Brain translocator protein occupancy by ONO-2952 in healthy adults: A Phase 1 PET study using [11 C]PBR28.

ONO-2952, a novel antagonist of translocator protein 18 kDa (TSPO), binds with high affinity to TSPO in rat brain and human tumor cell line membrane preparations. This study used the TSPO-specific PET radioligand [11 C]PBR28 to confirm binding of ONO-2952 to brain TSPO in human subjects, and evaluate brain TSPO occupancy and its relationship with ONO-2952 plasma concentration. Sixteen healthy subjects received a single oral dose of 200, 60, 20, or 6 mg ONO-2952 (n = 4 per dose). Two PET scans with [11 C]PBR28 were conducted ≤7 days apart: at baseline and 24 h after ONO-2952 administration. [11 C]PBR28 regional distribution volume (VT ) was derived with kinetic modeling using the arterial input function and a two tissue compartment model. Nonspecific binding (VND ) was obtained on an individual basis for each subject using linear regression as the x-intercept of the Lassen plot. The binding potential relative to VND (BPND ) was derived as the difference between VT in the ROI (VT ROI) and VND , normalized to VND ; BPND = (VT ROI - VND )/VND . TSPO occupancy was calculated as the change in BPND (ΔBPND ) from individual's baseline scan to the on-medication scan to the baseline BPND value. TSPO occupancy by ONO-2952 was dose dependent between 20-200 mg, approaching saturation at 200 mg both in the whole brain and in 15 anatomic regions of interest (ROI). Estimated Ki values ranged from 24.1 to 72.2 nM. This open-label, single-center, single-dose study demonstrated engagement of ONO-2952 to brain TSPO. The relationship between pharmacokinetics and TSPO occupancy observed in this study support the hypothesis that ONO-2952 could potentially modulate neurosteroid production by binding to brain TSPO.

Safety, Tolerability, and Pharmacokinetic Profile of the Novel Translocator Protein 18 kDa Antagonist ONO-2952 in Healthy Volunteers.

PURPOSE: To investigate safety, tolerability, and pharmacokinetic properties of single and multiple doses of novel translocator protein 18 kDa antagonist ONO-2952 in healthy subjects. METHODS: Double-blind, placebo-controlled single (SAD) and multiple (MAD) dose escalation studies were conducted. Healthy men and women aged 18 to 55 years inclusive and without history of psychiatric disorders were eligible. Forty-eight volunteers received single doses of ONO-2952 (3, 10, 30, 100, 200, or 400 mg) or placebo under fasted conditions (SAD study), and 36 received ONO-2952 (30, 60, or 100 mg/d) or placebo for 21 consecutive days under fed conditions (MAD study). ONO-2952 10 and 200 mg were administered under fasted and fed conditions in the SAD study to investigate the effect of food on the absorption of ONO-2952. Safety assessments included adverse events, vital signs, 12-lead ECGs, and clinical laboratory evaluations. Plasma and urine pharmacokinetic profiles of ONO-2952 were determined. FINDINGS: Across both studies, mean age ranged from 29.8 to 39.8 years, most participants were white, and the proportion of female volunteers was 52%. No treatment or dose-related trends in adverse events were observed. The most frequent adverse events were headache and presyncope (n = 2 each [SAD study]) and constipation and headache (n = 3 each [MAD study]). All headache and constipation episodes were possibly related to the study drug. Plasma ONO-2952 concentrations peaked 2.5 to 3.5 hours (SAD study) and 3.0 to 4.0 hours (MAD study) postdose. ONO-2952 systemic exposure increased less than dose proportionally under fasted conditions. Fed conditions significantly increased exposure compared with fasted conditions: geometric mean ratios of Cmax (90% CIs) were 229% (176-299 [10 mg]) and 778% (623-971 [200 mg]), and AUClast were 159% (131-192 [10 mg]) and 382% (288-506 [200 mg]). In the MAD study, the systemic exposure of ONO-2952 increased in a slightly greater than dose-proportional manner. Geometric mean accumulation ratios (95% CI) of AUC24 were 2.50 (2.09-2.98 [30 mg]), 2.23 (1.85-2.68 [60 mg]), and 2.73 (2.10-3.55 [100 mg]); and Cmax were 1.65 (1.43-1.90 [30 mg]), 1.56 (1.31-1.85 [60 mg]), and 1.85 (1.38-2.49 [100 mg]). IMPLICATIONS: ONO-2952 was safe and well tolerated in these early clinical studies investigating safety, tolerability, and pharmacokinetic properties of single and multiple doses. ONO-2952 systemic exposure increased in a less than dose-proportional manner under fasted conditions and in a slightly greater than dose-proportional manner under fed conditions. These results support the progression of ONO-2952 to further studies in humans. SAD study: ClinicalTials.gov identifier: NCT01364441. MAD study: ClinicalTrials.gov identifier: NCT01489345.

Effects of N-alkyl and ammonium groups on the hydrolytic cleavage of DNA with a Cu(II)TACH (1,3,5-triaminocyclohexane) complex. Speciation, kinetic, and DNA-binding studies for reaction mechanism.

cis,cis-1,3,5-Triaminocyclohexane (c-TACH), its N-alkyl-derivatives (alkyl = methyl, ethyl), and trans,cis-1,3,5-triaminocyclohexane (t-TACH) were prepared, and speciation and DNA cleaving property of Cu(II) complexes of these ligands were investigated. All of the complexes efficiently promote the hydrolytic cleavage of supercoiled plasmid DNA under physiological conditions without further additives. The DNA cleavage rate (V(obs)) trend at pH values between 8 and 9 is N-Me(3) = N-Et(1) < t-TACH < c-TACH < N-Et(2) < N-Et(3). At pH 7, the trend is c-TACH < N-Et(3) = N-Et(2) < N-Et(1) < N-Me(3) << t-TACH. The cleavage rate constants at 35 degrees C, for the c-TACH complex are 3 x 10(-1) h(-1) at pH 8.1 and 2 x 10(-1) h(-1) at pH 7.0 ([DNA] = 7 microM, [Cu(II)-complex] = 105 microM). The hydrolytically active species at pH > 8 is CuL(H(2)O)(OH)(+) in which L coordinates to Cu(II) as a tridentate ligand for all complexes except for t-TACH. The hydrolytically active species at pH 7 is CuLH(H(2)O)(3)(3+) or CuLH(H(2)O)(4)(3+) in which LH coordinates as bidentate ligand. DNA-binding constants of c-TACH and t-TACH complexes are presented and the effects of N-alkyl and ammonium groups are discussed in light of the proposed reaction mechanism.

Discovery of new chemical leads for selective EP1 receptor antagonists.

A series of 4-([2-[alkyl(phenylsulfonyl)amino]phenoxy]methyl)benzoic acids were identified as functional PGE(2) antagonists with selectivity for the EP1 receptor subtype starting from a chemical lead 1, which was found while screening our in-house compound library. Discovery of the optimized analogs 21-23 is presented here and structure-activity relationships (SAR) are also discussed.

Combined effects of prostaglandin E receptor subtype EP1 and subtype EP4 antagonists on intestinal tumorigenesis in adenomatous polyposis coli gene knockout mice.

Previous studies have shown that prostaglandin E(2) (PGE(2)) is involved in intestinal carcinogenesis through its binding to the PGE(2) receptor subtypes EP(1) and EP(4) and activation of downstream pathways. ONO-8711 and ONO-AE2-227, prostaglandin E receptor subtype EP(1)- and EP(4)-selective antagonists, respectively, are known to suppress formation of intestinal polyps in adenomatous polyposis coli gene-deficient mice. The present study was designed to investigate the combined effects of EP(1) and EP(4) antagonists on spontaneous polyp formation in APC1309 mice in order to determine the contribution of each receptor to intestinal tumorigenesis. APC1309 mice were treated with 400 ppm of ONO-8711 alone, 400 ppm of ONO-AE2-227 alone or both in combination in the diet for 6 weeks. The mean area of polyps found in the intestine, calculated as the longer diameter x the shorter diameter x pi, was reduced by 12%, 43% (P < 0.01) and 56% (P < 0.01) of the mean control value (8.8 mm(2)) in the ONO-8711 alone, ONO-AE2-227 alone and combination treatment groups, respectively, suggesting clear additive effects of the combination. The same additive tendency for suppression was also observed with respect to the numbers of polyps in the intestine. Polyp size reduction was more remarkable with the EP(4) antagonist, while the number reduction was more pronounced with the EP(1) antagonist. Our results indicate that EP(1) and EP(4) may have separate intrinsic roles and, to some extent, contribute to polyp formation independently. Thus, combination treatment has potential for the chemoprevention of colon carcinogenesis.

Host prostaglandin E(2)-EP3 signaling regulates tumor-associated angiogenesis and tumor growth.

Nonsteroidal antiinflammatories are known to suppress incidence and progression of malignancies including colorectal cancers. However, the precise mechanism of this action remains unknown. Using prostaglandin (PG) receptor knockout mice, we have evaluated a role of PGs in tumor-associated angiogenesis and tumor growth, and identified PG receptors involved. Sarcoma-180 cells implanted in wild-type (WT) mice formed a tumor with extensive angiogenesis, which was greatly suppressed by specific inhibitors for cyclooxygenase (COX)-2 but not for COX-1. Angiogenesis in sponge implantation model, which can mimic tumor-stromal angiogenesis, was markedly suppressed in mice lacking EP3 (EP3(-/-)) with reduced expression of vascular endothelial growth factor (VEGF) around the sponge implants. Further, implanted tumor growth (sarcoma-180, Lewis lung carcinoma) was markedly suppressed in EP3(-/-), in which tumor-associated angiogenesis was also reduced. Immunohistochemical analysis revealed that major VEGF-expressing cells in the stroma were CD3/Mac-1 double-negative fibroblasts, and that VEGF-expression in the stroma was markedly reduced in EP3(-/-), compared with WT. Application of an EP3 receptor antagonist inhibited tumor growth and angiogenesis in WT, but not in EP3(-/-). These results demonstrate significance of host stromal PGE(2)-EP3 receptor signaling in tumor development and angiogenesis. An EP3 receptor antagonist may be a candidate of chemopreventive agents effective for malignant tumors.

Involvement of prostaglandin E receptor subtype EP(4) in colon carcinogenesis.

Accumulating evidence indicates that overproduction of prostanoids attributable to overexpression of cyclooxygenase-2 (COX-2) plays an important role in colon carcinogenesis. We have shown recently that the prostaglandin (PG) E receptor, EP(1), but not EP(3), is involved in mouse colon carcinogenesis. In line with our previous study, here we examined the role of prostanoid receptors in colon carcinogenesis using six additional lines of knockout mice deficient in prostanoid receptors EP(2), EP(4), DP, FP, IP, or TP. The animals were treated with the colon carcinogen, azoxymethane (AOM), and examined for the development of aberrant crypt foci (ACFs), putative preneoplastic lesions in the colon. Formation of ACFs was decreased only in the EP(4)-knockout mice, to 56% of the wild-type level. To confirm these results, we also examined the inhibitory effects of an EP(4)-selective antagonist, ONO-AE2-227, in the diet on the formation of AOM-induced colon ACFs in C57BL/6Cr mice and on the development of intestinal polyps in Min mice. ONO-AE2-227 at a dose of 400 ppm reduced the formation of ACFs to 67% of the control level, and intestinal polyp numbers in Min mice receiving 300 ppm were decreased to 69% of the control level. Plating efficiency assays showed that addition of 1.0 microM ONO-AE1-329, an EP(4)-selective agonist, resulted in a 1.8-fold increase in the colony number of the human colon cancer cell line, HCA-7, similar to the effect of PGE(2). Moreover, EP(4) mRNA expression was clearly observed in normal colon mucosa and colon tumors in mice. Our previous and present results indicate that PGE(2) contributes to colon carcinogenesis through its actions mediated through EP(1) and EP(4) receptors; therefore, antagonists for these two receptors may be good candidates as chemopreventive agents against colon cancer.