Semaphorin signaling via MICAL3 induces symmetric cell division to expand breast cancer stem-like cells.

Cancer stem-like cells (CSCs) are expanded in the CSC niche by increased frequency of symmetric cell divisions at the expense of asymmetric cell divisions. The symmetric division of CSCs is important for the malignant properties of cancer; however, underlying molecular mechanisms remain largely elusive. Here, we show a cytokine, semaphorin 3 (Sema3), produced from the CSC niche, induces symmetric divisions of CSCs to expand the CSC population. Our findings indicate that stimulation with Sema3 induced sphere formation in breast cancer cells through neuropilin 1 (NP1) receptor that was specifically expressed in breast CSCs (BCSCs). Knockdown of MICAL3, a cytoplasmic Sema3 signal transducer, greatly decreased tumor sphere formation and tumor-initiating activity. Mechanistically, Sema3 induced interaction among MICAL3, collapsin response mediator protein 2 (CRMP2), and Numb. It appears that activity of MICAL3 monooxygenase (MO) stimulated by Sema3 is required for tumor sphere formation, interaction between CRMP2 and Numb, and accumulation of Numb protein. We found that knockdown of CRMP2 or Numb significantly decreased tumor sphere formation. Moreover, MICAL3 knockdown significantly decreased Sema3-induced symmetric divisions in NP1/Numb-positive BCSCs and increased asymmetric division that produces NP1/Numb negative cells without stem-like properties. In addition, breast cancer patients with NP1-positive cancer tissues show poor prognosis. Therefore, the niche factor Sema3-stimulated NP1/MICAL3/CRMP2/Numb axis appears to expand CSCs at least partly through increased frequency of MICAL3-mediated symmetric division of CSCs.

ErbB receptor tyrosine kinase/NF-κB signaling controls mammosphere formation in human breast cancer.

Breast cancer is one of the most common cancers in humans. However, our understanding of the cellular and molecular mechanisms underlying tumorigenesis in breast tissues is limited. Here, we identified a molecular mechanism that controls the ability of breast cancer cells to form multicellular spheroids (mammospheres). We found that heregulin (HRG), a ligand for ErbB3, induced mammosphere formation of a breast cancer stem cell (BCSC)-enriched population as well as in breast cancer cell lines. HRG-induced mammosphere formation was reduced by treatment with inhibitors for phosphatidyl inositol 3-kinase (PI3K) or NF-κB and by expression of IκBα-Super Repressor (IκBαSR), a dominant-negative inhibitor for NF-κB. Moreover, the overexpression of IκBαSR in breast cancer cells inhibited tumorigenesis in NOD/SCID mice. Furthermore, we found that the expression of IL8, a regulator of self-renewal in BCSC-enriched populations, was induced by HRG through the activation of the PI3K/NF-κB pathway. These findings illustrate that HRG/ErbB3 signaling appears to maintain mammosphere formation through a PI3K/NF-κB pathway in human breast cancer.

Effectiveness and safety of tegafur-gimeracil-oteracil potassium (TS-1) for metastatic breast cancer: a single-center retrospective study.

BACKGROUND: Tegafur-gimeracil-oteracil potassium (TS-1)is a drug that is used mainly as a third-line treatment or beyond for metastatic breast cancer(MBC). However, there is still insufficient evidence on its clinical effectiveness, and there are very few reports on clinical research using TS-1 up front. In this report, we examined the effectiveness and safety of TS-1 therapy for MBC. PATIENTS AND METHODS: The subjects were 46 patients with MBC who were treated with TS-1 between January 2005 and January 2013. These patients were retrospectively examined. RESULTS: The objective response rate to TS-1 therapy was 30.4%, clinical benefit rate (CBR)was 50.0%, and the median time to treatment failure was 10.7 months. When examined by site, the CBR was high locally (46.2%), in the lymph nodes (40.7%), in the bone (42.9%), and in the lungs and pleura (44.8%). However it was low in the liver(30.0%). The relationship was examined between clinicopathological factors and the effectiveness of TS-1 therapy. The objective response rate (ORR) was significantly higher for patients with disease-free interval (DFI) of 2 years or more (p=0.039), TS-1 therapy used as third-line treatment or earlier (p=0.022), negative HER2 status (p=0.020), and no history of capecitabine (CAP)therapy (p=0.049). The CBR was significantly higher for patients with no visceral metastasis (p=0.032), TS-1 used as third-line treatment or earlier (p=0.019), negative HER2 status (p= 0.045), no history of CAP therapy (p=0.006), and no history of tegafur-uracil/doxifluridine therapy (p=0.031). Multivariate analysis showed that DFI of 2 years or more (p=0.035, odds ratio:0.104)was an independent predictor of effectiveness assessed by ORR. There were only 4 patients in whom the treatment was discontinued due to adverse event, and TS-1 was generally well tolerated. CONCLUSION: TS-1 was highly effective and well tolerated by patients with MBC. Its up-front use might enable the maintenance of satisfactory QOL and the enhancement of its clinical effectiveness.

Efficacy of high-dose toremifene therapy in postmenopausal patients with metastatic breast cancer resistant to aromatase inhibitors:a retrospective, single-institution study.

BACKGROUND: Aromatase inhibitors(AI)have established efficacy as first-line therapy in postmenopausal patients with hormone-sensitive metastatic breast cancer(MBC). However,the use of endocrine therapy has not yet been established for second-line and later therapy. Our study examined the efficacy of high-dose toremifene therapy(HD-TOR)in patients with MBC resistant to AIs. PATIENTS AND METHODS: A retrospective analysis was carried out to determine outcomes in 85 postmenopausal patients with MBC resistant to AIs who began HD-TOR between May 2001 and October 2011. The patients received toremifene 120 mg once daily on consecutive days. RESULTS: The objective response rate(ORR)was 21.2%,the clinical benefit rate(CBR)was 41.2%,and the median time to treatment failure(TTF)was 7.3 months. The CBR was high in patients with ER-positive status(p=0.045),no visceral metastasis(p=0.037),HD -TOR as first- or second-line therapy(p=0.007),no history of tamoxifen(TAM)therapy(p=0.019),and no history of chemotherapy(p=0.017). Multivariate analysis showed that ER-positive status(p=0.005, odds ratio: 0.064)and no visceral metastasis(p=0.034, odds ratio: 0.323)were independent predictors of efficacy. The TTF was significantly longer in patients with ER-positive status(p=0.019)and no history of TAM therapy(p=0.015). Multivariate analysis showed that ER-positive status(p=0.025, hazard ratio: 0.377)and no history of TAM therapy(p=0.002, hazard ratio: 0.422)were independent predictors of efficacy. No patient discontinued HDTOR therapy due to adverse events. CONCLUSION: HD-TOR is an effective endocrine therapy for patients with MBC who have failed AIs.

A case of breast cancer in the axillary tail of Spence - enhanced magnetic resonance imaging and positron emission tomography for diagnostic differentiation and preoperative treatment decision.

BACKGROUND: The management of cancer in the axillary area depends on the etiology of the tumor. CASE REPORT: A 37-year-old woman presented with a 2 cm mass in the axillary fossa. Core needle biopsy revealed adenocarcinoma. There were no abnormal breast findings on physical examination, mammography, or ultrasonography. However, enhanced magnetic resonance imaging (MRI) and positron emission tomography (PET) showed a segmentally-distributed, abnormal area in the upper-outer quadrant, continuous with the axillary mass. Samples of this area obtained by vacuum-assisted biopsy showed intraductal carcinoma. These findings indicated that the axillary lesion was a part of primary breast cancer originating from the axillary tail. Based on these results, the patient underwent total mastectomy with sentinel lymph node biopsy. Pathological examination of the specimen showed invasive ductal carcinoma accompanied by intraductal carcinoma extending up to 8.5 cm. Our case suggests that enhanced MRI and PET can provide useful preoperative information for the management of axillary breast lesions.

Crystal structures of the apo and holo form of rat catechol-O-methyltransferase.

Catechol-O-methyltransferase (COMT, EC 2.1.1.6) is a monomeric enzyme that catalyzes the transfer of a methyl group from S-adenosyl-l-methionine (AdoMet) to the phenolic oxygen of substituted catechols. Although the inhibitor recognition pattern and AdoMet site have already been studied crystallographically, structural information on the catalytic cycle of COMT has not yet been obtained. In this study, comparison of the co-factor and inhibitor-bound structures revealed that the Apo form of COMT shows a conformational change and there was no cleft corresponding to the AdoMet-binding site; the overall structure was partially open form and the substrate recognition site was not clearly defined. The Holo form of COMT was similar to the quaternary structure except for the beta6-beta7 and alpha2-alpha3 ligand recognition loops. These conformational changes provide a deeper insight into the structural events occurring in reactions catalyzed by AdoMet.

Crystal structures of rat catechol-O-methyltransferase complexed with coumarine-based inhibitor.

In human, catechol-O-methyltransferase (COMT: E.C. 2.1.1.6) is responsible for metabolism of catechol neurotransmitter and xenobiotics. The main clinical interest in COMT results from the possibility of using COMT inhibitors as adjuncts in the therapy of Parkinson's disease (PD) with l-DOPA. COMT is therefore a target for inhibitor development aiming at PD treatment and has been submitted to extensive structure-based drug design. Recently reported inhibitors have nitrocatechol structure that may inhibit oxidative phosphorylation and uncouple mitochondrial energy production. This work reports the first crystallographic study of Rat COMT complexed with non-nitrocatechol inhibitor. Analysis of the structural differences among the previously reported inhibitor complexes, coumarine-based inhibitor (4-phenyl-7, 8-dihydroxycoumarine: 4PCM) bound structure provides the explanation for inhibitor binding and can be used for future inhibitor design.

Orally active factor Xa inhibitors: investigation of a novel series of 3-amidinophenylsulfonamide derivatives using an amidoxime prodrug strategy.

A series of novel and potent 3-amidinophenylsulfonamide derivatives of factor Xa inhibitors were designed and synthesized using an amidoxime prodrug strategy. We focused on systemic clearance of parent compounds in rats, and performed in vivo pharmacokinetic screening. Incorporation of a carboxymethoxy group markedly improved systemic clearance (compound 43), and the related amidoxime 44 showed sufficient prodrug conversion. Compound 45, the double prodrug of 43, exhibited practicable bioavailability after oral administration in rats. Among the various compounds under investigation, KFA-1982 was selected for clinical development.

The importance of CH/pi hydrogen bonds in rational drug design: An ab initio fragment molecular orbital study to leukocyte-specific protein tyrosine (LCK) kinase.

The interaction energy was calculated, by the ab initio FMO method, for complexes between LCK protein and four inhibitors (staurosporine, BMS compound 2, and our compounds 3 and 4). In every case a number of CH/pi hydrogen bonds have been disclosed in the so-called adenine pocket. In complexes of 2, 3, and 4, CH/pi and NH/pi hydrogen bonds have been observed in another pocket. In view of the above results, the aniline ring of 3 was replaced by 2,6-dimethyl aniline to increase the potency for LCK kinase. A 10-fold increase in the potency has been achieved for 4 over 3. We suggest that the concept of weak hydrogen bonds is useful in the rational design of drugs.

Structure-activity relationship studies of imidazo[1,2-c]pyrimidine derivatives as potent and orally effective Syk family kinases inhibitors.

Spleen tyrosine kinase (Syk) and zeta-associated protein kinase of 70k Da (ZAP-70) are members of the Syk family and non-receptor-type protein tyrosine kinases, which play crucial roles in B- and T-cell activation. Therefore, a Syk family tyrosine kinases inhibitor would be a useful therapeutic agent for the treatment of various allergic disorders and autoimmune diseases. Previously, we reported that 1,2,4-triazolo[4,3-c]pyrimidine derivative 1 and 1,2,4-triazolo[1,5-c]pyrimidine derivative 2 showed strong inhibitory activities against Syk family kinases. These compounds also exhibited high-level suppression of IL-2 in cellular assays. However, their oral efficacies were poor in a mouse model of IL-2 production. To improve oral effectiveness, we investigated a new series of Syk family kinases inhibitors. We found that imidazo[1,2-c]pyrimidine derivatives potently inhibited the Syk family kinases. Among these agents, compound 9f not only showed strong inhibitory activities against Syk and ZAP-70 kinases in vitro, but its oral administration resulted in the in vivo suppression of both the passive cutaneous anaphylaxis reaction and Concanavalin A-induced IL-2 production in a mouse model.

Preoperative evaluation of thyroid pathology in patients with primary hyperparathyroidism.

In parathyroidectomy, it has been recognized that a shift to a minimally invasive procedure may be accompanied by a possibility of missing thyroid pathology. However, only a few findings concerning preoperative thyroid evaluation have been reported. We investigated the prevalence of concomitant thyroid pathology by preoperative neck ultrasonography (US) in patients with primary hyperparathyroidism. There were 85 patients (66 women, 19 men; mean age 57 years) in the study group. The mean preoperative calcium level was 11.2mg/dL, and the mean intact parathyroid hormone level was 206 pg/mL. All patients underwent neck US following fine-needle aspiration biopsy (FNAB). Of the 85 patients, 21 (24.7%) had thyroid nodules. Among 21 patients with thyroid nodules, 9 (10.6%) had malignant thyroid tumors, while 12 (14.1%) patients had benign thyroid nodules including multinodular goiter. Of the 9 patients with malignant thyroid nodules, 4 had papillary carcinomas with lymph node metastases. The prevalence of thyroid disease associated with hyperparathyroidism is high, and evaluation of the thyroid pathology by US enables the shift from bilateral neck exploration to the minimally invasive parathyroid surgery.

The pathological findings of vasculitis simultaneously occurring with carcinoma, invasive breast carcinoma in a patient with Behçet's disease.

We present a rare case of invasive right breast carcinoma in a 72-year-old woman with Behçet's disease (BD). A radical modified right mastectomy and axillary lymphadenectomy were performed and postoperative hormonal therapy with the aromatase inhibitor anastrozole was administered for adjuvant therapy. At 10 months follow-up the patient remains disease free. Malignancies associated with BD are very uncommon. The pathological findings showed small vessel vasculitis and lobulitis of the breast in association with invasive carcinoma.

Efficacy of octreotide against chylothorax following lateral neck dissection for thyroid cancer: A case report.

INTRODUCTION: Postsurgical chylothorax is a rare complication of cervical dissection for thyroid cancer. We report that octreotide, a synthetic analog of somatostatin, is effective in treating chylothorax after thyroid carcinoma surgery. PRESENTATION OF CASE: The patient was a 48-year-old woman who presented to our institution complaining of a left anterior cervical mass. We diagnosed this as thyroid papillary carcinoma and performed a subtotal excision of the thyroid gland with left cervical lymph node dissection. The patient developed dyspnea, and a chest X-ray revealed bilateral chylothorax on Day 4 post-surgery. Octreotide was administered since bilateral chylothorax was noted. A marked decrease in chyle effusion was noted just 3 days after starting octreotide, and after a total of 9 days of treatment, there were no further signs of chylous effusion. DISCUSSION: Octreotide is effective against postsurgical chylothorax; however, if there are no signs of improvement, we believe surgical treatment should be considered. CONCLUSION: Octreotide should be administered first to treat postsurgical chylothorax before surgical treatment is considered.

Scedosporium prolificans Endocarditis: Case Report and Literature Review.

Scedosporium prolificans, a hyaline filamentous fungus, is widely distributed in the environment and is currently an emerging human pathogen, especially among immunocompromised patients. However, S. prolificans endocarditis is rare. We herein report a case of S. prolificans endocarditis in a 64-year-old patient with breast cancer in complete remission for 30 years after chemotherapy and radiation treatment who was not cured. Susceptibility testing showed resistance to all antifungal drugs, except echinocandin. A review of the literature revealed 10 cases of S. prolificans endocarditis; of these, only one involved an immunocompetent host with no risk factors and only two patients survived. In order to improve the mortality rate, it is necessary to establish rapid diagnostic methods and efficient therapeutic approaches.

Distribution of angiogenin and its gene message in colorectal cancer patients and their clinical relevance.

BACKGROUND: [corrected] The distribution of angiogenin (ANG) in normal colorectal and colorectal cancer tissues has not been precisely elucidated, while studies on the clinical significance of ANG have been scanty at best. PATIENTS AND METHODS: We investigated the distribution of ANG and its gene message in 58 colorectal cancer and 58 corresponding normal colorectal tissue pairs by immunohistochemistry and in situ hybridization, the circulating ANG levels in sera by ELISA from 94 colorectal cancer patients and 52 healthy volunteers, as well as the clinical significance of ANG expression. RESULTS: Strong, moderate and weak ANG immunoreactivity of cancer cells was observed in 22, 31 and 5 patients, respectively. ANG immunoreactivity was also observed in interstitial cells in the vicinity of and at the invasion front of cancer cells, as well as in normal superficial epithelial cells and in some interstitial cells. The distribution of the ANG gene message was coherent with the ANG protein localization. The mean serum ANG concentration was significantly (p =0.00005) higher in cancer patients as compared with that in the healthy volunteers and the degree of its elevation was more significant in the more progressed cancer subgroups. The degrees of ANG protein and mRNA expression correlated with serum ANG concentration (p<0.05). Survival analyses revealed a significant correlation between higher serum ANG concentrations (> or = 400ng/ml) and worse disease-free (p=0.003) or disease-specific (p=0.03) survivals. CONCLUSION: These results suggest that ANG is expressed in colorectal cancer cells and in normal colorectal superficial cells as well as in interstitial cells and that serum levels of ANG reflect the degree of tissue ANG expression that could be a predictor of postoperative outcome.

[An adult case of hypoplasia of the left lung disclosed by pneumonectomy for pulmonary M. avium complex infection associated with intractable pneumothorax].

A case of W-P-W syndrome complicated with pulmonary hypoplasia disclosed by pneumonectomy for pulmonary Mycobacterium avium complex infection associated with intractable pneumothorax was reported. A male patient aged 52 years consulted our clinic with chief complaints of cough and abnormal shadows on his chest radiogram, which was consistent with mycobacteriosis on his left lung. MAC infection was soon confirmed by sputum examination and he was treated with RFP, EB, INH combined with CAM. In spite of the chemotherapy, sputum examination of the patient remained positive. Furthermore, eleven months after initiating the treatment, an intractable pneumothorax concurrent with a large dead space at the left lower lung field was consistently observed on his chest radiogram. Therefore, he was first treated by video assisted thoracoscopic surgery, but soon relapsed which led to tention pneumothorax gradually. Consequently, a left pneumonectomy had to be performed and the following developmental abnormalities combined with pathological changes caused by MAC infection were disclosed: concerning the upper lobe, defect of lingula, formation of a peripheral type of congenital air-filled parenchymal cyst measuring 5 x 6 cm in S3, and atelectatic induration caused by MAC infection on the remaining part of the upper lobe where strong adhesion was seen between the chest wall and the lung. Concerning the lower lobe, congenital shortening of visceral pleura, mainly mediastinal surface, causing marked deformity of the lower lobe with elevation of margo inferior. This created a large dead space between the lower lobe and diaphragma, and formation of a walnut-sized nest of atelectatic induration caused by MAC infection in S6. The patient's post-operative clinical course was uneventful and his arterial blood gas was elevated from 76 torr to 99.2 torr. He was discharged three weeks after the operation. Several controversial issues relating to this case were discussed; the predisposition existing on the hypoplastic lung to MAC infection, the possible reason why the congenital pulmonary cyst was not involved in MAC infection, the location of perforation of the upper lobe that caused intractable pneumothorax, and the difficulty in diagnosing congenital air-filled bullous parenchymal cyst by current conventional chest radiogram.

Hepatotoxicity induced by trastuzumab used for breast cancer adjuvant therapy: a case report.

INTRODUCTION: Trastuzumab is generally considered a highly safe drug, but there have been cases of infusion reaction and cardiotoxicity. This report will present a rare case of hepatotoxicity induced by trastuzumab used for adjuvant therapy of human epidermal growth factor receptor type 2-positive breast cancer. CASE PRESENTATION: The patient was a 60-year-old Japanese postmenopausal woman with a non-contributory past medical history. She presented for detailed examination of an abnormality in her left breast. She had left breast cancer (T2N1M0, stage IIB) that was positive for estrogen receptor and progesterone receptor and was human epidermal growth factor receptor type 2 3+. She began receiving epirubicin and cyclophosphamide therapy but developed hepatotoxicity (aspartate aminotransferase 43 U/L, alanine aminotransferase 104 U/L, alkaline phosphatase 634 U/L, and γ-glutamyl transpeptidase 383 U/L). Thus, the therapy was discontinued after two cycles, and a weekly paclitaxel therapy was begun. After the absence of an adverse event was confirmed, she also began receiving trastuzumab (4 mg/kg) at the second cycle. However, hepatotoxicity (aspartate aminotransferase 267 U/L, alanine aminotransferase 246 U/L, alkaline phosphatase 553 U/L, and γ-glutamyl transpeptidase 240 U/L) developed again, and trastuzumab was discontinued. She received paclitaxel monotherapy for a total of four cycles and subsequently underwent partial mastectomy and axillary dissection. After completing adjuvant radiation therapy (breast, 50 Gy), she received trastuzumab administration (4 mg/kg) but hepatotoxicity (aspartate aminotransferase 47 U/L, alanine aminotransferase 102 U/L, alkaline phosphatase 377 U/L, and γ-glutamyl transpeptidase 91 U/L) recurred. Thus, it was discontinued again. There was no hepatitis B or C virus infection, and a drug-induced lymphocyte stimulation test revealed a positive reaction to trastuzumab (stimulation index: 227%). Thereafter she has used only oral letrozole (2.5mg/day) and no recurrent cancer has been observed. CONCLUSIONS: Although trastuzumab is a highly safe drug, one must be mindful of its risk for hepatotoxicity. Periodic monitoring of liver functions is necessary during trastuzumab therapy.

Aberrantly methylated genes in human papillary thyroid cancer and their association with BRAF/RAS mutation.

Cancer arises through accumulation of epigenetic and genetic alteration. Aberrant promoter methylation is a common epigenetic mechanism of gene silencing in cancer cells. We here performed genome-wide analysis of DNA methylation of promoter regions by Infinium HumanMethylation27 BeadChip, using 14 clinical papillary thyroid cancer samples and 10 normal thyroid samples. Among the 14 papillary cancer cases, 11 showed frequent aberrant methylation, but the other three cases showed no aberrant methylation at all. Distribution of the hypermethylation among cancer samples was non-random, which implied existence of a subset of preferentially methylated papillary thyroid cancer. Among 25 frequently methylated genes, methylation status of six genes (HIST1H3J, POU4F2, SHOX2, PHKG2, TLX3, HOXA7) was validated quantitatively by pyrosequencing. Epigenetic silencing of these genes in methylated papillary thyroid cancer cell lines was confirmed by gene re-expression following treatment with 5-aza-2'-deoxycytidine and trichostatin A, and detected by real-time RT-PCR. Methylation of these six genes was validated by analysis of additional 20 papillary thyroid cancer and 10 normal samples. Among the 34 cancer samples in total, 26 cancer samples with preferential methylation were significantly associated with mutation of BRAF/RAS oncogene (P = 0.04, Fisher's exact test). Thus, we identified new genes with frequent epigenetic hypermethylation in papillary thyroid cancer, two subsets of either preferentially methylated or hardly methylated papillary thyroid cancer, with a concomitant occurrence of oncogene mutation and gene methylation. These hypermethylated genes may constitute potential biomarkers for papillary thyroid cancer.