Somatic mutation of the MEN1 gene in parathyroid tumours.

Primary hyperparathyroidism is a common disorder with an annual incidence of approximately 0.5 in 1,000 (ref. 1). In more than 95% of cases, the disease is caused by sporadic parathyroid adenoma or sporadic hyperplasia. Some cases are caused by inherited syndromes, such as multiple endocrine neoplasia type 1 (MEN1; ref. 2). In most cases, the molecular basis of parathyroid neoplasia is unknown. Parathyroid adenomas are usually monoclonal, suggesting that one important step in tumour development is a mutation in a progenitor cell. Approximately 30% of sporadic parathyroid tumours show loss of heterozygosity (LOH) for polymorphic markers on 11q13, the site of the MEN1 tumour suppressor gene. This raises the question of whether such sporadic parathyroid tumours are caused by sequential inactivation of both alleles of the MEN1 gene. We recently cloned the MEN1 gene and identified MEN1 germline mutations in fourteen of fifteen kindreds with familial MEN1 (ref. 10). We have studied parathyroid tumours not associated with MEN1 to determine whether somatic mutations in the MEN1 gene are present. Among 33 tumours we found somatic MEN1 gene mutation in 7, while the corresponding MEN1 germline sequence was normal in each patient. All tumours with MEN1 gene mutation showed LOH on 11q13, making the tumour cells hemi- or homozygous for the mutant allele. Thus, somatic MEN1 gene mutation for the mutant allele. Thus, somatic MEN1 gene mutation contributes to tumorigenesis in a substantial number of parathyroid tumours not associated with the MEN1 syndrome.

G proteins in cellular control.

In the past year, cDNA cloning has revealed substantial diversity in G protein alpha, beta and gamma subunits. The number of cellular functions recognized to be controlled by G proteins is also increasing. Most G proteins are associated with the cytoplasmic surface of the plasma membrane, and molecular mechanisms for membrane association of certain G protein subunits have been defined recently. Mutations in G protein subunits, both artificially induced and naturally occurring, have provided important insights into G protein structure and function.

Amino- and carboxy-terminal deletion mutants of Gs alpha are localized to the particulate fraction of transfected COS cells.

To elucidate the structural basis for membrane attachment of the alpha subunit of the stimulatory G protein (Gs alpha), mutant Gs alpha cDNAs with deletions of amino acid residues in the amino and/or carboxy termini were transiently expressed in COS-7 cells. The particulate and soluble fractions prepared from these cells were analyzed by immunoblot using peptide specific antibodies to monitor distribution of the expressed proteins. Transfection of mutant forms of Gs alpha with either 26 amino terminal residues deleted (delta 3-28) or with 59 amino terminal residues deleted (delta 1-59) resulted in immunoreactive proteins which localized primarily to the particulate fraction. Similarly, mutants with 10 (delta 385-394), 32 (delta 353-384), or 42 (delta 353-394) amino acid residues deleted from the carboxy terminus also localized to the particulate fraction, as did a mutant form of Gs alpha lacking amino acid residues at both the amino and carboxy termini (delta 3-28)/(delta 353-384). Mutant and wild type forms of Gs alpha demonstrated a similar degree of tightness in their binding to membranes as demonstrated by treatment with 2.5 M NaCl or 6 M urea, but some mutant forms were relatively resistant compared with wild type Gs alpha to solubilization by 15 mM NaOH or 1% sodium cholate. We conclude that: (a) deletion of significant portions of the amino and/or carboxyl terminus of Gs alpha is still compatible with protein expression; (b) deletion of these regions is insufficient to cause cytosolic localization of the expressed protein. The basis of Gs alpha membrane targeting remains to be elucidated.

Mediation of growth factor induced DNA synthesis and calcium mobilization by Gq and Gi2.

A newly identified subclass of the heterotrimeric GTP binding regulatory protein family, Gq, has been found to be expressed in a diverse range of cell types. We investigated the potential role of this protein in growth factor signal transduction pathways and its potential relationship to the function of other G alpha subclasses. Recent biochemical studies have suggested that Gq regulates the beta 1 isozyme of phospholipase C (PLC beta 1), an effector for some growth factors. By microinjection of inhibitory antibodies specific to distinct G alpha subunits into living cells, we have determined that G alpha q transduces bradykinin- and thrombin-stimulated intracellular calcium transients which are likely to be mediated by PLC beta 1. Moreover, we found that G alpha q function is required for the mitogenic action of both of these growth factors. These results indicate that both thrombin and bradykinin utilize Gq to couple to increases in intracellular calcium, and that Gq is a necessary component of the mitogenic action of these factors. While microinjection of antibodies against G alpha i2 did not abolish calcium transients stimulated by either of these factors, such microinjection prevented DNA synthesis in response to thrombin but not to bradykinin. These data suggest that thrombin-induced mitogenesis requires both Gq and Gi2, whereas bradykinin needs only the former. Thus, different growth factors operating upon the same cell type use overlapping yet distinct sets of G alpha subtypes in mitogenic signal transduction pathways. The direct identification of the coupling of both a pertussis toxin sensitive and insensitive G protein subtype in the mitogenic pathways utilized by thrombin offers an in vivo biochemical clarification of previous results obtained by pharmacologic studies.

Positional cloning of the gene for multiple endocrine neoplasia-type 1.

Multiple endocrine neoplasia-type 1 (MEN1) is an autosomal dominant familial cancer syndrome characterized by tumors in parathyroids, enteropancreatic endocrine tissues, and the anterior pituitary. DNA sequencing from a previously identified minimal interval on chromosome 11q13 identified several candidate genes, one of which contained 12 different frameshift, nonsense, missense, and in-frame deletion mutations in 14 probands from 15 families. The MEN1 gene contains 10 exons and encodes a ubiquitously expressed 2.8-kilobase transcript. The predicted 610-amino acid protein product, termed menin, exhibits no apparent similarities to any previously known proteins. The identification of MEN1 will enable improved understanding of the mechanism of endocrine tumorigenesis and should facilitate early diagnosis.

The G protein connection: molecular basis of membrane association.

Two distinct types of lipid modification, myristoylation and isoprenylation, are critical for membrane association of heterotrimeric G proteins. Elucidation of the molecular basis for G protein membrane association has important implications for understanding G protein structure and function, and is relevant to potential therapeutic approaches to AIDS and cancer.

Deficient guanine nucleotide regulatory unit activity in cultured fibroblast membranes from patients with pseudohypoparathyroidism type I. a cause of impaired synthesis of 3',5'-cyclic AMP by intact and broken cells.

Deficient activity of the guanine nucleotide regulatory protein (G unit), an integral component of the membrane-bound adenylate cyclase complex, has been implicated as the biochemical lesion in many patients with pseudohypoparathyroidism (PHP) type I. In addition to renal resistance to parathyroid hormone in this disorder, there is decreased responsiveness of diverse tissues to hormones that act via 3',5'-cyclic AMP (cAMP). To assess whether a deficiency of G units could account for impaired adenylate cyclase activity, we studied cAMP production in intact cultured fibroblasts and fibroblast plasma membranes from five patients with PHP in response to several activators of adenylate cyclase. The number of G units in PHP fibroblast membranes, measured by cholera toxin-dependent [(32)P]ADP ribosylation of G-unit peptides, as well as the G-unit activity, determined by the ability of detergent extracts to reconstitute adenylate cyclase activity in G-unit-deficient S49 CYC(-) membranes, were found to be markedly reduced compared with control membranes (43 and 40%, respectively), The activation of fibroblast membrane adenylate cyclase by effectors that act directly through the G unit (guanosine triphosphate, guanosine 5'-0-[3-thiotriphosphate] [GTP-gamma-S], NaF) was significantly greater in control membranes than in membranes from patients with PHP. Moreover, we found that hormone (prostaglandin E(1)) stimulated adenylate cyclase activity was also greater in control membranes than in PHP membranes. Neither the apparent affinity of membrane adenylate cyclase for GTP-gamma-S (apparent K(m) =5 X 10(-8) M) nor the rate of enzyme activation by GTP-gamma-S was significantly different in fibroblast membranes from control subjects and patients with PHP. In contrast to the notable differences in hormone and G-unit-activated adenylate cyclase shown in fibroblast membranes from PHP patients and control subjects, the intrinsic catalytic activity of membranes, as determined by forskolin-stimulated adenylate cyclase, was not significantly different in the two groups. Intact fibroblasts derived from patients with PHP accumulated significantly (P 0.001) less cAMP (46+/-21 pmol cAMP/mcg DNA, n = 5) than cells from normal individuals (170+/-51 pmol cAMP/mcg DNA, n = 11) when stimulated with PGE(1). PGE(1)-stimulated accumulation of cAMP by intact fibroblast monolayers correlated closely with PGE(1) plus GTP-activated membrane adenylate cyclase activity in both patients and controls (r = 0.97, P < 0.001). Our data show that, in patients with PHP, (a) fibroblast membranes show a decreased complement of G units, (b) membrane catalytic activity is normal, but adenylate cyclase activity is reduced when stimulated by hormone or by effectors which activate the G unit, (c) the ability of cells to accumulate cAMP in response to hormone stimulation is reduced, and (d) reduced membrane adenylate cyclase activity correlates well with impaired cellular cAMP synthesis. These results, taken together, indicate that a deficiency of G-unit activity can impair synthesis of cAMP by both intact and broken cells, and may explain the resistance of multiple tissues to hormones that act via cAMP observed in PHP.

Deficient adenylate cyclase regulatory protein in renal membranes from a patient with pseudohypoparathyroidism.

Recent studies have established that some patients with pseudohypoparathyroidism have a deficiency of the adenylate cyclase regulatory protein (the G unit) in plasma membranes from erythrocytes, platelets, and fibroblasts. We have directly measured the activity of the G unit in renal membranes from a patient with pseudohypoparathyroidism who, in addition to parathyroid hormone resistance, has resistance to thyrotropin and gonadotropins. Erythrocyte membrane G unit activity was 57% that of control erythrocyte membranes. Lubrol PX extracts of renal membranes had only 30% of the G unit activity of control renal membrane extracts, whether assayed with sodium fluoride or guanosine-5'-O-(3-thiotriphosphate) (GTP-gamma-S). In cholate extracts, the G unit activity was 37 and 48% of control with fluoride or GTP-gamma-S, respectively. Cholera toxin-dependent incorporation of [32P]ADP-ribose into the 42,000-Mr subunit of the G unit was decreased in renal membranes from the patient compared with control renal membranes. The data demonstrate that the membrane G unit deficiency in pseudohypoparathyroidism extends to the cells of a clinically relevant parathyroid hormone target tissue.

Reproduction of human fibrous dysplasia of bone in immunocompromised mice by transplanted mosaics of normal and Gsalpha-mutated skeletal progenitor cells.

We have isolated progenitor cells from the stromal system of the fibrous dysplastic marrow of patients with McCune-Albright Syndrome. Analysis of the Gsalpha gene from individual colonies provided direct evidence for the presence of two different genotypes within single fibrous dysplastic lesions: marrow stromal cells containing two normal Gsalpha alleles, and those containing one normal allele and an allele with an activating mutation. Transplantation of clonal populations of normal cells into the subcutis of immunocompromised mice resulted in normal ossicle formation. In contrast, transplantation of clonal populations of mutant cells always led to the loss of transplanted cells from the transplantation site and no ossicle formation. However, transplantation of a mixture of normal and mutant cells reproduced an abnormal ectopic ossicle recapitulating human fibrous dysplasia and providing an in vivo cellular model of this disease. These results provide experimental evidence for the necessity of both normal and mutant cells in the development of McCune-Albright Syndrome fibrous dysplastic lesions in bone.

Urinary calcium excretion in familial hypocalciuric hypercalcemia. Persistence of relative hypocalciuria after induction of hypoparathyroidism.

Familial hypocalciuric hypercalcemia (FHH) is an autosomal dominant trait comprising hypercalcemia, hypophosphatemia, parathyroid hyperplasia, and unusually low renal clearance of calcium. We evaluated the role of parathyroid hormone in the relative hypocalciuria of FHH and characterized the renal transport of calcium in this disorder using three previously hypercalcemic FHH patients with surgical hypoparathyroidism and three controls with surgical hypoparathyroidism. Intravenous infusion of calcium chloride in two patients with FHH and in three controls increased serum calcium from a mean basal of 5.0 to a mean peak of 6.8 meq/liter in two FHH patients and from 4.2 to 5.7 in three control subjects. Urinary calcium in a third FHH patient was studied without calcium infusion during recovery from hypercalcemia of vitamin D intoxication. At all serum concentrations of calcium, calcium clearance was lower in FHH than in controls; at base-line serum calcium, the ratio of calcium clearance to inulin clearance (C(Ca)/C(IN)) in FHH subjects was 32% of that in controls and decreased to 19% during hypercalcemia. Calcium infusion increased the ratio of sodium clearance to inulin clearance in controls from a base line of 0.020 to 0.053 at peak concentrations of calcium in serum, but did not affect this parameter in FHH (0.017 at base-line serum calcium vs. 0.019 at peak). When calcium infusion studies were performed (in two patients with FHH and one control) during administration of acetazolamide, a drug whose principal renal action causes inhibition of proximal transport of solute, C(Ca)/C(IN) in the patients with FHH was 29 and 7% of that of the control at base-line and peak serum calcium, respectively. In contrast, ethacrynic acid, a diuretic that acts in the ascending limb of the loop of Henle, increased C(Ca)/C(IN) more in the FHH patients than in the control subject; C(Ca)/C(IN) was 65% at base-line and 47% at peak serum calcium, compared with that of the control subject. The greater calciuric response to ethacrynic acid than to acetazolamide or calcium infusion alone in FHH indicates that a major renal locus of abnormal calcium transport in this disorder may be the ascending limb of the loop of Henle.Decreased clearance of calcium in patients with FHH and hypoparathyroidism when compared with hypoparathyroid controls indicates that relative hypocalciuria in FHH is not dependent on hyperparathyroidism. Since the parathyroid glands in FHH are not appropriately suppressed by calcium, this implies that FHH represents a disorder of abnormal transport of, and/or response to, extracellular calcium in at least two organs, parathyroid gland and kidney.

Impaired formation of beta-adrenergic receptor-nucleotide regulatory protein complexes in pseudohypoparathyroidism.

Decreased activity of the guanine nucleotide regulatory protein (N) of the adenylate cyclase system is present in cell membranes of some patients with pseudohypoparathyrodism (PHP-Ia) whereas others have normal activity of N (PHP-Ib). Low N activity in PHP-Ia results in a decrease in hormone (H)-stimulatable adenylate cyclase in various tissues, which might be due to decreased ability to form an agonist-specific high affinity complex composed of H, receptor (R), and N. To test this hypothesis, we compared beta-adrenergic agonist-specific binding properties in erythrocyte membranes from five patients with PHP-Ia (N = 45% of control), five patients with PHP-Ib (N = 97%), and five control subjects. Competition curves that were generated by increasing concentrations of the beta-agonist isoproterenol competing with [125I]pindolol were shallow (slope factors less than 1) and were computer fit to a two-state model with corresponding high and low affinity for the agonist. The agonist competition curves from the PHP-Ia patients were shifted significantly (P less than 0.02) to the right as a result of a significant (P less than 0.01) decrease in the percent of beta-adrenergic receptors in the high affinity state from 64 +/- 22% in PHP-Ib and 56 +/- 5% in controls to 10 +/- 8% in PHP-Ia. The agonist competition curves were computer fit to a "ternary complex" model for the two-step reaction: H + R + N in equilibrium HR + N in equilibrium HRN. The modeling was consistent with a 60% decrease in the functional concentration of N, and was in good agreement with the biochemically determined decrease in erythrocyte N protein activity. These in vitro findings in erythrocytes taken together with the recent observations that in vivo isoproterenol-stimulated adenylate cyclase activity is decreased in patients with PHP (Carlson, H. E., and A. S. Brickman, 1983, J. Clin. Endocrinol. Metab. 56:1323-1326) are consistent with the notion that N is a bifunctional protein interacting with both R and the adenylate cyclase. It may be that in patients with PHP-Ia a single molecular and genetic defect accounts for both decreased HRN formation and decreased adenylate cyclase activity, whereas in PHP-Ib the biochemical lesion(s) appear not to affect HRN complex formation.

G-protein involvement in matrix-mediated motility and invasion of high and low experimental metastatic B16 melanoma clones.

Membranes from a B16 murine melanoma clone of high experimental metastatic capacity show increased amounts of pertussis toxin (PT) substrate when compared to a low metastatic counterpart. Using specific antibodies, we identified Gi2 as the PT-sensitive G-protein uniquely abundant in highly metastatic cells. ADP ribosylation of a G-protein alpha subunit by PT decreased both the migration of tumor cells through Matrigel (Collaborative Research, Bedford, MA) and the fibronectin-, laminin-, and collagen type IV-mediated motility of a high metastatic clone. Treatment of cells from a low metastatic clone with PT did not alter either the relatively low invasive capacity or lower motility of these cells. While cholera toxin treatment of cells resulted in decreased invasion and motility of both high and low metastatic clones, there were significant qualitative and quantitative differences, when compared to the PT effects, which indicated that the two toxins were acting on different second messenger systems. PT treatment of B16 clones of high or low experimental metastatic capacity does not result in any alteration in cellular cyclic AMP accumulation suggesting that the PT substrate is not linked with the adenylyl cyclase enzyme complex. The data suggest that a PT-sensitive G-protein, possibly Gi2, regulates second messenger pathways that contribute to the metastatic capacity of B16 melanoma cells.

Allelic loss from chromosome 11 in parathyroid tumors.

Parathyroid tumors may occur in a sporadic fashion or, more rarely, as part of a familial syndrome (such as familial multiple endocrine neoplasia type I). The MENI gene has been mapped by linkage analysis to chromosome 11 at band q11-q13, and presumably acts as a tumor suppressor gene. In the present study, which is an extension of our previous studies, we examined 41 parathyroid tumors from patients with familial multiple endocrine neoplasia type I and 61 sporadic parathyroid tumors with markers on chromosome 11, to assess the extent of allelic loss in those tumors. Twenty-four of the MENI-associated tumors (58%) and 16 of the sporadic parathyroid tumors (26%) displayed allelic loss from chromosome 11. The region of overlap of the allelic losses in the MENI-associated tumors enables us to place the MENI gene between PGA centromerically and INT2 telomerically, a region spanning about 7.5 cM. Taken together with locus ordering by linkage analysis, this clearly localizes the MENI gene telomeric to the PGA locus. Our inability to detect allelic loss on chromosome 11 in some parathyroid tumors suggests the existence of other genes involved in the development and/or progression of this subgroup of presumably monoclonal tumors; or that localized events involving the 11q tumor suppressor gene have occurred in some parathyroid tumors whose detection is beyond the sensitivity of our analysis; or that at least some of the specimens analyzed were in fact primarily hyperplastic parathyroid tissue.

Allelic deletions on chromosome 11q13 in multiple tumors from individual MEN1 patients.

Familial multiple endocrine neoplasia type 1 is an autosomal dominant hereditary disorder characterized by multiple parathyroid, pancreatic, duodenal, and pituitary tumors. The parathyroid tumors may arise as diffuse areas of hyperplasia, whereas the pancreatic and duodenal tumors usually form as discrete nodules. Except for a single report, tumor loss of heterozygosity (LOH) mapping of the putative MEN1 suppressor gene on chromosome 11q13 in the past has been restricted by analysis of a single tumor from individual patients and somatic cellular contamination. For this reason, it has not been possible to analyze the clonality of the emerging MEN1 neoplasms. Furthermore, it has been previously unknown whether the LOH pattern varies between individual MEN1 tumors in a given patient or among tumors of different histological origins within unrelated patients. To address these previous limitations, the present study introduces a refinement in microdissection in which endothelial cells are stained and selectively excluded. Tissue microdissection was applied to study LOH patterns on chromosome 11q13 using 8 polymorphic DNA markers in 44 different MEN1 tumors from parathyroid, pancreas, and duodenum in nine unrelated patients. In addition, X-chromosome inactivation clonal analysis was applied to 16 individual microdissected regions from seven parathyroid glands in three female patients. The LOH rates of parathyroid lesions (100%) and endocrine tumors of the pancreas (83%) were strikingly different from the LOH rate of gastrinomas (21%), suggesting that the mechanism that drives LOH may be influenced by the tissue context. Moreover, combined LOH and X-chromosome inactivation scoring of the same microdissected region revealed that parathyroid MEN1 neoplasms can consist of more than one clone. In this study, the centromeric boundary of the putative MEN1 gene was PYGM. Analysis of differential LOH patterns in multiple microdissected tumors in the same patient constitutes a novel approach to suppressor gene mapping.

Allelic deletions on chromosome 11q13 in multiple endocrine neoplasia type 1-associated and sporadic gastrinomas and pancreatic endocrine tumors.

Endocrine tumors (ETs) of pancreas and duodenum occur sporadically and as a part of multiple endocrine neoplasia type 1 (MEN1). The MEN1 tumor suppressor gene has been localized to chromosome 11q13 by linkage analysis but has not yet isolated. Previous allelic deletion studies in enteropancreatic ETs suggested MEN1 gene involvement in tumorigenesis of familial pancreatic ETs (nongastrinomas) and sporadic gastrinomas. However, only a few MEN1-associated duodenal gastrinomas and sporadic pancreatic nongastrinomas have been investigated. We used tissue microdissection to analyze 95 archival pancreatic and duodenal ETs and metastases from 50 patients for loss of heterozygosity (LOH) on 11q13 with 10 polymorphic markers spanning the area of the putative MEN1 gene. Chromosome 11q13 LOH was detected in 23 of 27 (85%) MEN1-associated pancreatic ETs (nongastrinomas), 14 of 34 (41%) MEN1-associated gastrinomas, 3 of 16 (19%) sporadic insulinomas, and 8 of 18 (44%) sporadic gastrinomas. Analysis of LOH on 11q13 showed different deletion patterns in ETs from different MEN1 patients and in multiple tumors from individual MEN1 patients. The present results suggest that the MEN1 gene plays a role in all four tumor types. The lower rate of 11q13 LOH in MEN1-associated and sporadic gastrinomas and sporadic insulinomas as compared to MEN1 nongastrinomas may reflect alternative genetic pathways for the development of these tumors or mechanisms of the MEN1 gene inactivation that do not involve large deletions. The isolation of the MEN1 gene is necessary to further define its role in pathogenesis of pancreatic and duodenal ETs.

Localization of the multiple endocrine neoplasia type I (MEN1) gene based on tumor loss of heterozygosity analysis.

Multiple endocrine neoplasia type I (MEN1) is an inherited syndrome that results in parathyroid, anterior pituitary, and pancreatic and duodenal endocrine tumors as well as foregut carcinoids in affected patients. The gene responsible for the disease has been linked to chromosome 11q13. We analyzed loss of heterozygosity (LOH) in 188 tumors from 81 patients in an attempt to further define the location of the MEN1 gene. Both tumors from MEN1 patients and corresponding sporadic tumors were analyzed. Tumor types included parathyroid, gastrinoma, pancreatic endocrine, pituitary, and lung carcinoid. Six tumors (three MEN1 and three sporadic tumors) were identified that provided important LOH boundaries. Four tumors (two parathyroid tumors, one gastrinoma, and one lung carcinoid tumor) showed allelic loss that placed the MEN1 gene distal to marker PYGM. Two tumors (one gastrinoma and one parathyroid tumor) showed an LOH boundary that placed the gene proximal to D11S449, one of which further moved the telomeric boundary to D11S4936. Taken together, the present data suggest that the MEN1 gene lies between PYGM and D11S4936, a region of approximately 300 kb on chromosome 11q13.

Haplotype analysis defines a minimal interval for the multiple endocrine neoplasia type 1 (MEN1) gene.

Multiple endocrine neoplasia type 1 (MEN1) is an inherited syndrome characterized by development of multiple endocrine tumors in affected individuals. The gene responsible for the disease has been mapped to chromosome 11q13 by linkage analysis, but the gene itself has not yet been identified. We allelotyped 33 affected individuals from an extensive MEN1 kindred using eight polymorphic markers located on chromosome 11q13, including two new markers (D11S4907 and D11S4908) that we derived and mapped to the SEA-D11S913 region. Analysis of affected individuals revealed two separate recombination events, providing new centromeric and telomeric boundaries for the MEN1 gene. The present data indicate the MEN1 gene is located between markers D11S1883 and D11S4907, an approximate 2 Mb region on chromosome 11q13.

Mutations of the MEN1 tumor suppressor gene in pituitary tumors.

Although pituitary adenomas are monoclonal proliferations, somatic mutations involving genes that govern cell proliferation or hormone production have been difficult to identify. The genetic etiology of most pituitary tumors, therefore, remains unknown. Pituitary adenomas can develop sporadically or as a part of multiple endocrine neoplasia type 1 (MEN1). Recently, the gene responsible for MEN1 was cloned. To elucidate the potential etiological role of the MEN1 gene in pituitary tumorigenesis, 39 sporadic pituitary adenomas from 38 patients and 1 pituitary adenoma from a familial MEN1 patient were examined for MEN1 gene mutations and allelic deletions. Four of 39 sporadic pituitary adenomas showed a deletion of one copy of the MEN1 gene, and a specific MEN1 gene mutation in the remaining gene copy was detected in 2 of these tumors. The corresponding germ-line sequence was normal in all sporadic cases. A specific MEN1 mutation was detected in a pituitary adenoma and corresponding germ-line DNA in a patient with familial MEN1. An allelic deletion of the remaining copy of the MEN1 gene was also found in the patient's tumor. Genetic alterations of the MEN1 gene represent a candidate pathogenetic mechanism of pituitary tumorigenesis. The data suggest that somatic MEN1 gene mutations and deletions play a causative role in the development of a subgroup of sporadic pituitary adenomas.

Somatic mutations of the MEN1 tumor suppressor gene in sporadic gastrinomas and insulinomas.

Gastrinomas and insulinomas are frequent in multiple endocrine neoplasia type 1 (MEN1). The MEN1 tumor suppressor gene was recently identified. To elucidate the etiological role of the MEN1 gene in sporadic enteropancreatic endocrine tumorigenesis, we analyzed tumors (28 gastrinomas and 12 insulinomas) from 40 patients for MEN1 gene mutations and allelic deletions. One copy of the MEN1 gene was found to be deleted in 25 of 27 (93%) sporadic gastrinomas and in 6 of 12 (50%) sporadic insulinomas. MEN1 gene mutations were identified in 9 of 27 (33%) sporadic gastrinomas and 2 of 12 (17%) insulinomas and were not seen in corresponding germ-line DNA sequence. A specific MEN1 mutation was detected in one gastrinoma and in the corresponding germ-line DNA of a patient who had no family history of MEN1. Somatic MEN1 gene mutations and deletions play a critical role in the tumorigenesis of sporadic gastrinomas and may also contribute to the development of a subgroup of insulinomas.