Junji Matsuoka

Gene Pathways Associated With Prognosis and Chemotherapy Sensitivity in Molecular Subtypes of Breast Cancer

BACKGROUND We hypothesized that distinct biological processes might be associated with prognosis and chemotherapy sensitivity in the different types of breast cancers. METHODS We performed gene set analyses with BRB-ArrayTools statistical software including 2331 functionally annotated gene sets (ie, lists of genes that correspond to a particular biological pathway or biochemical function) assembled from Ingenuity Pathway Analysis and Gene Ontology databases corresponding to almost all known biological processes. Gene set analysis was performed on gene expression data from three cohorts of 234, 170, and 175 patients with HER2-normal lymph node-negative breast cancer who received no systemic adjuvant therapy to identify gene sets associated prognosis and three additional cohorts of 198, 85, and 62 patients with HER2-normal stage I-III breast cancer who received preoperative chemotherapy to identify gene sets associated with pathological complete response to therapy. These analyses were performed separately for estrogen receptor (ER)-positive and ER-negative breast cancers. Interaction between gene sets and survival and treatment response by breast cancer subtype was assessed in individual datasets and also in pooled datasets. Statistical significance was estimated with permutation test. All statistical tests were two-sided. RESULTS For ER-positive cancers, from 370 to 434 gene sets were associated with prognosis (P ≤ .05) and from 209 to 267 gene sets were associated with chemotherapy response in analysis by individual dataset. For ER-positive cancers, 131 gene sets were associated with prognosis and 69 were associated with pathological complete response (P ≤.001) in pooled analysis. Increased expression of cell cycle-related gene sets was associated with poor prognosis, and B-cell immunity-related gene sets were associated with good prognosis. For ER-negative cancers, from 175 to 288 gene sets were associated with prognosis and from 212 to 285 gene sets were associated with chemotherapy response. In pooled analyses of ER-negative cancers, 14 gene sets were associated with prognosis and 23 were associated with response. Gene sets involved in sphingolipid and glycolipid metabolism were associated with better prognosis and those involved in base excision repair, cell aging, and spindle microtubule regulation were associated with chemotherapy response. CONCLUSION Different biological processes were associated with prognosis and chemotherapy response in ER-positive and ER-negative breast cancers.

Estrogen Receptor (ER) mRNA and ER-Related Gene Expression in Breast Cancers That Are 1% to 10% ER-Positive by Immunohistochemistry

PURPOSE We examined borderline estrogen receptor (ER) -positive cancers, defined as having 1% to 10% positivity by immunohistochemistry (IHC), to determine whether they show the same global gene-expression pattern and high ESR1 mRNA expression as ER-positive cancers or if they are more similar to ER-negative cancers. PATIENTS AND METHODS ER status was determined by IHC in 465 primary breast cancers and with the Affymetrix U133A gene chip. We compared expressions of ESR1 mRNA and a 106 probe set ER-associated gene signature score between ER-negative (n = 183), 1% to 9% (n = 25), 10% (n = 6), and more than 10% (n = 251) ER-positive cancers. We also assessed the molecular class by using the PAM50 classifier and plotted survival by ER status. RESULTS Among the 1% to 9%, 10%, and more than 10% ER IHC-positive patients, 24%, 67%, and 92% were also positive by ESR1 mRNA expression. The average ESR1 expression was significantly higher in the ≥ 10% ER-positive cohorts compared with the 1% to 9% or ER-negative cohort. The average ER gene signature scores were similar for the ER-negative and 1% to 9% IHC-positive patients and were significantly lower than in ≥ 10% ER-positive patients. Among the 1% to 9% ER-positive patients, 8% were luminal B and 48% were basal-like; among the 10% ER-positive patients, 50% were luminal. The overall survival rate of 1% to 9% ER-positive patients with cancer was between those of patients in the ≥ 10% ER-positive and ER-negative groups. CONCLUSION A minority of the 1% to 9% IHC ER-positive tumors show molecular features similar to those of ER-positive, potentially endocrine-sensitive tumors, whereas most show ER-negative, basal-like molecular characteristics. The safest clinical approach may be to use both adjuvant endocrine therapy and chemotherapy in this rare subset of patients.

Antibacterial Properties of Antimicrobial-Finished Textile Products

The antibacterial properties of five kinds of antimicrobial‐finished textile products (AFTPs) were examined against Staphylococcus aureus, including methicillin‐resistant (MRSA) strains and Pseudomonas aeruginosa, under wet and dry conditions. Textile products containing Ag. Zn. ammonium Zeolite and chitosan were found to be effective against methicillin‐sensitive S. aureus (MSSA) for up to 6 hr of incubation under wet and dry conditions, and effective against MRSA for up to 24 hr of incubation only under wet conditions. Under dry conditions, however, all AFTPs were ineffective against one MRSA strain. When organic matter was added to the incubation mixture, textile products containing Ag. Zn. ammonium Zeolite and chitosan still showed antibacterial activities, but not as strongly. The results of this study suggested the following: (1) There are differences in antibacterial properties among commercially available AFTPs; (2) Determining effectiveness requires several hours of incubation; (3) Water content as an environmental factor can affect effectiveness; and (4) Some bacterial species and strains are not affected by AFTPs. The antibacterial properties of AFTPs in the clinical setting may be of limited value.

Inhibition of inducible NF-κB activity reduces chemoresistance to 5-fluorouracil in human stomach cancer cell line

5-fluorouracil (5-FU) is used for the treatment of stomach and colon cancer, but many tumors are resistant to this chemotherapeutic agent. 5-FU induces apoptosis of several cancer cell lines, while some chemotherapeutic agents are known to activate the transcriptional factor NF-kappaB, which strongly suppresses apoptosis in vitro. In the present study, we investigated the relationship between activation of NF-kappaB and chemoresistance to 5-FU in human stomach cancer cell lines, NUGC3 (5-FU sensitive) and NUGC3/5FU/L (5-FU resistant). Treatment with 5-FU for 9-12 h caused activation of inducible NF-kappaB in NUGC3/5FU/L cells but not in NUGC3 cells. 5-FU also resulted in an increase in the number of TUNEL-positive cells and enhanced caspase-3 activity 3- to 5-fold in NUGC3 cells but not NUGC3/5FU/L cells. Moreover we also demonstrated that the inhibition of inducible NF-kappaB activation by using a NF-kappaB decoy could induce apoptosis and reduce chemoresistance against 5-FU. Our results suggest that 5-FU chemoresistance can be overcome by inhibition of inducible NF-kappaB activation, and that the use of the NF-kappaB decoy combined with 5-FU treatment is a new molecular and gene therapeutic strategy aimed at treatment of human stomach cancers resistant to 5-FU.

Localization of heparanase in esophageal cancer cells: Respective roles in prognosis and differentiation

In this study, we examined the distribution of heparanase protein in 75 esophageal squamous cell carcinomas by immunohistochemistry and analyzed the relationship between heparanase expression and clinicopathological characteristics. In situ hybridization showed that the mRNA expression pattern of heparanase was similar to that of the protein, suggesting that increased expression of the heparanase protein at the invasive front was caused by an increase of heparanase mRNA in tumor cells. Heparanase expression correlated significantly with depth of tumor invasion, lymph node metastasis, tumor node metastasis (TNM) stage and lymphatic invasion. Overexpression of heparanase in esophageal cancers was also associated with poor survival. In addition to its localization in the cytoplasm and cell membrane, heparanase was also identified in the nuclei of normal epithelial and tumor cells by immunohistochemistry. Furthermore, nuclear heparanase was detected in nuclear extract of cancer cell lines by Western blot and immunohistochemistry. Examination of the role of nuclear heparanase in cell proliferation and differentiation by double immunostaining for proliferating cell nuclear antigen (PCNA) and cytokeratin 10 (CK10) showed significant relationship between nuclear heparanase expression and differentiation (heparanase vs CK10), but not for proliferative state of esophageal cancer cells (heparanase vs PCNA). Our results suggest that cytoplasmic heparanase appears to be a useful prognostic marker in patients with esophageal cancer and that nuclear heparanase protein may play a role in differentiation. Inhibition of heparanase activity may be effective in the control of esophageal tumor invasion and metastasis.

Inhibition of focal adhesion kinase as a potential therapeutic strategy for imatinib-resistant gastrointestinal stromal tumor

Focal adhesion kinase (FAK) is often up-regulated in a variety of malignancies, including gastrointestinal stromal tumor (GIST), and its overexpression seems to be associated with tumor progressiveness and poor prognosis. GIST is well known to have a mutation to c-KIT; thus, a specific c-KIT inhibitor (imatinib) is recognized as the first-line chemotherapy for GIST, although a certain type of c-KIT mutation reveals a resistance to imatinib due to as yet uncertain molecular mechanisms. To assess the c-KIT mutation-related variation of cellular responses to imatinib, murine lymphocyte-derived Ba/F3 cells, which are stably transduced with different types of c-KIT mutation, were treated with either imatinib or a FAK inhibitor (TAE226), and their antitumor effects were determined in vitro and in vivo. A mutation at exon 11 (KITdel559-560) displayed a high sensitivity to imatinib, whereas that at exon 17 (KIT820Tyr) showed a significant resistance to imatinib in vitro and in vivo. KIT820Tyr cells appeared to maintain the activities of FAK and AKT under the imatinib treatment, suggesting that FAK might play a role in cell survival in imatinib-resistant cells. When FAK activity in those cells was inhibited by TAE226, cell growth was equally suppressed and the cells underwent apoptosis regardless of the c-KIT mutation types. Oral administration of TAE226 significantly diminished tumor growth in nude mice bearing KIT820Tyr xenografts. In summary, c-KIT mutation at exon 17 displayed a resistance to imatinib with maintained activations of FAK and subsequent survival signals. Targeting FAK could be a potential therapeutic strategy for imatinib-resistant GISTs. [Mol Cancer Ther 2009;8(1):127–34]

Heparanase expression correlates with malignant potential in human colon cancer

Purpose Heparanase cleaves carbohydrate chains of heparan sulphate proteoglycans and is an important component of the extracellular matrix. This study was designed to determine the relation between heparanase expression and prognosis of patients with colon cancer.Methods The study included 54 patients (35 males and 19 females) who underwent colorectal resection for colorectal cancer between January 1992 and December 1994. Expression of heparanase protein and mRNA were determined and correlated with various clinicopathological parameters. In vitro studies were also performed to examine tumor invasion and to test the effects of heparanase inhibition, and in vivo studies were performed to examine tumor metastasis and prognosis.Results Heparanase expression was detected in the invasion front of the tumor in 37 of 54 (69%) colon cancer samples, whereas 17 of 54 (31%) tumors were negative. Expression of heparanase was significantly more frequent in tumors of higher TNM stage (P=0.0481), higher Dukes stage (P=0.0411), higher vascular infiltration (P=0.0146), and higher lymph vessel infiltration (P=0.0010). Heparanase expression in colon cancers correlated significantly with poor survival (P=0.0361). Heparanase-transfected colon cancer cells exhibited significant invasion compared with control-transfected colon cancer cells (P=0.001), and the peritoneal dissemination model also showed the malignant potential of heparanase-transfected cells, as assayed by number of nodules (P=0.017) and survival (P=0.0062). Inhibition of heparanase significantly reduced the invasive capacity of cancer cells (P=0.003).Conclusions Heparanase is a marker for poor prognosis of patients with colon cancer and could be a suitable target for antitumor therapy in colon cancer.

Different gene expressions are associated with the different molecular subtypes of inflammatory breast cancer

The goal of this study was to determine whether gene expression differences exist between inflammatory breast cancers (IBC) and T stage-matched non-IBC patients stratified by hormone receptor and HER2 status. We used Affymetrix GeneChips to analyze 82 tumor samples (25 T4d patients, and 57 T4a-c patients) of newly diagnosed breast cancers. Genes that were differentially expressed between the IBC and non-IBC specimens were identified using the t test, and differential expression of gene sets was assessed using gene set analysis. Three distinct clinical subtypes of IBC and non-IBC were compared: ER-positive/HER2-normal, HER2-amplified, and ER-negative/HER2-normal. When we compared expression data from all IBC with all non-IBC, we found no significant differences after adjusting for multiple testing. When IBC and non-IBC tumors were compared by clinical subtype, however, significant differences emerged. Complement and immune system-related pathways were overexpressed in ER-positive/HER2-normal IBC. Protein translation and mTOR signaling were overexpressed in HER2-amplified IBC. Apoptosis-, neural-, and lipid metabolism-related pathways were overexpressed in ER-negative/HER2-normal IBC compared with non-IBC of the same receptor phenotype. In this T stage-matched case–control study, the survival curves of patients with IBC and non-IBC were similar for all three clinical subtypes. IBC tumors can be divided into molecular and clinical subtypes similar to those of non-IBC. Clinical subtypes of IBC show molecular differences compared with similar subtypes of non-IBC.

Dual-Tyrosine Kinase Inhibitor for Focal Adhesion Kinase and Insulin-like Growth Factor-I Receptor Exhibits Anticancer Effect in Esophageal Adenocarcinoma In vitro and In vivo

Purpose: Focal adhesion kinase (FAK) regulates integrin and growth factor–mediated signaling pathways to enhance cell migration, proliferation, and survival, and its up-regulation correlates malignant grade and poor outcome in several types of cancer. In this study, we aimed to raise a potential therapeutic strategy using a FAK inhibitor for Barretts esophageal adenocarcinoma. Experimental Design: The expression status of FAK in clinical Barretts esophageal adenocarcinoma tissues was determined by immunohistochemistry. Cultured esophageal adenocarcinoma cells were treated with TAE226, a specific FAK inhibitor with an additional effect of inhibiting insulin-like growth factor-I receptor (IGF-IR), to assess its anticancer effect in vitro. Western blot was carried out to explore a participating signaling pathway for TAE226-induced cell death. Furthermore, TAE226 was orally administered to s.c. xenograft animals to investigate its anticancer effect in vivo. Results: Strong expression of FAK was found in 94.0% of Barretts esophageal adenocarcinoma compared with 17.9% of Barretts epithelia, suggesting that FAK might play a critical role in the progression of Barretts esophageal adenocarcinoma. When esophageal adenocarcinoma cells were treated with TAE226, cell proliferation and migration were greatly inhibited with an apparent structural change of actin fiber and a loss of cell adhesion. The activities of FAK, IGF-IR, and AKT were suppressed by TAE226 and subsequent dephosphorylation of BAD at Ser136 occurred, resulting in caspase-mediated apoptosis. In vivo tumor volume was significantly reduced by oral administration of TAE226. Conclusions: These results suggest that TAE226, a dual tyrosine kinase inhibitor for FAK and IGF-IR, could become a new remedy for Barretts esophageal adenocarcinoma.

Free jejunal graft for hypopharyngeal and esophageal reconstruction

AimsThis study assessed the techniques of the free jejunal graft for the reconstruction of hypopharynx or cervical esophagus and discussed the main aspects related to those procedures.Methods and resultsBy using free jejunal grafts, we reconstructed 54 hypopharyngeal and cervical esophageal cancers. In this study, 23 out of 54 patients had a malignant tumor located in the hypopharynx and 31 in the cervical esophagus (27 primary cases and four secondary cases). Despite the multi-step and time-consuming procedure, we did not incur any trans-operative complication. Furthermore, we undertook the larynx preserving cervical esophagectomy and free jejunal graft reconstruction in six patients with cervical esophageal cancer, and those patients acquired a good quality of life.ConclusionFor the reconstruction of hypopharynx or cervical esophagus, the free jejunal graft is a very useful technique and improves the patient’s quality of life.