Identification of highly selective SIK1/2 inhibitors that modulate innate immune activation and suppress intestinal inflammation.

The salt-inducible kinases (SIK) 1-3 are key regulators of pro- versus anti-inflammatory cytokine responses during innate immune activation. The lack of highly SIK-family or SIK isoform-selective inhibitors suitable for repeat, oral dosing has limited the study of the optimal SIK isoform selectivity profile for suppressing inflammation in vivo. To overcome this challenge, we devised a structure-based design strategy for developing potent SIK inhibitors that are highly selective against other kinases by engaging two differentiating features of the SIK catalytic site. This effort resulted in SIK1/2-selective probes that inhibit key intracellular proximal signaling events including reducing phosphorylation of the SIK substrate cAMP response element binding protein (CREB) regulated transcription coactivator 3 (CRTC3) as detected with an internally generated phospho-Ser329-CRTC3-specific antibody. These inhibitors also suppress production of pro-inflammatory cytokines while inducing anti-inflammatory interleukin-10 in activated human and murine myeloid cells and in mice following a lipopolysaccharide challenge. Oral dosing of these compounds ameliorates disease in a murine colitis model. These findings define an approach to generate highly selective SIK1/2 inhibitors and establish that targeting these isoforms may be a useful strategy to suppress pathological inflammation.

Toward better understanding and management of chemobrain: the potential utilities of the MemTrax memory test.

OBJECTIVE: To study the effects of chemotherapy on cognitive function in breast cancer patients, and to investigate the relationship of MemTrax test of memory and related functions to the FACT-Cog functional self-assessment for the evaluation and management of chemobrain. METHODS: In this prospective cohort study, clinical information of pathologically confirmed female breast cancer patients who decided to receive chemotherapy were collected in a questionnaire which was developed for this study and provided as a supplementary file. The FACT-Cog self-assessment and MemTrax test were administered before and after the chemotherapy treatments. Patients with chemobrain were identified using published criteria based on FACT-Cog scores, and MemTrax scores from chemobrain patients were analyzed. RESULTS: Fifty-six patients participated in this study, of which 41 participants completed 4 or more cycles of chemotherapy and were included in the final analyses here. Using the reported high end of minimal clinical differences (10.6 points) of FACT-Cog before and after chemotherapy, 18 patients suffered from chemobrain in this study. In these 18 chemobrain patients, no cognitive impairments were detected by MemTrax, which paradoxically demonstrated an improvement in the normal cognitive range. CONCLUSION: The cognitive impairment induced by chemotherapy in breast cancer patients is detectable by the FACT-Cog in a Chinese cohort but is not detected by the MemTrax memory test. The fact that the more objective MemTrax could not detect the impairment could alleviate patients' concerns which in turn would be beneficial for patients' mental health.

LncRNA DSCAM-AS1 acts as a sponge of miR-137 to enhance Tamoxifen resistance in breast cancer.

OBJECTIVE: To investigate the influence of long noncoding RNA (lncRNA) DSCAM-AS1 on the propagation and apoptosis of Tamoxifen-resistant (TR) breast cancer cells via regulation of mircoRNA (miR)-137 and epidermal growth factor receptor pathway substrate 8 (EPS8). METHODS: Data of GSE5840 downloaded from the Gene Expression Omnibus database were utilized to screen out aberrantly expressed lncRNA and messenger RNA in breast cancer tissue samples. The expressions of DSCAM-AS1, miR-137, and EPS8 were determined by quantitative real time polymerase chain reaction (qRT-PCR). Cell lines were screened by half maximal inhibitory concentration (IC 50 ). 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay and the flow cytometry assay were used to detect cell proliferation, apoptosis, and cell cycle. The relationship among DSCAM-AS1, miR-137, and EPS8 was studied by miRcode, TargetScan, and Pearson correlation coefficient. A xenograft mouse model experiment was performed to demonstrate the effect of DSCAM-AS1 and EPS8 on tumor growth in vivo. RESULTS: LncRNA DSCAM-AS1 and EPS8 were significantly upregulated, whereas miR-137 was downregulated in TR tissues. DSCAM-AS1 could promote the Tamoxifen resistance of breast cancer, and it was negatively correlated with miR-137, whereas positively correlated with the expression of EPS8 in TR breast cancer tissues. Furthermore, miR-137 could inhibit tumor development and arrest cell cycle at the G0/G1 phase by targeting the 3'-UTR of EPS8. DSCAM-AS1 targeted miR-137 and EPS8 to promote propagation of TR breast cancer cells and inhibit cell apoptosis. CONCLUSION: LncRNA DSCAM-AS1 acts as a competing endogenous RNA of miR-137 and regulates EPS8 to promote cell reproduction and suppresses cell apoptosis in TR breast cancer.

The discovery of potent selective NPY Y(2) antagonists.

A series of small molecules with a piperidinyl core were synthesized and tested for binding affinity (IC50) at human Neuropeptide Y Y2 receptor. Various amide related analogs (ureas, reversed amides, and sulfonamides) were evaluated. Several potent and selective NPY Y2 antagonists were identified.

Translational evaluation of JNJ-18038683, a 5-hydroxytryptamine type 7 receptor antagonist, on rapid eye movement sleep and in major depressive disorder.

In rodents 5-hydroxytryptamine type 7 (5-HT(7)) receptor blockade has been shown to be effective in models of depression and to increase the latency to rapid eye movement (REM) sleep and decrease REM duration. In the clinic, the REM sleep reduction observed with many antidepressants may serve as a biomarker. We report here the preclinical and clinical evaluation of a 5-HT(7) receptor antagonist, (3-(4-chlorophenyl)-1,4,5,6,7,8-hexahydro-1-(phenylmethyl)pyrazolo[3,4-d]azepine 2-hydroxy-1,2,3-propanetricarboxylate) (JNJ-18038683). In rodents, JNJ-18038683 increased the latency to REM sleep and decreased REM duration, and this effect was maintained after repeated administration for 7 days. The compound was effective in the mouse tail suspension test. JNJ-18038683 enhanced serotonin transmission, antidepressant-like behavior, and REM sleep suppression induced by citalopram in rodents. In healthy human volunteers JNJ-18038683 prolonged REM latency and reduced REM sleep duration, demonstrating that the effect of 5-HT(7) blockade on REM sleep translated from rodents to humans. Like in rats, JNJ-18038683 enhanced REM sleep suppression induced by citalopram in humans, although a drug-drug interaction could not be ruled out. In a double-blind, active, and placebo-controlled clinical trial in 225 patients suffering from major depressive disorder, neither treatment with pharmacologically active doses of JNJ-18038683 or escitalopram separated from placebo, indicating a failed study lacking assay sensitivity. Post hoc analyses using an enrichment window strategy, where all the efficacy data from sites with an implausible high placebo response [placebo group Montgomery-Åsberg Depression Rating Scale (MADRS) < = 12] and from sites with no placebo response (MADRS > = 28) are removed, there was a clinically meaningful difference between JNJ-18038683 and placebo. Further clinical studies are required to characterize the potential antidepressant efficacy of JNJ-18038683.

lncRNA TUG1 inhibits the cancer stem cell­like properties of temozolomide­resistant glioma cells by interacting with EZH2.

Temozolomide (TMZ) is currently one of the first­line drugs used for the treatment of high­grade gliomas. However, TMZ resistance results in unsatisfactory therapeutic effects in gliomas. Cancer stem cells (CSCs) have recently been determined to serve a pivotal regulatory role in tumor metastasis, recurrence and chemoresistance. In addition, numerous reports have shown that long non­coding RNAs (lncRNAs) exert an essential role in the occurrence and development of tumors, and can be used as biomarkers for tumor diagnosis and treatment. Among them, studies have revealed that taurine upregulated gene 1 (TUG1) exhibits an important regulatory effect on the malignant biological behavior of glioma cells. Moreover, it has been reported that enhancer of Zeste homolog 2 polycomb repressive complex subunit 2 (EZH2) promotes tumorigenesis, including in glioma. However, the underlying mechanism of the interaction of TUG1 and EZH2 with CSCs of glioma remains elusive, and thus requires further clarification. The present study aimed to explore the role of TUG1 and EZH2 in TMZ resistance in glioma. Cell Counting Kit­8, colony formation,sphere formation and Annexin V­FITC/PI assays were used to detect the proliferation, clone formation efficiency, stemness and apoptosis of TMZ­resistant glioma cells. Xenograft tumor assay was used to detect the effect of TUG1 on the tumorigenesis of TMZ­resistant glioma cells. The present findings demonstrated that TUG1 exhibited a low expression in glioma cells, while EZH2 expression was the opposite. Moreover, it was observed that A172/TMZ cells possessed higher CSCs­like properties compared with parent cells, and that TUG1 and EZH2 were abnormally expressed in A172/TMZ cells. Knockdown of TUG1 or overexpression of EZH2 promoted A172/TMZ cell proliferation and CSCs­like properties, as well as inhibited their apoptosis, thereby enhancing the TMZ resistance of A172/TMZ cells. Furthermore, it was found that TUG1 alleviated the TMZ resistance of A172/TMZ cells by inhibiting EZH2 expression. Of note, overexpression of TUG1 inhibited the tumorigenicity of A172/TMZ cells by downregulating EZH2 expression in vivo. Collectively, the present study demonstrated that TUG1 served an essential regulatory role in TMZ resistance of gliomas.

Discovery of a Gut-Restricted JAK Inhibitor for the Treatment of Inflammatory Bowel Disease.

To identify Janus kinase (JAK) inhibitors that selectively target gastrointestinal tissues with limited systemic exposures, a class of imidazopyrrolopyridines with a range of physical properties was prepared and evaluated. We identified compounds with low intrinsic permeability and determined a correlation between permeability and physicochemical properties, clogP and tPSA, for a subset of compounds. This low intrinsic permeability translated into compounds displaying high colonic exposure and low systemic exposure after oral dosing at 25 mg/kg in mouse. In a mouse PK/PD model, oral dosing of lead compound 2 demonstrated dose-dependent inhibition of pSTAT phosphorylation in colonic explants post-oral dose but low systemic exposure and no measurable systemic pharmacodynamic activity. We thus demonstrate the utility of JAK inhibitors with low intrinsic permeability as a feasible approach to develop gut-restricted, pharmacologically active molecules with a potential advantage over systemically available compounds that are limited by systemic on-target adverse events.

Preparation and biological evaluation of indole, benzimidazole, and thienopyrrole piperazine carboxamides: potent human histamine h(4) antagonists.

Three series of H(4) receptor ligands, derived from indoly-2-yl-(4-methyl-piperazin-1-yl)-methanones, have been synthesized and their structure-activity relationships evaluated for activity at the H(4) receptor in competitive binding and functional assays. In all cases, substitution of small lipophilic groups in the 4 and 5-positions led to increased activity in a [(3)H]histamine radiolabeled ligand competitive binding assay. In vitro metabolism and initial pharmacokinetic studies were performed on selected compounds leading to the identification of indole 8 and benzimidazole 40 as potent H(4) antagonists with the potential for further development. In addition, both 8 and 40 demonstrated efficacy in in vitro mast cell and eosinophil chemotaxis assays.

The first potent and selective non-imidazole human histamine H4 receptor antagonists.

Following the discovery of the human histamine H4 receptor, a high throughput screen of our corporate compound collection identified compound 6 as a potential lead. Investigation of the SAR resulted in the discovery of novel compounds 10e and 10l, which are the first potent and selective histamine H4 receptor antagonists to be described.

Reconsideration of 5-hydroxytryptamine (5-HT)(7) receptor distribution using [(3)H]5-carboxamidotryptamine and [(3)H]8-hydroxy-2-(di-n-propylamino)tetraline: analysis in brain of 5-HT(1A) knockout and 5-HT(1A/1B) double-knockout mice.

The characterization and anatomical distribution of 5-hydroxytryptamine (5-HT)(7) receptor binding sites in brain tissue has been hampered by the lack of a specific radioligand. In the present autoradiographic study, we took advantage of 5-HT(1A) knockout and 5-HT(1A/1B) double-knockout mice to revisit the pharmacological characterization and anatomical localization of 5-HT(7) binding sites in mouse brain using [(3)H]5-carboxamidotryptamine (5-CT) and [(3)H]8-hydroxy-2-(di-n-propylamino)tetraline (8-OH-DPAT). The distribution pattern of [(3)H]5-CT binding sites (2 nM) in the brain of mice lacking the 5-HT(1A/1B) receptor was scarce and confined to the septum, globus pallidus, thalamus, hypothalamus, amygdala, cortex, and substantia nigra. The low densities of [(3)H]5-CT binding sites detected in septum, thalamus, hypothalamus, amygdala, and cortex were displaced by 10 microM of the selective 5-HT(7) receptor antagonist (R)-3-(2-(2-(4-methylpiperidin-1-yl) ethyl)pyrrolidine-1-sulfonyl) phenol (SB-269970). The SB-269970-insensitive [(3)H]5-CT binding sites detected in globus pallidus and substantia nigra of 5-HT(1A/1B) knockout mice were displaced by N-[3-(2-dimethylamino)ethoxy-4-methoxy-phenyl]-2'-methyl-4'- (5-methyl-1,2,4-oxadiazol-3-yl)-(1,1'-biphenyl)-4-carboxamide hydrochloride (SB-216641) (1 microM), demonstrating the 5-HT(1D) nature of these binding sites. In contrast to the low densities of [(3)H]5-CT binding sites, high-to-moderate densities of [(3)H]8-OH-DPAT binding sites (10 nM) were found throughout the brain of 5-HT(1A) and 5-HT(1A/1B) knockout mice (olfactory system, septum, thalamus, hypothalamus, amygdala, CA3 field of the hippocampus, cortical mantle, and central gray). These [(3)H]8-OH-DPAT binding sites were displaced by 10 microM SB-269970, risperidone, and methiothepin but not by pindolol, N-tert-butyl-3-[4-(2-methoxyphenyl)piperazin-1-yl]-2-phenylpropanamide (WAY- 100135), or citalopram. We conclude that despite its high affinity for the 5-HT(7) receptor in tissue homogenates, [(3)H]5-CT is not a good tracer for measuring 5-HT(7) receptor binding sites autoradiographically. Also, the lower affinity ligand [(3)H]8-OH-DPAT is a much better tracer for autoradiographic studies at the 5-HT(7) receptor binding sites.

Novel non-peptidic neuropeptide Y Y2 receptor antagonists.

Through SAR studies of a piperidinylindoline cinnamide HTS lead, the first potent, non-peptide, low molecular weight selective Neuropeptide Y Y2 (NPY Y2) antagonists have been synthesized. The SAR studies around the piperidinyl, the indolinyl, and the cinnamyl moieties are discussed.

Non-imidazole heterocyclic histamine H3 receptor antagonists.

Continued exploration of the SAR around the lead imidazopyridine histamine H(3) antagonist 1 has led to the discovery of several related series of heterocyclic histamine H(3) antagonists. The synthesis and SAR of indolizine, indole and pyrazolopyridine based compounds are now described.