Agneta Jansson

17β-Hydroxysteroid Dehydrogenase 14 Affects Estradiol Levels in Breast Cancer Cells and Is a Prognostic Marker in Estrogen Receptor–Positive Breast Cancer

Estrogens have an important role in the progression of breast cancer. The 17β-hydroxysteroid dehydrogenase (17HSD) family has been identified to be of significance in hormone-dependent tissues. 17HSD1 and 17HSD2 are the main 17HSD enzymes involved in breast cancer investigated this far, but it is possible that other hormone-regulating enzymes have a similar role. 17HSD5 and 17HSD12 are associated with sex steroid metabolism, and 17HSD14 is a newly discovered enzyme that may be involved in the estrogen balance. The mRNA expression of 17HSD5, 17HSD12, and 17HSD14 were analyzed in 131 breast cancer specimens by semiquantitative real-time PCR. The results were compared with recurrence-free survival and breast cancer-specific survival of the patients. The breast cancer cell lines MCF7, SKBR3, and ZR75-1 were transiently transfected with 17HSD14 to investigate any possible effect on estradiol levels. We found that high 17HSD5 was related to significantly higher risk of late relapse in estrogen receptor (ER)–positive patients remaining recurrence-free later than 5 years after diagnosis ( P = 0.02). No relation to 17HSD12 expression was found, indicating that 17HSD12 is of minor importance in breast cancer. Patients with ER-positive tumors with high expression levels of 17HSD14 showed a significantly better prognosis about recurrence-free survival ( P = 0.008) as well as breast cancer-specific survival ( P = 0.01), confirmed by multivariate analysis ( P = 0.04). Transfection of 17HSD14 in the human breast cancer cells MCF7 and SKBR3 significantly decreased the levels of estradiol, presenting an effect of high expression levels of the enzyme. (Cancer Res 2006; 66(23): 11471-7)

Decreased tumor cell proliferation as an indicator of the effect of preoperative radiotherapy of rectal cancer

BACKGROUND Rectal cancer is a common malignancy, with significant local recurrence and death rates. Preoperative radiotherapy and refined surgical technique can improve local control rates and disease-free survival. PURPOSE To investigate the relationship between the tumor growth fraction in rectal cancer measured with Ki-67 and the outcome, with and without short-term preoperative radiotherapy. METHOD Ki-67 (MIB-1) immunohistochemistry was used to measure tumor cell proliferation in the preoperative biopsy and the surgical specimen. MATERIALS Specimens from 152 patients from the Southeast Swedish Health Care region were included in the Swedish rectal cancer trial 1987-1990. RESULTS Tumors with low proliferation treated with preoperative radiotherapy had a significantly reduced recurrence rate. The influence on death from rectal cancer was shown only in the univariate analysis. Preoperative radiotherapy of tumors with high proliferation did not significantly improve local control and disease-free survival. The interaction between Ki-67 status and the benefit of radiotherapy was significant for the reduced recurrence rate (p = 0.03), with a trend toward improved disease-free survival (p = 0.08). In the surgery-alone group, Ki-67 staining did not significantly correlate with local recurrence or survival rates. CONCLUSION Many Ki-67 stained tumor cells in the preoperative biopsy predicts an increased treatment failure rate after preoperative radiotherapy of rectal cancer.

17Beta-hydroxysteroid dehydrogenase enzymes and breast cancer ☆

Sex steroids play an important role in the development and differentiation in several tissues. Biologically active hormones that are locally converted in endocrine organs in the tissue where they exert their effects without release into extracellular space is a field of endocrinology that has been called intracrinology. In pre-menopausal women the ovary is the main source of estrogens, but in post-menopausal women the estrogen production as main site of synthesis moves to peripheral tissues and almost all of the sex steroids are synthesised from precursors of adrenal origin. In breast cancer 60-80% of the tumors express high levels of oestrogen receptor (ER) alpha which gives estrogen a proliferative effect. Breast tumors tend to have a higher intratumoral estrogen concentration than normal breast tissue and plasma, and in situ synthesis and the metabolism of estrogens is believed to be of great importance for the development and progression of the disease. The activity of estrogen metabolizing enzymes in breast are mainly aromatase, estrone sulfatases and 17HSD enzymes. 17HSD1 and 17HSD2 are the family members known to be of main importance in breast cancer. High expression of 17HSD1 has been associated to poor prognosis in breast cancer and late relapse among patients with ER-positive tumors. One of the mechanisms behind high 17HSD1 expression is gene amplification. Low or absent expression of 17HSD2 is associated to decreased survival in ER-positive breast cancer. 17HSD14 is one of the latest discovered 17HSD enzymes, transfection of 17HSD14 in human breast cancer cells significantly decreased the levels of estradiol in the culture medium. Low expression of 17HSD14 mRNA expression in breast cancer was correlated to decreased survival. The understanding of intratumoral synthesis of sex steroids in breast cancer is crucial to understand the disease both in pre- and post-menopausal women. Further studies are desirable to state the direct role of these enzymes in breast cancer and which patients that may benefit from new therapeutic strategies targeting 17HSD enzymes. The new inhibitors targeting 17HSD1 have shown promising results in pre-clinical studies to have clinical potential in the future.

Ratio of 17HSD1 to 17HSD2 Protein Expression Predicts the Outcome of Tamoxifen Treatment in Postmenopausal Breast Cancer Patients

Purpose: Estrogens have great significance in the development of breast cancer. After menopause, most estrogen biosynthesis is done in peripheral tissue, and the main enzymes involved in balancing the amount of estrone against estradiol are 17β-hydroxysteroid dehydrogenases (17HSD). The aim of this study was to investigate the prognostic and tamoxifen predictive values of 17HSD1 and 17HSD2 expression. Experimental Design: Tumors from low-risk breast cancer patients randomized to adjuvant tamoxifen therapy or no adjuvant treatment were analyzed with immunohistochemistry to investigate protein expression of 17HSD1 and 17HSD2 in 912 cases. All patients had lymph node-negative breast cancer and were postmenopausal at the time of diagnosis. Results: Low 17HSD1 expression was associated with significant benefit from tamoxifen treatment among patients with estrogen receptor (ER)-positive tumors (P < 0.001). For patients with a 17HSD1 score not exceeding that of 17HSD2, tamoxifen increased the rate of distant recurrence-free survival (hazard ratio, 0.37; 95% confidence interval, 0.23-0.60) and breast cancer-specific survival (hazard ratio, 0.30; 95% confidence interval, 0.16-0.54), whereas no apparent effect was observed when the 17HSD1 score was higher than that of 17HSD2. The interaction was significant for both distant recurrence-free survival (P = 0.036) and breast cancer-specific survival (P = 0.014). In the cohort of systemically untreated patients, no prognostic importance was observed. Conclusions: This is the first report that clearly distinguishes between the prognostic and predictive importance of 17HSD1 and 17HSD2 in ER-positive breast cancer treated with or without tamoxifen. Our data suggest that the 17HSD1/17HSD2 ratio might be useful as a predictive factor for tamoxifen treatment in ER-positive breast cancer patients.

Heat shock protein 72/73 in relation to cytoplasmic p53 expression and prognosis in colorectal adenocarcinomas

Heat shock proteins (hsp) are molecular chaperones that are increased by various environmental and patho‐physiological stimuli. Hsp can bind to mutant/wild‐type p53 in tumors and, consequently, could not only regulate p53 accumulation or localization but also modulate its biological effects on cells. However, there is little information available on the significance of hsp expression in colorectal cancer. The aim of our study was to investigate the relationship of hsp to p53 expression, clinico‐pathological factors and prognosis in a series of 256 patients with colorectal adenocarcinomas, using immuno‐histochemistry. Seventy‐five cases exhibited hsp expression in the cytoplasm, with 11 presenting both cytoplasmic and nuclear staining. Hsp expression was related positively to cytoplasmic p53 expression but not to nuclear p53 expression. In the subgroup of rectal tumors, hsp over‐expression appeared to predict unfavorable survival, though its prognostic value diminished using multivariate analysis. There were no significant relationships of hsp with patient sex or age, tumor site, Dukes stage, growth pattern or differentiation. Int. J. Cancer 74:600–604, 1997.© 1997 Wiley‐Liss, Inc.

YAP1 is a potential biomarker for cetuximab resistance in head and neck cancer

OBJECTIVES Targeted therapy against the epidermal growth factor receptor (EGFR) only variably represents a therapeutic advance in head and neck squamous cell carcinoma (HNSCC). This study addresses the need of biomarkers of treatment response to the EGFR-targeting antibody cetuximab (Erbitux®). MATERIALS AND METHODS The intrinsic cetuximab sensitivity of HNSCC cell lines was assessed by a crystal violet assay. Gene copy number analysis of five resistant and five sensitive cell lines was performed using the Affymetrix SNP 6.0 platform. Quantitative real-time PCR was used for verification of selected copy number alterations and assessment of mRNA expression. The functional importance of the findings on the gene and mRNA level was investigated employing siRNA technology. The data was statistically evaluated using Mann-Whitney U-test and Spearmans correlation test. RESULTS Analysis of the intrinsic cetuximab sensitivity of 32 HNSCC cell lines characterized five and nine lines as cetuximab sensitive or resistant, respectively. Gene copy number analysis of five resistant versus five sensitive cell lines identified 39 amplified protein-coding genes, including YAP1, in the genomic regions 11q22.1 or 5p13-15. Assessment using qPCR verified that YAP1 amplification associated with cetuximab resistance. Amplification of YAP1 correlated to higher mRNA levels, and RNA knockdown resulted in increased cetuximab sensitivity. Assessment of several independent clinical data sets in the public domain confirmed YAP1 amplifications in multiple tumor types including HNSCC, along with highly differential expression in a subset of HNSCC patients. CONCLUSION Taken together, we provide evidence that YAP1 could represent a novel biomarker gene of cetuximab resistance in HNSCC cell lines.

Noxa in colorectal cancer: a study on DNA, mRNA and protein expression.

Noxa is a BH3-only member of the Bcl-2 family, upregulated by p53 as a response to DNA damage. Mutations in the BH3-only region of other BH3-only members lead to an inactive protein. We have investigated the mRNA expression of Noxa with real-time PCR in 94 unselected colorectal adenocarcinomas and the corresponding normal mucosa. Among them, Noxa protein expression was investigated with immunohistochemistry in 16 tumors and six corresponding normal mucosa samples. Further, we searched for Noxa mutations in all the cases using single-stranded conformation polymorphism and DNA sequencing. The mRNA expression of Noxa was weak in 9% and strong in 2% of the tumors, and decreased in 9% and increased in 16% of the tumors compared with the normal mucosa; however, these changes did not have any clinical or pathological significance. The protein level in most of the cases investigated was correlated with the mRNA level. We did not find any mutations in the Noxa gene. Thus, we suggest that Noxa may not be of importance in the development of colorectal cancer.

Predictive relevance of HOXB13 protein expression for tamoxifen benefit in breast cancer

IntroductionThe HOXB13:IL17BR index has been identified to predict clinical outcome in the setting of adjuvant tamoxifen monotherapy of breast cancer. Further studies have shown that HOXB13 in particular can indicate benefit of prolonged tamoxifen treatment. Patients with high-expressing tumors did not benefit from prolonged treatment, suggesting that HOXB13 might be involved in tamoxifen resistance. No studies have been made regarding the HOXB13 protein levels in breast cancer. The aim of our study was to investigate whether tamoxifen benefit can be correlated to different levels of HOXB13 protein expression.MethodsWe used immunohistochemistry to analyze protein levels of HOXB13 in tumor samples from 912 postmenopausal node-negative breast cancer patients randomized to adjuvant tamoxifen therapy or no endocrine treatment.ResultsTamoxifen-treated patients with estrogen receptor-positive tumors expressing none or low levels of HOXB13 had a clear benefit from tamoxifen in terms of longer distant recurrence-free survival (DRFS) (hazard ratio = 0.38, 95% confidence interval = 0.23 to 0.60, P = 0.000048). However, for patients with a high or intermediate HOXB13 tumor expression, tamoxifen did not prolong the DRFS compared with the untreated patients (hazard ratio = 0.88, 95% confidence interval = 0.47 to 1.65, P = 0.69). Interaction between HOXB13 expression and benefit from tamoxifen was statistically significant for DRFS (P = 0.035). No prognostic value could be ascribed to HOXB13 among systemically untreated patients.ConclusionsA high HOXB13 expression was associated with decreased benefit from tamoxifen, which indicates that HOXB13 protein level may be used as a predictive marker for tamoxifen treatment.

p53 Mutations are present in colorectal cancer with cytoplasmic p53 accumulation

Previous studies have shown that nuclear p53 over‐expression is an indicator of p53 mutations whereas cytoplasmic p53 accumulation is related to wild‐type p53 in several kinds of tumors. Cytoplasmic p53 accumulation has been demonstrated to be an independent prognostic factor in colorectal adenocarcinomas. The purpose was to examine whether mutations occur in cases with p53 accumulated in the cytoplasm and whether there are any differences in the frequency and characteristics of p53 mutations in different staining patterns. In the present study, we identified p53 mutations using PCR single‐strand conformation polymorphism (SSCP) and DNA sequencing in 75 primary colorectal adenocarcinomas with different staining patterns (negative, nucleus, cytoplasm, nucleus and cytoplasm). The results show that the frequency and nature of mutations in tumors with cytoplasmic p53 accumulation were similar to those with nuclear p53 expression. However, the tumors with accumulation in both the nucleus and cytoplasm demonstrated a higher mutation rate. We suppose that the role of cytoplasmic p53 accumulation in predicting prognosis in patients with colorectal cancer may be dependent on both mutational and non‐mutational mechanisms.

Androgen receptor expression predicts beneficial tamoxifen response in oestrogen receptor-α-negative breast cancer.

Background:Although the androgen receptor (AR) is frequently expressed in breast cancer, its relevance in the disease is not fully understood. In addition, the relevance of AR in determining tamoxifen treatment efficiency requires evaluation.Purpose:To investigate the tamoxifen predictive relevance of the AR protein expression in breast cancer.MethodsPatients were randomised to tamoxifen 40 mg daily for 2 or 5 years or to no endocrine treatment. Mean follow-up was 15 years. Hazard ratios were calculated with recurrence-free survival as end point.Results:In patients with oestrogen receptor (ER)-negative tumours, expression of AR predicted decreased recurrence rate with tamoxifen (hazard ratio (HR)=0.34; 95% confidence interval (CI)=0.14–0.81; P=0.015), whereas the opposite was seen in the AR− group (HR=2.92; 95% CI=1.16–7.31; P=0.022). Interaction test was significant P<0.001. Patients with triple-negative and AR+ tumours benefitted from tamoxifen treatment (HR=0.12; 95% CI=0.014–0.95 P=0.044), whereas patients with AR− tumours had worse outcome when treated with tamoxifen (HR=3.98; 95% CI=1.32–12.03; P=0.014). Interaction test was significant P=0.003. Patients with ER+ tumours showed benefit from tamoxifen treatment regardless of AR expression.Conclusions:AR can predict tamoxifen treatment benefit in patients with ER− tumours and triple-negative breast cancer.