Hyperplastic-like Colon Polyps That Preceded Microsatellite-Unstable Adenocarcinomas

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1 Anatomic Pathology / HYPERPLASTIC-LIKE COLON POLYPS PRECEDING MICROSATELLITE-UNSTABLE ADENOCARCINOMAS Hyperplastic-like Colon Polyps That Preceded Microsatellite-Unstable Adenocarcinomas Neal S. Goldstein, MD, Punam Bhanot, MD, Eva Odish, HTL(ASCP), and Susan Hunter, SI(ASCP) Key Words: Colon; Serrated; Adenoma; Hyperplastic; Polyp; Hereditary nonpolyposis colon cancer; HNPCC Abstract We compared hyperplastic-like polyps that preceded microsatellite-unstable adenocarcinomas to incidental hyperplastic polyps to identify distinguishing morphologic criteria. The study group included 106 hyperplastic-like, nonadenomatous, serrated polyps, most from the ascending colon in 91 patients; the control group included 106 rectosigmoid hyperplastic polyps from 106 patients in whom adenocarcinoma did not develop. Study group polyps had an expanded crypt proliferative zone, a serrated architectural outline that became apparent in the basilar crypt regions, basilar crypt dilation, inverted crypts, and a predominance of dysmaturational crypts (crypts with minimal cell maturation). In contrast, control group polyps had a proliferative zone confined to the basal crypt region, serrated architecture that became apparent in the superficial crypt region, rare to no basilar crypt dilation, and rare or no dysmaturational crypts. Hyperplastic-like polyps that preceded microsatellite-unstable adenocarcinomas had a distinctive constellation of morphologic features related to altered and decreased cell function and control that resulted in dysmaturational crypts. Dysmaturation constitutes a range of morphologic alterations, some of which overlap with incidental-type innocuous hyperplastic polyps. The morphologic features described herein provide initial guidelines to identify this potentially important subset of premalignant serrated-like polyps. Approximately 15% of sporadic, or nonsyndromic, colorectal adenocarcinomas are microsatellite unstable and have deficiencies in their mismatch repair enzyme complex. 1-9 Serrated adenomas are thought to be the most common precursor lesion in this group of adenocarcinomas Jass et al, Jass and Talbot, 46 and Jass proposed a serrated neoplasia precursor pathway concept for microsatellite unstable neoplasms. They conceptually linked hyperplastic polyps and serrated adenomas with microsatellite-unstable adenocarcinomas and suggested that some hyperplastic polyps were the initial lesion in the serrated neoplasia pathway. The morphologic features of these putative initial pathway hyperplastic-like polyps is not well characterized. Features that distinguish them from innocent hyperplastic polyps are poorly defined. This distinction may be important. As a group, serrated adenomas are precursor lesions that have a risk of progression to adenocarcinoma and require complete eradication and enrollment of patients into a surveillance colonoscopy program. Conversely, incidental-type hyperplastic polyps are not treated in this manner We studied hyperplastic-like serrated polyps that preceded sporadic microsatellite instability (MSI)-high adenocarcinomas to identify a preliminary set of morphologic criteria for this entity. The goal of the present study was limited to the identification of morphologic features that might distinguish this subset of potentially biologically important lesions. Materials and Methods Study group polyps were resected or biopsied hyperplastic polyps that preceded in the same site mismatch repair deficient or MSI-high adenocarcinomas. An initial 778 Am J Clin Pathol 2003;119: Downloaded 778 from

2 Anatomic Pathology / ORIGINAL ARTICLE William Beaumont Health Care system (Royal Oak, MI) computer search for the period January, 1, 1980, through December 30, 2000, identified 1,417 potential cases from patients older than 65 years with cecum-transverse colon adenocarcinomas and a preceding hyperplastic polyp from the same location as the adenocarcinoma. These criteria were chosen to maximize the yield of sporadic (nonhereditary) microsatellite-unstable adenocarcinomas. 3,25,35,57-71 Additional refined computer searches and a slide review excluded 870 cases based on chemotherapy, inflammatory bowel disease, a preceding polyp that was an adenoma (tubular or serrated), insufficient colon site information, or unavailable tissue blocks. 17,19 Sections of the remaining 547 adenocarcinomas were evaluated for mismatch repair enzyme deficiency (absent hmlh1 immunoreactivity) or microsatellite instability (MSI-high) using the 5 Bethesda consensus conference derived primers. 61,66,72,73 Ninetyone colon adenocarcinomas were determined to be mismatch repair deficient and/or microsatellite-unstable adenocarcinomas. The final study group was 106 hyperplastic-like polyps that preceded 91 same-site, microsatellite-unstable adenocarcinomas. None of the study group polyps had adenomatous epithelium. Control group polyps were selected randomly from a large pool of patients with rectal or sigmoid, solitary, hyperplastic polyps completely resected during the period January 1, 1984, through December 31, 1987, and who did not have colorectal adenocarcinoma before or after the sentinel colonoscopy; were alive on the follow-up date of January 1, 2002; and underwent a subsequent colonoscopy that did not yield a tubular adenoma from the same region of the colon as the hyperplastic polyp. One control hyperplastic polyp was selected for each serrated study polyp. Pathologic Evaluation The evaluation included 18 features: (1) polyp dimension; (2) collagen band thickness; (3) location of the initial serrated crypt architecture; (4) quantity of dilated crypt bases; (5) quantity of horizontal crypts; (6) presence of inverted crypts; (7) crypt branching; (8) mitoses; (9) predominant nuclear shape in the basilar, middle, and superficial or upper-third sections of the crypts; (10) prominence of nucleoli in the cells of the basilar, middle, and superficial crypt regions; (11) presence of goblet cells; (12) presence of distended, mucin-filled cells in superficial crypts; (13) presence of cells with abundant eosinophilic cytoplasm in superficial crypts; (14) nuclear maturation; (15) cell maturation; (16) percentage of dysmaturational crypts; (17) loss of hmlh1 nuclear immunoreactivity in crypt epithelial cells; and (18) mean nuclear area of cells in the basilar, middle, and superficial crypt regions. The morphologic features chosen to be evaluated were selected based on the findings of previous authors. 14,17,24,35 Identical or similar scoring systems were used when feasible. The polyp dimension was determined from the endoscopy report if resected by piecemeal excision in fragments or from the gross description in the pathology report if the specimen was a single-snare polypectomy specimen. The maximum thickness of the collagen band beneath the surface epithelium was measured (in micrometers). Architectural Features The location of the initial serrated crypt architecture was classified as the basilar, middle, or superficial region. Dilated crypt bases were classified as none (all crypt bases were a smaller diameter than the immediately suprabasilar crypt segment), rare isolated crypts or a cluster of fewer than 5 crypts, 10% to 33% of crypts, or more than 33% of crypts. Horizontal crypts oriented parallel to the muscularis mucosae along the base were classified as none, rare (<3 crypts), focal (3 crypts, or 33% of crypts), or prominent (> 33% of crypts). Inverted crypts (so-called misplaced crypts) below the muscularis mucosae were classified as present or absent. Crypt branching was classified as none, fewer than 33% of crypts, 33% to 67% of crypts, or more than 67% of crypts. Crypt Epithelial Cell Features Features pertaining to the superficial or upper one third of the crypts were evaluated only in intracryptal cells that had not breached the ostium. Surface epithelial cells were excluded from the evaluation. Mitoses were counted separately in the basilar, middle, and superficial crypt regions. Ten crypts were counted and reported as the mean mitotic index per crypt. The predominant nuclear shape in the basilar, middle, and superficial or upper third sections of the crypts were classified as round to oval, columnar or elongate, flattened, or mixed. The prominence of nucleoli in the cells of the basilar, middle, and superficial crypt regions was classified as inconspicuous, average, or prominent or eosinophilic. Goblet cells present in the crypt bases were classified as present or absent. Distended, mucin-filled cells in the superficial crypts were classified as present or absent. Cells with abundant eosinophilic cytoplasm in the superficial crypts were classified as present or absent. Cryptal Maturation Features The changes in nuclear maturation, cell maturation, and percentage of dysmaturational crypts were evaluated along the length of the crypt, base to ostium. Nuclear maturation was defined as decreased nuclear size; lighter stained, finer grained, and more homogeneous chromatin; and increased uniformity of nuclear shape and Downloaded from Am J Clin Pathol 2003;119:

3 Goldstein et al / HYPERPLASTIC-LIKE COLON POLYPS PRECEDING MICROSATELLITE-UNSTABLE ADENOCARCINOMAS chromatin among the adjacent cells. Nuclear maturation was classified as present in no crypts, 1% to 10% of the crypts, 11% to 50% of the crypts, 51% to 90% of the crypts, or maturation in more than 90% of the crypts. Cell maturation was defined as increased amounts of lightly eosinophilic or vacuolated, mucin-filled cytoplasm and decreased nuclear/cytoplasmic ratio from nuclear contraction and increased cytoplasm. Cell maturation was classified as present or absent. The percentage of dysmaturational crypts was identified from the conglomerate of architectural, nuclear, and cytologic features. Dysmaturational crypts had basilar crypt dilation, a serrated architecture that started in the basilar region, and a greater number of superficial crypt cells with predominantly round to oval nuclei and average or prominent nucleoli. The amount of nuclear and cell maturation along dysmaturational crypts was minimal to none. The percentage of dysmaturational crypts was classified as none, 1% to 10%, 11% to 50%, 51% to 90%, and more than 90%. Ancillary Studies, Including Immunohistochemical Analysis The loss of hmlh1 nuclear immunoreactivity in crypt epithelial cells was classified as no loss or decreased or absent staining in the basilar, middle, or superficial crypt region. The mean nuclear area of cells in the basilar, middle, and superficial crypt regions was determined in 10 cells from each crypt region in 10 well-oriented crypts from each polyp that were chosen randomly and outlined using a CAS 200 Image Analyzer (Becton Dickinson, Elmhurst, IL). For each region, 100 cells were measured. hmlh1 immunohistochemical staining was optimized. 74 We applied 4-µm-thick tissue sections to charged slides, and slides were dried for 45 minutes in a 60 C oven. Slides were immersed in a 1-mmol/L concentration of EDTA solution (ph 8), preheated to 95 C, in a commercial vegetable steamer for 30 minutes. The hmlh1 antibody (clone, G ; dilution, 1:250; Pharmingen, San Diego, CA) was incubated over the slides for 30 minutes using an autostainer (DAKO, Carpinteria, CA). The initial chromogen system was the Envision+ polymer detection system (DAKO). We switched to the 2-step AP900-9M supersensitive detection system (BioGenex, Table 1 Study Group Polyps and Subsequent Adenocarcinoma Polyp-Adenocarcinoma Interval 10 y 6 to <10 y 3 to <6 y <3 y No. of polyps Mean polyp dimension (mm) San Ramon, CA) midway through the study because it was determined to produce stronger nuclear reactivity and less cytoplasmic nonspecific granular staining. The chromogen in both systems was diaminobenzidine and hematoxylin counterstain. The Fisher exact (2-tailed) and χ 2 tests were used to compare the means of categorical variables, and logistic and linear regressions were used to compare continuous variables (Systat computer program, version 10, SPSS, Chicago, IL). Results Clinical Study Group Polyps The mean and median patient ages were 68.2 and 69.5 years, respectively (range, years; SD, 6.1 years). Of the 91 patients, 58 (64%) were women. The polyps and subsequent adenocarcinomas were located in the cecum in 21 patients (23%), the proximal ascending colon in 44 patients (48%), the distal ascending colon in 20 patients (22%), the proximal transverse colon in 5 patients (5%), and the distal transverse colon in 1 patient (1%). The mean and median intervals between polypectomy and subsequent adenocarcinoma were 7.3 and 6.5 years, respectively (range, years; SD, 4.4 years). The endoscopist noted multiple polyps in a close geographic distribution in 47 patients (52%). Of the 106 polyps, 34 (32.1%) were biopsied but not resected. Polyps resected 10 or more years before colectomy had a mean maximum dimension of 4.8 mm compared with a mean of 7.5 mm for polyps resected less than 3 years before the adenocarcinoma Table 1. A shorter interval between polypectomy and adenocarcinoma was associated significantly with a larger mean maximum polyp dimension (P <.01). Larger polyp dimension was associated significantly with increasing patient age (P =.023). Of the 91 patients, 15 (16%) had 2 metachronous hyperplastic-like polyps, all located in the proximal ascending colon. The mean interval between the first polypectomy and subsequent adenocarcinoma among these 15 patients was 11.9 years (range, years; SD, 3.2 years). The mean interval between the 2 polyps was 6.3 years (range, years; SD, 2.0 years). The second polyp was larger than the initial polyp in 14 (93%) of 15 patients. The mean increase in maximum dimension between the 2 polyps was 5.5 mm (range, 1-10 mm). Excluding the patient with a smaller second polyp, the mean increase in the maximum dimension of the second polyp was 0.95 mm per interval year (range, mm; SD, 0.6 mm). 780 Am J Clin Pathol 2003;119: Downloaded 780 from

4 Anatomic Pathology / ORIGINAL ARTICLE Control Group Polyps The mean and median ages at polypectomy were 64.1 and 64.0 years, respectively (range, years; SD, 5.4 years). Of the 106 patients, 24 (22.6%) were women. The mean and median follow-up intervals during which no adenocarcinoma developed was 16.5 years (range, years; SD, 0.87 years). Morphologic Features Architecture Study group polyps Image 1, Image 2, Image 3, Image 4, and Image 5 had a significantly larger mean dimension than control group polyps Image 6, Image 7, and Image 8 (5.9 vs 4.3 mm; P <.01) Table 2. A significantly greater percentage of crypts in study group polyps had serration that began in the basilar region and crypt basilar dilation than did control group polyps (Table 2). These features were present in study group polyps of all dimensions, including those with a dimension of 5 mm Table 3. Inverted crypts were seen only in study group polyps. Crypt Epithelial Cells The mean mitotic indices in the basilar crypt regions of both polyp groups were similar Table 4. They were significantly greater in the middle and superficial crypt regions of study group polyps than in the control group polyps. The largest difference between polyp groups (0.77) was in the middle crypt region. Mitoses were extremely uncommon in the superficial crypt regions of control group polyps. Of the Table 2 Architectural Features * Polyp Group Feature Study (n = 106) Control (n = 106) P Mean/median polyp size (range; SD), mm 5.9/5.0 ( ; 3.1) 4.3/4.0 ( ; 1.5) <.01 Mean/median collagen band thickness (range; SD), µm 1.8/1.7 ( ; 0.5) 4.6/4.7 ( ; 2.5).019 Inverted crypts subjacent to muscularis mucosae 28 (26.4) 0 (0.0) <.01 Serration began in basilar crypt region 82 (77.4) 13 (12.3) <.01 Basilar crypt dilation None 1 (0.9) 79 (74.5) 1-5 crypts 21 (19.8) 25 (23.6) 10%-33% of crypts 51 (48.1) 2 (1.9) >33% of crypts 33 (31.1) 0 (0.0) 10% of crypts 84 (79.2) 2 (1.9) <.01 Horizontal crypts.39 None 69 (65.1) 98 (92.5) Rare 29 (27.4) 8 (7.5) Focal 8 (7.5) 0 (0.0) Extensive 0 (0.0) 0 (0.0) Branched crypts.14 None 81 (76.4) 105 (99.1) <33% of crypts 25 (23.6) 1 (0.9) 33%-67% of crypts 0 (0.0) 0 (0.0) >67% of crypts 0 (0.0) 0 (0.0) * Data are given as number (percentage) unless otherwise indicated. Table 3 Architectural Features and Polyp Dimension * Polyp Dimension (mm) Feature/Group >5.0 Serrated architecture started in basilar crypt region Study 5/6 (83) 16/21 (76) 9/15 (60) 11/14 (79) 41/56 (73) 50/50 (100) Control 2/8 (25) 0/29 (0) 4/30 (13) 4/17 (24) 10/84 (12) 3/22 (14) Basilar crypt dilation Of rare crypts Study 4/6 (67) 10/21 (48) 7/15 (47) 0/14 (0) 21/56 (38) 0/50 (0) Control 6/8 (75) 4/29 (14) 7/30 (23) 5/17 (29) 22/84 (26) 3/22 (14) Of >10% of crypts Study 1/6 (17) 11/21 (52) 8/15 (53) 14/14 (100) 34/56 (61) 50/50 (100) Control 0/6 (0) 0/29 (0) 0/30 (0) 0/17 (0) 0/84 (0) 2/22 (9) * Data are given as number/total number in category (percentage). Downloaded from Am J Clin Pathol 2003;119:

5 Goldstein et al / HYPERPLASTIC-LIKE COLON POLYPS PRECEDING MICROSATELLITE-UNSTABLE ADENOCARCINOMAS A B C D E Image 1 Study group polyp. A, Two centrally located crypts have dilated bases. The serrated architecture begins in the deep basal regions of the crypts (H&E, 24). B, Mitoses are present at the middle crypt level. Three of the 4 crypts are dysmaturational. Maturation is present in the second crypt from the left (H&E, 64). C, Dilated crypt bases with proliferative-zone cells (H&E, 640). D, Middle crypt region. Cells are similar to basilar cells without maturational changes (H&E, 640). E, Superficial crypt region. Cells display some maturational changes, including increased mucus, decreased nuclear size, and less prominent nucleoli. However, nuclei are larger than nuclei in superficial crypt cells of control group polyps (Images 6D, 7C, and 8C) (H&E, 640). 782 Am J Clin Pathol 2003;119: Downloaded 782 from

6 Anatomic Pathology / ORIGINAL ARTICLE A B C D E F Image 2 Study group polyp. A, A 0.4-cm polyp. Distinction from a control group polyp is difficult at low magnification (H&E, 32). B, The majority of crypts are dysmaturational with dilated bases. Serrated outline begins in the basilar region of most crypts (H&E, 160). C, Dilated crypt bases with proliferative zone epithelial cells (H&E, 480). D, Lower middle crypt region. Mitoses are present in the middle crypt region. The cells are similar to those of the proliferative zone; significant maturational changes are absent (H&E, 480). E, Upper middle crypt and superficial crypt regions. Although the amount of mucinous cytoplasm has increased, most of the nuclei show minimal maturation and remain similar to those of the basilar proliferative crypt region. Substantial nuclear maturation is apparent in the far right side of the slide, in the superficial crypt region (H&E, 480). F, Superficial crypt region. The transition of nuclear maturation is apparent in the left-sided region of the lower crypt. The nuclei of the cells in the superficial crypt region are larger in study group polyps than in control group polyps (Images 6D, 7C, and 8C) (H&E, 480). Downloaded from Am J Clin Pathol 2003;119:

7 Goldstein et al / HYPERPLASTIC-LIKE COLON POLYPS PRECEDING MICROSATELLITE-UNSTABLE ADENOCARCINOMAS A B C D E Image 3 Study group polyp. A, Portion of a 1.5-cm polyp. Thickened, serrated mucosa produces a raised villiform lesion. The transition to normal mucosa is abrupt (H&E, 24). B, The majority of crypts are dysmaturational, and many have dilated bases (H&E, 64). C, Serration begins immediately in the deep, basilar crypt region (H&E, 64). D, Dilated crypt base lined by proliferative zone cells. Many cells have abundant mucus, which is a feature of dysmaturation (H&E, 640). E, Superficial crypt region. Many cells have too much cytoplasmic mucin producing redundant-appearing amounts of cytoplasm. The nuclei are slightly but not significantly smaller than the cells of the basilar proliferative zone (H&E, 640). 784 Am J Clin Pathol 2003;119: Downloaded 784 from

8 Anatomic Pathology / ORIGINAL ARTICLE A B C D Image 4 Study group polyp (serrated adenoma?). A, Focus of inverted crypts, located subjacent to the muscularis propria (H&E, 24). B, Pushing pattern of inverted crypts below the muscularis propria. Inverted regions usually were subjacent to crypts with basilar dilation (H&E, 64). C, Basilar crypt region with numerous mitoses (H&E, 640). D, Superficial crypt region. Epithelial cells have increased cytoplasm and slightly smaller nuclei than cells in the basilar region. However, the amount of nuclear size diminution is less than encountered in hyperplastic polyps (H&E, 640). 106 study group polyps, 104 (98.1%) had predominantly round to oval nuclei in the middle crypt region compared with 38 control group polyps (35.8%). The nuclear shape remained round to oval in the middle crypt regions of study group polyps followed by a rapid change in nuclear shape to columnar or flat in the superficial crypt region. Nucleolar prominence showed similar, significant differences for the middle and superficial crypt regions in the 2 polyp groups. Maturational Changes Along Crypts A significantly greater percentage of crypts in study group polyps had minimal nuclear maturation along their lengths Table 5. Crypts with minimal nuclear maturation were the majority in 105 study group polyps (99.1%) compared with 22 control group polyps (20.8%) (P <.001). Crypts with minimal nuclear maturation constituted more than 90% of crypts in 92 study group polyps (86.8%) but only 1 control group polyp (0.9%) (P <.001). These differences were present in study and control group polyps of all dimensions Table 6. The majority of polyp crypts were dysmaturational in all 106 study group polyps (100.0%; Table 5). Conversely, 79 control group polyps (74.5%) had no or rare dysmaturational crypts. Dysmaturational crypts were the slight majority in 36 study group polyps (34.0%) and the overwhelming majority Downloaded from Am J Clin Pathol 2003;119:

9 Goldstein et al / HYPERPLASTIC-LIKE COLON POLYPS PRECEDING MICROSATELLITE-UNSTABLE ADENOCARCINOMAS A B C Image 5 Study group polyp. A, Diminutive polyp with focus of inverted crypts and extensive crypt basilar dilation (H&E, 32). B, Inverted crypts have a stellate, expansive growth pattern below the muscularis mucosae (H&E, 64). C, Majority of crypts are dysmaturational (H&E, 64). in the other 70 (66.0%), whereas they were the overwhelming majority in only 14 control group polyps (13.2%) (P =.012). The percentage of dysmaturational crypts increased in polyps of larger dimension in both study and control group polyps (Table 6). Ancillary Studies Loss of hmlh1 immunoreactivity occurred almost exclusively in cells of dysmaturational crypts in a superficial crypt to basilar crypt distribution Table 7. Eighty-six study group polyps (81.1%) had loss of immunoreactivity in the superficial and middle crypt regions or near absent staining except for a few basilar crypt cells. The mean nuclear areas of cells in the basilar and middle crypt regions were similar in study and control group polyps. The mean nuclear area in the superficial crypt region of study group polyps was 7.8 µm 2 smaller than the mean nuclear area of basilar crypt cells, supporting the minimal maturation changes observed over the length of crypts. The mean nuclear area of cells in the superficial crypt region in control group polyps was 13.1 µm 2, which was a significant difference (P <.01). Reevaluation of Control Group Polyps Control group polyps were reviewed and reclassified using the features identified as being characteristic of study group polyps to reclassify the lesions. Although this line of investigation was circular, it functioned as a surrogate indicator of the specificity of the morphologic features identified in the previous phase of the study. The inclusion criteria of hyperplastic polyps as control group lesions were based on the polyp s location in the colon and the lack of a subsequent colorectal adenocarcinoma; it was possible that some of the control group lesions would be reclassified as study group type hyperplastic polyps. 786 Am J Clin Pathol 2003;119: Downloaded 786 from

10 Anatomic Pathology / ORIGINAL ARTICLE A B C D Image 6 Control group polyp. A, Serrated architecture becomes apparent in upper middle and superficial crypt regions (H&E, 32). B, Crypt bases are uniformly small, rounded, and undilated. Serrated architecture becomes apparent in the superficial crypt region (H&E, 64). C, Basilar crypt region, proliferative zone cells with occasional mitoses. Cell maturational changes become apparent in the basilar crypt region (H&E, 480). D, Superficial crypt region. Cells have abundant cytoplasm. Nuclei are substantially smaller than those of the basilar region, and nucleoli are inconspicuous. Compared with the cells of the superficial crypt region of study group polyps (Images 1E, 2F, 3E, 4D), the nuclei of control group are smaller and there are greater maturational changes (H&E, 480). Twenty-four control group polyps (22.6%) had the constellation of morphologic features characteristic of study group polyps Table 8. These 24 lesions were distinctly similar to study group polyps and were unlike any other control group polyps. When these 24 polyps were removed from the control group, the remaining 82 polyps were extremely homogeneous and distinctly different from study group polyps. The superficial crypt mitotic index of the 24 study group like control group polyps was 0.83 per crypt, which was similar to the value of 0.86 in the study group polyps and substantially greater than the 0.03 value among the 82 remaining control group polyps. The 24 study group polyps accounted for all of the control group polyps that had predominantly dysmaturational crypts. Discussion The hyperplastic-like polyps that preceded microsatelliteunstable adenocarcinomas had expanded crypt proliferation zones, crypt basilar dilation, a serrated crypt architecture that Downloaded from Am J Clin Pathol 2003;119:

11 Goldstein et al / HYPERPLASTIC-LIKE COLON POLYPS PRECEDING MICROSATELLITE-UNSTABLE ADENOCARCINOMAS A B C Image 7 Control group polyp. A, Slight crypt disarray and superficial crypt region dilation simulates dysmaturational crypts (H&E, 32). B, Crypt bases are tapered and small. Serrated crypt architectural outline becomes apparent in the middle crypt region. Cell maturation is apparent within the basilar crypt region (H&E, 64). C, Cells in the superficial crypt region have small, uniform nuclei that are smaller than the cells of the superficial crypt region of study group polyps (Images 1E, 2F, 3E, 4D) (H&E, 480). became apparent in the basilar region, and decreased or absent cell maturation along the length of the crypt. Alterations in the extent (persistence) and location of immature epithelial cells and an expanded proliferation crypt zone combined to form a distinctive appearing process that we term dysmaturation. Dysmaturation and altered architecture were detected easily at medium magnifications and were the features that distinguished between study and control group polyps. Hyperplastic-like serrated study group polyps had an expanded zone of crypt epithelial cell proliferation, a serrated architectural outline that became apparent in the basilar regions of crypts, and basilar dilation of a substantial minority or majority of crypts, whereas the proliferation zone was confined to the most basilar crypt region, serration became apparent in the middle crypt region, and only rare crypts had basal dilation in control group polyps, consistent with previous findings. 14,31,35,39,40,75-77 Many study group polyps were larger than 5 mm, whereas almost all control group polyps were smaller than 5 mm, as in previous studies. 14,16,28,29,75,78,79 Some study group polyps also had inverted crypts subjacent to the muscularis mucosae or horizontally oriented crypts, whereas they were not features of control group polyps, consistent with the findings of Torlakovic and Snover. 14 As in previous studies, 31,35 the majority of crypts in hyperplastic-like serrated study group polyps were dysmaturational, whereas dysmaturational crypts were absent or rare in control group polyps. The homogeneity of cells along the lengths of dysmaturational crypts was borne out by minimally smaller mean nuclear areas in the superficial crypt regions compared with basilar regions in study group polyps, whereas it was significantly smaller in superficial crypt regions of control group polyps. Our results are similar to and support the recent findings of Torlakovic et al. 80 These authors studied 289 serrated 788 Am J Clin Pathol 2003;119: Downloaded 788 from

12 Anatomic Pathology / ORIGINAL ARTICLE A B C Image 8 Control group polyp. A, One crypt in the left side of the photograph has basilar dilation. Serrated architectural outline becomes apparent in the superficial crypt region (H&E, 64). B, Basilar region, proliferative zone cells. Maturational changes are apparent within the basilar crypt region (H&E, 480). C, Superficial region epithelial cells have stratified nuclei that vary in shape between cuboidal and columnar. Their size is similar to the upper basilar region cell nuclei rather than the lower basilar region proliferative zone cell nuclei (H&E, 480). polyps and identified a constellation of morphologic changes, including abnormal proliferation, crypt distortion and dilatation, and decreased numbers of endocrine cells. The features these authors collectively termed abnormal proliferation are identical to the features we termed dysmaturational crypts. Serrated polyps with abnormal proliferation constituted 18% of the studied polyps in the study by Torlakovic et al, 80 which is similar to the 22.6% of control group polyps in our study. The results of the present study differ slightly from those of other authors. One study that compared serrated adenomas (with and without adenomatous epithelium) from 6 patients with hyperplastic polyposis syndrome and control hyperplastic polyps found that most of the serrated adenomas had numerous areas with horizontally oriented crypts. 14 Rare horizontally oriented crypts were present in a high percentage of study group polyps in the present study, but in only 8 (7.5%) were they present focally. This difference may be a function of polyp size. Most of the polyps in the aforementioned study were large (mean, 1.2 cm), and all of the study group polyps in the present study with focal horizontal crypts were larger than 8 mm. The other architectural features of study group polyps in the present study were not influenced by polyp dimension. The prevalence of polyps containing cells with abundant eosinophilic cytoplasm or distended, mucin-filled goblet cells in the superficial crypt regions in our study also was lower than reported by other authors. Some authors found that these cytoplasmic features were characteristic of most serrated adenomas. 14,17,35 The conceptual and morphologic road that led to the concept that some nonadenomatous hyperplastic-like polyps are preneoplastic precursor lesions is convoluted. Numerous small studies described or photographed hyperplastic-like lesions as an unusual change in hyperplastic polyps, 11,81-85 as Downloaded from Am J Clin Pathol 2003;119:

13 Goldstein et al / HYPERPLASTIC-LIKE COLON POLYPS PRECEDING MICROSATELLITE-UNSTABLE ADENOCARCINOMAS Table 4 Crypt Epithelial Cells * Polyp Group Feature Study (n = 106) Control (n = 106) P Mean mitotic index/crypt Basilar crypt region Middle crypt region <.01 Superficial crypt region <.01 Predominant nuclear shape Middle crypt region.032 Round to oval 104 (98.1) 38 (35.8) Mixed 2 (1.9) 2 (1.9) Columnar or flat 0 (0.0) 11 (10.4) Columnar or flat with 3 crypts with round to oval nuclei 0 (0.0) 55 (51.9) Superficial crypt region <.01 Round to oval 91 (85.8) 1 (0.9) Mixed 15 (14.2) 43 (40.6) Columnar or flat 0 (0.0) 39 (36.8) Columnar or flat with 3 crypts with round to oval nuclei 0 (0.0) 23 (21.7) Nucleolar prominence Basilar crypt region.46 Prominent 81 (76.4) 67 (63.2) Average 25 (23.6) 39 (36.8) Inconspicuous 0 (0.0) 0 (0.0) Middle crypt region <.01 Prominent 62 (58.5) 4 (3.8) Average 44 (41.5) 87 (82.0) Inconspicuous 0 (0.0) 15 (14.2) Superficial region <.01 Prominent and eosinophilic 36 (34.0) 1 (0.9) Average 68 (64.2) 44 (41.5) Inconspicuous 2 (1.9) 61 (57.5) Goblet cells in crypt bases 44 (41.5) 22 (20.8).75 Distended, mucin-filled goblet cells in superficial crypt region 39 (36.8) 4 (3.8).058 Cells with abundant eosinophilic cytoplasm in superficial crypt region 45 (42.5) 12 (11.3).084 * Data are given as number (percentage) unless otherwise indicated. part of mixed hyperplastic-adenomatous polyps, 18,81,82,85-91 or as lesions adjacent to adenocarcinomas The term serrated was first applied by Longacre and Fenoglio- Preiser 17 to architecturally serrated, mixed, adenomatous, and hyperplastic-like polyps. All of the polyps in their study had areas of adenomatous epithelium. Study of precursor polyps in patients with hereditary nonpolyposis colon cancer and hyperplastic polyposis syndromes expanded the definition of serrated adenomas to include nonadenomatous hyperplastic-like polyps. 13,14,17-19,22,24,35,41,42,44,45,47-49 Concomitant with this expanded definition of the serrated adenoma, Jass proposed the serrated neoplasia pathway concept of colorectal adenocarcinoma in nonsyndromic patients. This pathway is mechanistically distinct from the model proposed by Vogelstein et al 97 of a tubular adenoma adenocarcinoma tumor-suppressor gene pathway. 24,42-44,47,48,98,99 Evidence that supports serrated adenomas as mechanistically different from tubular adenomas includes the location of cells with the greatest dysplasia, location of crypt proliferative zones, prevalence of K-ras, p53, and APC gene mutations and MUC mucoprotein expression profiles. 15,21,27,40,77, The driving molecular mechanism in the serrated neoplasia pathway is the mutator, or mismatch repair deficiency phenomenon. Inactivation of the MLH1 gene of the DNA mismatch repair complex produces a progressively greater number of genome-wide mutations that eventuates in microsatellite-unstable colorectal adenocarcinomas. There seem to be several possibly related processes contributing to MLH1 promoter gene inactivation that leads to down-regulation of MLH1 gene expression, including silencing of the DNA repair enzyme O-6-methylguanine-DNA methyltransferase 20,106 and the CpG island methylator phenotype of the promoter region. 32,37,59,61,62,65, The serrated neoplasia pathway model included all serrated polyps. Iino et al, 24 Jass et al, 44 and Jass 48 suggested that hyperplastic-type polyps were the initial lesion in the serrated neoplasia pathway continuum and linked hyperplastic polyps with microsatellite-unstable adenocarcinomas. They also suggested that the idea that all hyperplastic polyps were innocent, benign, lesions was erroneous. 24,44,48 Other authors added their support to this concept; hyperplastic polyps and serrated adenomas are a single entity at different points of a histologic and molecular continuum of increased 790 Am J Clin Pathol 2003;119: Downloaded 790 from

14 Anatomic Pathology / ORIGINAL ARTICLE Table 5 Crypt Maturation Features * Polyp Group Features Study (n = 106) Control (n = 106) P Minimal epithelial cell nuclear maturation (percentage of crypts) 0-10 (>90% normal) 0 (0.0) 41 (38.7) (0.9) 43 (40.6) (12.3) 21 (19.8) < (55.7) 1 (0.9) 100 (0% normal) 33 (31.1) 0 (0.0) >90% (<10% normal) 92 (86.8) 1 (0.9) <.001 Dysmaturational crypts (%) 0 0 (0.0) 27 (25.5) (0.0) 52 (49.1) (0.0) 3 (2.8) (34.0) 10 (9.4) (47.2) 14 (13.2) (18.9) 0 (0.0) > (100.0) 24 (22.6).012 * Data are given as number (percentage). Table 6 Crypt Epithelial Cell Maturation and Polyp Dimension * Polyp Dimension (mm) Feature 4 5 to 8 >8 Minimal nuclear maturation in >90% of crypts Study group 40/42 (95) 39/46 (85) 13/18 (72) Control group 1/ 67 (1) 0/39 (0) 0 (0) Dysmaturational crypts (%) In study group polyps /42 (36) 16/46 (35) 5/18 (28) >90 27/42 (64) 30/46 (65) 13/18 (72) In control group polyps 0 27/67 (40) 0 (0) 0 (0) /67 (34) 7/39 (18) 0 (0) /67 (7) 20/39 (51) 0 (0) /67 (7) 5/39 (13) 0 (0) >90 7/67 (10) 7/39 (18) 0 (0) * Data are given as number (percentage). mutations, progressive loss of controlled cell function including proliferation and maturation, and clonal expansion of cells with abnormal maturation within hyperplastic polyps to produce serrated adenomas. 22,23,34,79,100,111 Evidence linking hyperplastic polyps and serrated adenomas comes from many avenues. Variable amounts of hyperplastic polyp, serrated adenoma, and villous adenoma typically are seen in larger polyps in the serrated neoplasia pathway. Identical clonal alterations have been identified in the hyperplastic and serrated adenoma components of mixed polyps. 24,34,112 The lesions have similar rates of genetic mutations, distributions and patterns of cell proliferation, gene mutations, levels of DNA microsatellite instability, patterns of absent hmlh1 expression in crypt cells, apoptotic dysregulation, cell antigen, and MUC mucoprotein expression profiles. 24,27,29,39,44,45,76,99,100,112,113 We also found that study group polyps had many crypts with absent hmlh1 nuclear immunoreactivity. 32,44-46,114 The present study provides morphologic criteria to identify the subgroup of serrated neoplastic polyps that preceded microsatellite-unstable colon adenocarcinomas and, therefore, may be true premalignant neoplasms. In our opinion, these results support the concept of Jass that some hyperplastic-like polyps are the initial lesion in the serrated neoplasia pathway. Progressive accumulation of mutations throughout the genome from altered mismatch repair function may result in a progressive decrease in the cell function, resulting in the morphologic change of dysmaturational crypts. In this vein, dysmaturation, similar to mutational events and progression along the serrated neoplasia pathway, constitutes a range of morphologic alterations, some of which overlap with incidental-type, innocuous hyperplastic polyps. Downloaded from Am J Clin Pathol 2003;119:

15 Goldstein et al / HYPERPLASTIC-LIKE COLON POLYPS PRECEDING MICROSATELLITE-UNSTABLE ADENOCARCINOMAS Table 7 hmlh1 Immunohistochemical and Image Morphometric Analysis Polyp Groups Feature Study (n = 106) Control (n = 106) P hmlh1 nuclear immunoreactivity in dysmaturational crypts * Entire crypt 11 (10.4) 67 (63.2) Basilar and middle crypt regions 38 (35.8) 8 (7.5).68 Basilar crypt region 28 (26.4) 15 (14.2) Nonreactive crypts 29 (27.4) 16 (15.1) Crypt epithelial cell mean nuclear area (µm 2 ) Basilar crypt region Middle crypt region Superficial crypt region <.01 * Data are given as number (percentage). Table 8 Reevaluation of Control Group Polyps * Control Group Polyps Feature Control Group like Study Group like Study Group Polyps No. of polyps 82 (77.3%) 24 (22.6%) 106 (100.0) Mean polyp size (mm) Mean collagen band thickness (µm) Superficial crypt region mitotic index Serrated architecture starts in basilar crypt region 0 (0.0) 13 (12.3) 82 (77.4) Dilation of 10% of crypt bases 0 (0.0) 2 (1.9) 84 (79.2) No epithelial cell nuclear maturation in >50% of crypts 0 (0.0) 22 (20.8) 105 (99.1) >50% dysmaturational crypts 0 (0.0) 24 (22.6) 106 (100.0) Decreased or absent hmlh1 nuclear reactivity in 0 (0.0) 16 (15.1) 29 (27.4) dysmaturational crypt epithelial cells of basilar, middle, and superficial crypt regions * Data are given as number (percentage) unless otherwise indicated. Cellular mechanisms that result in a serrated architecture are topics of active investigation. Jass et al 44 suggested that serrated architecture may not be a fundamental feature of the mutator pathway, but due to an accommodation of increased epithelial volume due to the combination of hyperplasia and cytoplasmic maturation. This may be partially correct. Affected basilar crypt epithelial cells in serrated adenomas proliferate and grow outward rather than upward toward the luminal surface and at a pace that is greater than admixed, normal epithelial cells and the accompanying crypt-related stroma. 115 Expansion by these aberrantly growing cells seems to be facilitated by decreased apoptosis, which leads to decreased surface epithelial cell loss (anoikis). 39,116 Outward proliferation without migration may be the cause of basilar crypt dilation and inverted crypt growth in serrated adenomas. Hyperproliferation compared with the adjacent stroma in this subgroup of crypt epithelial cells may explain the immediate basilar appearance of a serrated architectural outline. All study group polyps in the present study, including those smaller than 4 mm, had the constellation of characteristic morphologic features described herein. The diagnostic sensitivity of these features for identifying study group polyps seems high. However, their specificity was low in our first evaluation because 24 control group hyperplastic polyps (22.6%) also had these features. When these 24 polyps were excluded from the control group, the remaining control group polyps had repetitively similar morphologic features that were distinctly different from study group polyps and the other 24 control group polyps. This suggests these 24 control group polyps were left-sided study group type polyps that arose in the rectum of patients in whom adenocarcinoma did not develop subsequently. The sigmoid and rectum have been the most common sites for serrated adenomas in some studies. 17,29,99,117 The results of the present study support the opinions of previous authors about 2 types of hyperplastic-like colorectal polyps. 29,32,41,44,49,78 Torlakovic and Snover 14 made these same suggestions in a notably insightful study in One type is small, occurs predominantly in the sigmoid and rectum, is composed of thin, nondilated crypts with a serrated architectural outline that becomes apparent at the middle crypt level, and has no or rare individual dysmaturational crypts; this type seems to have no significant associated risk of progression to adenocarcinoma. The second type 792 Am J Clin Pathol 2003;119: Downloaded 792 from

16 Anatomic Pathology / ORIGINAL ARTICLE of hyperplastic-like polyp has the characteristic morphologic features of serrated adenoma listed previously, may arise in patients with a family history of a colorectal adenocarcinoma, and often is large and located in the cecum or ascending colon. This group has a risk of progression to adenocarcinoma. Small dimension or polyp location should not automatically exclude the diagnosis of serrated adenoma. A large percentage of the acquired genetic mutations necessary for progression occurs in serrated adenomas that are smaller than most of the polyps detected by endoscopy. 111 Small hyperplastic-like serrated adenomas in nonsyndromic patients have a progression risk that is similar to that for tubular adenomas. 75 Progression to adenocarcinoma may be rapid once the necessary mutations have been acquired, which may be unrelated to the presence of adenomatous epithelium. This phenomenon may explain many of the socalled interval or missed adenocarcinomas in which a recent preceding colonoscopy failed to detect the adenocarcinoma. 111,118 We, similar to other authors, believe that polyps in the serrated neoplasia pathway require complete eradication and enrollment of patients into a colonoscopic surveillance program. 44,50-56,78 The optimal term for polyps in the serrated neoplasia pathway is unclear, especially the subgroup that is devoid of adenomatous epithelium and is the focus of the present study. One option is to continue to name all nonadenomatous, hyperplastic-like, serrated polyps as hyperplastic polyps. Some authors have maintained the original study criterion that adenomatous epithelium must be present for the diagnosis of serrated adenoma, 38,86,119 calling polyps hyperplastic in its absence. 11,38,120 This diagnostic approach may cause harm to patients because it does not recognize the premalignant nature of the hyperplastic-like polyp in the serrated neoplasia pathway. The treating physician may not comprehend the need for complete polyp eradication, and the patient may not be counseled to undergo colonoscopy at a future time. In addition, serrated neoplasia polyps may have nonadenomatous, high-grade dysplastic epithelial cells within the deep regions of crypts that may evolve into adenocarcinoma in the absence of an adenomatous epithelial component. A second option is to term all polyps in the serrated neoplasia pathway, including those without adenomatous epithelium, as serrated adenomas. The advantage of this approach is that the premalignant nature of the lesion would be identified, and most treating physicians are cognizant of the appropriate therapy and follow-up parameters surrounding an adenoma diagnosis. The disadvantage of this approach is that it continues to perpetuate the untruth that only adenomatous polyps are premalignant. Torlakovic et al 80 recently recommended that this group of polyps be termed sessile serrated adenoma. We support their recommendation. It should not be inferred from the present discussion that all hyperplastic polyps are thought to be premalignant precursor lesions. 49 The numerous studies of preceding decades clearly show the benign, nonprogressive nature of most hyperplastic polyps ,56 Future studies will be needed to examine the questions engendered by the hyperplastic-serrated polyp precursor lesions of the serrated neoplasia pathway. The initial necessary step of examining the issues pertaining to whether some hyperplastic polyps are premalignant lesions is the establishment of morphologic or other criteria that can identify the subsets of polyps deserving additional study. From the Department of Anatomic Pathology, William Beaumont Hospital, Royal Oak, MI. Address reprint requests to Dr Goldstein: Dept of Anatomic Pathology, William Beaumont Hospital, 3601 W Thirteen Mile Rd, Royal Oak, MI Acknowledgments: We thank Ken Batts, MD, for reviewing the manuscript and for insightful comments. hmlh1 immunohistochemical staining was optimized with the assistance of Larry Burgart, MD, Pat Roche, PhD, Linda Murphy, and Barbara Crawford, Mayo Clinic, Rochester, MN. This article honors Klaus Lewin, MD, mentor to and teacher of many pathologists. His personage and ideas flourish in his students. References 1. Aaltonen LA, Peltomaki P, Mecklin JP, et al. Replication errors in benign and malignant tumors from hereditary nonpolyposis colorectal cancer patients. Cancer Res. 1994;54: Kim H, Jen J, Vogelstein B, et al. Clinical and pathological characteristics of sporadic colorectal carcinomas with DNA replication errors in microsatellite sequences. Am J Pathol. 1994;145: Cawkwell L, Li D, Lewis FA, et al. Microsatellite instability in colorectal cancer: improved assessment using fluorescent polymerase chain reaction. Gastroenterology. 1995;109: Liu B, Nicolaides NC, Markowitz S, et al. Mismatch repair gene defects in sporadic colorectal cancers with microsatellite instability. Nat Genet. 1995;9: Moslein G, Tester DJ, Lindor NM, et al. Microsatellite instability and mutation analysis of hmsh2 and hmlh1 in patients with sporadic, familial and hereditary colorectal cancer. Hum Mol Genet. 1996;5: Thibodeau SN, French AJ, Roche PC, et al. Altered expression of hmsh2 and hmlh1 in tumors with microsatellite instability and genetic alterations in mismatch repair genes. Cancer Res. 1996;56: Wu Y, Nystrom-Lahti M, Osinga J, et al. MSH2 and MLH1 mutations in sporadic replication error positive colorectal carcinoma as assessed by two-dimensional DNA electrophoresis. Genes Chromosomes Cancer. 1997;18: Halling KC, French AJ, McDonnell SK, et al. Microsatellite instability and 8p allelic imbalance in stage B2 and C colorectal adenocarcinomas. J Natl Cancer Inst. 1999;91: Downloaded from Am J Clin Pathol 2003;119:

17 Goldstein et al / HYPERPLASTIC-LIKE COLON POLYPS PRECEDING MICROSATELLITE-UNSTABLE ADENOCARCINOMAS 9. Jernvall P, Makinen MJ, Karttunen TJ, et al. Microsatellite instability: impact on cancer progression in proximal and distal colorectal cancers. Eur J Cancer. 1999;35: Fucci L, Pirrelli M, Caruso ML. Carcinoma and synchronous hyperplastic polyps of the large bowel. Pathologica. 1994;86: Warner AS, Glick ME, Fogt F. Multiple large hyperplastic polyps of the colon coincident with adenocarcinoma. Am J Gastroenterol. 1994;89: Lieverse RJ, Kibbelaar RE, Griffioen G, et al. Colonic adenocarcinoma in a patient with multiple hyperplastic polyps. Neth J Med. 1995;46: Jeevaratnam P, Cottier DS, Browett PJ, et al. Familial giant hyperplastic polyposis predisposing to colorectal cancer: a new hereditary bowel cancer syndrome. J Pathol. 1996;179: Torlakovic E, Snover DC. Serrated adenomatous polyposis in humans. Gastroenterology. 1996;110: Otori K, Oda Y, Sugiyama K, et al. High frequency of K-ras mutations in human colorectal hyperplastic polyps. Gut. 1997;40: Ajioka Y, Watanabe H, Jass JR, et al. Infrequent K-ras codon 12 mutation in serrated adenomas of human colorectum. Gut. 1998;42: Longacre TA, Fenoglio-Preiser CM. Mixed hyperplastic adenomatous polyps/serrated adenomas: a distinct form of colorectal neoplasia. Am J Surg Pathol. 1990;14: Urbanski SJ, Kossakowska AE, Marcon N, et al. Mixed hyperplastic adenomatous polyps: an underdiagnosed entity: report of a case of adenocarcinoma arising within a mixed hyperplastic adenomatous polyp. Am J Surg Pathol. 1984;8: Fenoglio-Preiser CM. Hyperplastic polyps, adenomatous polyps, and mixed hyperplastic adenomatous polyps of the colon: definitions. Prog Clin Biol Res. 1988;279: Whitehall VL, Walsh MD, Young J, et al. Methylation of O-6- methylguanine DNA methyltransferase characterizes a subset of colorectal cancer with low-level DNA microsatellite instability. Cancer Res. 2001;61: Hiyama T, Yokozaki H, Shimamoto F, et al. Frequent p53 gene mutations in serrated adenomas of the colorectum. J Pathol. 1998;186: Uchida H, Ando H, Maruyama K, et al. Genetic alterations of mixed hyperplastic adenomatous polyps in the colon and rectum. Jpn J Cancer Res. 1998;89: Ban S. Small hyperplastic polyps of the colorectum showing deranged cell organization: a lesion considered to be a serrated adenoma [letter]? Am J Surg Pathol. 1999;23: Iino H, Jass JR, Simms LA, et al. DNA microsatellite instability in hyperplastic polyps, serrated adenomas, and mixed polyps: a mild mutator pathway for colorectal cancer? J Clin Pathol. 1999;52: Samowitz WS, Slattery ML. Regional reproducibility of microsatellite instability in sporadic colorectal cancer. Genes Chromosomes Cancer. 1999;26: Stoler DL, Chen N, Basik M, et al. The onset and extent of genomic instability in sporadic colorectal tumor progression. Proc Natl Acad Sci U S A. 1999;96: Biemer-Huttmann AE, Walsh MD, McGuckin MA, et al. Mucin core protein expression in colorectal cancers with high levels of microsatellite instability indicates a novel pathway of morphogenesis. Clin Cancer Res. 2000;6: Hawkins NJ, Gorman P, Tomlinson IP, et al. Colorectal carcinomas arising in the hyperplastic polyposis syndrome progress through the chromosomal instability pathway. Am J Pathol. 2000;157: Rashid A, Houlihan PS, Booker S, et al. Phenotypic and molecular characteristics of hyperplastic polyposis. Gastroenterology. 2000;119: Abeyasundara H, Hampshire P. Hyperplastic polyposis associated with synchronous adenocarcinomas of the transverse colon. ANZ J Surg. 2001;71: Hamatani S, Wada R, Tsujimoto S, et al. Histomorphometric characteristics and cellular kinetics of colorectal polyps with epithelial serrated proliferation adjacent to carcinoma. Oncol Rep. 2001;8: Hawkins NJ, Ward RL. Sporadic colorectal cancers with microsatellite instability and their possible origin in hyperplastic polyps and serrated adenomas. J Natl Cancer Inst. 2001;93: Komura K, Masuda H, Esumi M. Distinct relationship between polypoid growth type and sporadic colorectal carcinomas with microsatellite instability. Hepatogastroenterology. 2001;48: Leggett BA, Devereaux B, Biden KG, et al. Hyperplastic polyposis: association with colorectal cancer. Am J Surg Pathol. 2001;25: Makinen MJ, George SMC, Jernvall P, et al. Colorectal carcinoma associated with serrated adenoma: prevalence, histologic features, and prognosis. J Pathol. 2001;193: Albuquerque C, Cravo M, Cruz C, et al. Genetic characterisation of patients with multiple colonic polyps. J Med Genet. 2002;39: Chan AO, Issa JP, Morris JS, et al. Concordant CpG island methylation in hyperplastic polyposis. Am J Pathol. 2002;160: Fogt F, Brien T, Brown CA, et al. Genetic alterations in serrated adenomas: comparison to conventional adenomas and hyperplastic polyps. Hum Pathol. 2002;33: Tateyama H, Li W, Takahashi E, et al. Apoptosis index and apoptosis-related antigen expression in serrated adenoma of the colorectum: the saw-toothed structure may be related to inhibition of apoptosis. Am J Surg Pathol. 2002;26: Wong WM, Mandir N, Goodlad RA, et al. Histogenesis of human colorectal adenomas and hyperplastic polyps: the role of cell proliferation and crypt fission. Gut. 2002;50: Hamilton SR. Origin of colorectal cancers in hyperplastic polyps and serrated adenomas: another truism bites the dust [editorial]. J Natl Cancer Inst. 2001;93: Jass JR, Cottier DS, Pokos V, et al. Mixed epithelial polyps in association with hereditary non-polyposis colorectal cancer providing an alternative pathway of cancer histogenesis. Pathology. 1997;29: Jass JR, Biden KG, Cummings MC, et al. Characterisation of a subtype of colorectal cancer combining features of the suppressor and mild mutator pathways. J Clin Pathol. 1999;52: Jass JR, Young J, Leggett BA. Hyperplastic polyps and DNA microsatellite unstable cancers of the colorectum. Histopathology. 2000;37: Jass JR, Iino H, Ruszkiewicz A, et al. Neoplastic progression occurs through mutator pathways in hyperplastic polyposis of the colorectum. Gut. 2000;47: Am J Clin Pathol 2003;119: Downloaded 794 from