Naoki Ogane

Therapeutic strategy targeting the mTOR-HIF-1α-VEGF pathway in ovarian clear cell adenocarcinoma

Malignant tumors usually involve a relatively hypoxic state, which induces overexpression of hypoxia‐inducible factor‐1α (HIF‐1α) to satisfactorily enable the tumor to survive. Thus, inhibition of the mammalian target of rapamycin (mTOR) pathway including HIF‐1α is expected to play a major role in suppression of tumor cell growth, having recently drawn much attention as an anti‐cancer therapeutic strategy for various malignant tumors. In the present study, which compared clear cell adenocarcinoma (CLA) of the ovary with serous adenocarcinoma (SEA), the immunohistochemical expression of mTOR, phosphorylated‐mTOR (p‐mTOR), HIF‐1α, and vascular endothelial growth factor (VEGF) was examined in surgically resected specimens of 29 SEA and 47 CLA. There were no significant differences in expression of mTOR, HIF‐1α and VEGF between SEA and CLA, but it was noted that p‐mTOR expression was more prominent in CLA than SEA. Then, using the cell lines of CLA (RMG‐1 and W3uF), an experimental study was designed to clarify whether tumor suppression due to downregulation of mTOR activity could represent a promising therapeutic strategy for CLA. After treatment of an analogue of rapamycin (everolimus), expression of mTOR, p‐mTOR, HIF‐1α and VEGF was examined on western blot. As a result, although mTOR expression remained unchangeable, expression of p‐mTOR, HIF‐1α and VEGF was shown to be sharply depressed. The same expression alterations were demonstrated in the xenograft model treated with everolimus. In conclusion, mTOR‐targeted therapy through usage of drugs such as everolimus may be more effective for CLA of the ovary because of its significant expression of p‐mTOR.

Granulosa cell tumor with activated mTOR-HIF-1α-VEGF pathway

The DNA‐binding activity of hypoxia‐inducible factor‐1 α (HIF‐1α) has been analyzed for various gynecological tumors. Among the tumors that were studied, there was a finding of a high level of DNA‐binding HIF‐1α activity, although it was limited to one case of adult type granulosa cell tumor (GCT). In this case a 60‐year‐old female had marked immunohistochemical expression of HIF‐1α. The expressions of the mammalian target of rapamycin (mTOR) and phosphorylated‐mTOR (p‐mTOR) were also marked, and vascular endothelial growth factor (VEGF) was moderately expressed. To compare the expression profiles, 11 consecutive cases with adult type GCT were used. All cases showed marked expressions of HIF‐1α and mTOR, but p‐mTOR expression was moderately to markedly observed in four of the 12 cases. VEGF was expressed in all cases in varying degrees. Based on the evidence that downregulation of the mTOR pathway due to treatment with rapamycin (everolimus) would suppress tumor cell growth, an experimental study using the GCT cell line was designed to clarify whether HIF‐1α and VEGF expressions decline. As a result, the expressions of p‐mTOR, HIF‐1α and VEGF were suppressed, but those of mTOR were not. It was concluded that mTOR‐targeted therapy may represent a promising strategy for some GCT with an activated mTOR‐HIF‐1α‐VEGF pathway.

Clinicopathological implications of expressions of hypoxia-related molecules in esophageal superficial squamous cell carcinoma.

This study was conducted to clarify whether or not expressions of hypoxia-related molecules would have clinicopathological significance in squamous cell carcinoma (SCC) of the esophagus. Expressions of hypoxia inducible factor-1 alpha (HIF-1alpha), glucose transporter 1 (GLUT-1) and RAC-1 were immunohistochemically analyzed in 96 surgically resected SCCs at pT1b (sm1, 12 cases; sm2, 35 cases; sm3, 49 cases). They were divided into a lymph node metastasis (LNM)-positive group composed of 44 cases and an LNM-negative group composed of 52 cases. Immunohistochemical profiles were estimated based on the staining extent (score: 1+, 2+, 3+) and intensity (score: 1+, 2+, 3+). A significant expression pattern was found in the nucleus for HIF-1alpha, cell membrane for GLUT-1 and cytoplasm for RAC-1. The cases were categorized into a high score group (total score of 4 or more) and a low score group (total score of 3 or less) in each maker, respectively. A comparison made between the LNM-positive group and the LNM-negative group showed that the proportion of cases with a high score was larger in the LNM-positive group than in the LNM-negative group (HIF-1alpha, P = .02; GLUT-1, P = .008; RAC-1, P = .001). Among them, HIF-1alpha was found to be significantly related to the disease-free survival (P = .019) and overall survival (P = .034) as well as LNM (disease-free survival, P = .030; overall survival, P = .030). The multivariate analysis demonstrated that the HIF-1alpha expression would be an independent indicator for prognosis. In the superficial SCCs of the esophagus, GLUT-1 and RAC-1 may be involved in LNM, and HIF-1alpha overexpression is expected to predict an unfavorable clinical outcome.

Association of hypoxia-inducible factor-1 expression with histology in epithelial ovarian tumors: a quantitative analysis of HIF-1

BackgroundHypoxia-inducible factor-1 (HIF-1) is an essential transcription factor that mediates cellular and systemic homeostatic responses to reduced oxygen availability in mammals. So far, using immunohistochemistry we have analyzed the association of HIF-1α expression with histological type among epithelial ovarian tumors. In the present study, quantitative analyses of activated HIF-1 level in the nucleus and of accumulated HIF-1α level in the cytoplasm were performed to clarify whether or not the hypoxic state would be correlated to histology, malignancy, and tumor size in epithelial ovarian tumors.MethodHIF-1 level in the nucleus was analyzed using DNA binding assay, and HIF-1α level in the cytoplasm was measured by ELISA for a total of 36 epithelial ovarian tumors as follows: 5 serous adenocarcinomas (SEAs), 7 clear cell adenocarcinomas (CLAs), 7 endometrioid adenocarcinomas (ENAs), 4 mucinous adenocarcinomas (MUAs), 2 mucinous borderline tumors (MBTs), and 11 mucinous adenomas.ResultsHIF-1 level (mg/ml) in the nucleus and HIF-1α level (mg/ml) in the cytoplasm were on average 0.116 and 0.178 for SEAs, 0.328 and 0.306 for CLAs, 0.171 and 0.305 for ENAs, 0.097 and 0.176 for MUAs, 0.224 and 0.180 for mucinous borderline tumors, 0.152 and 0.154 for mucinous adenomas. CLAs showed the highest levels for both of HIF-1 and HIF-1α, while MUAs showed the lowest levels of both. Mucinous adenomas were higher in HIF-1 than MUAs.ConclusionHypoxic state was considered to be closely related to histological type of epithelial ovarian tumors, suggesting that CLAs may be most hypoxic. In the comparison of mucinous tumors, malignancies would not always become most hypoxic. Tumor size may not be strongly associated with hypoxic state.

Immunohistochemical Study of Type-1 Blood Antigen Expressions in Thyroid Tumors: The Significance for Papillary Carcinomas

Immunohistochemical expressions of type 1 blood group antigens were studied for 95 cases of thyroid tumors, including 29 follicular adenomas, 23 follicular carcinomas, and 43 papillary carcinomas, applying monoclonal antibodies against DU-PAN-2, CA19–9, Lewisa (Lea), and Lewisb (Leb). Normal thyroid tissue invariably failed to express all four antigens. In follicular adenomas, DU-PAN-2 and CA19–9 were focally expressed in 7% and 21% of cases, and in follicular carcinomas, CA19–9 expression was limited to one case (4%); all cases were negative for DU-PAN-2. No or little expression of Lea or Leb was observed in these follicular tumors. In contrast, DU-PAN-2, CA19–9, Lea, and Leb were expressed in 98%, 84%, 33%, and 49% of 43 papillary carcinomas, respectively. The positive stainings were observed mainly on the luminal surface of the tumor cells. The number of tumor cells that expressed DU-PAN-2 generally was greater than that of tumor cells that expressed CA19–9, Lea, or Leb. There was no significant difference in antigen expressions in female papillary carcinomas between subjects who were younger and older than 50 years old. The results suggest that DU-PAN-2 would be a useful immunohistochemical marker for distinguishing papillary carcinomas from follicular tumors. These immunohistochemical profiles imply the following: the activity of α2–3 sialyltransferase, a specific glycosyltransferase, would be more strongly enhanced in papillary carcinomas than in follicular tumors; the antigen expressions in papillary carcinomas may not be related to the alteration of the female sex hormone environment.

Endometrial intraepithelial carcinoma in association with polyp: review of eight cases

BackgroundThe uterine endometrial polyp (EMP) has a potential risk of developing malignant tumors especially in postmenopausal women. These malignancies include endometrial intraepithelial carcinoma (EIC).Patients and methodsEight patients with EIC in the EMP, who were postmenopausal with ages ranging from 49 to 76 years (av. 62), were cytologically reviewed in comparison with histological findings.ResultsThe endometrial cytological findings were summarized as follows: mucous and watery diathesis as a background lacking or with little necrotic inflammatory changes; micropapillary cluster formation; abrupt transition between carcinoma cells and normal cells; nuclear enlargement; high N/C ratio; and single or a few prominent nucleoli. Histologically, one case had EIC alone in the EMP; three cases had EIC with stromal invasion confined to the EMP; and four cases had EIC in the atrophic endometrium in addition to EIC in the EMP. Seven patients have taken a disease-free course after surgical resection, but one patient died 44 months following the initial diagnosis because of the massive tumor extending over her peritoneal cavity.ConclusionsEndometrial cytology may be helpful for the detection of early endometrial adenocarcinomas with serous features including EIC. Some early stage endometrial adenocarcinomas represented by EIC exceptionally take an aggressive clinical course irrespective of a lack of extrauterine lesions.Virtual SlidesThe virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1651876760876449

Immunohistochemical expression profiles of solute carrier transporters in alpha‐fetoprotein‐producing gastric cancer

Alpha‐fetoprotein (AFP)‐producing gastric cancer (GC) is an aggressive tumour with high rates of liver metastasis and poor prognosis, and for which a validated chemotherapy regimen has not been established. Drug uptake by solute carrier (SLC) transporters is proposed as one of the mechanisms involved in sensitivity to chemotherapy. In this study, we aimed to develop important insights into effective chemotherapeutic regimens for AFP‐producing GC.

Association of histone deacetylase expression with histology and prognosis of ovarian cancer

Histone deacetylase (HDAC) inhibitor is known to have a cytotoxic effect on ovarian cancer cell lines. The present study analyzed the association between immunohistochemical HDAC expression and clinicopathological findings, in particular, the association with histological type and effect of chemotherapy. The histology of the 201 ovarian cancers addressed was as follows: Serous carcinoma (SEC), 100 cases; clear cell carcinoma (CCC), 56 cases; endometrioid carcinoma (EMC), 36 cases; and mucinous carcinoma (MUC), 9 cases. Immunohistochemical analyses of HDACs 1, 2, 3, 4, 5, 6 and 7 expression levels were performed using tissue microarrays, composed of 201 primary tumors and 38 tumors following chemotherapy. Overexpression of HDAC1 was detected in the nucleus of all cases with MUC, followed by CCC (80%), SEC (73%), and EMC (53%). CCC specifically demonstrated HDAC7 expression in both the nucleus (27%) and the cytoplasm (54%), and HDAC6 expression in the nucleus (34%). The comparison between prior to and following chemotherapy revealed a nuclear expression increase in HDAC1 (76% vs. 92%; P=0.03) and HDAC7 (0.0 vs. 16%; P=0.01), and cytoplasmic expression increase in HDAC6 (40 vs. 74%; P=<0.01) and HDAC7 (16 vs. 66%; P=<0.01). HDAC1 nuclear expression adversely affected overall survival in SEC (P=0.02) and EMC (P=0.03), and HDAC7 cytoplasmic expression in CCC was associated with a poor prognosis (P=0.06). In multivariate analysis, HDAC6 nuclear expression was determined as a poor prognostic factor (hazard ratio=3.51; 95% confidence interval, 1.49 to 8.27, P=<0.01). In the subgroup analysis, HDAC6 nuclear expression was associated with a poor prognosis in CCC (P=0.07), International Federation of Obstetrics and Gynecology stage III/IV (P=0.07), and suboptimal surgery (P=<0.01). In conclusion, HDACs may be associated with the prognosis of ovarian cancers, depending on the histological subtypes, and upregulated following chemotherapy. HDAC1, 6 and 7 may therefor act as promising therapeutic targets in the future.

Preponderance of endometrial carcinoma in elderly patients

Elderly patients with endometrial carcinoma (EMC) are considered to have a poor clinical outcome. The present study included 79 patients aged ≥70 years with EMC stage I or II according to the International Federation of Gynecology and Obstetrics classification, and it was conducted to analyse the clinicopathological significance of histological type (I or II), depth of myometrial invasion (<1/2 or ≥1/2), lymphovascular invasion (+ or -) and immunohistochemical profile. The aim of these analyses was to determine whether these factors may adversely affect the patient outcome and the underlying mechanisms. The immunohistochemical markers used were estrogen receptor (ER), Ki-67 and p53. The expression of these markers was evaluated as high (+) or low (−). Accordingly, the patients were divided into groups as follows: 54 cases type I vs. 25 cases type II; 48 cases with myometrial invasion <1/2 vs. 31 cases without myometrial invasion ≥1/2; 63 cases with lymphovascular invasion vs. 16 cases without lymphovascular invasion; 57 cases with ER (+) vs. 22 cases with ER (−); 24 cases with Ki-67 (+) vs. 55 cases with Ki-67 (−); and 29 cases with p53 (+) vs. 50 cases with p53 (−). In conclusion, close attention must be paid to elderly patients with EMC due to the tumors intrinsic aggressiveness, which may include the ER (−) and p53 (+) pattern as an independent poor prognostic factor.

Cleaved caspase-3 expression is a potential prognostic factor for endometrial cancer with positive peritoneal cytology

Positive peritoneal cytology (PPC) in endometrial cancer remains a controversial topic. Cleaved caspase‐3 (CC3) and Ki‐67 are excellent markers of apoptotic and proliferating cells, respectively. The objective of this study was to determine the significance of CC3 and Ki‐67 expression in peritoneal cytology samples as prognostic factors for endometrial cancer with PPC.