Barrett's Esophagus and Adenocarcinoma of the Gastroesophageal Junction: a Pathologic Perspective

Surg Oncol Clin N Am 15 (2006) 715–732

Barrett’s Esophagus and Adenocarcinoma of the Gastroesophageal Junction: a Pathologic Perspective Laura H. Tang, MD, PhD*, David S. Klimstra, MD Department of Pathology, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, Room C507, New York, NY 10021, USA

Barrett’s esophagus is defined clinically by the presence of endoscopically evident columnar mucosa in the distal esophagus with histopathologic confirmation of the presence of intestinal-type epithelium. The eponym ‘‘Barrett’s’’ esophagus represents a specialized intestinal metaplasia of normal squamocolumnar epithelium at the gastroesophageal junction (GEJ), and reflects the original observation of Norman Barrett of what he erroneously believed to be ectopic gastric mucosa. The etiology of Barrett’s esophagus is understood poorly, but chronic gastroesophageal reflux disease (GERD) is considered a major contributing factor among other factors, such as obesity and abnormal GEJ anatomy [1]. Barrett’s esophagus has been documented, from the historical perspective and the contemporary evidence, to be associated with the development of adenocarcinoma of the GEJ [2]. As with neoplasia elsewhere, this transformation from normal to a malignant mucosa occurs through a sequence of morphologic changes that is accompanied by specific molecular genetic events [3,4]. It is believed that the development of a Barrett-type mucosa with intestinal goblet-type cells is due to an altered process of differentiation of pluripotent epithelial stem cells in response to the local injury and repair process [5], which reflects acid and possibly bile-induced damage. Although the morphologic transformation of the Barrett-type epithelium has been well recognized, the exact sequence of molecular events that occurs in GEJ adenocarcinoma is not as well established as that identified in the colon. The potential identification

* Corresponding author. E-mail address: [email protected] (L.H. Tang). 1055-3207/06/$ - see front matter Ó 2006 Elsevier Inc. All rights reserved. doi:10.1016/j.soc.2006.07.007 surgonc.theclinics.com

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and isolation of markers for screening purposes and possibly prognostic information are areas of considerable clinical and scientific interest. Most instances of Barrett’s esophagus are identified in patients who have signs and symptoms of gastroesophageal reflux [6]. A subset of patients is diagnosed with the condition while being evaluated endoscopically for a nonreflux-related symptomatology [6]. Biopsy specimens that are taken from the columnar mucosa usually reveal evidence of intestinal metaplasia. Furthermore, this type of metaplastic change almost invariably is evident in the epithelium surrounding most cases of GEJ adenocarcinoma. As a result of such observations, Barrett’s esophagus has been considered as a major risk factor for adenocarcinoma at the GEJ. The similar recognition that adenocarcinoma of the GEJ has been increasing at a dramatic rate has led to the development of a high level of interest in the pathology, early identification, and treatment of the condition [6]. The precise relationship between Barrett’s esophagus and the development of GEJ adenocarcinoma is not understood completely. Approximately 10% to 20% of individuals who undergo upper gastrointestinal endoscopy carry a diagnosis of Barrett’s esophagus, and the prevalence and incidence of adenocarcinoma in this setting is only 5% to 10% and less than 1%, respectively [6–8]. Thus, the principal issue in Barrett’s esophagus is to identify and define its relationship with the incidence of adenocarcinoma of the distal esophagus, which is the most rapidly increasing cancer in the United States [9], and there exists a critical need to improve the therapeutic efficacy with which it is managed. The yield of endoscopic surveillance program for Barrett’s esophagus is low [10]. Cellular and molecular markers are more likely to facilitate the identification of individuals whose metaplasia is of pathologic significance in terms of the development of dysplastic and neoplastic transformation. Although several such candidate markers have been proposed, routine application of them has not been the practice. Thus, a combined endoscopic surveillance and histopathologic evaluation remain the gold standard for the assessment of the progression of Barrett’s esophagus [1]. The diagnosis of Barrett’s esophagus is not only limited to a wide spectrum of sensitive patient care issues but includes overtly emotive therapeutic responses from physicians. Following the diagnosis of Barrett’s esophagus, patients require cautious education to understand its clinical course, treatment options, the statistical risk for the development of adenocarcinoma, appropriate surveillance, and the importance of compliance. Most of these issues, at the early stage of the diagnosis, are handled by clinicians, usually gastroenterologists; however, given a wide spectrum of current treatment options [11–16], radiologists, surgeons, and oncologists may be involved at different stages of the disease. Because of the lack of definitive guidelines for treating Barrett’s esophagus and related adenocarcinoma, there is a plethora of complex diagnostic and therapeutic options with which patients need to become familiar (eg, which pills to take? how frequently

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should endoscopy be performed? when to have photodynamic or radiation therapy; is surgery necessary? what kind of surgery is required? if adenocarcinoma is documented, is neo-adjuvant or adjuvant chemotherapy required?). Thus, it is apparent that the appropriate clinical management of Barrett’s esophagus requires a multidisciplinary clinical team. In this particular disease setting, specialized gastrointestinal pathologists play a crucial role in the clinical decision-making. Therefore, this article outlines Barrett’s esophagus from the perspective of a pathologist with the focus on biology, pathobiology, histopathology, and the criteria that are involved in the diagnosis of dysplasia. In addition, it emphasizes issues that are related to communications between pathologists and gastroenterologists, surgeons, and oncologists. It is mandatory that every team member who is involved in the management of Barrett’s esophagus provides adequate information and delivers comprehensible messages to each other. This strategy assures the development of a comprehensive multifaceted management strategy for individual patients. To assure prompt therapeutic action and the safety of a patient, a specialized gastrointestinal pathologist should understand as much of the clinical issues that are relevant to the condition as the clinician should comprehend the statements that are contained in pathologic reports. History of Barrett’s esophagus In 1950, Norman Barrett published a report in the British journal Surgery in which he defined the esophagus as ‘‘that part of the foregut, distal to cricopharyngeal the sphincter, which is lined by squamous epithelium’’ [17]. He went on to describe several patients who had ulcerations in a tubular, intrathoracic organ that appeared to be the esophagus, except that its distal portion was lined extensively by a ‘‘gastric-type’’ mucosa. Because the esophagus was, by definition, a squamous-lined structure, Barrett believed that the columnar-lined structure was a tubular segment of the stomach generated by traction induced by a congenitally short esophagus and tethered within the chest. At that time, Barrett did not mention intestinal features (goblet cells) in this columnar lining, nor did he raise the question of intestinal metaplasia. Subsequently in 1951, Bosher and Taylor commented on the appearance of ‘‘heterotopic gastric mucosa’’ in the esophagus with ulceration and stricture formation. They noted that ‘‘the ‘gastric mucosa’ was composed of glands which contained intestinal type goblet cells but not parietal cells.’’ A year later Morson and Belcher commented upon the relationship of adenocarcinoma of the esophagus and the ‘‘ectopic gastric mucosa.’’ As pathologists they also noted that in an individual who has adenocarcinoma, the esophageal mucosa exhibited ‘‘atrophic changes with changes towards an intestinal type containing many goblet cells’’ [17]. In 1953, Allison and Johnstone [18] proposed that Barrett had misidentified the columnar-lined intrathoracic structure as stomach, and that in reality it

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was the portion of distal esophagus lined by ‘‘gastric mucosa.’’ They noted that in contrast to the stomach, the structure lacked a peritoneal covering, often harbored islands of squamous epithelium, and possessed the type of submucosal glands and muscularis propria that were characteristic of those of the esophagus. Thus, 7 years after his original report, Barrett [19] reversed himself and concurred that the columnar-lined organ that he had previously believed to be the stomach was, in fact, the esophagus; he suggested that the condition be called ‘‘lower esophagus lined by columnar epithelium.’’ Despite the clarification, the condition remains known as Barrett’s esophagus and continues to invoke as much confusion as when described initially.

Pathology of Barrett’s esophagus–associated adenocarcinoma Macroscopic pathology The GEJ can be defined physiologically, anatomically, and microscopically. Anatomic landmarks, such as the peritoneal reflection from the stomach onto the diaphragm or the incisura (angle of His), can be used to define the GEJ in surgically resected specimens [20]. Endoscopically, the proximal margin of gastric fold is used as a guide to define the GEJ [1]. Despite the different landmarks used, the gross appearance of Barrett’s mucosa in surgically resected specimens is correlated closely to that observed at endoscopy (Fig. 1). Examination of the distal esophagus at the GEJ reveals flat and salmon-pink mucosa above the squamocolumnar junction (Z-line), which is presented as single or multiple short, fingerlike protrusions, or a long segment of such mucosa well into the tubular squamous mucosa. The former is regarded as short-segment (!3 cm) Barrett’s esophagus and the latter is regarded as long-segment (R3 cm) Barrett’s esophagus [1]. This assessment is based on a well-delineated Z-line (see Fig. 1a), however, which is not always evident in many individuals because the squamocolumnar junctional mucosa becomes irregular with severe reflux or circumferential Barrett’s mucosa (Fig. 2). In particular, when the columnar-type mucosa is present in continuity with the Z-line, the distinction between columnar-type epithelium without or with scant intestinal metaplasia often is not possible by gross or endoscopic examination. In the absence of goblet cells, the flat columnar-type epithelium is considered as cardia-type mucosa by histologic definition (see Fig. 1), and is not a criterion for the diagnosis of Barrett’s esophagus [1]. Furthermore, the degree of glandular dysplasia within Barrett’s mucosa cannot be evaluated by macroscopic examination (see Fig. 2). Most cases of primary adenocarcinomas of the esophagus arise in the lower third or the esophagus within the segment of Barrett’s mucosa [21,22], which may be recognizable adjacent to the tumor as typical salmon-pink flat mucosa, particularly at the early stage of the carcinoma (see Fig. 2). Superficial adenocarcinoma, which usually is identified in the course of surveillance of Barrett’s esophagus, may exhibit subtle

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Fig. 1. Macro- and microanatomy of the GEJ. (A) Formalin-fixed specimen reveals a welldelineated squamocolumnar junction (Z-line). (B) Histology of the GEJ (original magnification  10), corresponding with the gross specimen in (A), reveals the squamous mucosa located on the top, the proximal gastric fundic-type mucosa in the lower portion, and a short segment of simple mucinous columnar-type mucosa situated in between. (C) A zoom view of a segment in (B) (original magnification  25), which demonstrates an epithelial transition from a columnar mucosa with loosely packed submucosal mucinous glands (thick arrow) to fundic mucosa with oxyntic glands (thin arrow). The phenotype of the surface mucinous epithelium appears to remain unchanged.

macroscopic alterations within Barrett’s mucosa, such as mucosal bumps or plaques. At the time of diagnosis, however, most tumors are at the advanced stage with an ulcerative or mass lesion infiltrating into the esophageal wall. In some of these advanced cases, Barrett’s mucosa may be displaced by overgrowth of the tumor and is no longer recognizable in the adjacent mucosa by gross or microscopic examination. In rare cases, when an adenocarcinoma of the esophagus arises truly independent of Barrett’s esophagus, the histogenesis of the tumor that arises from ectopic gastric glands (gastric inlet) or from peri-esophageal glands should be considered. These tumors also are found in the upper and middle portion of the esophagus [23]. Histopathology Definition of Barrett’s esophagus Barrett’s esophagus is defined histologically by the presence of intestinaltype goblet cells within the columnar-type epithelium that contain basophilic acid mucin and can be detected by Alcian Blue at pH 2.5 [1]. In contrast, the mucin that is present in the columnar-type epithelium is clear and slightly eosinophilic. Histochemically, the mucin is neutral, periodic acid-Schiff–positive, but Alcian Blue is negative at pH 2.5 or lower.

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Fig. 2. Specimens of esophagectomy with Barrett’s lesions. (A) A long segment of Barrett’s mucosa extends 6 cm up into the esophagus with a mixed low- and high-grade dysplasia on histopathologic examination. (B) Esophagectomy, after neoadjuvant radiation and chemotherapy, reveals treatment-related fibrosis (star) and residual Barrett’s esophagus (arrow); residual adenocarcinoma is identified only on microscopic examination. (C) Adenocarcinoma arises in distal esophagus and straddles the GEJ.

The columnar-type epithelium that is associated with Barrett’s esophagus is similar to surface mucous glands of gastric mucosa by morphology and by histochemistry. This type of columnar epithelium, present at the GEJ, is considered as cardia-type mucosa (see Fig. 1). Cardia-type mucosa and its association with Barrett’s mucosa Considerable controversy exists in regard to the location, extent, and even the existence of the gastric cardia [24]. According to anatomic description, the term ‘‘cardia’’ applies to the part of the stomach that lies around the orifice of the tubular esophagus, at the level of the angle of His. This is the point at which the tubular esophagus joins the saccular stomach (see Fig. 1). The American Joint Committee on Cancer describes the cardia as the first part of the stomach, which is located immediately below the diaphragm and the GEJ [25]. Endoscopically, there is no anatomic landmark for the distal margin of the so-called ‘‘cardia.’’ The squamocolumnar junction (Z-line) may occur at or above the anatomic GEJ. When the squamocolumnar junction is located above the GEJ, there is a columnar-lined segment of the esophagus. Histopathologists consider the cardia-type mucosa to be composed of shallow mucus-secreting epithelium with loosely packed or cystic mucinous glands beneath its surface (see Fig. 1). The histologic finding of cardiatype mucosa does not correlate necessarily with the anatomically defined cardia, because it can be present in the tubular esophagus. This discrepancy probably can be explained by the micro- and functional anatomy of each individual. Examinations of autopsy material from embryos, fetuses, and infants indicates that cardiac mucosa is present in all cases, but varies in

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length throughout gestation [26]. Cardia mucosa is present at birth as a normal structure, and is located in the distal esophagus above the angle of His. Thus, the cardia-type epithelium probably is not an abnormal finding in adults, and it may represent a physiologically accepted metaplastic process that has developed as a consequence of the dynamic function of the lower esophagus. Of particular interest is that the development of intestinal metaplasia occurs invariably within the columnar-type mucosa. Thus, it is an issue of debate as to whether Barrett’s esophagus and associated neoplasia may develop from the columnar/cardia-type mucosa [27,28]. Immunoprofile of Barrett’s mucosa Several studies have sought to identify sensitive and specific markers of intestinal-type mucosa in the esophagus. These markers include the MUC antigens and other mucin components, and different cytokeratin (CK) subtypes. CKs are the intermediate filaments that are characteristic of epithelial cells. Their expression has been identified in more than 20 distinct forms, with highly variable patterns among specific types of epithelium. Regarding Barrett’s esophagus, it has been proposed that there is a unique pattern of CK7/CK20 expression of Barrett’s intestinal metaplasia, with a strong CK7 staining at the surface and in deep glands, and a weak superficial CK20 positivity [29]. Although this sensitive and specific CK pattern has been observed in Barrett’s esophagus [30], most cases of Barrett’s esophagus are easily identifiable on routine hematoxylin and eosin (H&E)–stained sections, and additional immunohistochemical studies usually are not necessary. In respect to the significance of CK immunoreactivity in preneoplastic conditions, it seems unlikely that this pattern will be of great usefulness, because it usually provides information that is similar to that obtained with mucin histochemistry, and lacks sensitivity for preneoplasia. Other antibodies that are directed against MUC mucin gene products, especially MUC1 and MUC2 (intestinal mucin), also have been used to characterize intestinal-type epithelium of Barrett’s esophagus. These studies have demonstrated an abnormal MUC2 expression in Barrett’s intestinal mucosa, which is lost when the cells become neoplastic. MUC1 was absent in metaplastic and dysplastic epithelium, but was expressed in carcinomas, which suggests that it could differentiate dysplasia from carcinoma in mucosal biopsies [31,32]. Dysplasia in Barrett’s esophagus Because the major risk for patients who have Barrett’s esophagus is the development of an adenocarcinoma, there has been considerable interest in defining a subgroup of patients at risk in whom effective surveillance can be undertaken. At this time, only morphologic assessment, especially epithelial dysplasia, is of use in delineating this subpopulation. Definition. Dysplasia primarily is a morphologic term and is considered a carcinoma precursor and a marker of high risk for malignancy in several

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gastrointestinal sites. As such, it has been defined as an unequivocally neoplastic epithelium confined within the basement membrane of the gland from which it arises [33]. Although this definition initially was proposed in reference to premalignant morphologic alterations that developed in inflammatory bowel disease, it has been extended to apply to the entire gastrointestinal tract, including Barrett’s esophagus [34]. The morphologic criteria for the identification of dysplasia include a combination of architectural and cytologic abnormalities, each of which may occur in different quantity, quality, and intensity. Dysplasia is divided into a two-tiered system of low-grade dysplasia (LGD) and high-grade dysplasia (HGD) [35]. As a result of this degree of complexity in definition and classification of dysplasia, an assessment of dysplasia culminates in considerable subjectivity and less than optimal reproducibility among gastrointestinal pathologists. Unfortunately, there are no other reliable markers and practical solutions for defining dysplasia and for evaluating high cancer risk in a particular individual. Several criteria, guidelines, illustrations, and algorithms have been developed in an attempt to increase reproducibility in the assessment of Barrett’s dysplasia [1,4,36,37]; however, depending on the training and the experience, there is intra- and interobserver inconsistency even among the most experienced gastrointestinal pathologists [38]. Although observer variation in the assessment of Barrett’s dysplasia and related lesions is not likely to improve based on morphologic evaluation alone, the clinical impact of the issue is less significant because pathologists are extremely consistent when analyzing lesions of the highest and lowest risk in Barrett’s dysplasia. Those in the intermediate categories are in the minority, and, therefore, form a smaller subgroup [39]. Classification of dysplasia. Because dysplasia is a morphologic entity that is diagnosed in routine histopathologic examination, it has to be recognized on endoscopic biopsies with routinely H&E-stained sections. The most significant issue in diagnosis is to distinguish dysplasia from the two ends of the morphologic spectrum (eg, from reactive/regenerative nonneoplastic modifications, often called reactive atypia, and from invasive carcinoma, especially in its early stage or in a superficial fashion with invasion limited to the lamina propria). Barrett’s esophagus is classified as negative for dysplasia; indefinite for dysplasia; LGD; and HGD. This is based on a combination of the following morphologic features: (1) maturation of the surface epithelium, (2) overall architecture, (3) cytologic and nuclear features, and (4) field effect, such as inflammation and procedure-related epithelial atypia and regeneration. The general morphologic features are summarized in Table 1. Among the grading system, the category of ‘‘indefinite for dysplasia’’ usually elicits the most conflicting issues between pathologists and clinicians. The most commonly encountered situation in this category is the presence of active inflammation of the mucosa with associated reactive cytologic atypia,

Table 1 Morphologic criteria used in the classification of Barrett’s dysplasia Morphologic features

Negative for dysplasia (Fig. 3a)

Surface gland maturation Normal architectural ratio of glands to lamina propria Bland cytologic features No significant background inflammation Surface gland maturation Retained architectural ratio of glands to lamina propria Mild to moderate nuclear atypia, may be prominent within areas of active inflammatory infiltrate Background of active inflammation, erosion, or ulceration, or recent field radiation or invasive intervention. Loss of surface maturation with or without villiform configuration Mild architectural distortion, including glandular crowding or minimal papillary proliferation within gland lumen. Pseudo-nuclear stratification and increased nuclear to cytoplasmic ratio, but retained cellular polarity. No significant background inflammation Complete loss of surface maturation and cellular polarity Significant architectural complexity, including gland crowding (overrunning lamina propria) and prominent papillary extension or cribriforming within gland. Significant nuclear abnormality with nuclear pleomorphism and frequent mitoses Inflammation may be present, but usually not prominent.

Indefinite for dysplasia (Fig. 3b)

Low-grade dysplasia (Fig. 3c)

High-grade dysplasia (Fig. 3d)

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Classification of Barrett’s dysplasia

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which may be indistinguishable for the degree of cytologic atypia seen in association with true dysplasia. This also may be seen in mucosa that are subjected to radiation therapy or previous invasive procedures. Thus, to avoid an overinterpretation of dysplasia or overlooking a genuine dysplasia that is masked by inflammation, the category of ‘‘indefinite for dysplasia’’ seems to be most appropriate under the circumstances, provided there exists a mutual understanding between the team members that are involved in the management of Barrett’s esophagus. Some pathologists might further explicate the description by referring to ‘‘indefinite for dysplasia favor reactive or indefinite for dysplasia favor dysplasia’’ if relevant clinical information is provided. In either situation, the message to the gastroenterologist would be a recommendation of rebiopsy when the inflammation subsides. In the interest of facilitating clinical decision-making, a previous threetiered classification (mild–moderate–severe) for Barrett’s dysplasia is no longer used by most pathologists [33,38]. In the two-grade system, LGD includes the mild and moderate dysplasia referred to in the three-tiered system; HGD is reserved for severe dysplasia only. Dysplasia must be recognized as constituting a continuum, however. In the spectrum of the moderate dysplasia in the previous three-tiered system, the authors believe that the high end of this spectrum should be documented cautiously. Pathologists should maintain a high threshold before identifying HGD, because, depending on management strategies in different institutions, some patients may undergo surgical resection if labeled with this diagnosis [16,40]. Thus, in instances where there is LGD that is almost atypical enough to fall in the high-grade category, it is appropriate for pathologists to alert the clinician that there is more than the earliest stage of dysplasia with a recommendation of perhaps an earlier rebiopsy on the next schedule, or an immediate rebiopsy if there is a concern of insufficient sampling. This strategy should ensure that a HGD is not overlooked. Safety and danger in diagnosis of dysplasia for pathologists Evaluation of Barrett’s esophagus and related dysplasia may be intimidating for less experienced pathologists; nevertheless, familiarity can be acquired by following certain criteria for safety and by avoiding several potentially dangerous interpretations (Box 1). Furthermore, in view of accommodating clinical surveillance and intervention, it may be pertinent for pathologists, when possible, to provide quantitative and qualitative assessment of specific lesions (eg, the presence of rare goblet cells versus extensive intestinal metaplasia; focal versus diffuse dysplasia; superficial versus infiltrating adenocarcinoma). Such documentation can provide information on disease progression as well as guidance for future assessment of biopsy material. Barrett’s-associated adenocarcinoma When evaluating patients in surveillance cohorts, it has been established that the presence of dysplasia indicates an increased risk for

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Box 1. Guidance for the safe assessment of Barrett’s dysplasia and adenocarcinoma in biopsies Criteria for safety Awareness of clinical and gastrointestinal endoscopic history of the patient Understand clinical implications in pathologic diagnosis of dysplasia. Establish comprehensive communication with team members involved in management of Barrett’s esophagus. Take particular caution when diagnosing dysplasia for the first time, review previous biopsy material if available. Seek a second opinion when dealing with complex or difficult cases. Provide a clear and intelligible report; add a note to explain if necessary. Criteria for danger Contradiction of the safety criteria, and in addition: Diagnosing HGD or carcinoma in a patient for the first time without communicating with clinical team members Diagnosing dysplasia or carcinoma in a background of active inflammation, ulceration, or after treatment (although it may occur in some) Rendering a diagnosis with insufficient material Subjectively influenced by previously rendered diagnoses

adenocarcinoma; however, the natural history of this dysplasia, particularly LGD, is difficult to predict in individual patients [41]. The natural history of HGD is a matter of debate. According to studies from the last 2 decades, the detection of HGD in patients who have Barrett’s esophagus is the indication of a synchronous adenocarcinoma. The neoplasia has been claimed to remain undetected even by the most rigorous biopsy protocols, only to be discovered in surgically resected specimens. The reported prevalence of such adenocarcinoma ranges from 0% to 75% in some series, and it has been estimated that an average 40% occult adenocarcinoma occurs in HGD [42,43]. Such a figure requires more vigorous evaluation and the presentation of more robust data [8,44]. As a consequence of such conflicting data regarding the likelihood of occult adenocarcinoma, the reported biopsy protocols that lead to the surgical resection vary considerably between centers [14–16,40,45]. Adenocarcinoma arising in Barrett’s esophagus with HGD is defined by neoplastic cells that have penetrated through the basement membrane and infiltrated into the lamina propria or beyond. Most pathologists are unlikely

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to miss submucosal invasive carcinomas, particularly when provided with the clinical and endoscopic impression of a mass lesions; however, the diagnosis of a superficial adenocarcinoma may be a delicate issue in certain clinical settings. When the biopsy is superficial or the lesion itself is superficial, the impact of the diagnosis of carcinoma versus HGD should be taken into serous consideration. In view of the fact that even the presence of lamina propria invasion alone carries the risk for regional lymph node metastasis, considerable caution must be exercised. Thus, the diagnosis of a superficially invasive carcinoma usually leads to surgical intervention. The assessment of combined architectural and cytologic features is necessary in the identification of adenocarcinoma in biopsy specimens, in the same manner as in dysplasia. The malignant epithelium that is present in carcinoma exhibits a high level of architectural complexity and glandular proliferation has extended beyond crowding, as noted in HGD. The malignant glands begin to merge with each other and form an anatomizing pattern, in which the contour of individual glands is no longer apparent. Budding of small clusters or individual neoplastic cells from highly dysplastic glands also are helpful clues (Fig. 3). Nuclear pleomorphism usually is more apparent in carcinoma than in HGD with associated macronucleoli and atypical mitoses. Evaluation of Barrett’s adenocarcinoma in surgical specimens To ensure accurate tumor staging and postoperative management, the specimen should be processed with a particular focus on tumor location, depth of invasion, pathology of the surrounding mucosa, adequate lymph node dissection from the specimen, and treatment-related changes. When the tumor straddles the GEJ, the proportion of the lesion in the esophagus and in the stomach should be documented carefully. The tissue sections are oriented and assessed best after at least several hours of formalin fixation. If possible, gross photographs should be taken before and after fixation to assure best illustration of the lesion, which can be complementary to the histologic assessment. In the absence of a large tumor mass, it is best to block the GEJ involved by tumor and submit the entire area for histologic evaluation. The staging for adenocarcinoma of the GEJ is included in esophageal tumor staging in the sixth edition of the American Joint Committee on Cancer staging system, in which extensive lymph node assessment is not required [25]. The clinical prognosis of GEJ adenocarcinoma is predicted best by adequate lymph node evaluation, however; thus, the lymph node staging for gastric carcinoma, at least 15 lymph nodes, has been evaluated at the authors’ institution. The pathologic assessment of GEJ carcinoma in response to radiation/ chemotherapy involves the gross and the microscopic examination of the resected surgical specimen. The gross appearance of treated tumors varies from mucosal ulceration to a fibrous scar, or a prominent mass lesion in the case of a less than profound tumor regression. At the microscopic level,

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Fig. 3. Examples of Barrett’s esophagus and associated lesions. (A) Barrett’s esophagus reveals diffuse intestinal metaplasia without dysplasia. (B) Barrett’s esophagus–indefinite for dysplasia. The mucosa is inflamed actively with associated nuclear atypia; however, surface epithelial maturation is evident. (C) Barrett’s esophagus with low-grade glandular dysplasia; the surface epithelium exhibits nuclear stratification and lacks complete maturation, but the cells remain polarized. (D) Barrett’s esophagus with high-grade dysplasia; there is significant nuclear atypia and loss of surface epithelial maturation and polarity. (E) Barrett’s esophagus with severe glandular dysplasia and associated architectural complexity, suspicious for superficial adenocarcinoma with lamina propria invasion. (F) Superficially invasive adenocarcinoma in association with high-grade glandular dysplasia.

a positive treatment related-effect is observed as abolition of the malignant epithelium and replacement by reactive fibrosis or fibro-inflammation within the mucosa or the gastroesophageal wall. Thus, the ultimate pathologic response to treatment is determined by the amount of residual viable carcinoma in relation to areas of fibrosis or fibro-inflammation within the gross

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lesion, which is associated inversely with, and expressed as, percentage of a favorable treatment response. Thus, a 100% treatment response indicates fibrosis or fibro-inflammation within an entire gross lesion without microscopic evidence of carcinoma, and a 0% response represents an entirely viable tumor in the absence of any fibrosis of fibro-inflammation. Acellular mucin is regarded as a form of positive treatment response, not as residual/viable tumor. The pathologic stage of the residual carcinoma is determined by the presence of viable malignant epithelium in the deepest layer of the gastroesophageal wall. Positive lymph nodes are defined as having at least one focus of viable tumor cells in lymph nodes. Pathologists are confronted often with adenocarcinomas that straddle the GEJ, and there has been considerable debate regarding the tumor genesis and the relationship between GEJ and gastric cardia adenocarcinoma (see Fig. 2). Various criteria have been used to categorize tumors that are situated at the GEJ. In most classification systems, the anatomic location of the epicenter or predominant mass of the tumor is used to determine whether the neoplasm is esophageal or gastric (cardia) in origin [24,46,47]. It is almost impossible to document that in a given location, such as the GEJ, tumors will grow to the same extent in a proximal and distal direction, which allows us to conclude that the epicenter is the origin. There exists no consensus regarding the definition of cancer of the gastric cardia. Most data on cardia mucosa, its dysplasia, and cardiac cancer are not comparable because of the lack of diagnostic criteria. Uniformity in classification, terminology, and diagnostic criteria are required to clarify the issue of cardia, carditis, and cardiac adenocarcinoma and its relationship with adenocarcinoma of the GEJ [23,26,47]. It seems, however, that the similarities between adenocarcinoma of the GEJ or cardia and Barrett’s adenocarcinoma outnumber the dissimilarities.

Molecular pathology of Barrett’s esophagus and adenocarcinoma On morphologic examination it is apparent that the carcinogenetic process of Barrett’s mucosa progresses through increasing grades of epithelial dysplasia. In addition and parallel to the morphologic sequence of events that leads from metaplasia to carcinoma in Barrett’s mucosa, several changes in gene structure, gene expression, and protein structure are associated with the progression of Barrett’s esophagus to adenocarcinoma. Accumulation of these changes, rather than the exact sequence of these changes, seems to be essential. Although the frequency and timing of these alterations are not as well established as are those in colorectal carcinogenesis, some investigators have proposed a molecular cancer progression scheme of Barrett’s esophagus, in which multiple molecular pathways are involved and interact with each other [2,6,48–50]. Alterations in tumor suppressor genes, among which p53 and p16 are early events in the

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metaplasia-dysplasia-adenocarcinoma sequence, which is followed by subsequent alterations in cell cycle regulators, such as alterations in cyclin D1 and E, growth factors, and receptors (tumor growth factor-a, epidermal growth factor receptor, Her2-Neu, ERBB2). Ongoing genomic instability leads to cumulative genetic errors, and, thereby, the generation of multiple clones of transformed cells. Cytogenetic studies using G-banding, interphase fluorescence in situ hybridization, and comparative genomic hybridization have revealed a complex pattern of structural and numerical chromosomal aberrations in adenocarcinoma of the distal esophagus and gastric cardia [51–54]. On the molecular genetic level, microsatellite analyses revealed frequent loss of heterozygosity (LOH) as well as allelic imbalances on chromosomes 5q, 17p, and 18q [55] and on chromosomes 3q, 4q, 5q, 6q, 9p, 9q, 12p, 12q, 17p, and 18q [56]. The latter studies have provided support for the proposed metaplasiadysplasia-carcinoma sequence, and demonstrated a sequential accumulation of alterations and microsatellite changes in metaplasia and dysplasia. Some molecular alterations may be helpful in recognizing high-risk patients who have Barrett’s esophagus and are liable to develop cancer, although such markers are not available for routine diagnostic use. Alterations that seem to have potential in the surveillance of patients include changes in DNA ploidy (tetraploidy and aneuploidy), which can be evaluated by flow cytometry. In addition, abnormalities in p16 (methylation, mutation, and loss of heterozygosity or LOH), and p53 (mutation, LOH) can be assessed more reliably by molecular methods than by immunohistochemistry, which promise and merit further clinical research.

Summary Barrett’s esophagus is recognized clearly to possess potential for neoplastic progression that can be identified endoscopically and confirmed by histologic evaluation. A diagnosis of dysplasia in Barrett’s esophagus has major clinical and therapeutic consequences, although numerous studies have demonstrated that it is not completely reproducible. To ensure patient safety, it is imperative to establish a clear communication structure between individual team members who are involved in the management of Barrett’s esophagus. Several nonmorphologic markers that have been proposed to complement the valuation reflect recent advances in the genetic and molecular processes that are involved in the carcinogenesis of Barrett’s esophagus. The best ancillary tools are immunohistochemistry with antibodies directed against specific biomarkers, flow cytometry to assess gross DNA abnormalities, and fluorescence in situ hybridization for the detection of chromosomal aberrations. It is probable that in the near future techniques, such as global gene expression profiling with cDNA microarrays, identification of polymorphic genetic markers using single nucleotide

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