Introduction

Multiple endocrine neoplasia type 2 (MEN2) is an autosomal, dominantly inherited disorder manifesting various combinations of medullary thyroid carcinoma (MTC) and pheochromocytoma, with hyperparathyroidism (MEN2A) or neuromas of the enteric autonomic nerve cells (MEN2B). MEN2 is caused by the gain-of-function mutation in the RET protooncogene, encoding a transmembrane receptor tyrosine kinase [1, 2]. Most of the mutations, found in the cysteine-rich extracellular domain, give rise to ligand-independent receptor dimerization and cross-phosphorylation, leading to constitutive activation of the downstream signal of the receptor [3]. Mutation of the cysteine codon 634 constitutes 80–90 % of MEN2A cases, although those caused by mutations of the cysteine codon 611, 618, and 620 are also observed. Although they are a minor subset, RET mutations in MEN2A cases are also identified within the intracellular domain, including those originally reported as mutations of familial medullary thyroid carcinoma (FMTC). On the other hand, most cases with MEN2B carry M918T or A883F mutations in the tyrosine kinase domain, suggesting strong genotype–phenotype correlations.

The discovery of strong genotype–phenotype correlations that govern the development of MEN2A-associated endocrine neoplasia in MEN2 cases has prompted us to utilize the identified RET mutations for the prediction of prognosis, and for the determination of surgical concept. In particular, mutations at codons 609, 768, 790, 791, 804 and 891 are classified as level 1, having the lowest risk for aggressiveness among the three levels of MTC [4]. A rare mutation, S891A, has been associated solely with intermediate-risk FMTC [5–7]. For the carriers of such FMTC mutations, intensive screening for age-related development of pheochromocytoma need not be started until they are 20 years old [7]. However, a rare case with a S891A mutation expressing MTC and pheochromocytoma was recently reported, suggesting the limitation of genotype-based predictions [8].

In this manuscript, we report a rare case of a patient who was affected by bilateral pheochromocytomas as the first manifestation of MEN2A, whose subsequent screening for RET mutation identified S891A.

Case report

A 57-year-old woman visited a local hospital with cough and vomiting. She had episodes of periodic headaches and paroxysmal palpitations. Bilateral adrenal tumors were identified by abdominal CT scan (maximum diameter of 4 cm for right mass and 9 cm for left mass) (Fig. 1a). Pheochromocytomas were diagnosed by 131I-metaiodobenzylguanidine (MIBG) scan (Fig. 1b), along with elevated urinary catecholamine and metabolite concentrations (Table 1), and she was, therefore, transferred to the Fujita Health University Hospital for surgical management. On admission, blood pressure was 111/68 mmHg. Since her maternal aunt had adrenal disease and had died from cerebral vascular disease, MEN2A was strongly suspected. Although her basal serum calcitonin level was normal(37.0 pg/ml), elevated levels were recorded following stimulation with 2 mg/kg of Ca2+ infusion (330 pg/ml). However, neither ultrasonography nor 99mTc-methoxy-isobutyl-isonitrile (MIBI) scan showed any cervical lesions. Although serum PTH level was elevated (340.5 pg/ml), serum calcium and phosphorus were within normal range (9.3, 3.8 mg/dl), and urinary Ca excretion was not elevated (Ca/creatinine = 0.163). Finally, screening for the RET gene was performed after appropriate informed consent was obtained (approved by the Ethical Review Board for Human Genome Studies at Fujita Health University), and, unexpectedly, the S891A mutation was identified.

Fig. 1
figure 1

a Computed tomography of abdomen shows right (arrow) and left (open arrow) adrenal tumors. b 131I-MIBG scan reveals abnormal uptakes indicating bilateral pheochromocytomas

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Table 1 Urinary catecholamine and metabolite level
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Bilateral adrenal tumors were surgically resected, and the diagnosis of pheochromocytomas was confirmed by histological examination (Fig. 2a). One year later, the patient underwent prophylactic total thyroidectomy. Histological examination demonstrated that multiple nodal lesions were scattered, indicating the presence of C-cell hyperplasia without any evidence of MTC (Fig. 2b, c). Regarding the four resected parathyroid glands (Fig. 3a), two right glands were slightly enlarged (231 mg for superior and 118 mg for inferior glands) compared with the two left glands (48 mg for superior and 15 mg for inferior glands). However, no specific histological change was observed even in the enlarged glands (Fig. 3b). Approximately, one-third of the resected parathyroid glands were implanted into the muscle of the left forearm.

Fig. 2
figure 2

a Microscopy of left adrenal tumor shows pheochromocytoma (hematoxylin & eosin, ×100). B Microscopy of thyroid gland shows C-cell hyperplasia (hematoxylin &eosin, ×100). c Microscopy of thyroid gland shows C-cell hyperplasia (calcitonin immunostaining, ×100)

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Fig. 3
figure 3

a Macroscopy of the resected parathyroid glands shows slightly enlarged right upper and lower gland. (a right upper gland, b left upper gland, c right lower gland, d left lower gland) b Microscopy of right upper parathyroid gland shows no hyperplasic change (hematoxylin & eosin, ×100)

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Currently, the patient’s two brothers (59 and 53 years old, respectively) and three sisters (69, 64 and 61 years old, respectively) do not have any clinical symptoms associated with MEN2A, although screening for RET mutation has not been performed for them yet. The patient has two sons, 29 and 27 years old, neither of whom have clinical symptoms. The 27-year-old son requested RET mutation screening, and indeed, the S891A mutation was identified (Fig. 4). Routine chemical screening of the blood, including basal serum calcitonin levels, was all normal. Ultrasonography did not detect any mass within the thyroid. He is being followed up carefully as a presymptomatic MEN2A case.

Fig. 4
figure 4

Pedigree of the family. The arrow indicates the proband. The II-3, who was affected by adrenal disease and died from cerebral vascular disease, might possibly carry the S891A mutation

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Discussion

S891A mutation constitutes 2 % of all RET mutations identified in MEN2/FMTC cases [9]. Early reports stress the association of this mutation with FMTC, but accumulating evidence shows the mutant’s capacity to induce a wider spectrum of MEN2A [10]. S891A mutation causes MTC in 63.5 % of cases, pheochromocytoma in 4.1 % of cases and parathyroid hyperplasia in 4.1 % of cases [10]. Compilation of MEN2A-related clinical manifestation in patients with RETS891A mutation in previous reports [5, 6, 8, 10–17] is described in Table 2. Indeed, the management guideline of medullary thyroid cancer by the American Thyroid Association categorizes pheochromocytoma in S891A mutation as ‘rare’ [18]. None of ten Japanese cases with S891A mutation reported in a recent study had pheochromocytoma [17], but our case report combined with previous data indicates that S891A patients as well as other MEN2A patients require early detection of subclinical pheochromocytoma to prevent a potential hypertensive crisis In MEN2 patients, the gain-of-function mutation in the RET receptor tyrosine kinase gene constitutively activates the downstream signals, leading to transformation of the cells [3]. Although the mutations in the cysteine-rich extracellular domain all target cysteine codons, inducing ligand-independent RET dimerization, the mutations located in the intracellular tyrosine kinase domain do not target cysteine codons. These mutations, including S891A, are considered to give rise to structural changes in the protein facilitating the access of adenosine triphosphate and substrate to the catalytic site [19]. The less constitutive RET kinase activation relative to mutations in the cysteine-rich extracellular domain might result in less neoplastic transforming capacity [20, 21].

Table 2 MEN2A-related clinical manifestation in patients with RETS891A mutation in previous reports
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The S891A mutation is classified as level 1 [4] or level A [18], the lowest risk group among the three (level 1–3) or the four (level A–D) RET codon mutation stratification categories. The penetrance and aggressiveness of MTC arising in cases with S891A are variable, but MTC develops at a later age and grows more slowly than with the higher risk mutations. There has been little consensus concerning the management of patients with level 1/level A mutations. According to the guidelines, cases with S891A mutation still need prophylactic resection, and some experts recommended thyroidectomy by the age of 5 years, while others suggest that thyroidectomy by the age of 10 years is appropriate with careful follow-up and periodic calcitonin testing [4, 14, 18]. Since the timing of thyroidectomy should be determined considering the earliest finding of MTC in asymptomatic carriers, prophylactic thyroidectomy at an early age is generally recommended even in cases with level 1/level A mutations [14, 16, 18, 22]. In this regard, two S891A cases, whose MTCs were diagnosed at 17 years old, might be of help in determining the timing of surgery [10].

In this patient, serum levels of PTH were mildly elevated, although the urinary excretion of Ca2+ was not increased. One possibility for this is that the patient has primary hyperparathyroidism due to parathyroid gland hyperplasia or adenoma, as is observed in 20–30 % of MEN2A cases [23]. Since the resected right parathyroid glands of the patient were found to enlarge, we carefully examined the histology of the four resected parathyroid tissues, but there was no evidence of hyperplasia or adenoma. Alternatively, it is possible that the elevated serum PTH levels were due to secondary to decreased serum Ca2+ levels induced by excess calcitonin excretion from the C-cell hyperplasia. Furthermore, this hypothesis implies that the patient with the S891A mutation carried slow-growing C-cell hyperplasia for a long period, during which time the decreased Ca2+ was compensated for by the secondary hyperparathyroidism. These data lend support to the concept that C-cell hyperplasia in the patient with S891A mutation, in spite of its slow growth speed, has a high chance of obtaining another hit for malignant transformation. The patient’s two brothers and three sisters, all of whom are more than 50 years old, do not have any clinical symptoms associated with MEN2A at this moment. This implies that early surgical intervention is not required for S891A cases, although it is still possible that none of the brothers and sisters carries the S891A mutation. Careful follow-up of these older relatives is required even though they do not yet have any MEN2-related symptom.