Current and Proposed Molecular Diagnostics in a Genitourinary Service Line Laboratory at a Tertiary Clinical Institution

Abstract: The idea that detailed knowledge of molecular oncogenesis will drive diagnostic, prognostic, and therapeutic clinical decision making in an increasingly multidisciplinary practice of oncologic care has been anticipated for many years. With the recent rapid advancement in our understanding of the molecular underpinnings of genitourinary malignancies, this concept is now starting to take shape in the fields of prostate, kidney, bladder, testicular, and penile cancer. Such breakthroughs necessitate the development of robust clinical-grade assays that can be quickly made available for patients to facilitate diagnosis in challenging cases, risk-stratify patients for subsequent clinical management, select the appropriate targeted therapy from among increasingly diverse and numerous options, and enroll patients in advanced clinical trials. This rapid translation of basic and clinical cancer research requires a streamlined, multidisciplinary approach to clinical assay development, termed here the molecular diagnostics service line laboratory. In this review, we summarize the current state and explore the future of molecular diagnostics in genitourinary oncology to conceptualize a genitourinary service line laboratory at a tertiary clinical institution. Key Words: Prostate cancer, kidney cancer, bladder cancer, testicular cancer, penile cancer, immunohistochemistry (IHC), fluorescent in situ hybridization (FISH), in situ hybridization (ISH), whole-genome sequencing (Cancer J 2014;20: 29Y42) T he genomic era is rapidly revolutionizing the practice of health care, and in almost no area is this more apparent than oncology, where molecular data are increasingly driving patient care in terms of diagnosis, prognosis, and therapeutics. The promise of personalized medicine, wherein the therapeutic options for an individual patient are tailored to his/her specific tumor genetics and biology, requires robust clinical assays for the biomarker(s) of interest. Our evolving understanding of the molecular underpinnings of urologic malignancies provides an emerging role for molecular testing in the treatment of these common neoplasms. Here, we envision the concept of a genitourinary service line laboratory at a tertiary clinical institution, and using an organ-based approach, we review the current state and explore the future of molecular diagnostics in genitourinary oncology. Genitourinary Service Line Laboratory: Concept and Services With the advent of widespread whole-genome sequencing of tumors, going forward, we expect the pace of molecular discoveries to quicken rather than abate. This trend will undoubtedly be associated with the need to bring advances made in the laboratory rapidly into the realm of routine clinical practice. We believe there are several ways this is already happening and will further develop in the future. As the cost of sequencing decreases, one such possibility is routine whole-genome sequencing of clinical tumor specimens, with the selection of therapeutics based on the prevalent targetable molecular alterations. A clinical sequencing pilot project, termed MI-ONCOSEQ, has already been established at the University of Michigan Health System (UMHS) for patients with advanced tumors that are resistant to conventional histologybased therapies. 1 Based on the results of this novel project, it seems clear to us that there are tumors that would benefit from this whole-genome sequencing approach. It is also evident that cancers arising in different organsVas well as different cancer subtypes within the same organ systemVare not uniformly similar but instead house different genetic drivers. In this age of translational medicine, it is imperative that discoveries from technologies such as clinical sequencing or traditional clinical cancer research be quickly incorporated into the development of clinical-grade assays in a CLIA (Clinical Laboratory Improvement Amendments)Ycertified environment. Broader availability of such assays will facilitate patient enrollment in a new generation of clinical trials that incorporate these rapid molecular advances and will expand the available pool of clinical sites beyond specialized academic centers. Hence, we propose the concept of dedicated molecular diagnostic service line laboratories, initially purposed for and centered around specific organ systems (i.e., genitourinary, pulmonary, etc.

Genital verruciform xanthoma: lessons from a contemporary multi‐institutional series

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/163435/2/his14198.pdfhttp://deepblue.lib.umich.edu/bitstream/2027.42/163435/1/his14198_am.pd

Clinical Utility and Concordance of Upper Urinary Tract Cytology and Biopsy in Predicting Clinicopathologic Features of Upper Urinary Tract Urothelial Carcinoma

5% of urothelial carcinoma occurs in the upper urinary tract (UUT), a challenging location to biopsy. We aim to evaluate concordance between biopsy, cytology, and resection specimens in a large upper tract urothelial carcinoma (UTUC) cohort.117 UTUC resections with UUT biopsy and/or cytology specimens from 2000–2016 were retrieved; pathologic material was re-reviewed, evaluated for concordance, and correlated with clinical information. 14% pre-operative biopsies, including 8 from renal pelvis and 6 from ureter, lacked neoplastic diagnoses. 77% diagnostic biopsies included subepithelial tissue; 11% demonstrated reclassification of grade and 30% demonstrated reclassification of invasion status. 26% of renal pelvis UTUC and 36% ureter UTUC were invasive only on resection. Of 18 UTUC reclassified from noninvasive high-grade papillary urothelial carcinoma (HGPUC) to invasive HGPUC, 39% had prior radical cystectomy (versus 8% invasive UTUC and 11% noninvasive UTUC with concordant biopsies). Most high-grade UTUC (88%) and some low-grade UTUC (58%) resections had abnormal cytology results. Biopsy-resection pairs with concordant invasion status and pairs with discordant invasion status showed similar rates of recurrence (38% versus 38%) and metastasis (25% versus 27%). 14% of UUT biopsies lacked diagnostic neoplastic material. Grade concordance between biopsy and resection was high (89%), but 30% of cases showed invasion only on resection. Subepithelial tissue was less commonly present in ureter biopsies, particularly from mid or proximal ureter. UTUC in patients with prior cystectomy were more likely to show invasion on resection but not biopsy

Somatic mutations of CADM1 in aldosterone-producing adenomas and gap junction-dependent regulation of aldosterone production

Aldosterone-producing adenomas (APAs) are the commonest curable cause of hypertension. Most have gain-of-function somatic mutations of ion channels or transporters. Herein we report the discovery, replication and phenotype of mutations in the neuronal cell adhesion gene CADM1. Independent whole exome sequencing of 40 and 81 APAs found intramembranous p.Val380Asp or p.Gly379Asp variants in two patients whose hypertension and periodic primary aldosteronism were cured by adrenalectomy. Replication identified two more APAs with each variant (total, n = 6). The most upregulated gene (10- to 25-fold) in human adrenocortical H295R cells transduced with the mutations (compared to wildtype) was CYP11B2 (aldosterone synthase), and biological rhythms were the most differentially expressed process. CADM1 knockdown or mutation inhibited gap junction (GJ)-permeable dye transfer. GJ blockade by Gap27 increased CYP11B2 similarly to CADM1 mutation. Human adrenal zona glomerulosa (ZG) expression of GJA1 (the main GJ protein) was patchy, and annular GJs (sequelae of GJ communication) were less prominent in CYP11B2-positive micronodules than adjacent ZG. Somatic mutations of CADM1 cause reversible hypertension and reveal a role for GJ communication in suppressing physiological aldosterone production

Serial monitoring of genomic alterations in circulating tumor cells of ER-positive/HER2-negative advanced breast cancer: feasibility of precision oncology biomarker detection.

Nearly all estrogen receptor (ER)-positive (POS) metastatic breast cancers become refractory to endocrine (ET) and other therapies, leading to lethal disease presumably due to evolving genomic alterations. Timely monitoring of the molecular events associated with response/progression by serial tissue biopsies is logistically difficult. Use of liquid biopsies, including circulating tumor cells (CTC) and circulating tumor DNA (ctDNA), might provide highly informative, yet easily obtainable, evidence for better precision oncology care. Although ctDNA profiling has been well investigated, the CTC precision oncology genomic landscape and the advantages it may offer over ctDNA in ER-POS breast cancer remain largely unexplored. Whole-blood (WB) specimens were collected at serial time points from patients with advanced ER-POS/HER2-negative (NEG) advanced breast cancer in a phase I trial of AZD9496, an oral selective ER degrader (SERD) ET. Individual CTC were isolated from WB using tandem CellSearch® /DEPArray™ technologies and genomically profiled by targeted single-cell DNA next-generation sequencing (scNGS). High-quality CTC (n = 123) from 12 patients profiled by scNGS showed 100% concordance with ctDNA detection of driver estrogen receptor α (ESR1) mutations. We developed a novel CTC-based framework for precision medicine actionability reporting (MI-CTCseq) that incorporates novel features, such as clonal predominance and zygosity of targetable alterations, both unambiguously identifiable in CTC compared to ctDNA. Thus, we nominated opportunities for targeted therapies in 73% of patients, directed at alterations in phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA), fibroblast growth factor receptor 2 (FGFR2), and KIT proto-oncogene, receptor tyrosine kinase (KIT). Intrapatient, inter-CTC genomic heterogeneity was observed, at times between time points, in subclonal alterations. Our analysis suggests that serial monitoring of the CTC genome is feasible and should enable real-time tracking of tumor evolution during progression, permitting more combination precision medicine interventions

Metastatic renal cell carcinoma, clear cell type, of the parotid gland

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/109307/1/dc23103.pd

Double somatic mutations in CTNNB1 and GNA11 in an aldosterone-producing adenoma

Double somatic mutations in CTNNB1 and GNA11/Q have recently been identified in a small subset of aldosterone-producing adenomas (APAs). As a possible pathogenesis of APA due to these mutations, an association with pregnancy, menopause, or puberty has been proposed. However, because of its rarity, characteristics of APA with these mutations have not been well characterized. A 46-year-old Japanese woman presented with hypertension and hypokalemia. She had two pregnancies in the past but had no history of pregnancy-induced hypertension. She had regular menstrual cycle at presentation and was diagnosed as having primary aldosteronism after endocrinologic examinations. Computed tomography revealed a 2 cm right adrenal mass. Adrenal venous sampling demonstrated excess aldosterone production from the right adrenal gland. She underwent right laparoscopic adrenalectomy. The resected right adrenal tumor was histologically diagnosed as adrenocortical adenoma and subsequent immunohistochemistry (IHC) revealed diffuse immunoreactivity of aldosterone synthase (CYP11B2) and visinin like 1, a marker of the zona glomerulosa (ZG), whereas 11β-hydroxylase, a steroidogenic enzyme for cortisol biosynthesis, was mostly negative. CYP11B2 IHC-guided targeted next-generation sequencing identified somatic CTNNB1 (p.D32Y) and GNA11 (p.Q209H) mutations. Immunofluorescence staining of the tumor also revealed the presence of activated β-catenin, consistent with features of the normal ZG. The expression patterns of steroidogenic enzymes and related proteins indicated ZG features of the tumor cells. PA was clinically and biochemically cured after surgery. In conclusion, our study indicated that CTNNB1 and GNA11-mutated APA has characteristics of the ZG. The disease could occur in adults with no clear association with pregnancy or menopause

Frequent discordance between ERG gene rearrangement and ERG protein expression in a rapid autopsy cohort of patients with lethal, metastatic, castration‐resistant prostate cancer

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/108006/1/pros22836.pd