Integrated Epigenome, Exome, and Transcriptome Analyses Reveal Molecular Subtypes and Homeotic Transformation in Uterine Fibroids

George, Jitu W.; Fan, Huihui; Johnson, Benjamin K.; Carpenter, Tyler J.; Foy, Kelly K.; Chatterjee, Anindita; Patterson, Amanda L.; Koeman, Julie; Adams, Marie; Madaj, Zachary; Chesla, David; Marsh, Erica E.; Triche, Timothy J.; Shen, Hui; Teixeira, Jose

doi:10.1016/j.celrep.2019.11.077

Cited by 60 publications

(76 citation statements)

References 107 publications

(100 reference statements)

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“…Two samples, originally classified as HMGA1 overexpressing leiomyomas [ 14 ], displayed clear expression patterns of leiomyomas of the FH subtype. Exome sequencing revealed a somatic missense variant FH c.955G > A, p.(Asp319Asn) and a somatic intronic variant FH c.556-24T > G in one of the samples (MP169F, Figure 4 A).…”

Section: Resultsmentioning

confidence: 99%

“…Recent studies have also revealed that the majority of leiomyomas can be classified into three main molecular subtypes, reflecting mutations in MED12 , HMGA2 , and FH . Furthermore, we and others have shown that each subtype is characterized by a distinct global gene expression pattern [ 7 , 14 ], suggesting that leiomyomas could be classified by gene expression profiling. However, RNA-based studies on leiomyomas have largely been limited to fresh frozen tissue specimens.…”

Section: Discussionmentioning

confidence: 99%

“…In addition, we confirmed the accuracy of these 80 genes in a second independent dataset comprising fresh frozen samples that were sequenced using standard RNA sequencing. Furthermore, this led to the identification of biallelic loss of FH in two tumors that were originally classified as HMGA1 overexpressing tumors by George et al [ 14 ]. This motivated us to explore the expression pattern of HMGA1 in our FFPE dataset, revealing significant upregulation of HMGA1 in leiomyomas of the FH subtype, but not in leiomyomas of the MED12 and HMGA2 subtypes.…”

Section: Discussionmentioning

confidence: 99%

“…While the strongest evidence of each molecular subtype may be derived from whole-genome sequencing, it is not always the most feasible and cost-effective method for classifying leiomyomas. Most studies have therefore utilized cytogenetics, immunohistochemistry, qPCR, and Sanger sequencing [ 12 , 13 , 14 ]. Indeed, Sanger sequencing is the most common method for detecting hotspot mutations in exons 1 and 2 of MED12 , whereas cytogenetics, immunohistochemistry, and qPCR have all been utilized for detecting HMGA2 aberrations.…”

Section: Introductionmentioning

confidence: 99%

“…We and others have previously shown that MED12 , HMGA2 , and FH aberrations result in distinct global gene expression patterns, suggesting that leiomyomas can be classified by gene expression profiling [ 7 , 14 ]. However, the majority of molecular studies on leiomyomas have been performed using fresh frozen tissue.…”

Section: Introductionmentioning

confidence: 99%

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3′RNA Sequencing Accurately Classifies Formalin-Fixed Paraffin-Embedded Uterine Leiomyomas

Mehine

Khamaiseh

Ahvenainen

et al. 2020

Cancers

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Uterine leiomyomas are benign smooth muscle tumors occurring in 70% of women of reproductive age. The majority of leiomyomas harbor one of three well-established genetic changes: a hotspot mutation in MED12, overexpression of HMGA2, or biallelic loss of FH. The majority of studies have classified leiomyomas by complex and costly methods, such as whole-genome sequencing, or by combining multiple traditional methods, such as immunohistochemistry and Sanger sequencing. The type of specimens and the amount of resources available often determine the choice. A more universal, cost-effective, and scalable method for classifying leiomyomas is needed. The aim of this study was to evaluate whether RNA sequencing can accurately classify formalin-fixed paraffin-embedded (FFPE) leiomyomas. We performed 3′RNA sequencing with 44 leiomyoma and 5 myometrium FFPE samples, revealing that the samples clustered according to the mutation status of MED12, HMGA2, and FH. Furthermore, we confirmed each subtype in a publicly available fresh frozen dataset. These results indicate that a targeted 3′RNA sequencing panel could serve as a cost-effective and robust tool for stratifying both fresh frozen and FFPE leiomyomas. This study also highlights 3′RNA sequencing as a promising method for studying the abundance of unexploited tissue material that is routinely stored in hospital archives.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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3′RNA Sequencing Accurately Classifies Formalin-Fixed Paraffin-Embedded Uterine Leiomyomas

Mehine

Khamaiseh

Ahvenainen

et al. 2020

Cancers

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3′RNA and whole‐genome sequencing of archival uterine leiomyomas reveal a tumor subtype with chromosomal rearrangements affecting either HMGA2, HMGA1, or PLAG1

Jokinen

Mehine

Reinikka

et al. 2022

Genes Chromosomes & Cancer

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Uterine leiomyomas, or fibroids, are very common smooth muscle tumors that arise from the myometrium. They can be divided into distinct molecular subtypes. We have previously shown that 3′RNA‐sequencing is highly effective in classifying archival formalin‐fixed paraffin‐embedded (FFPE) leiomyomas according to the underlying mutation. In this study, we performed 3′RNA‐sequencing with 111 FFPE leiomyomas previously classified as negative for driver alterations in mediator complex subunit 12 (MED12), high mobility group AT‐hook 2 (HMGA2), and fumarate hydratase (FH) by Sanger sequencing and immunohistochemistry. This revealed 43 tumors that displayed expression features typically seen in HMGA2‐positive tumors, including overexpression of PLAG1. We explored 12 such leiomyomas by whole‐genome sequencing to identify their underlying genomic drivers and to evaluate the feasibility of detecting chromosomal driver alterations from FFPE material. Four tumors with significant HMGA2 overexpression at the protein‐level served as controls. We identified chromosomal rearrangements targeting either HMGA2, HMGA1, or PLAG1 in all 16 tumors, demonstrating that it is possible to detect chromosomal driver alterations in archival leiomyoma specimens as old as 18 years. Furthermore, two tumors displayed biallelic loss of DEPDC5 and one tumor harbored a COL4A5–COL4A6 deletion. These observations suggest that instead of only HMGA2‐positive leiomyomas, a distinct leiomyoma subtype is characterized by rearrangements targeting either HMGA2, HMGA1, or PLAG1. The results indicate that the frequency of HMGA2‐positive leiomyomas may be higher than estimated in previous studies where immunohistochemistry has been used. This study also demonstrates the feasibility of detecting chromosomal driver alterations from archival FFPE material.

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Integrative analysis of the DNA methylome and transcriptome in uterine leiomyoma shows altered regulation of genes involved in metabolism, proliferation, extracellular matrix, and vesicles

Carbajo-García

Corachán

Juárez‐Barber

et al. 2022

The Journal of Pathology

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Uterine leiomyomas (ULs) are the most common benign tumors in women of reproductive age. Despite the high prevalence, tumor pathology remains unclear, which hampers the development of safe and effective treatments. Epigenetic mechanisms appear to be involved in UL development, particularly via DNA methylation that regulates gene expression. We aimed to determine the relationship between DNA methylation and gene expression in UL compared with adjacent myometrium (MM) to identify molecular mechanisms involved in UL formation that are under epigenetic control. Our results showed a different DNA methylation profile between UL and MM, leading to hypermethylation of UL, and a different global transcriptome profile. Integration of DNA methylation and whole‐transcriptome RNA‐sequencing data identified 93 genes regulated by methylation, with 22 hypomethylated/upregulated and 71 hypermethylated/downregulated. Functional enrichment analysis showed dysregulated biological processes and molecular functions involved in metabolism and cell physiology, response to extracellular signals, invasion, and proliferation, as well as pathways related to uterine biology and cancer. Cellular components such as cell membranes, vesicles, extracellular matrix, and cell junctions were dysregulated in UL. In addition, we found hypomethylation/upregulation of oncogenes (PRL, ATP8B4, CEMIP, ZPMS2‐AS1, RIMS2, TFAP2C) and hypermethylation/downregulation of tumor suppressor genes (EFEMP1, FBLN2, ARHGAP10, HTATIP2), which are related to proliferation, invasion, altered metabolism, deposition of extracellular matrix, and Wnt/β‐catenin pathway dysregulation. This confirms that key processes of UL development are under DNA methylation control. Finally, inhibition of DNA methyltransferases by 5‐aza‐2'‐deoxycitidine increased the expression of hypermethylated/downregulated genes in UL cells in vitro. In conclusion, gene regulation by DNA methylation is implicated in UL pathogenesis, and reversion of this methylation could offer a therapeutic option for UL. © 2022 The Pathological Society of Great Britain and Ireland.

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Integrated Epigenome, Exome, and Transcriptome Analyses Reveal Molecular Subtypes and Homeotic Transformation in Uterine Fibroids

Cited by 60 publications

References 107 publications

3′RNA Sequencing Accurately Classifies Formalin-Fixed Paraffin-Embedded Uterine Leiomyomas

3′RNA Sequencing Accurately Classifies Formalin-Fixed Paraffin-Embedded Uterine Leiomyomas

3′RNA and whole‐genome sequencing of archival uterine leiomyomas reveal a tumor subtype with chromosomal rearrangements affecting either HMGA2, HMGA1, or PLAG1

Integrative analysis of the DNA methylome and transcriptome in uterine leiomyoma shows altered regulation of genes involved in metabolism, proliferation, extracellular matrix, and vesicles

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