Brain-penetrant Compounds Research Articles

Identification and Characterization of a Blood-Brain Barrier Penetrant Inositol Hexakisphosphate Kinase (IP6K) Inhibitor.

Inositol hexakisphosphate kinases (IP6Ks) have been studied for their role in glucose homeostasis, metabolic disease, fatty liver disease, chronic kidney disease, neurological development, and psychiatric disease. IP6Ks phosphorylate inositol hexakisphosphate (IP6) to the pyrophosphate, 5-diphosphoinositol-1,2,3,4,6-pentakisphosphate (5-IP7). Most of the currently known potent IP6K inhibitors contain a critical carboxylic acid which limits blood-brain barrier (BBB) penetration. In this work, the synthesis and testing of a variety of carboxylic acid isosteres resulted in several new compounds with improved BBB penetration. The most promising compound has an IP6K1 IC50 of 16 nM with an improved brain/plasma ratio and a favorable pharmacokinetic profile. This series of brain penetrant compounds may be used to investigate the role of IP6Ks in CNS disorders.

Anti-prion drugs do not improve survival in novel knock-in models of inherited prion disease.

Prion diseases uniquely manifest in three distinct forms: inherited, sporadic, and infectious. Wild-type prions are responsible for the sporadic and infectious versions, while mutant prions cause inherited variants like fatal familial insomnia (FFI) and familial Creutzfeldt-Jakob disease (fCJD). Although some drugs can prolong prion incubation times up to four-fold in rodent models of infectious prion diseases, no effective treatments for FFI and fCJD have been found. In this study, we evaluated the efficacy of various anti-prion drugs on newly-developed knock-in mouse models for FFI and fCJD. These models express bank vole prion protein (PrP) with the pathogenic D178N and E200K mutations. We applied various drug regimens known to be highly effective against wild-type prions in vivo as well as a brain-penetrant compound that inhibits mutant PrPSc propagation in vitro. None of the regimens tested (Anle138b, IND24, Anle138b + IND24, cellulose ether, and PSCMA) significantly extended disease-free survival or prevented mutant PrPSc accumulation in either knock-in mouse model, despite their ability to induce strain adaptation of mutant prions. Our results show that anti-prion drugs originally developed to treat infectious prion diseases do not necessarily work for inherited prion diseases, and that the recombinant sPMCA is not a reliable platform for identifying compounds that target mutant prions. This work underscores the need to develop therapies and validate screening assays specifically for mutant prions, as well as anti-prion strategies that are not strain-dependent.

Utilizing a Dual Human Transporter MDCKII-MDR1-BCRP Cell Line to Assess Efflux at the Blood Brain Barrier.

To facilitate the design of drugs readily able to cross the blood brain barrier (BBB), a Madin-Darby canine kidney (MDCK) cell line was established that over expresses both P-glycoprotein (Pgp) and breast cancer resistance protein (BCRP), the main human efflux transporters of the BBB. Proteomics analyses indicate BCRP is expressed at a higher level than Pgp in this cell line. This cell line shows good activity for both transporters [BCRP substrate dantrolene efflux ratio (ER) 16.3 ± 0.9, Pgp substrate quinidine ER 27.5 ± 1.2], and use of selective transporter inhibitors enables an assessment of the relative contributions to overall ERs. The MDCKII-MDR1-BCRP ER negatively correlates with rat unbound brain/unbound plasma ratio, Kpuu Highly brain penetrant compounds with rat Kpuu ≥ 0.3 show ERs ≤ 2 in the MDCKII-MDR1-BCRP assay while compounds predominantly excluded from the brain, Kpuu ≤ 0.05, demonstrate ERs ≥ 20. A subset of compounds with MDCKII-MDR1-BCRP ER < 2 and rat Kpuu < 0.3 were shown to be substrates of rat Pgp using a rat transfected cell line, MDCKII-rMdr1a. These compounds also showed ERs > 2 in the human National Institutes of Health (NIH) MDCKI-MDR1 (high Pgp expression) cell line, which suggests that they are weak human Pgp substrates. Characterization of 37 drugs targeting the central nervous system in the MDCKII-MDR1-BCRP efflux assay show 36 have ERs < 2. In drug discovery, use of the MDCKII-MDR1-BCRP in parallel with the NIH MDCKI-MDR1 cell line is useful for identification of compounds with high brain penetration. SIGNIFICANCE STATEMENT: A single cell line that includes both the major human efflux transporters of the blood brain barrier (MDCKII-MDR1-BCRP) has been established facilitating the rapid identification of efflux substrates and enabling the design of brain penetrant molecules. Efflux ratios using this cell line demonstrate a clear relationship with brain penetration as defined by rat brain Kpuu.

TMIC-33. UNDERSTAND AND TARGET ASTROCYTE REACTIVITY IN BRAIN TUMORS TO PREVENT RADIORESISTANCE

Abstract Glioblastoma multiforme (GBM) is the most aggressive type of glioma and deadliest brain tumor. Despite aggressive treatments including surgery, chemo-, and radiotherapy, tumors invariably recur as incurable lesions, and the median survival is approximately 15 months after diagnosis. Recurrence is tightly linked to tumor cell resistance to ionizing radiation (IR), a feature that in turn is linked to stem cell characteristics of tumor cells. Previous results from our lab indicate that stromal astrocytes respond to IR with a reactive phenotype that in turn promotes therapeutic resistance of neighboring tumor cells. Our objectives were to i) identify pathways involved in IR-induced astrocyte reactivity; ii) identify compounds able to inhibit IR-induced astrocyte reactivity. We used two complementary approaches to meet our objectives. Using an antibody array we found that JNK and ABL pathways were activated following IR in primary human astrocytes. To validate their involvement in the induction of astrocyte reactive phenotype we used an image-based readout combining the changes in the expression of reactivity markers and in the morphology of astrocytes. For the second approach, we used a similar readout to perform an image-based drug screen which aimed to identify compounds able to inhibit IR-induced astrocyte reactivity. The drug libraries used consisted of approved drugs to get candidates with potential for drug repurposing. We identified 20 compounds inhibiting IR-induced astrocyte reactivity and amongst them, 10 are brain-penetrant compounds that could make them good candidates for repurposing in GBM. An in vivo test is now ongoing with 2 of these compounds in a PDGFA/PDGFB-driven mouse model of GBM to validate their efficiency to increase radiosensitivity of the tumor cell by inhibiting astrocyte reactivity

Transformation of a Dopamine D2 Receptor Agonist to Partial Agonists as Novel Antipsychotic Agents.

Designed ligands of G protein-coupled receptors can exert a spectrum of modulating effects, varying from full agonists and partial agonists to antagonists and inverse agonists. For the dopamine D2 receptor (D2R), partial agonist activity is the pharmacological feature of the third-generation antipsychotics, including aripiprazole, brexpiprazole, and cariprazine. Started from a benzofuran-derived D2R full agonist O4LE6 (4), which was identified using a structure-based method by us in previous studies, a series of D2R partial agonists were designed and synthesized by introducing different tail groups. Among them, compound 10b showed excellent activity in D2R pharmacological assays. Further optimizations using a structural rigidification approach led to the discovery of brain-penetrant compounds 29c and 29d, which exhibited potent antipsychotic effects in the mouse hyperlocomotion model. Compound 29c also showed excellent drug-like pharmacokinetic properties in rats and qualifies as an antipsychotic agent that is worth further evaluations.

Discovery of azaspirocyclic 1H-3,4,5-Trisubstitued pyrazoles as novel G2019S-LRRK2 selective kinase inhibitors

Mutations in the Leucine Rich Repeat Protein Kinase 2 gene (LRRK2) are genetic predispositions for Parkinson's Disease, of which the G2019S (GS) missense mutation is the most common. GS-LRRK2 has a hyperactive kinase, and although numerous drug discovery programs have targeted the LRRK2 kinase, few have reached clinical trials. We recently reported on the discovery of a novel LRRK2 kinase inhibitor chemotype, 1H-pyrazole biaryl sulfonamides. Although both potent and selective GS-LRRK2 inhibitors, 1H-pyrazole biaryl sulfonamides are incapable of crossing the blood-brain barrier. Retaining the core 1H-pyrazole and focusing our efforts on a phenylsulfonamide bioisosteric replacement, we report the discovery and preliminary development of azaspirocyclic 1H-3,4,5-trisubstituted pyrazoles as potent and selective (>2000-fold) GS-LRRK2 kinase inhibitors capable of entering rodent brain. The compounds disclosed here present an excellent starting point for the development of more brain penetrant compounds.

Abstract 6369: Discovery of potent, orally bioavailable, brain penetrant RAD51 inhibitor as an anti-cancer agent

Abstract Proliferating cells undergo DNA damage, and this is more pronounced in cancer cells due to their high rate of proliferation. Cancer cells are thus more dependent on the DNA damage repair pathway for their survival. Double-strand breaks (DSBs) are amongst the most severe type of DNA damage and are predominantly repaired by homologous recombination (HR) in an error-free manner. RAD51 is a pivotal recombinase for DSB repair by the HR pathway. Binding of RAD51 with BRCA2 followed by its nuclear translocation, is one of the key repair mechanisms for RAD51 mediated DSB repair. We have identified novel and potent RAD51 inhibitors disrupting the RAD51:BRCA2 interaction, which can address BRCA2 WT patient population. Upon induction of exogenous DNA damage, our compounds inhibit nuclear RAD51 foci formation and demonstrate sustained γH2AX accumulation in the nucleus, suggesting persistent DNA damage. Our orally bioavailable &amp; brain penetrant lead compound shows anti-proliferative activity in multiple BRCA2 WT cancer cell lines across various indications, and excellent synergy when combined with PARP1 inhibitors such as Olaparib. Further, in a wound healing assay, treatment with our lead compound inhibits cell migration in a dose-dependent manner. TNBC patients with extensive brain metastasis have limited treatment options. Thus, a brain penetrant RAD51 inhibitor could provide a novel treatment option in this setting. Citation Format: Sanjita Sasmal, Sukanya Patra, Venkatesham Boorgu, Shankar Chithaluri, Mahesh Yanamadra, Deepika Hiremath, Hemashankar Pathange, Sandeep M. Girme, Amitkumar Pawar, Rutuja Narayan Shinde, Nilanjan Samanta. Discovery of potent, orally bioavailable, brain penetrant RAD51 inhibitor as an anti-cancer agent [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6369.

Leveraging Structure-Based Drug Design to Identify Next-Generation MAT2A Inhibitors, Including Brain-Penetrant and Peripherally Efficacious Leads.

Inhibition of the S-adenosyl methionine (SAM)-producing metabolic enzyme, methionine adenosyltransferase 2A (MAT2A), has received significant interest in the field of medicinal chemistry due to its implication as a synthetic lethal target in cancers with the deletion of the methylthioadenosine phosphorylase (MTAP) gene. Here, we report the identification of novel MAT2A inhibitors with distinct in vivo properties that may enhance their utility in treating patients. Following a high-throughput screening, we successfully applied the structure-based design lessons from our first-in-class MAT2A inhibitor, AG-270, to rapidly redesign and optimize our initial hit into two new lead compounds: a brain-penetrant compound, AGI-41998, and a potent, but limited brain-penetrant compound, AGI-43192. We hope that the identification and first disclosure of brain-penetrant MAT2A inhibitors will create new opportunities to explore the potential therapeutic effects of SAM modulation in the central nervous system (CNS).

Identification of an Orally Bioavailable, Brain-Penetrant Compound with Selectivity for the Cannabinoid Type 2 Receptor.

Modulation of the endocannabinoid system (ECS) is of great interest for its therapeutic relevance in several pathophysiological processes. The CB2 subtype is largely localized to immune effectors, including microglia within the central nervous system, where it promotes anti-inflammation. Recently, a rational drug design toward precise modulation of the CB2 active site revealed the novelty of Pyrrolo[2,1-c][1,4]benzodiazepines tricyclic chemotype with a high conformational similarity in comparison to the existing leads. These compounds are structurally unique, confirming their chemotype novelty. In our continuing search for new chemotypes as selective CB2 regulatory molecules, following SAR approaches, a total of 17 selected (S,E)-11-[2-(arylmethylene)hydrazono]-PBD analogs were synthesized and tested for their ability to bind to the CB1 and CB2 receptor orthosteric sites. A competitive [3H]CP-55,940 binding screen revealed five compounds that exhibited >60% displacement at 10 μM concentration. Further concentration-response analysis revealed two compounds, 4k and 4q, as potent and selective CB2 ligands with sub-micromolar activities (Ki = 146 nM and 137 nM, respectively). In order to support the potential efficacy and safety of the analogs, the oral and intravenous pharmacokinetic properties of compound 4k were sought. Compound 4k was orally bioavailable, reaching maximum brain concentrations of 602 ± 162 ng/g (p.o.) with an elimination half-life of 22.9 ± 3.73 h. Whether administered via the oral or intravenous route, the elimination half-lives ranged between 9.3 and 16.7 h in the liver and kidneys. These compounds represent novel chemotypes, which can be further optimized for improved affinity and selectivity toward the CB2 receptor.

Scaffold Hopping to Imidazo[1,2-a]pyrazin-8-one Positive Allosteric Modulators of Metabotropic Glutamate 2 Receptor.

Glutamate hyperfunction is implicated in multiple neurological and psychiatric diseases. Activation of the mGlu2 receptor results in reduced glutamate release and decreased excitability representing a promising novel therapeutic agent for the treatment of disorders such as epilepsy, schizophrenia, mood, anxiety, and other neuropsychiatric disorders. We have previously reported substantial efforts leading to potent and selective mGlu2 PAMs from different chemical series. Herein, the discovery and optimization of a novel series of imidazopyrazinone mGlu2 PAMs are reported. This new scaffold originated from computational searching of fragment databases and comparison with our previously explored scaffolds. Optimization guided by our robust understanding of SAR from former series led to potent, selective, and brain-penetrant compounds.

208-LB: Peripheral CB1 Inverse Agonism Improves Metabolism in DIO Mice Independent of Hepatic FGF21

Obesity is associated with an overactive endocannabinoid system, and selective blockade of CB1R in peripheral tissues, including the liver, reverses HFD-induced metabolic abnormalities by restoring normal lipid and glucose homeostasis and avoid unwanted adverse neuropsychiatric effects. The CB1R inverse agonist JD5037 has been shown equal effective with its brain-penetrant compound in reducing appetite, body weight, hepatic steatosis, and insulin resistance. Fibroblast growth factor-21 (FGF21) has emerged as a major endocrine regulator derived from the liver that reduces adiposity and hepatic steatosis and improves glucose tolerance and insulin sensitivity, with changes similar to those induced by CB1R blockade. Here we investigated whether FGF21 mediate the metabolic effects of CB1R blockade by JD5037 in DIO mice. In C57BL/6J wild-type mice, HFD caused a robust increase in hepatic Fgf21 mRNA and serum FGF21 levels, which were reversed by chronic peripheral CB1R antagonist JD5037 treatment to levels observed in STD or vehicle-treated hepatocyte-specific CB1R-/- (hCB1-/-) mice, indicating activation of CB1R in the liver is largely involved in HFD-induced “FGF21-resistant” state. In contrast, the expression of the FGF21 receptor Fgfr1 and co-receptor β-klotho (Klb) were dramatically reduced by HFD in both epididymal fat and brain tissue in wild-type mice, and these effects were reversed by JD5037 treatment. To address whether FGF21 mediated the metabolic effects of CB1R blockade, we repeated JD5037 treatment in liver-specific FGF21-/- (hFGF21-/-) mice. Surprisingly, JD5037 treatment was almost equally effective in both HFD-fed wild-type and in hFGF21-/- mice in reducing body weight and hepatic steatosis, attenuating hyperinsulinemia and hyperleptinemia. The current data suggest that peripheral CB1R blockade by inverse agonist in obese mice improves insulin sensitivity and energy expenditure independently of hepatic FGF21. Disclosure J. Liu: None. Funding National Institutes of Health

Full-length in meso structure and mechanism of rat kynurenine 3-monooxygenase inhibition

The structural mechanisms of single-pass transmembrane enzymes remain elusive. Kynurenine 3-monooxygenase (KMO) is a mitochondrial protein involved in the eukaryotic tryptophan catabolic pathway and is linked to various diseases. Here, we report the mammalian full-length structure of KMO in its membrane-embedded form, complexed with compound 3 (identified internally) and compound 4 (identified via DNA-encoded chemical library screening) at 3.0 Å resolution. Despite predictions suggesting that KMO has two transmembrane domains, we show that KMO is actually a single-pass transmembrane protein, with the other transmembrane domain lying laterally along the membrane, where it forms part of the ligand-binding pocket. Further exploration of compound 3 led to identification of the brain-penetrant compound, 5. We show that KMO is dimeric, and that mutations at the dimeric interface abolish its activity. These results will provide insight for the drug discovery of additional blood-brain-barrier molecules, and help illuminate the complex biology behind single-pass transmembrane enzymes.

Calculation of an Apical Efflux Ratio from P-Glycoprotein (P-gp) In Vitro Transport Experiments Shows an Improved Correlation with In Vivo Cerebrospinal Fluid Measurements in Rats: Impact on P-gp Screening and Compound Optimization.

P-glycoprotein (P-gp) is a major blood-brain barrier (BBB) efflux transporter. In vitro approaches, including bidirectional efflux ratio (ER), are used to measure P-gp-mediated transport, but findings can be inconsistent across models. We propose a novel, more physiologically relevant, in vitro model: unidirectional apical efflux ratio (AP-ER)-a ratio of permeability rates at the apical side of the BBB with and without P-gp inhibitor. To test our approach, ER and AP-ER were calculated for 3227 structurally diverse compounds in porcine kidney epithelial cells (LLC-PK1) overexpressing human or mouse P-gp and classified based on their passive transcellular P-gp permeability or charged properties. In vivo rat infusion studies were performed for selected compounds with high ER but low AP-ER. One-third of the 3227 compounds had bidirectional ER that was much higher than AP-ER; very few had AP-ER higher than ER. Compounds with a large difference between AP-ER and ER were typically basic compounds with low-to-medium passive permeability and high lipophilicity and/or amphiphilicity, leading to strong membrane binding. Outcomes in the human model were similar to those in mice, suggesting AP-ER/ER ratios may be conserved for at least two species. AP-ER predicted measured cerebrospinal fluid (CSF) concentration better than ER for the five compounds tested in our in vivo rat infusion studies. We report superior estimations of the CSF concentrations of the compounds when based on less resource-intensive AP-ER versus classic ER. Better understanding of the properties leading to high P-gp-mediated efflux in vivo could support more efficient brain-penetrant compound screening and optimization. SIGNIFICANCE STATEMENT: To address inconsistencies associated with the historical, bidirectional efflux ratio (ER) calculation of P-glycoprotein-mediated transport, we propose to use the novel, more physiologically relevant, unidirectional apical efflux ratio (AP-ER) model. In vitro experiments suggested that compounds with strong membrane binding showed the largest difference between AP-ER and ER, and in vivo infusion studies showed that AP-ER predicted cerebrospinal fluid concentrations of compounds better than ER; outcomes in the human model were similar to those in mice.

Evaluating the Pharmacokinetics and Systemic Effects of a Permeability Enhancer Sodium N-[8-(2-hydroxybenzoyl)amino] Caprylate in Rats

Oral administration is the preferred route for drug delivery and its success is highly dependent on a compound's ADME properties, of which, permeability plays a major role. Therefore, permeability enhancers are an attractive area of research in the pharmaceutical industry. Recent data suggest that sodium N-[8-(2-hydroxybenzoyl) amino] caprylate (SNAC) is an effective permeability enhancer, yet the pharmacokinetic (PK) and systemic effects of SNAC are poorly understood, specifically its oral bioavailability and systemic effects on distribution, which could influence the safety of certain drugs. To answer these questions, both in vitro and in vivo studies were conducted. Of 3 permeability enhancers (SNAC, 4-CNAB, and 5-CNAC), SNAC was found to have the greatest effect on the absorption of cyanocobalamin in rats. It was also found that SNAC is orally bioavailable (almost 40%) when dosed to rats. Based on these findings, tool compounds were co-dosed in rats to further evaluate the systemic effects of SNAC. Oral co-dosing of SNAC with an intravenous infusion of 2 poorly brain penetrant compounds, quinidine, and gabapentin, did not increase brain ISF: plasma ratio or total brain:plasma ratio for either of these compounds, implying that SNAC is effective only in the intestine at pharmacologically relevant concentrations.

PDTM-02. TARGETING THE RB PATHWAY IN MEDULLOBLASTOMA

Abstract Medulloblastoma (MB), the most common malignant pediatric brain tumor, is classified into four major molecularly and histopathologically distinct subgroups, among which MYC-driven Group 3 MBs confer a poor prognosis. The Cyclin D/CDK4/CDK6/RB pathway is frequently deregulated in MB leading to uncontrolled cell proliferation, but tumors express an intact RB protein (Northcott et al., Nature, 2017). Therefore, CDK4/6 inhibitor drugs offer a possible therapeutic approach to treat MBs. Because single agent therapy ultimately leads to drug resistance, we initiated in vitro combination drug screens to identify drug classes potentiating CDK4/6 inhibitors. We used Group 3 MB patient-derived orthotopic xenografts (PDOXs), a human cell line (HDMB03), and freshly dissociated tumor cells propagated only in the mouse brain. The drug screen included 90 compounds comprising targeted and cytotoxic drugs that are FDA approved or under active clinical investigation. Using a bioluminescence-based assay that measures ATP consumption (CellTiter-Glo) to evaluate the number of viable cells, these 90 compounds were screened in combination with a fixed concentration of ribociclib, one of the three FDA approved CDK4/6 inhibitors. The primary screen, carried out in HDMB03 cells, revealed several drugs with additive or synergistic potential when combined with ribociclib, including BET inhibitors, MEK inhibitors, PI3K/mTOR inhibitors and gemcitabine. We are currently evaluating the combination of brain penetrant compounds in Group 3 MB PDOXs. The identification of potent drug combinations should provide new therapeutic options for the treatment of Group 3 MB, one of the most difficult to treat.

Brain-penetrant PQR620 mTOR and PQR530 PI3K/mTOR inhibitor reduce huntingtin levels in cell models of HD

One of the pathological hallmarks of Huntington disease (HD) is accumulation of the disease-causing mutant huntingtin (mHTT), which leads to the disruption of a variety of cellular functions, ultimately resulting in cell death. Induction of autophagy, for example by the inhibition of mechanistic target of rapamycin (mTOR) signaling, has been shown to reduce HTT levels and aggregates. While rapalogs like rapamycin allosterically inhibit the mTOR complex 1 (TORC1), ATP-competitive mTOR inhibitors suppress activities of TORC1 and TORC2 and have been shown to be more efficient in inducing autophagy and reducing protein levels and aggregates than rapalogs. The ability to cross the blood-brain barrier of first generation catalytic mTOR inhibitors has so far been limited, and therefore sufficient target coverage in the brain could not be reached. Two novel, brain penetrant compounds – the mTORC1/2 inhibitor PQR620, and the dual pan-phosphoinositide 3-kinase (PI3K) and mTORC1/2 kinase inhibitor PQR530 - were evaluated by assessing their potential to induce autophagy and reducing mHTT levels. For this purpose, expression levels of autophagic markers and well-defined mTOR targets were analyzed in STHdh cells and HEK293T cells and in mouse brains. Both compounds potently inhibited mTOR signaling in cell models as well as in mouse brain. As proof of principle, reduction of aggregates and levels of soluble mHTT were demonstrated upon treatment with both compounds. Originally developed for cancer treatment, these second generation mTORC1/2 and PI3K/mTOR inhibitors show brain penetrance and efficacy in cell models of HD, making them candidate molecules for further investigations in HD.

Characterization of AJ5012 as a Novel Peripheral Cannabinoid 1 Receptor Antagonist in Mouse Models of Obesity

Obesity-induced adipose tissue inflammation, in which the NLRP3 inflammasome is a pivotal mediator, is implicated in the development of insulin resistance. Cannabinoid 1 receptor (CB1R) antagonists have been shown to improve insulin resistance and the associated metabolic abnormalities, but their therapeutic development was discontinued due to neuropsychiatric side effects. Although growing evidence suggests the role of CB1Rs in proinflammatory signaling, their direct effects on adipose tissue inflammation have not yet been evaluated. Here we report that the peripherally restricted CB1R antagonist AJ5012, which exhibits beneficial metabolic effects comparable with its brain-penetrant parent compound rimonabant, suppresses macrophage infiltration into white adipose tissue, activation of the NLRP3 inflammasome, and production of proinflammatory cytokines in diet-induced obese mice. Moreover, we identified the downstream signaling pathways by which CB1R regulates proinflammatory gene expression. These results suggest that peripheral CB1R blockade improves obesity-induced insulin resistance by suppressing adipose tissue inflammation via the NLRP3 inflammasome. Disclosure J. Kim: None. J. Yoon: None. E. Kim: None. S. Yoon: None. W. Kim: None.

Correction to Application of Structure-Based Design and Parallel Chemistry to Identify a Potent, Selective, and Brain Penetrant Phosphodiesterase 2A Inhibitor.

Phosphodiesterase 2A (PDE2A) inhibitors have been reported to demonstrate in vivo activity in preclinical models of cognition. To more fully explore the biology of PDE2A inhibition, we sought to identify potent PDE2A inhibitors with improved brain penetration as compared to current literature compounds. Applying estimated human dose calculations while simultaneously leveraging synthetically enabled chemistry and structure-based drug design has resulted in a highly potent, selective, brain penetrant compound 71 (PF-05085727) that effects in vivo biochemical changes commensurate with PDE2A inhibition along with behavioral and electrophysiological reversal of the effects of NMDA antagonists in rodents. This data supports the ability of PDE2A inhibitors to potentiate NMDA signaling and their further development for clinical cognition indications.

In Situ Tissue Labeling of Cerebral Amyloid Using HIV-Related Tat Peptide.

Delivering peptide-based drugs to the brain is a major challenge because of the existence of the blood-brain barrier (BBB). To overcome this problem, cell-penetrating peptides derived from proteins that are able to cross biological membranes have been used as cell-permeable and brain-penetrant compounds. An example is the transactivator of transcription protein transduction domain (Tat) of the human immunodeficiency virus. The basic domain of Tat is formed of arginine and lysine amino acid residues. Tat has been used as brain-penetrant carrier also in therapies for Alzheimer disease (AD), the most common form of dementia characterized by extracellular cerebral deposits of amyloid made up of Aβ peptide. The aim of our study was to assess whether Tat bind to amyloid deposits of AD and other amyloidoses. An in situ labeling using biotinylated Tat 48-57 peptide was employed in the brain tissue with amyloid deposits made up of Aβ (patients with AD and transgenic AD mice), of prion protein (patients with Gerstmann-Straussler-Scheinker disease), and other amyloidosis, processed by different fixations and pretreatments of histological sections. Our results showed that Tat peptide binds amyloid deposits made up of Aβ, PrP, and immunoglobulin lambda chains in the brain and other tissues processed by alcoholic fixatives but not in formalin-fixed tissue. The fact that biotinylated Tat peptide stains amyloid of different biochemical composition and the specific charge characteristics of the molecules suggests that Tat may bind to heparan sulfate glicosaminoglicans, that are present in amyloid deposits. Inhibition of the binding by Tat pre-incubation with protamine reinforces this hypothesis. Binding of Tat to amyloid deposits should be kept in mind in interpreting the results of studies employing this molecule as brain-penetrating compound for the treatment of cerebral amyloidoses. Our results also suggest that Tat may be helpful for the analysis of the mechanisms of amyloidogenesis, and in particular, the interactions between specific amyloid peptides and glicosaminoglicans.

Application of Structure-Based Design and Parallel Chemistry to Identify a Potent, Selective, and Brain Penetrant Phosphodiesterase 2A Inhibitor.

Phosphodiesterase 2A (PDE2A) inhibitors have been reported to demonstrate in vivo activity in preclinical models of cognition. To more fully explore the biology of PDE2A inhibition, we sought to identify potent PDE2A inhibitors with improved brain penetration as compared to current literature compounds. Applying estimated human dose calculations while simultaneously leveraging synthetically enabled chemistry and structure-based drug design has resulted in a highly potent, selective, brain penetrant compound 71 (PF-05085727) that effects in vivo biochemical changes commensurate with PDE2A inhibition along with behavioral and electrophysiological reversal of the effects of NMDA antagonists in rodents. This data supports the ability of PDE2A inhibitors to potentiate NMDA signaling and their further development for clinical cognition indications.