Multiple endocrine neoplasia type 1: extensive analysis of a large database of Florentine patients

Multiple endocrine neoplasia (MEN1) is a rare (prevalence 3–20/100,000) congenital endocrine syndrome, consisting of the development of multiple neuroendocrine tumours (NETs) in a single patient, principally at parathyroid glands, anterior pituitary and gastro-entero-pancreatic (GEP) and thoracic tracts. Other endocrine and non-endocrine tissues can be affected with less frequency, accounting for over 20 different combinations of tumours and lesions.

Principally affected organs are the parathyroids; parathyroid multiple adenomas affect up to 100% of patients by the age of 50, representing the first clinical manifestation in about 90% of cases with a mean age of onset of 20–25 years and rare described cases also by the age of 8 [1]. Parathyroid disease manifests principally as primary hyperparathyroidism (PHPT), which can be normocalcemic in some cases, and often remains asymptomatic in many patients. Symptomatic PHPT is due to the prolonged hypercalcemia and it usually results in loss of bone mass (osteopenia or osteoporosis) and/or nephrocalcinosis. Adenoma ablation by surgery is the treatment of choice, mostly for hypercalcemic and symptomatic cases, even if both the right time and the kind of intervention are still controversial. Pharmacological treatment with calcimimetic drugs demonstrated to be able to control hypercalcemia in patients who do not meet the criteria for parathyroid surgery or do not want to undergo the intervention [2].

The second most common manifestation in MEN1 are neuroendocrine tumours of the GEP tract (GEP-NETs), affecting about 30–70% of patients [1], usually manifesting as multiple adenomas and often recurrent after surgical resection. They present both as micro- or macro-adenomas (diameter over 0.5 cm), and can be non-functioning tumours (NFTs; 20% of GEP-NETs) or active tumours secreting excessive quantities of hormones. NFTs are located mainly in the pancreas and usually they remain asymptomatic till their excessive growth causes compression of adjacent tissues and/or their (frequent) malignant progression and metastases manifest. A continuous imaging surveillance is required and surgery ablation is suggested for NFTs over 0.5 cm in diameter. Recently, also non-functioning NETs (NF-NETs) of the stomach (previously referred as gastric carcinoid of type II enterochromaffin-like cells) have been included in the GEP-NETs group; they have an estimated penetrance of about 10% of MEN1 patients. Functioning GEP-NETs produce excessive hormones, such as gastrin, insulin, somatostatin, glucagon or vasoactive intestinal polypeptide (VIP), causing in many cases an associated endocrine syndrome, and they are, respectively, named gastrinomas, insulinomas, somatostatinomas, glucagonomas and VIPomas. Gastrinomas are the most common (over 50% of cases) MEN1 secreting GEP-NETs; about 90% are located in the duodenum and 10% in the pancreas. Pancreatic gastrinomas are more aggressive. Over secretion of gastrin is responsible for Zollister Ellison Syndrome (ZES) in many cases. Usually, MEN1 gastrinomas are small (less than 0.5 mm) and multiple, with a frequent rate of malignant progression and metastases development, being, together with severe ulcers, one of the most common causes of MEN1-related premature deaths. Insulinomas are the second most common GEP-NETs in MEN1, manifesting in 10–30% of patients, often as multiple tumours. Surgery is usually the therapeutic approach to GEP-NETs, but it is not always effective due to multiple nature of these tumours that are often scattered through the entire neuroendocrine tissue. In case of non-resectable tumours or advanced metastatic cancer, some pharmacological treatments demonstrated to be effective in increasing median progression-free survival [i.e somatostatin analogues (SSAs), cytotoxic chemotherapy (streptozocin and 5-fluorouracil, doxorubicin, temozolomide with capecitabine), inhibitors of thyrosin kinase receptors (sunitinib), and inhibitors of the mammalian target of rapamycin (everolimus)].

Adenomas of anterior pituitary gland affect about 30–40% of MEN1 patients and represent the third most common tumours in MEN1. They can be hormone-secreting or NFTs. Often they develop as single tumours, are non-invasive, and very rarely manifest a malignant progression. Signs and symptoms are principally due to macro-adenomas compressing cerebral structures, or specifically derived by the over-production of one or more pituitary hormones (i.e prolactin, PRL; somatotropin, GH; and corticotropin, ACTH). Functioning tumours are prolactinomas (PRLomas, 60% of pituitary adenomas), somatotropinomas (25%) or corticotropinomas (5%). Trans-sphenoidal or endoscopic resection or radioablation are the treatments of choice for macro-adenomas and NFTs. PRL-secreting micro-adenomas are pharmacologically treated by dopamine agonists, while somatotropinomas are treated with SSAs.

Other MEN1-associated NETs are thoracic carcinoids, of the thymus and the bronchopulmonary tract, in 3% of cases, and tumours/lesions of the adrenal glands in about 20–40% of patients.

Non-endocrine multiple skin lesions are also frequent (i.e collagenomas, angiofybromas, fibromas, angiomas, and lipomas), often manifesting even before of MEN1 neuroendocrine tumours and being, thus, useful in favoring an early diagnosis. Lipomas can manifest also at visceral level.

Germinal inactivating heterozygote mutations of the MEN1 tumour suppressor gene have been identified as responsible for the development of the syndrome, mostly through the loss of the second wild type copy of the gene at somatic level of specifically predisposed neuroendocrine tissues. To date, over 1500 different germinal and somatic mutations, spanning the entire coding region (exons 2–10) and splicing sites of MEN1, have been described, none of them being associated with a specific clinical phenotype and/or disease penetrance [3, 4]. The lack of a direct genotype-phenotype correlation does not allow to foresee the exact clinical course and tumour localization of the disease, to program personalised diagnostic screening or therapeutic plans.

The institution (and continuous updating) of patients’ large specific registries or databases is very useful in the management of rare diseases, like MEN1. Indeed, the collection of clinical, biochemical and genetic characteristics of unselected patients represents a good approach to increase the knowledge on epidemiological aspects of the disease, and natural course and prognosis of single manifestations of the syndrome.

During the last decade, important national MEN1 databases favoured the study of epidemiological, diagnostic, clinical and therapeutic aspects of MEN1-associated tumours, such as the DutchMEN1 Study Group (DMSG), the Groupe d’Etude des Tumeurs Endocrines (GTE), the multicentre database of the MEN1 Consortium of Japan [6], and the Italian MEN1 database [7].

Here, we reported the results of the analysis of a large database of Florentine MEN1 patients from the Referral Centre for Hereditary Endocrine Tumours of the “Regione Toscana”, comparing our data with the previously published. The availability of the Referral Centre allowed the strict interaction of healthcare specialists in different areas of NETs and, thus, granted the collection of extensive data and a continuous long-term follow-up of patients, as well as the possibility to perform genetic test and to associate genetic and clinical data.

Given the autosomal dominant partner of inheritance, MEN1 syndrome manifests an equal gender distribution, however, in our database a greater predominance (64.2% vs. 35.8%) of female patients was detected, confirming what had been previously shown in similar studies carried out in France, the Netherlands, Japan and Italy [6‐9].

The analysis of our MEN1 database confirmed PHPT as the most common manifestation of the syndrome, reaching a penetrance of over 95% after the age of 55, followed by GEP-NETs (about 60%) and pituitary tumours (about 52%), respectively. These data were in accordance with those previously reported by the Japan database [6], and found in the Italian MEN1 database [7], which includes also part of our Florentine patients.

Distribution of GEP-NETs in our database was in accordance with previously published data [1] for all tumour types; insulinoma resulted to be higher in Japanese patients (22%) with respect to Western countries (10%) [1] and our study (about 11%).

Regarding pituitary tumours, our patients showed a higher prevalence of PRLoma (over 41%) with respect to data reported in the MEN1 International guideline (20%) [1], in the DMSG database (16%) [8] and in the GTE cohort (30%) [10].

In our cohort PHPT was prevalently treated by TPT with parathyroid tissue autograft, conversely to a cohort study of the DMSG in which STP with bilateral trans-cervical thymectomy was the procedure of choice [11], and PHPT patients from the MEN1 Japan database which were operated, more than a half, by single gland parathyroidectomy [6]. TPT granted a lower rate of persistence and a longer PHPT-free post-operatory period, than SPT and PPT. A very low percentage of operated patients (5.1%) manifest post-surgical permanent hypoparathyroidism that has been pharmacologically mitigated over the years.

General mean age of diagnosis of the three main clinical MEN1 manifestation in our patients was about 15 years (reaching up to 20 years in case of gastrinomas) earlier than sporadic counterparts; due to natural characteristics of MEN1 but also favoured by the periodic tumour-surveillance screening program and constant follow-up of all MEN1 affected patients and mutation carriers. MEN1-associated tumours have an earlier age of onset, with respect to sporadic counterparts, and often they present a more aggressive course and behaviour; an early diagnosis, followed by an early surgical and/or pharmacological intervention, is the gold standard for reducing morbidity and mortality. In this light, the institution of patients’ database helps in better setting and programming a continuous monitoring and follow-up of patients and periodical screenings of both affected and asymptomatic mutation carriers, increasing the reduction of morbidity and mortality of MEN1. Morbidity in our patients was principally due to a hormone over-secretion (i.e gastrin leading to ZES and causing ulcers and bleeding; PRL responsible for amenorrhea, oligomenorrhea and/or galactorrhea in women and impotence, infertility, decreased libido in men; insulin provoking hypoglycemia, etc), while mortality was caused, prevalently, by malignant progression and metastases of not early detected gastrinomas and carcinoids. Data from the DMSG evidenced that half of patients, diagnosed with liver metastases from duodenopancreatic NETs, died precociously, in a median follow-up of 4 years since tumour discovery, prompting the importance of an early diagnosis and a constant periodic clinical surveillance [12]. Indeed, data from our database highlighted that all the precociously deceased patients were characterised by a late diagnosis of MEN1 (about 15 years after the occurrence of the first clinical manifestation), and they were already presenting, at the time of diagnosis, malignancies (with or without metastasis) and/or severe tumour-related consequences untreatable by current available surgical and/or medical options. Long-term unrecognised and untreated gastrinomas and MEN1-related malignancies were the most common causes of death among our patients, confirming data from a GTE study which indicated duodenopancreatic and thymic NETs as responsible for increased risk of death in MEN1 patients [13].

Data from our database confirmed that biochemical screenings for hormone over-production are able to anticipate the diagnosis of MEN1 of over 10 years with respect to imaging methods, for functioning tumours; radiological screenings are the only effective diagnostic method for NF-NETs. Given the high malignant potential of pancreatic NFTs and their frequency in MEN1 patients, routine radiological surveillance of entero-pancreatic tract is mandatory. Our screening protocol consists in performing abdominal RMI of CT scan every 3 years in mutation carriers and in MEN1 affected individuals by the age of 20, associated with an intermediate pancreatic eco-endoscopy at 18 months, or, alternatively, an abdominal RMI of CT scan every 2 years without the pancreatic eco-endoscopy. In our series of patients, these imaging approaches granted the early recognition of 39 NF pancreatic NETs and a gastric NF-NET, and the pre-operatory localization of tumours. None of our patients died for an undiscovered NF-GEP-NETs, confirming the importance of these applied diagnostic procedures, accordingly also with suggestion of the GTE group [14].

The genetic test is the only one that grants a real early diagnosis, allowing the identification of mutation carriers, within mutated pedigrees, at their still asymptomatic level, and decades before the appearance of any biochemical valuable alteration. The application of genetic test in our patients allowed, indeed, the identification of 34 asymptomatic mutation carriers, 14 of them manifested their first clinical manifestation of MEN1 a mean of about 4 years after the genetic diagnosis; the constant diagnostic screening after genetic test granted the early recognition of the clinical manifestations and favoured a very early therapeutic intervention. In the last two decades, thanks to early genetic diagnosis and identification of mutation carriers at a very young age it has been possible to analyse MEN1 features and manifestations also in young individuals. For years, evidence and data of MEN1 in childhood and adolescents had been derived mainly from case reports. Recently, thanks to the establishment of national databases and collection of large series of patients, two studies, one from the GTE database [15] and one from this Florentine database [16], have investigated clinical, therapeutic and genetic aspects of MEN1 in children and adolescents (respectively before the age of 21 and 20 years), favouring the collection of important data regarding the management of this syndrome in young individuals.

The progressive application of MEN1 genetic test worldwide notably decreased MEN1-associated morbidity and mortality. Unfortunately, MEN1 syndrome shows no direct correlation between genotype and phenotype [17, 18]. Only one study [19] on the GTE cohort showed that survival rate of mutated MEN1 patients was significantly lower among carriers of a MEN1 mutation affecting binding sites with JunD, presenting a 2-fold higher death risk of MEN1-associated tumours; no association was found between mutations and other peculiar phenotypic features. The analysis of our MEN1 patients’ database, as well as that of the Italian MEN1 database [18], confirmed the lack of a direct correlation between a specific mutation, mutation type or mutated gene region with clinical manifestations and MEN1 phenotypes, not allowing the setting of personalised screening and therapeutic programs. The detailed intra-familial analysis of clinical phenotype, age of tumour onset, multiple tumour association, disease penetrance, severity, course and prognosis in all our pedigrees with more than one affected member confirmed the total absence of correlation between these characteristics and the MEN1 mutation. Only a single positive association between development of a GEP-NET and frameshift mutations was reported, with respect to missense mutations. However, this association was not replicated for another mutation type with similar gap in percentage of tumour occurrence; this suggested that the positive association could be only an accidental statistical association. This datum needs to be verified or denied by further studies in broad and different MEN1 series of patients. The GTE research group demonstrated, in MEN1 pedigrees from its database, an intra-familial heritability for pituitary (64%), adrenal (65%) and thymic NETs (97%), progressively decreasing along parental degree distance [20]. The progressive decrease along generations of intra-familial clinical hereditability and, mostly of all, the absence of direct genotype-phenotype correlation prompt the hypothesis of an important, direct role of other modifying factors in the determination of individual MEN1 tumorigenesis. Since differences in MEN1 phenotype have been reported also in identical twins, epigenetics factors, such as microRNAs and histone modifications, are, currently, considered as the most probable responsible determinants, presumably triggered by environmental factors, to define the MEN1 phenotype in patients bearing the same MEN1 mutation. Identification and study of these factors, thanks to the availability of patients’ databases and tissue banks, are mandatory for a complete comprehension of MEN1 tumorigenesis, and for the development of new target diagnostic and therapeutic strategies.

In conclusion, our long-term clinical practice (as Referral Centre for endocrine inherited tumour syndromes), together with the institution of an over 25 year-lasting MEN1 patients’ database, highlighted that the collection of data (including a detailed family and personal clinical and therapeutic history), the performance of the genetic test in patients and first-degree relatives, as well as the continuous follow up are essential for a correct and early diagnosis and for granting patients the best available diagnostic and therapeutic management.