A Contemporary Update on Pathology Standards for Bladder Cancer: Transurethral Resection and Radical Cystectomy Specimens

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1 EUROPEAN UROLOGY 63 (2013) available at journal homepage: Collaborative Review Bladder Cancer A Contemporary Update on Pathology Standards for Bladder Cancer: Transurethral Resection and Radical Cystectomy Specimens Donna E. Hansel a, *, Mahul B. Amin b, Eva Comperat c, Richard J. Cote d, Ruth Knüchel e, Rodolfo Montironi f, Victor E. Reuter g, Mark S. Soloway h, Saleem A. Umar d, Theodorus H. Van der Kwast i a Department of Pathology, Cleveland Clinic, Cleveland, OH, USA; b Department of Pathology, Cedars-Sinai Medical Center, Beverly Hills, CA, USA; c Department of Pathology, Hopital La Pitié-Salpetriere, UPMC Paris VI, Paris, France; d Department of Pathology, University of Miami Miller School of Medicine, Miami, FL, USA; e Department of Pathology, RWTH Aachen University, Aachen, Germany; f Institute of Pathological Anatomy, Polytechnic University of the Marche Region School of Medicine, United Hospitals, Ancona, Italy; g Department of Pathology, Memorial Sloan Kettering, New York, NY, USA; h Department of Urology, University of Miami Miller School of Medicine, Miami, FL, USA; i Department of Pathology, University Health Network, Toronto, Canada Article info Article history: Accepted October 5, 2012 Published online ahead of print on October 12, 2012 Keywords: Bladder cancer Pathology Review Standards Abstract Context: Pathology standards for the diagnosis of bladder cancer (BCa) have recently evolved to better reflect patient diagnosis and clinical outcomes. Objective: To update pathology reporting standards for BCa. Evidence acquisition: We searched the international medical literature and reviewed all articles that addressed BCa gross dissection, pathologic diagnosis, staging, and reporting as of June 6, We also reviewed the proceedings from the recent Second International Consultation on Bladder Cancer (Vienna, Austria). The literature selected for review focuses on evidence-based studies that address histopathologic factors in BCa, with emphasis placed on factors that influence patient diagnosis and clinical outcomes. Evidence synthesis: We separated data into three main components for analysis based on the type of specimen obtained: (1) transurethral resection specimens, with an emphasis on pathologic staging, variants of urothelial carcinoma, angiolymphatic invasion, and relevant ancillary techniques such as immunohistochemistry in assessing these features; (2) cystectomy specimens, with an emphasis on pt0 disease, prostatic involvement by urothelial carcinoma and lymph node dissection and analysis; and (3) cytology correlates, with recommendations for the use of cytology paired with tissue-based sampling. Areas of controversy are described and recommendations based on existing guidelines are provided. The value of a multidisciplinary team is highlighted. Conclusions: Ongoing international collaborations amongst pathologists have led to emerging standards in the reporting and microscopic diagnosis of BCa specimens. Although some areas remain controversial, we present the most up-to-date data and guidelines relevant to neoplastic pathology of the urinary bladder. # 2012 European Association of Urology. Published by Elsevier B.V. All rights reserved. * Corresponding author Euclid Avenue, Desk L25, Cleveland Clinic, Cleveland, OH 44195, USA. Tel ; Fax: address: hanseld@ccf.org (D.E. Hansel) /$ see back matter # 2012 European Association of Urology. Published by Elsevier B.V. All rights reserved.

2 322 EUROPEAN UROLOGY 63 (2013) Introduction Bladder cancer (BCa) represents the fourth most common cause of cancer in men in the United States and the seventh most common cause of cancer in men worldwide, with a much lower incidence in women that reflects an approximate 3:1 male-to-female ratio [1,2]. Although most patients are older than 60 yr of age, BCa may affect younger patients. More than 90% of BCa instances represent urothelial carcinomas, which can be subdivided by grade, stage, and subtype (conventional urothelial carcinoma vs a variant morphology) [3]. From a diagnostic pathology perspective, BCa can be challenging because of subtleties in histologic interpretation and physical limitations of the specimens obtained by endoscopic means such as biopsy and transurethral resections (TURs). A more complete evaluation of a BCa instance can be performed on a cystectomy specimen, if available, and correlated with prior TUR or biopsy specimens. A cystectomy specimen and associated lymph node dissection (LND) may present a separate challenge. A separate issue is the role of cytologic evaluation in diagnosis and follow-up. Within the past few years, several changes in reporting and microscopic diagnosis have been made. We present a current update on the interpretation of BCa specimens. 2. Evidence acquisition We searched the international medical literature and reviewed all articles that addressed BCa gross dissection, pathologic diagnosis, staging, and reporting as of June 6, We also reviewed the proceedings from the recent Second International Consultation on Bladder Cancer (Vienna, Austria) [4]. The literature selected for review focuses on evidence-based studies that address histopathologic factors in BCa, with emphasis placed on factors that influence patient diagnosis and clinical outcomes. 3. Evidence synthesis 3.1. Pathology reporting for bladder cancer: transurethral resection specimens Cold cup biopsy or TUR can be performed upon the initial finding of a bladder tumor, depending on urologist intent, with processing of the specimen commonly consisting of formalin fixation, separate submission of tumor and base when designated, and submission of the entire specimen [5]. In most instances, margin status in a fragmented TUR specimen is not possible, although use of en bloc TUR resections for appropriately sized lesions may address some concerns of margin status and potentially improve staging [6,7]. For the purposes of this review, we focus on microscopy and reporting elements associated with TUR specimens (Table 1) [8] Pathologic staging in transurethral resection specimens Pathologic staging of BCa is clinically the most powerful determinant in regard to patient prognosis and treatment Table 1 Recommended reporting elements for bladder transurethral resection specimens [6] Reporting element Procedure Histologic type Superficial lesions Histologic grade * Muscularis propria Angiolymphatic invasion Depth of invasion or stage Final pathologic stage Description Biopsy, TUR, or other descriptor Urothelial carcinoma or a variant histology with description Describes the presence of in situ lesions Grade of urothelial carcinoma, squamous cell carcinoma, or adenocarcinoma Describes whether muscularis propria is present, absent, or indeterminate Present, absent, or indeterminate Includes staging for in situ or invasive disease Includes TNM criteria TUR = transurethral resection; CAP = College of American Pathologists; BCa = bladder cancer; ISUP = International Society of Urological Pathology; WHO = World Health Organization. * Adapted from CAP Cancer Protocols. Histologic grade has been shown to be relevant for BCa outcomes, and in the United States, ISUP 1998/WHO 2004 grading is recommended and used in the CAP template in this table. However, it is recognized that the use of this grading schema in Europe is varied, with approximately half of pathologists (50.9%) using ISUP 1998/ WHO 2004 and the remainder using either WHO 1973 (43.4%) or WHO 1999 (31.4%) [5]. Optional reporting elements include tumor configuration and additional pathologic findings. decision in addition to grading [9]. Pathologic staging is based on the presence or absence of invasion and, in the case of invasive tumors, on the extent of invasion into the bladder wall, with the layers of the bladder wall and adjacent organs serving as staging landmarks. In general, progressive invasion into the bladder wall by microscopic level increases stage, including involvement of the lamina propria (pt1), superficial or deep muscularis propria (pt2a and pt2b, respectively), perivesical fat either microscopically (pt3a) or macroscopically (pt3b), and adjacent organs (pt4a) or the pelvic/abdominal wall (pt4b). However, on TUR specimens, only invasion up to the pt2 stage can be diagnosed with certainty; involvement of fibroadipose tissue on small (noncystectomy) specimens may be misleading, as fibroadipose tissue can frequently be identified in the lamina propria and may lead to an erroneous diagnosis of perivesical fat involvement [10]. In addition, pt4 diagnosis at pathology may be made only in the presence of a full resection, in which transmural invasion through the bladder wall and into the prostate can be identified with certainty using appropriate orientation. Invasion into the detrusor muscle (muscularis propria; pt2 disease) will lead to a consideration for a cystectomy depending on a variety of clinical factors such as the patient s comorbidity [11,12]. Several factors may conspire to make correct pathologic staging difficult [13 16]. For example, in one study of seven experienced pathologists, agreement on lamina propria invasion could only be obtained in 61% of cases after three assessments [15]; in another study, re-review of 164 BCa cases resulted in up- or downstaging of 20% of pt1 BCa instances [14]. A summary of potential sources of over- and understaging are listed in Table 2. The following subsections address challenges in the pathologic staging of BCa by depth

3 Table 2 Common causes of pt2 staging errors on transurethral resection EUROPEAN UROLOGY 63 (2013) [(Fig._1)TD$FIG] Staging error Overstaging of pt2 disease Understaging of pt2 disease Cause Hypertrophic muscularis mucosae Tangential sectioning Thermal artifact Desmoplastic reaction Involvement of fibroadipose tissue Thermal artifact Superficial muscularis propria in the bladder neck Absence of detrusor muscle in the TUR chip with invasion Nested and large nested variants of urothelial carcinoma TUR = transurethral resection. of invasion and includes our recommendations for minimizing these challenges pt1 disease. Invasion into the lamina propria constitutes pt1 disease and includes a range of invasive tumor volumes, from scattered individual, invasive cells to confluent tumor nests that may obscure the background architecture. Relevant to the diagnosis of pt1 disease is (1) the extent of lamina propria present in the specimen to allow identification of invasion at the base of the lesion, (2) features that aid in the identification of small numbers of invasive urothelial carcinoma cells, and (3) whether substaging the depth of lamina propria invasion is of clinical value. The first consideration involves the extent of lamina propria for assessment, as in the case of TUR specimens, in which only the exophytic portion of the tumor is sampled. Although pt1 disease may be assigned in the presence of invasion into the fibrovascular stalk of a papillary frond, in most instances, invasion may be found at the base of the lesion. Frequently, the submitted tissue from a TUR has either limited lamina propria or has cautery artifact, both of which limit the ability to determine the depth of any tumor invasion beyond the basement membrane. One option in such a case is to report the tumor as stage ptx according to the ptnm staging system 2009 to reflect an insufficient specimen, although this may be somewhat confusing when a bladder neoplasm appears to be present and a minimal stage can be assigned. A second option recommended by these authors is to provide a description that relays the limited presence of lamina propria within the sampled material. Because the presence of detrusor muscle is required for appropriate staging, it is also recommended that within the report a suggestion to undergo a repeat TUR to obtain this required piece of staging information be considered. The second consideration regards the unequivocal identification of the invasive nature of individual or small clusters of urothelial carcinoma cells, which has been variably termed microinvasive disease, early lamina propria invasion, and focally invasive (Fig. 1). Microinvasive bladder carcinoma was first defined by Farrow, who defined the invasive component as < 5 mm in depth [17]. Amin et al. Fig. 1 Microinvasion can be diagnosed even on the presence of a single cell (arrow). In this case, overlying flat carcinoma in situ gives rise to early invasive or microinvasive disease. considered the 5-mm cut-off too liberal and suggested 2 mm [18]. Another publication proposed a cut-off of not more than 20 invading cells, measured from the stroma epithelial interface [19]. Immunohistochemistry using anticytokeratin antibodies (especially pan-keratins and cytokeratin 8 and 18) may be useful in identifying rare invasive cells, especially in a background of inflammation or cautery artifact [20], although nonspecific cross-reactivity with myofibroblasts and smooth muscle cells should be excluded with light microscopy [21]. Currently, no standard exists to define microinvasive disease or to define the extent of invasive tumor volume into the lamina propria. It is appropriate to provide a descriptive analysis of microinvasive disease, clarifying that rare to few invasive tumor cells were identified, especially in instances where ancillary studies or consensus conference was used to identify these cells. The third consideration is whether substaging of pt1 disease can provide prognostic information. In some instances, this may overlap with the above discussion of microinvasive disease, as individually invasive tumor cells are generally found near the surface urothelium rather than deep within the lamina propria. Several strategies have been proposed to substratify invasive tumors cells within the lamina propria. One substaging system employs the muscularis mucosae and the associated plexus venosus in the lamina propria as reference points for distinguishing pt1a (above muscularis mucosae), pt1b (muscularis mucosae to plexus venosus), and pt1c (beyond plexus venosus) substages [22]. A condensed and perhaps more robust substaging system incorporates pt1a and pt1b cancers in the substage of minimally invasive BCa [23]. Although these studies appear promising, it has been noted that substaging based on these microscopic landmarks may pose challenges, including considerable interobserver variability and the inability to assign a substage because of the absence of the muscularis mucosae/plexus venosus

4 324 EUROPEAN UROLOGY 63 (2013) [(Fig._2)TD$FIG] and poor orientation. Finally, the use of distance measurements to the deepest portion of the invasive carcinoma may provide some value; in addition to the studies cited earlier [17,18], two independent studies used maximum diameter of invasive carcinoma in any direction <1 high-power field (about 0.5 mm) as a cut-off between focal and extensive pt1 BCa, with independent prognostic significance for progression [24,25]; one recent study of 406 cases of high-grade pt1 BCa suggested that a cut-off of a 1.0-mm maximum diameter of invasion corresponds best with a greater risk for recurrence, progression, and cancer-specific mortality [26]. The challenges associated with current pt1 substaging based on depth of lamina propria invasion have somewhat limited its use and thus has not been included in the World Health Organization (WHO) 2004 classification of BCa types. However, we recommend that a brief qualitative description of the extent of invasion, such as focal versus extensive, be included in the pathology report to provide the urologist with an estimate of the extent of disease pt2 disease. Proper pt2 staging, a key driver of patient management, involves two major components: (1) the presence of muscularis propria in the specimen and (2) determination of muscularis propria involvement by tumor. As standard reporting of the presence of muscularis propria (independent of tumor invasion) has recently been recommended for inclusion in all pathology reports, the variability in pt2 staging has been dramatically reduced. However, the presence of true invasion into the muscularis propria can be challenging, especially in the context of a florid desmoplastic reaction, hypertrophic muscularis mucosae of the lamina propria, or superficial location of the muscularis propria in the bladder neck, where invasion may spuriously be deemed as invasion of the muscularis mucosae. Several morphologic features have been described to distinguish muscularis propria from desmoplasia and muscularis mucosae, although in practical terms these are somewhat subjective. Some pathologists have employed immunohistochemical markers or special stains to attempt to define an objective measure of invasion into the muscularis propria and to distinguish muscularis propria from muscularis mucosae. Recent attention has been given to smoothelin, which was identified as a marker for contractile smooth muscle [27] and which has been suggested to be differentially expressed between the muscularis mucosae of the lamina propria and the muscularis propria [28 31].In general, smoothelin shows an intense positive immunostaining in all detrusor muscle samples but negative or weak positivity in muscularis mucosae (Fig. 2). When combined with vimentin (positive in muscularis mucosae), smoothelin could improve the sensitivity of distinguishing muscularis mucosae from muscularis propria to >80% [32]. Two subsequent studies addressed the question of whether immunohistochemistry for smoothelin would help resolve smooth muscle subtyping in problematic TUR specimens [31,33]. Whereas one publication found smoothelin to have 97% specificity in distinguishing muscularis mucosae from detrusor muscle [31], the second publication noted Fig. 2 Smoothelin immunohistochemical stains primarily demonstrate robust staining in the detrusor muscle (arrow) but only weak staining of the muscularis mucosae within the lamina propria (arrowhead). Careful titration of the antibody and controls is essential when using this newly described antibody. occasional overlap in the intensity of smoothelin staining of these two muscle layers that might lead to spurious pathologic staging results [33]. Currently, use of immunohistochemistry to aid in pt2 diagnosis remains in the research realm and is not routinely employed in pathology reporting of BCa Artifacts that influence pathologic staging. Numerous factors may lead to difficulty in pathologic interpretation of TUR specimens, including cautery (thermal) and crush artifacts that preclude a good visual assessment of the available material (Fig. 3). Tangential sections and lack of spatial orientation caused by the random embedding of bladder tissue are confounders that may lead to problems in interpretation of depth of invasion. In instances in which [(Fig._3)TD$FIG] Fig. 3 A typical example of a specimen with cautery artifact even to the untrained eye, a diagnosis of grade or stage on this specimen would be an obvious challenge.

5 Table 3 Selected urothelial carcinoma variants at risk for understaging EUROPEAN UROLOGY 63 (2013) [(Fig._4)TD$FIG] Variant Nested/large nested Microcystic Small tubules Plasmacytoid Lymphoepithelioma-like Micropapillary Sarcomatoid Staging challenge Bland morphology that mimics von Brunn nests Bland morphology that mimics cystitis cystica Bland morphology that mimics cystitis cystica Infiltrative single cells that can be easily missed Obscuring inflammation may mask carcinoma cells Aggressive cancer; often understaged in cases that lack muscularis propria Aggressive cancer with spindled morphology; may obscure background muscularis propria artifact significantly impairs pathologic assessment, it is recommended that the most accurate staging information be provided and that a comment be made in the final report regarding the limitations of the material. Urologists can assist the pathologist by reducing the coagulation current during the TUR of a bladder tumor, submitting the base of the tumor resection separately, and using a cold cup biopsy forceps if the tumor area is small to limit cautery artifact Urothelial carcinoma variants and their impact on diagnosis and staging Urothelial carcinoma may develop unusual morphologic features ( variants ) that deviate from conventional appearance; for example, a common variant is represented by urothelial carcinoma with squamous or glandular differentiation [4]. Several studies have suggested that variant differentiation may alter disease course, although evidencebased studies have been limited because of the rarity of most variants. In this section, we have selected two examples of variant morphology that may commonly lead to potential understaging in TUR specimens: micropapillary carcinoma and the nested/large nested variant of urothelial carcinoma. Additional selected variants that may be problematic in TUR staging, in part because of recognition of these entities as neoplastic, are represented in Table 3. Amin et al. described the micropapillary variant in 1994 that resembles papillary serous carcinoma of the ovary [34]. Features include infiltrating papillary clusters of tumor cells polarized to the exterior surface that lack central vascular cores and are surrounded by prominent retraction artifact that may mimic vascular invasion (Fig. 4). From a clinical perspective, >95% of these tumors are muscle invasive at the time of detection [34], and lymph node metastases occur in 27 35% of patients [35,36]. Upstaging is common, with 52.7% of patients upstaged in one study [36] and 79% in a second study [35]. As such, the presence or absence of muscularis propria (detrusor muscle) in the TUR specimen, and its involvement, is critical when diagnosing this aggressive variant. Despite distinctive clinical features associated with patients who have this diagnosis, challenges exist in its pathologic diagnosis. A recent study from Sangoi et al. showed that although the recognition of a classic form of micropapillary is quite uniform Fig. 4 The micropapillary variant of urothelial carcinoma consists of small clusters of tumor cells that lack a fibrovascular core, are polarized to the outer surface of the cluster, and are surrounded by retraction artifact (arrow) that can mimic vascular spaces. (93% agreement), the inclusion of additional cases of urothelial carcinoma with retraction artifact reduced the ability to categorize micropapillary from nonmicropapillary (k: 0.54); these findings suggest that further study of this variant is needed to define the individual pathologic features required for diagnosis [37]. There is an agreement among pathologists, however, that every micropapillary component, even minor ones, should be reported because the percentage of the micropapillary component appears inversely related to survival [38]. One paper showed that survival was inversely linked to percentage of the micropapillary variant, with <10% micropapillary features resulting in a better mean survival [35], although this remains to be validated. We recommend that any micropapillary component be reported, with an emphasis on the presence or absence of muscularis propria in the specimen. The second frequently underdiagnosed entity is the nested variant of urothelial carcinoma. This variant had been initially described by Stern in 1979 as an unusual benign bladder tumor of von Brunn nest origin [39] but was redefined in 1989 as a carcinoma of the urinary bladder with deceptively benign-appearing foci [40]. This entity can be easily under-recognized, as the superficial components of the tumor are composed of small nests without major atypia and are easily confused with von Brunn nests (Fig. 5). When the nested variant was compared to high-grade urothelial carcinoma, it more frequently displayed muscleinvasive disease on TUR (70% vs 31%, respectively), more frequent extravesical disease (83% vs 33%, respectively), and increased rates of metastatic disease (67% vs 19%, respectively) [41]. Recently, a large nested variant of urothelial carcinoma was described that maintains the bland morphologic appearance of classic nested carcinoma but has medium to large nests rather than the smaller nests associated with the classic form [42] (Fig. 6). Although only 23 patients have been described in this recent series, it appears that this variant also harbors the potential for

6 326 [(Fig._5)TD$FIG] EUROPEAN UROLOGY 63 (2013) Table 4 Recommended reporting elements for cystectomy specimens [6] Element Specimen Procedure Tumor size Histologic type Superficial lesions Histologic grade Angiolymphatic invasion Depth of invasion or stage Margin status Final pathologic stage Description Bladder or other Cystectomy subtype or anterior exenteration Greatest dimension Urothelial carcinoma or a variant histology with description Describes the presence of in situ lesions Grade of urothelial carcinoma, squamous cell carcinoma, or adenocarcinoma Present, absent, or indeterminate Includes staging for in situ or invasive disease Specifies involvement by in situ or invasive carcinoma Includes TNM criteria Fig. 5 The nested variant of urothelial carcinoma is defined by small- to medium-sized nests of bland-appearing urothelial cells. Its aggressive nature, however, becomes apparent with invasion of the detrusor muscle (asterisk). metastatic spread, as 3 out of 17 patients with follow-up developed distant metastases, and 3 patients died of BCa [42]. In cases of extensive urothelial nests and the absence of muscularis propria invasion, it is recommended that the possibility of nested urothelial carcinoma be raised in the pathology report and an additional TUR be performed to determine the extent and depth of the lesion in question prior to a clinical decision Determination of vascular invasion Several studies suggest the clinical importance of lymphovascular invasion (LVI) as a predictive marker of outcomes for BCa, including local and distant recurrence, lymph node status, and overall survival (OS) [43,44]. The diagnosis of LVI, however, is treacherous, with retraction artifact readily mimicking LVI (as already discussed for the micropapillary [(Fig._6)TD$FIG] Fig. 6 A newly described variant of invasive urothelial carcinoma includes the large nested variant, which shows bland cytology. In this case, the tumor is shown invading the detrusor muscle. variant). Larsen et al. demonstrated that only 14% of their patients diagnosed with LVI by light microscopy actually had tumor within endothelium-lined spaces when validated by immunohistochemistry [45]. In contrast, Kunju et al. demonstrated a good concordance of LVI in TUR and cystectomy samples (41% vs 65%, respectively), suggesting good reproducibility of diagnostic criteria [46]. Given the conflicting data on the subject and the challenges inherent in assessment of LVI, further prospective validation is required [47]. However, the current recommendation is to report LVI whenever identified in the specimen, preferably with the support of immunohistochemistry in challenging cases Pathology reporting for bladder cancer: cystectomy specimens A summary of recommended reporting elements for cystectomy specimens is provided in Table 4. In this section, we highlight pt0 disease, prostatic involvement by urothelial carcinoma, and handling of LNDs as areas of active discussion in pathology pt0 disease By American Joint Committee on Cancer/Union Internationale Contre le Cancer TNM guidelines, stage pt0 is a condition in which there is no evidence of primary carcinoma in the cystectomy specimen following an initial cancer diagnosis in prior biopsy or TUR specimen [48]. The incidence of stage pt0 bladder carcinoma at cystectomy has been reported to be between 5% and 20% [49 51], with 5-yr recurrence-free survival, cancer-specific survival, and OS rates of 84%, 88%, and 84%, respectively [50], with a subset of patients remaining at risk for disease recurrence or progression [51,52]. In cases of non muscle-invasive tumor, some experts consider it reasonable to offer immediate (or early ) radical cystectomy (RC) to patients who are deemed to be at high risk of progression (ie, highgrade pt1 tumors, multiple recurrent high-grade tumors, and high-grade tumors with concomitant flat urothelial carcinoma in situ [CIS]) [53,54], thus potentially increasing the frequency of pt0 cystectomies. Furthermore, the

7 increasing use of neoadjuvant therapy has led to downstaging of many patients at RC [55 60]. From a pathologist s practical standpoint, the gross sampling of cystectomy specimens that are devoid of obvious gross lesions must be performed with care. Although no current guidelines exist on the gross dissection of these specimens, it is recommended that prior surgical sites or mucosal ulcerations be sampled in their entirety for residual tumor and adequate mapping performed. Furthermore, areas that appear red and regions of mucosal induration should be sampled to rule out CIS. There is no need for complete embedding of the entire bladder wall. Communication between the urologic surgeon and pathologist is key in cases where prior surgical sites, mucosal ulcerations, or alterations are not evident to ensure that appropriate tissue sampling occurs. EUROPEAN UROLOGY 63 (2013) [(Fig._7)TD$FIG] Changes in staging for prostate involvement by urothelial carcinoma Prostatic involvement may occur via multiple routes, including direct extension from an extravesical bladder mass, direct invasion from an in situ lesion within the prostatic urethra, or colonization of the prostatic ducts followed by invasion [61]. Outcomes by prostatic invasion subtype were evaluated in one series of 142 BCa instances with prostate involvement [62]; in this study, patients were divided into those with tumor invasion through the full thickness of the bladder wall and those with tumor extension from the prostatic urethra into the prostate. Five-year OS rates were 21% and 55% for these groups, respectively, and the presence of prostatic stromal invasion was associated with a worse prognosis than for patients in whom urothelial cancer was confined to the urethral mucosa only (CIS). Another study demonstrated 5-yr survival of 7% and 46%, respectively, for patients with urothelial carcinoma showing direct extension from the bladder versus those with invasion from the prostatic urethra [63]. Based in part on these findings, criteria that define prostate involvement by urothelial carcinoma have been streamlined. Current staging guidelines recommended that true extravesical (transmural) extension of a urothelial carcinoma arising in the bladder be classified as pt4a; in contrast, urothelial carcinoma arising with the prostatic urethra and the associated prostatic ducts be classified according to urethral carcinoma guidelines into ptis pu (CIS involving the prostatic urethra), ptis pd (CIS involving the prostatic ducts), pt1 (invasion from the urethral surface to the subepithelial connective tissue), and pt2 (invasion from the urethra or ducts into the prostatic stroma) stages (Fig. 7) [48]. Furthermore, attempts have been made to predict which patients with a history of urothelial carcinoma of the bladder will have prostatic involvement using a promising algorithm that includes previous recurrence, location, and number of foci of BCa [64] Lymph node dissection and analysis The extent and bilaterality of LND in patients with BCa as well as the number of dissected lymph nodes and lymph Fig. 7 Carcinoma in situ (CIS) involving prostatic ducts (arrow), as contrasted to normal-appearing prostate (arrowhead). The CIS remains confined within the normal structure and basement membrane of the prostatic ducts and, as such, would be categorized as ptis pd for staging purposes. node density have been the subject of numerous studies [65 75]. As a result of these studies, recent changes have been made to the staging of BCa that includes both positive lymph node counts (ie, single vs multiple ) and location of metastases within defined primary and secondary drainage sites (ie, true pelvis vs common iliac lymph nodes) [48]. Furthermore, the distinction between secondary drainage sites and otherwise metastatic disease has been more clearly delineated in the pathology classification schema, and the clarification between the inability to find lymph nodes, inability to assess identified lymph nodes, and the absence of lymph node metastases has also been made [48]. Based on the new staging criteria, the identification of even small lymph node metastases in individual lymph nodes appears critical, which is supported by one study in which the identification of only one versus two positive lymph nodes significantly affected outcomes [72], highlighting the need not only to carefully evaluate lymph node packets but also to take care in dissecting the perivesical soft tissue for additional lymph nodes in this region. In light of this finding, several key areas of controversy in pathology are highlighted, including gross dissection of lymph nodes and microscopic reporting of lymph node metastases Gross dissection of lymph nodes. Several historical challenges exist in dissecting lymph nodes at gross pathology, highlighted in a recent paper [76] that surveyed 10 different pathologists on the methods of LND employed and the assessment of hematoxylin and eosin (H&E) slides via both questionnaires and glass slides. Marked variability in lymph node handling and review was identified and included (1) whether additional fat is sectioned or submitted, (2) whether one can assess the number of lymph nodes in fragmented tissue accurately, (3) where to draw the distinction between one

8 328 EUROPEAN UROLOGY 63 (2013) versus two lymph nodes when closely apposed, and (4) how small a lymphoid collection actually begets a lymph node. In addition, the amount of lymph node tissue and fat submitted by the urologist may vary significantly, contributing to differences in the number of lymph nodes identified. In this latter instance, submission of lymph nodesasdiscretepacketsmaybeoptimal,asthismay better delineate the total number of nodes contained within one region of dissection (vs a larger, fragmented specimen) and also allow for better fixation and processing of a smaller specimen [77,78]. A few recent studies have addressed these issues. One study using 61 lymphadenectomy specimens compared the number of lymph nodes identified by manual and visual inspection following Carnoy s fixation to the additional number identified in residual fat tissue in which no nodes were identified [79]. In total, 61% of lymph nodes were identified by manual inspection compared to the remaining 39% of lymph nodes that were identified only after submission of the remaining fat. However, out of the 153 additional lymph nodes identified in the residual tissue, none harbored metastatic deposits, suggesting that full specimen submission may not yield any additional clinical information when assessing small lymph nodes not otherwise detectable. In general pathology practice, the handling of LNDs is generally performed via careful palpation and dissection of individual lymph nodes, with sectioning of larger lymph nodes (>4 mm). When processing is complete, review of H&E-stained sections is performed, which appears to yield reliable results in BCa patients on representative section without the requirement of additional deeper recuts [80], which is in line with current practice at most institutions Features of metastatic urothelial carcinoma deposits on microscopic report. Another area of active histopathologic research in BCa lymph node metastases involves assessment of the metastatic deposit, including the size of the metastatic deposit [67 69] and the presence of extranodal extension (ie, metastatic tumor deposit spread outside of the confines of the lymph node capsule) [66,68,81]. Currently, no consensus exists for which metastatic features affect outcome; however, it is recommended that gross and microscopic features be described as accurately as possible in the final pathology report to provide information to the treating physician regarding the extent of disease Cytology associated with paired tissue-based sampling Diagnostic tools that involve noninvasive or minimally invasive methods are appealing to patients, although these techniques require continual reassessment to guarantee appropriate patient care. Urine cytology for the detection of BCa cells has been considered a valid diagnostic method because its specificity and sensitivity are high when highgrade tumor cells are sought [82]. As such, high-quality specimen preparation of urine samples and bladder washings is required and, if performed properly, often renders other supportive techniques unnecessary [83]. Strong diagnostic training for the uropathologist and the cytologist, together with a supply of clinical information, are further requirements [84]. Cytology is especially relevant in a number of clinical settings, and recommendations for use include the following: (1) In patients with a positive urine cytology for tumor cells, TUR possibly with fluorescence cystoscopy should be performed; (2) the combined value of a bladder washing with a bladder biopsy is clinically useful, because a high-grade, positive bladder washing in parallel to a cystoscopically visible tumor, supports a high-grade diagnosis [85]; (3) a positive urine cytology paired with a negative biopsy specimen may suggest a small, flat, highgrade lesion (CIS), and a repeat assessment or evaluation of the upper tract should be performed, preferably with a bladder washing [86,87]; (4) a diagnosis of positive cytology from upper tract urine or washings of the ureter or renal pelvis may be an indicator of significant disease andshouldbefollowedupwithrepeatassessment,rereview of the specimen, and importantly ureteroscopy in the absence of radiographic changes consistent with a urothelial tumor. The urologist cannot reliably separate cells from the upper and lower urinary tract when a catheter is passed through the bladder into the ureter or renal collecting system. Tumor cells obtained from urine or washing may be from the bladder or prostate or from the upper tract [88]. Following intravesical therapy, urine and bladder washing cytology can be used to monitor for recurrence. The presence of atypical or suspicious findings is not uncommon, and fluorescence in situ hybridization studies may be considered to determine aneuploidy [54]. Although these four settings are considered the highest priority for cytologic diagnosis in urothelial cancer, we agree that improved detection of low-grade tumors and the ability to predict their biological behavior are also needed. Experimental and clinical history as well as current studies seem to clearly indicate that a multiparametric approach to urine or bladder washing specimens has to be defined to reach adequate sensitivity and specificity [89,90] Communication to enhance the care of bladder cancer patients Pathology functions not in a vacuum but as a dynamic specialty that requires knowledge of patient history, clinical findings, and dialogue with clinical colleagues [91]. Cooperative efforts to provide better patient care are important and, perhaps more than most other disease sites, are especially relevant in BCa, where cystoscopic findings and patient history are critically relevant in the diagnosis of any bladder lesion. As such, it is important for pathologists to receive relevant patient information from the treating clinician and to be comfortable enough to open a dialogue in any patient case that presents a challenge. Likewise, the ever-evolving pathology literature related to

9 EUROPEAN UROLOGY 63 (2013) BCa diagnosis requires frequent communication to keep the clinician aware of changes in this arena, and it is important for the pathologist to recognize his or her role in this important aspect of communication. Finally, the ability to share areas of needed improvement in BCa diagnosis as well as ideas on new research concepts and treatment modalities is vital to advancing a field that urgently requires new knowledge. As such, a team-based approach to BCa that involves not only the pathologist and urologist but also the urologic oncologist, radiation oncologist, primary care physician, and researchers is vital. Likewise, several recommendations have been made to enhance the quality of the specimen received by the diagnosing pathologist [92]. These include (1) familiarity with the past urologic history of the patient, (2) limiting trauma to the lower urinary tract when taking the initial biopsy, (3) removal of small papillary bladder tumors by cold cup biopsy forceps to avoid cautery artifact, (4) removal of larger papillary and all sessile or flat tumors with a resectoscope at a low coagulation current setting to minimize cautery artifact, (5) efforts to obtain muscularis propria (detrusor muscle) when high-grade carcinoma is suspected,and(6)performingaturoftheprostateif prostatic involvement is suspected. We encourage the urologist to sit down and review the histology with the pathologist whenever feasible and particularly when the pathologic grade or stage will lead to a critical treatment decision. In the future, this material will be readily available alongside the written report, enhancing the clinician s familiarity with the vagaries of urothelial pathology. 4. Conclusions Encouraging progress has been made in defining the standards of TUR and cystectomy specimen handling and diagnosis during the past several decades, although we are far from optimal care for these patients. Ongoing reevaluation of how we handle pathologic specimens is critical not only for patient diagnosis and guiding treatment but for providing accurate material for ongoing research and biological understanding of the disease. Importantly, the role of ongoing communication among the various specialties that manage patients with this disease is a crucial component and should be optimized whenever possible. Author contributions: Donna E. Hansel had full access to all the data in the study and takes responsibility for the integrity of the data and the accuracy of the data analysis. Study concept and design: None. Acquisition of data: None. Analysis and interpretation of data: Cote, Montironi, Amin. Drafting of the manuscript: Comperat, Umar, van der Kwast, Hansel, Knuchel, Soloway. Critical revision of the manuscript for important intellectual content: Reuter, Hansel. Statistical analysis: None. Obtaining funding: None. Administrative, technical, or material support: None. Supervision: None. Other (specify): None. Financial disclosures: Donna E. Hansel certifies that all conflicts of interest, including specific financial interests and relationships and affiliations relevant to the subject matter or materials discussed in the manuscript (eg, employment/affiliation, grants or funding, consultancies, honoraria, stock ownership or options, expert testimony, royalties, or patents filed, received, or pending), are the following: None. Funding/Support and role of the sponsor: This publication was made possible by the Clinical and Translational Science Collaborative of Cleveland, UL1TR from the National Center for Advancing Translational Sciences (NCATS) component of the National Institutes of Health and NIH roadmap for Medical Research. Its contents are solely the responsibility of the authors and do not necessarily represent the official views of the NIH. Acknowledgment statement: The authors would like to thank Dr. James Catto for his insightful recommendations in the preparation of this manuscript. References [1] American Cancer Society. Cancer facts and figures Atlanta, GA: American Cancer Society; [2] Jemal A, Bray F, Center MM, Ferlay J, Ward E, Forman D. Global cancer statistics. CA Cancer J Clin 2011;61: [3] Eble JN, Sauter G, Epstein JI, Sesterhenn IA, editors. Pathology and genetics of tumours of the urinary system and male genital organs. Lyon, France: IARC Press; [4] Soloway M, Khoury S, editors. 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