Loureirin C and Xanthoceraside Prevent Abnormal Behaviors Associated with Downregulation of Brain Derived Neurotrophic Factor and AKT/mTOR/CREB Signaling in the Prefrontal Cortex Induced by Chronic Corticosterone Exposure in Mice.

Brain derived neurotrophic factor (BDNF) is one of the most abundant neurotrophic factors, and its deficits are involved in the pathogenesis of major depressive disorders (MDD). Loureirin C (Lou C) is a compound derived from red resin extracted from the stems of Chinese dragon's blood. Xanthoceraside (Xan) is a triterpenoid saponin extracted from the husks of Xanthoceras sorbifolia Bunge. These compounds have neuroprotective effects through upregulation of BDNF. The present study aimed to evaluate whether Lou C and Xan attenuate abnormal behaviors induced by chronic corticosterone (CORT) administration. CORT was administered subcutaneously to mice for 3 weeks, and Lou C and Xan, dispensed orally once a day during the last 2 weeks of CORT administration. Chronic CORT administration induced abnormal behaviors such as prolonged starting latency in the open field test, decreased social interaction time in the social interaction test and prolonged latency to eat in the novelty suppressed feeding test. Chronic CORT administration decreased the expression levels of BDNF and the phosphorylation of protein kinase B (Akt), mammalian target of rapamycin (mTOR), and the cAMP response element binding protein (CREB) in the prefrontal cortex. Lou C and Xan dose-dependently prevented the abnormal behaviors and decreased the expression levels of BDNF and in phosphorylation of AKT, mTOR, and CREB in the prefrontal cortex of CORT mice. These results suggest that Lou C and Xan could be attractive candidates for pharmacotherapy of MDD at least in part, given their propensity to increase BDNF expression and phosphorylation of AKT, mTOR, and CREB.

Efficacy and safety of talazoparib in Japanese patients with germline BRCA-mutated locally advanced or metastatic breast cancer: results of the phase 1 dose-expansion study.

BACKGROUND: Talazoparib, a poly(ADP-ribose) polymerase enzyme inhibitor, is approved for the treatment of patients with germline BRCA1/2 (gBRCA1/2)-mutated HER2-negative advanced breast cancer. This two-part study, a recently published dose-escalation part followed by the dose-expansion part reported here, evaluated the efficacy and safety of talazoparib in Japanese patients with gBRCA1/2-mutated advanced breast cancer. METHODS: In this open-label, multicenter phase 1 study (NCT03343054), the primary endpoint of the dose-expansion part was confirmed objective response rate (ORR), determined by investigator assessment (RECIST 1.1). Secondary endpoints included progression-free survival (PFS), overall survival (OS), safety, and pharmacokinetics. Patients received the recommended phase 2 dose (1 mg/day; 0.75 mg/day moderate renal impairment). RESULTS: Nineteen Japanese patients with gBRCA1/2-mutated locally advanced or metastatic breast cancer were enrolled. Confirmed ORR was 57.9% (11/19; 90% confidence interval [CI] 36.8-77.0). Stable disease was observed in 36.8% (7/19) of patients. Per investigator assessment, median PFS was 7.2 months (95% CI 4.1-not estimable) and 12-month OS rate was 84.7% (90% CI 57.5-95.1). Median OS was not reached; 17/19 patients were alive and censored at 12 months. All patients experienced treatment-related adverse events (AEs); the majority were hematologic. The most common treatment-related AE was anemia (68.4%; [13/19]). Grade 3/4 treatment-related AEs were observed in 52.6% (10/19) of patients. During the safety period, there were no grade 5 treatment-emergent AEs, treatment-related serious AEs, or deaths. CONCLUSIONS: In Japanese patients with gBRCA mutations and locally advanced or metastatic breast cancer, talazoparib monotherapy was generally well tolerated and resulted in clinically meaningful ORRs. GOV IDENTIFIER: NCT03343054.

Safety, pharmacokinetics, and preliminary efficacy of the PARP inhibitor talazoparib in Japanese patients with advanced solid tumors: phase 1 study.

BACKGROUND: Talazoparib is a poly(ADP-ribose) polymerase enzyme inhibitor. This open-label, non-randomized, phase 1 study of talazoparib investigated the safety, pharmacokinetics, and preliminary antitumor activity in Japanese patients with locally advanced or metastatic solid tumors, regardless of mutations in DNA damage repair-related genes, who are resistant to/ineligible for standard therapies. METHODS: Patients received talazoparib dosed orally at 0.75 or 1 mg once daily using a modified 3 + 3 dose-escalation scheme. Primary endpoint was dose-limiting toxicities during the first cycle of talazoparib. RESULTS: Nine patients (median age 62.0 years) were included: 3 and 6 patients at the 0.75 and 1.0 mg once-daily dose levels, respectively. No dose-limiting toxicities were reported. The most commonly reported treatment-emergent adverse events (≥2 patients) were anemia, stomatitis, maculopapular rash, platelet count decreased, neutrophil count decreased, and alanine aminotransferase increased. Three patients had grade ≥ 3 treatment-emergent adverse events (anemia, brain metastases [1 patient each], and neutrophil and white blood cell count decreased [same patient]). Two patients temporarily discontinued treatment due to a treatment-emergent adverse event, and 1 patient required a dose reduction for neutrophil count decreased (all at 1 mg once daily). Talazoparib exposure (Cmax and AUC) after single and multiple dosing was slightly higher proportionally with talazoparib 1 mg than talazoparib 0.75 mg. The overall disease control rate was 44.4%, including 2 patients with stable disease. The recommended phase 2 dose of talazoparib was established as 1 mg once daily. CONCLUSIONS: Single-agent talazoparib was well tolerated and had preliminary antitumor activity in Japanese patients with advanced solid tumors. ClinicalTrials.gov identifier: NCT03343054 (November 17, 2017).

Evaluation of the Association of Polymorphisms With Palbociclib-Induced Neutropenia: Pharmacogenetic Analysis of PALOMA-2/-3.

BACKGROUND: The most frequently reported treatment-related adverse event in clinical trials with the cyclin-dependent kinase 4/6 (CDK4/6) inhibitor palbociclib is neutropenia. Allelic variants in ABCB1 and ERCC1 might be associated with early occurrence (i.e., end of week 2 treatment) of grade 3/4 neutropenia. Pharmacogenetic analyses were performed to uncover associations between single nucleotide polymorphisms (SNPs) in these genes, patient baseline characteristics, and early occurrence of grade 3/4 neutropenia. MATERIALS AND METHODS: ABCB1 (rs1045642, rs1128503) and ERCC1 (rs3212986, rs11615) were analyzed in germline DNA from palbociclib-treated patients from PALOMA-2 (n = 584) and PALOMA-3 (n = 442). SNP, race, and cycle 1 day 15 (C1D15) absolute neutrophil count (ANC) data were available for 652 patients. Univariate and multivariable analyses evaluated associations between SNPs, patient baseline characteristics, and early occurrence of grade 3/4 neutropenia. Analyses were stratified by Asian (n = 122) and non-Asian (n = 530) ethnicity. Median progression-free survival (mPFS) was estimated using the Kaplan-Meier method. The effect of genetic variants on palbociclib pharmacokinetics was analyzed. RESULTS: ABCB1 and ERCC1_rs11615 SNP frequencies differed between Asian and non-Asian patients. Multivariable analysis showed that low baseline ANC was a strong independent risk factor for C1D15 grade 3/4 neutropenia regardless of race (Asians: odds ratio [OR], 6.033, 95% confidence interval [CI], 2.615-13.922, p < .0001; Non-Asians: OR, 6.884, 95% CI, 4.138-11.451, p < .0001). ABCB1_rs1128503 (C/C vs. T/T: OR, 0.57, 95% CI, 0.311-1.047, p = .070) and ERCC1_rs11615 (A/A vs. G/G: OR, 1.75, 95% CI, 0.901-3.397, p = .098) were potential independent risk factors for C1D15 grade 3/4 neutropenia in non-Asian patients. Palbociclib mPFS was consistent across genetic variants; exposure was not associated with ABCB1 genotype. CONCLUSION: This is the first comprehensive assessment of pharmacogenetic data in relationship to exposure to a CDK4/6 inhibitor. Pharmacogenetic testing may inform about potentially increased likelihood of patients developing severe neutropenia (NCT01740427, NCT01942135). IMPLICATIONS FOR PRACTICE: Palbociclib plus endocrine therapy improves hormone receptor-positive/human epidermal growth factor receptor 2-negative advanced breast cancer outcomes, but is commonly associated with neutropenia. Genetic variants in ABCB1 may influence palbociclib exposure, and in ERCC1 are associated with chemotherapy-induced severe neutropenia. Here, the associations of single nucleotide polymorphisms in these genes and baseline characteristics with neutropenia were assessed. Low baseline absolute neutrophil count was a strong risk factor (p < .0001) for grade 3/4 neutropenia. There was a trend indicating that ABCB1_rs1128503 and ERCC1_rs11615 were potential risk factors (p < .10) for grade 3/4 neutropenia in non-Asian patients. Pharmacogenetic testing could inform clinicians about the likelihood of severe neutropenia with palbociclib.

Kynurenine 3-monooxygenase deficiency induces depression-like behavior via enhanced antagonism of α7 nicotinic acetylcholine receptors by kynurenic acid.

Tryptophan (TRP) is metabolized via the kynurenine (KYN) pathway, which is related to the pathogenesis of major depressive disorder (MDD). Kynurenine 3-monooxygenase (KMO) is a pivotal enzyme in the metabolism of KYN to 3-hydroxykynurenine. In rodents, KMO deficiency induces a depression-like behavior and increases the levels of kynurenic acid (KA), a KYN metabolite formed by kynurenine aminotransferases (KATs). KA antagonizes α7 nicotinic acetylcholine receptor (α7nAChR). Here, we investigated the involvement of KA in depression-like behavior in KMO knockout (KO) mice. KYN, KA, and anthranilic acid but not TRP or 3-hydroxyanthranilic acid were elevated in the prefrontal cortex of KMO KO mice. The mRNA levels of KAT1 and α7nAChR but not KAT2-4, α4nAChR, or ß2nAChR were elevated in the prefrontal cortex of KMO KO mice. Nicotine blocked increase in locomotor activity, decrease in social interaction time, and prolonged immobility in a forced swimming test, but it did not decrease sucrose preference in the KMO KO mice. Methyllycaconitine (an α7nAChR antagonist) antagonized the effect of nicotine on decreased social interaction time and prolonged immobility in the forced swimming test, but not increased locomotor activity. Galantamine (an α7nAChR allosteric agonist) blocked the increased locomotor activity and prolonged immobility in the forced swimming test, but not the decreased social interaction time in the KMO KO mice. In conclusion, elevation of KA levels contributes to depression-like behaviors in KMO KO mice by α7nAChR antagonism. The ameliorating effects of nicotine and galantamine on depression-like behaviors in KMO KO mice are associated with the activation of α7nAChR.

Investigation on the Epoxidation of Piperitenone, and Structure-activity Relationships of Piperitenone Oxide for Differentiation-inducing Activity.

Piperitenone oxide, a major chemical constituent of the essential oil of spearmint, Mentha spicata, induces differentiation in human colon cancer RCM-1 cells. In this study, piperitenone oxide and trans-piperitenone dioxide were prepared as racemic forms by epoxidation of piperitenone. The relative configuration between two epoxides in piperitenone dioxide was determined to be trans by 1H NMR analysis and nuclear Overhauser effect spectroscopy (NOESY) in conjunction with density functional theory (DFT) calculations. Optical resolution of (±)-piperitenone oxide by high-performance liquid chromatography (HPLC) using a chiral stationary phase (CSP) afforded both enantiomers with over 98% enantiomeric excess (ee). Evaluation of the differentiation-inducing activity of the synthetic compounds revealed that the epoxide at C-1 and C-6 in piperitenone oxide is important for the activity, and (+)-piperitenone oxide has stronger activity than (-)-piperitenone oxide. The results obtained in this study provide new information on the application of piperitenone oxide and spearmint for differentiation-inducing therapy. Furthermore, natural piperitenone oxide was isolated from M. spicata. The enantiomeric excess of the isolated natural piperitenone oxide was 66% ee. Epoxidation of piperitenone with hydrogen peroxide proceeded in a phosphate buffer under weak basic conditions to give (±)-piperitenone oxide. These results suggest that the nonenzymatic epoxidation of piperitenone, which causes a decrease in the enantiomeric excess of natural piperitenone oxide, is accompanied by an enzymatic epoxidation in the biosynthesis of piperitenone oxide.

Correction to: Neutropenia management with palbociclib in Japanese patients with advanced breast cancer.

The correct name of the last author should be "Masakazu Toi", and not ''Masakuzu Toi" as given in the original publication of the article.

Palbociclib Plus Letrozole as First-Line Therapy in Postmenopausal Asian Women With Metastatic Breast Cancer: Results From the Phase III, Randomized PALOMA-2 Study.

PURPOSE: In PALOMA-2, palbociclib plus letrozole significantly improved progression-free survival (PFS) as initial treatment of estrogen receptor-positive/human epidermal growth factor receptor 2-negative advanced breast cancer. We assessed the benefit of palbociclib plus letrozole in Asians. PATIENTS AND METHODS: Of 666 enrolled postmenopausal women with estrogen receptor-positive/human epidermal growth factor receptor 2-negative advanced breast cancer (no prior treatment of advanced disease), 95 were Asian. Patients were randomly assigned 2:1 to receive palbociclib plus letrozole or placebo plus letrozole. The primary end point was investigator-assessed PFS. Secondary end points were overall survival, objective response, patient-reported outcomes, pharmacokinetics, and safety. RESULTS: Median PFS was significantly longer in Asian patients who received palbociclib plus letrozole versus placebo plus letrozole (25.7 months [95% CI, 19.2 months to not estimable] v 13.9 months [95% CI, 7.4 to 22.0 months]; hazard ratio, 0.49; 95% CI, 0.27 to 0.87; P = .007). The most common toxicities with palbociclib were hematologic and more frequent among Asians versus non-Asians: neutropenia (any grade, 95.4% v 76.8%; grade 3/4, 89.2% v 62.5%), leukopenia (43.1% v 38.3%; 32.3% v 23.5%), and thrombocytopenia (27.7% v 13.5%; 4.6% v 1.1%). No Asians had febrile neutropenia. Discontinuation rates as a result of adverse events were similar among Asian and non-Asian patients who received palbociclib plus letrozole (10.8% and 9.5%). In Asians, quality of life (QOL) was maintained with no significant differences observed between treatments from baseline in breast cancer-specific QOL and general health status scores. Change from baseline in EuroQol five dimensions index scores was significantly higher with palbociclib plus letrozole (0.013 v -0.069; P = .0132). Geometric mean palbociclib trough concentration values were higher in Asians versus non-Asians (93.8 v 61.7 ng/mL). CONCLUSION: Consistent with the overall study population, the addition of palbociclib to letrozole significantly improved PFS in Asians. Hematologic toxicities were more frequent in Asians versus non-Asians but manageable with early dose modifications while maintaining QOL.

Neutropenia management with palbociclib in Japanese patients with advanced breast cancer.

BACKGROUND: The cyclin-dependent kinase 4/6 (CDK4/6) inhibitor palbociclib, in combination with endocrine therapy (ET), significantly prolonged progression-free survival in women with hormone receptor-positive, human epidermal growth factor receptor 2-negative advanced breast cancer (HR+/HER2- ABC) in PALOMA-2 and PALOMA-3. Neutropenia and palbociclib dose reductions/interruptions occurred more frequently in the Japanese versus overall populations. We evaluated neutropenia patterns, palbociclib dose management, and clinical responses after dose reduction in Japanese patients in PALOMA-2 and PALOMA-3 and a single-arm Japanese phase 2 study. METHODS: PALOMA-2 and the Japanese phase 2 study enrolled postmenopausal women with estrogen receptor-positive, HER2- ABC who had not received prior systemic therapy for advanced disease; PALOMA-3 enrolled women with HR+/HER2- ABC, regardless of menopausal status, whose disease had progressed after prior ET. Palbociclib (125 mg/day) was administered 3 weeks on/1 week off. Dose reduction/interruption, cycle delay, tumor response, and laboratory-assessed neutropenia were analyzed in Japanese patients who received palbociclib. RESULTS: A total of 101 Japanese patients received palbociclib + ET. Among Japanese patients in the 3 studies, the frequency of all-grade/grade 3/grade 4 neutropenia was 94%/53%/34%, 100%/69%/21%, and 100%/67%/26%, respectively. Twenty (63%), 28 (67%), and 15 (56%) patients required palbociclib dose reduction. Dose interruption or reduction did not affect palbociclib treatment duration, and durable tumor response was observed despite dose reduction. CONCLUSION: Neutropenia was manageable with dose modifications, without affecting palbociclib treatment duration or efficacy. TRIAL REGISTRATION: Pfizer (NCT01740427, NCT01684215, NCT01942135).

Palbociclib in combination with fulvestrant in patients with hormone receptor-positive, human epidermal growth factor receptor 2-negative advanced breast cancer: PALOMA-3 subgroup analysis of Japanese patients.

BACKGROUND: In the double-blind, phase 3 PALOMA-3 study, palbociclib-fulvestrant significantly prolonged progression-free survival versus placebo-fulvestrant in patients with hormone receptor-positive (HR+), human epidermal growth factor receptor 2-negative (HER2-) metastatic breast cancer (MBC) whose disease had progressed on prior endocrine therapy. The present study evaluated the efficacy, safety, and pharmacokinetics of palbociclib plus fulvestrant in Japanese patients enrolled in PALOMA-3. METHODS: Pre/peri/postmenopausal women with HR+/HER2- MBC were randomized 2:1 to fulvestrant (500 mg) and either palbociclib (125 mg/day; 3 weeks on/1 week off; n = 347) or placebo (n = 174). Prespecified exploratory analyses compared the efficacy (data cutoff: October 23, 2015), safety, and pharmacokinetics (data cutoff: December 5, 2014) in Japanese women versus the overall population. RESULTS: A total of 35 Japanese women were randomized to palbociclib-fulvestrant (n = 27) or placebo-fulvestrant (n = 8). Median progression-free survival was 13.6 months (95% CI, 7.5-not estimable) in the Japanese palbociclib-fulvestrant group and 11.2 months (95% CI, 5.6-not estimable) in the placebo-fulvestrant group. The most common adverse event (AE) in Japanese patients was neutropenia (all grades, 93%); no discontinuations were due to an AE. Geometric mean trough concentration values (within-subject mean steady state) for palbociclib were similar for Japanese Asian (excluding Japanese), and non-Asian patients (84.4 ng/mL, 86.3 ng/mL, and 74.8 ng/mL, respectively). CONCLUSION(S): The results for the overall population and Japanese patients in PALOMA-3 suggest that palbociclib plus fulvestrant was effective and well tolerated in Japanese patients with HR+/HER2‒ MBC whose disease had progressed on prior endocrine therapy (Pfizer; NCT01942135).

Palbociclib in combination with letrozole in patients with estrogen receptor-positive, human epidermal growth factor receptor 2-negative advanced breast cancer: PALOMA-2 subgroup analysis of Japanese patients.

BACKGROUND: In PALOMA-2, palbociclib-letrozole significantly improved progression-free survival (PFS) vs placebo-letrozole in women with estrogen receptor-positive, human epidermal growth factor receptor 2-negative (ER+/HER2-) advanced breast cancer (ABC) in the first-line setting. We evaluated the efficacy, safety, and pharmacokinetics of palbociclib in Japanese women in PALOMA-2. METHODS: In this phase 3 study, 666 postmenopausal women with ER+/HER2- ABC were randomized 2:1 to palbociclib (125 mg/day [3 weeks on/1 week off]) plus letrozole (2.5 mg daily) or placebo plus letrozole. A prespecified, exploratory, subgroup analysis of Japanese patients (n = 46) was conducted to compare results with those of the overall population. RESULTS: At the February 26, 2016 cutoff, median PFS among the 46 Japanese patients was 22.2 months (95%CI, 13.6‒not estimable) with palbociclib-letrozole vs 13.8 months (5.6‒22.2) with placebo-letrozole (hazard ratio, 0.59 [95%CI, 0.26-1.34]). The most common adverse events (AEs) were hematologic and more frequent among Japanese patients than the overall population (neutropenia: 93.8% [87.5% grade 3/4] vs 79.5% [66.4%]; leukopenia: 62.5% [43.8%] vs 39.0% [24.8%]); no Japanese patients had febrile neutropenia. Palbociclib dose reductions due to toxicity (mainly neutropenia) were more common in Japanese patients (62.5% vs 36.0%); few permanently discontinued due to AEs. Although mean palbociclib trough concentration was higher in Japanese patients vs non-Asians (95.4 vs 61.7 ng/mL), the range of individual values of the Japanese patients was within that of non-Asians. CONCLUSIONS: These results from PALOMA-2 suggest that palbociclib-letrozole merits consideration as a first-line treatment option for postmenopausal Japanese patients with ER+/HER2‒ ABC. ClinicalTrials.gov: NCT01740427.

Palbociclib in combination with letrozole as first-line treatment for advanced breast cancer: A Japanese phase II study.

This single-arm, open-label, phase II study in 42 Japanese postmenopausal patients with estrogen receptor-positive/human epidermal growth factor receptor 2-negative (ER+/HER2-) advanced breast cancer evaluated the efficacy, safety, and pharmacokinetics of first-line palbociclib (125 mg once daily, 3 weeks on/1 week off) coadministered with letrozole (2.5 mg once daily). Primary endpoint of investigator-assessed 1-year progression-free survival (PFS) probability was 75.0% (90% CI, 61.3%-84.4%), far surpassing the 40% lower limit of the 90% CI supporting efficacy. Median duration of treatment was 438 days. Among secondary efficacy measures, median PFS was not reached (95% CI, 16.7: not estimable), 17/42 patients (40.5%) had an objective response, 36/42 (85.7%) maintained disease control, and 27/42 (64.3%) remained in follow-up. Median overall survival was not reached, and 1-year survival probability was 92.9% (95% CI, 79.5%-97.6%). Results of intensive pharmacokinetics in a subset of 6 patients showed palbociclib steady-state mean area under the plasma concentration-time curve over the dosing interval [τ] and mean maximum plasma concentration were 1979 ng·h/mL and 124.7 ng/mL, respectively. For day 15 plasma samples from cycles 1 and 2, geometric mean of the within-patient mean trough concentration was 90.1 ng/mL. The most common treatment-related adverse events were neutropenia (100%) and stomatitis (73.8%). There was 1 case of treatment-related febrile neutropenia. Toxicities were generally tolerated and manageable by dose modifications and/or medical care. Efficacy and safety of first-line palbociclib plus letrozole therapy is supported in Japanese postmenopausal patients with treatment-naive ER+/HER2- advanced breast cancer.

Phase I study of palbociclib, a cyclin-dependent kinase 4/6 inhibitor, in Japanese patients.

This phase I study in Japanese patients evaluated the safety, pharmacokinetics, and preliminary efficacy of palbociclib, a highly selective and reversible oral cyclin-dependent kinase 4/6 inhibitor, as monotherapy for solid tumors (part 1) and combined with letrozole as first-line treatment of postmenopausal patients with estrogen receptor-positive, human epidermal growth factor receptor 2-negative advanced breast cancer (part 2). Part 1 evaluated palbociclib 100 and 125 mg once daily (3 weeks on/1 week off; n = 6 each group) to determine the maximum tolerated dose. Part 2 evaluated palbociclib maximum tolerated dose (125 mg) plus letrozole 2.5 mg (n = 6). The most common treatment-related adverse event was neutropenia (all grades/grade 3/4): 100 mg, 83%/67%; 125 mg, 67%/33%; and palbociclib plus letrozole, 100%/83%. Heavier pretreatment with chemotherapy may have resulted in higher neutropenia rates observed with the 100-mg dose. Palbociclib exposure was higher with 125 vs 100 mg (mean area under the plasma concentration-time curve over dosing interval [τ]: 1322 vs 547.5 ng·h/mL [single dose], 2838 vs 1276 ng·h/mL [multiple dose]; mean maximum plasma concentration: 104.1 vs 41.4 ng/mL [single dose], 185.5 vs 77.4 ng/mL [multiple dose]). Half-life was 23-26 h. No drug-drug interactions between palbociclib and letrozole occurred. Four patients had stable disease (≥24 weeks in one patient with rectal cancer [100 mg] and one with esophageal cancer [125 mg]) in part 1; two patients had partial response and two had stable disease (both ≥24 weeks) in part 2. Palbociclib at the 125-mg dose (schedule 3/1) was tolerated and is the recommended dose for monotherapy and letrozole combination therapy in Japanese patients. The trials are registered with www.ClinicalTrials.gov: A5481010 and NCT01684215.

Increased de novo riboflavin synthesis and hydrolysis of FMN are involved in riboflavin secretion from Hyoscyamus albus hairy roots under iron deficiency.

Riboflavin secretion by Hyoscyamus albus hairy roots under Fe deficiency was examined to determine where riboflavin is produced and whether production occurs via an enhancement of riboflavin biosynthesis or a stimulation of flavin mononucleotide (FMN) hydrolysis. Confocal fluorescent microscopy showed that riboflavin was mainly localized in the epidermis and cortex of the root tip and, at the cellular level, in the apoplast. The expressions of three genes involved in the de novo biosynthesis of riboflavin (GTP cyclohydrolase II/3,4-dihydroxy-2-butanone 4-phosphate synthase; 6,7-dimethyl-8-ribityllumazine synthase; riboflavin synthase) were compared between Fe-starved and Fe-replete roots over a time-course of 7 days, using RT-PCR. All three genes were found to be highly expressed over the period 1-7 days in the roots cultured under Fe deficiency. Since riboflavin secretion began to be detected only from 3 days, there was a lag phase observed between the increased transcript accumulations and riboflavin secretion. To determine whether FMN hydrolysis might contribute to the riboflavin secretion in Fe-deficient root cultures, FMN hydrolase activity was determined and was found to be substantially increased after 3 days, when riboflavin secretion became detectable. These results suggested that not only de novo riboflavin synthesis but also the hydrolysis of FMN contributes to riboflavin secretion under conditions of Fe deficiency. Respiration activity was assayed during the time-course, and was also found to be enhanced after 3 days under Fe deficiency, suggesting a possible link with riboflavin secretion. On the other hand, several respiratory inhibitors were found not to affect riboflavin synthase transcript accumulation.

Resolution of vocal fold polyps with conservative treatment.

OBJECTIVES: Vocal fold polyp is generally thought to require surgical removal. However, a certain proportion of polyps resolve with conservative treatment. This study was performed to clarify the frequency of spontaneous resolution of vocal fold polyp and identify features associated with polyps that are likely to resolve without surgery. STUDY DESIGN: Retrospective study. METHODS: A review of the medical records of patients diagnosed with vocal fold polyps in Tokyo Voice Center from January 2001 to December 2008. RESULTS: Of 644 patients with the diagnosis of vocal fold polyp, 132 received conservative treatment, 433 were treated surgically, and 79 dropped out without attending for further consultation after the initial visit. Of those treated conservatively, 55 experienced complete resolution after a mean of 5.1 months of follow-up from the outset, and 29 showed lesion shrinkage after a mean of 4.1 months of follow-up. Polyps that resolved with conservative therapy were more likely than those that remained unchanged or enlarged to occur in women, be smaller, and have a shorter duration of symptoms. We could not determine the superiority of voice therapy. CONCLUSIONS: At least 9.7% of vocal fold polyps might resolve without surgery. Conservative treatment should be considered as an option for selected patients with smaller and more recent-onset polyps.

Iron deficiency induces changes in riboflavin secretion and the mitochondrial electron transport chain in hairy roots of Hyoscyamus albus.

Hyoscyamus albus hairy roots secrete riboflavin under Fe-deficient conditions. To determine whether this secretion was linked to an enhancement of respiration, both riboflavin secretion and the reduction of 2,3,5-triphenyltetrazolium chloride (TTC), as a measure of respiration activity, were determined in hairy roots cultured under Fe-deficient and Fe-replete conditions, with or without aeration. Appreciable TTC-reducing activity was detected at the root tips, at the bases of lateral roots and in internal tissues, notably the vascular system. TTC-reducing activity increased under Fe deficiency and this increase occurred in concert with riboflavin secretion and was more apparent under aeration. Riboflavin secretion was not apparent under Fe-replete conditions. In order to examine which elements of the mitochondrial electron transport chain might be involved, the effects of the respiratory inhibitors, barbiturate, dicoumarol, malonic acid, antimycin, KCN and salicylhydroxamic acid (SHAM) were investigated. Under Fe-deficient conditions, malonic acid affected neither root growth, TTC-reducing activity nor riboflavin secretion, whereas barbiturate and SHAM inhibited only root growth and TTC-reducing activity, respectively, and the other compounds variously inhibited growth and TTC-reducing activity. Riboflavin secretion was decreased, in concert with TTC-reducing activity, by dicoumarol, antimycin and KCN, but not by SHAM. In Fe-replete roots, all inhibitors which reduced riboflavin secretion in Fe-deficient roots showed somewhat different effects: notably, antimycin and KCN did not significantly inhibit TTC-reducing activity and the inhibition by dicoumarol was much weaker in Fe-replete roots. Combined treatment with KCN and SHAM also revealed that Fe-deficient and Fe-replete roots reduced TTC in different ways. A decrease in the Fe content of mitochondria in Fe-deficient roots was confirmed. Overall, the results suggest that, under conditions of Fe deficiency in H. albus hairy roots, the alternative NAD(P)H dehydrogenases, complex III and complex IV, but not the alternative oxidase, are actively involved both in respiration and in riboflavin secretion.

[Clinical experience with silicon pharyngeal tube for pharyngocutaneous fistula and cervical esophagus stenosis].

Total laryngectomy or laryngopharyngectomy are commonly performed for the treatment of laryngeal cancer or hypopharyngeal cancer. However pharyngocutaneous fistula and cervical esophageal stenosis have been reported as postoperative complications of these procedures. We used a silicon pharyngeal tube in cases that developed pharyngocutaneous fistula and cervical esophageal stenosis. The pharyngeal tube was useful for controlling aspiration pneumonia and for starting oral feeding in a case of pharyngocutaneous fistula after a total laryngectomy. It was also helpful for starting oral feeding in a case with cervical esophageal stenosis after total laryngopharyngectomy and free jejunum interposition. This patient was able to maintain a good quality of life until re-operation. Adverse effects from the insertion of the tube included a foreign body sensation and pharyngeal pain that was tolerable with the use of NSAIDs for a short time. Silicon pharyngeal tubes are useful for the treatment of pharyngocutaneous fistula and cervical esophageal stenosis.

Adenoviral gene transfer of BDNF and GDNF synergistically prevent motoneuron loss in the nucleus ambiguus.

We have previously shown that neuroprotective effects of an adenoviral glial cell line-derived neurotrophic factor (GDNF) gene transfer on the lesioned adult rat motoneurons in the nucleus ambiguus. In the present study, we examined neuroprotective effects of adenoviral gene transfer of brain-derived neurotrophic factor (BDNF) or/and GDNF to motoneurons in nucleus ambiguus using an adult rat vagal nerve avulsion model. The animals avulsed and inoculated with adenoviral vectors encoding BDNF (AxCAmBDNFME) or/and GDNF (AxCAhGDNF) showed immunolabeling for BDNF or/and GDNF in the nucleus ambiguus on the treated side, respectively, and expression of virus-induced BDNF or/and GDNF mRNA transcripts in the brainstem tissue that contained the nucleus ambiguus of the treated side. The treatment with AxCAhGDNF or AxCAmBDNFME significantly prevented the loss of vagal motoneurons in comparison to the control; the protective effect of AxCAmBDNFME was greater than that of AxCAhGDNF. The combined treatment with AxCAmBDNFME and AxCAhGDNF acted synergistically and significantly larger number of vagal motoneurons was preserved as compared to either AxCAmBDNFME treatment or AxCAhGDNF treatment. The treatment with AxCAmBDNFME or/and AxCAhGDNF after avulsion also suppressed the activity of nitric oxide synthase in lesioned motoneurons in the nucleus ambiguus. These results indicate that adenovirus-mediated BDNF and GDNF gene transfer may prevent the degeneration of motoneurons in humans after either vagal nerve injury or recurrent laryngeal nerve injury.

Influence of genetic polymorphisms of styrene-metabolizing enzymes and smoking habits on levels of urinary metabolites after occupational exposure to styrene.

Here we evaluate the influence of individual genetic polymorphisms of drug-metabolizing enzymes as well as body mass index (BMI) and lifestyle (smoking, alcohol consumption) on urinary metabolites after occupational exposure to styrene. Seventy-three workers exposed to styrene in a reinforced-plastics workplace were studied. The personal styrene exposure in the air and the urinary styrene metabolites mandelic acid and phenylglyoxylic acid were measured. The subjects' genetic polymorphisms in the genes that encode the styrene-metabolizing enzymes CYP2E1, CYP2B6, EPHX1, GSTM1, GSTT1 and GSTP1 were determined. Neither genotype nor lifestyle significantly affected urinary metabolites. There was, however, an interaction between the CYP2E1 genotype and smoking. Among non-smokers, urinary styrene metabolites were significantly decreased in subjects with c1/c1 alleles of CYP2E1 as compared with those with the c1/c2 genotype. There was no significant difference in urinary metabolites among smokers. When the combined influence of the CYP2B6 genotype and the predicted activity of EPHX1 were examined, urinary metabolites in subjects with low enzyme activity were lower than in those with medium or high activity after high styrene exposure (>or=50 ppm). The results suggest that genetic susceptibility and lifestyle should be considered in biological monitoring of exposure to styrene.

Abnormal beta-catenin expression in oral cancer with no gene mutation: correlation with expression of cyclin D1 and epidermal growth factor receptor, Ki-67 labeling index, and clinicopathological features.

Beta-Catenin not only acts as a regulator of E-cadherin-mediated cell-cell adhesion but also plays an important role in Wnt signaling. To assess the prevalence of Wnt signaling, we examined beta-catenin mutation and its immunohistochemical protein expression in oral cancers. The results were linked with expression of cyclin D1, one of the target genes of Wnt signaling, expression of epidermal growth factor receptor (EGFR) relevant to beta-catenin tyrosine phosphorylation, Ki-67 labeling index, clinicopathological features, and survival. In the analysis based on membranous expression of beta-catenin, 75 (68.2%) of 110 cases showed a reduced membranous pattern, and the remaining 35 (31.8%) had a preserved membranous pattern similar to that in oral epithelium. In the analysis of another category of beta-catenin expression, a cytoplasmic/nuclear pattern was observed in 21 (19.1%) of the 110 tumors. Most (19/21, 90.5%) of these tumors had a concomitant reduction of membranous expression of beta-catenin. The reduced membranous or cytoplasmic/nuclear pattern of beta-catenin was significantly associated with an invasive growth pattern, EGFR expression, an increased Ki-67 labeling index, and shorter survival but not with cyclin D1 expression. Mutational analyses of beta-catenin were performed for 39 cases, including the 21 tumors with a cytoplasmic/nuclear pattern, but no mutations in the beta-catenin gene exon 3 were detected in these samples. Our data indicate that altered expression of beta-catenin may play an important role in tumor progression through increased proliferation and invasiveness under EGFR activation. However, mutations of beta-catenin do not appear to be responsible for tumor development and abnormal expression of beta-catenin in oral cancers.