Role of squalene synthase in prostate cancer risk and the biological aggressiveness of human prostate cancer.

BACKGROUND: We previously conducted a genome-wide linkage analysis of Japanese nuclear families affected with prostate cancer and showed that the susceptibility to prostate cancer was closely linked to D8S550 at 8p23. The role of farnesyl diphosphate farnesyltransferase (FDFT1), which is located under the peak marker D8S550 at 8p23, and squalene synthase, the enzyme encoded by FDFT1, in prostate cancer was studied. METHODS: The association among common variants of FDFT1 with prostate cancer risk, the promoter activities of FDFT1 with different genotypes and the effects of inhibition of squalene synthase were studied, and the FDFT1 transcript levels of human prostate samples were quantified. RESULTS: The A allele of rs2645429 was significantly associated with prostate cancer risk in a Japanese familial prostate cancer population. Rs2645429 was located in the promoter region of FDFT1, and the AA genotype showed significantly increased promoter activity. The knockdown of FDFT1 mRNA expression or squalene synthase inhibition led to a significant decrease in prostate cancer cell proliferation. Additionally, human prostate cancer specimens expressed significantly higher levels of FDFT1 mRNA compared with noncancerous specimens. Finally, aggressive cancers showed higher transcript levels. CONCLUSIONS: FDFT1 and its encoded enzyme, squalene synthase, may play an important role in prostate cancer development and its aggressive phenotypes.

Assimilation and translocation of nitrogen and carbon in Curcuma alismatifolia Gagnep.

Curcuma or Siam tulip (Curcuma alismatifolia Gagnep.) is an ornamental flowering plant with two underground storage organs, rhizomes and storage roots. Characteristics of N and C assimilation and transport in curcuma were investigated. The plants were treated with (15)NH(4) (+) + (15)NO(3) (-) and (13)CO(2) at 10, 13 or 21 weeks after planting. Plants were sampled at several stages up to 32 weeks. The C stored in old storage roots was used rapidly during the first 10 weeks; after which N stored in old rhizomes and old storage roots were used. The daily gain in C depending on photosynthesis was remarkably high between 10 and 21 weeks. However, the daily gain in N was relatively constant throughout the growth period. The (15)N absorbed at 10 weeks was initially accumulated in leaves and roots, but some was transported to flowering organs at 13 weeks. At harvest, 41% of (15)N was recovered in new rhizomes and 17% in new storage roots. After (13)CO(2) exposure at 10 and 13 weeks, the distribution of (13)C among organs was relatively constant in subsequent stages. When given (13)CO(2) at 21 weeks, a large amount of labelled C was recovered in new storage roots and new rhizomes at harvest. Both new rhizomes and new storage roots stored N and C, however, rhizomes played a more important role in supplying N, while storage roots provided C.

Role of genetic polymorphisms of the RNASEL gene on familial prostate cancer risk in a Japanese population.

The RNASEL gene on chromosome 1q25 has been identified as a prostate cancer susceptibility gene. We screened for RNASEL germline mutations in familial prostate cancer patients, and performed a case-control study to examine the association of specific variants with prostate cancer risk in the Japanese. Three variants within the RNASEL gene, G282A, G1385A and T1623G were identified. G1385 and T1623G variants result in previously reported Arg462Gln and Asp541Glu variants, respectively. The novel G282A variant does not cause amino-acid substitution. A case-control study consisting of 101 familial prostate cancer cases and 105 noncancer controls showed that the Gln/Gln genotype of codon462 was observed in 7.6% of controls. However, the Gln/Gln genotype was not observed in cases, and reduced prostate cancer risk (odds ratio (OR)=0.061, P=0.014). The Asp/Asp genotype of codon541 increased the familial prostate cancer risk (OR=7.37, P=0.0004). In subset analysis, a significant association was observed in patients with more than two affected members (OR=3.15, P=0.028), and weak associations were found in patients with metastatic disease (OR=2.40, P=0.11) and high-grade disease (Gleason score >or=7) (OR=3.07, P=0.14). These findings suggested that the polymorphic changes within the RNASEL gene may be associated with familial prostate cancer risk in a Japanese population.

Novel mutations in GJB2 encoding connexin-26 in Japanese patients with keratitis-ichthyosis-deafness syndrome.

BACKGROUND: Germline missense mutations in the GJB2 gene that encodes connexin-26 (Cx26) have recently been found to be the cause of the keratitis-ichthyosis-deafness (KID) syndrome. OBJECTIVES: To define the GJB2 mutations in three Japanese patients with KID syndrome. METHODS: Genomic DNA was extracted from peripheral blood and used to amplify the GJB2 gene. Direct sequencing and endonuclease digestion were used for mutation analysis and DNA-based diagnosis. RESULTS: We identified two heterozygous mis-sense mutations (D50Y, D50N) in the GJB2 gene in three Japanese patients with KID syndrome. All mutations were located on the first extracellular domain of Cx26. CONCLUSIONS: These data expand the GJB2 mutation database and show that a dominant mutation of Cx26 can cause KID syndrome in Japanese patients.

Malignant fibrous histiocytoma of the right spermatic cord: a case report.

The patient was a 47-year-old male, who visited Hidaka Hospital with a chief complaint of swelling in the right inguinal region and the scrotum. With a diagnosis of a right spermatic cord tumor, right high orchiectomy was performed. Since an inflammatory type of malignant fibrous histiocytoma (MFH) was diagnosed from histopathological findings, chemotherapy and radiation therapy were performed as postoperative treatment. Malignant fibrous histiocytoma with the primary focus of the spermatic cords is a rare disease. To our knowledge, this is the 20th case of MFH of the spermatic cord in Japan (the 42nd in the world) and it is the second case of inflammatory type of MFH in Japan.

Dicationic dithiocarbamate carbapenems with anti-MRSA activity.

A new class of 1 beta-methylcarbapenems bearing a doubly quaternarized 1,4-diazabicyclooctane (DABCO) substituted dithiocarbamate moiety at the C-2 side chain was prepared, and the biological profiles of the compounds, including in vitro and in vivo anti-MRSA activity and DHP-I susceptibility, were evaluated to identify a carbapenem derivative that was superior to BO-3482 (1). As a result, we discovered a 1 beta-methyl-2-[4-(4-carbamoylmethyl-1,4-diazabicyclo[2,2,2]octanediium-1-yl)methyl-1,2,3,6-tetrahydropyridinylthiocarbonylthio]carbapenem, 14a showing greater than 2-fold better anti-MRSA activity in a mouse infection model and 3-fold better DHP-I susceptibility as compared with BO-3482 (1).

Discovery of a novel CCR3 selective antagonist.

In searching for a novel CCR3 receptor antagonist, we designed a library that included a variety of carboxamide derivatives based on the structure of our potent antagonists for human CCR1 and CCR3 receptors, and screened the new compounds for inhibitory activity against 125I-Eotaxin binding to human CCR3 receptors expressed in CHO cells. Among them, two 2-(benzothiazolethio)acetamide derivatives (1a and 2a) showed binding affinities with IC50 values of 750 and 1000 nM, respectively, for human CCR3 receptors. These compounds (1a and 2a) also possessed weak binding affinities for human CCR1 receptors. We selected la as a lead compound for derivatization to improve in vitro potency and selectivity for CCR3 over CCRI receptors. Derivatization of la by incorporating substituents into each benzene ring of the benzothiazole and piperidine side chain resulted in the discovery of a compound (1b) exhibiting 820-fold selectivity for CCR3 receptors (IC50 = 2.3 nM) over CCR1 receptors (IC50 = 1900 nM). This compound (1b) also showed potent functional antagonist activity for inhibiting Eotaxin (IC50 = 27 nM)- or RANTES (IC50 = 13 nM)-induced Ca2+ increases in eosinophils.

Pharmacological properties of (2R)-N-[1-(6-aminopyridin-2-ylmethyl)piperidin-4-yl]-2-[(1R)-3,3-difluorocyclopentyl]-2-hydroxy-2-phenylacetamide: a novel mucarinic antagonist with M(2)-sparing antagonistic activity.

We evaluated the pharmacological profiles of (2R)-N-[1-(6- aminopyridin-2-ylmethyl)piperidin-4-yl]-2-[(1R)-3,3-difluorocyclopentyl]-2-hydroxy-2-phenylacetamide(compound A), which is a novel muscarinic receptor antagonist with M(2)-sparing antagonistic activity. Compound A inhibited [(3)H]NMS binding to cloned human muscarinic m1, m2, m3, m4, and m5 receptors expressed in Chinese hamster ovary cells with K(i) values (nM) of 1.5, 540, 2.8, 15, and 7.7, respectively. In isolated rat tissues, compound A inhibited carbachol-induced responses with 540-fold selectivity for trachea (K(B) = 1.2 nM) over atria (K(B) = 650 nM). In in vivo rat assays, compound A inhibited acetylcholine-induced bronchoconstriction and bradycardia with intravenous ED(50) values of 0.022 mg/kg and >/=10 mg/kg, respectively. Furthermore, in dogs, compound A (0.1-1 mg/kg p.o.) dose dependently shifted the methacholine concentration-respiratory resistance curves. In mice, compound A (10 mg/kg i.v.) did not inhibit oxotremorine-induced tremor. The brain/plasma ratio (K(p)) of compound A (3 mg/kg i.v.) was 0.13 in rats; this K(p) was less than that of scopolamine (1.7) and darifenacin (0.24). The inhibition of compound A (3 mg/kg i.v.) on ex vivo binding in rat cerebral cortex was almost similar to that of NMS. These findings demonstrate that compound A has high selectivity for M(3) receptors over M(2) receptors, displays a potent, oral M(3) antagonistic activity without inhibition of central muscarinic receptors because of low brain penetration. It is well known that central muscarinic antagonists may have diverse CNS effects, and M(2) receptors regulate cardiac pacing and act as autoreceptors in the lung and bladder. Thus, compound A may have fewer cardiac or CNS side effects than nonselective compounds.

Rapid N transport to pods and seeds in N-deficient soybean plants.

Non-nodulated soybean (Glycine max (L.) Merr.) plants were cultivated hydroponically under N-sufficient (5 mM NaNO(3)) or N-deficient (0.5 mM NaNO(3)) conditions. (13)N- or (15)N- labelled nitrate was fed to the cut end of the stems, and the accumulation of nitrate-derived N in the pods, nodes and stems was compared. Real-time images of (13)N distribution in stems, petioles and pods were obtained using a Positron Emitting Tracer Imaging System for a period of 40 min. The results indicated that the radioactivity in the pods of N-deficient plants was about 10 times higher than that of N-sufficient plants, although radioactivity in the stems and nodes of N-deficient versus N-sufficient plants was not different. A similar result was obtained by supplying (15)NO(3) to cut soybean shoots for 1 h. The fact that the N translocation into the pods from NO(3) fed to the stem base was much faster in N-deficient plants may be due to the strong sink activity of the pods in N-deficient plants. Alternatively, the redistribution of N from the leaves to the pods via the phloem may be accelerated in N-deficient plants. The temporal accumulation of (13)NO(3) in nodes was suggested in both N-sufficient and N-deficient plants. In one (13)NO(3) pulse-chase experiment, radioactivity in the stem declined rapidly after transferring the shoot from the (13)NO(3) solution to non-labelled NO(3); in contrast, the radioactivity in the node declined minimally during the same time period.

Design, synthesis, and discovery of a novel CCR1 antagonist.

The CC chemokines may play an important role in the pathogenesis of chronic inflammatory diseases including rheumatoid arthritis, and their effects are thought to be mediated through CCR1 receptors. Several nonpeptide CCR1 receptor antagonists that showed high affinity for human CCR1 receptors have been identified; however, their effectiveness in animal models of inflammatory diseases has been scarcely demonstrated, probably due to species selectivity of the antagonists. To elucidate the pathophysiological role of CCR1 receptors in murine models of disease, we looked for a potent antagonist for both murine and human CCR1 receptors. Screening of our chemical collection for inhibition of (125)I-MIP-1alpha binding to human CCR1 receptors transfected in CHO cells led to the identification of xanthene-9-carboxamide 1a as the lead compound. Derivatization of 1a by quaternarizing the piperidine nitrogen with various alkyl groups and by installing substituents into the xanthene moiety dramatically improved the inhibitory activity against both human and murine CCR1 receptors. As a result, 2q-1 showing IC(50) values of 0.9 and 5.8 nM for human and murine CCR1 receptors, respectively, was discovered. This compound is the first murine CCR1 receptor antagonist and may be a useful tool for clarifying the role of CCR1 receptors in murine models of disease.

Identification of a potent and nonpeptidyl ccr3 antagonist.

CCR3 is expressed in a variety of leukocyte subsets, especially eosinophils, and may be involved in allergic disorders such as atopic asthma. To clarify the pathophysiological roles of CCR3 in allergic disorders, we developed a nonpeptidyl CCR3 antagonist. This antagonist, which is referred to as "Compound X," that inhibited the binding of [(125)I]Eotaxin to CHO cells transfected with human CCR3 with an IC(50) value of 2.3 nM. In human eosinophils, Compound X also inhibited Eotaxin-induced increases in intracellular Ca(2+) concentrations and chemotaxis. Thus, Compound X appears to be a highly potent CCR3 antagonist. These findings suggest that Compound X may be a useful tool for elucidating the pathophysiological roles of CCR3 in a variety of allergic disorders.

Discovery of novel trans-3,5-disubstituted pyrrolidinylthio-1beta-methylcarbapenems.

Novel trans-3,5-disubstituted pyrrolidinylthio-1beta-methylcarbapenems were designed and synthesized to provide J-111,347 (1a) as the first example of an exceptionally broad-spectrum antibiotic, showing activity against methicillin-resistant Staphyloccocus aureus (MRSA) as well as Pseudomonas aeruginosa. Further derivation of 1a afforded J-111,225 (2a), J-114,870 (3a), and J-114,871 (3b). which showed improved safety profiles and retained broad-spectrum antibacterial activities.

[Hereditary prostate cancer].

Familial prostate cancer patients are sometimes encountered. Hereditary prostate cancer is a more specific form of familial prostate cancer that is inherited by a susceptibility gene consistent with Mendelian inheritance. Early age at onset is the most important characteristic. No clear differences in either stage, grade or prognosis have been found between hereditary and sporadic prostate cancer. No susceptibility genes have been isolated yet, but several genes may exist. In Japan, doctors are not generally aware of hereditary and familial prostate cancer. Family history is one of the most important risk factors of prostate cancer. We should make an effort to find prostate cancer patients at an early stage in the high risk families.

Trends and characteristics in prostate cancer mortality in Japan.

BACKGROUND: In North America, the incidence and mortality of prostate cancer has been declining since the early 1990s. We calculated the age-adjusted death rates, age-specific death rates and standardized mortality ratio (SMR) for prostate cancer in Japan and analyzed their features. METHODS: Yearly age-adjusted death rates for prostate cancer were calculated by dividing the number of events by the population at risk, with direct standardization to the world population. Age-specific death rates were calculated for the 1970s, 1980s and 1990s and which age group showed the highest rate of increase was examined. The SMR in each prefecture was also calculated for each period. RESULTS: The respective number of deaths and the age-adjusted death rate was 1107 and 2.29 in 1973 and 6251 and 5.15 in 1997. The age-specific death rates showed an exponential increase with age in all three periods and the rate of increase was higher in older age groups. The distribution of SMR showed the same tendency in all three periods. The prefectures with significantly high or low SMR were distributed in clusters. CONCLUSIONS: The prostate cancer death rate is increasing rapidly in Japan. However, the age-adjusted death rate has remained stable from 1996 to 1997. How this figure will change and whether the prostate cancer death rate in Japan will begin to decline, like in North America, is of interest. The prefectures with significantly high or low SMR showed a characteristic clustered distribution pattern.

Discovery of a muscarinic M3 receptor antagonist with high selectivity for M3 over M2 receptors among 2-[(1S,3S)-3-sulfonylaminocyclopentyl]phenylacetamide derivatives.

In the course of developing a metabolically stable M3 receptor antagonist from the prototype antagonist, J-104129 (1), introduction of certain substituents into the cyclopentane ring of 1 was found to be effective not only in improving metabolic stability but also in greatly enhancing the subtype selectivity. Among the cyclopentane analogues, sulfonamide derivatives (10f) and (10g) displayed 160- and 310-fold selectivity for M3 over M2 receptors, and both were significantly more selective than the prototype antagonist (120-fold). Subsequent derivatization of the sulfonamide series led to the highly selective M3 receptor antagonists (10h, 10i and 10j) with >490-fold selectivity for M3 over M2 receptors. Among them, p-nitrophenylsulfonamide (J-107320, 10h) exhibited 1100-fold selectivity for M3 receptors (Ki = 2.5 nM) over M2 receptors (Ki = 2800 nM) in the human muscarinic receptor binding assay using [3H]-NMS as a radio ligand.

Modulation of lung local immune responses by oral administration of a herbal medicine Sho-saiko-to.

Sho-saiko-to (SST), a Chinese/Japanese herbal medicine (Kampo medicine) widely used to treat chronic hepatitis in Japan, is known to modulate immune responses, and thus its immunomodulating activity may be responsible for its bi-directional effects on the lungs as therapeutic efficacy in various lung diseases and involvement in development of interstitial pneumonia. We administered SST to BALB/c mice orally and examined the lung tissue levels of pro/anti-inflammatory cytokines, interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6), and the effects of SST on acute lung injury induced by instillation of lipopolysaccharide (LPS) or IL-1. Although SST had no effect on lung TNF-alpha or IL-1beta level, it increased IL-6. Investigation of active fractions of SST suggested that multiple ingredients were supposed to be responsible for IL-6-inducing activity. Liquiritigenin, a metabolite of liquiritin which is one of the major ingredients in SST enhanced in vitro IL-6 production in anti-CD3 monoclonal antibody (anti-CD3 mAb)-stimulated lung mononuclear cells in a cell-type specific and dose-dependent manner. SST suppressed LPS-induced lung injury at the later phase when lung leak was evident while being ineffective on initial neutrophil sequestration to the lung in these models. These findings suggest that SST modulates lung inflammation by regulating local immune response.

Synthesis of J-111,347, a novel 1 beta-methylcarbapenem with broad-spectrum antibacterial activity.

Synthesis of J-111,347 (1), a new 1 beta-methylcarbapenem with broad-spectrum antibacterial activity including that against methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa, was achieved via diastereoselective preparation of a side-chain thiol 3 from an optically active (R)-3,4-dihydroxybutanal 4.

Structure-activity relationships of trans-3,5-disubstituted pyrrolidinylthio-1beta-methylcarbapenems. Part 1: J-111,347 and related compounds.

1Beta-methylcarbapenems having various 3,5-disubstituted pyrrolidinylthio-side chains at C-2 were designed and synthesized. Evaluation of their antibacterial activities indicated that J-111,347 (1a) is the first example of an extremely broad spectrum antibiotic showing activity against methicillin-resistant Staphylococcus aureus (MRSA) as well as Pseudomonas aeruginosa.