Marsh, DJ (Deborah J)
Latest papers:
Goswin Y Meyer-Rochow,
Janine M Smith,
Anne-Louise Richardson,
Deborah J Marsh,
Stan B Sidhu,
Bruce G Robinson,
Diana E Benn
Cancer Genetics, Hormones, and Cancer Group, Kolling Institute of Medical Research, Royal North Shore Hospital and University of Sydney, Sydney, Australia; University of Sydney Department of Endocrine Surgery, Royal North Shore Hospital, St Leonards, Australia.
BACKGROUND: Pheochromocytomas are neuroendocrine tumors of chromaffin cell origin which arise from the adrenal medulla and less commonly the extra-adrenal sympathetic paraganglia. Pheochromocytomas are component tumors of the familial syndromes multiple endocrine neoplasia Type 2, von Hippel Lindau disease, Neurofibromatosis Type 1, and the pheochromocytoma/paraganglioma syndromes caused by mutations in the RET, VHL, NF1, SDHB, and SDHD genes, respectively. The aim of this study was to evaluate denaturing high performance liquid chromatography (dHPLC) as a screening tool for the detection of germline mutations within VHL, SDHB, and SDHD in pheochromocytoma patients. METHODS: Polymerase chain reaction of all exons of VHL, SDHB, and SDHD genes was performed on leukocyte DNA extracted from stored blood samples of 74 unrelated patients treated for pheochromocytoma. After dHPLC analysis, all samples demonstrating variance were selected for sequencing. RESULTS: Of the 74 patients, 12 mutations and 16 polymorphisms were identified by dHPLC and confirmed on sequencing. More specifically, a total of 5 mutations and 15 polymorphisms were detected in SDHB and 7 mutations and 1 polymorphism were identified in VHL. No SDHD mutations or polymorphisms were identified. By sequencing only dHPLC variants, the total amount of DNA sequencing required was reduced by approximately 88%. CONCLUSIONS: dHPLC is an effective screening tool for the detection of germline mutations in SDHB, SDHD, and VHL and has application for diagnostic germline mutation analysis in pheochromocytoma patients.
Viive M Howell,
Anthony Gill,
Adele Clarkson,
Anne E Nelson,
Robert Dunne,
Leigh W Delbridge,
Bruce G Robinson,
Bin Tean Teh,
Oliver Gimm,
Deborah J Marsh
Kolling Institute of Medical Research, University of Sydney, Royal North Shore Hospital, St Leonards, Australia; Department of Anatomical Pathology, Royal North Shore Hospital, St Leonards, Australia and University of Sydney; Commonwealth Scientific and Industrial Research Organisation, Mathematical and Information Sciences, North Ryde, Australia; University of Sydney Endocrine Surgical Unit, Royal North Shore Hospital, St. Leonards, Australia; Van Andel Research Institute, Grand Rapids, MI, USA; Department of Surgery, University Hospital Linköping, Linköping, Sweden.
Context: Parafibromin, encoded by HRPT2, is the first marker with significant benefit in the diagnosis of parathyroid carcinoma. However, as parafibromin is only involved in up to 70% of parathyroid carcinomas and loss of parafibromin immunoreactivity may not be observed in all cases of HRPT2 mutation, a complementary marker is needed. Objective: We sought to determine the efficacy of increased expression of protein gene product 9.5 (PGP9.5), encoded by UCHL1 (Ubiquitin carboxyl-terminal esterase L1) as an additional marker to loss of parafibromin immunoreactivity for the diagnosis of parathyroid carcinoma. Design: In total, 146 parathyroid tumor and 9 normal tissues were analyzed for the expression of parafibromin and PGP9.5 using immunohistochemistry and for UCHL1 by quantitative RT-PCR. These samples included 6 hyperparathyroidism-jaw tumor syndrome related tumors and 24 sporadic carcinomas. Results: In tumors with evidence of malignancy, strong staining for PGP9.5 had a sensitivity of 78% for the detection of parathyroid carcinoma and/or HRPT2 mutation and a specificity of 100%. Complete lack of nuclear parafibromin staining had a sensitivity of 67% and a specificity of 100%. PGP9.5 was positive in a tumor with the HRPT2 mutation L64P that expressed parafibromin. Further, UCHL1 was highly expressed in the carcinoma/hyperparathyroidism-jaw tumor syndrome group compared to normal (p < 0.05) and benign specimens (p < 0.001). Conclusion: These results suggest that positive staining for PGP9.5 has utility as a marker for parathyroid malignancy, with a slightly superior sensitivity (p=0.03) and similar high specificity to that of parafibromin.
Most cited papers:
Diana E Benn,
Anne-Paule Gimenez-Roqueplo,
Jennifer R Reilly,
Jérôme Bertherat,
John Burgess,
Karen Byth,
Michael Croxson,
Patricia L M Dahia,
Marianne Elston,
Oliver Gimm,
David Henley,
Philippe Herman,
Victoria Murday,
Patricia Niccoli-Sire,
Janice L Pasieka,
Vincent Rohmer,
Kathy Tucker,
Xavier Jeunemaitre,
Deborah J Marsh,
Pierre-François Plouin,
Bruce G Robinson
Cancer Genetics, Kolling Institute of Medical Research, Royal North Shore Hospital and University of Sydney, Sydney, Australia (DE Benn PhD, JR Reilly BComm, DJ Marsh PhD, BG Robinson MD, MSc, FRACP) and Department of Endocrinology, Royal North Shore Hospital, Sydney, Australia (BG Robinson); Hereditary Cancer Clinic, Prince of Wales Hospital and School of Medicine, University of New South Wales, Sydney, New South Wales, Australia (K Tucker MBBS FRACP); NHMRC Clinical Trials Centre, University of Sydney, Australia (K Byth PhD); Department of Endocrinology, Royal Hobart Hospital, Tasmania, Australia (J Burgess MD FRACP); Department of Endocrinology and Diabetes, Sir Charles Gairdner Hospital, Nedlands, Western Australia, Australia (D Henley MBBS (Hons); Department of Endocrinology, Greenlane Clinical Centre, Auckland, New Zealand (M Croxson MBChB FRACP); Department of Endocrinology, Waikato Hospital, Hamilton, New Zealand (M Elston MBChB); Department of Genetics (AP Gimenez-Roqueplo MD PhD, X Jeunemaitre MD PhD) and Department of Hypertension (PF Plouin MD PhD), Hôpital Européen Georges Pompidou, Assistance Publique des Hôpitaux de Paris, INSERM U36, Collège de France, University Paris 5, France; Department of Otorhinolaryngology and Head and Neck Surgery, Hôpital Lariboisière, Assistance Publique des Hopitaux de Paris, France (P Herman, MD PhD); Department of Endocrinology, INSERM U567, Hôpital Cochin Assistance Publique des Hôpitaux de Paris, University Paris 5, France (J Bertherat MD PhD); Department of Endocrinology, Hôpital de la Timone, Assistance Publique des Hôpitaux de Marseille, France (P Niccoli-Sire MD); Department of Endocrinology, Hôpital d'Angers, Angers, France (V Rohmer MD); PGL. NET network (AP Gimenez-Roqueplo, J Bertherat, X Jeunemaitre, P Niccoli-Sire, V Rohmer, P Herman, PF Plouin); COMETE network (AP Gimenez-Roqueplo, J Bertherat, X Jeunemaitre, PF Plouin); Department of General, Visceral and Vascular Surgery, Martin-Luther-University Halle-Wittenberg, Halle, Germany (O Gimm MD); West of Scotland, Regional Genetics Services, Yorkhill, Scotland (V Murday BSc FRCP); Department of Surgery, Faculty of Medicine, University of Calgary, Calgary, Canada (J Pasieka MD FRCSC FACS); Departments Medicine and Cellular & Structural Biology, University of Texas Health Science Center, San Antonio, Texas, USA (P Dahia MD PhD).
Context: The identification of succinate dehydrogenase (SDH) gene mutations in Pheochromocytoma/Paraganglioma syndromes has necessitated clear elucidation of genotype-phenotype associations. Objective: To determine genotype-phenotype associations in a cohort of patients with Pheochromocytoma/Paraganglioma syndromes and succinate dehydrogenase subunit B (SDHB) or subunit D (SDHD) mutations. Design, Setting and Participants: The International SDH Consortium studied 116 individuals (83 affected and 33 clinically unaffected) from 62 families with Pheochromocytoma/Paraganglioma syndromes and SDHB or SDHD mutations. Clinical data were collected between August 2003 to September 2004 from tertiary referral centers in Australia, France, New Zealand, Germany, United States of America, Canada and Scotland. Main Outcome Measures: Data were collected on patients with pheochromocytomas and/or paragangliomas, with respect to onset of disease, diagnosis, genetic testing, surgery, pathology and disease progression. Clinical features were evaluated for evidence of genotype-phenotype associations and penetrance was determined. Results: SDHB mutation carriers were more likely than SDHD mutation carriers to develop extra-adrenal pheochromocytomas and malignant disease, whereas SDHD mutation carriers had a greater propensity to develop head and neck paragangliomas and multiple tumors. For the index cases there was no difference between 43 SDHB and 19 SDHD mutation carriers in the time to first diagnosis (34 yr vs. 28 yr respectively, P = 0.3). However, when all mutation carriers were included (n = 112), the estimated age-related penetrance was different for SDHB vs. SDHD mutation carriers (P = 0.008). Conclusions: For clinical follow up, features of SDHB mutation associated disease include a later age of onset, extra-adrenal (abdominal or thoracic) tumors and a higher rate of malignancy. In contrast, SDHD mutation carriers in addition to head and neck paragangliomas should be observed for multifocal tumors, infrequent malignancy and the possibility of extra-adrenal pheochromocytoma.
Diana E Benn,
Michael S Croxson,
Kathy Tucker,
Christopher P Bambach,
Anne Louise Richardson,
Leigh Delbridge,
Peter T Pullan,
Jeremy Hammond,
Deborah J Marsh,
Bruce G Robinson
Cancer Genetics, Kolling Institute of Medical Research, Royal North Shore Hospital, Syney, NSW, Australia.
Phaeochromocytomas arising in adrenal or extra-adrenal sites and paragangliomas of the head and neck, in particular of the carotid bodies, occur sporadically and also in a familial setting. In addition to mutations in RET and VHL in familial disease, germline mutations in SDHD and SDHB genes that encode subunits of mitochondrial complex II have also been associated with the development of familial phaeochromocytomas. To further investigate the role of SDHD and SDHB in the development of these tumours we determined the occurrence of germline SDHD and SDHB mutations in four patients with a family history of phaeochromocytoma with associated head and neck paraganglioma, one patient with a family history of phaeochromocytoma only and two patients with apparently sporadic extra-adrenal phaeochromocytoma, one of whom had early onset disease. Secondly, we investigated whether somatic SDHB mutations correlated with loss of heterozygosity at 1p36 in a subgroup of 11 sporadic and three MEN 2-associated RET-mutation-positive phaeochromocytomas. Novel SDHB mutations were identified in the probands from four families and two apparently sporadic cases (six of seven probands studied), including two missense mutations, a single nonsense and frameshift mutation, as well as two splice site mutations, one of which was shown to have partial penetrance resulting in 'leaky' splicing. Further, five intronic polymorphisms in SDHB were found. No SDHD mutations were identified. In addition, no somatic SDHB mutations were found in the remaining allele of the 11 sporadic adrenal phaeochromocytomas with allelic loss at 1p36 or the three MEN 2-associated RET-mutation-positive phaeochromocytomas. Therefore, we conclude that SDHB has a major role in the pathogenesis of familial phaeochromocytomas, but the possible role of SDHB in sporadic tumours showing allelic loss at 1p36 has yet to be ascertained.
Mesh-terms: Adolescent; Adrenal Gland Neoplasms :: enzymology; Adrenal Gland Neoplasms :: genetics; Adult; Age of Onset; Australia :: epidemiology; Child; Chromosomes, Human, Pair 1 :: genetics; DNA Mutational Analysis; DNA, Neoplasm :: genetics; Electron Transport Complex II; Frameshift Mutation; Germ-Line Mutation; Head and Neck Neoplasms :: enzymology; Head and Neck Neoplasms :: epidemiology; Head and Neck Neoplasms :: genetics; Human; Introns :: genetics; Loss of Heterozygosity; Male; Middle Aged; Multienzyme Complexes :: genetics; Multiple Endocrine Neoplasia :: enzymology; Multiple Endocrine Neoplasia :: epidemiology; Multiple Endocrine Neoplasia :: genetics; Mutation, Missense; Neoplasm Proteins :: genetics; Neoplasm Proteins :: physiology; Neoplastic Syndromes, Hereditary :: enzymology; Neoplastic Syndromes, Hereditary :: genetics; Oxidoreductases :: genetics; Paraganglioma :: enzymology; Paraganglioma :: epidemiology; Paraganglioma :: genetics; Pedigree; Pheochromocytoma :: enzymology; Pheochromocytoma :: epidemiology; Pheochromocytoma :: genetics; Protein Subunits :: deficiency; Protein Subunits :: genetics; Protein Subunits :: physiology; RNA Splice Sites :: genetics; Retroperitoneal Neoplasms :: enzymology; Retroperitoneal Neoplasms :: genetics; Succinate Dehydrogenase :: deficiency; Succinate Dehydrogenase :: genetics; Succinate Dehydrogenase :: physiology; Support, Non-U.S. Gov't;
Department of Molecular Medicine, Kolling Institute of Medical Research, Royal North Shore Hospital, University of Sydney, NSW 2065, Australia.
Parafibromin is a putative tumor suppressor encoded by HRPT2, mutations in which have been implicated in the familial tumor syndrome hyperparathyroidism jaw tumor syndrome (HPT-JT), and sporadic parathyroid carcinoma. Recently, parafibromin has been shown to be an accessory factor for RNA polymerase II as part of the human Paf 1 complex, suggesting, as has been shown for its yeast homologue (Cdc 73), that it may have a role as an important regulator of transcription. Parafibromin has also been shown to interact with a histone methyltransferase complex that methylates histone H3 and to inhibit proliferation when overexpressed in mammalian cell lines. Despite these findings, the cellular localization of parafibromin has been controversial, with reports of both nuclear and nucleocytoplasmic localization. We have expressed wild-type and mutant parafibromin tagged with enhanced green fluorescent protein and have identified a functional bipartite nuclear localization signal (NLS) at residues 125-139 (nucleotides 373-417), KRAADEVLAEAKKPR, that is evolutionarily conserved and critical for the nuclear localization of parafibromin. We have also shown that the C-terminal arm of this bipartite NLS plays the primary role in nuclear localization. In support of these findings, specific HRPT2 mutations identified in HPT-JT or sporadic parathyroid carcinoma predicted to truncate parafibromin upstream of or within this NLS disrupt nuclear localization.
Mesh-terms: Amino Acid Sequence; Animals; Base Sequence; Cell Line; DNA Primers; Genes, Tumor Suppressor; Humans; Molecular Sequence Data; Mutagenesis, Site-Directed; Nuclear Localization Signals; Parathyroid Neoplasms :: genetics; Research Support, Non-U.S. Gov't; Sequence Homology, Amino Acid; Tumor Suppressor Proteins :: genetics;
Patricia L M Dahia,
Ken N Ross,
Matthew E Wright,
César Y Hayashida,
Sandro Santagata,
Marta Barontini,
Andrew L Kung,
Gabriela Sanso,
James F Powers,
Arthur S Tischler,
Richard Hodin,
Shannon Heitritter,
Francis Moore,
Robert Dluhy,
Julie Ann Sosa,
I Tolgay Ocal,
Diana E Benn,
Deborah J Marsh,
Bruce G Robinson,
Katherine Schneider,
Judy Garber,
Seth M Arum,
Márta Korbonits,
Ashley Grossman,
Pascal Pigny,
Sérgio P A Toledo,
Vania Nosé,
Cheng Li,
Charles D Stiles
Department of Cancer Biology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts, United States of America.
Pheochromocytomas are neural crest-derived tumors that arise from inherited or sporadic mutations in at least six independent genes. The proteins encoded by these multiple genes regulate distinct functions. We show here a functional link between tumors with VHL mutations and those with disruption of the genes encoding for succinate dehydrogenase (SDH) subunits B (SDHB) and D (SDHD). A transcription profile of reduced oxidoreductase is detected in all three of these tumor types, together with an angiogenesis/hypoxia profile typical of VHL dysfunction. The oxidoreductase defect, not previously detected in VHL-null tumors, is explained by suppression of the SDHB protein, a component of mitochondrial complex II. The decrease in SDHB is also noted in tumors with SDHD mutations. Gain-of-function and loss-of-function analyses show that the link between hypoxia signals (via VHL) and mitochondrial signals (via SDH) is mediated by HIF1alpha. These findings explain the shared features of pheochromocytomas with VHL and SDH mutations and suggest an additional mechanism for increased HIF1alpha activity in tumors.
Centre for Hormone Research, Department of Endocrinology & Diabetes, Royal Children's Hospital, Parkville, Victoria, Australia.
OBJECTIVE: Germline mutations in succinate dehydrogenase (SDH)B, SDHC and SDHD, encoding three of the four subunits of mitochondrial complex II, have been implicated in the tumourigenesis of familial paragangliomas and phaeochromocytomas. Twenty-three SDHB mutations have been identified to date. PATIENTS: We present a novel missense SDHB exon 2 mutation (c.118 A > G; K40E) identified in an Australian family. The proband was diagnosed with phaeochromocytoma at an early age following an unexpected hypertensive crisis and was found to be SDHB mutation-positive. Subsequent genetic screening of 26 family members has identified 17 mutation-positive relatives. In addition to the proband, four mutation positive relatives were found to have clinical symptoms or a lesion and/or catecholamine excess after the identification of the mutation led to further evaluation. Both the proband and an uncle have required surgical removal of a tumour. CONCLUSIONS: This family indicates the importance of germline screening of first-degree relatives when a patient presents with an apparently sporadic extra adrenal phaeochromocytoma at a young age or whenever a patient with a nonsecretory paraganglioma is found.
Mesh-terms: Adrenal Gland Neoplasms :: genetics; Adrenal Gland Neoplasms :: surgery; Child; DNA Mutational Analysis; Female; Humans; Iron-Sulfur Proteins; Male; Mutation, Missense; Paraganglioma :: genetics; Paraganglioma :: surgery; Pedigree; Pheochromocytoma :: genetics; Pheochromocytoma :: surgery; Protein Subunits :: genetics; Succinate Dehydrogenase :: genetics;
Trisha Dwight,
Anne E Nelson,
George Theodosopoulos,
Anne Louise Richardson,
Diana L Learoyd,
Jeanette Philips,
Leigh Delbridge,
Jan Zedenius,
Bin T Teh,
Catharina Larsson,
Deborah J Marsh,
Bruce G Robinson
Cancer Genetics Unit, Royal North Shore Hospital, Sydney, Australia. trisha.dwight@cmm.ki.se
Multiple parathyroid tumors, as opposed to hyperplasia, have been reported in a subset of patients with sporadic primary hyperparathyroidism (PHPT). It is not clear whether these multiple tumors are representative of a neoplastic process or whether they merely represent hyperplasia that has affected the parathyroid glands differentially and resulted in asynchronous growth. The molecular genetic techniques of comparative genomic hybridization (CGH), loss of heterozygosity (LOH), and MEN1 mutation analysis were performed on a series of five patients with multiglandular PHPT, each of which had two parathyroid tumors removed. Analysis of these multiple parathyroid tumors from patients with PHPT revealed that independent genetic events were associated with the development of a subset of these tumors. The DNA sequence copy number changes, identified by CGH analyses, either involved different chromosomal regions in the paired glands of a patient (two patients), or those regions implicated in one gland were not changed in a second gland from the same patient (two patients). Each of the three patients exhibiting LOH demonstrated different changes between the paired glands. Where LOH was detected in one gland from a patient, the other gland from the same patient either exhibited no allelic loss or the loss detected was in another region. Each of the three tumors exhibiting LOH at 11q13 was found to contain a somatic MEN1 mutation in the remaining allele, however these mutations were not present in the germline or in the paired gland from the same patient. Although it is possible that a separate series of genetic changes has arisen randomly in two separate glands within the same individual, it seems more likely that the development of these multiple tumors has arisen because of the involvement of other unknown factors. These factors may be genetic [such as the involvement of one or more germline mutations in an unknown low-penetrance gene(s), germline mosaicism or alterations in calcium-sensing receptor gene(s)], epigenetic, physiological, or environmental.
Mesh-terms: Cell Transformation, Neoplastic :: genetics; Chromosome Aberrations; Chromosome Mapping; Disease Progression; Female; Human; Hyperparathyroidism :: genetics; Hyperparathyroidism :: pathology; Hyperparathyroidism :: surgery; Loss of Heterozygosity; Male; Middle Aged; Multiple Endocrine Neoplasia Type 1 :: genetics; Mutation; Nucleic Acid Hybridization; Parathyroid Neoplasms :: genetics; Parathyroid Neoplasms :: pathology; Parathyroid Neoplasms :: surgery; Support, Non-U.S. Gov't;
Laboratory of Cellular and Diagnostic Proteomics, Kolling Institute of Medical Research, University of Sydney, Royal North Shore Hospital, St. Leonards, New South Wales, Australia.
Ovarian cancer has the highest mortality among the gynecologic malignancies. The phosphatidylinositol 3-kinase (PI3K) pathway is frequently activated, leading to increased cell survival. This study aimed to identify secreted proteins regulated by the PI3K pathway in ovarian cancer cell lines. Surface-enhanced laser desorption-ionization time-of-flight mass spectrometry with cation-exchange protein-chips was used to analyze secreted proteins from five ovarian cancer cell lines (SKOV-3, PE01, OVCAR-3, OV167, and OV207). To activate the PI3K pathway, cells were treated with 50 ng/mL epidermal growth factor (EGF) with or without 10 mumol/L LY294002, a PI3K inhibitor. Proteins induced by EGF and inhibited by LY294002, in the m/z range 7,500 to 9,500, were purified chromatographically, identified by peptide mass fingerprinting and NH(2)-terminal sequencing, and confirmed by immunodepletion. Two immunologically related proteins, m/z approximately 8,385 and 8,922, were identified as truncated and intact forms, respectively, of interleukin 8, a chemokine previously shown to be elevated in serum of ovarian cancer patients. Another protein, m/z 7,866, was identified as CXC chemokine ligand 1 (CXCL1) or GRO-alpha, a chemokine associated with melanoma formation and some epithelial cancers. EGF-stimulated CXCL1 levels were variably decreased by mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase kinase and p38 MAPK inhibition in the five cell lines, but only LY294002 fully reversed the EGF effect in all cell lines. Immunoreactive CXCL1 levels in 160 conditioned media were highly correlated with corresponding peak intensities at m/z 7,866 by mass spectrometry, indicating the quantitative nature of these analyses. We conclude that proteomic analysis of cell models of human disease may facilitate the discovery of pathway-dependent proteins.(Cancer Res 2006; 66(3): 1376-83).
Cancer Genetics, Kolling Institute of Medical Research, Royal North Shore Hospital, St Leonards, NSW 2065, Australia. bgr@med.usyd.edu.au.
Genetic understanding of pheochromocytoma (PHEO) and paraganglioma (PGL) syndromes has recently expanded with the identification of the involvement of the mitochondrial complex II peptides, namely the succinate dehydrogenase subunit B (SDHB), subunit C (SDHC), and subunit D (SDHD). In patients with PHEO and/or PGL genetic testing for germline mutations in SDHD and SDHB has been recommended, in addition to the PHEO susceptibility genes VHL and RET. After careful clinical assessment of the patient, suspected familial disease may direct the clinician to the appropriate gene for testing. In the absence of obvious features of familial disease, the decision regarding the appropriate gene for testing is more difficult. Such testing can be costly and time consuming, but a rational prioritization of gene testing can streamline the process. Therefore in order to achieve this for apparently sporadic cases we propose a decision matrix based on site of tumor, functionality, and age at presentation.
Parafibromin is a putative tumor suppressor encoded by HRPT2 and implicated in parathyroid tumorigenesis. We previously reported a functional bipartite nuclear localization signal (NLS) at residues 125-139. We now demonstrate that parafibromin exhibits nucleolar localization, mediated by three nucleolar localization signals (NoLS) at resides 76-92, 192-194 and 393-409. These NoLS represent clusters of basic amino acids arginine and lysine, similar to those found in other nucleolar proteins, as well as being characteristic of NLSs. While parafibromin's bipartite NLS is the primary determinant of nuclear localization, it does not mediate nucleolar localization. In contrast, the three identified NoLSs play only a minor role in nuclear localization, but are critical for the nucleolar localization of parafibromin.
Viive M Howell,
John W Cardinal,
Anne-Louise Richardson,
Oliver Gimm,
Bruce G Robinson,
Deborah J Marsh
Kolling Institute of Medical Research, Royal North Shore Hospital, St. Leonards, NSW 2065, Australia. debbie_marsh@med.usyd.edu.au.
Familial hyperparathyroidism, a disease of the parathyroid glands, may occur in conjunction with pituitary and pancreatic tumors (multiple endocrine neoplasia type I), kidney and bone tumors (hyperparathyroidism jaw tumor syndrome), or alone (familial isolated hyperparathyroidism). This study describes the development and validation of rapid scanning for mutations in two tumor suppressor genes linked to familial hyperparathyroidism-MEN1 and HRPT2. Denaturing high-performance liquid chromatography mutation scanning for MEN1 was performed using a set of 10 amplicons covering the nine coding exons and flanking intronic regions and for HRPT2 using a set of three amplicons for exons 1, 2, and 7 and flanking intronic regions, in which 80% of the mutations identified to date are located. All 52 MEN1 mutations or polymorphisms, 46 known and six unknown, were successfully detected. Mutation detection in exon 9 was not confounded by the presence of the common polymorphism D418D. In addition, all 10 HRPT2 mutations were successfully detected, and a two-step approach was able to distinguish IVS2 common polymorphisms from exon 2 mutations. The development of rapid denaturing high performance liquid chromatography mutation scanning of MEN1 and HRPT2 facilitates a molecular diagnosis of the associated familial syndromes for both clinically affected and at-risk family members.