Anna Sankila

Dual role of FoxA1 in androgen receptor binding to chromatin, androgen signalling and prostate cancer

High androgen receptor (AR) level in primary tumour predicts increased prostate cancer‐specific mortality. However, the mechanisms that regulate AR function in prostate cancer are poorly known. We report here a new paradigm for the forkhead protein FoxA1 action in androgen signalling. Besides pioneering the AR pathway, FoxA1 depletion elicited extensive redistribution of AR‐binding sites (ARBs) on LNCaP‐1F5 cell chromatin that was commensurate with changes in androgen‐dependent gene expression signature. We identified three distinct classes of ARBs and androgen‐responsive genes: (i) independent of FoxA1, (ii) pioneered by FoxA1 and (iii) masked by FoxA1 and functional upon FoxA1 depletion. FoxA1 depletion also reprogrammed AR binding in VCaP cells, and glucocorticoid receptor binding and glucocorticoid‐dependent signalling in LNCaP‐1F5 cells. Importantly, FoxA1 protein level in primary prostate tumour had significant association to disease outcome; high FoxA1 level was associated with poor prognosis, whereas low FoxA1 level, even in the presence of high AR expression, predicted good prognosis. The role of FoxA1 in androgen signalling and prostate cancer is distinctly different from that in oestrogen signalling and breast cancer.

Long-term Efficacy of Maintenance Bacillus Calmette-Guérin versus Maintenance Mitomycin C Instillation Therapy in Frequently Recurrent TaT1 Tumours without Carcinoma In Situ: A Subgroup Analysis of the Prospective, Randomised FinnBladder I Study with a 20-Year Follow-up

BACKGROUND The long-term prospective data on bacillus Calmette-Guérin (BCG) and mitomycin C (MMC) instillation therapy are limited. OBJECTIVE To compare the long-term benefit of BCG and MMC maintenance therapy in patients with recurrent bladder carcinoma. DESIGN, SETTING, AND PARTICIPANTS Eighty-nine patients with frequently recurrent TaT1 disease without carcinoma in situ (CIS) were eligible. Originally, the patients were enrolled in the prospective FinnBladder I study between 1984 and 1987 and randomised to receive BCG or MMC. Both regimens involved five weekly instillations, followed by monthly instillations for 2 yr. Because of alkalinising the urine and adjusting the dose to bladder capacity, the average concentration of MMC was low: 30-40 mg in 150-200 ml of phosphate buffer. Overall median follow-up time was 8.5 yr, whereas the median follow-up time of the patients who were still alive was 19.4 yr. MEASUREMENTS Primary end points were time to first recurrence and overall mortality. Secondary end points were progression and disease-specific mortality. RESULTS AND LIMITATIONS Thirty-six of 45 patients (80.0%) in the MMC group experienced recurrence in contrast to 26 of 44 patients (59.1%) in the BCG group. This finding was reflected in significantly lower cumulative incidence estimates in the BCG group (p=0.005). There was a weak trend for fewer progressions (p=0.1) and cancer-specific deaths (p=0.2) in the cumulative incidence analysis, as 4 patients versus 10 patients progressed and 4 patients versus 9 patients died from the disease in the BCG group versus the MMC group, respectively. No difference existed in the overall mortality. The study population, however, was too small for conclusive evidence about progression or survival. CONCLUSIONS An intensive intravesical BCG immunotherapy results in a sustained and significant long-term reduction in recurrence in frequently recurrent bladder carcinoma. The relatively low progression rate during the long follow-up suggests that it may be difficult to show significant differences in overall mortality with a substantially larger but otherwise similar study population. TRIAL REGISTRATION Registration was not considered to be necessary at this stage of the follow-up because the study was initiated as early as 1984 and the last randomisation took place in July 1987, that is, long before the current requirements concerning study registrations were implemented.

Systematic knockdown of epigenetic enzymes identifies a novel histone demethylase PHF8 overexpressed in prostate cancer with an impact on cell proliferation, migration and invasion

Our understanding of key epigenetic regulators involved in specific biological processes and cancers is still incomplete, despite great progress in genome-wide studies of the epigenome. Here, we carried out a systematic, genome-wide analysis of the functional significance of 615 epigenetic proteins in prostate cancer (PrCa) cells. We used the high-content cell-spot microarray technology and siRNA silencing of PrCa cell lines for functional screening of cell proliferation, survival, androgen receptor (AR) expression, histone methylation and acetylation. Our study highlights subsets of epigenetic enzymes influencing different cancer cell phenotypes. Plant homeo domain (PHD) finger proteins have a key role in cell survival and histone methylation, whereas histone deacetylases were primarily involved in regulating AR expression. In contrast, JumonjiC-domain (JmjC) containing histone lysine demethylases (KDMs) mainly had an impact on cell proliferation. Our results show that the KDMs JARID1B, PHF8, KDM3A, KDM3B and KDM4A were highly expressed in clinical PrCa samples. The PHD-finger protein 8 (PHF8), a transcriptional coactivator with both PHD- and JmjC-domains, was moderately to strongly expressed in 80% of clinical PrCa samples, whereas 76% of normal and benign samples were negative or only showed weak PHF8 expression. Strong PHF8 expression correlated significantly with high Gleason grade and was borderline significant for poor prognosis. The results of functional PHF8 knockdown implicate a role in cell migration and invasion, as shown by cell motility and 3-D invasion assays. Our study suggests that various cellular phenotypes are regulated by distinct subsets of epigenetic enzymes. Proteins interpreting and modifying histone methylation, such as JmjC-domain and particularly PHD-finger proteins like PHF8, are activated in subsets of PrCas and promote cancer relevant phenotypes.

DNA Damage Recognition via Activated ATM and p53 Pathway in Nonproliferating Human Prostate Tissue

DNA damage response (DDR) pathways have been extensively studied in cancer cell lines and mouse models, but little is known about how DNA damage is recognized by different cell types in nonmalignant, slowly replicating human tissues. Here, we assess, using ex vivo cultures of human prostate tissue, DDR caused by cytotoxic drugs (camptothecin, doxorubicin, etoposide, and cisplatin) and ionizing radiation (IR) in the context of normal tissue architecture. Using specific markers for basal and luminal epithelial cells, we determine and quantify cell compartment-specific damage recognition. IR, doxorubicin, and etoposide induced the phosphorylation of H2A.X on Ser(139) (γH2AX) and DNA damage foci formation. Surprisingly, luminal epithelial cells lack the prominent γH2AX response after IR when compared with basal cells, although ATM phosphorylation on Ser(1981) and 53BP1 foci were clearly detectable in both cell types. The attenuated γH2AX response seems to result from low levels of total H2A.X in the luminal cells. Marked increase in p53, a downstream target of the activated ATM pathway, was detected only in response to camptothecin and doxorubicin. These findings emphasize the diversity of pathways activated by DNA damage in slowly replicating tissues and reveal an unexpected deviation in the prostate luminal compartment that may be relevant in prostate tumorigenesis. Detailed mapping of tissue and cell type differences in DDR will provide an outlook of relevant responses to therapeutic strategies.

Treatment of prostate cancer with Ad5/3Δ24hCG allows non-invasive detection of the magnitude and persistence of virus replication in vivo

Hormone refractory metastatic prostate cancer is a deadly disease that currently lacks curative treatments. Conditionally replicating adenoviruses (CRAds) are promising new agents against cancer due to their innate capability to cause oncolysis of tumor cells. Their antitumor effect is determined in part by their capacity for infecting cancer cells. However, the respective primary receptor, the coxsackie-adenovirus receptor (CAR), is variably expressed in many cancer types. We created Ad5/3Δ24hCG, a novel CRAd retargeted to the adenovirus serotype 3 receptor, which has been reported to be highly expressed in tumors. Furthermore, we added a transgene for the β-chain of human chorionic gonadotropin (hCGβ), whose expression was tightly coupled to virus replication. Ad5/3Δ24hCG was found effective in killing prostate cancer cells, and oncolysis was seen in concordance with hCGβ production. In a s.c. in vivo model of hormone refractory prostate cancer, Ad5/3Δ24hCG treatment resulted in statistically significant tumor growth inhibition. Moreover, i.v. injection of Ad5/3Δ24hCG prolonged the survival of mice with hormone refractory prostate cancer metastatic to the lung. Detection of hCGβ in serum samples confirmed viral replication in vivo. Infection of human clinical samples of cancerous and normal prostatic tissue resulted in effective hCGβ production in cancer tissue, whereas it remained low in nonmalignant tissue, suggesting cancer-specific replication. These results suggest that Ad5/3Δ24hCG is a potent virus for the treatment of hormone refractory prostate cancer in vitro and in vivo. These preclinical data set the stage for translation into clinical studies. [Mol Cancer Ther 2007;6(2):742–51]

PSA forms complexes with α1-antichymotrypsin in prostate.

PSA is the most useful prostate cancer marker. However, its levels are increased also in some non‐malignant conditions. In circulation, the majority of PSA is complexed with protease inhibitors, including α1‐antichymotrypsin (ACT). The proportion of the PSA‐ACT complex is higher in patients with prostate cancer than in controls without cancer. The expression of ACT has been shown to be higher in prostate cancer than in benign prostatic hyperplasia. However, results regarding the extent which PSA forms complexes within the prostate and whether there are differences in complex formation between normal and malignant prostatic tissue are inconsistent and limited.

High expression of antizyme inhibitor 2, an activator of ornithine decarboxylase in steroidogenic cells of human gonads

High activity of ornithine decarboxylase (ODC), the rate-limiting enzyme of polyamine synthesis, is typically present in rapidly proliferating normal and malignant cells. The mitotically inactive steroidogenic cells in rodent testis and ovaries, however, also display high ODC activity. The activity of ODC in these cells responds to luteinizing hormone, and inhibition of ODC reduces the production of steroid hormones. Polyamines and ODC also control proliferation of germ cells and spermiogenesis. The activity of ODC, especially in proliferating cells, is regulated by antizyme inhibitor (AZIN). This protein displaces ODC from a complex with its inhibitor, antizyme. We have previously identified and cloned a second AZIN, i.e. antizyme inhibitor 2 (AZIN2), which has the highest levels of expression in brain and in testis. In the present study, we have used immunohistochemistry and in situ hybridization to localize the expression of AZIN2 in human gonads. We found a robust expression of AZIN2 in steroidogenic cells: testicular Leydig cells and Leydig cell tumors, in ovarian luteinized cells lining corpus luteum cysts, and in hilus cells. The results suggest that AZIN2 is not primarily involved in regulating the proliferation of the germinal epithelium, indicating a different role for AZIN1 and AZIN2 in the regulation of ODC. The localization of AZIN2 implies possible involvement in the gonadal synthesis and/or release of steroid hormones.

Expression of human chorionic gonadotropin in testicular germ cell tumors

BACKGROUND We have shown that most patients with seminomas have elevated serum concentrations of the free β subunit of human chorionic gonadotropin (hCGβ) and that in nonseminomatous testicular cancer, most of the hCG in the serum is hyperglycosylated (hCG-h). However, the tissue expression of hCG-h or hCGβ in germ cell tumors (GCTs) has not been reported. Our objective was to study the expression and diagnostic value of hCG-h and hCGβ in testicular GCTs. METHODS We studied the immunohistochemical expression of hCG, hCG-h, hCGβ, and the free α subunit of hCG (hCGα) in GCTs from 154 patients. We compared the tissue expression with serum concentrations and evaluated the correlation between staining intensity, established prognostic variables, and outcome. RESULTS The expression varied between tumor types. All forms of hCG, including hCG-h, were detected in embryonal carcinomas (22%) and mixed GCTs (48%). Polyclonal hCG and monoclonal hCGβ antibodies detected immunoreactivity in some seminomas (7%). No form of hCG was found in spermatocytic seminomas, pure teratomas, or a yolk sac tumor. The serum concentrations correlated with the corresponding tumor expression. The staining intensities of hCG, hCGβ, hCG-h, and hCGα correlated with disease stage but not significantly with relapse, disease-related mortality, or progression-free survival. CONCLUSION Trophoblastic tissue expresses hCG, hCG-h, and free subunits together whereas seminoma tissue occasionally expresses hCGβ. This difference might aid in differential diagnosis of some difficult-to-classify cases.

Laparoscopic radical prostatectomy: Surgical, oncological and functional outcomes

Objective. To report the first results of laparoscopic radical prostatectomy (LRP) at our institution. Material and methods. The surgical, functional and oncological outcomes of all patients who underwent LRP at Helsinki University Central Hospital between May 2002 and May 2006 were prospectively evaluated. The first eight patients were operated transperitoneally and the next 72 extraperitoneally. Results. The mean operative time was 328±73 min (range 210–510 min). The mean estimated blood loss was 769±906 ml (range 50–5500 ml), and 18 patients (22.5%) had transfusions. The mean catheterization time was 13.2±4.0 days (range 9–35 days). Mean hospital stay was 5.7±3.1 days (range 3–15 days). Nineteen patients (23.8%) had perioperative complications. The conversion rate to open surgery was 11.3% and 6.3% required an immediate re-operation. Three anastomotic strictures (5.7%) and two cases of ileus (3.8%) were observed. The cancer was intracapsular (pT1–2) in 68 patients (85.0%) and extracapsular (pT3–4) in 12 (15.0%). Positive surgical margins were noted in 20 patients (25.0%) in total: 17.6% of pT1–2 cases and 66.7% of pT3–4 cases. Prostate-specific antigen recurrence (≥0.2 ng/ml) was noted in 5/53 patients (9.4%), who were followed for >12 months. The continence rate (no daily pad use) was 86.8% and the potency rate was 34.6% at 12 months. Conclusions. The surgical, functional and oncological results of our first LRPs compare fairly well with the early experience of others. LRP is feasible outside high-volume centers but the learning curve is expected to be long.

Comparison of intermittent and continuous androgen deprivation and quality of life between patients with locally advanced and patients with metastatic prostate cancer: a post hoc analysis of the randomized FinnProstate Study VII

Abstract Objective. The aim of the study was to compare intermittent (IAD) and continuous (CAD) androgen deprivation therapy (ADT) between locally advanced (M0) and metastatic (M1) prostate cancer, and the effect of ADT on the quality of life. Material and methods. In total, 852 men with advanced prostate cancer were enrolled to receive goserelin acetate for 24 weeks. Of these, 554 patients whose prostate-specific antigen (PSA) decreased to less than 10 ng/ml or by at least 50% (<20 ng/ml at baseline) were randomized to IAD or CAD. In the IAD arm, ADT was resumed for at least 24 weeks whenever PSA increased to greater than 20 ng/ml or above baseline. Results. Median follow-up time was 65 months. Median times from randomization to progression, death, prostate cancer death and treatment failure in M0 and M1 patients were 46.8 and 21.4, 57.6 and 40.3, 59.5 and 40.7, and 41.9 and 20.0 months, respectively (p < 0.001). No significant differences emerged between IAD and CAD. ADT showed a beneficial effect on pain, activity limitation and social functioning in M1 patients, and a deleterious effect on physical capacity in M0 patients and on sexual functioning in both groups. IAD offered extra benefit for activity limitation, social functioning and recovery of sexual functioning. Conclusions. IAD is as efficient as CAD in treatment of locally advanced and metastatic prostate cancer. ADT improves quality of life in M1 patients, with IAD offering extra benefit.