Pathological criteria and practical issues in papillary lesions of the breast a review

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1 Histopathology 2016, 68, DOI: /his REVIEW Pathological criteria and practical issues in papillary lesions of the breast a review Yun-Bi Ni & Gary M Tse Department of Anatomical and Cellular Pathology, Prince of Wales Hospital, The Chinese University of Hong Kong, Hong Kong Ni Y-B & Tse G M (2016) Histopathology 68, DOI: /his Pathological criteria and practical issues in papillary lesions of the breast a review Papillary lesions of the breast include a broad spectrum of lesions, ranging from benign papilloma, papilloma with atypical ductal hyperplasia (ADH) or ductal carcinoma in situ (DCIS) to papillary carcinoma. The accurate diagnosis of mammary papillary lesions is a challenge for pathologists, owing to the overlapping features among these lesions. In this review, some of the diagnostic criteria of papillary lesions are discussed, with special emphasis on some Keywords: biopsy, breast, papillary lesions key morphological features, namely fibrovascular cores, epithelial proliferation in a solid pattern, intraductal papilloma complicated by ADH or DCIS, and invasion and its mimics. The roles of immunohistochemistry, and the interpretation of myoepithelial cell markers, hormone receptors, and high molecular weight cytokeratin, are addressed. Finally, novel biomarkers and genetic aberrations in papillary lesions are summarized. Introduction Papillary lesions of the breast can present with nipple discharge when centrally located, or as palpable masses when they are located near the surface or are relatively large. Mammography, ultrasound and magnetic resonance imaging may reveal the deeper lesions. Sometimes the smaller lesions can also be identified incidentally in breast biopsies obtained for other reasons. Papillary lesions include a broad spectrum of lesions, ranging from benign papilloma, papilloma with atypical ductal hyperplasia (ADH) or ductal carcinoma in situ (DCIS) to papillary carcinoma. In a large cohort of benign breast biopsies, papillomas account for 5.3% of the cases. 1 In the Netherlands Cancer Registry, papillary carcinomas are seen in 0.7% of the malignant breast tumours. 2 Address for correspondence: G M Tse, Department of Anatomical and Cellular Pathology, Prince of Wales Hospital, Ngan Shing Street, Shatin, NT, Hong Kong. garytse@cuhk.edu.hk Morphologically, papillary lesions of the breast are characterized by the presence of arborizing fibrovascular cores derived from the wall of the ducts, which are often distended by the papillary lesions. These fibrovascular cores are, in turn, lined by layers of epithelial cells, with or without a complete layer of intervening myoepithelial cells. The accurate diagnosis of papillary lesions continues to be a challenge for pathologists. The underlying causes of this diagnostic difficulty are multiple. Different papillary lesions may possess overlapping morphological and immunohistochemical (IHC) features; there are limitations regarding the usual diagnostic samples [core needle biopsy (CNB) or fine needle aspiration cytology]; and there is a lack of consensus on the subsequent management strategy based on CNB diagnosis. Studies have been performed to determine whether specific biomarkers could improve the accuracy of pathological diagnosis. The pathogenesis of papillary lesions is enigmatic, with only a few studies having characterized their molecular aberrations. This rather scanty information is still helpful in 2015 John Wiley & Sons Ltd.

2 Papillary lesion of the breast 23 understanding their pathogenesis and biological behaviour, and may shed light on the differential diagnosis. In this review, the diagnostic criteria of papillary lesions are discussed, with emphasis on some practical diagnostic issues, including differentiating different papillary lesions on the basis of histological and IHC features. Management options based on CNB diagnosis will also be evaluated. Finally, new adjunct biomarkers and the genetic aberrations of breast papillary lesions are also summarized. Pathological classification of papillary lesions of the breast In the World Health Organization (WHO) classification of breast tumours, papillary lesions are classified as intraductal papilloma, intraductal papillary carcinoma, encapsulated papillary carcinoma, and solid papillary carcinoma. Invasive papillary carcinoma is now grouped under rare tumours, but not under papillary lesions. 3 7 INTRADUCTAL PAPILLOMA Intraductal papillomas can be divided into central (solitary) and peripheral (multiple), which share similar structures of arborescent fibrovascular cores covered by myoepithelial cells and epithelial cells; the former may be attenuated and become inconspicuous. The periphery of the lesion also shows a layer of myoepithelial cells (Figure 1). In benign papillomas, the epithelial cells may show usual ductal hyperplasia (UDH) or squamous or apocrine metaplasia. Nevertheless, epithelial cell mitoses are absent or extremely rare. Other histological features that may be superimposed on intraductal papillomas include haemorrhage, infarction, stromal fibrosis (in extreme examples, the papillary architecture may be obscured; this is termed sclerosed papillomas or ductal adenomas), and mucinous, clear cell and sebaceous metaplasia. 8,9 Intraductal papillomas may also be complicated by ADH or DCIS. These are characterized by focal abnormal epithelial proliferation with cytological and architectural features of low-grade ductal neoplasia. In these areas, myoepithelial cells may be scanty or absent. The accurate separation between these two diagnoses remains a contentious issue. Some authors have proposed using a percentage criterion (30% or 90%), 10,11 whereas others have proposed an extent criterion ( 3 mm), 12 for a diagnosis of papilloma with DCIS to be made. Although the extent criterion (ADH, atypical focus of <3 mm; DCIS, atypical focus of 3 mm) has been recommended by the WHO working group, it is also acknowledged to be a pragmatic guideline lacking scientific evidence. 3,13,14 When the abnormal epithelial proliferation shows intermediate or high nuclear grade features, DCIS should be diagnosed regardless of the extent containing atypical cells, and the 3-mm rule does not apply. 15 Intraductal papillomas without atypia but with surrounding proliferative changes incur a two-fold relative risk for subsequent carcinoma as compared with the general population, and this is similar to other forms of proliferative breast disease without atypia. The risk is higher in multiple papillomas than in solitary papilloma. 1,16 The relative risk of developing breast cancer in patients with papillomas with atypia is 5- to 7.5-fold greater than in patients with papillomas without atypia. 1,12,16 INTRADUCTAL PAPILLARY CARCINOMA Figure 1. Intraductal papilloma. The inset highlights fibrovascular cores covered by a layer of myoepithelial cells with overlying epithelial cells within a duct, and the peripheral myoepithelial cell layer. In intraductal papillary carcinomas, the ducts and the terminal ductal lobular units are filled with slender, branching fibrovascular stalks, covered by a single population of neoplastic cells. The neoplastic cells are usually columnar, with low-grade to intermediate-grade nuclei (Figure 2). They are arranged into one to several layers, and may form different geometric patterns, including micropapillary, cribriform or solid. There are absent or scanty myoepithelial cells separating the papillae and the epithelial proliferation.

3 24 Y-B Ni & G M Tse A myoepithelial cell layer is usually identified at the periphery of the lesions. Sometimes, these lesions may show a dimorphic cell population, with a second population of clear epithelial cells that may sometimes be mistaken for myoepithelial cells. 17 Intraductal papillary carcinomas are usually multifocal and peripheral in distribution. ENCAPSULATED PAPILLARY CARCINOMA Figure 2. Intraductal papillary carcinoma. The inset highlights fibrovascular cores covered by neoplastic cells with low-grade nuclei without intervening myoepithelial cells. Encapsulated papillary carcinomas possess a thick fibrous capsule, and, within the capsule, there is proliferation of delicate fibrovascular stalks covered by monotonous neoplastic cells, in mostly solid to cribriform patterns. Typically, these lesions lack myoepithelial cells both around the fibrovascular cores and at the periphery (Figure 3). 18,19 The exact nature of encapsulated papillary carcinomas is not known. The lack of myoepithelial cells has led some authors to propose an invasive nature, 18,19 but others have suggested encapsulated papillary carcinomas to be in-situ lesions, by demonstrating the presence of intact basement membrane with collagen type IV staining. 20 Genetic studies have also shown variable results, with some showing similar genetic changes in encapsulated papillary carcinomas and invasive carcinomas, 21 and others showing that encapsulated papillary carcinomas were closer to carcinomas in situ. 22 Thus, although the true genetic nature of encapsulated papillary carcinomas remains unsettled, they may be considered either as indolent invasive carcinomas (because of the lack of an outer myoepithelial cell layer) or carcinomas in transition from in situ to invasive. A minority of encapsulated papillary carcinomas may be associated with an invasive component. The invasive component is characterized by tumour cells showing an infiltrative pattern extending beyond the fibrous capsule of the lesion and inducing a stromal reaction. This invasive component may be papillary, or may adopt the pattern of infiltrating duct carcinoma, no special type. Staging of encapsulated papillary carcinomas has been controversial. If there is component of conventional invasive carcinoma, the stage should follow only the invasive component. In the absence of these invasive foci, the staging is suggested to be Tis. 5 The goal is to prevent overtreatment of such lesions. 23,24 Although most encapsulated papillary carcinomas are of low and intermediate nuclear grade, a distinct proportion may show high-grade cytonuclear features. This group of encapsulated papillary carcinomas are more likely to be oestrogen receptor (ER)-negative and of larger size, and are more frequently associated with stromal invasion. 25 It is suggested that this group should be managed in a similar fashion to conventional forms of invasive breast carcinoma, based on established clinicopathological parameters. 25 A B Figure 3. A, Encapsulated papillary carcinoma. B, p63 highlights the lack of myoepithelial cells both around the fibrovascular cores and at the periphery. The inset shows the positive myoepithelial staining in the adjacent normal breast tissue as a control.

4 Papillary lesion of the breast 25 SOLID PAPILLARY CARCINOMA Solid papillary carcinomas are composed of multiple nodular masses arising from duct-like structures, and form a geographical pattern. Although the neoplastic cell proliferation is usually cellular, and the papillary structure is not prominent, an underlying fibrovascular stromal network can still be observed. The neoplastic cells are oval or spindle-shaped, with low-grade to intermediate-grade nuclei. Occasionally, these epithelial cells may show streaming (and hence be confused with florid epithelial hyperplasia) or accumulation of extracellular mucin. Very often, they also show IHC expression of neuroendocrine markers. Frequently, these tumour cell nests lack surrounding myoepithelial cells (Figure 4), 26 and they may be diagnosed as intraductal papillary carcinoma (with florid epithelial hyperplasia), particularly if a peripheral myoepithelial cell layer can be demonstrated. The categorization of solid papillary carcinomas is still controversial. In some cases, an obvious invasive component may be seen, and these tend to show mucinous or neuroendocrine differentiation, although other histological subtypes have also been described For staging purposes, it is recommended that solid papillary carcinoma without definite invasion be staged as Tis 6 ; and when invasive foci are present, they should be staged on the basis of the invasive foci only. INVASIVE PAPILLARY CARCINOMA Primary invasive papillary carcinomas are extremely rare, 30 and their diagnosis should follow stringent criteria of invasive carcinomas harbouring a papillary architecture with papillae formed by malignant cells closely related to fibrovascular cores, and possessing a permeative front. 7 An in-situ component is rare, and, when present extensively, should prompt a diagnosis of papillary carcinoma (in situ) with focal invasion. In some of the previous studies reporting on clinical outcomes of invasive papillary carcinomas, the diagnostic criterion was not clear, so little is known of the clinical behaviour of invasive papillary carcinoma when a stringent diagnostic criterion is used. Survival data from the Netherlands Cancer Registry have demonstrated the absence of a survival difference between in-situ and invasive papillary lesions. 2 It should be noted that metastatic papillary carcinomas, including those from the ovary, thyroid, and lung, need to be fully ruled out when a diagnosis of primary invasive papillary carcinoma of the breast is made. Second, in terms of terminology, invasive papillary carcinoma should also be distinguished from invasive carcinoma arising in conjunction with encapsulated and solid papillary carcinomas. In the latter setting, the term papillary carcinoma (in situ) with invasion should be used. Third, invasive papillary carcinoma should not be confused with invasive micropapillary carcinoma, which is an aggressive form of invasive breast carcinoma that is more likely to be associated with lymphovascular invasion, axillary lymph node metastases, 31,32 higher cyclin D1 expression, a higher proliferation rate and MYC (8q24) amplification than invasive carcinoma of no special type. 33 Morphologically, invasive micropapillary carcinomas are devoid of true fibrovascular cores, and are characterized by the presence of hollow or morula-like clusters of neoplastic cells surrounded by clear spaces. The apical portion of the cells faces the stroma around the tumour clusters instead of facing the central lumen of the acinar structure (inside-out pattern). Electron microscopy studies 34 and mucin studies 35 support the concept of reverse orientation of tumour cells in this lesion. Practical issues with papillary lesions of the breast In the characterization and differentiation of papillary lesions, there are many practical issues that are A B Figure 4. A, Solid papillary carcinoma. B, Synaptophysin highlights the neuroendocrine differentiation.

5 26 Y-B Ni & G M Tse problematic for the practising pathologists in routine handling and diagnosis. These problems mainly arise in several areas, as follows: (i) overlapping morphological features among benign, atypical and malignant papillary lesions; (ii) overlapping IHC staining patterns among benign, atypical and malignant papillary lesions; and (iii) further management of breast papillary lesions based on diagnosis in CNBs. OVERLAPPING MORPHOLOGICAL FEATURES Fibrovascular cores The cytological and architectural patterns of benign and malignant papillary lesions show significant overlap. Although papillomas have been described as possessing broad, pinkish and fibrotic fibrovascular cores as compared with papillary carcinomas, 24,36 it was recently reported that these broad fibrovascular cores may also be seen in malignant (and atypical) papillary lesions (Figure 5). 37 This highlighted the fact that the size of fibrovascular cores may not be an infallible criterion for differentiation of papillary lesions, especially between intraductal papillomas and intraductal papillary carcinomas. Epithelial proliferation with spindled nuclei and nuclear streaming in a solid configuration Papillary lesions are commonly complicated by epithelial proliferation. Problems in diagnosis arise when the proliferative epithelial cells show spindled nuclei and nuclear streaming in a solid configuration. At the benign end of the spectrum, florid epithelial hyperplasia tends to show spindled nuclei with streaming, whereas the less common lesion showing Figure 5. Broad fibrovascular cores in solid papillary carcinoma. an alarmingly similar morphology is solid papillary carcinoma with neuroendocrine differentiation (Figure 6). The latter lesion characteristically occurs in older women, and tends to be slowly growing. 28 The haematoxylin and eosin morphologies of these entities can sometimes be totally indistinguishable, and it may be necessary to resort to immunohistochemistry (discussed below) to achieve reliable differentiation. Intraductal papillomas complicated by ADH or DCIS Intraductal papillomas may be complicated by superimposed ADH or DCIS, with the latter diagnosis having more sinister implications. Nevertheless, concrete evidence of the difference in outcome has not been well reported. The WHO panel recommended use of the quantitative criterion of 3 mm. 3 This differentiation based on a quantitative difference creates a potential problem in assigning diagnostic labels and in communication with surgeons and patients, as a biopsy from a papilloma with DCIS may only show a <3-mm focus of atypical epithelial hyperplasia, resulting in a biopsy diagnosis of papilloma with ADH. To the unwary, this may be seen erroneously as an undercall/false negative on the part of the pathologist. Invasion and its mimics True/frank invasion in a papillary lesion needs to be differentiated from mimics, including benign glandular entrapment and mechanical dislodgement during a previous biopsy. 38,39 Stromal fibrosis is commonly seen in papillomas, and epithelial components may become entrapped in the fibrotic area and may mimic invasive carcinoma. However, a genuine diagnosis of malignancy should not be based only on an apparently infiltrative pattern. Instead, the diagnosis of carcinoma must rest on the cytological and architectural characteristics of the papillary tumour itself. Only after having determined that a papillary tumour shows features of malignancy should one consider the question of whether the irregular epithelial clusters in the adjacent fibrotic tissue reflect entrapment or invasion. Several features are helpful, as follows (Table 1) (Figure 7): (i) in benign glandular entrapment, the benign epithelial cells are arranged in irregular but smoothly contoured clusters, and tend to flow in the direction of the fibroblasts and bundles of collagen, whereas the invasive foci are seen traversing the normal stroma in different directions; (ii) the benign entrapped glandular component typically shows a polygonal shape that is compressed into long and slender cords, whereas the invasive foci retain an open and sometimes angulated glandular

6 Papillary lesion of the breast 27 A B Figure 6. Epithelial proliferation with spindled nuclei and nuclear streaming. A, Florid hyperplasia in intraductal papilloma. B, Solid papillary carcinoma. Table 1. Features that are helpful in differentiating benign glandular entrapment and invasive foci Features Arrangement of epithelial cells Architectures of epithelial cell clusters Myoepithelial cells Benign glandular entrapment Irregular but smoothly contoured clusters, flow in the direction of the fibroblasts and bundles of collagen Polygonal shape compressed into long and slender cords Present Invasive foci Traversing the normal stroma in different directions Open and sometimes angulated lumen Absent lumen; and (iii) myoepithelial cells are always retained, albeit sometimes attenuated in the entrapped benign glands, and these can be highlighted by IHC staining, whereas the invasive foci are devoid of myoepithelial cells. Mechanical displacement of the epithelium is particularly prone to occur in encapsulated papillary carcinoma when the capsule is punctured by the core biopsy needle, as friable tumour fragments escape together with cyst fluid. 40 Its differentiation from true/frank invasion is best determined by histological examination, as IHC staining for myoepithelial cells will not be helpful in this setting, 38,41 because both differentials are negative for mypepithelial cells. The possible clues are the associated haemorrhage, haemosiderin-laden macrophages, fat necrosis, inflammation, granulation tissue, and possible needle tract reaction. However, one should also keep in mind the possibility of stromal invasion in an area that was adjacent, by chance, to the needle tract. OVERLAPPING IHC FEATURES Figure 7. Benign glandular entrapment in intraductal papilloma. The epithelial cells are arranged in irregular but smoothly contoured clusters, and are compressed by the collagenized stroma. They tend to flow in the direction of bundles of collagen. The inset shows that p63 highlights myoepithelial cells surrounding the entrapped epithelial cells. The use of IHC staining in the evaluation of papillary lesions has become routine. The most frequently used groups of markers include p63, high molecular weight cytokeratins (HMWCKs), and hormone receptors, mostly ER. The IHC phenotypes of papillary lesions are summarized by the WHO panel (Table 2). 4 It is generally accepted that, in benign papillary lesions, p63 and HMWCKs are positive, and ER shows heterogeneous staining. For malignant lesions, p63 and HMWCKs are negative, and ER shows homogeneous staining. There are, however, many exceptions to these rules, and sometimes these rules can be difficult to apply. For hormone receptors such as ER, homogeneous or heterogeneous staining has not been well defined, and thus can be difficult to differentiate; as a result, this criterion alone may not be very reliable. In a study focusing on the IHC panel of cases with inconsistent diagnosis on CNB and excisional specimens, ER homogeneous staining was arbitrarily defined as uniform strong nuclear reactivity of >60% of the

7 28 Y-B Ni & G M Tse Table 2. Immunohistochemical features of papillary lesions of the breast (adapted from World Health Organization ) p63 Papillary fronds Periphery of lesion HMWCKs Hormonal receptors Intraductal papilloma Positive Positive Positive: Myoepithelial cells UDH (heterogeneous positivity) Negative: Apocrine metaplasia Positive (patchy): Luminal cells UDH (heterogeneous positivity) Negative: Apocrine metaplasia Papilloma with ADH or DICS Positive in papilloma; may be scant in the ADH/DCIS component Positive Positive: Myoepithelial cells UDH (heterogeneous positivity) Negative: Apocrine metaplasia ADH/DCIS Positive (patchy): Luminal cells UDH (heterogeneous positivity) Negative: Apocrine metaplasia Positive, strong and diffuse: ADH/DCIS Intraductal papillary carcinoma Encapsulated papillary carcinoma Negative Positive Negative: Neoplastic cell population Negative Usually negative Negative: Neoplastic cell population Positive, strong and diffuse: Neoplastic cell population Positive, strong and diffuse: Neoplastic cell population Solid papillary carcinoma Solid-papillary areas are negative May be negative or positive Negative Positive, strong and diffuse ADH, Atypical ductal hyperplasia; DCIS, Ductal carcinoma in situ; HMWCK, High molecular weight cytokeratin; UDH, Usual ductal hyperplasia. epithelial cells, and 30% (3/10) of papillary carcinomas were underdiagnosed as benign lesions, whereas 67% (2/3) of benign papillary lesions were overdiagnosed as malignant in CNB specimens. 42 Strong and diffuse ER staining has also been noted in 5% (1/20) of benign papillary lesions in another cohort. 43 In our routine diagnostic work, homogeneous ER staining has also been encountered in papillomas, notably when there is a columnar cell-type change seen in the lining epithelium (Figure 8). p63 is usually used as a myoepithelial marker. Unlike other myoepithelial markers, p63 shows nuclear staining and minimal cross-reactivity with stromal cells or myofibroblastas, and is thus considered to be a superior myoepithelial marker for diagnosis. 44 When applying a myoepithelial marker for characterization of papillary lesions, one has to be meticulous about the location of the myoepithelial cells. In general, myoepithelial cells in papillary lesions can be considered to reside in two different compartments, either within the lesion around the fibrovascular cores, or around the lesion at the duct wall. It is by assessing myoepithelial cells around the fibrovascular cores inside the lesion that one can differentiate between benign and malignant papillary Figure 8. Homogeneous staining for oestrogen receptor (inset) in intraductal papilloma.

8 Papillary lesion of the breast 29 lesions. In general, papillomas have a complete, readily demonstrable myoepithelial layer around fibrovascular cores, whereas papillary carcinomas tend to lose this layer. In contrast, whereas benign papillary lesions usually show a surrounding myoepithelial cell layer at the duct wall, malignant papillary lesions may or may not show an outer myoepithelial cell layer and the absence of an outer myoepithelial cell layer does not automatically imply invasion. Lesions that are categorized as in-situ carcinoma, including encapsulated papillary carcinomas and some solid papillary carcinomas, can be devoid of an outer myoepithelial lining (Figure 3). 5,6 High molecular weight cytokeratins play an important role in the evaluation of the nature of any of the solid epithelial proliferations that frequently complicate papillary lesions. 45 In general, benign epithelial proliferations (UDH and florid hyperplasia) express HMWCKs, whereas atypical epithelial proliferations (ADH and DCIS) do not. This is a well-documented practice, and the only potential problem is that HMWCKs should only be evaluated in the solid areas, and not in the papillary areas. Benign luminal cells lining the papillary fibrovascular cores do not always express HMWCKs, so a casual observation may result in an erroneous diagnosis of atypical/malignant papillary lesion in a benign papilloma with only scanty luminal epithelium dotting the fibrovascular cores and not expressing HMWCKs. Some authors have also suggested the use of combination of biomarkers, as this may enhance the sensitivity and specificity in making a specific diagnosis: the combination of cytokeratin (CK) 5/6 and ER has been recommended for identifying atypical papillary lesions 43 ; another study has recommended CK5/6, p63, and neuroendocrine markers 46 ; and others have suggested ER and MUC3 in papillary DCIS, but CK5/6 and p63 in intraductal papillomas. 47 A study used CK8/18, a luminal cytokeratin, as a component of a CK5/CK8/18/p63 cocktail, and this resulted in 100% sensitivity 48 ; another study showed that a combination of HMWCKs, namely CK5/6, CK14, and 34bE12, gave the best overall specificity and sensitivity for identifying malignant papillary lesions based only on their staining of different morphological patterns, without identifying cell types. 49 Thus, the use of more than one marker is recommended. FURTHER MANAGEMENT OF BREAST PAPILLARY LESIONS BASED ON DIAGNOSIS IN CNB Core needle biopsy has gained wide acceptance in the diagnosis of breast papillary lesions. The subsequent management of CNB-diagnosed atypical or malignant papillary lesions is well established. Malignant papillary lesions need to be surgically excised. Atypical papillary lesions also need to be excised to determine whether a more significant lesion is present. However, the management of CNB-diagnosed benign papillary lesions is more controversial. A meta-analysis rescreening studies conducted between January 1985 and March 2012 demonstrated a substantial risk of upgrading after surgical excision for CNB-diagnosed non-malignant breast papillary lesions, with a pooled underestimation rate of 15.7%. 50 Subsequent studies reported a range of 3 33% of CNB-diagnosed benign papillary lesions being upgraded to atypical or malignant categories in surgical excision or imaging follow-up A topographical and histopathological study found that ~25% of DCISs associated with papilloma had a potential risk of sampling error in small samples of CNBs, owing to an eccentric distribution or a low proportion of DCIS within the papilloma, whereas radiology showed segmental abnormalities in 83% of these cases. 63 Thus, for this group of CNB-diagnosed non-malignant papillary lesions, some authors have recommended surgical excision, whereas others have suggested close imaging follow-up, rather than invasive surgical procedures. Others have recommended surgical excision when segmental abnormalities are seen radiologically, because this might suggest coexisting DCIS. 63 Larger tissue samples significantly improved the predictive value of benign histology on CNB. 61 Papillomas sampled with a 12-gauge or larger needle, seven or more cores, or >96 mm may retain their benign features upon excision. 56 The new adjunct biomarkers and genetic aberrations in differentiating benign and malignant papillary lesions Owing to the diagnostic challenge posed by papillary lesions, recent studies have evaluated new biomarkers to differentiate benign papillary lesions from atypical or malignant ones. Cell cycle markers, including cyclin B1 and cyclin D1, are independently associated with malignancy in papillary lesions. Cyclin B1 was found to be useful for identifying malignant papillary lesions (sensitivity, 80%; specificity, 72.7%), whereas cyclin D1 showed lower specificity (sensitivity, 86.4%; specificity, 32.6%). 64 Cancer stem cell markers have also been investigated. In an early small cohort, CD44 expression in papillomas was significantly

9 30 Y-B Ni & G M Tse higher than that in papillary carcinomas. 65 Subsequently, in a larger cohort of 160 cases, the absence of CD44 expression was useful for identifying malignant papillary lesions (sensitivity, 49.1%; specificity, 90%). 66 Recently, the absence of CD133 expression was also found to be useful for identifying malignant papillary lesions (sensitivity, 96.8%; specificity, 91.4%). 67 The pathogenesis of breast papillary lesions is still enigmatic. Relatively few studies have characterized molecular or cell-biological aberrations in papillary lesions. However, the genetic alterations that have been identified may be helpful in understanding the pathogenesis and in differential diagnosis. Alterations in chromosomes 3, 7, 17 and X have been detected in papillary carcinoma but not in papilloma. 68 Chromosome 16 mutations have been reported in the early steps of breast papillary tumorigenesis. A high frequency of loss of heterozygosity (LOH) at chromosome 16p13 and 16q21 was detected both in intraductal papillomas and papillary carcinomas, whereas LOH at locus 16q23 was limited to malignant lesions. LOH at the TP53 locus is also significantly associated with the malignant phenotype Additionally, a higher PIK3CA or AKT mutation frequency was identified in papillomas than in papillary carcinomas, indicating that papillomas could be driven by mutations in the PIK3CA AKT pathway, whereas papillary carcinomas might develop along a molecular pathway that is somewhat divergent from that of most benign papillary proliferations. 72 Gene profiling on papillary carcinomas demonstrated that most of them (93.8%, 15/16) could be classified as the luminal subtype, with the few remaining cases being of the basal-like subtype. On comparison with grade-matched and ER-matched invasive carcinomas of no special type, papillary carcinomas show similar patterns of DNA copy number alterations, but different transcriptomic profiles, with down-regulation of proliferation-related, cell assembly and organization, cellular movement and migration genes, and overexpression of genes related to homeostasis and angiogenesis, suggesting a less invasive phenotype. 73 Regarding the distinct papillary histological subtypes, encapsulated papillary carcinoma, solid papillary carcinoma and invasive papillary carcinoma showed similar array-based comparative genomic hybridization profiles, 21 suggesting that they may be histological variants of the same entity. However, differences in the transcriptomic profiles related to cell migration have been found among them, and may account for their different histological features. 73 Thus, at present, our knowledge about the genetic changes of papillary lesions is still limited, and the exact molecular pathogenesis and whether the current histology-based classification has any molecular basis remain to be determined. Conclusions Papillary lesions include a broad spectrum of lesions, ranging from benign to malignant. Their differential diagnoses remain difficult. Although the histology of these lesions is now better characterized, there are still many overlapping features, such as broad fibrovascular cores, epithelial proliferation with spindled nuclei and nuclear streaming in a solid configuration, intraductal papillomas complicated by ADH or DCIS, and invasion and its mimics. The use of immunohistochemistry is extremely useful, but one should be meticulous in the interpretation, especially over the issues of heterogeneous or homogeneous staining of ER, the location of the myoepithelial cells, and the interpretation of HMWCK expression within the lesions. Some novel biomarkers have been proposed to be useful for differentiating between benign and malignant papillary lesions. Different genetic aberrations have been identified in different papillary lesions, but their clinical application requires further validation. Pre-excision samples obtained with CNB can be difficult to interpret, and may result in unavoidable undercall, as the full extent of the lesion may not be assessable in CNBs. When a difficult CNB is reported, the potential for underdiagnosis should be communicated to the surgeons. The optimal management of patients with papillary lesions of the breast requires full cooperation and understanding among the surgeons, radiologists, and pathologists. 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