OlympiAD final overall survival and tolerability results: Olaparib versus chemotherapy treatment of physician's choice in patients with a germline BRCA mutation and HER2-negative metastatic breast cancer.

BACKGROUND: In the OlympiAD study, olaparib was shown to improve progression-free survival compared with chemotherapy treatment of physician's choice (TPC) in patients with a germline BRCA1 and/or BRCA2 mutation (BRCAm) and human epidermal growth factor receptor 2 (HER2)-negative metastatic breast cancer (mBC). We now report the planned final overall survival (OS) results, and describe the most common adverse events (AEs) to better understand olaparib tolerability in this population. PATIENTS AND METHODS: OlympiAD, a Phase III, randomized, controlled, open-label study (NCT02000622), enrolled patients with a germline BRCAm and HER2-negative mBC who had received ≤2 lines of chemotherapy for mBC. Patients were randomized to olaparib tablets (300 mg bid) or predeclared TPC (capecitabine, vinorelbine, or eribulin). OS and safety were secondary end points. RESULTS: A total of 205 patients were randomized to olaparib and 97 to TPC. At 64% data maturity, median OS was 19.3 months with olaparib versus 17.1 months with TPC (HR 0.90, 95% CI 0.66-1.23; P = 0.513); median follow-up was 25.3 and 26.3 months, respectively. HR for OS with olaparib versus TPC in prespecified subgroups were: prior chemotherapy for mBC [no (first-line setting): 0.51, 95% CI 0.29-0.90; yes (second/third-line): 1.13, 0.79-1.64]; receptor status (triple negative: 0.93, 0.62-1.43; hormone receptor positive: 0.86, 0.55-1.36); prior platinum (yes: 0.83, 0.49-1.45; no: 0.91, 0.64-1.33). Adverse events during olaparib treatment were generally low grade and manageable by supportive treatment or dose modification. There was a low rate of treatment discontinuation (4.9%), and the risk of developing anemia did not increase with extended olaparib exposure. CONCLUSIONS: While there was no statistically significant improvement in OS with olaparib compared to TPC, there was the possibility of meaningful OS benefit among patients who had not received chemotherapy for metastatic disease. Olaparib was generally well-tolerated, with no evidence of cumulative toxicity during extended exposure. Please see the article online for additional video content.

Responder analysis of the effects of denosumab on bone mineral density in men receiving androgen deprivation therapy for prostate cancer.

BACKGROUND: Denosumab, a fully human monoclonal antibody against RANK ligand, increased bone mineral density (BMD) and reduced fracture risk vs placebo in a phase 3 trial in men with prostate cancer on androgen deprivation therapy (ADT). The present analysis of this study evaluated BMD changes after 36 months in responder subgroups and in individual patients for three key skeletal sites (lumbar spine (LS), femoral neck (FN) and total hip (TH)) and the distal radius. METHODS: Men with nonmetastatic prostate cancer receiving ADT were treated with subcutaneous denosumab 60 mg (n=734) or placebo (n=734) every 6 months for up to 36 months in a phase 3, randomized, double-blind study. Patients were instructed to take supplemental calcium and vitamin D. For this BMD responder analysis, the primary outcome measure was the percentage change in BMD from baseline to month 36 at the LS, FN and TH as measured by dual-energy X-ray absorptiometry. BMD at the distal 1/3 radius at 36 months was measured in a substudy of 309 patients. RESULTS: At 36 months, significantly more patients in the denosumab arm had increases of >3% BMD from baseline at each site studied compared with placebo (LS, 78 vs 17%; FN, 48 vs 13%; TH, 48 vs 6%; distal 1/3 radius, 40 vs 7% (P<0.0001 for all)). BMD loss at the LS, FN and TH occurred in 1% of denosumab-treated patients vs 42% of placebo patients, and BMD gain at all three sites occurred in 69% of denosumab patients vs 8% of placebo patients. Lower baseline BMD was associated with higher-magnitude BMD responses to denosumab at the LS, FN and TH. CONCLUSIONS: In men with prostate cancer receiving ADT, significantly higher BMD response rates were observed with denosumab vs placebo. Patients with lower baseline T-scores benefited the most from denosumab treatment.

Detection of germ-cell-tumor-specific gene products in peripheral blood by immunomagnetic tumor cell enrichment followed by RT-PCR.

Therapeutic procedures in patients with testicular germ cell tumors (GCT) are determined by the histopathology of the primary tumor and the tumor extension. The aim of our study was to determine whether conventional staging could be supplemented by combining enrichment of disseminated testicular GCT cells from peripheral blood with subsequent detection of germ-cell-specific gene products. Blood samples from 46 patients with GCT of different clinical stages (CS) were examined by RT-PCR before therapy and >/=8 weeks thereafter for alpha-fetoprotein, beta-human chorionic gonadotropin and germ-cell-specific alkaline phosphatase mRNA. In addition, we performed titration experiments to evaluate whether the sensitivity can be improved by previous immunomagnetic tumor cell enrichment with anti-epithelial HEA-125 microbeads. No positive results were found in controls (n=15; specificity 100%). The overall ratio of positive PCR results in the group of patients with GCTs was 28.26%. The ratio was 35.7% for CS >IIb (n=5/14 patients), 20.0% for CS IIa-b (n=4/20) and 33.3% for CS I (n=4/12). FACS analysis in titration experiments with GCT cell lines showed that previous immunomagnetic tumor cell enrichment achieved a significant increase ranging up to 185.6 times the initial ratio and thus improved the measuring conditions for detection of tumor-specific transcripts. The sole qualitative RT-PCR of tumor-specific gene products in peripheral blood is not sensitive enough to improve staging in GCT patients. Immunomagnetic enrichment of GCT cells in peripheral blood seems a promising approach for increasing the sensitivity of RT-PCR.

Hyperphosphaturia after kidney transplantation in syngeneic rats: effects on nephrocalcinosis and bone metabolism?

BACKGROUND: Studies on kidney transplantation have thus far mainly dealt with surgical techniques, immunology, and transplant tolerance. Disturbed mineral metabolism after renal denervation has not received much attention. Basic physiological research in short-term experiments has shown that experimental renal denervation in rats leads to parathormone (PTH)-independent hyperphosphaturia (HPU). HPU and other metabolic complications also have been described after clinical kidney transplantation. Furthermore, there is an unexpected increase in the risk of bone fracture. However, these studies have examined an organism pre-damaged with regard to the parathyroid and immunosuppression. Experimental investigations in syngeneic rats were performed to see whether HPU also occurs after transplantation and thus after denervation and which target organs are involved. METHODS: Thirty-six male Lewis rats subjected to laparotomy (n = 12), unilateral nephrectomy (n = 12), or unilateral transplantation and bilateral nephrectomy (n = 12) were observed for 18 weeks. RESULTS: Animals that underwent transplantation had a significant loss of phosphate in the urine not associated with decreased calcium, phosphate, or magnesium in bone. Stability test showed no deterioration, despite a slight increase in the bone parameters of alkaline phosphatase, cyclic AMP, and hydroxyproline with unchanged calciotropic hormones. Nephrocalcinosis was not observed. Parallel to HPU, there was a compensatory reduction in fecal phosphate excretion. CONCLUSIONS: The loss of phosphate after clinical kidney transplantation in the predamaged parathyroid hormone control system as well as immunosuppression and a surprising increase in the incidence of bone fractures may be explained by the denervation-related loss of phosphate. The lack of intestinal counter-regulation could be an important pathomechanism.

[Molecular diagnostics in urologic oncology. Detection of nucleic acids in urine samples]. / Molekulare Diagnostik in der urologischen Onkologie. Detektion von Nukleinsäuren in Urinproben.

The goal of molecular diagnostics in oncology is the early diagnosis of malignant disease processes during initial work-up or as part of follow-up. Body fluids serve as the primary material for non-invasive diagnostic methods. Besides actual tumor cells, the examination of urine can yield evidence of secreted proteins or even free nucleic acids. In principle, all of the methods available for the detection of tumor markers in tissue or blood samples can be successfully applied to the examination of urine samples. However, molecular biological examination of urine samples is associated with important problems because the cells in such samples are exposed to significant degradation and regression effects and because certain components of the urine act to inhibit the polymerase chain reaction. The present overview discusses the respective strengths and weakness of the available technology as applied to the diagnosis of urologic malignancies. Experimental studies conducted to date have reported high sensitivities and specificities for molecular diagnostics using urine samples. It is important to note that not only carcinomas of the urinary bladder can be diagnosed from material obtained in urine samples: in fact, the method can be used to diagnose entities such as renal cell and prostate carcinomas and, due to renal filtration of DNA, even non-urologic malignancies. The diagnostic application of these methods, however, remains in an experimental stage and must still clear several hurdles before becoming available for routine clinical use.

Intestinal and renal handling of oxalate in magnesium-deficient rats. Evaluation of intestinal in vivo 14C-oxalate perfusion.

OBJECTIVE: To clarify in vivo, using isolated small intestinal loops perfused with radioactive 14C-oxalate, whether intestinal hyperabsorption or reduced secretion is important in magnesium deficiency (MgD), as this is a potential cause of calcium oxalate urolithiasis. MATERIALS AND METHODS: Twenty-four Sprague-Dawley rats were either fed a standard diet (control, 12 rats) or a magnesium-deficient diet (MgD, 12 rats) for 19 weeks. One hour before the animals were killed, a defined length of a small intestinal loop was isolated and filled with 5 mL of 0.9% NaCl and a defined amount of intravenous 14C-oxalate applied. Using this method it was possible to determine the secretion of unlabelled oxalate into the intestinal lumen, from the specific activity in plasma. RESULTS: Plasma oxalate levels doubled under MgD; urinary calcium and phosphorus also increased significantly, while urine oxalate tended to decrease. The secretion of oxalate into the intestinal lumen of MgD animals increased significantly, by five times that of the control. The relative supersaturation for calcium oxalate remained constant. Elementary analysis of renal tissue showed an increase in calcium and phosphorus under MgD, in the sense of nephrocalcinosis, but no concretions were detected (no nephrolithiasis). CONCLUSION: In contrast to earlier studies, there is no evidence that hyperoxaluria is responsible for the possible development of urolithiasis in MgD. This was confirmed by calcium phosphate deposits in renal tissue, even though there was no evidence of oxalate urolithiasis. The increase in plasma oxalate seems to be completely compensated by strongly increased oxalate secretion into the intestinal lumen.

DNA-based detection of prostate cancer in blood, urine, and ejaculates.

BACKGROUND: Methylation-specific polymerase chain reaction (MSP) targeting promoter hypermethylation of the glutathione S transferase P1 gene (GSTP1), as the most frequent DNA alteration in prostatic carcinoma, was used for the molecular detection of cell-bound and cell-free prostate tumor DNA in various human bodily fluids. MATERIALS AND METHODS: We investigated GSTP1 promoter hypermethylation in DNA isolated from plasma, serum, nucleated blood cells, ejaculates, urines after prostate massage, and prostate tissue from 33 patients with prostate cancer and 26 control patients with benign prostatic hyperplasia (BPH). Using a viral DNA extraction kit specifically recommended for DNA isolation from urine samples, GSTP1 promoter hypermethylation in urine sediments after prostatic massage was investigated in a cohort of 29 patients with prostate cancer and 40 controls with BPH. Fluorescently labeled MSP products were analyzed on an automated gene sequencer. RESULTS: GSTP1 promoter hypermethylation was found in 90% of tumors (18 of 20), 72% of plasma or serum samples (23 of 32), 50% of ejaculates (4 of 8), and between 36% (4 of 11; normal DNA isolation kit) and 76% (22 of 29; viral kit) of exprimated urines from patients with prostate cancer. Also, MSP identified circulating tumor cells in 30% (10 out of 33) of prostate cancer patients. Except for one urine sample, GSTP1 promoter hypermethylation was not found in tissue and body fluids from patients with BPH. CONCLUSION: GSTP1 promoter methylation analysis provides a highly specific tool for DNA-based diagnosis of prostate cancer in body fluids.

Detection of prostate-specific antigen RNA before and after radical retropubic prostatectomy and transurethral resection of the prostate using "Light-Cycler"-based quantitative real-time polymerase chain reaction.

OBJECTIVES: To report our initial experience gained in establishing real-time reverse transcriptase-polymerase chain reaction (RT-PCR) detection of prostate-specific antigen (PSA) mRNA using the quantitative online PCR system LightCycler. Many studies have thus far failed to provide the desired proof that the detection of circulating PSA-expressing tumor cells with RT-PCR in the blood samples of patients with prostate cancer (PCa) is a highly sensitive prognostic and course marker. One of the possible reasons is the lack of reliable quantification methods. METHODS: Blood samples before and after surgery were obtained from 87 patients who underwent radical prostatectomy for locally confined PCa and 27 patients who underwent transurethral resection of the prostate for benign prostatic hyperplasia. Eight days postoperatively, additional blood samples were obtained from the patients with PCa. Quantitative no-nested RT-PCR for PSA mRNA (291 bp) was performed using the LightCycler system applying the SYBR Green protocol. The number of circulating LNCaP tumor cell-equivalents per sample was estimated from the mean amplification value measured in a given number of LNCaP cells. RESULTS: PSA mRNA was detected preoperatively in 19 patients with Stage pT2 tumor (40%) and in 28 patients with tumor greater than Stage pT2 (72%), but in only 2 patients with benign prostatic hyperplasia (8%; analysis of variance, P <0.001). Significant quantitative differences were observed among Stage pT2 disease (1034 LNCaP tumor cell-equivalents/mL), greater than Stage pT2 disease (7830 cells/mL), and benign prostatic hyperplasia (58 cells/mL; analysis of variance for all groups, P <0.001). The correlation between the detection of PSA expression by RT-PCR and the Gleason score and serum PSA value was statistically significant. CONCLUSIONS: Our results show that the initial experience with the LightCycler system for PSA-assisted detection of circulating PSA mRNA in PCa by RT-PCR may be a promising preoperative prognostic marker for organ-confined or locally advanced PCa. Long-term follow-up of these patients with PCa must demonstrate the clinical value of molecular diagnostics with quantitative RT-PCR systems.

Molecular staging of surgical margins after radical prostatectomy by detection of telomerase activity.

BACKGROUND: The further course of prostate cancer (PC) after radical prostatectomy (RPX) is decisively influenced by the local tumor stage. Although it is thus far possible to assess the risk of local recurrence from the pathohistology, precise predictions cannot be made. A more precise evaluation would be desirable, mainly for early planning of adjuvant therapy. Other authors have shown that telomerase activity may be a marker for malignant potential. We assessed the detection of telomerase activity using the telomeric repeat amplification protocol (TRAP) in surgical margins compared to conventional histopathological examination. METHODS: Ninety-two patients with local PC who underwent RPX were examined. After RPX biopsies were obtained from four defined areas of the prostatic fossa and processed by TRAP assay for telomerase activity using a standard protocol. RESULTS: In 5 of 48 patients (10.4%) with organ-confined prostate carcinoma (pT2) telomerase activity could be detected. Seven of 47 patients (14.9%) with locally advanced PC (> pT2) had at least one positive specimen. CONCLUSIONS: The results obtained in our study indicate that detection of telomerase activity by TRAP assay may be a suitable parameter for molecular staging of surgical margins, because of the high tumor-specificity. Further follow-up must clarify whether patients with positive molecular detection have an increased risk of local recurrence.

DNA-based detection of prostate cancer in urine after prostatic massage.

OBJECTIVES: Promoter hypermethylation of the glutathione-S-transferase P1 (GSTP1) gene is a specific feature of prostate cancer. This epigenetic DNA alteration served as the target for molecular detection of prostate cancer cells in urine sediments after prostatic massage. METHODS: Bisulfite treatment followed by methylation-specific polymerase chain reaction was used to detect GSTP1 promoter hypermethylation in DNA isolated from urine sediments obtained after prostatic massage of men with and without prostate cancer. RESULTS: GSTP1 promoter hypermethylation was demonstrated in the sediments of 1 (2%) of 45 patients diagnosed with benign prostatic hyperplasia, 2 (29%) of 7 patients with prostatic intraepithelial neoplasia, 15 (68%) of 22 patients with early, intracapsular cancer, and 14 (78%) of 18 patients with locally advanced or systemic prostate cancer, resulting in a specificity of 98% and an overall sensitivity of 73% for the detection of prostate cancer. CONCLUSIONS: Urinalysis for GSTP1 promoter hypermethylation constitutes a sensitive and highly specific DNA-based marker for molecular detection of prostate cancer, including early stages.

Increased incidence of luteinizing hormone-releasing hormone receptor gene messenger RNA expression in hormone-refractory human prostate cancers.

There are few options for treating hormone-refractory prostate cancer (PC). Various studies indicate that luteinizing hormone-releasing hormone (LHRH) agonists may have a direct inhibitory effect on prostate tumors mediated by specific LHRH receptors. One study evaluated LHRH receptors in hormone-dependent PC tissue, but no data have thus far been obtained on the presence of LHRH receptors in benign prostatic hyperplasia (BPH) and especially hormone-refractory PC in patients. Thus, it is not yet clear whether LHRH receptors indicate tumor-related differentiation or even hormone-refractory dedifferentiation or are likewise associated with BPH. The aim of this study was to determine the rate of LHRH receptor mRNA expression in BPH and in primary, potentially androgen-dependent and in hormone-refractory PC with clinical progression. Multiplex reverse transcription-PCR was used to simultaneously detect the expression of mRNA for LHRH receptors and beta-actin in 48 patients with BPH, 14 with a primary, possibly hormone-dependent, prostate carcinoma (PPC), and 18 with a hormone-refractory prostate carcinoma (HRPC). Sixteen of 18 samples with HRPC showed intact RNA and expressed mRNA for LHRH receptors (100%). However, the RNA-intact PPC and BPH showed significantly lower expression of mRNA for LHRH receptors (46.2 and 55.3%, respectively; variance analysis: P = 0.0017). The significantly higher expression of mRNA for LHRH receptors in HRPC indicates that therapeutic concepts should be developed that target this site of action. In addition to possible direct effects of LHRH agonists or antagonists demonstrated previously in vitro, it seems useful to apply targeted cytotoxic LHRH analogues or monoclonal antibodies.

Molecular markers in testicular germ cell tumors--objects of clinical research or close to becoming clinical tools?

The aim of this short review is to critically evaluate hitherto investigated molecular markers for testicular germ cell tumors. Molecular parameters have been clinically established as diagnostic and prognostic markers for a number of tumors; this has not yet been achieved for germ cell tumors. There are interesting prospects, however. Studies on the ribonucleoprotein telomerase, for example, have demonstrated a correlation between enzyme activity and chemotherapeutic drug sensitivity. Moreover, innovative treatment approaches target this reverse transcriptase via telomerase antisense RNA. Another potential diagnostic marker is the detection of circulating tumor cells, which correlated with an increased relapse rate in initial studies. There are also interesting possibilities for the germ-cell-tumor-specific isochromosome [i(12)p], which is helpful in the differential diagnosis of mediastinal masses. Here initial studies demonstrated a correlation between the copy number and resistance against chemotherapeutic drugs. Without prospective studies to validate data obtained thus far, neither these nor other parameters can be assessed as diagnostic and prognostic factors. Irrespective of their immediate clinical applicability, however, investigations on molecular alterations in testicular germ cell tumors will become the basis for a molecular-diagnostically oriented subclassification of tumors as well as for novel therapeutic approaches.

Reverse transcriptase-polymerase chain reaction for prostate-specific antigen in the molecular staging of pelvic surgical margins after radical prostatectomy.

OBJECTIVES: To examine the application of reverse transcriptase-polymerase chain reaction (RT-PCR) to assist in prostate-specific antigen (PSA) detection in the surgical margins after radical prostatectomy (RP). The risk of local recurrence increases considerably in the presence of extracapsular tumor growth and/or positive surgical margins at RP. Although this makes it possible to identify patients with an increased risk of local recurrence, precise predictions cannot be made. A more precise assessment is desirable mainly for early planning of adjuvant therapy. METHODS: Ninety-five patients with clinically organ-confined prostate cancer (CaP) underwent RP. After removing the gland, biopsies were obtained from four defined areas of the prostatic fossa and processed for RT-PCR for PSA detection. Sixteen patients with muscle-invasive bladder carcinoma who underwent radical cystoprostatectomy served as controls. RESULTS: Thirty-two of 95 patients with CaP (35%) had at least one positive molecular margin indicating an expression for PSA; 19 of 48 (39%) of these had an organ-confined tumor stage according to conventional histology and 13 of 47 (28%) had tumor growth beyond the prostate. A statistically significant correlation between the frequency of positive molecular margins and clinical data was only observed in the group with disease greater than Stage pT2. All controls had negative molecular margins (P = 0.012). CONCLUSIONS: On the basis of the results obtained, molecular diagnostic RT-PCR for PSA detection in the surgical margins after RP seems to be an interesting supplementary tool for monitoring the course and establishing the prognosis. Long-term follow-up of these patients is needed to demonstrate the clinical value of molecular diagnostics of surgical margins during RP.

Fluorescent methylation-specific polymerase chain reaction for DNA-based detection of prostate cancer in bodily fluids.

Promoter hypermethylation of the glutathione S-transferase P1 gene (GSTP1) is the most frequent DNA alteration in prostatic carcinoma. Because this epigenetic DNA alteration can be reliably detected by methylation-specific PCR (MSP), we applied this new technique for molecular detection of prostate cancer in various human bodily fluids. We investigated GSTP1 promoter hypermethylation in DNA isolated from plasma, serum, ejaculate, and urine after prostate massage and from prostate carcinoma tissues from 33 patients with prostate cancer and 26 control patients with benign prostatic hyperplasia (BPH). Fluorescently labeled MSP products were analyzed on an automated gene sequencer. Whereas GSTP1 promoter hypermethylation was not detectable by MSP in prostate tissue and bodily fluids from patients with BPH, we found it in 94% of tumors (16 of 17), 72% of plasma or serum samples (23 of 32), 50% of ejaculate (4 of 8) and 36% of urine (4 of 11) from patients with prostate cancer. Additionally, MSP identified circulating tumor cells in 30% (10 of 33) of prostate cancer patients. Analysis of GSTP1 promoter hypermethylation by MSP thus provides a specific tool for molecular diagnosis of prostate cancer in bodily fluids.

Efficacy and tolerability of tolterodine in children with detrusor hyperreflexia.

OBJECTIVES: To investigate the urodynamic effects and tolerability of the new antimuscarinic drug tolterodine in children with detrusor hyperreflexia. METHODS: Twenty-two children (12 boys and 10 girls; age range 3 months to 15 years, mean age 5.7 years) with detrusor hyperreflexia resulting in maximum detrusor pressures exceeding 40 cm H(2)O during filling cystotonometry were enrolled to receive tolterodine tartrate (a total of 0.1 mg/kg orally daily, divided into two doses) either as a first-line therapy (n = 12, group 1) or replacing oxybutynin chloride therapy (n = 10, group 2). Within 3 months, all patients underwent urodynamic re-evaluation during ongoing tolterodine treatment. RESULTS: In group 1, the mean maximum bladder capacity increased from 120.2 to 173.0 mL (+44%), the mean detrusor compliance increased from 8.7 to 13.5 mL/cm H(2)O (+55%), and the mean maximum detrusor pressures decreased from 70.1 to 37.9 cm H(2)O (-46%); the differences were significant (P < 0.001). In group 2, no differences in the urodynamic effects of oxybutynin versus tolterodine were noted. Only 1 patient experienced a transient and moderately adverse effect with tolterodine. CONCLUSIONS: Although based on a limited number of subjects, these data indicate that in pediatric patients with detrusor hyperreflexia, tolterodine may be better tolerated than and equally effective as the standard drug oxybutynin chloride.