS: CD80 and CD86 are upregulated in an LPS-induced local inflammation, indicating invasion of activated APCs. 64Cu-NODAGA-abatacept PET allowed following APC activation over time.
Subject(s)
B7-1 Antigen/metabolism , B7-2 Antigen/metabolism , Inflammation/diagnostic imaging , Inflammation/metabolism , Abatacept/administration & dosage , Abatacept/pharmacokinetics , Animals , Copper Radioisotopes/pharmacokinetics , Immune Checkpoint Inhibitors/administration & dosage , Immune Checkpoint Inhibitors/pharmacokinetics , Inflammation/drug therapy , Inflammation/pathology , Lipopolysaccharides/administration & dosage , Male , Mice , Mice, Inbred C57BL , Models, Animal , Molecular Imaging/methods , Positron Emission Tomography Computed Tomography/methods , Radiopharmaceuticals/administration & dosage , Radiopharmaceuticals/pharmacokinetics , Tissue Distribution , Up-Regulation/drug effectsABSTRACT
The costimulatory molecule CD80 is an early marker for immune activation. It is upregulated on activated antigen-presenting cells. We aimed at developing a tracer for imaging CD80 by positron emission tomography (PET). Novel CD80 ligands were synthesized and tested by SPR for affinity to human CD80 (hCD80) and displacement of endogenous ligands. Several compounds bound with one-digit nanomolar affinity to hCD80 and displaced CTLA-4 and CD28 at nanomolar concentrations. A structure-affinity relationship study revealed relevant moieties for strong affinity to hCD80 and positions for further modifications. Lead compound MT107 (7f) was radiolabeled with carbon-11. In vitro, [11C]MT107 showed specific binding to hCD80-positive tissue and high plasma protein binding. In vivo, [11C]MT107 accumulated in liver, gall bladder, and intestines but only scarcely in hCD80-positive xenografts. The unfavorable in vivo performance may result from high plasma protein binding and extensive biliary excretion.
Subject(s)
B7-1 Antigen/analysis , Positron Emission Tomography Computed Tomography , Small Molecule Libraries/chemistry , Animals , Binding Sites , Humans , Mice , Mice, SCID , Molecular Structure , Neoplasms, Experimental/diagnostic imaging , Small Molecule Libraries/chemical synthesisABSTRACT
Physiologically based pharmacokinetic modelling (PBPK) is a powerful tool to predict in vivo pharmacokinetics based on physiological parameters and data from in vivo studies and in vitro assays. In vivo PBPK modelling in laboratory animals by noninvasive imaging could help to improve the in vivo-in vivo translation towards human pharmacokinetics modelling. We evaluated the feasibility of PBPK modelling with PET data from mice. We used data from two of our PET tracers under development, [11C]AM7 and [11C]MT107. PET images suggested hepatobiliary excretion which was reduced after cyclosporine administration. We fitted the time-activity curves of blood, liver, gallbladder/intestine, kidney, and peripheral tissue to a compartment model and compared the resulting pharmacokinetic parameters under control conditions ([11C]AM7 n = 2; [11C]MT107, n = 4) and after administration of cyclosporine ([11C]MT107, n = 4). The modelling revealed a significant reduction in [11C]MT107 hepatobiliary clearance from 35.2 ± 10.9 to 17.1 ± 5.6 µl/min after cyclosporine administration. The excretion profile of [11C]MT107 was shifted from predominantly hepatobiliary (CLH/CLR = 3.8 ± 3.0) to equal hepatobiliary and renal clearance (CLH/CLR = 0.9 ± 0.2). Our results show the potential of PBPK modelling for characterizing the in vivo effects of transporter inhibition on whole-body and organ-specific pharmacokinetics.
