Francesca Khani

Evidence for Molecular Differences in Prostate Cancer between African American and Caucasian Men

Purpose: The aim of this study was to compare the frequency of ERG rearrangement, PTEN deletion, SPINK1 overexpression, and SPOP mutation in prostate cancer in African American and Caucasian men. Experimental design: Dominant tumor nodules from radical prostatectomy specimens of 105 African American men (AAM) were compared with 113 dominant nodules from Caucasian men (CaM). Clinical and pathologic characteristics of the two groups were similar. SPINK1 overexpression was evaluated by immunohistochemistry, ERG rearrangement and PTEN deletion by FISH, and SPOP mutation by Sanger sequencing. Results: ERG rearrangement was identified in 48 of 113 tumors (42.5%) in CaM and 29 of 105 tumors (27.6%) in AAM (P = 0.024). PTEN deletion was seen in 19 of 96 tumors (19.8%) in CaM and 7 of 101 tumors (6.9%) in AAM (P = 0.011). SPINK1 overexpression was present in 9 of 110 tumors (8.2%) in CaM and 25 of 105 tumors (23.4%) in AAM (P = 0.002). SPOP mutation was identified in 8 of 78 (10.3%) tumors in CaM and 4 of 88 (4.5%) tumors in AAM (P = 0.230). When adjusted for age, body mass index, Gleason score, and pathologic stage, ERG rearrangement and SPINK1 overexpression remain significantly different (P = 0.018 and P = 0.008, respectively), and differences in PTEN deletion and SPOP mutation approach significance (P = 0.061 and P = 0.087, respectively). Conclusions: Significant molecular differences exist between prostate cancers in AAM and CaM. SPINK1 overexpression, an alteration associated with more aggressive prostate cancers, was more frequent in AAM, whereas ERG rearrangement and PTEN deletion were less frequent in this cohort. Further investigation is warranted to determine whether these molecular differences explain some of the disparity in incidence and mortality between these two ethnic groups. Clin Cancer Res; 20(18); 4925–34. ©2014 AACR.

Insulin-like growth factor messenger RNA-binding protein 3 expression helps prognostication in patients with upper tract urothelial carcinoma.

BACKGROUND Upper tract urothelial carcinoma (UTUC) is a clinically heterogeneous disease that lacks high-quality trials that provide definitive prognostic markers. Insulin-like growth factor messenger RNA binding protein 3 (IMP3) has been associated with outcomes in urothelial carcinoma of the bladder but was not yet studied in UTUC. OBJECTIVE To evaluate the association of the oncofetal protein IMP3 with oncologic outcomes in patients with UTUC treated with radical nephroureterectomy (RNU). DESIGN, SETTING, AND PARTICIPANTS We investigated the expression of IMP3 and its association with clinical outcomes using tissue microarrays constructed from 622 patients treated with RNU at seven international institutions between 1991 and 2008. INTERVENTION All patients were diagnosed with UTUC and underwent RNU. OUTCOME MEASUREMENT AND STATISTICAL ANALYSIS Uni- and multivariable Cox regression analyses evaluated the association of IMP3 protein expression with disease recurrence, cancer-specific mortality, and all-cause mortality. RESULTS AND LIMITATIONS IMP3 was expressed in 12.2% of patients with UTUC (n=76). The expression was tumor specific and correlated with higher stages/grades. Within a median follow-up of 27 mo (interquartile range [IQR]: 12-53), 191 patients (25.4%) experienced disease recurrence, and 165 (21.9%) died of the disease. Patients with IMP3 demonstrated significantly worse recurrence-free survival (27.4% vs 75.1%; p<0.01), cancer-specific survival (34.5% vs 78.9%; p<0.01), and overall survival (15.6% vs 64.8%; p<0.01) at 5 yr compared with those without IMP3. In multivariable Cox regression analyses, which adjusted for the effects of standard clinicopathologic features, IMP3 expression was independently associated with disease recurrence (hazard ratio [HR]: 1.87; p<0.01), cancer-specific mortality (HR: 2.15; p<0.01), and all-cause mortality (HR: 2.07; p<0.01). Major limitations include the retrospective design and relatively short follow-up time. CONCLUSIONS IMP3 expression is independently associated with disease recurrence, cancer-specific mortality, and all-cause mortality in UTUC. IMP3 may help improve risk stratification and prognostication of UTUC patients treated with RNU.

Exome Sequencing of African-American Prostate Cancer Reveals Loss-of-Function ERF Mutations

African-American men have the highest incidence of and mortality from prostate cancer. Whether a biological basis exists for this disparity remains unclear. Exome sequencing (n = 102) and targeted validation (n = 90) of localized primary hormone-naïve prostate cancer in African-American men identified several gene mutations not previously observed in this context, including recurrent loss-of-function mutations in ERF, an ETS transcriptional repressor, in 5% of cases. Analysis of existing prostate cancer cohorts revealed ERF deletions in 3% of primary prostate cancers and mutations or deletions in ERF in 3% to 5% of lethal castration-resistant prostate cancers. Knockdown of ERF confers increased anchorage-independent growth and generates a gene expression signature associated with oncogenic ETS activation and androgen signaling. Together, these results suggest that ERF is a prostate cancer tumor-suppressor gene. More generally, our findings support the application of systematic cancer genomic characterization in settings of broader ancestral diversity to enhance discovery and, eventually, therapeutic applications.Significance: Systematic genomic sequencing of prostate cancer in African-American men revealed new insights into prostate cancer, including the identification of ERF as a prostate cancer gene; somatic copy-number alteration differences; and uncommon PIK3CA and PTEN alterations. This study highlights the importance of inclusion of underrepresented minorities in cancer sequencing studies. Cancer Discov; 7(9); 973-83. ©2017 AACR.This article is highlighted in the In This Issue feature, p. 920.

Association of Oncofetal Protein Expression with Clinical Outcomes in Patients with Urothelial Carcinoma of the Bladder

PURPOSE Oncofetal proteins are expressed in the developing embryo. Oncofetal protein expression correlates with the clinical outcome of nonmuscle invasive bladder urothelial carcinoma. IMP3, MAGE-A, glypican-3 and TPBG are oncofetal proteins that have not been well characterized in urothelial carcinoma of the bladder. MATERIALS AND METHODS We investigated the expression of these 4 proteins and their association with clinical outcomes using tissue microarrays from 384 consecutive patients treated with radical cystectomy between 1988 and 2003 at 1 academic center. We stained for IMP3, MAGE-A, glypican-3 and TPBG. Univariable and multivariable Cox regression analyses were done to evaluate the association of oncofetal protein expression with disease recurrence and cancer specific mortality. RESULTS IMP3, MAGE-A, glypican-3 and TPBG were expressed in 39.5%, 45%, 6% and 85% of urothelial bladder carcinomas, respectively. Expression was tumor specific and did not correlate with pathological features except for TPBG. At a median followup of 128 months 176 patients (46%) experienced disease recurrence, 175 (45.5%) had died of the disease and 96 (27.5%) had died of another cause. On univariable analysis IMP3 and MAGE-A expression was associated with an increased risk of disease recurrence (p <0.001 and 0.03) and cancer specific mortality (p = 0.004 and 0.03, respectively). On multivariable Cox regression analysis adjusted for the effects of standard clinicopathological features IMP3 and MAGE-A expression was independently associated with disease recurrence (p = 0.004, HR 1.55, 95% CI 1.15-2.11 and p = 0.02, HR 1.44, 95% CI 1.05-1.99, respectively) but not with cancer specific mortality. CONCLUSIONS Oncofetal proteins are commonly and differentially expressed in urothelial carcinoma of the bladder compared to normal urothelium. IMP3 and MAGE-A expression was associated with disease recurrence and cancer specific mortality but glypican-3 and TPBG expression was not.

A “Chicken or Egg” Conundrum: Race, Molecular Subtype, and Tumor Location in Prostate Cancer

It is well known that differences in prostate cancer (PCa) incidence and prognosis exist among men of different ethnicities, and African American (AA) men have the highest PCa incidence and mortality compared to all other ethnic groups in the USA. While it has been shown that a variety of socioeconomic factors contribute to disparities in clinical outcomes, such external factors do not entirely account for these disparities [1]. Underlying differences in tumor biology among men of different ethnicities have been increasingly demonstrated in recent research, with comparative studies of tumors from AA cohorts having been most frequently investigated to date. In the era of new and evolving molecular diagnostic modalities and targeted treatments for PCa, identification of ethnic differences in the molecular classification of this disease is an essential step. In this issue of European Urology, Faisal and colleagues [2] provide further validation of known molecular differences in PCa between AA and Caucasian American (CA) men in a multi-institutional cohort of radical prostatectomy specimens from 121 AA and 179 CA men. Several studies have shown that ERG rearrangements are approximately half as frequent in tumors from AAmen as in tumors CA men, and the current findings further support the growing body of literature in this area [1,3]. In addition to ERG rearrangements, non-ERG ETS rearrangementswere also evaluated, including ETV1, ETV4, ETV5, and FLI1. The authors previously reported a higher frequency of these non-ERG ETS rearrangements in low-risk PCa samples from AA men compared to CA men (14% vs 2%) [4]. It is important to point out that the higher prevalence of

YBX2 Dysregulation is a Potential Cause for Late Maturation Arrest in Men with Non-Obstructive Azoospermia

Introduction YBX2 protein binds to Y-box promotors and to mRNAs in the cytoplasm of pachytene spermatocytes and round spermatids in rodents. Knock-out of YBX2 leads to maturation arrest in animal models. YBX2 binds PRM1 and 2 mRNA, which is transcribed early and sequestrated for translation during late spermatogenesis. Objective This study aimed to determine if the loss of YBX2 is associated with MA arrest due to the loss of sequestration of protamines in the human testis. As a second aim, we examined the expression of YBX2, and its transcription factors in maturation arrest (MA)(early and late) and normal controls in men. Methods RNAseq was performed using RNA extracted from human testis samples from 44 men with non-obstructive azoospermia and ten from healthy controls. Differential expression was performed using JMPgenomics, FDR<0.001. FANTOM5 was used to predict enhancers and inhibitors of YBX2 expression. Immunofluorescence (IF) was used to stain testis tissue sections with antibodies against YBX2, SYCP3, and PRM2 in normal and MA samples. Flow cytometry was utilized to characterize YBX2 positive cells. Results Expression of YBX2 mRNA was significantly downregulated in early and late MA compared to controls. Surprisingly, PRM1&2 mRNAs were also depleted in men with MA. Multifactorial regression analysis demonstrated a decrease in YBX2 expression in MA is due to decrease in COMP levels (p<0.0001). IF localized YBX2 protein in spermatocytes and round spermatids among fertile men, with rare YBX2 positive spermatocytes stained in LMA. PRM1&2 proteins were absent or abnormally sequestrated within spermatocytes. Conclusions Decrease in YBX2 protein expression in men with LMA leads to loss of translational suppression and lack of PRM1 and PRM2 necessary to complete spermatogenesis.

PD46-08 CLEAR CELL PAPILLARY RENAL CELL CARCINOMA: GENOMIC CHARACTERIZATION OF A DISTINCT AND INDOLENT TUMOR

INTRODUCTION AND OBJECTIVES: Clear cell papillary renal cell carcinoma (CCP-RCC) represents the fourth most common renal cell carcinoma (RCC) after conventional clear cell, papillary and chromophobe RCC. Although patients with CCP-RCC behave in an indolent fashion in reported cases, long-term clinical outcomes remain unclear. Also, only limited molecular information is available in CCP-RCC, which has primarily been focused on VHL alterations. In this study we aimed to better understand the mutational landscape of CCP-RCC and provide extended follow up information. METHODS: A retrospective search of our institutional database from 1997-2017 yielded 64 cases of CCP-RCC treated by surgical resection. Follow up was obtained for all patients. 17 cases that showed classic morphology and immunoprofile of CCP-RCC were selected for molecular testing. We used the Oncomine Comprehensive Assay v3 (ThermoFisher) that covers 161 cancer-related genes including 51 fusion drivers, 91 copy number variants, 48 full-length genes and 87 hotspot mutations. HDx positive controls were used and 90% of expected variants were detected. Average read depth per sample was 1418x. RESULTS: For the cohort of 64 CCP-RCC, the mean patient age was 60 years (range 34-82) with a 1.6:1 male:female ratio. CCPRCC was identified in the context of end-stage renal disease in 29.7% of the cases. No recurrence or metastasis was detected in any case after a mean follow up of 57 months (range 1-240 months). In the subset of 17 CCP-RCC that underwent sequencing, seven cases (41.1%) showed no mutations in the cancer-related genes tested, eight (47.1%) cases had a single mutation, and only two (11.8%) cases had 2 mutations (Table 1). No fusion drivers were detected. CONCLUSIONS: All 64 patients with CCP-RCC had indolent behavior after a maximum follow up of twenty years. Targeted sequencing of 17 cases showed recurrent somatic mutations in ARID1A, NOTCH3, KIT and TSC1 in subsets of tumors. Nearly half of the cases had no detectable mutation at all. This study is the largest and most comprehensive molecular characterization of CCP-RCC to date and reports the longest available follow up. Our findings support the notion that CCP-RCC harbors a unique mutational landscape and follows an indolent clinical course.

Grading, Staging, and Morphologic Risk Stratification of Bladder Cancer

Histopathologic assessment of bladder tissue remains the gold standard for diagnosis of urothelial neoplasms. The histologic grade and stage that are assigned to a tumor not only dictate the patient’s treatment but also provide important prognostic information. As will be discussed in detail elsewhere in this book, the grade and morphologic appearance of a tumor are also frequently associated with important underlying molecular alterations that may indicate sensitivity or resistance to certain treatment regimens. This chapter will discuss the histopathologic features of urothelial neoplasia, including grade and stage, which are critical to understanding the clinical and molecular aspects of the disease.

Pathology of Prostate Cancer

Although molecular pathology is a fast-growing and important area in the world of prostate cancer, the diagnosis of prostate cancer still requires histologic confirmation, and the foundation for virtually all treatment and research is still based on the tumor’s histomorphology. This chapter focuses on the pathology of prostate cancer, beginning with the diagnostic features of prostate cancer under the microscope. We also discuss the Gleason grading system as well as the newly adopted Grade Groups, which are prognostic groups that incorporate the modifications made to the Gleason system over the decades, provide a more meaningful framework for research, and allow for better patient understanding of their disease. The more common and clinically significant histologic variants of acinar and non-acinar prostate cancer are also reviewed, and applications of immunohistochemistry in prostate cancer diagnosis are also discussed. Treatment-related changes, such as those following radiation or hormone therapy, are examined as these can drastically change the morphology of the tumor as well as provide prognostic information. Finally, key aspects related to pathologic specimens, including their handling and the reporting of prostate cancer therein, are reviewed.

A Precious Diagnostic “Pearl”: The Necklace Pattern in Germ Cell Tumors of the Testis

Diffuse embryoma is a rare pattern of nonseminomatous germ cell tumor of the testis originally described in 1983. We report a case with this predominant pattern in an 18-year-old male with a painless palpable testicular mass. Although it is relatively common to see a diffuse embryoma pattern focally in mixed nonseminomatous germ cell tumors of the testis, it is rarely the predominant pattern and can represent a diagnostic pitfall on routine hematoxylin and eosin stain. We emphasize the importance of recognizing the individual components within the diffuse embryoma pattern, review the literature, and briefly discuss the ancillary immunohistochemical stains that may be utilized to help support the diagnosis.