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Westin, Gunnar
Publications (10 of 71) Show all publications
Barazeghi, E., Gill, A. J., Sidhu, S., Norlen, O., Dina, R., Palazzo, F. F., . . . Westin, G. (2017). A role for TET2 in parathyroid carcinoma. Endocrine-Related Cancer, 24(7), 329-338.
Open this publication in new window or tab >>A role for TET2 in parathyroid carcinoma
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2017 (English)In: Endocrine-Related Cancer, ISSN 1351-0088, E-ISSN 1479-6821, Vol. 24, no 7, 329-338 p.Article in journal (Refereed) Published
Abstract [en]

Primary hyperparathyroidism (pHPT) is rarely caused by parathyroid carcinoma (PC, <1-5% of pHPT cases). The TET proteins oxidize the epigenetic mark 5-methylcytosine to 5-hydroxymethylcytosine (5hmC) and inactivation by mutation or epigenetic deregulation of TET1 and TET2 play important roles in various cancers. Recently, we found that 5hmC was severely reduced in all of the analyzed PCs and with deranged expression of TET1 for the majority of PCs. Here, we have examined the expression of the TET2 protein in 15 5hmC-negative PCs from patients who had local invasion or metastases. Cell growth and cell migratory roles for TET2 as well as epigenetic deregulated expression were addressed. Immunohistochemistry revealed very low/undetectable expression of TET2 in all PCs and verified for two PCs that were available for western blotting analysis. Knockdown of TET2 in the parathyroid cell line sHPT-1 resulted in increased cell growth and increased cell migration. DNA sequencing of TET2 in PCs revealed two common variants and no obvious inactivating mutations. Quantitative bisulfite pyrosequencing analysis of the TET2 promoter CpG island revealed higher CpG methylation level in the PCs compared to that in normal tissues and treatment of a PC primary cell culture with the DNA methylation inhibitor 5-aza-2'-deoxycytidine caused increased expression of the methylated TET2 gene. Hence, the data suggest that deregulated expression of TET2 by DNA hypermethylation may contribute to the aberrantly low level of 5hmC in PCs and further that TET2 plays a cell growth and cell migratory regulatory role and may constitute a parathyroid tumor suppressor gene.

Keyword
5-hydroxymethylcytosine, TET2, primary hyperparathyroidism, parathyroid carcinoma, promoter hypermethylation, tumor suppressor
National Category
Endocrinology and Diabetes Cancer and Oncology
Identifiers
urn:nbn:se:uu:diva-330022 (URN)10.1530/ERC-17-0009 (DOI)000404978400007 ()
Funder
Swedish Cancer Society
Available from: 2017-10-09 Created: 2017-10-09 Last updated: 2017-10-09Bibliographically approved
Edfeldt, K., Daskalakis, K., Bäcklin, C., Norlén, O., Tiensuu Janson, E., Westin, G., . . . Stålberg, P. (2017). DcR3, TFF3 and Midkine are Novel Serum Biomarkers in Small Intestinal Neuroendocrine Tumors. Neuroendocrinology, 105(2), 170-181.
Open this publication in new window or tab >>DcR3, TFF3 and Midkine are Novel Serum Biomarkers in Small Intestinal Neuroendocrine Tumors
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2017 (English)In: Neuroendocrinology, ISSN 0028-3835, E-ISSN 1423-0194, Vol. 105, no 2, 170-181 p.Article in journal (Refereed) Published
Abstract [en]

Small intestinal neuroendocrine tumors (SI-NETs) are amine- and peptide producing neoplasms. Most patients display metastases at the time of diagnosis, they have an unpredictable individual disease course and the tumors are often therapy resistant. Chromogranin A (CgA) and 5-hydroxyindoleacetic acid (5-HIAA) are the clinically most used biomarkers today, but there is a great need for novel diagnostic and prognostic biomarkers and new therapeutic targets. Sixty-nine biomarkers were screened in serum from 23 SI-NET patients and 23 healthy controls using multiplex PLA (proximity ligation assay). A refined method, PEA (proximity extension assay), was used to analyze 76 additional biomarkers. Statistical testing and multivariate classification were performed. Immunohistochemistry and ELISA assays were performed in an extended cohort. Using PLA, 19 biomarkers showed a significant difference in serum concentrations between patients and controls, and PEA revealed difference in concentrations in 13 proteins. Multivariate classification analysis revealed decoy receptor 3 (DcR3), trefoil factor 3 (TFF3) and Midkine to be good biomarkers for disease, which was confirmed by ELISA analysis. All three biomarkers were expressed in tumor tissue. DcR3 concentrations were elevated in patients with stage IV disease. High concentrations of DcR3 and TFF3 were correlated to poor survival. DcR3, TFF3 and Midkine exhibited elevated serum concentrations in SI-NET patients compared to healthy controls, and DcR3 and TFF3 were associated with poor survival. DcR3 seems to be a marker for liver metastases while TFF3 and Midkine may be new diagnostic biomarkers for SI-NETs.

National Category
Endocrinology and Diabetes
Identifiers
urn:nbn:se:uu:diva-307769 (URN)10.1159/000452891 (DOI)000407672700008 ()27829249 (PubMedID)
Funder
Swedish Cancer Society
Available from: 2016-11-21 Created: 2016-11-21 Last updated: 2017-11-17Bibliographically approved
Barazeghi, E., Gill, A. J., Sidhu, S., Norlen, O., Dina, R., Palazzo, F. F., . . . Westin, G. (2016). 5-Hydroxymethylcytosine discriminates between parathyroid adenoma and carcinoma. Clinical Epigenetics, 8, Article ID 31.
Open this publication in new window or tab >>5-Hydroxymethylcytosine discriminates between parathyroid adenoma and carcinoma
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2016 (English)In: Clinical Epigenetics, E-ISSN 1868-7083, Vol. 8, 31Article in journal (Refereed) Published
Abstract [en]

Background: Primary hyperparathyroidism is characterized by enlarged parathyroid glands due to an adenoma (80-85 %) or multiglandular disease (similar to 15 %) causing hypersecretion of parathyroid hormone (PTH) and generally hypercalcemia. Parathyroid cancer is rare (<1-5 %). The epigenetic mark 5-hydroxymethylcytosine (5hmC) is reduced in various cancers, and this may involve reduced expression of the ten-eleven translocation 1 (TET1) enzyme. Here, we have performed novel experiments to determine the 5hmC level and TET1 protein expression in 43 parathyroid adenomas (PAs) and 17 parathyroid carcinomas (PCs) from patients who had local invasion or metastases and to address a potential growth regulatory role of TET1. Results: The global 5hmC level was determined by a semi-quantitative DNA immune-dot blot assay in a smaller number of tumors. The global 5hmC level was reduced in nine PCs and 15 PAs compared to four normal tissue samples (p < 0.05), and it was most severely reduced in the PCs. By immunohistochemistry, all 17 PCs stained negatively for 5hmC and TET1 showed negative or variably heterogeneous staining for the majority. All 43 PAs displayed positive 5hmC staining, and a similar aberrant staining pattern of 5hmC and TET1 was seen in about half of the PAs. Western blotting analysis of two PCs and nine PAs showed variable TET1 protein expression levels. A significantly higher tumor weight was associated to PAs displaying a more severe aberrant staining pattern of 5hmC and TET1. Overexpression of TET1 in a colony forming assay inhibited parathyroid tumor cell growth. Conclusions: 5hmC can discriminate between PAs and PCs. Whether 5hmC represents a novel marker for malignancy warrants further analysis in additional parathyroid tumor cohorts. The results support a growth regulatory role of TET1 in parathyroid tissue.

Keyword
5-hydroxymethylcytosine, 5hmC, Parathyroid cancer, Primary hyperparathyroidism, TET1
National Category
Cancer and Oncology
Identifiers
urn:nbn:se:uu:diva-282795 (URN)10.1186/s13148-016-0197-2 (DOI)000371782000002 ()26973719 (PubMedID)
Funder
Swedish Cancer Society
Available from: 2016-04-14 Created: 2016-04-07 Last updated: 2017-11-30Bibliographically approved
Edfeldt, K., Hellman, P., Westin, G. & Stålberg, P. (2016). A plausible role for actin gamma smooth muscle 2 (ACTG2) in small intestinal neuroendocrine tumorigenesis. BMC Endocrine Disorders, 16(19).
Open this publication in new window or tab >>A plausible role for actin gamma smooth muscle 2 (ACTG2) in small intestinal neuroendocrine tumorigenesis
2016 (English)In: BMC Endocrine Disorders, ISSN 1472-6823, E-ISSN 1472-6823, Vol. 16, no 19Article in journal (Refereed) Published
Abstract [en]

BACKGROUND: Small intestinal neuroendocrine tumors (SI-NETs) originate from the enterochromaffin cells in the ileum and jejunum. The knowledge about genetic and epigenetic abnormalities is limited. Low mRNA expression levels of actin gamma smooth muscle 2 (ACTG2) have been demonstrated in metastases relative to primary SI-NETs. ACTG2 and microRNA-145 (miR-145) are aberrantly expressed in other cancers and ACTG2 can be induced by miR-145. The aim of this study was to investigate the role of ACTG2 in small intestinal neuroendocrine tumorigenesis.

METHODS: Protein expression was analyzed in SI-NETs (n = 24) and in enterochromaffin cells by immunohistochemistry. The cell line CNDT2.5 was treated with the histone methyltransferase inhibitor 3-deazaneplanocin A (DZNep), the selective EZH2 inhibitor EPZ-6438, or 5-aza-2'-deoxycytidine, a DNA hypomethylating agent. Cells were transfected with ACTG2 expression plasmid or miR-145. Western blotting analysis, quantitative RT-PCR, colony formation- and viability assays were performed. miR-145 expression levels were measured in tumors.

RESULTS: Eight primary tumors and two lymph node metastases displayed variable levels of positive staining. Fourteen SI-NETs and normal enterochromaffin cells stained negatively. Overexpression of ACTG2 significantly inhibited CNDT2.5 cell growth. Treatment with DZNep or transfection with miR-145 induced ACTG2 expression (>10-fold), but no effects were detected after treatment with EPZ-6438 or 5-aza-2'-deoxycytidine. DZNep also induced miR-145 expression. SI-NETs expressed relatively low levels of miR-145, with reduced expression in metastases compared to primary tumors.

CONCLUSIONS: ACTG2 is expressed in a fraction of SI-NETs, can inhibit cell growth in vitro, and is positively regulated by miR-145. Theoretical therapeutic strategies based on these results are discussed.

Keyword
SI NET; ACTG2; miR 145; Epigenetic regulation
National Category
Endocrinology and Diabetes
Identifiers
urn:nbn:se:uu:diva-287949 (URN)10.1186/s12902-016-0100-3 (DOI)000374739800001 ()27107594 (PubMedID)
Available from: 2016-04-27 Created: 2016-04-27 Last updated: 2017-11-30Bibliographically approved
Stålberg, P., Westin, G. & Thirlwell, C. (2016). Genetics and epigenetics in small intestinal neuroendocrine tumours. Journal of Internal Medicine, 280(6), 584-594.
Open this publication in new window or tab >>Genetics and epigenetics in small intestinal neuroendocrine tumours
2016 (English)In: Journal of Internal Medicine, ISSN 0954-6820, E-ISSN 1365-2796, Vol. 280, no 6, 584-594 p.Article in journal (Refereed) Published
Abstract [en]

Neuroendocrine tumour of the small intestine (SI-NET), formerly known as midgut carcinoid tumour, is the most common small intestinal malignancy. The incidence is rising, with recent reports of 0.67 per 100 000 in the USA and 1.12 per 100 000 in Sweden. SI-NETs often present a challenge in terms of diagnosis and treatment, as patients often have widespread disease and are beyond cure by surgery. Somatostatin analogues provide the mainstay of medical treatment to control hormonal excess and increase the time to progression. Despite overall favourable prognosis (5-year overall survival of 65%), there is a need to find markers to identify both patients with worse outcome and new targets for therapy. Loss on chromosome 18 has been reported in 60-90% of SI-NETs, but mutated genes on this chromosome have failed detection. Recently, a putative tumour suppressor role has been suggested for TCEB3C occurring at 18q21 (encoding elongin A3), which may undergo epigenetic repression. CDKN1B has recently been revealed as the only recurrently mutated gene in SI-NETs but, with a frequency as low as 8%, its role as a driver in SI-NET development may be questioned. Integrated genomewide analysis including exome and whole-genome sequencing, gene expression, DNA methylation and copy number analysis has identified three novel molecular subtypes of SI-NET with differing clinical outcome. DNA methylation analysis has demonstrated that SI-NETs have significant epigenetic dysregulation in 70-80% of tumours. In this review, we focus on understanding of the genetic, epigenetic and molecular events that lead to development and progression of SI-NETs.

Keyword
DNA methylation, epigenetics, genetics, midgut carcinoid, molecular profiling, small intestinal neuroendocrine tumour
National Category
Cancer and Oncology Surgery
Identifiers
urn:nbn:se:uu:diva-307770 (URN)10.1111/joim.12526 (DOI)000388573300006 ()27306880 (PubMedID)
Available from: 2016-11-21 Created: 2016-11-21 Last updated: 2017-11-29Bibliographically approved
Daskalakis, K., Edfeldt, K., Norlén, O., Karakatsanis, A., Backlin, C., Tiensuu Janson, E., . . . Stålberg, P. (2016). Midkine Is a New Novel Serum Biomarker in Small Intestinal Neuroendocrine Tumors (SI-NETs). Paper presented at 13th Annual ENETS Conference for the Diagnosis and Treatment of Neuroendocrine Tumor Disease, MAR 09-11, 2016, Barcelona, SPAIN. Neuroendocrinology, 103, 45-45.
Open this publication in new window or tab >>Midkine Is a New Novel Serum Biomarker in Small Intestinal Neuroendocrine Tumors (SI-NETs)
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2016 (English)In: Neuroendocrinology, ISSN 0028-3835, E-ISSN 1423-0194, Vol. 103, 45-45 p.Article in journal (Refereed) Published
Keyword
Biomarker, SI-NET
National Category
Cancer and Oncology Endocrinology and Diabetes Neurology
Identifiers
urn:nbn:se:uu:diva-313702 (URN)000386481600119 ()
Conference
13th Annual ENETS Conference for the Diagnosis and Treatment of Neuroendocrine Tumor Disease, MAR 09-11, 2016, Barcelona, SPAIN
Available from: 2017-01-23 Created: 2017-01-23 Last updated: 2017-11-29Bibliographically approved
Westin, G. (2016). Molecular genetics and epigenetics of nonfamilial (sporadic) parathyroid tumours. Paper presented at Symposium on New Genetics with Impact on Treatment of Endocrine Tumour Disease, Uppsala, SWEDEN, JUN 04-05, 2015. Journal of Internal Medicine, 280(6), 551-558.
Open this publication in new window or tab >>Molecular genetics and epigenetics of nonfamilial (sporadic) parathyroid tumours
2016 (English)In: Journal of Internal Medicine, ISSN 0954-6820, E-ISSN 1365-2796, Vol. 280, no 6, 551-558 p.Article in journal, Meeting abstract (Refereed) Published
Abstract [en]

Primary hyperparathyroidism (pHPT) is a common endocrine disease characterized by excessive secretion of parathyroid hormone and an increased level of serum calcium. Overall, 80-85% of pHPT cases are due to a benign, single parathyroid adenoma (PA), and 15% to multiglandular disease (multiple adenomas/hyperplasia). Parathyroid carcinoma (PC) is rare, accounting for <0.5-1% of pHPT cases. Secondary hyperparathyroidism (sHPT) is a complication of renal failure, with the development of parathyroid tumours and hypercalcaemia. Recurrent mutations in the MEN1 gene have been confirmed by the whole-exome sequencing in 35% of PAs, suggesting that non-protein-coding genes, regulatory elements or epigenetic derangements may also have roles in the majority of PAs. DNA translocations with cyclin D1 overexpression occur in PAs (8%). In PCs, mutations in CDC73/HRPT2 are common. Activation of the WNT/β-catenin signalling pathway (accumulation of nonphosphorylated β-catenin) by an aberrantly truncated LRP5 receptor has been seen for the majority of investigated PAs and sHPT tumours, and possibly by APC inactivation through promoter methylation in PCs. Promoter methylation of several other genes and repressive histone H3 lysine 27 trimethylation by EZH2 of the HIC1 gene may also contribute to parathyroid tumorigenesis. It is possible that a common pathway exists for parathyroid tumour development. CCND1 (cyclin D1) and EZH2 overexpression, accumulation of nonphosphorylated β-catenin and repression of HIC1 have all been observed to occur in PAs, PCs and sHPT tumours. In addition, hypermethylation has been observed for the same genes in PAs and PCs (e.g. SFRP1, CDKN2A and WT1). Whether β-catenin represents a 'hub' in parathyroid tumour development will be discussed.

Keyword
adenoma, beta-catenin, cancer, H3K27me3, hypermethylation and hyperparathyroidism
National Category
Cancer and Oncology Surgery
Identifiers
urn:nbn:se:uu:diva-307772 (URN)10.1111/joim.12458 (DOI)000388573300003 ()27071708 (PubMedID)
Conference
Symposium on New Genetics with Impact on Treatment of Endocrine Tumour Disease, Uppsala, SWEDEN, JUN 04-05, 2015
Available from: 2016-11-21 Created: 2016-11-21 Last updated: 2017-11-29Bibliographically approved
Delgado, A. V., Crona, J., Maharjan, R., Hellman, P., Westin, G. & Björklund, P. (2015). Exome Sequencing and CNV Analysis on Chromosome 18 in Small Intestinal Neuroendocrine Tumors: Ruling Out a Suspect?. Hormone and Metabolic Research, 47(6), 452-455.
Open this publication in new window or tab >>Exome Sequencing and CNV Analysis on Chromosome 18 in Small Intestinal Neuroendocrine Tumors: Ruling Out a Suspect?
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2015 (English)In: Hormone and Metabolic Research, ISSN 0018-5043, E-ISSN 1439-4286, Vol. 47, no 6, 452-455 p.Article in journal (Refereed) Published
Abstract [en]

The genetic background in small intestinal neuroendocrine tumors is poorly understood, but several studies have revealed numerical imbalances. Loss of one copy of chromosome 18 is the most frequent genetic aberration in this tumor type, which indirectly suggests that a driver mutation may be present in the remaining allele. The aim of this study was to evaluate the mutation status on chromosome 18 in small intestinal neuroendocrine tumors. DNAs from 7 small intestinal neuroendocrine tumors were subjected to whole exome capture, followed by next generation sequencing and high resolution SNP array followed by copy number variation analysis. Exome capture sequencing generated an average coverage of 50.6-138.2. Only 19 genes were covered less than 8X. No tumor-specific somatic mutation was identified. Genomic profiling revealed loss of chromosome 18 in 5 out of 7 small intestinal neuroendocrine tumors and a number of other aberrancies. Loss of chromosome 18 is the most frequent genetic aberration in small intestinal neuroendocrine tumors, but no evidence for eventual mutations in the remaining allele. This suggests involvement of other mechanisms than point mutations in small intestinal neuroendocrine tumors tumorigenesis.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-244677 (URN)10.1055/s-0034-1389992 (DOI)000354881200009 ()25354328 (PubMedID)
Available from: 2015-02-19 Created: 2015-02-19 Last updated: 2017-12-04
Cromer, M. K., Choi, M., Nelson-Williams, C., Fonseca, A. L., Kunstman, J. W., Korah, R. M., . . . Lifton, R. P. (2015). Neomorphic effects of recurrent somatic mutations in Yin Yang 1 in insulin-producing adenomas. Proceedings of the National Academy of Sciences of the United States of America, 112(13), 4062-4067.
Open this publication in new window or tab >>Neomorphic effects of recurrent somatic mutations in Yin Yang 1 in insulin-producing adenomas
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2015 (English)In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 112, no 13, 4062-4067 p.Article in journal (Refereed) Published
Abstract [en]

Insulinomas are pancreatic islet tumors that inappropriately secrete insulin, producing hypoglycemia. Exome and targeted sequencing revealed that 14 of 43 insulinomas harbored the identical somatic mutation in the DNA-binding zinc finger of the transcription factor Yin Yang 1 (YY1). Chromatin immunoprecipitation sequencing (ChIP-Seq) showed that this T372R substitution changes the DNA motif bound by YY1. Global analysis of gene expression demonstrated distinct clustering of tumors with and without YY1(T372R) mutations. Genes showing large increases in expression in YY1(T372R) tumors included ADCY1 (an adenylyl cyclase) and CACNA2D2 (a Ca2+ channel); both are expressed at very low levels in normal beta-cells and show mutation-specific YY1 binding sites. Both gene products are involved in key pathways regulating insulin secretion. Expression of these genes in rat INS-1 cells demonstrated markedly increased insulin secretion. These findings indicate that YY1(T372R) mutations are neomorphic, resulting in constitutive activation of cAMP and Ca2+ signaling pathways involved in insulin secretion.

Keyword
exome sequencing, insulinoma, YY1, cAMP, adenylyl cyclase
National Category
Endocrinology and Diabetes
Identifiers
urn:nbn:se:uu:diva-251996 (URN)10.1073/pnas.1503696112 (DOI)000351914500068 ()25787250 (PubMedID)
Available from: 2015-05-08 Created: 2015-04-28 Last updated: 2017-12-04
Åkerström, T., Willenberg, H. S., Cupisti, K., Ip, J., Backman, S., Moser, A., . . . Hellman, P. (2015). Novel somatic mutations and distinct molecular signature in aldosterone-producing adenomas.. Endocrine-Related Cancer, 22(5), 735-744.
Open this publication in new window or tab >>Novel somatic mutations and distinct molecular signature in aldosterone-producing adenomas.
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2015 (English)In: Endocrine-Related Cancer, ISSN 1351-0088, E-ISSN 1479-6821, Vol. 22, no 5, 735-744 p.Article in journal (Refereed) Published
Abstract [en]

Aldosterone-producing adenomas (APAs) are found in 1.5-3.0% of hypertensive patients in primary care and can be cured by surgery. Elucidation of genetic events may improve our understanding of these tumors and ultimately improve patient care. Approximately 40% of APAs harbor a missense mutation in the KCNJ5 gene. More recently, somatic mutations in CACNA1D, ATP1A1 and ATP2B3, also important for membrane potential/intracellular Ca(2) (+) regulation, were observed in APAs. In this study, we analyzed 165 APAs for mutations in selected regions of these genes. We then correlated mutational findings with clinical and molecular phenotype using transcriptome analysis, immunohistochemistry and semiquantitative PCR. Somatic mutations in CACNA1D in 3.0% (one novel mutation), ATP1A1 in 6.1% (six novel mutations) and ATP2B3 in 3.0% (two novel mutations) were detected. All observed mutations were located in previously described hotspot regions. Patients with tumors harboring mutations in CACNA1D, ATP1A1 and ATP2B3 were operated at an older age, were more often male and had tumors that were smaller than those in patients with KCNJ5 mutated tumors. Microarray transcriptome analysis segregated KCNJ5 mutated tumors from ATP1A1/ATP2B3 mutated tumors and those without mutation. We observed significant transcription upregulation of CYP11B2, as well as the previously described glomerulosa-specific gene NPNT, in ATP1A1/ATP2B3 mutated tumors compared to KCNJ5 mutated tumors. In summary, we describe novel somatic mutations in proteins regulating the membrane potential/intracellular Ca(2) (+) levels, and also a distinct mRNA and clinical signature, dependent on genetic alteration.

Keyword
ATP1A1; CACNA1D; KCNJ5; primary aldosteronism; aldosterone-producing adenoma
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-266639 (URN)10.1530/ERC-15-0321 (DOI)000364022400010 ()26285814 (PubMedID)
Funder
Swedish Cancer SocietySwedish Research Council
Available from: 2015-11-10 Created: 2015-11-10 Last updated: 2017-12-01
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