Quantitative Model Analysis and Simulation of Pharmacokinetics and RNA Knockdown Effect After Systemic Administration of Cholesterol-Conjugated DNA/RNA Heteroduplex Oligonucleotide Crossing Blood-Brain Barrier of Mice.

The cholesterol-conjugated heteroduplex oligonucleotide (Chol-HDO) is a double-stranded complex; it comprises an antisense oligonucleotide (ASO) and its complementary strand with a cholesterol ligand. Chol-HDO is a powerful tool for achieving target RNA knockdown in the brains of mice after systemic injection. Here, a quantitative model analysis was conducted to characterize the relationship between the pharmacokinetics (PK) and pharmacodynamics (PD), non-coding RNA metastasis-associated lung adenocarcinoma 1 () RNA, of Chol-HDO, in a time-dependent manner.

Utility of Göttingen minipigs for the prediction of human pharmacokinetic profiles after intravenous drug administration.

Göttingen minipigs are increasingly used to evaluate the pharmacokinetic (PK) profiles of drug candidates. However, their accuracy in predicting human PK parameters is unclear. In this study, we investigated the utility of Göttingen minipigs for predicting human PK profiles.

Development of a bioanalytical method for circulating human T cells in animals using -based quantitative polymerase chain reaction and its application in preclinical biodistribution studies.

Introduction: In the development of cell therapy products for human use, studies on the biodistribution of transplanted cells in animals are important for assessing the safety and efficacy of these products. Although a few reports have described the biodistribution of human cells in animals using -based-polymerase chain reaction (Alu-PCR), most have used genomic DNA or synthetic oligonucleotide as calibrators, as opposed to actual cells. In addition, bioanalytical variability in the quantification of cells with respect to specificity, selectivity, accuracy, and precision, has not been evaluated.

Highly specific, quantitative polymerase chain reaction probe for the quantification of human cells in cynomolgus monkeys.

Quantification of human cells may be performed using quantitative polymerase chain reaction (qPCR). In preclinical studies, the human Alu sequence is widely used as biomarker for human DNA. However, because the Alu gene is shared by primates, its use is limited to non-primate studies.

Quantitative PCR methodology with a volume-based unit for the sophisticated cellular kinetic evaluation of chimeric antigen receptor T cells.

Although the cellular kinetics of chimeric antigen receptor T (CAR T) cells are expressed in units of copies/μg gDNA, this notation carries the risk of misrepresentation owing to dramatic changes in blood gDNA levels after lymphocyte-depleting chemotherapy and rapid expansion of CAR T cells. Therefore, we aimed to establish a novel qPCR methodology incorporating a spike-in calibration curve that expresses cellular kinetics in units of copies/μL blood, as is the case for conventional pharmacokinetic studies of small molecules and other biologics. Dog gDNA was used as an external control gene.

Quantitative application of flow cytometry for the analysis of circulating human T cells: A preclinical pharmacokinetic study.

As the application of flow cytometry to a quantitative pharmacokinetic study with adoptive T cell therapy is new, we aimed to investigate the quantitativity of flow cytometry-based analysis for the pharmacokinetic assessment of circulating human T cells in a preclinical study. We evaluated the selectivity, linearity, accuracy, precision, and sensitivity of flow cytometry-based analysis for human CD8 T cells in immunodeficient mouse blood. The CD3/8/45-positive cell population was successfully distinguished from the negative population.

A new class of pentapeptide KISS1 receptor agonists with hypothalamic-pituitary-gonadal axis activation.

The kisspeptin (Kp, Kp-54, metastin)/KISS1R system plays crucial roles in regulating the secretion of gonadotropin-releasing hormone. Continuous administration of nonapeptide Kp analogs caused plasma testosterone depletion, whereas bolus administration caused strong plasma testosterone elevation in male rats. To develop a new class of small peptide drugs, we focused on stepwise N-terminal truncation of Kp analogs and discovered potent pentapeptide analogs.

A quantitative LC/MSMS method for determination of edoxaban, a Xa inhibitor and its pharmacokinetic application in patients after total knee arthroplasty.

Edoxaban was extracted from human plasma by simple protein precipitation with acetonitrile, followed by quantitative determination using a liquid chromatography-mass spectrometry method. The recoveries of edoxaban and the internal standard (ticlopidine) from human plasma were >85%, and the within- and between-day coefficients of variation were within 15%. The limit of quantification in human plasma was 1 ng/mL.

Design and synthesis of a novel series of orally active, selective somatostatin receptor 2 agonists for the treatment of type 2 diabetes.

The discovery of a novel series of β-methyltryptophan (β MeTrp) derivatives as selective and orally active non-peptide somatostatin receptor 2 (SSTR2) agonists for the treatment of Type 2 diabetes is described. In our previous research, Compound A, β-MeTrp derivative with highly potent and selective SSTR2 agonistic activity IC (SSTR2/SSTR5)=0.3/>100 (nM), was identified asa drug candidate for treatment of Type 2 diabetes which lowers significantly plasma glucose level in Wistar fatty rats in its oral administrations.

Identification of cytidine-5-triphosphate synthase1-selective inhibitory peptide from random peptide library displayed on T7 phage.

Cytidine triphosphate synthase 1 (CTPS1) is an enzyme expressed in activated lymphocytes that catalyzes the conversion of uridine triphosphate (UTP) to cytidine triphosphate (CTP) with ATP-dependent amination, using either L-glutamine or ammonia as the nitrogen source. Since CTP plays an important role in DNA/RNA synthesis, phospholipid synthesis, and protein sialyation, CTPS1-inhibition is expected to control lymphocyte proliferation and size expansion in inflammatory diseases. In contrast, CTPS2, an isozyme of CTPS1 possessing 74% amino acid sequence homology, is expressed in normal lymphocytes.

Identification of AHCY inhibitors using novel high-throughput mass spectrometry.

S-adenosylhomocysteine hydrolase (AHCY) catalyzes the reversible hydrolysis of S-adenosylhomocysteine (SAH) to adenosine and l-homocysteine. This enzyme is frequently overexpressed in many tumor types and is considered to be a validated anti-tumor target. In order to enable the development of small molecule AHCY inhibitors as targeted cancer therapeutics we developed an assay based on a RapidFire high-throughput mass spectrometry detection system, which allows the direct measurement of AHCY enzymatic activity.

Nanophase-separated NiNb as an automobile exhaust catalyst.

Catalytic remediation of automobile exhaust has relied on precious metals (PMs) including platinum (Pt). Herein, we report that an intermetallic phase of Ni and niobium (Nb) (, NiNb) exhibits a significantly higher activity than that of Pt for the remediation of the most toxic gas in exhaust (, nitrogen monoxide (NO)) in the presence of carbon monoxide (CO). When subjected to the exhaust-remediation atmosphere, NiNb spontaneously evolves into a catalytically active nanophase-separated structure consisting of filamentous Ni networks (thickness < 10 nm) that are incorporated in a niobium oxide matrix (, NbO ( < 5/2)).

Discovery and characterization of selective human sphingomyelin synthase 2 inhibitors.

Sphingomyelin synthase (SMS) is a membrane enzyme that catalyzes the synthesis of sphingomyelin, is required for the maintenance of plasma membrane microdomain fluidity, and has two isoforms: SMS1 and SMS2. Although these isoforms exhibit the same SMS activity, they are different enzymes with distinguishable subcellular localizations. It was reported that SMS2 KO mice displayed lower inflammatory responses and anti-atherosclerotic effects, suggesting that inhibition of SMS2 would be a potential therapeutic approach for controlling inflammatory responses and atherosclerosis.

High-Throughput Screening to Identify Inhibitors of DEAD Box Helicase DDX41.

The human DEAD (Asp-Glu-Ala-Asp) box protein DDX41, a member of the DEXDc helicase family, has nucleic acid-dependent ATPase and RNA and DNA translocase and unwinding activities. DDX41 is affected by somatic mutations in sporadic cases of myeloid neoplasms as well as in a biallelic fashion in 50% of patients with germline DDX41 mutations. The R525H mutation in DDX41 is thought to play important roles in the development of hereditary myelodysplastic syndrome and acute myelocytic leukemia.

Identification of PARP14 inhibitors using novel methods for detecting auto-ribosylation.

Poly(ADP-ribose) polymerases (PARPs) use nicotinamide adenine dinucleotide (NAD) as a co-substrate to transfer ADP-ribose when it releases nicotinamide as the metabolized product. Enzymes of the PARP family play key roles in detecting and repairing DNA, modifying chromatin, regulating transcription, controlling energy metabolism, and inducing cell death. PARP14, the original member of the PARP family, has been reported to be associated with the development of inflammatory diseases and various cancer types, making it a potential therapeutic target.

Discovery of GPX4 inhibitory peptides from random peptide T7 phage display and subsequent structural analysis.

The phospholipid hydroperoxidase glutathione peroxidase (GPX4) is an enzyme that reduces lipid hydroperoxides in lipid membranes. Recently, GPX4 has been investigated as a target molecule that induces iron-dependent cell death (ferroptosis) selectively in cancer cells that express mutant Ras. GPX4 inhibitors have the potential to become novel anti-cancer drugs.

Design and Synthesis of an Investigational Nonapeptide KISS1 Receptor (KISS1R) Agonist, Ac-d-Tyr-Hydroxyproline (Hyp)-Asn-Thr-Phe-azaGly-Leu-Arg(Me)-Trp-NH (TAK-448), with Highly Potent Testosterone-Suppressive Activity and Excellent Water Solubility.

Metastin/kisspeptin is an endogenous ligand of KISS1 Receptor (KISS1R). Metastin and KISS1R are suggested to play crucial roles in regulating the secretion of gonadotropin-releasing hormone (GnRH), and continuous administration of metastin derivatives attenuated the plasma testosterone levels in male rats. Our optimization studies of metastin derivatives led to the discovery of 1 (Ac-d-Tyr-d-Trp-Asn-Thr-Phe-azaGly-Leu-Arg(Me)-Trp-NH, TAK-683), which suppressed plasma testosterone in rats at lower doses than those of leuprolide.

Physicochemically and pharmacokinetically stable nonapeptide KISS1 receptor agonists with highly potent testosterone-suppressive activity.

Modifications of metastin(45-54) produced peptide analogues with higher metabolic stability than metastin(45-54). N-terminally truncated nonapeptide 4 ([D-Tyr46,D-Pya(4)47,azaGly51,Arg(Me)53]metastin(46-54)) is a representative compound with both potent agonistic activity and metabolic stability. Although 4 had more potent testosterone-suppressant activity than metastin, it possessed physicochemical instability at pH 7 and insufficient in vivo activity.

A fragment-based approach to identifying S-adenosyl-l-methionine -competitive inhibitors of catechol O-methyl transferase (COMT).

Catechol O-methyl transferase belongs to the diverse family of S-adenosyl-l-methionine transferases. It is a target involved in the treatment of Parkinson's disease. Here we present a fragment-based screening approach to discover noncatechol derived COMT inhibitors which bind at the SAM binding pocket.

Pharmacologic profiles of investigational kisspeptin/metastin analogues, TAK-448 and TAK-683, in adult male rats in comparison to the GnRH analogue leuprolide.

Kisspeptin/metastin, a hypothalamic peptide, plays a pivotal role in controlling gonadotropin-releasing hormone (GnRH) neurons, and we have shown that continuous subcutaneous administration of kisspeptin analogues suppresses plasma testosterone in male rats. This study examined pharmacologic profiles of investigational kisspeptin analogues, TAK-448 and TAK-683, in male rats. Both analogues showed high receptor-binding affinity and potent and full agonistic activity for rat KISS1R, which were comparable to natural peptide Kp-10.

Design, synthesis, and biological evaluation of novel investigational nonapeptide KISS1R agonists with testosterone-suppressive activity.

Metastin/kisspeptin is a 54 amino acid peptide ligand of the KISS1R receptor and is a critical regulator of GnRH secretion. The N-terminally truncated peptide, metastin(45-54), possesses a 10-fold higher receptor-binding affinity than full-length metastin and agonistic KISS1R activity but is rapidly inactivated in rodent plasma. We have developed a decapeptide analog [D-Tyr(45),D-Trp(47),azaGly(51),Arg(Me)(53)]metastin(45-54) with improved serum stability compared with metastin(45-54) but with decreased KISS1R agonistic activity.

Discovery of TAK-960: an orally available small molecule inhibitor of polo-like kinase 1 (PLK1).

Using structure-based drug design, we identified and optimized a novel series of pyrimidodiazepinone PLK1 inhibitors resulting in the selection of the development candidate TAK-960. TAK-960 is currently undergoing Phase I evaluation in adult patients with advanced solid malignancies.