A pilot surrogate end point biomarker trial of perillyl alcohol in breast neoplasia.

PURPOSE: Efficient strategies to screen promising agents in early phase development are essential for rapid progress in breast cancer chemoprevention. We report our experience with the natural compound perillyl alcohol (POH) administered in a short-term surrogate end point biomarker (SEB) protocol, using the "window" between diagnostic and definitive surgery. EXPERIMENTAL DESIGN: Eligible patients included those with a diagnosis of atypical ductal hyperplasia, ductal carcinoma in situ, lobular carcinoma in situ, or invasive carcinoma (<3 cm in size) that required further surgery. Thirty-seven of 267 women screened were enrolled in the study (14%). Five women received single-dose POH (1.5 g/m2) 2 days before surgery, 16 received escalating doses of POH (1.2 g/m2 to 4.8 g/m2/day) for 2 days before surgery, and 16 served as untreated controls. Exploratory SEB analysis [estrogen receptor, progesterone receptor, proliferation, apoptosis, M6P/insulin-like growth factor (IGF)-2R, IGF1, IGF2 and transforming growth factor beta] was conducted before and after POH. RESULTS: Only a small portion of the population screened entered the study. Reasons for nonparticipation included protocol ineligibility, conflict of timing of surgery, miscellaneous logistical reasons, or patient's choice. POH administration was well tolerated and did not interfere with surgical management. The power to observe changes in candidate SEB was diminished by a 44% incidence of cases in which the index lesion was not present in the definitive surgical specimen. CONCLUSIONS: Preoperative POH exposure was safe and suitable for a more definitive phase II SEB study. Further investigations must overcome logistical obstacles to accrual, and they must focus on approaches to maximize tissue collection and to incorporate genomic analysis of target lesions.     

Immunotherapy with autologous dendritic cells and tumor-specific synthetic peptides for synovial sarcoma

Synovial sarcoma in an 11-year-old Japanese girl relapsed 5 months after autologous stem cell transplantation. Autologous dendritic cells (DCs) were generated from her peripheral blood mononuclear cells using granulocyte/macrophage colony-stimulating factor and IL-4. Dendritic cells were pulsed with synthetic peptides containing a junctional region of SYT-SSX2 fusion protein generated by t(X;18) and were administered once per week. No side effects were observed. Growth of metastatic nodules in the lung was temporally suppressed. The delayed-type hypersensitivity responses in skin were enhanced to tumor lysate but not to peripheral blood mononuclear cell lysate. The CD3+ cells cultured with pulsed DCs lysed tumor cells in vitro. Immunotherapy using DCs and tumor-specific peptides may be a safe approach in the treatment of childhood cancer.     

Ki16425, a subtype-selective antagonist for EDG-family lysophosphatidic acid receptors

Lysophosphatidic acid (LPA) exerts a variety of biological responses through specific receptors: three subtypes of the EDG-family receptors, LPA1, LPA2, and LPA3 (formerly known as EDG-2, EDG-4, and EDG-7, respectively), and LPA4/GPR23, structurally distinct from the EDG-family receptors, have so far been identified. In the present study, we characterized the action mechanisms of 3-(4-[4-([1-(2-chlorophenyl)ethoxy]carbonyl amino)-3-methyl-5-isoxazolyl] benzylsulfanyl) propanoic acid (Ki16425) on the EDG-family LPA receptors. Ki16425 inhibited several responses specific to LPA, depending on the cell types, without any appreciable effect on the responses to other related lipid receptor agonists, including sphingosine 1-phosphate. With the cells overexpressing LPA1, LPA2, or LPA3, we examined the selectivity and mode of inhibition by Ki16425 against the LPA-induced actions and compared them with those of dioctyl glycerol pyrophosphate (DGPP 8:0), a recently identified antagonist for LPA receptors. Ki16425 inhibited the LPA-induced response in the decreasing order of LPA1 >/= LPA3 > LPA2, whereas DGPP 8:0 preferentially inhibited the LPA3-induced actions. Ki16425 inhibited LPA-induced guanosine 5'-O-(3-thio)triphosphate binding as well as LPA receptor binding to membrane fractions with a same pharmacological specificity as in intact cells. The difference in the inhibition profile of Ki16425 and DGPP 8:0 was exploited for the evaluation of receptor subtypes involved in responses to LPA in A431 cells. Finally, Ki16425 also inhibited LPA-induced long-term responses, including DNA synthesis and cell migration. In conclusion, Ki16425 selectively inhibits LPA receptor-mediated actions, especially through LPA1 and LPA3; therefore, it may be useful in evaluating the role of LPA and its receptor subtypes involved in biological actions.     

Overexpression of V-1 prevents nitric oxide-induced cell death: involvement of enhanced tetrahydrobiopterin biosynthesis

Previously we reported that the synthesis of catecholamines, dopamine, and noradrenaline was enhanced by overexpression of V-1 protein, a neuronal protein active in the initial stage of development of the rat cerebellum, in the neuronal cell line PC12D, a model of dopamine cells (Yamakuni et al. [1998] J. Biol. Chem. 273:27051-27054). To investigate the physiological role of this protein, we examined the effect of V-1 overexpression on cell toxicity induced by nitric oxide (NO) used at low concentrations. Two clones of PC12D cells overexpressing V-1, transfectants termed V1-46 and V1-69, were significantly more resistant to NOR3 (an NO donor) but not to etoposide (an inhibitor of topoisomerase II)-induced apoptotic cell death than the control cells (termed C-7 and C-9) that had been transfected with the vector alone. The addition of L-DOPA, dopamine, or noradrenaline to the medium did not abolish NOR3-induced cell death in PC12D cells. Moreover, pretreatment of V1-46 and V1-69 cells with L-alpha-methyl-p-tyrosine (alpha-MPT), an inhibitor of tyrosine hydroxylase, to inhibit catecholamine biosynthesis did not affect the resistance to NO toxicity. These results indicate that the catecholamine levels increased by V-1 overexpression did not produce the protection against NOR3-induced toxicity. We further showed that overexpression of V-1 enhanced the synthesis of (6R)-L-erythro-5,6,7,8-tetrahydrobiopterin (BH(4)). In addition, pretreatment with BH(4) or with sepiapterin, which is converted to BH(4) intracellularly, significantly protected PC12D cells in a dose-dependent manner. The increased BH(4) synthesis by V-1 overexpression was dose dependently inhibited by pretreatment with diaminohydroxypyrimidine (DAHP), an inhibitor of GTP-cyclohydrolase I, which is the rate-limiting enzyme for the biosynthesis of BH(4), concomitantly with the loss of protective effect afforded by V-1 overexpression. Furthermore, the addition of BH(4) or sepiapterin to DAHP-pretreated V146 and V1-69 cells restored cell viability. Taken together, these results indicate that V1 protein plays an important role in protection against cell death induced by NO at low levels by promoting the synthesis of BH(4). Moreover, these findings suggest the up-regulation of V1 expression as a possible therapeutic target for protection against the insult of NO-induced oxidative stress.     

DNA-adduct formation in lungs,nasal mucosa,and livers of rats exposed to urban roadside air in Kawasaki City,Japan

The potency of ambient air for DNA-adduct formation was estimated using Wistar rats. The animals were maintained in a small-animal facility located beside a main highway intersection in Kawasaki City, Japan, for up to 60 weeks and were exposed to roadside air contaminated mainly with automobile emission (exposure group, EG) or to clean air (control group, CG). Compared to CG, the relative adduct levels (RAL) were increased significantly in EG lungs (17.1-fold (P<0.05)), nasal mucosa, and livers after exposure for 4 weeks. However, there were no significant differences in RAL between EG and CG after exposure for 12 weeks, but they were elevated again in EG after exposure for 48 or 60 weeks. These results suggest that roadside air in this region can cause the generation of DNA adducts. This activity of ambient roadside air can be estimated using experimental animals, indicating that biological monitoring of DNA-adduct formation may be a powerful tool to assess the effect of ambient air on human health.     

A comparison of heart rate during rest and work in shift workers with different work styles

To determine if the type of work performed should be considered in research on shift work and cardiovascular disease, we compared the heart rates, total number of steps walked, and blood pressures of 12 shift workers on the same rotating 3-shift schedule in a pulp and paper mill. Six workers were selected from the paper manufacturing section (group 1) and six workers from the chemical products section (group 2). Average heart rate (in beats per min) monitored during duty time was 84.3 in group 1 and 87.4 in group 2. Average heart rate during work was not significantly higher than that during rest in both groups 1 (work 85.8, rest 75.3) and 2 (work 87.9, rest 83.1). There was no significant difference in the total number of steps walked. A non-significant decrease in systolic blood pressure value was found in group 1 compared with that in group 2. Although future studies will be needed to explain the relation between different work styles and their effects on the health of shift workers, our results suggest no significant difference in heart rates among workers engaged in different kinds of work on the same shift work schedule.     

Comparison between cultures of urine sediment and of the vagina for detection of carriers of group B Streptococcus (GBS) among pregnant women

An attempt was made to detect carriers of group B Streptococcus (GBS) among pregnant women by vaginal culture. Reported carrier rates have varied from 3% to 15% and one study has reported that cultures of urine sediment showed a very high positivity rate of 30% among non-pregnant women. In this study, we cultured urine sediment from 110 non-pregnant and 415 pregnant women. Among non-pregnant women, 26% were found to be GBS carriers by culture of urine sediment. Among pregnant women, the carrier rate was 3% by vaginal culture and 15% by culture of urine sediment. The serotypes of cases positive by vaginal vulture were identical with those positive by urine sediment culture. For detection of GBS carriers among pregnant women, culture of urine sediment is easy and reduces the rate of false negativity.     

Changing pattern in Reserve Capacity of Gonatropin Secretion

Reserve capacity of Lh and FSH secretion was examined by standarized LHRH loading rest in 30 endocrinologically normal children, whose age ranged from 3 to 13 years. These sunjects were divided by sex. Both male and female subjects were divided into three groups by their chronological age, that is, Group A: under 6 years, Group B: 7 to 10 years and Group C: over 11 years of age. In the male the peakl values of LH were 10.86 ± 1.9 (mean ± SD) for Group a, 23,3 ± 7.3 for Group B, and 73.13 ± 6.4 mIU/ml for Group C. Corresponding values in the female were 8.58 ± 1.3, 20.56 ± 3, and 60.38 ± 8.5 mIU/ml, respectively. These intergroup differences in the peak LH values weer statistically significant (p<0.01)in both sexes. Regarding peak FSH values, no difference was seen among groups of either male or female cases. These findings indicate that even in prepuberty the reserve capacity of LH secretion begins to increase around the age of about 7 years.     

Contrast-enhanced sonography of pancreatic ductal carcinoma using agent detection imaging

Purpose The aim of this study was to evaluate the enhancement behavior of pancreatic ductal carcinoma by contrast-enhanced sonography with agent detection imaging (ADI), and to clarify the origin of microbubble signals by comparisons with histological findings of resected specimens. Methods The subjects were 21 patients with resectable pancreatic carcinoma. The final histological diagnosis was tubular adenocarcinoma in 20 cases, and anaplastic carcinoma in one case. Ultrasound examinations were performed using an Acuson Sequoia 512 series system, and the contrast agent (Levovist) was injected intravenously in doses of 7 ml (300 mg/ml). The ADI signals (in the tumor) were recorded continuously for 30 s after an injection of Levovist (vascular image) and then obtained intermittently (30 s time-intervals) until the signal had diminished in pancreatic tissue (perfusion image). Results Contrast enhancement of the tumor was observed in 71.4% of subjects on the vascular image and 76.3% of subjects on the perfusion image. Enhancement patterns on the vascular image were classified into three types: VI-1 (linear enhancement), VI-2 (spotty enhancement), and VI-3 (no enhancement). VI-1, VI-2, and VI-3 were seen in 9 (42.8%), 6 (28.6%), and 6 (28.6%) of the 21 cases, respectively. Enhancement patterns on the perfusion image were classified into four types: PI-1 (diffuse uneven enhancement), PI-2 (spotty enhancement), PI-3 (peripheral enhancement), and PI-4 (negative enhancement). The incidence of PI-1, PI-2, PI-3, and PI-4 was 4.8%, 42.9%, 28.6%, and 23.8%, respectively. With respect to resectable cases, these enhancement patterns were compared with histological findings, i.e., the distribution of blood vessels in the tumor, remaining pancreatic tissues in the tumor, differentiation of types of adenocarcinoma, volume of stroma, and invasion types of carcinoma. The enhanced patterns consequently corresponded to either the distribution of the blood vessels or the remaining pancreatic tissues in the tumor. Conclusion This study indicated that pancreatic ductal carcinoma is frequently enhanced by microbubbles, and the signals seem to originate from fine blood vessels and the remaining pancreatic tissues in the tumor.     

Effect of clenbuterol on contractile response in periurethral striated muscle of rabbits.

The effect of clenbuterol, a selective beta 2-adrenoceptor agonist, on isolated periurethral striated muscle preparations from rabbits has been investigated. The periurethral striated muscle produced a contraction in response to field stimulation. An application of clenbuterol resulted in a dose-dependent potentiation of the field stimulation-induced contraction. This potentiation was antagonized by propranolol and was greater than that of isoproterenol, suggesting a beta 2-agonistic action.