Up-regulation of HDAC6 Results in Poor Prognosis and Chemoresistance in Patients With Advanced Ovarian High-grade Serous Carcinoma.

BACKGROUND: Ovarian high-grade serous carcinoma (HGSC) gradually acquires chemoresistance after recurrence. Our previous study on ovarian clear-cell carcinoma found histone deacetylase 6 (HDAC6) overexpression led to chemoresistance. This study aimed to evaluate HDAC6 as a predictor of chemoresistance and a therapeutic target for ovarian HGSC. PATIENTS AND METHODS: The clinical significance of HDAC6 as a predictor of prognosis and chemoresistance in HGSC was immunohistochemically evaluated. In addition, expression of programmed cell death ligand-1 (PD-L1), and hypoxia-inducible factor-1α (HIF1α) were analyzed using clinical samples from 88 patients with ovarian HGSC, and their clinicopathological characteristics were reviewed. RESULTS: Twenty-three patients had high HDAC6 expression, 10 positive PD-L1 expression, and 33 high HIF-1α expression. HDAC6 up-regulation was correlated with not undergoing interval debulking surgery (p<0.001), incomplete surgical resection (p=0.002), and frequent occurrence of stable disease/progressive disease according to the Response Evaluation Criteria in Solid Tumors (p=0.005) criteria. On Kaplan-Meier analysis, high HDAC6 expression was significantly associated with reduced progression-free (p=0.001) and overall (p=0.008) survival. On multivariate analysis, high HDAC6 expression (hazard ratio=1.65, 95% confidence interval 1.03-2.66; p=0.039) and surgery status were independent prognostic factors of progression-free survival. PD-L1 and HIF1α expression positively correlated with that of HDAC6. CONCLUSION: HDAC6 may become a potential therapeutic target in patients with ovarian HGSC since its up-regulation is considered to be associated with a poor prognosis in patients with this cancer.

Hypoxia-inducible Factor-1α Suppression in Ovarian Clear-cell Carcinoma Cells by Silibinin Administration.

BACKGROUND/AIM: Advanced ovarian clear-cell carcinoma (CCC) fails to respond to standard chemotherapy, and has a poor prognosis. Since hypoxia-inducible factor-1 (HIF-1) stimulates various genes involved in cancer, we aimed to examine the efficacy of silibinin, an active component of milk thistle belonging to Asteraceae, in suppressing HIF-1 activity, and elucidate the underlying mechanism in human CCC cell lines. MATERIALS AND METHODS: Human ovarian CCC cell lines HAC-2, OVISE, and RMG-1 were treated with 500 µM silibinin for 4 h under normoxic and hypoxic conditions. Using DNA microarray, we analysed genes whose expression modulated more than 2-fold in response to hypoxia, whereas HIF-1α expression was measured using ELISA. RESULTS: Silibinin treatment decreased HIF-1α protein in all cell lines, and eIF4E2 and RPS6 mRNA in HAC-2 and RMG-1 cells. CONCLUSION: Silibinin suppressed HIF-1α protein under hypoxic conditions in CCC cell lines and could be a potential anti-cancer drug.

Clinicopathological correlation of ARID1A status with HDAC6 and its related factors in ovarian clear cell carcinoma.

Ovarian clear cell carcinoma (OCCC) is associated with a frequent loss in ARID1A function. ARID1A reportedly suppresses histone deacetylase (HDAC)6 in OCCC directly. Here, we evaluated the clinical significance of HDAC6 expression and its related factors in terms of ARID1A status. Immunohistochemical expression of HDAC6, hypoxia inducible factors-1α (HIF-1α), programmed death-1 ligand (PD-L1), CD44 (cancer stem cell marker), and ARID1A was analysed for 106 OCCC patients. High nuclear HDAC6 expression correlated with patient death (p = 0.038). In the multivariate analysis of overall survival, surgical status (complete or incomplete resection) (hazard ratio (HR) = 17.5; p = <0.001), HDAC6 nuclear expression (HR = 1.68; p = 0.034), and PD-L1 expression (HR = 1.95; p = 0.022) were the independent prognostic factors. HDAC6 upregulation and ARID1A loss did not necessarily occur simultaneously. High HDAC6 expression was associated with poor prognosis in OCCC with ARID1A loss; this was not observed without ARID1A loss. HDAC6 expression showed a significant positive correlation with HIF-1α, PD-L1, and CD44. In OCCC, HDAC6 involvement in prognosis depended on ARID1A status. HDAC6 also led to immuno- and hypoxia- tolerance and cancer stem cell phenotype. HDAC6 is a promising therapeutic target for OCCC with loss of ARID1A.

Impact of TP53 immunohistochemistry on the histological grading system for endometrial endometrioid carcinoma.

Endometrial endometrioid carcinoma is usually divided into three histological subgroups: grade 1 (G1), grade 2 (G2), and grade 3 (G3). Most cases of endometrial endometrioid carcinoma G1/2 have a favorable prognosis, although some can have unfavorable outcomes, especially when they involve elderly patients, with similarities to endometrioid carcinoma G3 and serous carcinoma. This retrospective study evaluated whether TP53 abnormalities in endometrial endometrioid carcinoma could be used to supplement the current grading system and improve its ability to predict clinical outcomes. Immunohistochemical expression of TP53 was analyzed using tissue microarrays from the surgically resected specimens of 475 patients with endometrial endometrioid carcinoma. Weak or moderate expression was defined as TP53-normal expression, while absent or strongly positive expression was defined as TP53-aberrant expression. The endometrial endometrioid carcinomas had originally been diagnosed as G1 (69%), G2 (18%), and G3 (13%). Univariate analyses revealed that TP53-aberrant expression was associated with poor survival in G1 and G2 cases, but not G3 cases. In addition, age (<60 years vs. ≥60 years) was correlated with TP53-aberrant expression in G1 cases (3% vs. 16%, p = 0.001), but not in G2 or G3 cases. Based on immunohistochemical TP53 expression, the endometrial endometrioid carcinomas were reclassified using a prognostic grading system as high-grade (G1 or G2 with TP53- aberrant expression, and G3 with TP53-normal or -aberrant expression) or low-grade (G1 or G2 with TP53-normal expression). The multivariate analyses revealed that the prognostic grading system (using histological grade and TP53 expression) could independently predict poor progression-free survival (hazard ratio: 2.91, p < 0.001) and overall survival (hazard ratio: 3.62, p < 0.001). Therefore, combining immunohistochemical TP53 expression with the traditional histological grading system for endometrial endometrioid carcinoma may help improve its ability to accurately predict the patient's prognosis.

Association of the hypoxia-inducible factor-1α (HIF-1α) gene polymorphisms with prognosis in ovarian clear cell carcinoma.

BACKGROUND: Ovarian clear cell carcinoma (OCCC) is the second most common ovarian cancer after serous carcinoma in Japan. OCCC has a more unfavorable clinical outcome due to a poor response to platinum-based chemotherapy, compared with serous carcinoma. Hypoxia inducible factor-1α (HIF-1α) is a key regulator of cellular response to hypoxia and plays an important role in tumor growth, and HIF-1α gene single-nucleotide polymorphisms (SNPs) adversely affect the outcome in some cancers. Herein, we investigated the association of the HIF-1α gene SPNs with clinical outcome in OCCCs. Eighty-nine patients with OCCC were recruited in whom pathological diagnosis was confirmed with surgically resected specimen. RESULTS: The SNPs of C1772T and G1790A in the HIF-1α gene occurred in 23.6 and 3.3% of the patients, respectively. In the univariate analysis, overall survival was associated with stage and surgical residual tumor but not with the SNPs C1772T, G1790A, C1772T and/or G1790A. In the multivariate survival analysis, a significant association was observed between outcome and FIGO stage and/or surgical residual tumor; however, no association was obtained between HIF-1α gene SNPs and these factors. CONCLUSION: In conclusion, unlike the other cancers in which HIF-1α gene SNPs were demonstrated to be associated with the outcome, OCCC prognosis may not be affected by HIF-1α gene SNPs. Further studies need to be performed to clarify the association of HIF-1α expression with the unfavorable prognosis in OCCCs, in terms of transcriptional/translational activity, nuclear translocation of the protein, and protein degradation.

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 tumor's intrinsic aggressiveness, which may include the ER (-) and p53 (+) pattern as an independent poor prognostic factor.

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.

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

AIMS: 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. METHODS AND RESULTS: We evaluated immunohistochemically the expression levels of a panel of SLC transporters in 20 AFP-producing GCs and 130 conventional GCs. SLC transporters examined were human equilibrative nucleoside transporter 1 (hENT1), organic anion transporter 2 (OAT2), organic cation transporter (OCT) 2, OCT6 and organic anion-transporting polypeptide 1B3 (OATP1B3). The rates of high expression levels of hENT1 (hENT1high ) and OAT2 (OAT2high ) were statistically higher in AFP-producing GC, compared with conventional GC. When analysing hENT1 and OAT2 in combination, hENT1high /OAT2high was the most particular expression profile for AFP-producing GC, with a greater significance than hENT1 or OAT2 alone. However, no significant differences in OCT2, OCT6 or OATP1B3 levels were detected between AFP-producing and conventional GCs. However, immunoreactivity for hENT1, OAT2 and OCT6 tended to be increased in GC tissues compared with non-neoplastic epithelia. CONCLUSIONS: Because hENT1 and OAT2 are crucial for the uptake of gemcitabine and 5-fluorouracil, respectively, our results suggest that patients with AFP-producing GC could potentially benefit from gemcitabine/fluoropyrimidine combination chemotherapy. Increased expression of hENT1, OAT2 and OCT6 may also be associated with the progression of GC.

Organic cation transporter 2 for predicting cisplatin-based neoadjuvant chemotherapy response in gastric cancer.

Some studies have shown the usability of neoadjuvant chemotherapy (NAC) in gastric cancer (GC). Nevertheless there are a few predictive markers of the effectiveness of NAC in GC. The aim of this study is to assess the predictive impact of organic cation transporter 2 (OCT2) expression on response to neoadjuvant chemotherapy (NAC) in gastric cancer. We retrospectively assessed 66 patients with advanced gastric cancer received NAC with S-1/cisplatin or paclitaxel/cisplatin. Expression levels of OCT2 were assessed by immunohistochemistry in pre-chemotherapy biopsies and correlated with clinicopathologic parameters including pathologic response. High expression level of OCT2 (OCT2(high)) was significantly associated with intestinal type according to Laurén classification (P = 0.03) and low histologic grade (P = 0.03). In univariate analysis of the entire cohort, no variables showed any significant association with a response, although intestinal type (P = 0.09), low histologic grade (P = 0.09), and OCT2(high) (P = 0.07) tended to be more frequent in responders compared with non-responders. When the two treatment groups were separately assessed in the univariate analysis, a significantly higher rate of OCT2(high) was observed in responders compared with non-responders in the S-1/cisplatin group (P = 0.001). In addition, multivariate analysis identified OCT2(high) as the sole independent predictor of response (P = 0.04). However, in the paclitaxel/cisplatin group, no variables were associated with response. Taken together, our results suggest that OCT2(high) may represent a potential predictor of response to NAC with S-1/cisplatin in gastric cancer.

Impact of Expression Levels of Platinum-uptake Transporters Copper Transporter 1 and Organic Cation Transporter 2 on Resistance to Anthracycline/Taxane-based Chemotherapy in Triple-negative Breast Cancer.

Adding platinum drugs to anthracycline/taxane (ANC-Tax)-based neoadjuvant chemotherapy (NAC) improves pathological complete response (pCR) rates in triple-negative breast cancer (TNBC). Copper transporter 1 (CTR1) and organic cation transporter 2 (OCT2) critically affect the uptake and cytotoxicity of platinum drugs. We immunohistochemically determined CTR1 and OCT2 levels in pre-chemotherapy biopsies from 105 patients with HER2-negative breast cancer treated with ANC-Tax-based NAC. In the TNBC group, Ki-67(high) [pathological good response (pGR), P = 0.04] was associated with response, whereas CTR1(high) (non-pGR, P = 0.03), OCT2(high) (non-pGR, P = 0.01; non-pCR, P = 0.03), and combined CTR1(high) and/or OCT2(high) (non-pGR, P = 0.005; non-pCR, P = 0.003) were associated with non-response. In multivariate analysis, Ki-67(high) was an independent factor for pGR and CTR1 for non-pGR. Combined CTR1/OCT2 was a strong independent factor for non-pGR. However, no variables were associated with response in luminal BC. These results indicate that platinum uptake transporters are predominantly expressed in ANC-Tax-resistant TNBCs, which implies that advantage associated with adding platinum drugs may depend on high drug uptake.

Expression of organic anion-transporting polypeptide 1A2 and organic cation transporter 6 as a predictor of pathologic response to neoadjuvant chemotherapy in triple negative breast cancer.

Organic anion-transporting polypeptide 1A2 (OATP1A2) and organic cation transporter 6 (OCT6) are involved in the uptake of taxanes and anthracyclines, respectively. The aim of this study was to evaluate expression levels of OATP1A2 and OCT6 as a predictor of response to neoadjuvant chemotherapy (NAC) in breast cancer. A total of 124 patients who received anthracycline/taxane-based NAC were included. Expression levels of OATP1A2 and OCT6 were immunohistochemically assessed in core needle biopsies obtained prior to NAC. A pathologic good response (pGR) and a pathologic complete response (pCR) were achieved in 24 and 10 % of patients, respectively. In univariate analysis of the entire cohort, negative hormone receptor (HR) status (pGR and pCR, P < 0.001), high Ki-67 level (pGR, P = 0.03; pCR, P = 0.02), triple negative (TN) subtype (pGR, P = 0.001; pCR, P < 0.001), and high OCT6 (pGR, P = 0.003) were associated with the response. In combined analysis, high OATP1A2/high OCT6 level was also a significant factor for pGR (P = 0.001) and pCR (P = 0.001). Two separate multivariate analyses showed that HR status, TN subtype and combined high OATP1A2/high OCT6 level were significant independent predictors. When TN and non-TN tumors were assessed separately in univariate analysis, high Ki-67 level (P = 0.04) were associated with pGR and combined high OATP1A2/high OCT6 level was associated with both pGR (P = 0.005) and pCR (P = 0.03) in the TN group. Multivariate analysis identified the combined high OATP1A2/high OCT6 level as the sole independent predictor of pGR. In the non-TN group, negative HR status (P = 0.03) and positive HER2 status (P = 0.005) were associated with pGR, but HER2 status was the sole independent predictor of pGR. These results suggest that response-associated predictors may differ between the TN and non-TN tumors. Combined high OATP1A2/high OCT6 may be a potential predictor of response to anthracycline/taxane-based chemotherapy in breast cancer, especially in TN tumors.

Prognostic value of organic anion transporting polypeptide 1B3 and copper transporter 1 expression in endometrial cancer patients treated with paclitaxel and carboplatin.

Paclitaxel and carboplatin (TC) chemotherapy is an effective and well-tolerated regimen against advanced endometrial cancer. Organic anion transporting polypeptide 1B3 (OATP1B3) and copper transporter 1 (CTR1) are critical for the uptake of paclitaxel and carboplatin, respectively. This study aimed to address the prognostic impact of OATP1B3 and CTR1 in endometrial cancer patients treated with adjuvant TC chemotherapy. We immunohistochemically evaluated the expressions of OATP1B3 and CTR1 in 47 stage III endometrial cancers. The high expression levels of OATP1B3 were significantly correlated with type I tumor (P = 0.0005). In univariate analysis, high expression levels of OATP1B3 (P = 0.047) and CTR1 (P = 0.009) were significantly associated with longer disease-free survival (DFS) and longer overall survival (OS), respectively. The patients with tumors showing high expression levels of at least one of OATP1B3 and CTR1 had potentially longer DFS (P = 0.058) and significantly longer OS (P = 0.003) sin the univariate analysis. Combined OATP1B3/CTR1 expression was the sole independent prognostic factor for longer OS in the multivariate analysis (P = 0.013). Our findings suggest that combined OATP1B3/CTR1 expression is a possible predictive/prognostic factor for a good outcome in stage III endometrial cancer patients treated with adjuvant TC chemotherapy.

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

BACKGROUND: The 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 METHODS: Eight 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. RESULTS: The 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. CONCLUSIONS: Endometrial 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 SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1651876760876449.

Diagnostic accuracy of fine-needle aspiration cytology of the breast in Japan: report from the Working Group on the Accuracy of Breast Fine-Needle Aspiration Cytology of the Japanese Society of Clinical Cytology.

The Working Group of the Japanese Society of Clinical Cytology was assembled to assess the current status of breast cytology in Japan by conducting a large-scale survey regarding the accuracy of fine-needle aspiration biopsy (FNAB) in Japan. We collected data and investigated the status of breast cytological diagnosis at 12 different cooperating facilities in Japan, and re-evaluated their false-negative and false-positive cases. Among 30,535 individuals who underwent a breast cytological examination, analyses were conducted on 10,890 individuals (35.7%) in whom cytological diagnoses were confirmed by histology. Among these patients, the cytological diagnosis had an inadequate rate of 17.7%, an indeterminate rate of 7.8%, a positive predictive value of 'malignancy suspected' cells of 92.4%, an absolute sensitivity of 76.7%, a complete sensitivity of 96.7%, a specificity of 84.3%, a positive predictive value of 'malignant' cells of 99.5%, a false-negative value of 3.31%, a false-positive value of 0.25% and an accuracy rate of 88.0%. Subsequently, 297 false-negative and 23 false-positive cases were re-evaluated and several factors were characterized (i.e. histological type, tumor size and misread points). This survey collected data from a large number of cases for breast FNAB. Based on our survey, the accuracy of FNAB in Japan was relatively high compared with the goal of assessment of diagnostic accuracy. However, there were some false-negative and false-positive cases. Improvements in accuracy resulting from the learning points in the present study will lead to more useful and reliable diagnostic tools in clinical practice.

Granulosa cell tumor with activated mTOR-HIF-1alpha-VEGF pathway.

The DNA-binding activity of hypoxia-inducible factor-1 alpha (HIF-1alpha) 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-1alpha 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-1alpha. 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-1alpha 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-1alpha and VEGF expressions decline. As a result, the expressions of p-mTOR, HIF-1alpha 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-1alpha-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.

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

Malignant tumors usually involve a relatively hypoxic state, which induces overexpression of hypoxia-inducible factor-1alpha (HIF-1alpha) to satisfactorily enable the tumor to survive. Thus, inhibition of the mammalian target of rapamycin (mTOR) pathway including HIF-1alpha 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-1alpha, 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-1alpha 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-1alpha and VEGF was examined on western blot. As a result, although mTOR expression remained unchangeable, expression of p-mTOR, HIF-1alpha 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.

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

BACKGROUND: Hypoxia-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-1alpha 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-1alpha 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. METHOD: HIF-1 level in the nucleus was analyzed using DNA binding assay, and HIF-1alpha 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. RESULTS: HIF-1 level (mg/ml) in the nucleus and HIF-1alpha 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-1alpha, while MUAs showed the lowest levels of both. Mucinous adenomas were higher in HIF-1 than MUAs. CONCLUSION: Hypoxic 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.

Glucose transporter-1 expression in the thyroid gland: clinicopathological significance for papillary carcinoma.

Glucose transporter-1 (GLUT-1) expression was immunohistochemically analysed in a total of 268 cases of thyroid gland disease, including 129 cases of papillary carcinoma (PC), 60 cases of follicular carcinoma (FC), 57 cases of follicular adenoma, and 22 cases of adenomatous goitre. Seventy-one percent (91/129) of PC cases showed GLUT-1 expression, semi-quantitatively evaluated as: +, 21 cases (16%); 2+, 37 cases (29%); 3+, 33 cases (26%); and negative, 38 cases (29%). These positive cases were divided into two groups: 'membrane-like' pattern in 24 cases (19%), and 'cytoplasm-predominance' pattern in 67 cases (52%). GLUT-1 expression was observed in 5% (3/60) of FC cases, but all follicular adenomas and adenomatous goitres were negative for GLUT-1 (PC vs. FC, p<0.0001). Membrane-like expression was observed more frequently in non-organ-confined PCs (pT4) than in organ-confined PCs (pT1, 2, and 3) (p=0.0056). Seventy-five percent (18/24) of PC cases showing membrane-like expression were non-organ-confined. The membrane-like pattern was observed more frequently in PCs with lymph node (LN) metastasis compared to those without (p=0.0036). Ninety-two percent (22/24) of PC cases showing the membrane-like pattern were not organ-confined. Semi-quantitative analysis of glut-1 mRNA by RT-PCR showed a tendency toward higher expression in PCs compared to FCs, follicular adenomas and adenomatous goitres, and the mRNA expression in PCs with a membrane-like pattern was higher than those showing cytoplasm-predominance. We concluded that: 1) GLUT-1 is immunohistochemically useful in distinguishing PC from FC and benign diseases; 2) GLUT-1 may play an important role in the advancement of PC and LN metastasis, and its membrane-like expression is of more clinical significance than the cytoplasm-predominance pattern; and 3) glut-1 mRNA expression corresponds with the immunohistochemical expression profile.