It is rare that a patient with Graves’ disease will undergo biopsy of the thyroid except in the event of a cold nodule developing within the hyperplastic gland. However, in that setting, it is possible that nonnodular tissue is included in the specimen, and it is important to distinguish the papillary hyperplasia from the pathology of the nodule in question. Surgery is an option for patients with this disorder, and total thyroidectomy provides an opportunity for the surgical pathologist to understand the morphology of this entity.

The pathology of Graves’ disease varies with the clinical status and therapy. In its florid form, the gland develops a diffuse increase in cellularity with epithelial hyperplasia forming papillary invaginations into the lumen of follicles that are devoid of colloid or contain only scant colloid with peripheral scalloping (Figs. 9.1 and 9.2, e-Fig. 9.1). The papillae have fibrovascular cores but usually are lined by tall columnar cells with basally oriented nuclei that are crowded but round and regular (Fig. 9.3, e-Fig. 9.2), without the irregularity of contour or the clearing with peripheral margination of chromatin that are the hallmarks of papillary carcinoma. Inflammatory cells are scarce and usually are not found in biopsies but rather are only found scattered in foci of the gland at surgical resection (Fig. 9.4).

The features of this disorder resemble papillary carcinoma superficially, since there is complex papillary architecture that in biopsies can mimic malignancy. In larger specimens, it is evident that the papillae in Graves’ disease are organized within follicles rather than having the haphazard pattern of papillary carcinoma. More critically, the cytologic features are not those of papillary carcinoma; the nuclei are crowded but usually retain their basal orientation. They do not have the convoluted membrane with grooves and inclusions that characterize papillary carcinoma, and they are usually evenly dark and basophilic rather than clear. The differential diagnosis based on these features is the hyperplastic or adenomatous nodule with papillary architecture discussed in the next section. In fine needle aspiration (FNA) specimens, marginal vacuoles or fireflare appearance in May-Grünwald-Giemsa stained smears was initially described as a distinctive feature of thyrotoxic goiter in hyperthyroidism. These vacuoles are thought to represent dilated endoplasmic reticulum, a manifestation of active pinocytosis containing colloid, or diffusion of thyroid hormone [1]. However, marginal vacuoles are found in various
nontoxic thyroid lesions [2], including papillary carcinoma and are therefore of little diagnostic assistance.

With the administration of antithyroid mediations (Tapazole, propylthiouracil), the hyperplasia recedes, follicles resume their more typical appearance, and colloid accumulates within their lumens (Fig. 9.5, e-Fig. 9.3). Radioactive iodine administration results in additional changes, including fibrosis, moderate focal chronic inflammation, and the presence of conspicuous “smudged” nuclei and nuclei that are large, hyperchromatic, and cytologically atypical (Fig. 9.6, e-Fig. 9.4).

A potential mimic of Graves’ disease is type I amiodarone-induced thyroiditis (AIT). This entity is characterized by diffuse or nodular hyperplasia with only focal inflammation [3] rather than the destructive thyroiditis of type I AIT (see Chapter 8).

The “papillary hyperplastic nodule” of the thyroid is usually identified in girls, often in teenagers, in and around the age of menarche. These lesions present as solitary nodules, and they may be associated with clinical or subclinical hyperfunction; they are usually hot on radioiodine scan. These lesions are characterized by a unique architecture that is best recognized at low magnification. Although they do have true papillae with fibrovascular cores, the papillae are usually organized and have a centripedal pattern
within enlarged, distorted follicles. These nodules are distinguished from papillary carcinoma in that they are totally encapsulated or at least very well delineated without evidence of invasion, they often have central cystic change, they usually have at least focal subfollicle formation in the centers of broad edematous papillae, and most importantly, they do not have the nuclear features of papillary carcinoma (Figs. 9.7, 9.8, 9.9, 9.10, 9.11). Although one analysis of clonality has suggested that these are polyclonal hyperplasias [4], the detection of GNAS (also known as Gsα, gsp) or TSH receptor activating mutations in such nodules suggests that they are neoplasms [5, 6, 7, 8, 9]. These specific mutations that activate TSH signaling account for their functional hyperactivity as well as their structural morphology that resembles localized Graves’ disease. Their behavior is almost always benign. Some have advocated the name “papillary adenoma” for these tumors; while scientifically appropriate, this term carries historical connotations that some feel are unacceptable [10].

In adults, one can have a similar histologic appearance in a “hot” nodule, that is, a thyroid nodule which is associated with clinical toxicity or subclinical hyperthyroidism and iodine uptake on scan. These lesions may be solitary, but in adults, they are more often seen in the setting of sporadic nodular goiter.

On fine needle aspiration and on histologic evaluation, particularly at frozen section, papillary hyperplastic nodules or adenomas can be very alarming and lead to a false-positive diagnosis of papillary carcinoma. Indeed, these entities give rise to well-formed papillae, but on higher magnification, they lack the cytologic criteria for the diagnosis of papillary carcinoma, including powdery nuclear chromatin, multiple micro- and/or macronucleoli, intranuclear cytoplasmic inclusions, and linear chromatin grooves [11, 12, 13]. Immunohistochemical studies have shown that HBME-1 can be useful to distinguish these lesions in histologic samples, since papillary carcinoma is frequently positive, but benign lesion are negative for this marker [14]. Molecular analysis GNAS or TSH receptor-activating mutations may be used but is usually not required.

Papillary carcinoma represents the most common thyroid epithelial malignancy diagnosed in regions of the world where goiters are not endemic. Although malignant, these lesions are usually indolent and most have an excellent prognosis with a 20-year survival rate of 90% or better [15, 16]. They are often infiltrative with local invasion and even extrathyroidal extension. When these lesions do metastasize, they most often do so initially via lymphatics with initial regional lymph node metastases. Metastases beyond the neck are unusual in common papillary carcinoma.

Papillary carcinomas may be multifocal. This has been interpreted as reflective of intraglandular lymphatic dissemination. However, the identification of microcarcinomas in up to 24% of the population [17] and the detection of different clonal rearrangements [18] or X-chromosome inactivation patterns [19] in the different lesions from a single patient support the interpretation of multifocal primary lesions in most patients.

The terminology applied to this entity has been confused and confusing. Initially, papillary carcinoma was described as lesion with papillary architecture (e-Fig. 9.5). These lesions were known to also have follicular (e-Fig. 9.6) and mixed papillary and follicular patterns [10, 20, 21, 22, 23, 24, 25, 26]. With the recognition that papillary carcinoma is characterized by what the WHO has described as “a distinctive set of nuclear characteristics” [27],
the histologic diagnosis of papillary carcinoma of the thyroid became a cytologic diagnosis based on these nuclear features. While this recognition has enhanced the utility of FNA in the diagnosis of thyroid nodules, it has also led to controversy about the classification of papillary carcinomas with pure follicular architecture [28, 29].

Molecular studies have expanded the understanding of this controversy. Exclusive, nonoverlapping activating genetic alterations involving BRAF, RET, and NTRK are found in the majority of cases of papillary carcinoma with papillary architecture (Figs. 9.12, 9.13, 9.14, 9.15, 9.16) including multiple variants discussed in next sections with two exceptions [30, 31]. The activating BRAF^V600E mutation is the most common, followed by other genetic alterations, including rearrangements such as RET/PTC or involving TRK; these yield a gene expression profile that has been described as “BRAF like” [30]. Rare reports of mutations in STK11, the gene responsible for Peutz-Jeghers syndrome, may also be implicated in the pathogenesis of papillary carcinoma [32], but this was associated with a BRAF^V600E mutation as well. In contrast, tumors with nuclear atypia and pure follicular architecture harbor RAS mutations and other genetic alterations that are also found in follicular adenomas and carcinomas [30]. Thus, tumors classified as “follicular variant papillary carcinoma” are more akin to follicular neoplasms genetically. The impact of nuclear morphology in this field should be reconsidered for follicular lesions [33], and this subject will be covered in Chapter 10.

BRAF, a proto-oncogene on 7q24, encodes a serine/threonine kinase that transduces regulatory signals through the RAS/RAF/MEK/ERK cascade. Gain-of-function BRAF mutation results in aberrant activation of ERK signaling implicated in tumorigenesis of several human cancers such
as melanoma and colon carcinoma [34]. Several point mutations of exon 15 have been identified in thyroid cancers; BRAF^V600E is the most common in sporadic papillary thyroid carcinoma [34, 35, 36] and is characteristic of papillary carcinomas with papillary architecture. This BRAF mutation occurs in papillary microcarcinomas and is considered to be an early event in thyroid follicular cell transformation. Although some studies suggested that BRAF is a predictor of aggressive behavior [37], more recent data suggest that the information obtained by these molecular studies actually reflected segregation of papillary from follicular variant papillary carcinomas; in pure groups of papillary lesions, BRAF mutation does not add further prognostic information alone [30, 38], and it has become clear that papillary lesions are more aggressive than follicular variant papillary carcinoma [39]. As discussed under “Prognostic features of papillary carcinoma,” (p. 155) additional molecular events are required to support more aggressive behavior [30, 40, 41]. The clinical application of BRAF inhibitors that would provide predictive value for patients with advanced disease who may benefit from this therapy remains to be proved [42, 43, 44, 45].

RET was the first activated receptor tyrosine kinase identified in thyroid cancer. The proto-oncogene on 10q11.2 encodes a transmembrane receptor tyrosine kinase with four cadherin-related motifs in the extracellular domain [34]. RET is normally expressed in the developing central and peripheral nervous systems and is essential for renal organogenesis and enteric neurogenesis. Glial-derived neurotrophic factor (GDNF) ligands and GDNF family receptor-α (GFRα) bind the extracellular domain of
RET to form a trimeric complex that induces tyrosine kinase autophosphorylation and activates several signaling pathways including ERK, PI3K, p38, and JUNK. Gain-of-function mutations of RET are involved in sporadic and familial C-cell-derived medullary thyroid carcinoma including multiple endocrine neoplasia (MEN) 2A, MEN 2B, and familial medullary thyroid carcinoma [46, 47, 48]. In contrast, more than 15 chimeric oncogenes, designated RET/PTC, are implicated in the development of papillary thyroid carcinoma [49]. Somatic chromosomal rearrangement leads to fusion of the 3′-terminal sequence of RET encoding the tyrosine kinase domain and 5′-terminal sequences of heterologous genes [49, 50, 51]. Although wildtype RET is not normally expressed in follicular cells, RET/PTC chimeric oncoproteins lacking a signal peptide and transmembrane domain are expressed in the cytoplasm of follicular cells under the control of the newly acquired promoters. The constitutive activation of the tyrosine domain in
the carboxyl-terminal end of RET/PTC induces signaling pathways within thyrocytes and causes cellular transformation in transgenic mice [52, 53, 54]. The high frequency of RET rearrangements in subclinical papillary thyroid microcarcinomas is consistent with these changes representing early events in the neoplastic process [18]. RET/PTC has been implicated in the development of the characteristic nuclear features of papillary carcinoma through alteration of the nuclear envelope and chromatin structure [55]. Heterogeneity of RET rearrangements in a single tumor has been identified and was interpreted as indicative of a relatively late event; however, an alternative interpretation is that multiple and distinct rearrangements signify multifocal transforming events in benign clonal neoplasms [56]. RET/PTC rearrangements have also been reported in thyroids with Hashimoto’s thyroiditis [57], and this has been the major criticism hampering the application of this molecular marker to biopsy diagnosis; however, these findings are controversial and may reflect either PCR contamination or the presence of common microscopic nodules of papillary thyroid carcinoma [17], since many studies that have excluded these possibilities do not confirm the data [18, 58].

There is wide variation in the reported frequency of RET/PTC rearrangements in papillary thyroid carcinomas, related to geographic location, radiation exposure, and detection methods [59]. The high incidence of RET rearrangements in childhood papillary thyroid carcinomas following the Chernobyl accident suggests a role for radiation damage in the genesis of these paracentric inversions [60, 61]. Analysis of chromatin patterns in thyroid follicular cells identified physical proximity of the partners involved in the illegitimate recombination of these rearrangements [62], providing a plausible scenario for radiation-related susceptibility. In nonradiation induced sporadic papillary carcinomas, RET/PTC-1 and RET/PTC-3 are the most common types of RET rearrangements, identified in approximately 40% and 15% of cases, respectively [59, 63, 64]. These fusions represent the second most frequent genomic alterations in classical papillary thyroid carcinomas [30]; however, the incidence of these rearrangements has been decreasing over time [65].

The receptor tyrosine kinase NTRK1 was the second identified subject of chromosomal rearrangement in thyroid tumorigenesis. The NTRK1 (TRK, TRKA) proto-oncogene on 1q22 encodes the transmembrane tyrosine kinase receptor for nerve growth factor. NTRK1 is typically restricted to neurons of the sensory spinal and cranial ganglia of neural crest origin and regulates neuronal growth and survival. NTRK1 rearrangements, showing ectopic expression and constitutive activation of the tyrosine kinase analogous to RET rearrangements, have been noted in 5% to 13% of sporadic but in only 3% of post-Chernobyl childhood papillary thyroid carcinomas [34]. To date, TPM3, TPR, and TFG have been identified as fusion partners forming chimeric oncogenes designated as TRK, TRK-T1 and T2, TRK-T3, respectively. The prevalence of each fusion type is nearly equal in sporadic papillary thyroid carcinomas, while TPM3-NTRK is more
frequent than other NTRK1 rearrangements in post-Chernobyl childhood papillary thyroid carcinomas. The relatively lower incidence of these rearrangements makes them less valuable as routine markers for diagnostic biopsies [30].