| 1. |
- Bausch, Birke, et al.
(author)
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Clinical Characterization of the Pheochromocytoma and Paraganglioma Susceptibility Genes SDHA, TMEM127, MAX, and SDHAF2 for Gene-Informed Prevention
- 2017
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In: JAMA Oncology. - : AMER MEDICAL ASSOC. - 2374-2437 .- 2374-2445. ; 3:9, s. 1204-1212
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Journal article (peer-reviewed)abstract
- IMPORTANCE Effective cancer prevention is based on accurate molecular diagnosis and results of genetic family screening, genotype-informed risk assessment, and tailored strategies for early diagnosis. The expanding etiology for hereditary pheochromocytomas and paragangliomas has recently included SDHA, TMEM127, MAX, and SDHAF2 as susceptibility genes. Clinical management guidelines for patients with germline mutations in these 4 newly included genes are lacking. OBJECTIVE To study the clinical spectra and age-related penetrance of individuals with mutations in the SDHA, TMEM127, MAX, and SDHAF2 genes. DESIGN, SETTING, AND PATIENTS This study analyzed the prospective, longitudinally followed up European-American-Asian Pheochromocytoma-Paraganglioma Registry for prevalence of SDHA, TMEM127, MAX, and SDHAF2 germline mutation carriers from 1993 to 2016. Genetic predictive testing and clinical investigation by imaging from neck to pelvis was offered to mutation-positive registrants and their relatives to clinically characterize the pheochromocytoma/paraganglioma diseases associated with mutations of the 4 new genes. MAIN OUTCOMES AND MEASURES Prevalence and spectra of germline mutations in the SDHA, TMEM127, MAX, and SDHAF2 genes were assessed. The clinical features of SDHA, TMEM127, MAX, and SDHAF2 disease were characterized. RESULTS Of 972 unrelated registrants without mutations in the classic pheochromocytoma- and paraganglioma-associated genes (632 female [65.0%] and 340 male [35.0%]; age range, 8-80; mean [SD] age, 41.0 [13.3] years), 58 (6.0%) carried germline mutations of interest, including 29 SDHA, 20 TMEM127, 8 MAX, and 1 SDHAF2. Fifty-three of 58 patients (91%) had familial, multiple, extra-adrenal, and/or malignant tumors and/or were younger than 40 years. Newly uncovered are 7 of 63 (11%) malignant pheochromocytomas and paragangliomas in SDHA and TMEM127 disease. SDHA disease occurred as early as 8 years of age. Extra-adrenal tumors occurred in 28 mutation carriers (48%) and in 23 of 29 SDHA mutation carriers (79%), particularly with head and neck paraganglioma. MAX disease occurred almost exclusively in the adrenal glands with frequently bilateral tumors. Penetrance in the largest subset, SDHA carriers, was 39% at 40 years of age and is statistically different in index patients (45%) vs mutation-carrying relatives (13%; P amp;lt; .001). CONCLUSIONS AND RELEVANCE The SDHA, TMEM127, MAX, and SDHAF2 genes may contribute to hereditary pheochromocytoma and paraganglioma. Genetic testing is recommended in patients at clinically high risk if the classic genes are mutation negative. Gene-specific prevention and/or early detection requires regular, systematic whole-body investigation.
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| 2. |
- Harnevik, Lotta, 1977-
(author)
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Molecular genetic studies on cystinuria
- 2007
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Doctoral thesis (other academic/artistic)abstract
- Cystinuria is defined as an inherited disorder characterized by increased urinary excretion of cystine and the dibasic amino acids arginine, lysine and ornithine. The only clinical manifestation of cystinuria is renal cystine stone formation due to the low solubility of cystine in the urine. Cystinuria can be attributed to mutations in the SLC3A1 and SLC7A9 genes in the majority of all cases and it has been a common expectation that molecular genetic studies of cystinuria would aid in understanding of the varying clinical outcome seen in the disease. Besides human, the disease has been most extensively studied in the domestic dog.The present study was undertaken to investigate the molecular genetic basis of cystinuria in patients from Sweden and to correlate genetic findings with phenotypes produced regarding cystine and dibasic amino acid excretion. Further, attempts were made to elucidate the molecular genetics of cystinuria in the dog.The entire coding sequences of the SLC3A1 and SLC7A9 genes were analysed by means of SSCA and DNA sequencing in 53 cystinuria patients and genetic findings were related to urinary excretion of cystine and dibasic amino acids in a subset of the patient group. We detected a total number of 22 different mutations in the SLC3A1 and SLC7A9 genes, 18 of which were described for the first time. We have found a probable genetic cause of cystinuria in approximately 74 % of our patients and a possible contribution to the disease in another 19 %. Mutations in the SLC3A1 gene is the major cause of cystinuria in our group, with only a minor contribution of SLC7A9 mutations. The group of patients presenting SLC3A1 mutations in a heterozygous state or lacking mutations in both genes had higher values of total urinary cystine and dibasic amino acids compared to patients homozygous for SLC3A1 mutations. The reason for this discrepancy remains unclear, but the possible impact of medical treatment with sulfhydryl compounds on total cystine values was ruled out.Sequencing of the full-length canine SLC7A9 cDNA was accomplished using the RACE technology and results from mutation analyses of SLC7A9 and SLC3A1 in cystinuric dogs showed that only two out of 13 dogs have mutations with possible impact on protein function in these genes. DNA sequencing was used for all exons of both genes in the dog, and in human cystinuria patients, all samples lacking mutations or showing heterozygosity after SSCA screening were sequenced in both genes as well. This implies that all point mutations present have been detected, but the possibility of mutations escaping PCR based methods as well as mutations in regulatory parts of the SLC3A1 and SLC7A9 genes remains in cases lacking a full molecular genetic explanation of the disease.Finally, clinical and genetic data from our study of cystinuria both in man and dog exemplifies that manifestation and clinical severity of cystinuria is not determined by genetic alterations in the SLC3A1 and SLC7A9 alone. Environmental factors, congenital malformations and modulating genetic factors are all possible contributors to the clinical outcome of cystinuria.
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| 3. |
- Kugelberg, Johan, et al.
(author)
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Role of SDHAF2 and SDHD in von Hippel-Lindau Associated Pheochromocytomas
- 2014
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In: World Journal of Surgery. - : Springer Verlag (Germany). - 0364-2313 .- 1432-2323. ; 38:3, s. 724-732
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Journal article (peer-reviewed)abstract
- Background Pheochromocytomas (PCCs) develop from the adrenal medulla and are often part of a hereditary syndrome such as von Hippel-Lindau (VHL) syndrome. In VHL, only about 30 % of patients with a VHL missense mutation develop PCCs. Thus, additional genetic events leading to formation of such tumors in patients with VHL syndrome are sought. SDHAF2 (previously termed SDH5) and SDHD are both located on chromosome 11q and are required for the function of mitochondrial complex II. While SDHAF2 has been shown to be mutated in patients with paragangliomas (PGLs), SDHD mutations have been found both in patients with PCCs and in patients with PGLs. Materials and methods Because loss of 11q is a common event in VHL-associated PCCs, we aimed to investigate whether SDHAF2 and SDHD are targets. In the present study, 41 VHL-associated PCCs were screened for mutations and loss of heterozygosity (LOH) in SDHAF2 or SDHD. Promoter methylation, as well as mRNA expression of SDHAF2 and SDHD, was studied. In addition, immunohistochemistry (IHC) of SDHB, known to be a universal marker for loss of any part the SDH complex, was conducted. Results and conclusions LOH was found in more than 50 % of the VHL-associated PCCs, and was correlated with a significant decrease (p less than 0.05) in both SDHAF2 and SDHD mRNA expression, which may be suggestive of a pathogenic role. However, while SDHB protein expression as determined by IHC in a small cohort of tumors was lower in PCCs than in the surrounding adrenal cortex, there was no obvious correlation with LOH or the level of SDHAF2/SDHD mRNA expression. In addition, the lack of mutations and promoter methylation in the investigated samples indicates that other events on chromosome 11 might be involved in the development of PCCs in association with VHL syndrome.
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| 4. |
- Rossitti, Hugo, et al.
(author)
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Activation of RAS Signalling is Associated with Altered Cell Adhesion in Phaeochromocytoma
- 2020
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In: International Journal of Molecular Sciences. - : MDPI. - 1661-6596 .- 1422-0067. ; 21:21
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Journal article (peer-reviewed)abstract
- Phaeochromocytomas and paragangliomas (PPGLs) are neuroendocrine catecholamine-producing tumours that may progress into inoperable metastatic disease. Treatment options for metastatic disease are limited, indicating a need for functional studies to identify pharmacologically targetable pathophysiological mechanisms, which require biologically relevant experimental models. Recently, a human progenitor phaeochromocytoma cell line named "hPheo1" was established, but its genotype has not been characterised. Performing exome sequencing analysis, we identified a KIF1B T827I mutation, and the oncogenic NRAS Q61K mutation. While KIF1B mutations are recurring somatic events in PPGLs, NRAS mutations have hitherto not been detected in PPGLs. Therefore, we aimed to assess its implications for the hPheo1 cell line, and possible relevance for the pathophysiology of PPGLs. We found that transient downregulation of NRAS in hPheo1 led to elevated expression of genes associated with cell adhesion, and enhanced adhesion to hPheo1 cells extracellular matrix. Analyses of previously published mRNA data from two independent PPGL patient cohorts (212 tissue samples) revealed a subcluster of PPGLs featuring hyperactivated RAS pathway-signalling and under-expression of cell adhesion-related gene expression programs. Thus, we conclude that NRAS activity in hPheo1 decreases adhesion to their own extracellular matrix and mirrors a transcriptomic RAS-signalling-related phenomenon in PPGLs.
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| 5. |
- Stenman, Adam, et al.
(author)
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HRAS mutation prevalence and associated expression patterns in pheochromocytoma
- 2016
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In: Genes, Chromosomes and Cancer. - : WILEY-BLACKWELL. - 1045-2257 .- 1098-2264. ; 55:5, s. 452-459
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Journal article (peer-reviewed)abstract
- Pheochromocytomas (PCC) and abdominal paragangliomas (PGL) display a highly diverse genetic background and recent gene expression profiling studies have shown that PCC and PGL (together PPGL) alter either kinase signaling pathways or the pseudo-hypoxia response pathway dependent of the genetic composition. Recurrent mutations in the Harvey rat sarcoma viral oncogene homolog (HRAS) have recently been verified in sporadic PPGLs. In order to further establish the HRAS mutation frequency and to characterize the associated expression profiles of HRAS mutated tumors, 156 PPGLs for exon 2 and 3 hotspot mutations in the HRAS gene was screened, and compared with microarray-based gene expression profiles for 93 of the cases. The activating HRAS mutations G13R, Q61R, and Q61K were found in 10/142 PCC (7.0%) and a Q61L mutation was revealed in 1/14 PGL (7.1%). All HRAS mutated cases included in the mRNA expression profiling grouped in Cluster 2, and 21 transcripts were identified as altered when comparing the mutated tumors with 91 HRAS wild-type PPGL. Somatic HRAS mutations were not revealed in cases with known PPGL susceptibility gene mutations and all HRAS mutated cases were benign. The HRAS mutation prevalence of all PPGL published up to date is 5.2% (49/950), and 8.8% (48/548) among cases without a known PPGL susceptibility gene mutation. The findings support a role of HRAS mutations as a somatic driver event in benign PPGL without other known susceptibility gene mutations. HRAS mutated PPGL cluster together with NF1- and RET-mutated tumors associated with activation of kinase-signaling pathways.
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| 6. |
- Stenman, Adam, et al.
(author)
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Molecular Profiling of Pheochromocytoma and Abdominal Paraganglioma Stratified by the PASS Algorithm Reveals Chromogranin B as Associated With Histologic Prediction of Malignant Behavior
- 2019
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In: American Journal of Surgical Pathology. - : LIPPINCOTT WILLIAMS & WILKINS. - 0147-5185 .- 1532-0979. ; 43:3, s. 409-421
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Journal article (peer-reviewed)abstract
- Pheochromocytomas (PCCs) and abdominal paragangliomas (PGLs), collectively abbreviated PPGL, are believed to exhibit malignant potential-but only subsets of cases will display full-blown malignant properties. The Pheochromocytoma of the Adrenal Gland Scaled Score (PASS) algorithm is a proposed histologic system to detect potential for aggressive behavior, but little is known regarding the coupling to underlying molecular genetics. In this study, a total of 92 PPGLs, previously characterized for susceptibility gene status and mRNA expressional profiles, were histologically assessed using the PASS criteria. A total of 32/92 PPGLs (35%) exhibited a PASS score amp;gt;= 4, including all 8 cases with malignant behavior (7 with known metastases and 1 with extensively infiltrative local recurrence). Statistical analyzes between expressional data and clinical parameters as well as individual PASS criteria yielded significant associations to Chromogranin B (CHGB), BRCA2, HIST1H3B, BUB1B, and RET to name a few, and CHGB had the strongest correlation to both PASS and metastasis/local recurrence of all analyzed genes. Evident CHGB downregulation was observed in PPGLs with high PASS and overtly malignant behavior, and was also associated with shorter disease-related survival. This finding was validated using quantitative real-time polymerase chain reaction, in which CHGB expression correlated with both PASS and metastasis/local recurrence with consistent findings obtained in the TCGA cohort. Moreover, immunohistochemical analyses of subsets of tumors showed a correlation between high PASS scores and negative or weak CHGB protein expression. Patients with PPGLs obtaining high PASS scores postoperatively, also exhibited low preoperative plasma levels of CHGB. These data collectively point out CHGB as a possible preoperative and postoperative marker for PPGLs with potential for aggressive behavior.
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| 7. |
- Welander, Jenny, et al.
(author)
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Frequent EPAS1/HIF2 alpha exons 9 and 12 mutations in non-familial pheochromocytoma
- 2014
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In: Endocrine-Related Cancer. - : BioScientifica. - 1351-0088 .- 1479-6821. ; 21:3, s. 495-504
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Journal article (peer-reviewed)abstract
- Pheochromocytomas are neuroendocrine tumors arising from the adrenal medulla. While heritable mutations are frequently described, less is known about the genetics of sporadic pheochromocytoma. Mutations in genes involved in the cellular hypoxia response have been identified in tumors, and recently EPAS1, encoding HIF2 alpha, has been revealed to be a new gene involved in the pathogenesis of pheochromocytoma and abdominal paraganglioma. The aim of this study was to further characterize EPAS1 alterations in non-familial pheochromocytomas. Tumor DNA from 42 adrenal pheochromocytoma cases with apparently sporadic presentation, without known hereditary mutations in predisposing genes, were analyzed for mutations in EPAS1 by sequencing of exons 9 and 12, which contain the two hydroxylation sites involved in HIF2a degradation, and also exon 2. In addition, the copy number at the EPAS1 locus as well as transcriptome-wide gene expression were studied by DNA and RNA microarray analyses, respectively. We identified six missense EPAS1 mutations, three in exon 9 and three in exon 12, in five of 42 pheochromocytomas (12%). The mutations were both somatic and constitutional, and had no overlap in 11 cases (26%) with somatic mutations in NF1 or RET. One sample had two different EPAS1 mutations, shown by cloning to occur in cis, possibly indicating a novel mechanism of HIF2a stabilization through inactivation of both hydroxylation sites. One of the tumors with an EPAS1 mutation also had a gain in DNA copy number at the EPAS1 locus. All EPAS1-mutated tumors displayed a pseudo-hypoxic gene expression pattern, indicating an oncogenic role of the identified mutations.
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| 8. |
- Welander, Jenny, et al.
(author)
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Integrative genomics reveals frequent somatic NF1 mutations in sporadic pheochromocytomas
- 2012
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In: Human Molecular Genetics. - : Oxford University Press (OUP): Policy B. - 0964-6906 .- 1460-2083. ; 21:26, s. 5406-5416
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Journal article (peer-reviewed)abstract
- Pheochromocytomas are neuroendocrine tumors of the adrenal medulla which can occur either sporadically or in the context of hereditary tumor syndromes. Whereas the genetic background of hereditary pheochromocytomas is becoming rather well-defined, very little is known about the more common sporadic form of the disease which constitutes approximate to 70 of all cases. In this study, we elucidate some of the molecular mechanisms behind sporadic pheochromocytoma by performing a comprehensive analysis of copy number alterations, gene expression, promoter methylation and somatic mutations in the genes RET, VHL, NF1, SDHA, SDHB, SDHC, SDHD, SDHAF2, KIF1B, TMEM127 and MAX, which have been associated with hereditary pheochromocytoma or paraganglioma. Our genomic and genetic analyses of 42 sporadic pheochromocytomas reveal that a large proportion (83) has an altered copy number in at least one of the known susceptibility genes, often in association with an altered messenger RNA (mRNA) expression. Specifically, 11 sporadic tumors (26) displayed a loss of one allele of the NF1 gene, which significantly correlated with a reduced NF1 mRNA expression. Subsequent sequencing of NF1 mRNA, followed by confirmation in the corresponding genomic DNA (gDNA), revealed somatic truncating mutations in 10 of the 11 tumors with NF1 loss. Our results thus suggest that the NF1 gene constitutes the most frequent (24) target of somatic mutations so far known in sporadic pheochromocytomas.
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| 9. |
- Welander, Jenny, et al.
(author)
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Rare Germline Mutations Identified by Targeted Next-Generation Sequencing of Susceptibility Genes in Pheochromocytoma and Paraganglioma
- 2014
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In: Journal of Clinical Endocrinology and Metabolism. - : Endocrine Society. - 0021-972X .- 1945-7197. ; 99:7, s. E1352-E1360
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Journal article (peer-reviewed)abstract
- Context: Pheochromocytomas and paragangliomas have a highly diverse genetic background, with a third of the cases carrying a germline mutation in 1 of 14 identified genes. Objective: This study aimed to evaluate next-generation sequencing for more efficient genetic testing of pheochromocytoma and paraganglioma and to establish germline and somatic mutation frequencies for all known susceptibility genes. Design: A targeted next-generation sequencing approach on an Illumina MiSeq instrument was used for a mutation analysis in 86 unselected pheochromocytoma and paraganglioma tumor samples. The study included the genes EGLN1, EPAS1, KIF1B beta, MAX, MEN1, NF1, RET, SDHA, SDHB, SDHC, SDHD, SDHAF2, TMEM127, and VHL. Results were verified in tumor and constitutional DNA with Sanger sequencing. Results: In all cases with clinical syndromes or known germline mutations, a mutation was detected in the expected gene. Among 68 nonfamilial tumors, 32 mutations were identified in 28 of the samples (41%), including germline mutations in EGLN1, KIF1B beta, SDHA, SDHB, and TMEM127 and somatic mutations in EPAS1, KIF1B beta, MAX, NF1, RET, and VHL, including one double monoallelic EPAS1 mutation. Conclusions: Targeted next-generation sequencing proved to be fast and cost effective for the genetic analysis of pheochromocytoma and paraganglioma. More than half of the tumors harbored mutations in the investigated genes. Notably, 7% of the apparently sporadic cases carried germline mutations, highlighting the importance of comprehensive genetic testing. KIF1B beta, which previously has not been investigated in a large cohort, appears to be an equally important tumor suppressor as MAX and TMEM127 and could be considered for genetic testing of these patients.
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