Kanishka Sircar

Expression of RANKL/RANK/OPG in primary and metastatic human prostate cancer as markers of disease stage and functional regulation.

Late‐stage prostate cancer patients are refractory to hormone therapy and exhibit a high propensity to develop skeletal metastasis. In this regard, the role of a novel cytokine system belonging to the tumor necrosis factor (TNF) family that is critical for osteoclastic osteolysis and that consists of receptor activator of NF‐κB ligand (RANKL), its receptor (RANK), and decoy receptor osteoprotegerin (OPG) is of potential interest.

PTEN genomic deletion is associated with p-Akt and AR signalling in poorer outcome, hormone refractory prostate cancer

PTEN haploinsufficiency is common in hormone‐sensitive prostate cancer, though the incidence of genomic deletion and its downstream effects have not been elucidated in clinical samples of hormone refractory prostate cancer (HRPC). Progression to androgen independence is pivotal in prostate cancer and mediated largely by the androgen receptor (AR). Since this process is distinct from metastatic progression, we examined alterations of the PTEN gene in locally advanced recurrent, non‐metastatic human HRPC tissues. Retrospective analyses of PTEN deletion status were correlated with activated downstream phospho‐Akt (p‐Akt) pathway proteins and with the androgen receptor. The prevalence of PTEN genomic deletions in transurethral resection samples of 59 HRPC patients with known clinical outcome was assessed by four‐colour FISH analyses. FISH was performed using six BAC clones spanning both flanking PTEN genomic regions and the PTEN gene locus, and a chromosome 10 centromeric probe. PTEN copy number was also evaluated in a subset of cases using single nucleotide polymorphism (SNP) arrays. In addition, the samples were immunostained with antibodies against p‐Akt, p‐mTOR, p‐70S6, and AR. The PTEN gene was deleted in 77% of cases, with 25% showing homozygous deletions, 18% homozygous and hemizygous deletions, and 34% hemizygous deletions only. In a subset of the study group, SNP array analysis confirmed the FISH findings. PTEN genomic deletion was significantly correlated to the expression of downstream p‐Akt (p < 0.0001), AR (p = 0.025), and to cancer‐specific mortality (p = 0.039). PTEN deletion is common in HRPC, with bi‐allelic loss correlating to disease‐specific mortality and associated with Akt and AR deregulation. Copyright

Targeting MET and AXL overcomes resistance to sunitinib therapy in renal cell carcinoma.

Antiangiogenic therapy resistance occurs frequently in patients with metastatic renal cell carcinoma (RCC). The purpose of this study was to understand the mechanism of resistance to sunitinib, an antiangiogenic small molecule, and to exploit this mechanism therapeutically. We hypothesized that sunitinib-induced upregulation of the prometastatic MET and AXL receptors is associated with resistance to sunitinib and with more aggressive tumor behavior. In the present study, tissue microarrays containing sunitinib-treated and untreated RCC tissues were stained with MET and AXL antibodies. The low malignant RCC cell line 786-O was chronically treated with sunitinib and assayed for AXL, MET, epithelial–mesenchymal transition (EMT) protein expression and activation. Co-culture experiments were used to examine the effect of sunitinib pretreatment on endothelial cell growth. The effects of AXL and MET were evaluated in various cell-based models by short hairpin RNA or inhibition by cabozantinib, the multi-tyrosine kinases inhibitor that targets vascular endothelial growth factor receptor, MET and AXL. Xenograft mouse models tested the ability of cabozantinib to rescue sunitinib resistance. We demonstrated that increased AXL and MET expression was associated with inferior clinical outcome in patients. Chronic sunitinib treatment of RCC cell lines activated both AXL and MET, induced EMT-associated gene expression changes, including upregulation of Snail and β-catenin, and increased cell migration and invasion. Pretreatment with sunitinib enhanced angiogenesis in 786-0/human umbilical vein endothelial cell co-culture models. The suppression of AXL or MET expression and the inhibition of AXL and MET activation using cabozantinib both impaired chronic sunitinib treatment-induced prometastatic behavior in cell culture and rescued acquired resistance to sunitinib in xenograft models. In summary, chronic sunitinib treatment induces the activation of AXL and MET signaling and promotes prometastatic behavior and angiogenesis. The inhibition of AXL and MET activity may overcome resistance induced by prolonged sunitinib therapy in metastatic RCC.

Everolimus Versus Sunitinib Prospective Evaluation in Metastatic Non–Clear Cell Renal Cell Carcinoma (ESPN): A Randomized Multicenter Phase 2 Trial

BACKGROUND Sunitinib and everolimus are standard first-line and second-line therapies, respectively, in clear cell renal cell carcinoma (ccRCC). OBJECTIVE To conduct a randomized phase 2 trial comparing sunitinib and everolimus in non-clear cell RCC (non-ccRCC). DESIGN, SETTING, AND PARTICIPANTS Patients with metastatic, non-ccRCC, or ccRCC with >20% sarcomatoid features (ccSRCC) were randomized to receive sunitinib or everolimus with crossover at disease progression. OUTCOME MEASUREMENT AND STATISTICAL ANALYSIS Primary end point was progression-free survival (PFS) in first-line therapy; 108 patients were needed to show improvement in median PFS (mPFS) from 12 wk with sunitinib to 20 wk with everolimus. RESULTS AND LIMITATIONS Interim analysis of 68 patients (papillary [27], chromophobe [12], unclassified [10], translocation [7], ccSRCC [12]) prompted early trial closure. The mPFS in first-line therapy was 6.1 mo with sunitinib and 4.1 mo with everolimus (p=0.6); median overall survival (mOS) was not reached with sunitinib and was 10.5 mo with everolimus, respectively (p=0.014). At final analysis, mOS was 16.2 and 14.9 mo with sunitinib and everolimus, respectively (p=0.18). There were four partial responses (PRs) in first-line therapy (sunitinib: 3 of 33 [9%]; everolimus, 1 of 35 [2.8%]) and four PRs in second-line therapy (sunitinib: 2 of 21 [9.5%]; everolimus, 2 of 23 [8.6%]), with mPFS of 1.8 mo and 2.8 mo, respectively. In patients without sarcomatoid features in their tumors (n=49), mOS was 31.6 mo with sunitinib and 10.5 mo with everolimus (p=0.075). Genomic profiling of a chromophobe RCC from a patient with a PR to first-line everolimus revealed a somatic TSC2 mutation. CONCLUSIONS In this trial, everolimus was not superior to sunitinib. Both agents demonstrated modest efficacy, underscoring the need for better therapies in non-ccRCC. PATIENT SUMMARY This randomized phase 2 trial provides the first head-to-head comparison of everolimus and sunitinib in patients with metastatic non-clear cell renal cell carcinoma (non-ccRCC). The observed very modest efficacy underscores the need to develop more effective therapies for non-ccRCC.

Modeling a lethal prostate cancer variant with small-cell carcinoma features

Purpose: Small-cell prostate carcinoma (SCPC) morphology predicts for a distinct clinical behavior, resistance to androgen ablation, and frequent but short responses to chemotherapy. We sought to develop model systems that reflect human SCPC and can improve our understanding of its biology. Experimental Design: We developed a set of castration-resistant prostate carcinomas xenografts and examined their fidelity to their human tumors of origin. We compared the expression and genomic profiles of SCPC and large-cell neuroendocrine carcinoma (LCNEC) xenografts to those of typical prostate adenocarcinoma xenografts. Results were validated immunohistochemically in a panel of 60 human tumors. Results: The reported SCPC and LCNEC xenografts retain high fidelity to their human tumors of origin and are characterized by a marked upregulation of UBE2C and other mitotic genes in the absence of androgen receptor (AR), retinoblastoma (RB1), and cyclin D1 (CCND1) expression. We confirmed these findings in a panel of samples of CRPC patients. In addition, array comparative genomic hybridization of the xenografts showed that the SCPC/LCNEC tumors display more copy number variations than the adenocarcinoma counterparts. Amplification of the UBE2C locus and microdeletions of RB1 were present in a subset, but none displayed AR nor CCND1 deletions. The AR, RB1, and CCND1 promoters showed no CpG methylation in the SCPC xenografts. Conclusion: Modeling human prostate carcinoma with xenografts allows in-depth and detailed studies of its underlying biology. The detailed clinical annotation of the donor tumors enables associations of anticipated relevance to be made. Future studies in the xenografts will address the functional significance of the findings. Clin Cancer Res; 18(3); 666–77. ©2011 AACR.

Adrenocortical carcinoma: Clinical outcomes and prognosis of 330 patients at a tertiary care center

OBJECTIVE Adrenocortical carcinoma (ACC) is a rare malignancy with a poor prognosis. Herein, we describe the clinical features and outcomes for a large series of ACC patients. DESIGN AND METHODS Retrospective review of ACC patients seen at The University of Texas MD Anderson Cancer Center from 1998 through 2011. RESULTS A total of 330 patients with median age at diagnosis of 48.5 years; 12 (3.6%) patients were under 18 years. Hormonally functioning tumors represented 41.8% (n=138) of all cases. Surgical resection for the primary tumor was done in 275 (83.3%) patients (45 at MD Anderson (16.4%)). For those who had surgical resection, the median local-recurrence-free time was 1.04 years. Factors associated with local recurrence included positive surgical margins (P=0.007) and advanced disease stage (P=0.026). Median overall survival time for all patients was 3.21 years. Median survival times were 24.1, 6.08, 3.47, and 0.89 years for stages I, II, III, and IV respectively. In multivariable analysis, older age, functioning tumors, and higher disease stage remained significant prognostic factors associated with poor survival. CONCLUSION ACC prognosis remains poor with the use of currently available treatments. Older age, functioning tumors, and incomplete resections are clinical factors associated with poor survival. Surgical expertise is important to achieve complete resections and to improve outcome.

Bladder neck involvement in pathological stage pT4 radical prostatectomy specimens is not an independent prognostic factor.

PURPOSE Bladder neck invasion by prostate cancer in radical prostatectomy specimens is uncommon and, thus, its influence on disease recurrence has not been well defined. Consequently the classification of bladder neck invasion in the TNM staging system is controversial. We studied our cohort of patients with stage pT4 disease and bladder neck invasion to clarify the true clinical behavior and prognostic significance of bladder neck invasion in radical prostatectomy specimens. MATERIALS AND METHODS The study group consisted of 4,090 consecutive patients treated with radical prostatectomy at one of our institutions between 1983 and 2001. Median followup was 53.1 months (range 1 to 189). After excluding from analysis patients treated with neoadjuvant androgen withdrawal or preoperative irradiation 72 of the remaining 2,571 (2.8%) with bladder neck invasion were classified with stage pT4 disease and their specimens were reviewed. Progression-free probability was determined by Kaplan-Meier analysis. Using the Cox proportional hazards model the independent prognostic significance of bladder neck invasion was assessed after controlling for pretreatment prostate specific antigen, final Gleason sum, extracapsular extension, surgical margins status, seminal vesicle invasion and lymph node involvement. RESULTS Of the 72 patients categorized with stage pT4 disease 14 (19%) had poorly differentiated Gleason sum 8 to 10 cancer, 38 (53%) had established extracapsular extension, 24 (33%) had seminal vesicle invasion and 8 (11%) had lymph node involvement. However, 26 patients (36%) had cancer confined to the prostate and 28 (39%) had negative surgical margins except for the bladder neck site. The mean 5-year progression-free probability plus or minus SD in all stage pT4 cases was 68% +/- 7%, which was better than in cases of seminal vesicle invasion (52% +/- 5%, log rank test p = 0.0156) but worse than in those of extracapsular extension (84% +/- 4.1%). Univariate analysis of the stage pT4 cohort revealed that higher prostatectomy Gleason sum, more extensive extracapsular extension and seminal vesicle invasion were significantly associated with an adverse prognosis. However, in a multivariate model that included all radical prostatectomy cases the finding of bladder neck invasion or stage pT4 disease did not independently predict prostate specific antigen recurrence. CONCLUSIONS Stage pT4 disease comprises a heterogeneous group of tumors with various pathological features and inconsistent outcomes. Assigning the pT4 stage to cases of microscopic bladder neck invasion provides no independent ability for predicting disease progression after adjusting for other adverse disease features. Due to this and previously reported data the definition of stage pT4 disease should be modified in the next version of the TNM staging system.

Plasmacytoid Urothelial Carcinoma, a Chemosensitive Cancer with Poor Prognosis, and Peritoneal Carcinomatosis

PURPOSE Plasmacytoid urothelial carcinoma is a rare variant histology with poorly defined clinical behavior. We report clinical outcome information on patients with predominant plasmacytoid urothelial carcinoma. MATERIALS AND METHODS We retrospectively analyzed treatments and outcomes in patients with predominant plasmacytoid urothelial carcinoma seen at our institution from 1990 through 2010. The Kaplan-Meier method was used to calculate overall and progression-free survival. RESULTS We identified 31 patients with a median age of 63.5 years, of whom 83.3% were male. TNM stage was cT1N0 in 4 patients, cT2N0 in 7, cT3b-4aN0 in 5 and cT4b, N+ or M+ in 15. Median overall survival was 17.7 months (stage I-III vs IV 45.8 vs 13.3). Five of the 16 patients with potentially surgically resectable plasmacytoid urothelial carcinoma (pT4aN0M0 or less) received neoadjuvant chemotherapy, 10 underwent initial surgery and 1 was treated only with transurethral resection of bladder tumor. Despite pathological down staging in 80% of the patients who received neoadjuvant chemotherapy, relapses were common. There was no survival difference between patients treated with neoadjuvant chemotherapy or initial surgery, although 7 received adjuvant chemotherapy. Surgical up staging with positive margins was also common for surgery alone. The most common site of recurrence was in the peritoneum (19 of 23 patients) with relapses even in those with a pathological complete response at surgery. In patients who presented with metastatic disease and were treated with chemotherapy median survival was 12.6 months. CONCLUSIONS Plasmacytoid urothelial carcinoma is an aggressive subset with overall poor outcomes. Although down staging is seen with neoadjuvant chemotherapy, there are few long-term survivors. There is a strong predilection for recurrence along the peritoneal lining.

Fascin regulates prostate cancer cell invasion and is associated with metastasis and biochemical failure in prostate cancer.

Purpose: Prostate cancer metastasis to secondary organs is considered an initial event in the development of hormone refractory disease and remains the major cause of death among prostate cancer patients. In this study, we investigated the role of fascin, a cytoskeleton actin–bundling protein involved in the formation of filopodia and cell migration, in prostate cancer progression. Experimental Design: Fascin protein expression was examined by immunohistochemistry in a cohort of 196 patients with localized prostate cancer and across several stages of disease progression, including hormone refractory disease. Cellular changes were also assessed in vitro and in vivo in DU145 prostate cancer cell line using fascin gene silencing. Results: Fascin epithelial expression was significantly up-regulated in localized and hormone refractory prostate cancer compared with benign prostate tissue (P < 0.05). Furthermore, high fascin expression was associated with an increased rate of prostate-specific antigen recurrence following radical prostatectomy (P = 0.075), signifying more aggressive clinical course, thus supporting a function for fascin in prostate cancer progression. In cellular models, fascin gene silencing using small interfering RNA in the androgen-independent prostate cancer cell line DU145 decreased cell motility and invasiveness while increasing cell adhesive properties. In addition, fascin small interfering RNA–expressing DU145 cells implanted orthotopically in mouse prostate showed significantly decreased growth (P < 0.005) and drastically prevented the formation of lymph node metastases (P < 0.001) compared with their matched controls. Conclusions: Our data show a function of fascin in the regulation of prostate cancer progression and emphasize the importance of fascin as a prognostic marker for aggressive disease and as a potential therapeutic target for advanced androgen independent disease.

Somatic Mosaicism and Cancer: A Micro-Genetic Examination into the Role of the Androgen Receptor Gene in Prostate Cancer

Recent evidence has shown that the androgen receptor (AR) plays a major role in all prostate cancer stages, including both androgen-dependent and -independent tumors. A large number of studies have examined the possible effects of a functional polymorphism in the AR gene, a variable-length CAG repeat, on the development of prostate cancer, but the results to date have been inconclusive. We have considered the fact that the tissue heterogeneity present in almost all prostate cancer tumors has rarely been regarded as an indicator of AR genetic heterogeneity. To determine if genetic heterogeneity exists and is a significant event in prostate cancer development, we have examined prostate cancer tumors for somatic shortening of the AR gene CAG repeat. All 72 laser capture microdissected samples from archival prostate cancer tissues, as well as samples from freshly prepared prostate cancer tissues, showed some genetic heterogeneity (somatic mosaicism) for AR CAG repeat length. Cancerous tissues showed a much greater degree of genetic heterogeneity than adjacent benign tissues, as well as a very significant shortening of their CAG repeat lengths. However, CAG repeat length heterogeneity was not observed in normal prostate tissues. It is hypothesized that somatic mosaicism of the AR CAG repeat in prostate cancer tumors may be found to be an important genetic event in precancerous tissue, which may subsequently lead to the development of prostate cancer.