Cancer Letters

Cancer Letters

Volume 235, Issue 1, 8 April 2006, Pages 136-140
Cancer Letters

No evidence for epigenetic inactivation of fumarate hydratase in leiomyomas and leiomyosarcomas

https://doi.org/10.1016/j.canlet.2005.04.017Get rights and content

Abstract

Germline mutations in Fumarate Hydratase (FH) cause the development of leiomyomas and leiomyosarcomas in the syndromes Multiple Cutaneous and Uterine Leiomyomata (MCUL1) and Hereditary Leiomyomatosis and Renal Cell Cancer (HLRCC). There is little evidence, however, that FH mutation plays a role in the development of sporadic leiomyomas or leiomyosarcomas. Such observations do not, however, exclude a role for FH in tumour development outside the context of MCUL1/HLRCC, as it is possible that FH expression could be silenced by epigenetic mechanisms. To explore this possibility we have developed a highly specific antibody to FH and analysed a series of forty-five fresh-frozen uterine leiomyomas and nine leiomyosarcomas for FH expression.

Introduction

Fibroids (uterine leiomyomas or myomas) are benign tumours originating in the smooth muscle of the uterus (myometrium). They are the commonest tumour in women during the reproductive years, occurring in up to 50% of women over the age of 30 years [1], increasing with age, and being more common in certain ethnic populations, especially the Afro-Caribbean [2], [3]. While 50% remain asymptomatic, fibroids can cause menorrhagia, pelvic pain/discomfort, bladder and bowel compression symptoms, and may contribute to infertility and miscarriage [4], [5]. They are the most common indication for hysterectomy in England [6], [7], while in the USA by the age of 60 years 30% of women will have had a hysterectomy, of which 60% will have been performed to treat fibroids [8].

The malignant counterparts of fibroids (leiomyosarcomas) are rare—an incidence of 0.67/1,00,000 women contrasting sharply with the benign lesion [9]. Leiomyosarcomas principally develop in the retroperitoneum, superficial soft tissues, and deep compartments of the extremities [10], [11]. The risk of malignant transformation of myomas has been reported to range from 0.3 to 1%. There are no data to show that the rate of growth of a myoma is related to the risk of malignant transformation [12]. Karyotypically leiomyosarcomas are characterised by a high-degree of chromosome instability and a wide range of anomalies [11], [13], [14].

Little is known about the molecular basis of sporadic leiomyomas or leiomyosarcomas. Recently, we have shown that germline mutations in Fumarate hydratase (FH) at chromosome 1q43 cause the rare autosomal dominant syndromes of Multiple Cutaneous and Uterine Leiomyomata (MCUL1) (MIM 150800) (http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=150800) and Hereditary Leiomyomatosis and Renal Cell Cancer (HLRCC) (MIM 605839) (http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=605839) [15]. MCUL1/HLRCC is typified by multiple skin leiomyomata and multiple early-onset uterine leiomyomas, and an increased risk of malignancy is also a feature, notably leiomyosarcoma and type II papillary renal cell carcinoma [16]. Fumarate hydratase, which is a component of the tricarboxylic acid cycle, acts as a tumour suppressor, its activity being very low or absent in tumours from individuals with MCUL1/HLRCC [15].

The identification of FH as the gene causing familial leiomyoma development in the MCUL1/HLRCC syndrome has instigated examination of whether or not FH mutation plays a role in the development of sporadic leiomyomas. To date there is little evidence that this is the case [17], [18], [19]. Such observations do not, however, exclude a role for FH in tumour development outside the context of MCUL1/HLRCC, as it is possible that expression could be silenced by epigenetic mechanisms. There are examples of such mechanisms with other tumours: germline mutations of E-cadherin and MLH1 cause familial gastric cancer and colorectal cancer, respectively [20], [21], whereas promoter hypermethylation rather than mutation leading to inactivation of these genes is commonly seen in sporadic forms of these tumours [22], [23]. To evaluate the possibility that FH may be epigenetically silenced in sporadic leiomyomas and leiomyosarcomas we have analysed its expression in a series of tumours.

Section snippets

Patient details and tissue collection

A series of 45 fresh leiomyomas and matched myometrium samples from 45 patients were obtained from the Gynaecology clinic, St George's Hospital, London, UK (n=40) and from the University of Nebraska Medical Center, Omaha, Nebraska, USA (n=5), average age at diagnosis was 42 years (range 32–53 years). Nine leiomyosarcoma samples were obtained from patients attending the Royal Marsden Hospital NHS Trust, London, UK, all were extra-uterine. Histological confirmation of the nine leiomyosarcoma

Antibody specificity

The antibody specificity was validated by western blotting. Fig. 1 demonstrates that the antiserum generated recognises a single band of approximately 50 kDa in a whole human fibroblast cell extract and a band of a much lower intensity in a whole cell extract of fibroblasts from fumarase deficient individual. Fumarase deficiency is a recessive condition (MIM 606812) (http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=606812), whereby affected individuals are homozygous/compound heterozygous for

Discussion

Three studies have previously reported the prevalence of somatic mutation of FH in sporadic leiomyomas. Kiuru et al. [18] analysed 41 uterine leiomyomas and no mutations were identified. We examined 129 leiomyomas from 21 individuals and similarly detected no mutations [17]. Lehtonen et al. [19] identified two somatic mutations in a study of 153 tumours from 46 individuals. Collectively this data indicates that mutation of FH is rare outside the context of MCUL1/HLRCC. Similarly FH mutations do

Acknowledgements

We are grateful to the patients who participated in this study. This study was supported by grants from Cancer Research UK, the Wellcome Trust and the National Institutes of Health/National Cancer Institute (P30 CA 36727).

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