Global metabolomic profiling of uterine leiomyomas.

BACKGROUND: Uterine leiomyomas can be classified into molecularly distinct subtypes according to their genetic triggers: MED12 mutations, HMGA2 upregulation, or inactivation of FH. The aim of this study was to identify metabolites and metabolic pathways that are dysregulated in different subtypes of leiomyomas. METHODS: We performed global metabolomic profiling of 25 uterine leiomyomas and 17 corresponding myometrium specimens using liquid chromatography-tandem mass spectroscopy. RESULTS: A total of 641 metabolites were detected. All leiomyomas displayed reduced homocarnosine and haeme metabolite levels. We identified a clearly distinct metabolomic profile for leiomyomas of the FH subtype, characterised by metabolic alterations in the tricarboxylic acid cycle and pentose phosphate pathways, and increased levels of multiple lipids and amino acids. Several metabolites were uniquely elevated in leiomyomas of the FH subtype, including N6-succinyladenosine and argininosuccinate, serving as potential biomarkers for FH deficiency. In contrast, leiomyomas of the MED12 subtype displayed reduced levels of vitamin A, multiple membrane lipids and amino acids, and dysregulation of vitamin C metabolism, a finding which was also compatible with gene expression data. CONCLUSIONS: The study reveals the metabolomic heterogeneity of leiomyomas and provides the requisite framework for strategies designed to target metabolic alterations promoting the growth of these prevalent tumours.

MED12 mutation frequency in unselected sporadic uterine leiomyomas.

OBJECTIVE: To determine the frequency of mediator complex subunit 12 (MED12) mutations in well-documented, prospectively collected, unselected series of sporadic uterine leiomyomas to better understand the contribution of MED12 mutations in leiomyoma genesis. DESIGN: Mutation analysis of two prospectively collected sample series. SETTING: Department of gynecology in university hospital and medical genetics research laboratory. PATIENT(S): 164 uterine leiomyomas from 28 patients (13 consecutive and 15 unselected patients) undergoing hysterectomy. INTERVENTION(S): MED12 mutation screening by direct sequencing, and clinical data collection. MAIN OUTCOME MEASURE(S): MED12 mutation status and various clinical variables. RESULT(S): MED12 mutations were found in 73 (83.0%) of 88 and 65 (85.5%) of 76 of uterine leiomyomas from the consecutive and unselected patient series, respectively. Smaller tumor size and a larger number of tumors correlated with positive MED12 mutation status. CONCLUSION(S): The frequency of MED12 mutations in our prospectively collected uterine leiomyoma sets was higher than in previous works. This is in keeping with the concept that MED12 mutation-positive tumors tend to be smaller in size than MED12 mutation-negative tumors. The results highlight the central role of MED12 mutations in uterine leiomyoma genesis.

Inhibition of FOSL1 overexpression in antiestrogen-resistant MCF-7 cells decreases cell growth and increases vacuolization and cell death.

Elevated activator protein-1 (AP-1) activity in breast cancer cells has been linked to Tamoxifen (TAM) resistance. Fos-like antigen-1 (FOSL1) is a member of the AP-1 transcription factor and is overexpressed in a variety of human cancers including breast tumors. We have previously established an estrogen-independent and antiestrogen Toremifene (TOR)-resistant subline of MCF-7 breast cancer cells. In these cells, the expression of FOSL1 is upregulated when compared to the parental cells. In the present study, partial inhibition of FOSL1 expression in these cells by small interfering RNA resulted in a marked decrease of cell growth. The inhibition of cell growth paralleled with changes in cell morphology such as increased formation of vacuoles followed by an increase in the number of dead cells. The inhibition of FOSL1 expression in these cells also restored sensitivity to TOR. Our results suggest that chemotherapy targeting overexpression of FOSL1 could be a potent strategy for treating endocrine resistant breast cancers.

Gene expression changes during the development of estrogen-independent and antiestrogen-resistant growth in breast cancer cell culture models.

We have established estrogen-independent and antiestrogen-resistant cell lines from hormone-dependent MCF-7 breast cancer cells by long-term culture in the absence of estrogen, or in the presence of antiestrogen toremifene, respectively. By using a cDNA microarray we compared gene expression profiles among estrogen-independent, antiestrogen-resistant and long-term estrogen-treated MCF-7 cells. We also determined how the expression of the differentially expressed genes has developed during the long-term culture of the cell lines. Of the screened 1176 cancer-related genes, FOSL1, TIMP1, L1CAM, GDF15, and MYBL2 were found to be differentially expressed between the cell lines. A change in FOSL1 and TIMP1 expression could be attributed to the development of antiestrogen resistance, whereas induced L1CAM expression was implicated in the development of estrogen-independent growth of the cells. Estrogen regulated genes GDF15 and L1CAM became regulated by toremifene in the later passage number of toremifene-resistant cells, which might be an indication of the developed estrogen-agonistic activity of toremifene in these cells. Our findings suggest a pattern where the hormone-responsive cancer cells, which survive E2 deprivation and/or antiestrogen treatment, first acquire necessary changes in gene expression for transition to maximal growth in the new hormonal environment. Then, after prolonged treatment with antiestrogen, the antiestrogen-resistant cells may eventually generate an E2-agonistic response to antiestrogen, probably acquiring additional growth advantage.

Changes in protein tyrosine phosphatase type IVA member 1 and zinc finger protein 36 C3H type-like 1 expression demonstrate altered estrogen and progestin effect in medroxyprogesterone acetate-resistant and estrogen-independent breast cancer cell models.

Estrogen stimulates proliferation in hormone-responsive breast cancer cells. Progestins inhibit the estrogen-mediated growth in these cells and are used in the treatment of mammary carcinomas. We applied cDNA microarray and real-time RT-PCR methods to reveal 17beta-estradiol- and medroxyprogesterone acetate (MPA)-regulated genes in MCF-7 breast cancer cells. We identified six genes, two of which were novel MPA and/or 17beta-estradiol-regulated genes: protein tyrosine phosphatase type IVA, member 1 (PTP4A1) and zinc finger protein 36, C3H type-like 1 (ZFP36L1). PTP4A1 expression was upregulated by 17beta-estradiol and this was opposed by MPA treatment of the cells. ZFP36L1 proved to be a direct target of MPA. Since MPA has also been shown to bind to glucocorticoid- and androgen receptors, we studied the regulation of the genes with progesterone, synthetic progestin R5020, dexamethasone and dihydrotestosterone. We also assessed the expression and hormonal regulation of PTP4A1 and ZFP36L1 mRNA in MCF-7-derived MPA-resistant and estrogen-independent sublines. The regulation of PTP4A1 expression upon 17beta-estradiol and combined 17beta-estradiol and MPA treatment was completely reversed in the estrogen-independent and MPA-resistant sublines, respectively. This study suggests an important role for PTP4A1 in cell growth, and shows that MPA may potentiate the effect of 17beta-estradiol in the MPA-resistant breast cancer cells.