Cytoplasmic HAX1 Is an Independent Risk Factor for Breast Cancer Metastasis.

HAX1 is an antiapoptotic factor involved in the regulation of cell migration and calcium homeostasis, overexpressed in several cancers, including breast cancer. It has been suggested that HAX1 is also implicated in metastasis. Herein we report the results of meta-analysis of HAX1 expression, based on publicly available data, which confirms its significant overexpression in breast cancer and demonstrates copy number gain and prognostic value of HAX1 overexpression for metastatic relapse in ER+ tumors. IHC analysis reported here also reveals its significant overexpression in breast cancer samples from primary tumors, indicating significantly higher HAX1 protein levels in a group of patients who developed distant metastases in a disease course. Moreover, we demonstrate that HAX1 localization is important for the prediction of metastatic relapse and that cytoplasmic but not nuclear HAX1 is an independent risk factor for breast cancer metastasis.

Beer consumption negatively regulates hormonal reproductive status and reduces apoptosis in Leydig cells in peripubertal rats.

Beer is one of the most popular alcoholic beverages consumed by young people. Ethanol intake is associated with harmful effects to the reproductive system. Bioactive compounds present in beer may diminish the toxics effect of ethanol. However, there is still little knowledge about the effect of beer consumption on hormonal regulation of male reproduction in organisms exposed to alcohol after the peripubertal age. Therefore, the aim of this study was to determine the influence of beer intake on plasma reproductive hormones, immunolocalization of cleaved caspase-3 (casp-3), and the level of the neuronal isoform of nitric oxide synthase (nNOS) in Leydig cells (LCs) in adolescent male Wistar rats. The animals, beginning at the age of 30 days, drank beer (10% ethanol; B2 group [2 weeks' exposure] and B4 group [4 weeks' exposure]), 10% ethanol solution (CE2 group [2 weeks' exposure] and CE4 group [4 weeks' exposure]), or water (C2 group [2 weeks' exposure] and C4 group [4 weeks' exposure]). Rats drinking beer for 4 weeks showed higher phenolic acid intake compared to rats drinking beer for 2 weeks. Rats exposed to beer for 4 weeks showed decreased plasma levels of follicle-stimulating hormone (FSH) and 17ß-estradiol (E2) (3.173 ng/mL and 11.49 pg/mL, respectively), compared to the CE4 (5.293 ng/mL and 43.912 pg/mL, respectively) and the C4 groups (5.002 ng/mL and 41.121 pg mL, respectively). Expression of cleaved caspase-3 in LCs was lower in the B4 group rats, compared to the CE4 group rats (ID score: 1.676 vs. 2.190). No changes in nNOS expression were observed. Beer consumption revealed a similar negative effect on hormonal regulation of male reproductive function, but lower apoptosis in LCs may be beneficial for steroidogenic activity.

miR-7 expression in serous ovarian carcinomas.

BACKGROUND/AIM: miR-7 has recently been linked to cancer. Some miR-7 targets, including B-cell lymphoma 2 (BCL2) and epidermal growth factor receptor (EGFR), are involved in ovarian cancer (OC) pathogenesis. The majority of OCs display TP53 mutations, which are critically important for OC development. We aimed to study the expression level of miR-7 and of two of its postulated target genes, BCL2 and EGFR, in serous ovarian carcinomas of different TP53 status and tumour grade. MATERIALS AND METHODS: Gene and miR expression was assessed by real-time reverse transcription polymerase chain reaction in 45 clinical samples of low- (G1+G2) and high- (G3) grade primary serous OC with wild-type (wt) or mutated TP53, as well as in three OC cell lines, each representing a different TP53 status. The results obtained in patients with OC were analysed against their disease-free survival (DFS). RESULTS: In high-grade OC with TP53 mutations, the level of miR-7 expression significantly exceeded (by several fold) that in wtTP53 cancer (p<0.01). Within the wtTP53 tumour series, the level of miR-7 expression was significantly higher (by over 10-fold) in high-grade than in low-grade OC (p<0.01). miR-7 expression was not found to influence DFS. The differences in miR-7 expression depending on TP53 status found in clinical OC samples were not observed in OC cell lines. miR-7 overexpression correlated with diminished BCL2 expression, but there was no relationship between miR-7 and EGFR expression, neither in tumour samples nor in the cell lines. CONCLUSION: There is a link between miR-7 expression and TP53 status and tumour grade in serous OC. Molecular mechanisms of these relationships need to be elucidated. Of the two postulated miR-7 target genes we examined, BCL2, but not EGFR, seems to be a possible miR-7 target in OC.

Cyclooxygenase-2 immunohistochemical expression in serrated polyps of the colon.

AIM OF THE STUDY: Cyclooxygenase-2 (COX-2) expression has been observed in a substantial percentage of classical adenomas of the large bowel. The aim of the study was to assess and compare the expression of COX-2 in serrated polyps of the colon. MATERIAL AND METHODS: One hundred and nineteen serrated polyps were analyzed. There were 83 hyperplastic polyps (HP), 19 sessile serrated polyps (SSP) and 17 traditional serrated adenomas (TSA). COX-2 expression was assessed semi-quantitatively (0-2) and each lesion was fully characterized in terms of anatomical location, size, histology, age and sex of the patient. The general estimating equation (GEE) model with logit link was used in the statistical analysis. RESULTS: Epithelial expression of COX-2 was found in 85/119 serrated polyps (71.43%): 57/83 (68.67%) HP, 16/19 (84.21%) SSP, and 12/17 (70.59%) TSA. In HP and SSP it was predominantly of weak (49/83 HP, 12/19 SSP), whereas in TSA it was mainly of medium/strong intensity (8/17). The TSA category was associated with more frequent COX-2 expression (OR = 7.00, 95% CI: 1.49-32.88, p = 0.014) than HP, but such relation was not found for SSP vs. HP (p > 0.1). No associations between COX-2 expression and clinical parameters were found. CONCLUSIONS: Immunohistochemical COX-2 expression cannot serve as a diagnostic adjunct to differentiate HP and SSP.

p19(INK4d) mRNA and protein expression as new prognostic factors in ovarian cancer patients.

p19(INK4d) (CDKN2D) is a negative regulator of the cell cycle. Little is known of its role in cancer development and prognosis. We aimed to evaluate the clinical significance of p19(INK4d) expression in ovarian carcinomas with respect to the TP53 accumulation status, as well as the frequency of CDKN2D mutations. p19(INK4d) and TP53 expression was evaluated immunohistochemically in 445 ovarian carcinomas: 246 patients were treated with platinum-cyclophosphamide (PC/PAC), while 199 were treated with taxane-platinum agents (TP). CDKN2D gene expression (mRNA) was examined in 106 carcinomas, while CDKN2D mutations in 68 tumors. Uni- and multivariate statistical analyses (logistic regression and the Cox proportional hazards model) were performed for patient groups divided according to the chemotherapeutic regimen administered, and in subgroups with and without TP53 accumulation. High p19(INK4d) expression increased the risk of death, but only in patients with the TP53-negative carcinomas (HR 1.61, P = 0.049 for PC/PAC-treated patients, HR 2.00, P = 0.015 for TP-treated patients). This result was confirmed by the mRNA analysis (HR 4.24, P = 0.001 for TP-treated group). High p19(INK4d) protein expression associated with adverse clinicopathological factors. We found no alterations in the CDKN2D gene; the c.90C>G (p.R30R; rs1968445) polymorphism was detected in 10% of tumors. Our results suggest that p19(INK4d) expression is a poor prognostic factor in ovarian cancer patients. Analyses of tumor groups according to the TP53 accumulation status facilitate the identification of cancer biomarkers.

Nuclear survivin expression is a positive prognostic factor in taxane-platinum-treated ovarian cancer patients.

BACKGROUND: Survivin is an inhibitor of apoptosis and a regulator of mitotic progression. TP53 protein is a negative transcriptional regulator of survivin. The aim of our study was to evaluate the clinical significance of survivin expression in advanced stages ovarian cancer with respect to the TP53 status. METHODS: Survivin and TP53 expression was evaluated immunohistochemically in 435 archival samples of ovarian carcinomas (244 patients were treated with platinum/cyclophosphamide-PC/PAC; 191-with taxane-platinum (TP) agents). Univariate and multivariate statistical analyses were performed in patients groups divided according to the administered chemotherapeutic regimen, and in subgroups with and without TP53 accumulation (TP53+ and TP53-, respectively). RESULTS: Nuclear and cytoplasmic survivin expression was observed in 92% and 74% of the carcinomas, respectively. In patients treated with TP, high nuclear survivin expression decreased the risk of disease recurrence and death, and increased the probability of high platinum sensitivity (p < 0.01), but only in the TP53(+) group, and not in the TP53(-) group. CONCLUSIONS: It appears that TP53 status determines the clinical importance of nuclear survivin expression in taxane-platinum treated ovarian cancer patients.

HAX-1 overexpression, splicing and cellular localization in tumors.

BACKGROUND: HAX-1 has been described as a protein potentially involved in carcinogenesis and especially metastasis. Its involvement in regulation of apoptosis and cell migration along with some data indicating its overexpression in cancer cell lines and tumors suggests that HAX-1 may play a role in neoplastic transformation. Here we present the first systematic analysis of HAX-1 expression in several solid tumors. METHODS: Using quantitative RT-PCR, we have determined the mRNA levels of HAX1 splice variant I in several solid tumors. We have also analyzed by semiquantitative and quantitative RT-PCR the expression of five HAX-1 splice variants in breast cancer samples and in normal tissue from the same individuals. Quantitative PCR was also employed to analyze the effect of estrogen on HAX1 expression in breast cancer cell line. Immunohistochemical analysis of HAX-1 was performed on normal and breast cancer samples. RESULTS: The results reveal statistically important HAX1 up-regulation in breast cancer, lung cancer and melanoma, along with some minor variations in the splicing pattern. HAX-1 up-regulation in breast cancer samples was confirmed by immunohistochemical analysis, which also revealed an intriguing HAX-1 localization in the nuclei of the tumor cells, associated with strong ER status. CONCLUSION: HAX-1 elevated levels in cancer tissues point to its involvement in neoplastic transformation, especially in breast cancer. The connection between HAX-1 nuclear location and ER status in breast cancer samples remains to be clarified.

PIK3CA amplification associates with resistance to chemotherapy in ovarian cancer patients.

PI3K/AKT signalling pathway controls important cellular processes such as the cell proliferation and apoptosis. PIK3CA gene encoding a catalytic subunit of the PI3K is mutated and/or amplified in various neoplasms, including ovarian cancer. We aimed to evaluate PIK3CA alterations and their clinical importance in ovarian cancer patients. Molecular analysis was performed on 117 ovarian carcinomas with the use of qPCR, SSCP and sequencing. In a group of 98 patients with complete clinical data, 62 patients were treated with standard taxane-platinum regimens and 36 patients with platinum-cyclophosphamide regimens. A multivariate analysis was performed by the Cox's and logistic regression models. PIK3CA mutations occurred in 5/117 (4.3%) carcinomas, exclusively in the endometrioid and clear cell types (p = 0.0002); they were also associated with low FIGO stage (p = 0.0003), low tumor grade (p = 0.045) and early patient's age at diagnosis (p = 0.0005). The PIK3CA amplification (predominantly a low-level) was found in 28/117 (24%) ovarian carcinomas. It was more frequent in TP53 mutant tumors (p = 0.012) and tended to associate with high pAKT expression (p = 0.061). The PIK3CA amplification strongly diminished odds of complete remission (OR = 0.25, p = 0.033) and platinum sensitive response (PS, OR = 0.12, p = 0.004) in the taxane-platinum treated patients. The odds of PS were also much lower in all patients with the PIK3CA amplification evaluated together, regardless of the treatment applied (OR = 0.18, p = 0.001). Our results suggest that PIK3CA amplification may be a marker predicting ovarian cancer response to chemotherapy.

TP53, BCL-2 and BAX analysis in 199 ovarian cancer patients treated with taxane-platinum regimens.

OBJECTIVE: In cell line studies, BCL-2 and BAX proteins interfere with cancer response to taxanes. This issue has not received much attention with regard to taxane-platinum (TP)-treated ovarian cancer patients. METHODS: We evaluated prognostic/predictive significance of BCL-2 and BAX with regard to TP53 status. Immunohistochemical analysis was performed on 199 ovarian carcinomas FIGO stage IIB-IV treated with TP; the results were analyzed by the Cox and logistic regression models. RESULTS: Clinicopathological parameters (residual tumor size, FIGO stage and/or tumor grade, but not patient's age) were the only or the strongest predictors of patient's outcome. Platinum highly sensitive response showed a positive association with TP53 accumulation (p=0.045). As in our previously published analysis on platinum-cyclophosphamide-treated group, complete remission showed a borderline negative (paradoxic) association with high BAX expression in the whole group (p=0.058) and with BCL-2 expression in the TP53(-) group (p=0.058). CONCLUSION: Our results suggest that TP53, BCL-2 and BAX proteins carry some predictive potential in taxane-platinum-treated ovarian cancer patients, auxiliary to clinicopathological factors. We have confirmed on another patient group that clinical importance of BCL-2 may depend on TP53 status.

P27(Kip1) and Ki-67 expression analysis in transitional cell carcinoma of the bladder.

P27(Kip1) protein is a cell cycle inhibitor which blocks the transition of cells from G1 to S phase, while Ki-67 is the most specific marker of proliferative activity. Both proteins are independent predictors of clinical outcome in various neoplasms. The aim of the study was to assess the prognostic value of p27(Kip1) and Ki-67 expression in urothelial bladder tumours. P27(Kip1) and Ki-67 expressions were evaluated immunohistochemically in archival samples of 45 superficial and 26 invasive transitional cell carcinomas obtained by a transurethral resection. In the patients with superficial tumours, disease-free survival (DFS) was positively influenced by good histological differentiation as well as by concurrent high p27(Kip1) and low Ki-67 expression. Multivariate analysis has confirmed that tumour grade and p27(Kip1)/Ki-67 status were independent predictors of DFS ( p=0.028 and p=0.029, respectively). P27(Kip1) or Ki-67 expressions did not influence overall survival. We conclude that a variable combined of p27(Kip1) and Ki-67 expressions is a better predictor of DFS in superficial bladder tumours than either protein alone.