Epithelial Neoplasms of the Large Intestine

Progression from Adenoma to Carcinoma

Most if not all CRCs arise from adenomas, either conventional adenomas, sessile serrated lesions (SSLs), or traditional serrated adenomas (TSAs) (in decreasing order of frequency). ^, Residual adenoma is identified in approximately 10% to 30% of CRCs; in the remainder, the adenomas are presumably overgrown by cancer. There are distinct associations between the histological type of precursor lesion and the CRC. These indicate that there are two broad pathways involved in neoplastic progression in the colorectum: the conventional adenoma pathway and the serrated adenoma/lesion pathway.

The conventional adenoma pathway accounts for approximately 70% to 80% of all CRCs and is more prevalent in the left colon and rectum than in the right colon. Conventional adenomas typically precede cancer by approximately 15 years. The prevalence of conventional adenomas in the U.S. population is approximately 25% by 50 years of age and 50% by 70 years of age, and these adenomas have a high lifetime risk of progression if not removed. Exact risks of progression are not known; one study estimated a 10% to 15% chance of progression during 10 years for a 1-cm conventional adenoma, whereas another review puts the risk of developing cancer at 3% to 5%. Endoscopic removal of conventional adenomas decreases the incidence of subsequent CRC. ^, The cumulative incidence of new adenomas within 3 years after normal endoscopy averages 27% at follow-up colonoscopy.

The serrated pathway has been increasingly recognized in the past 15 years, and it is estimated to account for approximately 20% to 30% of all CRCs. Most CRCs arising in the serrated pathway develop from SSLs, particularly those located in the right colon. The exact progression risk of SSL to CRC is unknown; however, it is unlikely to be higher than for a similar-sized conventional adenoma, and progression is likely to take at least 15 years on average. CRC is typically preceded by the development of dysplasia within these polyps, which marks a rapid acceleration in carcinogenesis. ^, Historically, conventional endoscopic screening programs have been less effective at reducing right-sided CRC, in large part because the risk of progression of serrated polyps, almost all of which were previously diagnosed as hyperplastic polyps, was not recognized. The progression risk of these polyps is sufficient to warrant their complete endoscopic removal. ^, Challenges remain for endoscopic surveillance programs because serrated polyps are difficult to recognize and completely remove endoscopically, ^, and the underlying genetic events that drive these lesions allow for development of CRCs in relatively small polyps (see later discussion). Furthermore, SSLs are overrepresented as the precursor lesion of interval cancers (cancers identified between screening intervals). ^, TSAs also precede CRC, with typically aggressive features; however, these lesions are much less common. Conventional hyperplastic polyps, particularly those in the left colon, rarely, if ever, progress to cancer.

Aberrant crypt foci represent the earliest stages of colorectal neoplasms and are present before the development of grossly apparent adenomatous polyps. ^, Aberrant crypt foci are microscopic lesions most readily identified by examination of methylene blue–stained, stripped mucosal sheets under a dissecting microscope. They are characterized by a localized collection of crypts that show an increase in crypt diameter and an increased number of lining epithelial cells, which imparts a serrated or slitlike appearance. Histological sections of aberrant crypt foci reveal a range of findings, such as normal or only mildly hyperplastic epithelium, features more typical of serrated polyps, or, rarely, true dysplasia (the latter being similar to microscopic adenomas incidentally identified in patients with familial adenomatous polyposis). Aberrant crypt foci may also be visualized endoscopically, although the technical challenges of these methods have prohibited routine clinical applications. ^,

Reports of very small (<1 cm) carcinomas that lack any evidence of residual adenoma have raised the possibility that some cancers may arise de novo. However, this is a theory that has lost credibility in recent years. These cancers represent less than 5% of all CRCs. Some studies suggest that they are more likely to be of higher grade than ex-adenoma carcinomas, with a higher risk of lymphatic and blood vessel invasion. ^, However, other studies do not support this hypothesis. Hornick and colleagues reported that small carcinomas without a dysplastic component shared clinical and molecular characteristics with small carcinomas that contained a minimal dysplastic component, and with larger carcinomas. It is also possible that some of these lesions represent rapid progression from a small adenoma to cancer because of the early acquisition of high-grade genetic alterations (e.g., aneuploidy, TP53 mutations).

Molecular Basis of Colorectal Cancer Progression

Human cancers are characterized by an accumulation of a variety of genetic alterations, including mutations that either activate oncogenes or inactivate tumor suppressor genes. Accumulation of genetic alterations is critical in the progression from adenoma to carcinoma and likely begins in aberrant crypt foci and other precursor cells that may not manifest morphological features of a neoplasm. To accumulate the array of genetic alterations typical of most CRCs, tumor cells must acquire mutations and epigenetic alterations at an increased rate compared with normal crypt epithelial cells. ^, Increased acquisition and tolerance of mutations is a hallmark of CRC development and is referred to as genome instability . ^, Genes involved in the maintenance of the genome have been likened to “caretakers” of the genome. There are three main patterns of genome instability important to the development of colorectal neoplasia: chromosomal instability (CIN), DNA mismatch repair (MMR) defects that result in microsatellite instability (MSI), and CpG island methylator phenotype (CIMP). Additional mechanisms of genome instability result from base excision repair defects and mutations in DNA polymerase proofreading function (POLE and POLD1). Genome instability is in most cases the result of somatic mutations and is important to the two major morphological pathways to CRC evident to histopathologists, the conventional adenoma to carcinoma (CIN predominant) and the serrated pathway (CIMP, DNA MMR–deficiency predominant) ( Fig. 27.3 ).

DNA Mismatch Repair Defects

MSI is characterized by widespread alterations in the size of repetitive DNA sequences. It is present in approximately 15% of CRCs. ^, MSI is caused by defective DNA MMR ( Fig. 27.4 ) (see Lynch Syndrome and Other Causes of Hereditary Nonpolyposis Colorectal Cancer). In addition to alterations in the size of repetitive DNA sequences, MSI results in a markedly increased rate of mutations of coding sequences (somatic hypermutation). In general, CRCs with MSI do not harbor abnormalities in chromosomal number or the focal regions of subchromosomal gains or losses that typify cancers with CIN. In most CRCs with MSI, the underlying defect in MMR function is caused by epigenetic CpG island hypermethylation–induced silencing of the MLH1 gene. This is a characteristic feature of many CRCs that arise in the serrated neoplastic pathway, and most of these cancers are high-frequency CIMP (see later discussion). ^{, ,} MSI is also the mechanism that underlies the progression of Lynch syndrome cancers, which are caused by inherited defects in DNA MMR (see later discussion). In Lynch syndrome, MSI develops in conventional adenomas and drives rapid progression to cancer.

Molecular Signaling Pathways Involved in Colorectal Cancer

The preceding text described the major mechanisms by which genomic instability develops in colorectal cancer cells. Molecular events at various points in this development of progressive genomic instability are associated with activation of various cell proliferation–associated signaling pathways. The signaling pathways provide a potential target for personalized therapy (see later discussion).

Wingless and Int-1 Signaling Pathway

The Wingless and Int-1 (Wnt) signaling pathway (also known as the Wnt/β-catenin signaling pathway ) is an evolutionarily conserved signaling cascade that is critical to embryonic development and intestinal epithelial renewal. Two main mechanisms of Wnt pathway activation are encountered in colorectal cancer. By far the most common is biallelic inactivation of the APC gene. The APC gene was first identified as the gene mutated in most individuals with familial adenomatous polyposis. In this syndrome, affected individuals inherit one mutant copy of APC that is functionally inactive. In tumorigenesis, the second allelic copy of the APC gene is also inactivated, which fulfills Knudson’s paradigm for tumor suppressor gene inactivation. APC mutations are also present in 70% to 80% of sporadic CRCs, developing at an early stage during neoplastic development, and they are found in dysplastic aberrant crypt foci. ^{, ,} In normal cells, APC forms a complex with glycogen synthase kinase-3β (GSK3β) and Axin, which degrades β-catenin. APC mutations result in inability of the APC complex to bind β-catenin, thereby releasing β-catenin and allowing it to accumulate in the nucleus, where it becomes involved in activating the transcription of a number of other downstream targets, such as cyclin D and Myc. The apparent necessity of APC inactivation for the development of early adenomas has resulted in its designation as a “gatekeeper” gene of colorectal neoplasia.

The other main mechanism of Wnt pathway activation is gain-of-function mutations of the gene encoding β-catenin (CTNNB1) ^, In contrast with the inactivating APC mutations, CTNNB1 mutations target amino acid residues integral to phosphorylation, resulting in persistent activation of Wnt signaling. Finally, mutations have been rarely identified in other Wnt-signaling pathway genes which may result in Wnt pathway activation in CRCs with wild-type APC . These mutations involve genes encoding for AXIN1, AXIN2, and TCF4 ( Fig. 27.5 ).

Mitogen-Activated Protein Kinase Pathway Activation

The mitogen-activated protein kinase (MAPK) pathway is the major mechanism by which extracellular proliferation factors (mitogens) promote cell growth and proliferation. This is accomplished by binding of the mitogen to a cell surface receptor (e.g., epidermal growth factor receptor [EGFR]), which triggers a phosphorylation cascade that incorporates the upstream transducer protein KRAS and downstream transducer proteins BRAF and ERK. ERK activation regulates nuclear targets such as cyclin D1 and CDK4 ^, ( Fig. 27.6 ). Mutations in components of the pathway are important to tumor growth, dissemination, and resistance to drug therapy in many human cancers. KRAS mutation is present in 30% to 40% of CRCs, while BRAF mutation is identified in 10% to 20%. ^{, ,} Importantly, MAPK pathway activation is a critical feature of the serrated pathway to CRC. KRAS mutation is also commonly found in the CIN pathway, where it occurs late in adenoma development and results in constitutive activation of the gene and hence the MAPK signaling pathway.

Other Pathways

Approximately 3% to 7% of CRCs have an HER2 somatic mutation or gene amplification. This offers a potential target for personalized therapy. The family of transforming growth factor (TGF)-β cytokines represents at least 30 proteins that signal via serine/threonine kinase transmembrane receptors that activate the SMAD family of proteins to regulate proliferation, differentiation, adhesion, migration, and other functions. Surface receptors include TGFBR2, TGFBR1, BMPR2, BMPR1A/1B, ACVR2A/2B, and ACVR1A/1B ( Fig. 27.7 ) Inactivation of the signaling pathway allows for unregulated proliferation. In the setting of colorectal carcinogenesis, this inactivation is important for the transition from low-grade dysplasia to high-grade dysplasia in an adenoma. Common causes for inactivation include germline mutations in SMAD4 and BMPR1A in the setting of juvenile polyposis, and somatic mutations in TGF-β receptor and SMAD family proteins. ^, Inactivating mutations in TGF-β receptors often occur in a background of MSI, and more than 50% of colon cancers with MSI contain mutations in ACVR2A and TGFBR2 .

Molecular Classification of Colorectal Cancer

A molecular-pathological classification of CRC has been proposed that is based on microsatellite status (MSI vs. microsatellite stable [MSS]), CIMP status (CIMP-H vs. non-CIMP-H), and the presence of a BRAF or KRAS mutation ( Table 27.4 ). ^{, ,} This classification results in five subtypes of CRC that have shown to have different prognostic significance. In addition, several of the subtypes have distinct morphology (e.g., MSI-high, see later discussion) and different therapeutic implications in the evolving era of personalized therapy (e.g., presence of a KRAS or BRAF mutation or MSI-high status).

TABLE 27.4

Molecular Pathological Classification of Colorectal Carcinoma

Type	MSI Status	CIMP	BRAF	KRAS	% of CRCs ∗
1	MSI	CIMP-H	Mutated	Wild-type	7
2	MSS	CIMP-H	Mutated	Wild-type	4
3	MSS	Non–CIMP-H	Wild-type	Mutated	26
4	MSS	Non–CIMP-H	Wild-type	Wild-type	47
5	MSI	Non–CIMP-H	Wild-type	Wild-type	4

CIMP, CpG island methylator phenotype; CIMP-H, high-frequency CpG island methylator phenotype; CRC, colorectal cancer; MSI, microsatellite instable; MSS, microsatellite stable.

∗ Phipps AI, Limburg PJ, Baron JA, et al. Association between molecular subtypes of colorectal cancer and patient survival. Gastroenterology . 2015;148(1):77–87.e2.

More recently, pure molecular classifications have been developed based on whole-genome sequencing (The Cancer Genome Atlas [TCGA]) and RNA sequencing (transcriptomic profiling; the Colorectal Cancer Subtyping Consortium [CRCSC]) of a large number of CRCs ( Table 27.5 ). The TCGA classification divides tumors into two broad categories: hypermutated or ultramutated (15%) and nonhypermutated (85%). The hypermutated subgroup shows mostly MSI. In 2% to 3% of cases, the degree of mutation is extreme and is the result of failure of the DNA polymerase proofreading proteins POLE or POLD1. ^{, , ,} The nonhypermutated cases are MSS with low levels of mutations, and they correspond mostly to tumors of the chromosome instability pathway. ^{, ,} Transcriptomic analysis of compiled RNA expression data from multiple studies has identified four main CRC subtypes referred to as consensus molecular subtypes (CMSs): CMS1 (MSI-immune, 14%) corresponds to most MSI-high cancers, CMS2 (canonical, 37%) includes most cases with Wnt pathway activation, CMS3 (metabolic, 13%) includes tumors with KRAS mutation, and CMS4 (mesenchymal, 23%) includes tumors with TGF-β pathway activation. ^, A further 13% of tumors show mixed RNA expression that does not clearly fit any of the preceding four subtypes. It is said that the CMS classification has the potential to provide for personalized medicine because it better reflects the innate biology of the tumor. However, at present, the molecular classifications for CRC remain a research tool. It remains to be seen what role they will play, if any, in the diagnostic or therapeutic approach to CRC in clinical practice.

TABLE 27.5

Comparison of Molecular Classifications of Colorectal Carcinoma

Modified from Muller MF, Ibrahim AE, Arends MJ. Molecular pathological classification of colorectal cancer. Virchows Arch . 2016;469(2):125-134.

	The Cancer Genome Atlas	CMS
Method	Whole-genome sequencing	RNA sequencing
Subtypes/molecular abnormality	Hypermutated (16%) (MSI, CIMP-H, BRAF- mutated, SCNA low)	CMS1: MSI-immune (14%) (MSI, CIMP-H, BRAF- mutated, immune infiltration)
	Nonhypermutated (84%) (MSS, SCNA high, Wnt pathway activation)	CMS2: Canonical (37%) (MSS, SCNA high, Wnt pathway activation)
		CMS3: Metabolic (13%) (non–CIMP-H, KRAS- mutated, SCNA low, MSS)
		CMS4: Mesenchymal (23%) (MSS, SNCA high, epithelial-to-mesenchymal transition, stromal alteration)
		Mixed features (13%)

CIMP, CpG island methylator phenotype; CIMP-H, high-frequency CpG island methylator phenotype; CMS, consensus molecular subtypes; MSI, microsatellite instable; MSS, microsatellite stable; SCNA, somatic copy number alteration.

Techniques to Facilitate Lymph Node Detection

Several techniques have been developed to increase the yield of lymph nodes found during tissue dissection. Essentially, these techniques serve two purposes, to make the lymph nodes more visible within the fat or to dissolve the fat around the nodes. Visualization methods are cheaper and simpler. One such technique is the use of GEWF solution, a mixture of glacial acetic acid, absolute ethanol, distilled water, and formaldehyde, all of which are found in most pathology laboratories. GEWF is a lymph node–highlighting solution resulting in lymph nodes becoming more readily identified as chalky-white nodules in fat after immersion in the solution for at least 24 hours. Studies that have evaluated the use of GEWF in CRC specimens have shown mixed results, with some demonstrating increased lymph node yields ^, and others showing no benefit related to this technique. ^, Another visualization method involves injection of methylene blue into the arterial supply of the fresh specimen before fixation. After fixation, the lymph nodes are highlighted by the blue dye. Again, the literature is divided as to whether there is any benefit in lymph node procurement via this procedure, and the dye may create unwanted staining should it inadvertently escape from the vessels on injection.

Fat-clearing techniques involve immersion of the specimen in graded alcohol solutions, followed by xylene to dissolve the fat. Although such methods do increase the yield of lymph nodes, they are time-consuming and expensive (as a result of reagent costs), and there is also the potential of exposure to noxious solvents.

Sentinel Lymph Nodes

The development of distant metastases in 20% to 30% of patients who have a primary tumor confined to the bowel wall is often cited as evidence of undetected metastasis at the time of surgical resection and pathological evaluation. Meticulous pathological evaluation of sentinel lymph nodes (lymph nodes that have the most direct drainage from the tumor) has been investigated in numerous studies. At least three procedures have been used to identify the sentinel node in patients with CRC: injection of methylene blue dye, intraoperative near-infrared fluorescence (NIR) imaging with indocyanine green (ICG), or injection of radiotracer material near the primary tumor. ^, The first one to four lymph nodes that change color or exhibit the highest radiation emission are considered the sentinel lymph nodes. Techniques used to evaluate sentinel lymph nodes include intraoperative frozen section, or thin slicing, submission in total, multiple tissue levels, and immunohistochemical analysis of paraffin-embedded material. Unlike the more common situation with breast carcinoma and melanoma in which knowledge of the sentinel node status aims to reduce the extent of surgery, sentinel node status in colorectal cancer is used only to direct more extensive resection. There are some data to suggest sentinel node sampling could also guide a decision toward formal resection in T1 colorectal cancers, which typically have a low likelihood of lymph node metastases; however, at present the data are inconclusive on the benefit of this procedure in early invasive tumors. A meta-analysis of the sentinel lymph node procedure for CRC showed a low sensitivity for sentinel lymph node detection, regardless of T stage, localization, or pathological technique used. Hence, many questions remain regarding the practical utility of the sentinel lymph node biopsy procedure in the management of CRC, and as a result, it remains infrequently performed.

Criteria for Malignancy and Biopsy Diagnosis

Tumors in which there is invasion confined to the lamina propria and muscularis mucosae (termed intramucosal adenocarcinoma ) are almost never associated with lymph node metastases. This observation is usually attributed to the paucity of lymphatics in the colorectal mucosa. Therefore colorectal tumors are considered to be “malignant” only if they have invaded through the muscularis mucosae into the submucosa (stage pT1). Although this may be sensible for the management of CRC, it has created some controversy about the appropriate nomenclature for neoplasms that do not invade into the submucosa. In the AJCC classification, invasion confined to the mucosa (i.e., lamina propria invasion without submucosal invasion, or “intramucosal” adenocarcinoma) is still classified as stage Tis because of the retention of the term intramucosal adenocarcinoma in certain cancer registries. Unfortunately, with this approach, high-grade dysplasia (intraepithelial neoplasia) is also classified as pTis. At a practical, diagnostic level, because of the lack of malignant potential of lamina propria invasion, use of the diagnostic term intramucosal carcinoma is not recommended, and the term carcinoma should be used only in the context of invasive adenocarcinoma, which refers to tumors that have infiltrated into or beyond the submucosa.

Biopsy specimens of sessile or flat lesions are usually superficial, are often poorly oriented, and may be ulcerated. The most important aspect of pathological examination is to determine whether invasion is present. Although the presence of single infiltrating cells or small, markedly irregular glands readily establishes a diagnosis of “invasion,” some better-differentiated cancers may not show this pattern of growth. In the absence of definitive pathological features of invasion, a diagnosis of invasion relies heavily on identification of desmoplasia, which is usually present with invasive adenocarcinoma and almost never present in intramucosal cancers. However, differentiation between malignancy-associated desmoplasia and the stromal reaction in an ulcerated adenoma can be difficult. The stroma of an ulcerated adenoma is often more cellular, with more abundant microvasculature, inflammatory cells, and plump fibroblasts. If the muscularis mucosae can be identified, it is helpful to note whether invasion penetrates this layer, but in many biopsy fragments, the muscularis mucosae is difficult to identify clearly. Furthermore, the suspicious tissue may be too small, or the orientation of the suspicious area too inadequate to allow certainty that invasion beyond the mucosa exists. In these cases, the presence of desmoplasia is a helpful indicator that the tumor has penetrated into the submucosa, in which case the carcinoma is at least pT1 ( Fig. 27.11 ). Desmoplasia is rarely, if ever, associated with lamina propria invasion alone. Box 27.2 provides a list of additional features that are suggestive of invasive adenocarcinoma. If one or more of these features is present in an equivocal case, the pathology report should convey that the appearance is suspicious for invasive adenocarcinoma. For those cancers that arise within a polyp, the requirement for surgical resection as opposed to polypectomy is determined on the basis of completeness of excision and the presence of high-risk factors for lymph node metastases (see later).

Adenocarcinoma, NOS

Most of these adenocarcinomas are moderately to well differentiated (low grade). Typically, there are medium-to large-sized glands, with moderate variability in gland size and configuration and a moderate amount of stroma ( Fig. 27.12 ). In well-differentiated tumors, the epithelial cells are usually tall and columnar and become increasingly cuboidal or polygonal, with decreasing degrees of differentiation. Mitotic figures and apoptosis are usually abundant. Glandular lumina are usually filled with inspissated eosinophilic material as well as nuclear and cellular debris, so-called “dirty necrosis” ( Fig. 27.13 ). When dirty necrosis is present in a metastasis of unknown primary origin, this feature is frequently used to infer a colorectal primary tumor. In general, there tends to be little difference between the superficial and deeper portions of the tumor, although the leading edge is often associated with gland rupture and more frequent foci of small, irregular, and infiltrative-appearing glands. Some tumors have a prominent papillary component, particularly at the surface. Desmoplasia can be prominent, as in cancers of the pancreas and biliary tract. In some instances, the stromal reaction is more collagenous and may show keloidal areas. The nature of the stromal reaction may be of prognostic significance, with myxoid desmoplastic stroma being associated with poorer survival than a collagen-rich stroma. ^, In addition to glandular cells, a variable number of Paneth cells, neuroendocrine cells, squamous cells, melanocytes, and trophoblasts can be found in adenocarcinoma NOS. Typically, the presence of these other cell types has no prognostic significance. Osseous metaplasia may also be present, possibly related to expression of bone morphogenetic protein by the carcinoma cells.

Mucinous Adenocarcinoma

The World Health Organization (WHO) defines mucinous tumors by the amount of extracellular mucin (i.e., arbitrarily defined as tumor composed of >50% mucin). The terms adenocarcinoma with mucinous features or adenocarcinoma with mucinous differentiation are often used to describe tumors that have a significant mucinous component (>10% but <50%). Most mucinous adenocarcinomas contain free-floating strips of neoplastic epithelium, or individual tumor cells, in the mucin ( Fig. 27.14 ). A variable number of signet ring cells also may be seen ( Fig. 27.15 ). If more than 50% of the tumor cells have a signet ring cell morphology, the tumor is best classified as a signet ring cell carcinoma, even if more than 50% of the tumor is composed of extracellular mucin.

Mucinous adenocarcinomas represent approximately 10% of all CRCs. They are more common in patients with Lynch syndrome and with IBD, but are less common in Asian populations. Mucinous adenocarcinoma is more likely to be diagnosed at an advanced stage. The type of genetic alterations identified in these tumors suggests that the molecular pathogenesis is different from that of adenocarcinoma NOS. Higher rates of mutation are found in genes of the MAPK and PI3K/Akt/mTOR pathways. Mucinous colorectal adenocarcinoma is also more likely to demonstrate CIMP-H and to have higher rates of MSI. ^, MSI-H mucinous tumors are found more often in younger individuals and are more likely to be exophytic and have an expanding growth pattern compared with non–MSI-H mucinous cancers. Immunohistochemical staining for TP53 is less frequently positive (30% vs. >50% in adenocarcinoma NOS), which suggests that mucinous carcinomas are less likely to have a stabilizing point mutation in TP53 . Expression of HATH1, a transcription factor that activates MUC2 expression in intestinal epithelium, is maintained in both mucinous and signet ring cell carcinomas, but is suppressed in nonmucinous carcinomas, which indicates a possible biological basis for mucinous neoplasms. Expression of MUC2 and MUC5AC mucin proteins are both increased; however, there is reduced expression of MUC1 (EMA).

Mucinous adenocarcinoma makes up a greater proportion of right colon tumors and is also more common in females. The cut surface is typically soft and gelatinous with little fibrous tissue, which imparts a “colloid” appearance to the tumor. The tumor is often composed of small nodules of mucin.

On microscopic examination, mucinous adenocarcinomas contain glandular structures or individual tumor cells embedded in pools of mucin; the mucin can be highlighted if necessary, with periodic acid–Schiff or Alcian blue stains. The margins of the tumor can be smooth and expansile or dissecting and infiltrative. In one series of 132 mucinous carcinomas, adjacent precursor adenomas were identified in 31%, a figure similar to that found in adenocarcinoma NOS.

The association between mucinous subtype and survival has been controversial historically. Compared with adenocarcinoma NOS, mucinous adenocarcinoma typically presents at a higher stage at the time of diagnosis, ^, is more likely to have peritoneal implants ^, and invade adjacent viscera, and is less likely to be cured by surgical resection. Mucinous adenocarcinoma is also more likely to show lymph node involvement beyond the pericolonic region. Two recent meta-analyses of the prognostic significance of mucinous adenocarcinoma concluded that when adjusted for stage, mucinous carcinomas do not manifest an overall poorer prognosis than usual-type adenocarcinoma. ^, Furthermore, it does not appear that MSI-H status in mucinous adenocarcinoma is of any prognostic value versus MSI-L or MSS status. Hence, the latest WHO guidelines recommend grading mucinous adenocarcinoma on the degree of glandular differentiation without regard for MSI status. However, there is some evidence that mucinous adenocarcinoma occurring in patients younger than 40 years of age or those developing in the rectum do represent a poorer prognostic group when compared with usual-type CRCs. ^, The adverse outcome of rectal location appears to relate to the poorer response of rectal mucinous carcinomas to neoadjuvant therapies, such as chemoradiotherapy, and to the high rates of incomplete resection that have been reported. ^, At present, mucinous differentiation is not viewed as a factor in treatment decisions for colorectal cancer. Immunotherapy and therapies directed against increased MUC2 protein expression hold promise for future treatment of this CRC subtype ( Box 27.3 ).

Signet Ring Cell Carcinoma

Signet ring cell carcinoma (SRCC) is defined as a tumor that is composed of at least 50% signet ring cells. For classification purposes, this feature supersedes the presence and amount of extracellular mucin. These tumors represent approximately 0.5% to 1.0% of all CRCs. They are slightly more common in men (male-to-female ratio, 1.3:1) and occur at a younger age (mean 63.5 years) than adenocarcinoma NOS. In some studies, more than 50% of signet ring cell adenocarcinomas were detected in individuals younger than 40 years of age. SRCC is also more common in ulcerative colitis, with up to one-third of all signet ring cell carcinomas occurring in patients who have this form of IBD. There is also a strong association with Lynch syndrome and more broadly with MSI. ^, They frequently exhibit CIMP-H (48%), and BRAF mutations are identified in 30% to 33% of cases. ^,

Although some reports suggest right-sided predominance, the overall literature is not consistent as to the site of predilection. ^{, ,} Synchronous tumors are found in 14% of patients. SRCCs are usually ulcerating, and approximately two-thirds have an infiltrative gross appearance. ^, A linitis plastica growth pattern occurs in up to 20% of cases.

Histologically, tumor cells show a characteristic mucin vacuole that pushes the nucleus to the periphery of the cell ( Fig. 27.16 ). A subset of signet ring cells are round and contain centrally located nuclei without an apparent mucin vacuole. Compared with gastric signet ring cell carcinomas, those in the colorectum are more likely to be associated with abundant extracellular mucin and less commonly result in diffuse infiltration of the tissues. It can be difficult to differentiate colorectal signet ring cell carcinoma from metastatic gastric signet ring carcinoma on morphological grounds, and immunohistochemistry is not helpful because tumors from both sites usually are negative for MUC1 and thyroid transcription factor 1 (TTF1) and positive for MUC2. CDX2 may not be expressed as well. However, the tumor cells are usually reactive for SATB2. In common with most gastric signet ring cell carcinomas, there is loss of E-cadherin expression in most cases.

Similar to mucinous adenocarcinomas, signet ring cell carcinomas are more likely to be diagnosed at an advanced stage. ^, Stage III or IV disease is found at the time of diagnosis in approximately 80% of patients. ^, Full-thickness penetration of the muscularis propria, vascular invasion, perineural invasion, and peritoneal seeding are more common than in adenocarcinoma NOS. ^, Some studies have reported distant metastases, often in atypical sites, in as many as 60% of patients at the time of diagnosis. ^{, ,} As a result, surgical resection is less likely to be curative. SRCCs that are MSI-L/MSS are particularly aggressive, ^, with 5-year survival rates less than 10% reported. ^, Mucin-rich cases may have a more favorable prognosis than mucin-poor cases. Peritoneal carcinomatosis is present in virtually all patients who die of their disease, although liver metastases are present in less than 50% of cases. Cytoreductive surgery with hyperthermic intraperitoneal chemotherapy (HIPEC) is not associated with improvement in prognosis in patients with peritoneal carcinomatosis. The frequent association with PDL1 positivity raises the prospect that immunotherapy may become more important in the treatment of this subgroup in the near future ( Box 27.4 ).

Undifferentiated Carcinoma

The term undifferentiated carcinoma is, essentially, restricted to cancers that contain evidence of epithelial differentiation but no obvious gland formation, although most undifferentiated tumors probably represent extremely poorly differentiated adenocarcinoma NOS. For instance, some authors accept this designation for tumors with a very small component (<5%) of gland formation.

Undifferentiated carcinomas tend to be bulky and soft because of their high degree of cellularity and relative lack of desmoplasia, and there is often extensive necrosis. There are typically sheets of cells, cords, or trabecular structures, frequently with an infiltrative growth pattern ( Fig. 27.17 ). The degree of pleomorphism is variable; some tumors show relatively uniform cytological features, whereas others show marked nuclear variability. The distinction between undifferentiated carcinoma and medullary carcinoma can be problematic. Differences from medullary carcinoma include the presence of a more infiltrative border, absence of a syncytial growth pattern, and absence of a heavy lymphocytic infiltrate within undifferentiated carcinomas.

Although pure undifferentiated carcinomas are rare, many adenocarcinomas NOS contain an undifferentiated component ( Box 27.5 ). These tumors are best classified as adenocarcinoma but would be considered high-grade in the WHO grading system based on the undifferentiated component. The presence of any undifferentiated component increases the probability that the tumor contains a DNA MMR deficiency, particularly if it is associated with tumor-infiltrating lymphocytes (see Carcinomas with DNA Mismatch Repair Deficiency).

Medullary Adenocarcinoma

This tumor is more often and simply called medullary carcinoma, and the WHO classification uses both terms. There is wide variation in the incidence of medullary carcinoma in the literature, which is a reflection of the different diagnostic thresholds used by the reporting pathologists. Overall, between 2% and 4% of CRCs are considered to be the medullary type. ^, These tumors are more common in women and typically occur in the cecum or proximal colon. ^, Medullary carcinoma has also been referred to as large cell minimally differentiated carcinoma.

Most medullary carcinomas are associated with a characteristic genomic profile. These tumors are less likely than usual CRCs to show KRAS and TP53 mutations, and they are more likely to harbor defects in DNA MMR and have BRAF mutation ^, (see Carcinomas with DNA Mismatch Repair Deficiency). Even when it is present as a small subcomponent of the tumor, a medullary pattern is often predictive of an underlying DNA MMR deficiency. Some authors consider demonstration of MSI to be a requirement for the diagnosis of medullary carcinoma ; however, this is not an absolute requirement in the fifth edition WHO classification. Medullary carcinomas are overrepresented in patients with Lynch syndrome.

Macroscopically, medullary carcinomas often present as large, bulky tumors. Histologically, they are characterized by sheets of polygonal-shaped cells with vesicular nuclei, prominent nucleoli, and abundant cytoplasm, and they are associated with numerous tumor-infiltrating lymphocytes ^, ( Fig. 27.18 ). Tumor cells may have an organoid or a trabecular architecture, and focal mucin production may also be present. An important diagnostic feature is the presence of a pushing border rather than an infiltrative margin, where the latter is better designated as an undifferentiated carcinoma. Immunophenotypically, these tumors are frequently CK20–, are occasionally CK7+, and often show reduced CDX2 expression. ^, Differentiation from other nonglandular carcinomas is important because medullary carcinomas have a more favorable outcome. Use of immunohistochemistry for calretinin may be helpful in this regard; in one study, calretinin was positive in 73% of medullary carcinomas but in only 12% of poorly differentiated adenocarcinomas. Also, SATB2 expression is usually retained. Neuroendocrine stains are typically negative. ^, PD-L1 expression is more frequent in medullary carcinomas, particularly in tumors that reveal MSI.

Overall, medullary carcinoma has a more favorable prognosis than adenocarcinoma NOS, even though it is more likely to be diagnosed at a higher T stage and have a higher frequency of vascular invasion ( Box 27.6 ).

Serrated Adenocarcinoma

The term serrated adenocarcinoma has been used in two main settings: in the first setting, it is used to describe CRCs that arise from serrated precursor lesions, including both SSLs and TSAs; in the second, it is used to describe CRCs that have a distinctive serrated morphology. Consistent with their origin in the serrated pathway, serrated adenocarcinomas are more often proximal than adenocarcinomas NOS in location. Using these definitions, up to 12% of all colorectal adenocarcinomas and up to 17% of all proximal adenocarcinomas can be considered to be of the serrated type. ^, Morphologically, these tumors are characterized by a variety of features as well. Architecturally, the tumor has epithelial serrations, and some degree of mucin production is common. In mucinous areas, tumor cells are often present in the form of aggregated balls and pseudopapillary rods that project into mucin. Trabecular areas are common in poorly differentiated cases. Tumor cells have abundant clear or eosinophilic cytoplasm, and the nuclei are vesicular with peripheral condensation of chromatin ( Fig. 27.19 ). Necrosis is uncommon or sometimes focal.

Studies of the molecular pathogenesis of serrated carcinomas have revealed significant heterogeneity, although most of these tumors are associated with a high degree of methylation (CIMP). ^, There are at least two major subtypes of serrated carcinomas: proximal MSI cancers that arise from SSL and distal MSS cancers that arise from TSAs. In one study, only 16.1% of all serrated carcinomas were MSI, but 8.2% of the nonserrated adenocarcinomas were also MSI. Therefore most proximal MSI cancers do not show histologically identifiable serrated features, even though they are believed to arise from serrated precursor lesions. In another study of more than 900 CRCs, 9.1% were classified as serrated adenocarcinomas, and about one-half of these had an identifiable serrated polyp precursor lesion. In this series, there was a trend toward decreased 5-year survival, particularly for left-sided cancers. Although the presence of a serrated precursor lesion should always be reported, the fifth edition WHO classification reserves the term serrated adenocarcinoma for tumors that show serrated features in the malignant component of the tumor ( Box 27.7 ).

Carcinoma with Squamous Metaplasia (Adenoacanthoma)

Adenoacanthomas are rare. In one report, they were defined by the presence of adenocarcinoma elements with abundant admixed areas of benign-appearing squamous metaplasia ( Fig. 27.20 ). The natural history and treatment are similar to those of pure adenocarcinomas.

Adenosquamous Carcinoma

Adenosquamous carcinomas are rare neoplasms, accounting for about 0.06% of all CRCs in Surveillance, Epidemiology, and End Results (SEER) data, although they were reported to be three times more common than pure squamous cell carcinomas in one series. ^{, ,} An association with paraneoplastic hypercalcemia and parathyroid hormone–related protein has been reported with sufficient frequency that a colorectal adenosquamous primary tumor should be considered in the differential diagnosis of patients with this clinical presentation. ^{, ,} This tumor type has also been seen in patients with chronic ulcerative colitis. ^, There is an even distribution between the right and left colon. Morphologically, the tumor is composed of both malignant squamous and glandular elements, either as separate components or admixed ( Fig. 27.21 ).

These cancers generally have a higher stage at presentation than adenocarcinomas NOS; in one study, 50% had metastases at the time of diagnosis ( Box 27.8 ). Although the overall survival rate in stage I/II disease is the same as comparably staged adenocarcinoma NOS, the survival rate for stage III/IV disease is significantly lower. ^, The overall 5-year survival rate for all adenosquamous carcinomas is 31%.

Squamous Cell Carcinoma

Primary squamous cell carcinomas of the colorectum are exceedingly rare. The etiology and histogenesis are unknown. Most authorities favor an origin from pluripotent stem cells capable of multidirectional differentiation. Some authors have suggested derivation from foci of squamous metaplasia associated with chronic mucosal irritation. Supporting this hypothesis are reported associations with ulcerative colitis and schistosomiasis. In two separate studies, all 31 cases were negative for human papillomavirus (HPV). ^, However, there is some limited evidence that HPV may play an etiological role in squamous cell carcinoma of the rectum. ^, Most primary squamous cell carcinomas manifest clinically at an advanced pathological stage. The initial presentation is often in elderly patients.

The pathological features are similar to those of squamous cell carcinomas in other organs ( Fig. 27.22 ). Diagnosis of primary squamous cell carcinoma requires (1) exclusion of metastasis from other sites (particularly lung), (2) exclusion of an associated squamous-lined fistula tract (which, if present, is likely the site of origin), and (3) differentiation from carcinomas of the anus that extend proximally into the lower rectum. ^, The 5-year survival rate for lymph node–negative squamous cell carcinomas is reported to be 85%. However, squamous cell carcinomas have been reported to have a poorer prognosis than stage-matched adenocarcinoma NOS ( Box 27.9 ). ^{, ,}

Carcinoma with Sarcomatoid Components (Sarcomatoid Carcinoma, Spindle Cell Carcinoma, Carcinosarcoma)

Sarcomatoid carcinomas, also termed carcinosarcomas or spindle cell carcinomas, are more common in elderly patients, are often bulky or fleshy, and show abundant hemorrhage. Microscopically, these tumors reveal a biphasic growth pattern that combines both epithelial and mesenchymal elements ( Fig. 27.23 ). Histological areas of transition may be observed in some cases. The spindle cell component may be entirely undifferentiated, or it may show osseous, cartilaginous, or smooth muscle differentiation.

Similar to sarcomatoid carcinomas in other anatomic locations, these tumors usually express keratins and epithelial membrane antigen (EMA) in both the carcinoma and the spindle cell components. Carcinoembryonic antigen (CEA) positivity is usually limited to the adenocarcinoma element. Focal S100 and myoglobin positivity has also been described in some of these tumors. Metastases may show either one or both of these cellular components. There is evidence in favor of a common progenitor cell origin as a result of the finding of shared TP53 mutations in both the epithelial and sarcomatoid components. Further subtyping into sarcomatoid carcinomas (which are keratin positive throughout the tumor) and carcinosarcomas (which are keratin negative in the mesenchymal component) has no apparent clinical or prognostic utility. These neoplasms are typically fast-growing and aggressive, with a mean survival of approximately 6 months ( Box 27.10 ).

Carcinoma with Rhabdoid Features

These tumors are considered a variant of carcinomas with sarcomatoid components in the fifth edition of the WHO classification. Most of these tumors occur in the right colon in patients in their seventh decade of life. This carcinoma is characterized by cells with abundant intracytoplasmic eosinophilic inclusions (“rhabdoid cells”) ( Fig. 27.24 ). The characteristic tumor cells may be a component of an otherwise typical adenocarcinoma NOS (“composite”) or represent a “pure” process. The tumor cells are arranged in diffuse sheets within a variable myxoid background. Spindle and pleomorphic tumor cells may also be encountered, and abortive glandular elements may be present as well. Keratin and vimentin are both expressed in the rhabdoid cells. A diagnostically useful feature is the loss of nuclear expression of SMARCB1 (INI1), a core subunit of the SWI/SNF chromatin remodeling complex. However, INI1 loss is not entirely specific because it may be seen in up to 11% of adenocarcinoma NOS patients. Abnormalities in centrosome structure (ciliary rootlet coiled coil [CROCC]) have also been described. Most colorectal rhabdoid carcinomas display a CIMP-H, BRAF -mutated, MSI molecular phenotype. ^, Carcinomas with rhabdoid features are highly aggressive, with an overall survival rate of 7.9 months ( Box 27.11 ).

Adenoma-Like Adenocarcinoma (Villous Adenocarcinoma, Invasive Papillary Adenocarcinoma)

Adenoma-like adenocarcinoma is a rare subtype of tumor characterized by ≥50% of the invasive component having an adenoma-like appearance with low-grade villous architecture ( Fig. 27.25 ). This subtype accounted for 8.6% of all CRCs in one series ; however, lower incidence rates in the order of 1% to 3% are more generally reported. ^, Fewer than one-half of the tumors have severe cytological atypia, and differentiation from a benign adenoma is difficult, either with or without prolapse changes, particularly on biopsy or limited resection material. The distinction is based on the presence of the neoplastic epithelium associated with a desmoplastic stromal response, which is present in almost all malignant cases. ^, The invasive margin is typically pushing rather than infiltrating in appearance. KRAS mutation is frequently present (58% in one study), and the prognosis is favorable ( Box 27.12 ).

Micropapillary Adenocarcinoma

Micropapillary adenocarcinoma is characterized histologically by small nested clusters of tumor cells that are characteristically retracted from the surrounding stroma ( Fig. 27.26 ). The incidence of this morphological pattern varies from 5% to 20%. Although not required to establish a diagnosis, micropapillary adenocarcinoma has a characteristic immunohistochemical finding referred to as an “inside-out” pattern of reaction for epithelial membrane antigen EMA (MUC1) whereby expression is on the external aspect of the epithelial nests rather than on the luminal aspect, as typically seen with adenocarcinoma NOS. This corresponds to the finding on electron microscopy of microvilli on the outer surface of the epithelium and secretory activity into the surrounding stroma, which presumably accounts for the separation of tumor nests from the surrounding stroma. Micropapillary adenocarcinoma also exhibits loss of MUC2 and loss or reduced E-cadherin staining. Tumor cell expression of vimentin and nuclear expression of SMAD4 is also present, which suggests that this morphological pattern represents a form of epithelial-to-mesenchymal transition. Micropapillary adenocarcinoma frequently shows mutation of TP53, KRAS, or BRAF, but they are MSS. Stem cell markers are often expressed. Adenocarcinomas containing a ≥5% component of micropapillary pattern are more aggressive than adenocarcinomas NOS, with an increased proportion of lymph node positivity (up to 80%), reflecting the higher levels of vascular invasion typically identified in this tumor type. In view of this, the WHO classification designates any adenocarcinoma with a ≥ 5% micropapillary component as a micropapillary adenocarcinoma ( Box 27.13 ).

Choriocarcinoma

Primary choriocarcinoma of the colorectum is rare. In reported cases, patients have been younger than those with adenocarcinoma NOS, with nearly one-half being younger than 50 years of age. These cancers may manifest with massive bleeding from the gastrointestinal tract or from liver metastases. ^, Microscopically, the tumor reveals a variable admixture of choriocarcinoma and adenocarcinoma elements. The choriocarcinoma component stains with SALL 4, human chorionic gonadotropin (β-hCG), and α-fetoprotein (AFP). On occasion, choriocarcinoma elements predominate in metastases when the primary tumor was only focally β-hCG+. Increased serum levels of β-hCG and α-fetoprotein have also been reported and serve as useful biomarkers of treatment response. Several theories exist to explain the choriocarcinoma component. The favored theory is dedifferentiation of the concomitant adenocarcinoma. These cancers generally show metastasis to the liver at the time of clinical presentation and follow an aggressive clinical course despite the availability of germ cell tumor–directed chemotherapy ( Box 27.14 ).

Tumors with Neuroendocrine Differentiation