Patient-matched analysis identifies deregulated networks in prostate cancer to guide personalized therapeutic intervention.

Prostate cancer (PrCa) is the second most common malignancy in men. More than 50% of advanced prostate cancers display the TMPRSS2-ERG fusion. Despite extensive cancer genome/transcriptome data, little is known about the impact of mutations and altered transcription on regulatory networks in the PrCa of individual patients. Using patient-matched normal and tumor samples, we established somatic variations and differential transcriptome profiles of primary ERG-positive prostate cancers. Integration of protein-protein interaction and gene-regulatory network databases defined highly diverse patient-specific network alterations. Different components of a given regulatory pathway were altered by novel and known mutations and/or aberrant gene expression, including deregulated ERG targets, and were validated by using a novel in silico methodology. Consequently, different sets of pathways were altered in each individual PrCa. In a given PrCa, several deregulated pathways share common factors, predicting synergistic effects on cancer progression. Our integrated analysis provides a paradigm to identify druggable key deregulated factors within regulatory networks to guide personalized therapies.

[The level of evidence for the use of biomarkers in the early detection of prostate cancer]. / Les niveaux de preuve des biomarqueurs utilisés pour la détection précoce des cancers de la prostate.

To systematically review the evidence for the use of PSA and other biomarkers in the early detection of prostate cancer, we searched PubMed for clinical trials and studies assessing PSA and other biomarkers in the early detection of prostate cancer, published between 2000 and May 2013 that included >200 subjects. The level of evidence (LOE) for clinical utility was evaluated using the tumor marker utility grading system. A total of 84 publications, corresponding to 70 trials and studies were selected for inclusion in this review. We attributed a level of evidence (LoE) of IA to PSA for early PCa detection, but we do not recommend its use in mass screening. Emerging biomarkers were assessed in prospective case-control and cohort studies: PCA3 (n=3); kallikreins (n=3); [-2]proPSA (n=5); fusion oncogenes (n=2). These studies used biopsy results for prostate cancer to determine specificity and sensitivity, but they did not assess the effect on PCa mortality. The LoE attributed was III-C. PSA can be used for early prostate cancer detection but mass screening is not recommended. Studies on other biomarkers suggest that they could be used, individually or in combination, to improve the selection of patients with elevated PSA levels for biopsy, but RCTs assessing their impact on prostate cancer management and mortality are needed. A better use of available tests is possible for men at risk in order to maximize the risk-benefit ratio.

Virilizing oncocytic adrenocortical carcinoma: clinical and immunohistochemical studies.

CONTEXT: Oncocytic tumors of the adrenal cortex are rare, mostly nonfunctioning and benign. SETTING: Report virilizing oncocytic adrenocortical carcinoma in a 50-year-old woman. PATIENT: She presented a recent and progressive virilization syndrome, associated with high blood pressure. Hormonal evaluation showed elevated serum testosterone and delta-4-androstenedione levels, normal urinary free cortisol level and incomplete suppression of cortisol at the 1 mg dexamethasone suppression test. CT scan of the abdomen revealed a 35 mm left adrenal mass. INTERVENTION: The patient underwent a left adrenalectomy, and the histological study showed a 3 cm oncocytic adrenocortical carcinoma with signs of malignancy. RESULTS: Immunohistochemical study revealed that tumor cells expressed the steroidogenic enzymes involved into androgen synthesis (3ßHSD and P450c17α), P450 aromatase and luteinizing hormone (LH) receptors. Post-operatively, signs of virilization improved rapidly, serum testosterone and delta-4-androstenedione levels returned to normal, as did the dexamethasone suppression test. During follow-up CT-scan and 18-FDG PET/CT showed a right ovary mass, corresponding to a follicular cyst associated with hyperthecosis. The patient is alive with no recurrence 48 months after adrenal surgery. CONCLUSION: Oncocytic adrenocortical carcinomas, although extremely rare, should be considered in women with a virilization syndrome. In this woman immunohistochimical studies revealed the presence of steroidogenic enzymes involved into androgen synthesis and aromatization, and LH receptors could be implicated in this pathology.

Periprostatic adipocytes act as a driving force for prostate cancer progression in obesity.

Obesity favours the occurrence of locally disseminated prostate cancer in the periprostatic adipose tissue (PPAT) surrounding the prostate gland. Here we show that adipocytes from PPAT support the directed migration of prostate cancer cells and that this event is strongly promoted by obesity. This process is dependent on the secretion of the chemokine CCL7 by adipocytes, which diffuses from PPAT to the peripheral zone of the prostate, stimulating the migration of CCR3 expressing tumour cells. In obesity, higher secretion of CCL7 by adipocytes facilitates extraprostatic extension. The observed increase in migration associated with obesity is totally abrogated when the CCR3/CCL7 axis is inhibited. In human prostate cancer tumours, expression of the CCR3 receptor is associated with the occurrence of aggressive disease with extended local dissemination and a higher risk of biochemical recurrence, highlighting the potential benefit of CCR3 antagonists in the treatment of prostate cancer.

Sphingosine Kinase 1 urothelial expression is increased in patients with neurogenic detrusor overactivity

Objectives: To evaluate the expression of sphingosine kinase 1 (SPK1) in the bladder wall in patients with neurogenic lower urinary tract dysfunction and its association with clinical, urodynamic and pathological features. Materials and Methods: The expression of SPK1 was studied in bladder wall specimens obtained from cystectomy using immunohistochemistry in ten patients with spinal cord injury (n=8) or multiple sclerosis (n=2) with urodynamically proven neuropathic bladder dysfunction, and in controls (n=5). Inflammation and fibrosis were analysed with histological criteria and SPK1 expression was determined by individual immunohistochemical staining. Results: Significant increased SPK1 urothelial immunoreactivity was shown in patients compared to control group (p=0.03). By contrast, SPK1 immunoreactivity in patients was significantly decreased in the sub-urothelium, muscles and nerves, p=0.02; 0.01 and 0.003, respectively. Patients with neurogenic detrusor overactivity (NDO) had higher SPK1 urothelium expression than those without any DO (p=0.04). Conclusions: SPK1 is expressed in the human bladder wall, specifically the urothelium, in bladder specimens from patients with NDO. The role of SPK1 in the pathophysiology of NDO needs further elucidation.

Mannose-6-phosphate receptor: a target for theranostics of prostate cancer.

The development of personalized and non-invasive cancer therapies based on new targets combined with nanodevices is a major challenge in nanomedicine. In this work, the over-expression of a membrane lectin, the cation-independent mannose 6-phosphate receptor (M6PR), was specifically demonstrated in prostate cancer cell lines and tissues. To efficiently target this lectin a mannose-6-phosphate analogue was synthesized in six steps and grafted onto the surface of functionalized mesoporous silica nanoparticles (MSNs). These MSNs were used for in vitro and ex vivo photodynamic therapy to treat prostate cancer cell lines and primary cell cultures prepared from patient biopsies. The results demonstrated the efficiency of M6PR targeting for prostate cancer theranostic.

Sphingosine Kinase 1 urothelial expression is increased in patients with neurogenic detrusor overactivity.

UNLABELLED: To evaluate the expression of sphingosine kinase 1 (SPK1) in the bladder wall in patients with neurogenic lower urinary tract dysfunction and its association with clinical, urodynamic and pathological features. MATERIALS AND METHODS: The expression of SPK1 was studied in bladder wall specimens obtained from cystectomy using immunohistochemistry in ten patients with spinal cord injury (n=8) or multiple sclerosis (n=2) with urodynamically proven neuropathic bladder dysfunction, and in controls (n=5). Inflammation and fibrosis were analysed with histological criteria and SPK1 expression was determined by individual immunohistochemical staining. RESULTS: Significant increased SPK1 urothelial immunoreactivity was shown in patients compared to control group (p=0.03). By contrast, SPK1 immunoreactivity in patients was significantly decreased in the sub-urothelium, muscles and nerves, p=0.02; 0.01 and 0.003, respectively. Patients with neurogenic detrusor overactivity (NDO) had higher SPK1 urothelium expression than those without any DO (p=0.04). CONCLUSIONS: SPK1 is expressed in the human bladder wall, specifically the urothelium, in bladder specimens from patients with NDO. The role of SPK1 in the pathophysiology of NDO needs further elucidation.

Normotensive incidentally discovered pheochromocytomas display specific biochemical, cellular, and molecular characteristics.

CONTEXT: A number of incidentally discovered pheochromocytomas are not associated with hypertension. The characteristics of normotensive incidentally discovered pheochromocytomas (NIPs) are poorly known. OBJECTIVE: The purpose of this work was to assess the clinical, hormonal, histological, and molecular features of NIPs. DESIGN: This was a retrospective cohort recruited from 2001 to 2011 in 2 tertiary care medical departments. PATIENTS AND METHODS: Clinical, biological, and radiological investigations performed in 96 consecutive patients with sporadic unilateral pheochromocytomas were examined; 47 patients had overt pheochromocytomas responsible for hypertension. Among the patients with incidental pheochromocytomas, 28 had hypertension and 21 were normotensive (NIPs). A total of 62 tumors were examined to determine the Pheochromocytoma of the Adrenal Gland Scale Score, and 29 were studied for the expression of 16 genes involved in chromaffin cell function. RESULTS: Tumor size and metaiodobenzylguanidine (MIBG) scintigraphy results were similar for hypertensive pheochromocytomas (HPs) and NIPs. Patients with NIPs displayed reduced summed levels of urinary catecholamines and metanephrines and, more specifically, reduced levels of adrenaline and metadrenaline compared with those of patients with HPs (P < .001). Urinary metanephrines had 98% diagnostic sensitivity in patients with HPs and only 75% in patients with NIPs (P < .01). Tumor diameter positively correlated with the total amount of urinary concentrations of metanephrines in patients with HPs (P < .001) but not in patients with NIPs. NIPs displayed global decreased chromaffin gene expression (reaching significance for 5 of them) and 2 corresponding proteins (phenylethanolamine N-methyltransferase and secretogranin II) and a significant increase in the cellularity, mitotic activity, and presence of atypical mitosis (P < .05). CONCLUSIONS: NIPs differ from pheochromocytomas responsible for hypertension and display features of altered chromaffin differentiation. These tumors may be misdiagnosed with the use of the usual biological diagnostic tools.

Vaginal lubrication after cervicovaginal stimulation is facilitated by phosphodiesterase type 5 inhibition in ovariectomized mice.

INTRODUCTION: Nitric oxide synthases (NOSs) and estrogen receptors are expressed in the vagina. AIM: We aimed to assess the impact of sildenafil on vaginal lubrication according to the hormonal status and to determine the role of the neuronal isoform of NOS (nNOS). METHODS: Four-week-old C57/BL6 female mice were sham operated or ovariectomized. At 10 weeks of age, they were injected intraperitoneally by any combination of sildenafil, 7-nitroindazole (7-NI)--a potent selective nNOS inhibitor--or the corresponding vehicles. Vaginal lubrication was induced in a physiological manner by cervical vaginal probing and quantified depending on the hormonal and pharmacological conditions. The animals were then sacrificed for vaginal histomorphometry. MAIN OUTCOME MEASURES: The main outcome measure is the quantification of vaginal transudate after cervicovaginal stimulation and vaginal histomorphometry. RESULTS: Sildenafil increased cervicovaginal probing-induced vaginal lubrication in ovariectomized and sham-operated animals. Ovariectomized mice exhibited decreased vaginal lubrication as compared with sham-operated mice. When taking into account the presence of severe vaginal atrophy, a threefold increase in transudate per gram of vagina wet weight was revealed in ovariectomized animals. Castration markedly reduced the thickness of the vaginal wall. nNOS inhibition by 7-NI had no impact on vaginal lubrication. CONCLUSIONS: Irrespective of the hormonal status, sildenafil increased vaginal lubrication. The vaginal effect of sildenafil was independent of the nNOS pathway and more pronounced in ovariectomized animals.

Prognostic interest in discriminating muscularis mucosa invasion (T1a vs T1b) in nonmuscle invasive bladder carcinoma: French national multicenter study with central pathology review.

PURPOSE: Predictive factors of T1 nonmuscle invasive bladder cancer evolution that could guide treatment decision making are lacking. We assessed the prognostic value of muscularis mucosa invasion in nonmuscle invasive bladder cancer. MATERIALS AND METHODS: In a national multicenter study patients with primary T1 nonmuscle invasive bladder cancer were recruited from 6 French hospitals. All patients had undergone transurethral resection of bladder tumor. All T1 tumors were substaged according to muscularis mucosa invasion as T1a-no invasion beyond the muscularis mucosa or T1b-invasion beyond the muscularis mucosa with muscle preservation. Subsequent central pathology review was then done by a single referent uropathologist. Muscularis mucosa invasion was tested as a prognostic factor for survival on univariate and multivariate analysis. RESULTS: A total of 587 patients were enrolled in the study, including 388 (66%) with T1a and 199 (34%) with T1b tumors. Median followup after transurethral resection of bladder tumor was 35 months (IQR 14-54). There was no significant difference between groups T1a and T1b except high tumor grade in T1b cases (p <0.0001). After central review, initial pathological substaging was confirmed in 84% of cases. On multivariate analysis muscularis mucosa invasion (T1b substage) was significantly associated with recurrence-free (p = 0.03), progression-free (p = 0.0002) and cancer specific (p = 0.02) survival. The main study limitation was absent systematic subsequent transurethral resection of bladder tumor. CONCLUSIONS: Muscularis mucosa invasion appears to be highly predictive of T1 nonmuscle invasive bladder cancer behavior. Consequently, systematic T1a vs T1b discrimination should be highly advocated by urologists and pathologists. We believe that it could aid in crucial decision making when choosing between conservative management and radical cystectomy remains a moot point.

First evidence of sphingosine 1-phosphate lyase protein expression and activity downregulation in human neoplasm: implication for resistance to therapeutics in prostate cancer.

This is the first report of sphingosine 1-phosphate lyase (SPL) protein expression and enzymatic activity in human neoplasm. This enzyme drives irreversible degradation of sphingosine 1-phosphate (S1P), a bioactive lipid associated with resistance to therapeutics in various cancers, including prostate adenocarcinoma. In fresh human prostatectomy specimens, a remarkable decrease in SPL enzymatic activity was found in tumor samples, as compared with normal adjacent tissues. A significant relationship between loss of SPL expression and higher Gleason score was confirmed in tissue microarray (TMA) analysis. Moreover, SPL protein expression and activity were inversely correlated with those of sphingosine kinase-1 (SphK1), the enzyme producing S1P. SPL and SphK1 expressions were independently predictive of aggressive cancer on TMA, supporting the relevance of S1P in prostate cancer. In human C4-2B and PC-3 cell lines, silencing SPL enhanced survival after irradiation or chemotherapy by decreasing expression of proteins involved in sensing and repairing DNA damage or apoptosis, respectively. In contrast, enforced expression of SPL sensitized cancer cells to irradiation or docetaxel by tilting the ceramide/S1P balance toward cell death. Interestingly, the S1P degradation products failed to sensitize to chemo- and radiotherapy, supporting the crucial role of ceramide/S1P balance in cancer. Of note, the combination of SPL enforced expression with a SphK1 silencing strategy by further decreasing S1P content made prostate cancer cells even more sensitive to anticancer therapies, suggesting that a dual strategy aimed at stimulating SPL, and inhibiting SphK1 could represent a future approach to sensitize cancer cells to cancer treatments.

Adrenocortical tumors: improving the practice of the Weiss system through virtual microscopy: a National Program of the French Network INCa-COMETE.

The Weiss score is the reference method to distinguish between a benign and a malignant adrenocortical tumor (ACT). A program was initiated to improve the reproducibility of the pathologic diagnosis of ACTs in France through the National INCa-COMETE Network. Twelve pathologists from all Reference Centers of the Network analyzed 50 selected ACTs using a web-based virtual microscopy approach in a blind design, allowing to determine the intraobserver and interobserver reproducibilities of the Weiss system. All ACTs were read twice in random order before and after a coaching meeting organized to harmonize and improve analyses and create an online tutorial. The validity of the virtual approach was first established by comparing the 2 consensuses (virtual and microscopic) obtained for each tumor by 3 pathologists who performed the 2 approaches in a blinded manner. For the "dichotomized Weiss score" (separating malignant ≥3 from benign ≤2 tumors) interobserver reproducibility was "substantial" at the first "virtual" reading (κ = 0.70) and increased at the second "virtual" reading (κ = 0.75). In parallel, 7 of the 9 items of the Weiss system showed improvement. The diagnostic accuracy of the observers as a group, using the modal group score approach, showed an improved sensitivity from 86% to 95% for the diagnosis of malignant ACTs. We show the validity of the virtual microscopy approach and that the program improved the practice of the Weiss system reading and therefore the diagnosis of ACT. This tool can now be extended for other research and/or routine purposes in this rare cancer.

Sphingosine kinase-1 activity and expression in human prostate cancer resection specimens.

PURPOSE: Sphingosine kinase-1 (SphK1) was shown in preclinical models and non-genitourinary cancers to be instrumental in cancer progression, adaptation to hypoxia and in tumour angiogenesis. No data were available in human prostate cancer. The present study was designed to assess SphK1 expression and activity in radical prostatectomy specimens and to research correlations with clinical features. MATERIALS AND METHODS: Transverse section of fresh tissue was obtained from 30 consecutive patients undergoing laparoscopic prostatectomy. SphK1 enzymatic activities of tumour and normal counterpart were determined. Relationships with PSA, Gleason sum, pathological stage, resection margin status and treatment failure were researched. SphK1 pattern of expression was then assessed on tissue microarray. RESULTS: A significant 2-fold increase in SphK1 enzymatic activity(11.1 ± 8.4 versus 5.9 ± 3.2 (P<0.04)) was observed in cancer. The upper quartile of SphK1 activity was associated with higher PSA (16.7 versus 6.4 ng/ml, P = 0.04), higher tumor volumes (20.7 versus 9.8, P = 0.002), higher rates of positive margins (85.7% versus 28.6%, P = 0.01) and surgical failure (71.4% versus 9.5%, P = 0.003) than the lower three quartiles. Odds ratios (OR) for treatment failure showed a strong relationship with SphK1 activity (OR: 23.7, P = 0.001), positive resection margins (OR: 15.0, P = 0.007) and Gleason sum (≥4+3, OR: 8.0, P = 0.003). Tissue microarrays showed discrete epithelial expression that varied with Gleason sum with significant relationship between SphK1 expression and higher Gleason sum. CONCLUSION: In complement to preclinical literature, the demonstrated relationships between SphK1-increased activity in cancer and relevant clinical features confirm a central role for SphK1 in prostate cancer that herald promising avenues in risk-assessment and treatment.

Activation of sphingosine kinase-1 in cancer: implications for therapeutic targeting.

Sphingolipid metabolites are critical to the regulation of a number of fundamental biological processes including cancer. Whereas ceramide and sphingosine mediate and trigger apoptosis or cell growth arrest, sphingosine 1-phosphate promotes proliferation, cell survival and angiogenesis. The delicate equilibrium between the intracellular levels of each of these sphingolipids is controlled by the enzymes that either produce or degrade these metabolites. Sphingosine kinase-1 is a crucial regulator of this two-pan balance, because its produces the pro-survival and pro-angiogenic sphingosine 1-phosphate and decreases the amount of both ceramide and sphingosine, the pro-apoptotic sphingolipids. Moreover, its gene is oncogenic, its mRNA is overproduced in several solid tumors, its overexpression protects cells from apoptosis, and its activity is down-regulated by anti-cancer treatments. Therefore, the sphingosine kinase-1/sphingosine 1-phosphate signaling pathway appears to be a target of interest for therapeutic manipulation.

Urothelial carcinoma of the bladder, lipid cell variant: clinicopathologic findings and LOH analysis.

In this report, we present the clinicopathologic features of 27 cases of the lipid cell variant of urothelial bladder carcinoma. This is a rare variant of bladder cancer recognized by the current WHO classification of urologic tumors. The lipid cell component varied from 10% to 50% of the tumor specimen; in 11 cases the lipid cell component composed greater than 30% of the tumor. The architectural pattern of the tumor varied from solid expansile to infiltrative nests. The large epithelial tumor cells had an eccentrically placed nucleus and abundant vacuolated cytoplasm resembling signetring lipoblasts. Mucin stains were negative in all the cases. Typical features of high grade conventional urothelial carcinoma were present in all the cases with micropapillary or plasmacytoid carcinoma in 2 and 1 cases, respectively; extensive squamous or glandular differentiation was present in 2 additional cases. Most neoplastic cells had nuclei of intermediate nuclear grade with occasional nuclear pleomorphism. Immunohistochemical staining showed that the lipid cell component was positive for cytokeratins 7, 20, CAM 5.2, high molecular weight (34ssE12) and AE1/AE3, epithelial membrane antigen, and thrombomodulin; vimentin and S100 protein were negative. The loss of heterozygosity (LOH) analysis was done on 8 cases using 4 polymorphic microsatellite markers (D9S171, D9S177, IFNA, and TP 53); LOH at least in 1 marker was present in 6 cases. The LOH results were the same for lipid variant and conventional urothelial carcinoma. Pathologic stage was Ta (n=1), T1 (=2), T2, at least (n=7), T3a (n=4), T3b (n=8), and T4a (n=5). Electron microcopy analysis based on 2 cases supported lipid content in tumor cells. Follow-up information was available in all the cases, ranging from 6 to 58 months (mean, 28 mo). Sixteen of the patients died of disease at 16 to 58 months (mean, 33 mo) and 8 patients were alive with disease at 8 to 25 months (mean, 22 mo). Another 3 patients died of other causes at 6 to 15 months (mean, 10 mo). In summary, lipid cell urothelial bladder carcinoma is typically associated with advanced stage high-grade urothelial carcinoma, in which the prognosis is poor and clonally related to the concurrent conventional urothelial carcinoma. In limited samples, it may be misdiagnosed as liposarcoma, sarcomatoid carcinoma (carcinosarcoma), or signetring cell carcinoma. Morphologic distinction from other malignant neoplasms with lipid cell phenotype is critical for its clinical management.

Sphingosine kinase-1 is central to androgen-regulated prostate cancer growth and survival.

BACKGROUND: Sphingosine kinase-1 (SphK1) is an oncogenic lipid kinase notably involved in response to anticancer therapies in prostate cancer. Androgens regulate prostate cancer cell proliferation, and androgen deprivation therapy is the standard of care in the management of patients with advanced disease. Here, we explored the role of SphK1 in the regulation of androgen-dependent prostate cancer cell growth and survival. METHODOLOGY/PRINCIPAL FINDINGS: Short-term androgen removal induced a rapid and transient SphK1 inhibition associated with a reduced cell growth in vitro and in vivo, an event that was not observed in the hormono-insensitive PC-3 cells. Supporting the critical role of SphK1 inhibition in the rapid effect of androgen depletion, its overexpression could impair the cell growth decrease. Similarly, the addition of dihydrotestosterone (DHT) to androgen-deprived LNCaP cells re-established cell proliferation, through an androgen receptor/PI3K/Akt dependent stimulation of SphK1, and inhibition of SphK1 could markedly impede the effects of DHT. Conversely, long-term removal of androgen support in LNCaP and C4-2B cells resulted in a progressive increase in SphK1 expression and activity throughout the progression to androgen-independence state, which was characterized by the acquisition of a neuroendocrine (NE)-like cell phenotype. Importantly, inhibition of the PI3K/Akt pathway--by negatively impacting SphK1 activity--could prevent NE differentiation in both cell models, an event that could be mimicked by SphK1 inhibitors. Fascinatingly, the reversability of the NE phenotype by exposure to normal medium was linked with a pronounced inhibition of SphK1 activity. CONCLUSIONS/SIGNIFICANCE: We report the first evidence that androgen deprivation induces a differential effect on SphK1 activity in hormone-sensitive prostate cancer cell models. These results also suggest that SphK1 activation upon chronic androgen deprivation may serve as a compensatory mechanism allowing prostate cancer cells to survive in androgen-depleted environment, giving support to its inhibition as a potential therapeutic strategy to delay/prevent the transition to androgen-independent prostate cancer.

Maffucci syndrome and adrenal cortex tumor.

We report the second known case of Maffucci syndrome associated with an adrenal cortex tumor. Endocrine tumors have been reported in patients with multiple enchondromas, although the underlying mechanism of this combination is unknown. Maffucci syndrome is characterized by abnormalities of several mesodermal derivates. Therefore, routine evaluation for involvement of the adrenal cortex may be warranted to improve our knowledge of this syndrome and of its pathophysiology.