Efficacy and safety of rebamipide liquid for chemoradiotherapy-induced oral mucositis in patients with head and neck cancer: a multicenter, randomized, double-blind, placebo-controlled, parallel-group phase II study.

BACKGROUND: Recent preclinical and phase I studies have reported that rebamipide decreased the severity of chemoradiotherapy-induced oral mucositis in patients with oral cancer. This placebo-controlled randomized phase II study assessed the clinical benefit of rebamipide in reducing the incidence of severe chemoradiotherapy-induced oral mucositis in patients with head and neck cancer (HNC). METHODS: Patients aged 20-75 years with HNC who were scheduled to receive chemoradiotherapy were enrolled. Patients were randomized to receive rebamipide 2% liquid, rebamipide 4% liquid, or placebo. The primary endpoint was the incidence of grade ≥ 3 oral mucositis determined by clinical examination and assessed by central review according to the Common Terminology Criteria of Adverse Events version 3.0. Secondary endpoints were the time to onset of grade ≥ 3 oral mucositis and the incidence of functional impairment (grade ≥ 3) based on the evaluation by the Oral Mucositis Evaluation Committee. RESULTS: From April 2014 to August 2015, 97 patients with HNC were enrolled, of whom 94 received treatment. The incidence of grade ≥ 3 oral mucositis was 29% and 25% in the rebamipide 2% and 4% groups, respectively, compared with 39% in the placebo group. The proportion of patients who did not develop grade ≥ 3 oral mucositis by day 50 of treatment was 57.9% in the placebo group, whereas the proportion was 68.0% in the rebamipide 2% group and 71.3% in the rebamipide 4% group. The incidences of adverse events potentially related to the study drug were 16%, 26%, and 13% in the placebo, rebamipide 2%, and rebamipide 4% groups, respectively. There was no significant difference in treatment compliance among the groups. CONCLUSIONS: The present phase II study suggests that mouth washing with rebamipide may be effective and safe for patients with HNC receiving chemoradiotherapy, and 4% liquid is the optimal dose of rebamipide. TRIAL REGISTRATION: ClinicalTrials.gov under the identifier NCT02085460 (the date of trial registration: March 11, 2014).

Indocyanine green angiography for intra-operative assessment in vascular surgery.

OBJECTIVES: Indocyanine green (ICG) angiography is used for the intra-operative assessment of the graft vessel in coronary artery bypass grafting to enable immediate revision if necessary. We report the feasibility and implications of an ICG colour imaging system, HyperEye Medical System (HEMS), in surgeries for arteriosclerosis obliterans (ASO) and abdominal aortic aneurysm (AAA) which carry risk of mesenteric ischaemia. METHODS: HEMS ICG angiography was used for the intra-operative assessment of 12 ASO patients and 10 AAA patients. RESULTS: In the ASO patients, HEMS angiography enabled visualisation of the graft and native artery. The fluorescent lucent region in the artery distal to the anastomosis was shown in 1 of 12 ASO patients. There was a 3-s time lag in the increase of intensity between the proximal artery and distal stenotic region. In AAA patients, HEMS angiography clearly showed the perfusion in the mesenteric arteries and intestinal wall as opaque. One AAA patient had segmental ischaemia due to thromboembolism and another one had diffuse ischaemia due to systemic malperfusion. The ischaemic region of the intestine was visualised as a fluorescent lucent area by HEMS angiography. CONCLUSION: HEMS angiography can accurately assess peripheral arterial perfusion in surgical cases with ASO and AAA.

Influence of 70 nm silica particles in mice with cisplatin or paraquat-induced toxicity.

In the pharmaceutical industry, nano-size materials are designed as drug carriers and diagnosis probes. Interactions between nano-size materials and chemicals need investigating. Here, we investigated whether nano-size materials affect chemical-induced toxicity using silica particles, which have been widely used in cosmetics and drug delivery and have diameters of 70 (SP70), 300 (SP300) and 1000 (SP1000) nm, a popular anti-tumor agent, cisplatin, and a widely used herbicide, paraquat. Mice were treated with either cisplatin (100 micromol/kg, intraperitoneally) or paraquat (50 mg/kg, intraperitoneally), with or without intravenous silica particle administration. All treatments were non-lethal and did not show severe toxicity, except for injection with both cisplatin and SP70, which were lethal. When mice received with paraquat and/or the silica particles, synergistic enhanced toxicity was observed in both paraquat- and SP70-treated mice. These synergic effects were not observed with either Si300 or 1000 treatment. Our findings suggest that further evaluation on the interaction between nano-size materials and chemicals is critical for the pharmaceutical application of nanotechnology.

Antiangiogenic activity of BAI1 in vivo: implications for gene therapy of human glioblastomas.

Glioblastomas are the most common primary brain tumors in adults. These tumors exhibit a high degree of vascularization, and malignant progression from astrocytoma to glioblastoma is often accompanied by increased angiogenesis and the upregulation of vascular endothelial growth factor and its receptors. In this study, we investigated the in vivo antiangiogenic and antitumor effects of brain-specific angiogenesis inhibitor 1 (BAI1) using human glioblastoma cell lines. Glioblastoma cells were transduced with an adenoviral vector encoding BAI1 (AdBAI1), and Northern and Western blot analyses, respectively, demonstrated BAI1 mRNA and protein expression in the transduced tumor cells. Using an in vivo neovascularization assay, we found that angiogenesis surrounding AdBAI1-transduced glioblastoma cells transplanted into transparent skinfold chambers of SCID mice was significantly impaired compared to control treated cells. Additionally, in vivo inoculation with AdBAI1 of established subcutaneous or intracerebral transplanted tumors significantly impaired tumor growth and promoted increased mouse survival. Morphologically, the tumors exhibited signs of impaired angiogenesis, such as extensive necrosis and reduced intratumoral vascular density. Taken together, these data strongly indicate that BAI1 may be an excellent gene therapy candidate for the treatment of brain tumors, especially human glioblastomas.

Metastases-related genes in the classification of liver and peritoneal metastasis in human gastric cancer.

INTRODUCTION: With the aim of identifying metastases-related genes in gastric cancer, we performed a broad analysis of differential gene expression between low-metastatic parental cell lines and established highly metastatic sublines. MATERIALS AND METHODS: We established novel cell lines, AZ-H5c, NUGC-3H5, and TMK-1H7, with a high potential of liver metastasis, and AZ-P7a, NUGC-3P4T, and TMK-1P4a, with a high potential of peritoneal metastasis. These cell lines were derived from low-metastatic parental AZ-521, NUGC-3, and TMK-1 cell lines, respectively. Furthermore, to investigate different levels of gene expression implicated in metastatic potentials in gastric cancer, we investigated approximately 2000 expressed genes in each cell line using a DNA microarray. RESULTS: Varieties of genes were up-regulated or down-regulated in highly metastatic liver and peritoneal cell lines. Fifty-eight genes, including the transferrin receptor, ras-related rho, and osteopontin, and 22 genes, including apolipoprotein E and inhibin A-submit, were up-regulated and down-regulated in two or three liver metastatic sublines. On the other hand, 19 genes, the transferrin receptor, c-fos, and RANTES, and 26 genes, including MAC25, PISSLRE, and RNA polymerase, were up-regulated and down-regulated in two or three peritoneal metastatic sublines. CONCLUSION: How gene expression is implicated in gastric cancer metastasis has never been thoroughly explained, and further studies are necessary to understand the involvement of genes in cancer metastasis more thoroughly. We hope that our highly metastatic liver and peritoneal experimental models are helpful for further study and gene therapy of human gastric cancer.

Profiling analysis of differential gene expression between hematogenous and peritoneal metastatic sublines of human pancreatic cancer using a DNA chip.

We established the novel sublines HPC-1H5, HPC-3H4, HPC-4H4, and Panc-1H5, which have a high potential of liver metastasis, and HPC-1P5a, HPC-3P4a, HPC-4P4a, and Panc-1P5a, which have a high potential of peritoneal dissemination, derived from low metastatic HPC-1, HPC-3, HPC-4, and Panc-1cell lines, respectively. To clarify the molecular mechanisms of cancer metastasis and of the different levels of gene expression in a variety of metastatic potentials in pancreatic cancer, we performed a broad analysis of differential gene expression analysis between parental cell lines and metastatic sublines. In comparison with the parental cell lines, 65 and 36 genes were overexpressed and underexpressed in highly liver-metastatic sublines. On the other hand, 43 and 45 genes were overexpressed and underexpressed in highly peritoneal-metastatic sublines. uPAR and Serin protease were overexpressed, and E2A and IGF1R were underexpressed in both metastatic sublines. Hierarchical clustering analysis revealed 22 genes classifying liver, peritoneal metastatic sublines and low-metastatic parental cell lines. These genes might be targeted genes separating those two major metastatic forms after surgery. A greater number of cell line samples and more genes will have to be utilized in future studies in order to understand the involvement of genes in cancer metastasis more thoroughly. However, these results will help to clarify the molecular mechanisms of pancreatic cancer metastasis.

Percutaneous radiofrequency ablation for unresectable large hepatic tumours during hepatic blood flow occlusion in four patients.

AIM: To evaluate percutaneous radiofrequency (RF) ablation therapy for unresectable large hepatic tumours combined with regional interruption of hepatic blood flow, and to assess the safety and efficacy of this procedure. MATERIALS AND METHODS: Four patients with hepatic tumours were enrolled in this study. Patients were treated by a single session of RF ablation during occlusion of both hepatic artery and hepatic vein. Tumour size ranged from 45-57 mm (mean 50.2 mm). Initial therapeutic efficacy was evaluated with helical computed tomography (CT) performed within 9 days after the treatment. CT or magnetic resonance imaging (MRI) was performed every 2-3 months thereafter. RESULTS: The largest axis of coagulated lesions after the ablation was 50-60 mm (mean 56.5 mm) in diameter. The ablation therapy was considered complete in three patients; after a mean follow-up of 12.7 months, CT and MRI revealed complete destruction of their tumours. One patient required further treatment. No severe complications occurred. CONCLUSION: Although further studies are needed, in this limited clinical trial a local ablation area exceeding 50 mm in diameter was achieved safely.

Differential gene expression screening between parental and highly metastatic pancreatic cancer variants using a DNA microarray.

To clarify the difference in genes expressed in hematogenous metastasis and peritoneal dissemination, a broad analysis of differential gene expression analysis between parental cell lines and established metastatic sublines was performed. Using an oligonucleotide array (Gene Chip, Affymetrix), approximately 2,000 genes involved in cancer were analyzed for each of the cell lines. HPC-4H4 (highly metastatic lines to the liver) compared with HPC-4 (low metastatic parental lines), in which 20 overexpressed genes and 5 underexpressed genes were recognized. HPC-4P4a (highly metastatic to the peritoneum) compared with HPC-4, in which 12 overexpressed genes and 15 underexpressed genes were also recognized. Analysis of HPC-4H4 and HPC-4P4a showed comparative up-regulation of 20 genes and down-regulation of 13 in the former, HPC-4H4. Further studies are needed to validate our hypothesis that some of the resulting differentially expressed genes might be implicated in the development of metastasis in pancreatic cancer. In conclusion, this genome-wide expression analysis will help to clarify the molecular mechanisms of cancer metastasis and of the different levels of gene expression in a variety of metastatic potentials in pancreatic cancer.

Metastatic-associated biological properties and differential gene expression profiles in established highly liver and peritoneal metastatic cell lines of human pancreatic cancer.

To elucidate metastasis mechanisms, we established a Panc-1H5 subline with a highly liver metastatic cell line and a Panc-1P4a with a highly peritoneal metastatic cell line, which were sequentially selected from the parental pancreatic cancer cell line Panc-1. Using these three cell lines, we investigated several biological properties and mRNA levels of differentially-expressed genes involved in cancer metastasis with a cDNA macroarray. The tumorigenicity, motile activity, adhesive activity and cytokine production of metastatic sublines were higher than those of parental Panc-1 cells. Particularly, in Panc-1H5 cells, adhesive activity to the extracellular matrix and angiogenetic factors increased, whereas in Panc-1P4a cells, motile activity was extremely enhanced compared with Panc-1 cells. Histopathological findings for the three cell lines were the same. In cDNA macroarray analysis of Panc-1H5 cells, 11 genes were up-regulated and 20 genes were down-regulated compared with parental Panc-1 cells. In Panc-1P4a cells, 7 genes were up-regulated and 13 genes were down-regulated compared with parental Panc-1 cells. This study provides a demonstration of global gene expression analysis of pancreatic cancer cells with liver and peritoneal metastasis and these results provide new insight into the study of human pancreatic cancer metastasis.

Adenosquamous carcinoma of the colorectum: report of two cases.

Adenosquamous carcinomas of the colorectum are rare neoplasms. Our experience with two cases is presented in this paper. One patient, who complained of bloody stool, was found to have adenocarcinoma in the sigmoid colon. He received a laparoscopy-assisted sigmoidectomy. The histological examination revealed that the tumor was adenosquamous carcinoma. To date, he has survived six months post operatively without evidence of recurrence. The other patient, who complained of anal bleeding, was found to have rectal adenocarcinoma and received a low anterior resection. Histological examination revealed that the tumor was an adenosquamous carcinoma. He remains alive, with no evidence of recurrence, nine years post operatively. Both cases showed paracolic lymph node metastasis. Because of its very low incidence, the histogenesis, malignancy and prognosis of this disease remain unclear. Thus, further clinical and histological study of this disease entity is required.

A novel experimental mouse model of peritoneal dissemination of human gastric cancer cells: analysis of the mechanism of peritoneal dissemination using cDNA macroarrays.

We established a new cell line, NUGC-3P4T, with high peritoneal metastatic disseminating potential in nude mice. NUGC-3P4T cells were derived from the human gastric carcinoma line NUGC-3, which has low capacity for peritoneal dissemination. NUGC-3P4T cells developed peritoneal dissemination in 10 / 10 (100%) mice, whereas the parental NUGC-3 cells developed dissemination in 1 / 5 (20.0%) mice. The metastatic foci in the peritoneum showed essentially the same histological appearance as those induced by parental cells. The tumorigenicity, the motile activity and the adhesive activity to the laminin of NUGC-3P4T cells were stronger than those of NUGC-3 cells. Production of IL-8 was significantly higher in NUGC-3P4T than in NUGC-3. cDNA macroarrays analysis showed that a variety of cytokines, interleukins, and other immunomodulators and their receptors were up- or down-regulated at the mRNA level in NUGC-3P4T cells, compared with NUGC-3 cells. Thus, this unique cell line and in vivo model might be useful to study the biology of peritoneal dissemination of human gastric cancer.

A new peritoneal dissemination model established from the human pancreatic cancer cell line.

We established a new cell line, HPC-3P4a, with high peritoneal disseminated potential in nude mice. HPC-3P4a was derived from a human pancreatic carcinoma cell line (HPC-3) that had low capacity for peritoneal dissemination. HPC-3P4a developed peritoneal dissemination in 10 of 11 (90.9%) cases, whereas parental HPC-3 developed peritoneal dissemination in one of six (16.7%) cases. The metastatic foci in the peritoneum showed essentially the same histologic appearance of parental involvement. The tumorigenicity, motility, and adhesive activity of HPC-3P4a to the extracellular matrix were stronger than were those of the HPC-3. In FACS analysis, HPC-3P4a significantly increased the expression of alpha6 and alpha(v)beta5 integrins, while it decreased alpha2 integrin, hCD44H, and hCD44v 10, as compared with HPC-3. The VEGF production of HPC-3P4a was significantly lower than that of HPC-3. Analysis of gene macroarrays showed a variety of cytokines, interleukin, and other immunomodulatory, and their receptors were up-regulated and down-regulated on an mRNA level in HPC-3P4a cells, compared with HPC-3 cells. Intrasplenic injection of HPC-3P4a produced no liver metastasis. We named our original highly liver metastatic cell line HPC-3H4 (previously reported). This HPC-3H4 cell was established by repeated intrasplenic injection from parental cell HPC-3; thus, it developed high liver metastasis. Moreover, HPC-3H4 developed peritoneal dissemination by intra-abdominal injection. In contrast, HPC-3P4a did not develop liver metastasis by intrasplenic injection. These findings are very interesting and might suggest that the process of hematogenous metastasis differed from that of peritoneal dissemination. Thus, this cell line may be useful for investigating the mechanism of peritoneal dissemination in human pancreatic cancer.

p53AIP1, a potential mediator of p53-dependent apoptosis, and its regulation by Ser-46-phosphorylated p53.

Through direct cloning of p53 binding sequences from human genomic DNA, we have isolated a novel gene, designated p53AIP1 (p53-regulated Apoptosis-Inducing Protein 1), whose expression is inducible by wild-type p53. Ectopically expressed p53AIP1, which is localized within mitochondria, leads to apoptotic cell death through dissipation of mitochondrial A(psi)m. We have found that upon severe DNA damage, Ser-46 on p53 is phosphorylated and apoptosis is induced. In addition, substitution of Ser-46 inhibits the ability of p53 to induce apoptosis and selectively blocks expression of p53AIP1. Our results suggest that p53AIP1 is likely to play an important role in mediating p53-dependent apoptosis, and phosphorylation of Ser-46 regulates the transcriptional activation of this apoptosis-inducing gene.

Tubular adenoma of the breast in a 73-year-old woman.

A 73-year-old woman presenting with a right breast mass is described. The patient underwent lumpectomy under a diagnosis of breast cancer. However, histopathologically the surgical specimen was tubular adenoma of the breast. This is a rare benign tumor that is difficult to differentiate from breast cancer clinically, especially in elderly patients. We describe two reported cases of tubular adenoma in patients older than 65-years in Japan, as well as the present case.

A novel experimental mouse model of peritoneal dissemination of human gastric cancer cells: different mechanisms in peritoneal dissemination and hematogenous metastasis.

We established a new cell line, AZ-P7a, with high peritoneal-metastatic potential in nude mice. AZ-P7a cells were derived from the human gastric carcinoma line AZ-521, which has low capacity for peritoneal dissemination. AZ-P7a cells developed peritoneal metastasis in 11 / 14 (78.6%) mice, whereas the parental AZ-521 cells developed metastasis in 2 / 6 (33.3%) mice. The metastatic foci in the peritoneum showed essentially the same histological appearance as those induced by parental cells. The tumorigenicity and the motile activity of AZ-P7a cells were stronger than those of the parental AZ-521 cells; in contrast, adhesion to the extracellular matrix and the production of vascular endothelial growth factor by AZ-P7a cells were decreased. In fluorescence-activated cell sorter (FACS) analysis, AZ-P7a cells expressed significantly greater levels of integrins alpha2, alpha3, alpha5, alpha6 and alphavbeta5, as compared with AZ-521 cells. However, alpha1, alpha4, alphavbeta3, hCD44H, hCD44v3, hCD44v6 and hCD44v10 were not expressed in either cell line. AZ-P7a cells developed no liver metastasis when administered by the intrasplenic injection method, though the highly liver metastatic cell line AZ-H5c showed the same rate of peritoneal dissemination as that exhibited by AZ-P7a cells after intraabdominal injection. These findings suggested that the mechanism of peritoneal dissemination differed from that of hematogenous metastasis. Moreover, the latter appears to be controlled by more complex mechanisms than the former. Thus, this cell line might be useful for investigating the mechanism of peritoneal dissemination of human gastric cancer.

Analysis of the p300/CBP-Associated Factor (PCAF) gene in astrocytic tumors.

The PCAF gene encodes the p300/CBP-Associated Factor (PCAF), a histone acetyltransferase, which regulates p53 by acetylation of Lys320 in the C-terminal portion of p53. While the p53 gene is one of the most frequently mutated tumor suppressor genes in human tumors, such mutations occur in only 30% of astrocytic tumors. Since PCAF can regulate p53 activity, abrogation of PCAF function by PCAF gene mutation could be an alternate mechanism to inactivate the p53 pathway in tumors lacking p53 mutations. To test this hypothesis, we determined the nucleotide sequence of the entire PCAF coding region in 37 astrocytic tumors (17 glioblastomas, 10 anaplastic astrocytomas, 7 low-grade astrocytomas, and 3 pilocytic astrocytomas). We detected two single-nucleotide alterations that represented non-deleterious polymorphisms (GAG > GAA Glu103Glu, AAT > AGT Asn386Ser) but no obvious functional mutations. Moreover, the frequency of the Asn386Ser allele that contained Ser386 in glioma patients was not statistically different from its frequency in individuals without disease, and no significant association was observed between the PCAF polymorphisms and the presence or absence of p53 mutations in the tumors. We conclude that the PCAF gene is not mutated during the development of the astrocytic tumors studied here.

Recombinant adenovirus expressing wild-type p53 is antiangiogenic: a proposed mechanism for bystander effect.

Angiogenesis is required for the growth and progression of malignancies. Recent studies have demonstrated that genetic alterations may accompany acquisition of the angiogenic phenotype. The tumor suppressor gene p53 is most frequently mutated in human cancers and is also known to be a transcriptional regulator of a variety of genes. Here, we investigated the antiangiogenic effect of the wild-type p53 (wt-p53) gene transfer on a human non-small cell lung cancer cell line. Mutant p53-expressing H226Br non-small cell lung cancer cells were transduced with the wt-p53 gene using a recombinant adenoviral vector (Ad5CMVp53) and applied to semiquantitative reverse transcription-PCRs for the detection of altered mRNA expression of angiogenic and/or antiangiogenic factors. In vivo neovascularization assay of Ad5CMVp53-infected cells was then performed using a membrane-diffusion chamber system s.c. transplanted in nu/nu mice. We also evaluated the effect of Ad5CMVp53-infected H226Br cells on nontransduced tumor cells in vivo by s.c. inoculating mixture of cells into nu/nu mice. Ad5CMVp53 infection markedly inhibited the expression of an angiogenic factor, vascular endothelial growth factor, and increased the expression of a novel antiangiogenic factor, brain-specific angiogenesis inhibitor 1, resulting in reduced neovascularization in vivo. Mixing experiments showed that tumor cells transduced with the wt-p53 gene inhibited the in vivo tumor growth of adjacent nontransduced cells. Our data suggest that a recombinant adenovirus expressing the wt-p53 gene is antiangiogenic, which may explain, in part, the mechanism of the bystander effect induced by the wt-p53 gene transfer on adjacent tumor cells.

Identification of BAIAP2 (BAI-associated protein 2), a novel human homologue of hamster IRSp53, whose SH3 domain interacts with the cytoplasmic domain of BAI1.

BAI1 (brain-specific angiogenesis inhibitor 1) was originally isolated as a p53-target gene specifically expressed in brain. To clarify its function, we have been searching for cellular proteins that associate with the cytoplasmic domain of BAI1. Using its intracellular carboxyl terminus as "bait" in a yeast two-hybrid system, we isolated a cDNA clone named BAIAP2 whose nucleotide sequence would encode a 521-amino acid protein showing significant homology to a 58/53-kDa substrate of insulin-receptor kinase in the hamster. As the expression profile of BAIAP2 examined by Northern blot analysis was almost identical to that of BAI1, BAIAP2 appears to be active mainly in neurons. In vitro binding assays confirmed that a proline-rich cytoplasmic fragment of BAI1 interacted with the Src homology 3 (SH3) domain of BAIAP2. Double-color immunofluorescent analysis revealed that BAIAP2 was localized at the cytoplasmic membrane when it was coexpressed with BAI1 in COS-7 cells; BAIAP2 not associated with BAI1 was diffused in the cytoplasm. Predominant localization of BAI1 protein in a sub-cellular fraction enriched in growth cones indicated a possible role of BAI1 as a cell adhesion molecule inducing growth cone guidance. As a protein partner of BAI1, BAIAP2 may represent an important link between membrane and cytoskeleton in the process of neuronal growth.