Evolution of Our Understanding of the Hyperparathyroid Syndromes: A Historical Perspective.

We examined manifestations of the HPT syndromes in the context of 5 broad and partly overlapping stages ( Figure 1 ). We conducted Medline searches for each syndrome, each gene, and each encoded protein, as well as for the term hyperparathyroid.

In 1903 Erdheim reported the autopsy of an acromegalic individual with a pituitary adenoma and four enlarged parathyroids ( 6 ). In 1926, Cushing reported the autopsy of a case with tumors in each of the 3 principal tissues of MEN1: parathyroid, pituitary, and pancreatico-duodenal ( 18 ). By 1953, Underdahl could review 14 MEN1 cases ( 19 ). In 1954 Wermer described heritability of MEN1 ( 20 ). In 1955, Zollinger speculated about an ulcerogenic hormone from a pancreatic islet tumor in MEN1 ( 21 ). In 1961, Sipple reported a case with pheochromocytoma and carcinoma of the thyroid; he also found this association in the literature ( 22 ). In 1962, Cushman reported a man with medullary thyroid cancer (MTC), parathyroid adenoma, and pheochromocytoma ( 23 ). In 1958, Jackson reported a kindred with autosomal dominant transmission of hypercalcemia. It included 7 likely PHPT members, of whom 4 had fibro-osseous jaw tumors ( 24 ). In 1966 and 1967 Jackson reported a family with 19 hypercalcemic members, normal PTH values in the presence of hypercalcemia, and failed subtotal parathyroidectomy in each of 3 operated members ( 25 , 26 ). In 1947 and 1948, two neonates were reported with severe PHPT. Serum calciums were 5.5 and 5.5 mM; they died at 5 and 11 months ( 27 , 28 ). The second neonate had parental consanguinity; a similarly affected sibling was born subsequently with serum calcium of 7.4 mM; after an urgent parathyroidectomy at 20 days, he became hypoparathyroid and developed normally ( 29 ). Recessive transmission was likely. In 1936 Goldman reported provisional FIHP ( FOOTNOTE 3 ) in two siblings with severe PHPT at young ages of 17 and 23 years ( 30 ). Serum calcium was 4.75 and 4.5 mM respectively. One large parathyroid tumor (12 and 7 grams) was resected from each. In 1954, Frohner reported provisional FIHP in 5 siblings with severe hypercalcemia between ages 25-30 years ( 31 ). In 1964, Ballard provided an extended characterization of MEN1, describing a family with 16 probably affected members and a review of 74 published cases ( 32 ). PHPT was the most prevalent feature and usually the earliest. The traits included multiplicity of parathyroid tumors, severe peptic ulcer, watery diarrhea, islet tumor sometimes malignant and occasionally insulin-secreting, pituitary tumor, bronchial carcinoid tumor, and lipoma. 16 of 33 deaths had occurred from peptic ulcer complications. The new RIA for gastrin in 1968 implicated gastrin as Zollinger’s speculated ulcerogenic hormone in MEN1 ( 21 , 33 ). Similarly, the prolactin RIA introduced in 1971 established prolactinoma as a common feature of MEN1 ( 34 , 35 ). In 1965, Schimke reported a family with MTC and pheochromocytoma; he collected 5 similar families from the literature ( 36 ). Williams reported similar families including proven or likely PHPT in 4 of 17 cases ( 37 ). Both authors distinguished this phenotype from MEN1. And Chong distinguished MEN2B as a separate familial syndrome, comprising MTC, pheochromocytoma, plus the mucosal neuroma phenotype, but without PHPT ( 38 ). In 1969, Clark used the new calcitonin RIA to reveal increased serum calcitonin levels in cases with MTC ( 39 ). Melvin described provoked serum calcitonin as more sensitive for detection of early MTC ( 40 ). Keiser described findings in a family in which 25 members had MTC, 11 had pheochromocytoma, and 16 had PHPT ( 41 ). In 1977, Dinnen reported a family with the three principal features of HPT-JT: benign parathyroid tumor, malignant parathyroid tumor, and ossifying fibroma of the jaw ( 42 ). The “jaw” tumors of HPT-JT were in the mandible or maxilla. They are currently diagnosed as ossifying fibromas or cementifying fibromas. They have few if any osteoclasts and do not vary with parathyroid activity; these features distinguish them from osteitis fibrosa of PHPT ( 42 ). There have been no formal guidelines for diagnosis of FHH, although many kindreds were reported ( 43 , 44 , 45 , 46 ). The diagnosis depended on a pattern of traits in multiple members of a family: autosomal dominant transmission; high penetrance of hypercalcemia even below age 10 years and approaching 100% in most families; hypercalcemia without hypercalciuria; hypercalcemia with normal serum PTH; and failure of subtotal parathyroidectomy. These features represent an atypical form of PHPT ( 47 ). Occasional cases developed pancreatitis or chondrocalcinosis. Rare FHH kindreds included cases of NSHPT ( 48 ). Urinary excretion of calcium could be expressed as a ratio of calcium clearance to creatinine clearance to provide a useful distinction of hypercalcemic cases with FHH from those with typical PHPT ( 49 ). The hypocalciuria was the PTH-independent expression of the FHH trait in the kidney ( 50 ). NSHPT is neonatal severe PHPT with multiple enlarged parathyroids, hypercalcemia (often over 4 mM), hypotonia, severe HPT bone disease often with bell-shaped thorax and rib fractures, each of which could contribute to respiratory compromise. Three kindreds with FHH included NSHPT in 1-2 members ( 48 ). Subsequently parental consanguinity in 2 other kindreds and mild hypercalcemia in parents suggested that the offspring with NSHPT could sometimes be a homozygous expression of the FHH trait ( 51 , 52 ). After the earliest reports, later descriptions of provisional FIHP focused on milder PHPT and larger families. Most of the largest families eventually manifested characteristics of MEN1, FHH, or HPT-JT ( 24 , 26 , 53 - 55 ). Most other kindreds with provisional FIHP were small and consistent with autosomal dominant transmission. One suggested recessive transmission ( 56 ). Parathyroid tumors in MEN1 are multiple, oligo- or monoclonal, and enlarged asymmetrically ( 57 - 59 ). Gastrinoma in MEN1 is usually in the duodenal mucosa, occurring as small multifocal tumors ( 60 ). Since the introduction of H2-histamine antagonists in 1976 and then proton pump inhibitors in 1989, death from oversecretion of stomach acid in MEN1 decreased almost completely ( 61 ). Thus, foregut neuroendocrine cancer (thymus, bronchus, duodenum, or pancreas) became the principal cause of death ( 62 ). Trait testing among many cases and relatives resulted in derivation of age-related penetrance values of MEN1; for example, penetrance was already 43% by age 20 ( 57 ). Hormone non-secreting tumors were recognized as a common feature: foregut neuroendocrine (30%), facial angiofibroma (90%), truncal collagenoma (70%), lipoma (10%), and leiomyoma in women (unknown prevalence) ( 63 , 64 ). Genetic linkage testing was used to identify carriers and non-carriers in some large kindreds ( 65 ). Diffuse or nodular parathyroid hyperplasia was sometimes recognized at thyroid surgery in MEN2A and was usually accompanied by normal serum calcium. Often only one parathyroid gland was removed ( 66 ). The age-dependent penetrance of MTC could be calculated from basal or penta-gastrin-simulated calcitonin; for example, it was zero at age 20 in one large family with MEN2A ( 67 ). Cutaneous lichen amyloidosis, a pruritic skin lesion in the interscapular region, was described in many MEN2A families ( 68 ). Hirschsprung disease or congenital megacolon was reported several times in MEN2A, including several families with multiple cases ( 69 - 71 ). Farndon and Lombardo described large kindreds (10 or more affected members) with isolated MTC ( 72 , 73 ). These families are now considered as a variant of MEN2A with low MTC aggressiveness ( 74 ). Genetic linkage testing was used for carrier detection in some large kindreds ( 75 ). Carpten tabulated traits in affected members of 14 selected families with HPT-JT. There were benign parathyroid tumors (84%), malignant parathyroid tumors (16%), jaw tumors (44%), and kidney lesions (25%) ( 76 ). Due to the sometimes early and aggressive growth of the parathyroid tumors in HPT-JT, severe hypercalcemia was reported as early as at 10 years ( 77 ). Frequently there was just one parathyroid adenoma or occasionally an atypical adenoma. One case had a PTH non-oversecreting parathyroid cancer ( 78 ). The benign or malignant parathyroid tumors often have a microcystic histology ( 79 ). Genitourinary lesions were frequent in HPT-JT ( 80 ), including benign and malignant uterine tumors, renal hamartomas, Wilms tumor, and polycystic kidney. Uterine tumors occurred in up to 80 % of affected women. The tumors were frequently accompanied by menorrhagia, often required hysterectomy. The collected pathology reports of uterine tumors showed adenomyosis and adenofibroma as the most frequent diagnoses ( 80 ). Further histologic analyses of the uterine tumors are needed. Genetic linkage analysis allowed the diagnosis of some asymptomatic carriers of HPT-JT ( 55 ). High serum calcium values cluster about an average that is consistent within a family ( 44 ). 1,25(OH)2D in serum is not elevated ( 81 ), and ulnar bone mineral density is normal. The parathyroid glands vary from mostly normal size and cellularity to mild enlargement with borderline hyperplasia ( 82 , 83 ). Rarely, FHH presented as a solitary parathyroid adenoma ( 84 ). NSHPT cases in 4 kindreds with parental consanguinity reflected a double dosage at the FHH chromosomal locus, presumably a homozygous mutation ( 85 ). Huang reported 14 families with provisional FIHP ( 86 ). 1 family had 3 affected cases, and the rest had only 2. Mean serum calcium was 3.46 mM, including 5 patients with hypercalcemic crisis. Barry reported 30 families with a median of 3 affected members ( 87 ). Mean serum calcium was 2.96 mM. Simonds reported 36 families. Median size of the kindreds was 2 affected members. Mean serum calcium was 3.06 mM ( 77 ). In 1988, the principal gene ( FOOTNOTE 4 ) for MEN1 was localized to chromosome 11q13 by genetic linkage and loss of heterozygosity (LOH) mapping in tumors from MEN1 ( 88 ). MEN1 was identified by positional cloning [ FOOTNOTE 5 ] in 1997 ( 89 ). Its encoded protein was named menin. MEN1 loss of function germline mutations have been found in 80-95% of probands with a family history of MEN1 ( 90 - 94 ). Sporadic MEN1 cases show a lower frequency of mutations at 6-30% ( 91 , 92 , 94 ). The mutations are scattered across the encoded 610 amino acids. Most of the mutations result in nonsense codons or frameshifts, consistent with menin inactivation ( 95 ). 1-13% of MEN1 probands have had heterozygous gross deletions of MEN1 ( 92 , 95 ). There are no genotype-phenotype correlations with mutations or gross deletion of MEN1 . However, sporadic MEN1 cases without MEN1 mutation have a higher frequency (40-60%) of growth-hormone secreting pituitary tumor. ( 96 , 97 , 94 ). Most of the tumors in MEN1 have LOH at 11q13, representing a second hit at the MEN1 gene. Those without 11q13 LOH often have a small mutation in the second MEN1 allele ( 98 ). With the exception of uterine leiomyomas, sporadic tumors of the same types as seen in MEN1 have both a somatic MEN1 mutation and 11q13 LOH in 5-40% ( 64 ). For example, MEN1 mutations occur in 30% of sporadic parathyroid adenomas ( 99 ) but in only 0-2% of uremic parathyroid tumors ( 100 , 101 ). A rearranged oncogene was identified by expression cloning in 1985 [ FOOTNOTE 6 ] ( 102 ). It was named RET , short for RE arranged during T ransfection. In 1987, the MEN2A gene was localized to chromosome 10 ( 103 ). In 1993, RET was shown to be the gene for MEN2A ( 104 , 105 ). Virtually 100% of probands with MEN2A, MEN2B, or familial MTC have a small germline missense activating mutation in RET ( 74 ). The prevalence of mild PHPT in MEN2A is highest at about 20% for mutations of Cysteine634 ( 106 ). RET has a distinct genotype-phenotype correlation. 85% of the RET mutations for MEN2A are at extracellular codon Cysteine634; 95% of the RET mutations for MEN2B encode Met918Thr ( 74 ). RET mutation testing for carrier detection was superior to provoked calcitonin in sensitivity and specificity. In fact, although 86 cases in a Dutch registry of families with MEN2A had undergone total thyroidectomy based on pentagastrin-provoked calcitonin testing, DNA testing later showed that 6 of 86 had no RET mutation ( 107 ). Thus, false positive (insufficient specificity) calcitonin tests had resulted in 6 inappropriate thyroidectomies. About 50% of sporadic MTCs and 5% of sporadic pheochromocytomas have a somatic mutation in RET ( 74 , 108 ). However, sporadic parathyroid tumors have not shown a RET mutation ( 108 , 109 ). The C-terminal half of RET can fuse with at least 10 other genes to give a rearranged oncogene termed RET/PTC . This somatic mutation is found in 20-70% of sporadic papillary thyroid cancers ( 111 ), in 1-2% of non-small cell lung cancers, and in fewer of other neoplasms ( 112 - 114 ). By inactivation, RET is the gene most frequently mutated in sporadic congenital megacolon at about 10%. The RET inactivating mutations causing megacolon are distributed across the open reading frame; however, some others are also activating mutations that simultaneously cause MEN2A ( 115 ). In 1995, Szabo localized the gene for HPT-JT to 1q21-q31 ( 116 ). In 2002, by positional cloning, Carpten identified the gene for HPT-JT as HRPT2 (later named CDC73 ) and named its encoded product parafibromin ( 76 ). 60-70% of HPT-JT kindreds have germline mutations in CDC73 . Most of the mutations have been in exon 1 that encodes the amino-terminal half of parafibromin ( 117 ). Germline or somatic mutations are virtually all frameshifts or stop codons ( 118 ). Gross deletions were reported in few single cases ( 119 ), but in as many as 33% ( 120 , 121 ). The tumors in HPT-JT have LOH around 1q21-q31, the CDC73 locus. There is no genotype-phenotype correlation for CDC73 ( 118 ). The mutation dependent penetrance of HPT-JT in adults has been 35-65% ( 122 , 123 ). Shattuck analyzed 15 cases with parathyroid cancer, presumably sporadic; they found heterozygous or compound hemizygous mutation of CDC73 in 10. Surprisingly and importantly, 3 of 8 informative cases had a germline mutation of CDC73 ( 124 , 125 ). Somatic mutation of CDC73 is rare in benign parathyroid adenomas without HPT-JT ( 117 , 126 ). Parafibromin is a biomarker for CDC73 . Parafibromin immunostaining is absent in benign or malignant parathyroid tumors of HPT-JT. It is also absent in 60-70% of sporadic parathyroid cancers, in 20% of sporadic atypical parathyroid adenomas, and <1-5% of sporadic parathyroid adenomas ( 120 , 127 - 129 ). The CASR gene, encoding the cell surface calcium-sensing receptor (CaSR), was isolated in 1993 by expression cloning ( 130 ). In the same year, Pollak identified CASR as the principal gene mutated in FHH, FHH1, and NSHPT ( 131 ). The CASR inactivating mutation is heterozygous in FHH1, with one mild case of FHH from homozygous mutation ( 132 ). Most mutations for FHH1 predict missense; those for homozygous NSHPT are mostly truncations or frameshifts ( 133 ). Somatic CASR mutation has not been found in sporadic parathyroid adenoma ( 134 , 135 ). Activating mutations in the CASR result in autosomal dominant hypoparathyroidism type 1 (ADH1) ( 133 ). Most FHH1 mutations and most ADH1 mutations are in the extracellular Venus fly-trap domain of the CaSR. They cluster in its cleft, which is predicted to be a calcium binding domain ( 133 , 136 , 137 ). The CASR mutations causing hypercalcemia or those causing hypocalcemia have a similar distribution in the CaSR and can even modify the same codon, albeit to differing amino acids ( 138 , 139 ). One unique family of 20 affected members with autosomal dominant PHPT, parathyroid adenoma, and hypercalciuria (i.e. not FHH) had a CaSR missense mutation in the cytoplasmic tail of the CaSR ( 140 ). Determination of its CASR status improved the understanding of each case of NSHPT. The CASR mutation is homozygous or compound heterozygous in the severest cases of NSHPT, but it is heterozygous in NSHPT cases of intermediate severity, sometimes termed neonatal primary hyperparathyroidism ( 141 - 143 ). Successful treatment of NSHPT with biallelic CASR mutation often requires total parathyroidectomy and maintenance for permanent hypoparathyroidism ( 144 ). Two enlarged parathyroid glands were analyzed from 1 neonate with homozygous NSHPT. Both glands showed polyclonality, indicating hyperplasia and not adenoma ( 145 ). Homozygotes or heterozygotes with NSHPT can have similarly elevated PTH and similarly severe HPT bone disease, but they have almost completely distinct serum calcium values. Maximal serum calcium is usually 3.6-8 mM in homozygotes but 2.75-3.5 mM in heterozygotes ( 85 , 141 , 142 ). An FHH heterozygote gestating in a wild type mother might sense the maternal normocalcemia as insufficient and thus develop severe secondary HPT in utero, causing one form of NSHPT ( 48 , 143 ). Though many heterozygous NSHPT cases gestated in a wild type mother, this pairing alone was not sufficient to cause NSHPT in many other cases ( 142 ). Most heterozygotes with intermediate NSHPT have Arg185Glu CaSR substitution; its neonatal severity has been suggested to result from a dominant negative effect ( 143 ). A heterozygote with or without calcimimetics, pamidronate, or subtotal parathyroidectomy may later outgrow its presumed secondary HPT and develop into typical FHH ( 141 , 143 , 146 ). A complete panel of the 3 recently identified MEN1, CASR, and CDC73 principal genes could be analyzed after 2002 to help characterize kindreds with provisional FIHP. A small or even a large kindred with provisional FIHP could hold a mutation, proving incomplete expression of another more complex syndrome ( 118 ). This was eventually found in large families for the MEN1 gene in a kindred with 14 affected ( 147 ), for the CASR gene with 20 affected ( 140 ), and for the CDC73 gene with 8 affected ( 148 , 149 ). Three groups screened 15 or more probands with provisional FIHP for mutations in MEN1, CASR, and CDC73 . Total of proband cases was 73. There were 7 with MEN1 mutation, 9 with CASR mutation, 2 with CDC73 mutation, and 55 with no mutation. ( 77 , 92 , 150 , 151 ). In other reports, 2 probands with provisional FIHP had germline mutation of CDKN1B ( 152 , 153 ). The number of affected members per kindred in each of those 3 recent large series was a median of only 2. However, 2 kindreds with FIHP have had 8 affected cases ( 150 , 154 ). The MEN1 gene encodes menin that is widely distributed and expressed principally in the nucleus ( 89 ). Some 30 binding-partners for menin have been found without final definition of menin’s endocrine tumorigenic pathway ( 155 ). Menin binds junD ( 156 , 157 ). JunD is a growth suppressor, but it became a growth promoter when its binding to menin was blocked ( 158 ). Separately, a menin in-frame deletion mutant was found to result in selective loss of the cytostatic activity of transforming growth factor beta (TGF-β) ( 159 ). Menin also binds to a histone methyl transferase complex containing MLL and 4 other proteins, homologous to those in the yeast COMPASS complex ( 160 , 161 ). Menin can recruit MLL to the promoters of 2 cyclin dependent kinase inhibitors (CDKIs) CDKN1B/p27 and CDKN2C/p18 , resulting in transcriptional activation of these CDKIs ( 162 , 163 ). Any of the above, as well as other pathways, represent potential tumor suppressor mechanisms. The carboxy terminal half of MLL undergoes a fusion mutation with one of some 40 different partners and is implicated in many leukemias and lymphomas; MLL oncogenic activity is dependent on its retained binding to menin ( 164 ). This is one of several potential oncogenic mechanisms for menin in tumorigenesis ( 165 ). Blockers of menin in oncogenesis are being developed ( 166 ). Crystal structures showed a deep cleft in menin ( 167 ). There is an amino acid similarity across 5 amino acids in the cleft-binding domains of MLL1 and junD. This sequence may contribute to their binding to the cleft; furthermore, MLL1 and junD modulated menin-regulated transcription in opposite directions ( 167 ). Mouse models with germline heterozygous loss of MEN1 showed many of the same tumors as in MEN1 of man, with the addition of pheochromocytoma (7%), male leydig cell tumors (60%), and diverse ovarian tumors (80%) ( 168 , 169 ). This implicated any of the latter 3 as possibly rare tumors in MEN1 of man ( 170 , 171 ). The RET gene has 21 exons. By alternative splicing, it encodes 3 protein variants with differing C-termini ( 172 ). It is expressed in thyroid C-cells, adrenal medulla, testis germ cells, in the urogenital system during development, enteric neural crest stem cells, and spinal motor neurons ( 173 , 174 ). RET has an extracellular n-terminal cadherin repeat, followed by a cysteine rich domain, followed by a hydrophobic transmembrane domain, and lastly a cytoplasmic domain, containing two tyrosine kinase subdomains and 10 phosphorylation sites ( 175 , 176 ). The transmembrane domain can bind through a glycosyl-phospatidyl inositol intermediate to a membrane coreceptor molecule in a 4 member family (GFRα1-4). Each coreceptor has an extracellular receptor domain that can bind GDNF, nurturin, artemin, or persephin respectively ( 177 ). Tyrosine autophosphorylated RET is a docking site for entry into several cytoplasmic signaling pathways, such as RAS, mitogen activated protein kinase (MAPK), PI3 kinase, AKT, and JNK. Activating mutations in RET increase its endogenous tyrosine kinase activity via RET dimerization at the cysteine rich domain (in MEN2A) or via direct increase of its cytoplasmic tyrosine kinase (in MEN2B) ( 178 ). Some tumors of MEN2A also show RET activation via a RET dosage effect (amplification of mutant RET or loss of wild type RET ) ( 179 ). RET is overexpressed in several cancer types ( 178 ). Multikinase inhibitors and RET-specific tyrosine kinase inhibitors are in use or under development for pharmacotherapies of RET driven neoplasms ( 180 - 182 ). Most of the RET mutations in congenital megacolon are inactivations ( 183 , 184 ). It is not clear how congenital megacolon can occur with either inactivating (in sporadic cases) or activating (in MEN2A) RET mutations ( 115 ). CDC73 encodes the 531 amino acid parafibromin molecule. Parafibromin is widely expressed and located mostly in the nucleus. The C-terminus of parafibromin is homologous to yeast cdc73, that is part of a yeast Polymerase Associated Factor 1 (PAF1) complex of 5 proteins, each with homology in man. In Hela cells, the PAF1 complex binds to a histone methyl transferase and to the POL2A subunit of RNA Polymerase II ( 185 ). In addition to activating transcription of many genes, PAF1 downregulates expression of cyclin D1 and c-myc ( 186 , 187 ). This is a potential tumor suppressor mechanism. In NIH 3T3 cells, SHP2 tyrosine-dephosphorylase primes parafibromin to bind beta catenin, upregulate targets such as cyclin D1 and c-myc, and thereby switch parafibromin from a growth suppressor to a growth promoter ( 188 ). Most of the many N-terminal missense mutations of CDC73 are predicted to disrupt its hydrophobic N-terminal core ( 189 , 190 ). PAF1 may interact directly with the COMPASS complex that contains menin ( 191 ). The CASR encodes the 1078 amino acid extracellular calcium sensing receptor (CaSR) (sometimes termed CaR). It is expressed principally on parathyroid cells and kidney and less on thyroid c-cells, pancreatic islets, gastrin secretory cells, osteoblasts, osteoclasts, bone marrow, etc ( 137 , 192 - 194 ). The CaSR is a class C G-protein coupled receptor (GPCR) with a large extracellular domain in a Venus fly-trap configuration ( 136 , 195 ). Increase of extracellular calcium is an agonist. The downstream result of calcium agonism is a decrease of PTH secretion within seconds. Small molecule agonists (calcimimetics) and antagonists (calcilytics) have been developed for possible pharmacotherapy; in particular, calcimimetics are widely used to treat secondary and primary HPT, including parathyroid cancer ( 193 , 196 ). In the parathyroid cell, the CaSR couples with Gα-11 and perhaps other G-proteins to direct a response to hypercalcemia through an increase of cytoplasmic calcium from intracellular stores and a MAPK induced increase of phosphatidylinositol-triphosphate ( 197 ). Some CASR mutations can bias the signaling selectively against either one of those two pathways ( 198 , 199 ). The normal CaSR is downregulated in the parathyroids of primary or secondary HPT ( 200 ). In the renal tubule the CaSR decreases calcium reabsorption in response to hypercalcemia ( 201 ). In mice with deleted PTH, the CaSR alone can guard against hypercalcemia, via its effects on kidney, thyroid c-cells, etc ( 202 ). The CASR mutations in FHH1 and NSHPT are inactivating. They decrease the sensitivity of the CaSR to calcium and thereby result in a higher set-point for calcium sensing by the parathyroid; this contributes downstream to a rise in serum calcium ( 197 , 202 - 204 ). They also decrease CaSR trafficking to the plasma membrane ( 205 ), thereby decreasing calcium sensing by a second mechanism. A small but undetermined fraction of hypocalciuric hypercalcemia cases is caused by an antibody to the calcium-sensing receptor and termed autoimmune hypocalciuric hypercalcemia ( 206 ). In 2000, Franklin engineered mice with knockout in 3 of 4 alleles from the CDKN2C/p18 and CDKN1B/p27 genes, encoding 2 cyclin dependent kinase inhibitors (CDKIs). The mice developed hyperplasia or neoplasia in 6 different hormone-secreting tissues, representing a simultaneous combination of MEN1 and MEN2A ( 207 ). In 2006 Pellegata studied rats with a similar syndrome and found a homozygous germline frameshift mutation of CDKN1B/p27 ( 208 ). Molatore found similar tumors in heterozygous rats, but tumors developed later than in homozygotes ( 209 ). Pellegata also reported a heterozygous germline inactivating mutation of CDKN1B/p27 in affected members of a kindred with MEN1-like features ( 208 ). Probands or kindreds with MEN1-like features with CDKN1B/p27 germline mutation were named MEN4 (OMIM 610755). The prevalence of MEN4 among all MEN1 probands plus MEN1-like probands is approximately 1% ( 152 ). MEN4 has been reported in 20 probands; 13 had no affected relatives, and only one had more than 3 affected relatives, specifically 12 mutation carriers ( 92 , 210 , 211 ). The overall features among probable MEN4 carriers differ mildly from MEN1: HPT-70%, prolactinoma-2%, GH tumor-9%, ACTH-tumor-5%, nonfunctioning pituitary tumor-9%, gastrinoma-5%, foregut neuroendocrine tumor other than gastrinoma-13% ( 210 , 211 , 212 ). Somatic mutation of CDKN1B/p27 has been implicated in various tumors of man. Somatic heterozygous inactivating mutations were found in 11-16% of hairy cell leukemia, 8% of small intestinal carcinoids, 4% of breast cancers, 4% of prostate cancers, 0-1% of parathyroid adenomas, and 0% of uremic parathyroids ( 153 , 213 - 219 ). Half of the CDKN1B/p27 germline and somatic mutations encode missense and are likely to be inactivations ( 152 ). While 1 allele of CDKN1B may be mutated or deleted in a neoplasm, the other allele is rarely mutated or deleted, suggesting a haplo-insufficient mechanism of tumorigenesis ( 209 ). Agarwal and Costa-Guda analyzed MEN1-like patients for germline mutations in multiple CDKIs, an implicated molecular pathway [ FOOTNOTE 7 ] ( 152 , 153 , 220 ). Agarwal found germline mutation in 1.5% for CDKN1B/p27 , 1% CDKN2B/p15 , 0.5% CDKN2C/p18 , and 0.5% CDKN1A/p21 ( 152 ). Among cases with sporadic parathyroid adenoma, Costa-Guda found germline mutations in CDKN1B/p27 in 2%, in CDKN1A/p21 in 1%, and in CDKN2C/p18 in 1% ( 153 , 220 ). The numbers of affected cases in these two studies were too few to estabish a phenotype from mutation of CDKN2B/p15, CDKN2C/p18, or CDKN1A/p21 that differs from MEN1 or MEN4. We term their phenotypes as MEN1-like. CDKN2B/p15 germline mutations were also found in 4% of familial renal cancer ( 221 ). Inactivating somatic mutations of CDKN2B/p15, CDKN1A/p21, and CDKN2C/p18 have also been implicated in several neoplasms. CDKN2C/p18 somatic mutation was found together with germline RET mutation in 10-20% of MTC or pheochromocytoma from MEN2A and in sporadic MTC together with somatic RET mutations ( 222 , 223 ). CDKN1A/p21 somatic mutations were found in 14% of bladder cancers ( 224 ) and 7% of Hurthle cell thyroid cancers ( 225 ). In 1992, Chou found genetic linkage of FHH to chromosome 3q; FHH at that locus is also termed FHH1 ( 226 ). In 1993, Heath found linkage to 19p13.3 in one kindred (termed FHH2) ( 227 ). And, in 1999, Lloyd found linkage to 19q13 in another kindred (termed FHH3) ( 228 ). In 2013, GNA11 was identified as the gene at 19p13.3 for FHH2 ( 229 ). Four probands have been reported with FHH2 and with differing germline inactivating mutations of GNA11 ( 137 , 230 , 231 ). Hypercalcemia may be milder in FHH2 than in FHH1 ( 137 ). In contrast to the inactivating mutations of GNA11 in FHH2, germline activating mutations of GNA11 cause autosomal dominant hypoparathyroidism type 2 (ADH2) ( 229 ). And somatic activating mutations, are frequent in certain melanomas and in certain vascular neoplasms ( 232 - 236 ). In 2013, Nesbit used whole exome sequencing to identify the FHH3 gene at 19q13.32 as AP2S1 ( FOOTNOTE 8 ) ( 237 ). Based on probands and their syndromal mutations, the approximate prevalence ratios are 64:1:10:25 for FHH1: FHH2: FHH3: and other cause(s), such as autoimmune ( 206 ). Approximately 50 FHH3 probands have had loss of function missense mutation in the same arginine-15 codon: R15C, R15H, R15L ( 238 - 240 ). Average serum calcium was higher in FHH3 than FHH1 in 1 of 2 reports ( 137 , 239 ). The R15L mutation in FHH3 was associated with the severest hypercalcemia, with cognitive impairment at times, and with decreased bone mass ( 137 ). Gorvin searched large databases of germline sequences. She found several novel nonsynonymous changes in AP2S1 . In particular, M117I was associated with mild hypercalcemia in one subject and with predicted loss of function change in AP2S1 ( 241 ). Because alternate mutations of the CASR or GNA11 genes can cause the hypoparathyroid ADH1 or ADH2 syndromes respectively, alternate mutations of AP2S1 were sought in hypoparathyroid probands ( 242 , 243 ). No “ADH3” mutation of AP2S1 was found. Warner mapped a locus for FIHP to chromosome 2p13.3-14 ( 244 ). In 2016, Guan used whole exome sequencing to select candidates for the FIHP gene He identified GCM2 (or GCMB ) at chromosome 6p24.2 as the mutated gene in 7 of 40 probands ( 245 ). Greben confirmed this by finding mutation in 4 of 24 probands ( 246 ). This does not exclude the possibility of another gene for FIHP at 2p13.3-14 ( 244 ). Most operations in cases with activating GCM2 mutation showed multiple parathyroid tumors. A member of one kindred showed a homozygous mutation of GCM2 without a unique phenotype ( 245 ). The diameter of the largest parathyroid tumor was greater in FIHP probands with mutation of GCM2 than in those without ( 245 ). In contrast, inactivating mutations of GCM2 cause familial isolated hypoparathyroidism ( 247 ). There are two CDKI gene families. The INK4/ARF family contains CDKN2A/p16, CDKN2B/p15, CDKN2C/p18, and CDKN2D/p19, each with 4 N-terminal ankyrin repeats. The Cip/Kip family contains CDKN1A/p21, CDKN1B/p27, and CDKN1C/p57 each with shared homology across a 60 amino-acid N-terminus. INK4 proteins bind to and inhibit cyclin dependent kinases (CDKs). Cip/Kip proteins bind to and inhibit both a cyclin and a CDK. CDKIs inhibit CDKs and thereby inhibit the cell cycle. Loss of function in CDKIs lead to increased CDK activities, phosphorylation of retinoblastoma protein and other substrates, release of E2F transcription factors, increased cell proliferation, and sometimes tumor formation ( 248 ). CDKN1B/p27 is nuclear and widely expressed. It is under-expressed and largely cytoplasmic in many neoplasms ( 214 , 249 ). CDKN1B/p27 binds to and inhibits the kinase of cyclin E/CDK2 and cyclin A/CDK2 ( 250 ). CDKN1B/p27 concentration is regulated at the levels of transcription, translation, and degradation ( 249 , 251 , 252 ). The CDKN1B/p27 mutations in MEN4 are inactivations. Like CDKN1B, the CDKN2B, CDKN2C, and CDKN1A CDKI knockouts also cause selected tumors in mice. CDKN2B/p15 binds to and inhibits CDK4 or CDK6, that are regulated by D-type cyclins ( 253 ). CDKN2B/p15 knockout mice have a mild tumor diathesis, with mostly angiosarcomas that occur in 8% ( 254 ). CDKN2C/p18 also binds directly to and inhibits CDK4 or CDK6 ( 255 ). Among CDKN2C/p18 knockout mice, 40% develop large pituitary tumors, 10% testicular tumors, and 8% pheochromocytoma ( 207 , 254 ). GNA11 encodes a widely expressed guanine nucleotide binding protein (G-protein) alpha subunit (Gα-11) ( 256 ). Gα-11 belongs to a 21 member family of G-alpha subunits of the heterotrimeric membrane-associated G-protein signaling complex ( 257 ). A G-protein transduces information from extracellular GPCRs to multiple cytoplasmic signaling pathways. Gα-11 is believed to be the principal alpha subunit that transduces the signal from the activated CaSR in the parathyroid cell ( 137 ). It facilitates signaling both via cytoplasmic inositol 1,4,5-triphosphate/cytoplasmic Ca++ and via the MAPK cascade. The 4 known inactivating mutations of GNA11 in FHH2 are dispersed and inframe. They result in a 30% upward shift of the set-point for calcium in vitro versus the more pathologic upward shift of 50% in FHH1 ( 137 ). The abnormalities have been normalized by treatment with a calcimimetic in vitro or in vivo ( 231 ). Mice with a parathyroid gland specific double knockout of GNA11 and GNAQ are a parathyroid selective model for GNA11 inactivation; they have NSHPT ( 258 ). AP2S1 encodes the sigma subunit 1 (AP2σ) of the adaptor-related protein complex 2. Adapter protein-2 (AP2) is a heterotetramer of α, β, υ and σ subunits. There are 5 complexes AP1-AP5. Each adapter complex has its own homologs for its 4 subunits. The sigma subunit 1 has a mw of 17kD and 142 amino acids and is the smallest of the 4 subunits of AP2 ( 259 ). AP2 is critical for clathrin mediated endocytosis, which internalizes membrane cargo components such as CaSRs. It links clathrin to membranes and binds to tyrosine- or acidic dileucine motifs of diverse membrane-associated cargo proteins ( 260 ). The 3 different AP2S1 mutations in FHH3 are each at arginine-15; each decreases the sensitivity of CaSRs to extracellular calcium. Molecular modeling suggested loss of function interaction of AP2σ with the acidic dileucine motif in the tail of the CaSR ( 261 ). Each of the 3 observed mutants (as well as 3 non-observed but engineered arginine-15 mutants) shifted the calcium set-point upwards similarly in vitro. Those abnormal set-points could be corrected by a calcimimetic in vitro and/or in vivo ( 238 , 261 ). In the drosophila CNS, the glial cells missing gene ( gcm ) is transiently expressed in glial precursors to switch their fate from neuronal to glial. Gcm binds with high specificity to a consensus DNA sequence. There are homologous genes from human ( GCM1 and GCM2 ) and mouse. Their products share a highly conserved N-terminal DNA-binding region with drosophila gcm ( 262 ). In mouse embryos. Gcm1 was expressed only in the placenta; gcm2 had low levels in neural tissue but higher levels only in the parathyroids ( 263 ). Gcm2 probably has many target genes. ( 264 ); for example, it binds to and activates the promoters of the PTH gene and of the CASR gene ( 265 , 266 ). An assay of transcriptional activation by GCM2 revealed a C-terminal conserved inhibitory domain (CCID) of approximately 20 amino acids. Transcription was activated by deletion of the CCID domain from GCM2. 5 missense polymorphisms of GCM2 from 11 FIHP cases were in the CCID and activated transcription, indicating that they represent gain of function mutations ( 245 , 246 ). This was the inverse of inactivations in the N-terminus of GCM2 that were reported as a cause of familial isolated hypoparathyroidism and that usually inactivate transcription by GCM2 ( 247 ). Similarly, germline knockout of gcm2 in mice caused absence of the parathyroids, albeit with residual PTH secretion from the thymic region ( 267 ).

Examination of the evolution of our understanding of the HPT syndromes highlights breakthroughs and innovations, shared among the syndromes. For example, during a period of over 100 years there have been stepwise advances in methods for detection of the carrier of each HPT syndrome. This has progressed from autopsies, to serum calcium, to basal level of a hormone, to provoked level of a hormone, to genetic linkage, and finally to gene mutation. In stage 1 (1903-1967), the early descriptions of the HPT syndromes used the limited tools available. For example, the earliest report of MEN1 was a case with PTH-secreting and GH-secreting tumors identified in an autopsy in 1903 ( 6 ). This was decades before the discovery of either hormone. Early diagnoses of the HPT syndromes depended on symptoms (such as nephrolithiasis, fracture, bone pain) and the available laboratory analyses (specifically serum chemistry, bone radiographs, and pathology analyses from surgery or autopsy). The introduction of practical measurements of serum calcium around 1925 was a major breakthrough shared by each of the HPT syndromes ( 12 ). Hypercalcemia provided the first laboratory insight about a disorder in many patients with a HPT syndrome. Its archival value in the HPT syndromes reflects our current knowledge that PHPT can often be suspected by hypercalcemia without the PTH RIA, that hypercalcemia is highly penetrant in most of these syndromes, and that hypercalcemia is the earliest expression in most of these syndromes ( 268 ). Possible inheritability was reported for each hyperparathyroid syndrome. There was a necessary bias towards more severe and/or more striking features of any syndrome. Consequently, some early interpretations warranted changes later on. For example, in 2 early kindreds with provisional FIHP, the young ages at onset and the severity of PHPT differed markedly from provisional FIHP as seen today ( 30 , 31 , 77 ). Either or both of those kindreds may have had an incomplete expression of HPT-JT that can have early aggressive parathyroid features ( 55 , 77 , 269 , 270 , 271 ). Similarly, Zollinger implicated pancreatic tumor as the source of his hypothesized ulcerogenic hormone in MEN1, but duodenal tumors were more likely the source ( 20 , 60 ). In stage 2 (1959-1985), RIAs for relevant hormones (PTH, gastrin, insulin, calcitonin, human prolactin, etc) were introduced ( 17 , 33 , 34 , 39 , 272 ). Rosalyn Yalow received the Nobel Prize in Medicine for this invention ( 272 ). Hormone RIA could sometimes be applied to identify a symptomatic and even an asymptomatic carrier of MEN1 or MEN2A ( 40 , 41 , 273 ); however, hormone RIA could not reliably identify a non-carrier. The RIAs were also useful for monitoring of the emergence of a hormonal tumor in a syndrome carrier, for characterizing a tumor, and for following the status of a tumor. In fact, annual immunoassay of multiple hormones in an MEN1 carrier remains a central recommendation in the current MEN1 guidelines ( 274 ). In stage 3 (1981-2006), the pathology of the parathyroid glands was found to be largely distinct in several HPT syndromes ( 275 ). The central advance during stage 3 was the assembly of a large case series - in one unique institution, in a collaboration, in a consortium, or in a registry ( 63 , 76 , 107 , 276 ). This supported calculation of age-related penetrance of a syndrome or of a specific trait within a syndrome ( 57 , 67 ). This also enabled the discovery of non-hormonal traits of the skin in MEN1 and in MEN2A ( 63 , 68 ). Genetic linkage testing was now possible in some large kindreds with MEN1, MEN2A, or HPT-JT to identify carriers and non-carriers of that syndrome ( 55 , 65 , 75 , 107 , 279 ). In stage 4 (1985 to the present), genetic mapping defined the approximate chromosomal location of a syndromal gene before identification of the gene ( 88 , 103 , 116 , 226 - 228 ). Subsequently, each of the 4 principal genes ( MEN1, RET, CASR, CDC73 ) from 5 syndromes was identified and then studied. Identification of these genes eventually became possible during only 9 years because of past assembly of many kindreds and the breakthroughs of gene cloning methods, particularly as accelerated by the Human Genome Project ( 280 - 283 ). Identification of a principal gene pointed to the significant fraction of cases without mutation or deletion in that gene. It thus served as an impetus to identification of other genes for that syndrome in stage 5. In stage 5 (1993 to the present), 7 more syndromal genes were identified and then studied. Central methods included animal models ( CDKN1B in MEN4), candidate selection as a neighbor in a pathway, cloning of genes as in stage 4, and, in particular, the innovation of whole exome sequencing for FHH3 and FIHP ( 208 , 237 , 245 , 284 , 285 ). Whole exome sequencing could promote identification of a mutated gene from small numbers of probands and without preliminary need for chromosome mapping information. In stages 4 and 5, the normal and mutated functions of a gene could be studied. An identified gene could be used immediately for robust detection of carriers and non-carriers of its mutations. This resulted in important advances in clinical management of each HPT syndrome. Cases with newly identified mutations led to more precision in the phenotypes for each syndrome. In stages 4 and 5 animal models could be used to explore a syndromal gene ( MEN1, CDKN1B, CASR, GNA11, AP2S1, GCM2 ), even before its molecular pathway was fully understood ( 168 , 207 , 258 , 267 , 287 - 289 ). After its molecular pathway was partly understood, a gene ( RET, CASR, GNA11 ) could be used to help develop pharmacotherapies ( 180 , 181 , 196 , 288 , 290 ). Similar benefits should also accrue from MEN1 and CDC73 genes, but first these genes await further clarification of their detailed mechanisms. Questions persist about how many syndromal genes remain to be identified. The answer can be approached by examining the percent of probands with a syndrome but without a mutation or a large deletion of its known gene(s). As part of this process, the percent of probands with large deletions of the identified genes should be explored in more detail ( 92 , 95 , 120 , 121 ). Rare if any probands lack identified mutation in MEN2A or NSHPT ( 74 , 142 ). However, approximately 20% of probands lack a mutation or deletion among familial MEN1, HPT-JT, or FHH ( 76 , 90 - 92 , 137 , 239 ). Furthermore, 50% of probands lack a mutation or deletion among sporadic MEN1 ( 91 , 92 ); 83% lack this in FIHP ( 245 , 246 ). In conclusion, our understandings about the HPT syndromes have advanced during serial stages. Examination of the stages has helped to clarify the roles of central advances, specifically measurement of serum calcium, RIA, assembly of a large series of cases, gene cloning, and whole exome sequencing. Each central advance impacted several HPT syndromes simultaneously. The accomplishments have been dramatic, and advances will continue. In particular, additional syndromal genes will surely be identified and their mechanisms elucidated in the near future.

Primary hyperparathyroidism (PHPT) has a population prevalence of approximately 1 in 1000 ( 1 ). 10% or more of PHPT cases are part of a syndrome ( FOOTNOTE 1 ) ( 2 ). We review expressions of 6 syndromes that often include PHPT. These are multiple endocrine neoplasia type 1 (MEN1) ( FOOTNOTE 2 ), multiple endocrine neoplasia type 2A (MEN2A), hyperparathyroidism-jaw tumor syndrome (HPT-JT), familial hypocalciuric hypercalcemia (FHH), neonatal severe primary hyperparathyroidism (NSHPT), and familial isolated hyperparathyroidism (FIHP) ( 3 ). We omit transient neonatal hyperparathyroidism and type 1 Bartter syndrome ( 4 , 5 ). The earliest knowledge about PHPT syndromes dates back more than 100 years ( 6 ), and knowledge about PHPT dates back even further ( 7 ). We cite several landmarks in understanding PHPT to serve as background for understanding the HPT syndromes; these landmarks overlap with stage 1 for each of the HPT syndromes (see below). The parathyroid glands were discovered during an autopsy of a rhinocerous in 1852 ( 7 , 8 ). The skeletal pathology of osteitis fibrosa cystica, later attributed to severe primary or secondary HPT, was described in 1891 ( 9 ). In 1906, Erdheim reported that tetany was caused by loss of the parathyroids ( 10 ). MacCallum treated tetany with infusion of calcium in 1909 ( 11 ). Practical measurement of serum calcium was developed by several groups after 1925 ( 12 ). Parathyroid hormone (PTH) was discovered in 1925 ( 13 ). The first parathyroidectomy for PHPT was performed by Mandl also in 1925 ( 14 , 15 ). In 1934 Albright described multigland primary parathyroid hyperplasia among 3 of the first 17 operations for PHPT at Massachusetts General Hospital ( 16 ). The radioimmunoassay (RIA) for PTH was introduced in 1963 ( 17 ).