Deficiency in PHD2-mediated hydroxylation of HIF2α underlies Pacak-Zhuang syndrome.

Pacak-Zhuang syndrome is caused by mutations in the EPAS1 gene, which encodes for one of the three hypoxia-inducible factor alpha (HIFα) paralogs HIF2α and is associated with defined but varied phenotypic presentations including neuroendocrine tumors and polycythemia. However, the mechanisms underlying the complex genotype-phenotype correlations remain incompletely understood. Here, we devised a quantitative method for determining the dissociation constant (Kd) of the HIF2α peptides containing disease-associated mutations and the catalytic domain of prolyl-hydroxylase (PHD2) using microscale thermophoresis (MST) and showed that neuroendocrine-associated Class 1 HIF2α mutants have distinctly higher Kd than the exclusively polycythemia-associated Class 2 HIF2α mutants. Based on the co-crystal structure of PHD2/HIF2α peptide complex at 1.8 Å resolution, we showed that the Class 1 mutated residues are localized to the critical interface between HIF2α and PHD2, adjacent to the PHD2 active catalytic site, while Class 2 mutated residues are localized to the more flexible region of HIF2α that makes less contact with PHD2. Concordantly, Class 1 mutations were found to significantly increase HIF2α-mediated transcriptional activation in cellulo compared to Class 2 counterparts. These results reveal a structural mechanism in which the strength of the interaction between HIF2α and PHD2 is at the root of the general genotype-phenotype correlations observed in Pacak-Zhuang syndrome.

Measuring Oxygenation in Newborn Infants with Targeted Oxygen Ranges (MONITOR): a randomised crossover pilot study.

OBJECTIVE: The Neonatal Oxygenation Prospective Meta-analysis (NeOProM) Collaboration showed that high (91-95%) versus low (85-89%) SpO2 targets reduced mortality. Trials of higher targets are needed to determine whether any more survival advantage may be gained. This pilot study explored the achieved oxygenation patterns observed when targeting SpO2 92-97% to facilitate the design of future trials. DESIGN: Single-centre prospective randomised crossover pilot study. Manual FiO2 adjustment. Study time 12 hours per infant. 6 hours targeting SpO2 90-95% and 6 hours targeting SpO2 92-97%. PATIENTS: Twenty preterm infants born <29 weeks' gestation, greater than 48 hours old, receiving supplemental oxygen. OUTCOMES: Primary outcome was percentage time with SpO2 above 97% and below 90%. Pre-defined secondary outcomes included percentage time spent within, above or below transcutaneous PO2 (TcPO2) 6.7-10.7 kPa (50-80 mm Hg). Comparisons were made using paired-samples t-test (2-tailed). RESULTS: With SpO2 target 92-97% versus 90-95%, the mean (IQR) percentage time above SpO2 97% was 11.3% (2.7-20.9) versus 7.8% (1.7-13.9), p=0.02. Percentage time with SpO2 <90% was 13.1% (6.7-19.1) versus 17.9% (11.1-22.4), p=0.003. Percentage time with SpO2 <80% was 1% (0.1-1.4) versus 1.6% (0.4-2.6), p=0.119. Percentage time with TcPO2 <6.7 kPa (50 mm Hg) was 49.6% (30.2-66.0) versus 55% (34.3-73.5), p=0.63. Percentage time above TcPO2 10.7 kPa (80 mm Hg) was 1.4% (0-1.4) versus 1.8% (0-0), p=0.746. CONCLUSIONS: Targeting SpO2 92-97% produced a right shift in SpO2 and TcPO2 distribution, with reduced time at SpO2 <90% and increased time at SpO2 >97%, without increasing time with TcPO2 >10.7 kPa (80 mm Hg). Clinical trials targeting this higher SpO2 range could be conducted without significant hyperoxic exposure. TRIAL REGISTRATION NUMBER: NCT03360292.

Mitochondrial damage activates the NLRP10 inflammasome.

Upon detecting pathogens or cell stress, several NOD-like receptors (NLRs) form inflammasome complexes with the adapter ASC and caspase-1, inducing gasdermin D (GSDMD)-dependent cell death and maturation and release of IL-1ß and IL-18. The triggers and activation mechanisms of several inflammasome-forming sensors are not well understood. Here we show that mitochondrial damage activates the NLRP10 inflammasome, leading to ASC speck formation and caspase-1-dependent cytokine release. While the AIM2 inflammasome can also sense mitochondrial demise by detecting mitochondrial DNA (mtDNA) in the cytosol, NLRP10 monitors mitochondrial integrity in an mtDNA-independent manner, suggesting the recognition of distinct molecular entities displayed by the damaged organelles. NLRP10 is highly expressed in differentiated human keratinocytes, in which it can also assemble an inflammasome. Our study shows that this inflammasome surveils mitochondrial integrity. These findings might also lead to a better understanding of mitochondria-linked inflammatory diseases.

Hypoxia-inducible factor underlies von Hippel-Lindau disease stigmata.

von Hippel-Lindau (VHL) disease is a rare hereditary cancer syndrome that causes a predisposition to renal clear-cell carcinoma, hemangioblastoma, pheochromocytoma, and autosomal-recessive familial polycythemia. pVHL is the substrate conferring subunit of an E3 ubiquitin ligase complex that binds to the three hypoxia-inducible factor alpha subunits (HIF1-3α) for polyubiquitylation under conditions of normoxia, targeting them for immediate degradation by the proteasome. Certain mutations in pVHL have been determined to be causative of VHL disease through the disruption of HIFα degradation. However, it remains a focus of investigation and debate whether the disruption of HIFα degradation alone is sufficient to explain the complex genotype-phenotype relationship of VHL disease or whether the other lesser or yet characterized substrates and functions of pVHL impact the development of the VHL disease stigmata; the elucidation of which would have a significant ramification to the direction of research efforts and future management and care of VHL patients and for those manifesting sporadic counterparts of VHL disease. Here, we examine the current literature including the other emergent pseudohypoxic diseases and propose that the VHL disease-phenotypic spectrum could be explained solely by the varied disruption of HIFα signaling upon the loss or mutation in pVHL.

Direct H2O2 Synthesis, without H2 Gas.

We report here the direct hydrogenation of O2 gas to form hydrogen peroxide (H2O2) using a membrane reactor without H2 gas. Hydrogen is sourced from water, and the reactor is driven by electricity. Hydrogenation chemistry is achieved using a hydrogen-permeable Pd foil that separates an electrolysis chamber that generates reactive H atoms, from a hydrogenation chamber where H atoms react with O2 to form H2O2. Our results show that the concentration of H2O2 can be increased ∼8 times (from 56.5 to 443 mg/L) by optimizing the ratio of methanol-to-water in the chemical chamber, and through catalyst design. We demonstrate that the concentration of H2O2 is acutely sensitive to the H2O2 decomposition rate. This decomposition rate can be minimized by using AuPd alloy catalysts instead of pure Pd. This study presents a new pathway to directly synthesize H2O2 using water electrolysis without ever using H2 gas.

A self-driving laboratory advances the Pareto front for material properties.

Useful materials must satisfy multiple objectives, where the optimization of one objective is often at the expense of another. The Pareto front reports the optimal trade-offs between these conflicting objectives. Here we use a self-driving laboratory, Ada, to define the Pareto front of conductivities and processing temperatures for palladium films formed by combustion synthesis. Ada discovers new synthesis conditions that yield metallic films at lower processing temperatures (below 200 °C) relative to the prior art for this technique (250 °C). This temperature difference makes possible the coating of different commodity plastic materials (e.g., Nafion, polyethersulfone). These combustion synthesis conditions enable us to to spray coat uniform palladium films with moderate conductivity (1.1 × 105 S m-1) at 191 °C. Spray coating at 226 °C yields films with conductivities (2.0 × 106 S m-1) comparable to those of sputtered films (2.0 to 5.8 × 106 S m-1). This work shows how a self-driving laboratoy can discover materials that provide optimal trade-offs between conflicting objectives.

A C-Terminally Truncated Variant of Neurospora crassa VDAC Assembles Into a Partially Functional Form in the Mitochondrial Outer Membrane and Forms Multimers in vitro.

The voltage-dependent anion-selective channel (VDAC) is a porin in the mitochondrial outer membrane (MOM). Unlike bacterial porins, several mitochondrial ß-barrels comprise an odd number of ß-strands, as is the case for the 19-ß-stranded VDAC. Previously, a variant of a VDAC from Neurospora crassa, VDAC-ΔC, lacking the predicted 19th ß-strand, was found to form gated, anion-selective channels in artificial membranes. In vivo, the two C-terminal ß-strands (ß18 and ß19) in VDAC form a ß-hairpin necessary for import from the cytoplasm into mitochondria and the ß-signal required for assembly in the mitochondrial outer membrane resides in ß19. The current study demonstrated that the putative 18-stranded ß-barrel formed by VDAC-ΔC can be imported and assembled in the MOM in vivo and can also partially rescue the phenotype associated with the deletion of VDAC from a strain of N. crassa. Furthermore, when expressed and purified from Escherichia coli, VDAC-ΔC can be folded into a ß-strand-rich form in decyl-maltoside. Size exclusion chromatography (SEC) alone or combined with multi-angle light scattering (SEC-MALS) and analytical ultracentrifugation revealed that, unlike full-length VDACs, VDAC-ΔC can self-organize into dimers and higher order oligomers in the absence of sterol.

"What if I'm not trans enough? What if I'm not man enough?": Transgender young adults' experiences of gender-affirming healthcare readiness assessments in Aotearoa New Zealand.

Background: A growing body of transgender health research reports that transgender people often feel pressure to conform to a dominant narrative during gender-affirming readiness assessments. In New Zealand, however, no study to date has specifically examined transgender people's experiences of readiness assessments for gender-affirming healthcare. Aims: This study aimed to explore the experiences of transgender young adults (aged 16-30) during gender-affirming readiness assessments in New Zealand. We also sought participants' views on improving transgender healthcare provision. Methods: Thirteen transgender young adults took part in individual interviews or focus groups. Participants were asked to describe how they felt about the questions asked during readiness assessments and how the readiness assessment process could be improved. We used thematic analysis to identify patterns of meaning across the dataset. Results: We identified two themes. Firstly, proving gender explores participants' views of readiness assessments as designed to establish whether they were "trans enough" or "truly" transgender, and why readiness assessments are conducted in this manner. Secondly, the trans narrative describes the pressure participants felt to adhere to a dominant transgender narrative in order to gain access to the healthcare they needed. Discussion: Our findings call attention to the importance of a trans-affirmative approach and the need to clarify the purpose of gender-affirming healthcare readiness assessments in New Zealand.

First-line therapy, autologous stem-cell transplantation, and post-transplantation maintenance in the management of newly diagnosed mantle cell lymphoma.

Background: In Ontario, no clearly defined standard of care for the management of mantle cell lymphoma (mcl) has been developed, and substantial variability from centre to centre is evident. This guidance document was prompted by the need to harmonize practice in Ontario with respect to first-line, conditioning, and post-transplantation maintenance therapy for patients newly diagnosed with transplantation-eligible mcl. Methods: The medline and embase databases were systematically searched from January 2013 to January 2020 for evidence, and the best available evidence was used to draft recommendations relevant to first-line therapy, autologous stem-cell transplantation, and post-transplantation maintenance in the management of transplantation-eligible newly diagnosed mcl. Final approval of this guidance document was obtained from the Stem Cell Transplant Advisory Committee. Recommendations: These recommendations apply to all cases of transplantation-eligible newly diagnosed mcl:■ Alternating cycles of r-chop (rituximab plus cyclophosphamide-doxorubicin-vincristine-prednisolone) and r-dhap [rituximab plus dexamethasone-high-dose cytarabine-cisplatin] is the recommended first-line treatment for symptomatic patients newly diagnosed with mcl before autologous stem-cell transplantation (asct).■ Rituximab plus hyperfractionated cyclophosphamide-vincristine-doxorubicin-dexamethasone (r-hypercvad), alternating with methotrexate and cytarabine, is not recommended for the treatment of patients with newly diagnosed mcl.■ beam (carmustine-etoposide-cytarabine-melphalan), beac (carmustine-etoposide-cytarabine-cyclophosphamide), and total-body irradiation-based regimens are reasonable conditioning options for patients with mcl who have responded to first-line therapy and who are undergoing asct.■ Maintenance therapy with rituximab is recommended for patients with newly diagnosed mcl who have undergone asct.

Blocking of efflux transporters in rats improves translational validation of brain radioligands.

BACKGROUND: Positron emission tomography (PET) is a molecular imaging technique that can be used to investigate the in vivo pharmacology of drugs. Initial preclinical evaluation of PET tracers is often conducted in rodents due to the accessibility of disease models as well as economic considerations. Compared to larger species, rodents display a higher expression and/or activity of efflux transporters such as the P-glycoprotein (P-gp). Low brain uptake could, therefore, be species-specific and uptake in rodents not be predictive for that in humans. We hypothesized that a better prediction from rodent data could be achieved when a tracer is evaluated under P-gp inhibition. Consequently, we compared the performance of eight neuroreceptor tracers in rats with and without P-gp inhibition including a specific binding blockade. This data set was then used to predict the binding of these eight tracers in pigs. METHODS: PET tracers targeting serotonin 5-HT2A receptors ([18F]MH.MZ, [18F]Altanserin, [11C]Cimbi-36, [11C]Pimavanserin), serotonin 5-HT7 receptors ([11C]Cimbi-701, [11C]Cimbi-717 and [11C]BA-10) and dopamine D2/3 receptors ([18F]Fallypride) were used in the study. The brain uptake and target-specific binding of these PET radiotracers were evaluated in rats with and without inhibition of P-gp. Rat data were subsequently compared to the results obtained in pigs. RESULTS: Without P-gp inhibition, the amount of target-specific binding in the rat brain was sufficient to justify further translation for three out of eight evaluated tracers. With P-gp inhibition, results for five out of eight tracers justified further translation. The performance in pigs could correctly be predicted for six out of eight tracers when rat data obtained under P-gp inhibition were used, compared to four out of eight tracers without P-gp inhibition. CONCLUSIONS: P-gp strongly affects the uptake of PET tracers in rodents, but false prediction outcomes can be reduced by evaluating a tracer under P-gp inhibition.

A Diverse Specialty: What Students Teach Us About Neurology and "Neurophobia".

OBJECTIVE: To explore what elective students learn about the specialty of Neurology. METHODS: A prospective qualitative study using pre- and post-elective written questionnaires. RESULTS: Analysis concentrated on three main themes: What did students learn about the specialty of Neurology? What would they change about their experience? Did their opinions change? Major findings were (i) pre- and post-elective the most frequent response for "what is the best thing about Neurology?" was the "process of localization" and (ii) post-elective students were less likely to cite the challenge or problem-solving aspect of Neurology as the best thing while more emphasized the importance of the physical exam and the variety of cases. (iii) Students were most surprised by the scope of neurological practice. (iv) They would diversify the setting of their elective to include less time spent in the emergency room and more time in clinic. (v) The perception of Neurology as a specialty in which patients have a poor prognosis was the opinion that changed the most. CONCLUSIONS: Showcasing the diversity of cases and careers in Neurology may be a useful strategy to increase interest in the specialty and reduce neurophobia. Lectures or small groups early in medical school should concentrate on clear examples of common neurological conditions and emphasize the role of general neurologists and subspecialists involved in patient care. Whenever possible students should rotate through different clinics and not concentrate exclusively on emergency room and in-patient cases.

Rhenium Complexes of Pyridyl-Mesoionic Carbenes: Photochemical Properties and Electrocatalytic CO2 Reduction.

Mesoionic carbenes have found wide use as components of homogeneous catalysts. Recent discoveries have, however, shown that metal complexes of such ligands also have huge potential in photochemical research and in the activation of small molecules. We present here three ReI complexes with mesoionic pyridyl-carbene ligands. The complexes display reduction steps which were investigated via UV-vis-NIR-IR spectro-electrochemistry, and these results point toward an EC mechanism. The ReI compounds emit in the visible range in solution at room temperature with excited state lifetimes that are dependent on the substituents of the mesoionic carbenes. These complexes are also potent electrocatalysts for the selective reduction of CO2 to CO. Whereas the substituents on the carbenes have no influence on the reduction potentials, the electrocatalytic efficiency is strongly dependent on the substituents. This fact is likely a result of catalyst instability. The results presented here thus introduce mesoionic carbenes as new potent ligands for the generation of emissive ReI complexes and for electrocatalytic CO2 reduction.

Solution structure and oligomeric state of the E. coliglycerol facilitator.

Protein dynamics at atomic resolution can provide deep insights into the biological activities of proteins and enzymes but they can also make structure and dynamics studies challenging. Despite their well-known biological and pharmaceutical importance, integral membrane protein structure and dynamics studies lag behind those of water-soluble proteins mainly owing to solubility problems that result upon their removal from the membrane. Escherichia coli glycerol facilitator (GF) is a member of the aquaglyceroporin family that allows for the highly selective passive diffusion of its substrate glycerol across the inner membrane of the bacterium. Previous molecular dynamics simulations and hydrogen-deuterium exchange studies suggested that protein dynamics play an important role in the passage of glycerol through the protein pore. With the aim of studying GF dynamics by solution and solid-state nuclear magnetic resonance (NMR) spectroscopy we optimized the expression of isotope-labelled GF and explored various solubilizing agents including detergents, osmolytes, amphipols, random heteropolymers, lipid nanodiscs, bicelles and other buffer additives to optimize the solubility and polydispersity of the protein. The GF protein is most stable and soluble in lauryl maltose neopentyl glycol (LMNG), where it exists in a tetramer-octamer equilibrium. The solution structures of the GF tetramer and octamer were determined by negative-stain transmission electron microscopy (TEM), size-exclusion chromatography small-angle X-ray scattering (SEC-SAXS) and solid-state magic-angle spinning NMR spectroscopy. Although NMR sample preparation still needs optimization for full structure and dynamics studies, negative stain TEM and SEC-SAXS revealed low-resolution structures of the detergent-solubilized tetramer and octamer particles. The non-native octamer appears to form from the association of the cytoplasmic faces of two tetramers, the interaction apparently mediated by their disordered N- and C-termini. This information may be useful in future studies directed at reducing the heterogeneity and self-association of the protein.

Expression of CD1d by astrocytes corresponds with relative activity in multiple sclerosis lesions.

The CD1 protein family present lipid antigens to the immune system. CD1d has been observed in the CNS of MS patients, yet no studies have quantitatively characterized this expression and related it to inflammatory demyelinative activity in MS plaques. In this study, we set out to localize and quantify the presence of CD1d expression by astrocytes in MS brain tissue lesions. Formalin-fixed, paraffin-embedded MS and control brain tissues were examined. Lesions were classified as active, chronic active or chronic silent. Using immunofluorescence, the density of CD1d-positive cells was determined in active lesions, chronic active lesion edges and chronic active lesion centers. The percentage of CD1d-positive cells that were GFAP-positive was also determined in each of these regions. CD1d immunoreactivity was significantly increased in MS compared to control tissue, was significantly more prevalent in areas of active demyelination, and colocalized with GFAP-positive reactive astrocytes. Increases of CD1d immunoreactivity in the CNS of MS patients being greatest in areas of active demyelination and localized to GFAP-positive astrocytes lend support to the hypothesis of a lipid-targeted autoimmune process contributing to the pathogenesis of MS.

Radiosynthesis and preclinical evaluation of [11 C]Cimbi-701 - Towards the imaging of cerebral 5-HT7 receptors.

The serotonin 7 (5-HT7 ) receptor is suggested to be involved in a broad variety of CNS disorders, but very few in vivo tools exist to study this important target. Molecular imaging with positron emission tomography (PET) would enable an in vivo characterization of the 5-HT7 receptor. However, no clinical PET radiotracer exists for this receptor, and thus we aimed to develop such a tracer. In this study, we present the preclinical evaluation of [11 C]Cimbi-701. Cimbi-701 was synthesized in a one-step procedure starting from SB-269970. Its selectivity profile was determined using an academic screening platform (NIMH Psychoactive Drug Screening Program). Successful radiolabeling of [11 C]Cimbi-701 and subsequent in vivo evaluation was conducted in rats, pigs and baboon. In vivo specificity was investigated by 5-HT7 and σ receptor blocking studies. P-gp efflux transporter dependency was investigated using elacridar. [11 C]Cimbi-701 could successfully be synthesized. Selectivity profiling revealed high affinity for the 5-HT7 (Ki = 18 nM), σ-1 (Ki = 9.2 nM) and σ-2 (Ki = 1.6 nM) receptors. In rats, [11 C]Cimbi-701 acted as a strong P-gp substrate. After P-gp inhibition, rat brain uptake could specifically be blocked by 5-HT7 and σ receptor ligands. In pig, high brain uptake and specific 5-HT7 and σ-receptor binding was found for [11 C]Cimbi-701 without P-gp inhibition. Finally, low brain uptake was found in baboons. Both the specific σ-receptor binding and the low brain uptake of [11 C]Cimbi-701 displayed in baboon discouraged further translation to humans. Instead, we suggest exploration of this structural class as results indicate that selective 5-HT7 receptor imaging might be possible when more selective non-P-gp substrates are identified.

Towards selective CNS PET imaging of the 5-HT7 receptor system: Past, present and future.

Since its discovery in 1993, the serotonin receptor subtype 7 (5-HT7) has attracted significant attention as a potential drug target; due to its elucidated roles in conditions such as insomnia, schizophrenia, and more. Therefore, it is unsurprising that there has been relatively early efforts undertaken to develop a positron emission tomography (PET) imaging agent for said receptor system. PET can be clinically used to probe receptor systems in vivo, permitting information such as a drug's occupancy against this system to be investigated. This review focuses on the efforts towards the development of a 5-HT7R selective PET CNS tracer over the last 20 years, critically reflecting on applied strategies and commonly employed chemical frameworks and suggests future considerations that are needed to successfully develop a PET tracer for this clinically relevant target. This article is part of the special issue entitled 'Serotonin Research: Crossing Scales and Boundaries'.

Characterisation of the Selective Reduced Uteroplacental Perfusion (sRUPP) Model of Preeclampsia.

Preeclampsia is a complication of pregnancy characterised by gestational hypertension, proteinuria and/or end organ disease. The reduced uteroplacental perfusion (RUPP) model, via partial occlusion of the lower abdominal aorta, mimics insufficient placental perfusion as a primary causal characteristic of preeclampsia. However, a major limitation of the RUPP model is that perfusion is reduced to the entire hindquarters of the rat resulting in hindlimb ischemia. We hypothesised that clipping the uterine and ovarian arteries in the selective (s)RUPP model would provoke signs of preeclampsia while avoiding systemic ischemia. Sham, RUPP or sRUPP procedures were performed in pregnant Sprague Dawley rats on gestational day (GD)14. On GD21 uterine blood flow was significantly reduced in both the RUPP and sRUPP models while aortic flow was reduced only in RUPP. Both models resulted in increased MAP, increased vascular oxidative stress (superoxide generation), increased pro-inflammatory (RANTES) and reduced pro-angiogenic (endoglin) mediators. Vascular compliance and constriction were unaltered in either RUPP or sRUPP groups. In summary, refinements to the RUPP model simultaneously maintain the characteristic phenotype of preeclampsia and avoid peripheral ischemia; providing a useful tool which may be used to increase our knowledge and bring us closer to a solution for women affected by preeclampsia.

Studying Conformational Changes of the Yersinia Type-III-Secretion Effector YopO in Solution by Integrative Structural Biology.

The type-III secretion effector YopO helps pathogenic Yersinia to outmaneuver the human immune system. Injected into host cells, it functions as a Ser/Thr kinase after activation by actin binding. This activation process is thought to involve large conformational changes. We use PELDOR spectroscopy and small-angle X-ray scattering in combination with available crystal structures to study these conformational transitions. Low-resolution hybrid models of the YopO/actin structure in solution were constructed, where the kinase domain of YopO is tilted "backward" compared with the crystal structure, thus shortening the distance between actin and the kinase active site, potentially affecting the substrate specificity of YopO. Furthermore, the GDI domain of the hybrid models resembles a conformation that was previously observed in a crystal structure of the isolated GDI domain. We investigate possible structural reasons for the inactivity of the apo state, analyze its flexibility and discuss the biological implications.

Development and Evaluation of Two Potential 5-HT7 Receptor PET Tracers: [18F]ENL09 and [18F]ENL10.

The latest addition to the serotonin (5-HT) receptor family is the 5-HT7 receptor (5-HT7R). This receptor has gained interest as a drug target due to its involvement in various disorders such as depression or schizophrenia. There is currently no clinically validated positron emission tomography (PET) tracer for the 5-HT7R available. But, the (arylpiperazinyl-butyl)oxindole scaffold provides a promising lead structure for this purpose. Here, we synthesized 12 (arylpiperazinyl-butyl)oxindole derivatives and in vitro affinity screening identified two structures with suitable affinity and selectivity to be radiolabeled and tested as 5-HT7R selective PET tracers. Next, the radiolabeled products [18F]ENL09 and [18F]ENL10 were evaluated as PET tracers in rats. Both tracers were found to be P-gp substrates, but after P-gp inhibition the brain uptake showed a regional distribution in line with the known 5-HT7R distribution.  The [18F]ENL10 brain binding was displaceable with a 5-HT7R selective ligand, whereas [18F]ENL09 was not. We find that [18F]ENL10 is a promising 5-HT7R selective PET tracer candidate that should be investigated in higher species.