5-aza-2'-deoxycitidine inhibits cell proliferation, extracellular matrix formation and Wnt/ß-catenin pathway in human uterine leiomyomas.

BACKGROUND: Uterine leiomyoma is a benign tumor with unclear pathogenesis and inaccurate treatment. This tumor exhibits altered DNA methylation related to disease progression. DNMT inhibitors as 5-aza-2'-deoxycytidine (5-aza-CdR), have been suggested to treat tumors in which DNA methylation is altered. We aimed to evaluate whether DNA methylation reversion with 5-aza-CdR reduces cell proliferation and extracellular matrix (ECM) formation in uterine leiomyoma cells to provide a potential treatment option. METHODS: Prospective study using uterine leiomyoma and adjacent myometrium tissues and human uterine leiomyoma primary (HULP) cells (n = 16). In tissues, gene expression was analyzed by qRT-PCR and DNMT activity by ELISA. Effects of 5-aza-CdR treatment on HULP cells were assessed by CellTiter, western blot, and qRT-PCR. RESULTS: DNMT1 gene expression was higher in uterine leiomyoma vs myometrium. Similarly, DNMT activity was greater in uterine leiomyoma and HULP cells (6.5 vs 3.8 OD/h/mg; 211.3 vs 63.7 OD/h/mg, respectively). After 5-aza-CdR treatment on HULP cells, cell viability was reduced, significantly so at 10 µM (85.3%). Treatment with 10 µM 5-aza-CdR on HULP cells significantly decreased expression of proliferation marker PCNA (FC = 0.695) and of ECM proteins (COLLAGEN I FC = 0.654; PAI-1, FC = 0.654; FIBRONECTIN FC = 0.733). 5-aza-CdR treatment also decreased expression of Wnt/ß-catenin pathway final targets, including WISP1 protein expression (10 µM, FC = 0.699), c-MYC gene expression (2 µM, FC = 0.745 and 10 µM, FC = 0.728), and MMP7 gene expression (5 µM, FC = 0.520 and 10 µM, FC = 0.577). CONCLUSIONS: 5-aza-CdR treatment inhibits cell proliferation, ECM formation, and Wnt/ß-catenin signaling pathway targets in HULP cells, suggesting that DNA methylation inhibition is a viable therapeutic target in uterine leiomyoma.

Vitamin D as an effective treatment in human uterine leiomyomas independent of mediator complex subunit 12 mutation.

OBJECTIVE: To study whether vitamin D (VitD) inhibits cell proliferation and Wnt/ß-catenin and transforming growth factor-ß (TGFß) signaling pathways in uterine leiomyomas independent of mediator complex subunit 12 (MED12) mutation status. DESIGN: Prospective study comparing leiomyoma vs. myometrial tissues and human uterine leiomyoma primary (HULP) cells treated with or without VitD and analyzed by MED12 mutation status. SETTING: Hospital and university laboratories. PATIENT(S): Women with uterine leiomyoma without any treatment (n = 37). INTERVENTION(S): Uterine leiomyoma and myometrium samples were collected from women undergoing surgery because of symptomatic leiomyoma pathology. MAIN OUTCOME MEASURE(S): Analysis of Wnt/ß-catenin and TGFß pathways and proliferation by quantitative real-time polymerase chain reaction in leiomyoma and myometrial tissue as well as in VitD-treated HULP cells analyzed by Sanger sequencing. RESULTS: Sequencing data showed that 46% of leiomyomas presented MED12 mutation, whereas no mutations were detected in adjacent myometrium. Expression of Wnt/ß-catenin and TGFß pathway genes was significantly increased in MED12-mutated leiomyomas compared to matched myometrium; no significant differences were found in wild-type (WT) leiomyomas. In HULP cells, VitD significantly decreased PCNA expression of both MED12-mutated and WT groups. VitD treatment decreased WNT4 and ß-catenin expression in both groups compared to controls, with significance for WNT4 expression in MED12-mutated samples. Similarly, VitD significantly inhibited TGFß3 expression in cells from both groups. MMP9 expression also decreased. CONCLUSION: Despite molecular differences between MED12-mutated and WT leiomyomas, VitD inhibited Wnt/ß-catenin and TGFß pathways in HULP cells, suggesting VitD as an effective treatment to reduce proliferation and extracellular matrix formation in different molecular subtypes of uterine leiomyomas.

Magnetic resonance imaging and ultrasound for assessing parametrial infiltration in cervical cancer. A systematic review and meta-analysis.

AIMS: To provide information on the current evidence regarding the diagnostic performance of ultrasound and MRI for assessing parametrial involvement in cervical cancer using the histological report as the reference standard. MATERIAL AND METHODS: Meta-analysis. An extensive search of papers comparing ultrasound and MRI in assessing parametrial infiltration in cervical cancer using pathologic analysis as a reference standard was performed in Medline (Pubmed) and Web of Science from January 1990 to September 2019. Quality was assessed using the QUADAS-2 tool. RESULTS: Our extended search identified 205 citations but after exclusions we finally included 9 articles in the meta-analysis. The risk of bias for most studies was low for four domains were assessed in QUADAS-2. Overall, for ultrasound pooled estimated sensitivity and specificity for diagnosing parametrial infiltration was 78% (95% confidence interval [CI]:48%-93%) and 96% (95% CI=89%-99%), respectively. For MRI these figures were 68% (95% CI=54%-80%) and 91% (95% CI=84%-95%), respectively. No statistical differences were found when comparing both methods (p=0.548). Heterogeneity was low/moderate for MRI and high for ultrasound. CONCLUSION: Ultrasound and MRI have similar diagnostic performance for detecting parametrial infiltration in women with cervical cancer. This might have relevance from the clinical point of view, since ultrasound is cheaper than MRI.

Long-term vitamin D treatment decreases human uterine leiomyoma size in a xenograft animal model.

OBJECTIVE: To study the effects of short- and long-term vitamin D treatment on uterine leiomyomas in vivo through cell proliferation, extracellular matrix (ECM) degradation, and apoptosis. DESIGN: Preclinical study of human leiomyoma treatment with vitamin D in an nonhuman animal model. SETTING: Hospital and university laboratories. PATIENT(S)/ANIMAL(S): Human leiomyomas were collected from patients and implanted in ovariectomized NOD-SCID mice. INTERVENTION(S): Mice were treated with vitamin D (0.5 µg/kg/d or 1 µg/kg/d) or vehicle for 21 or 60 days. MAIN OUTCOME MEASURE(S): Vitamin D effect in xenograft tissue was assessed by monitoring tumor size (18F-FDG positron-emission tomography/computerized tomography and macroscopic examination), cell proliferation (immunohistochemistry and quantitative real-time polymerase chain reaction [qRT-PCR]), ECM (Western blot), transforming growth factor (TGF) ß3 (qRT-PCR), and apoptosis (Westrn blot and TUNEL). RESULT(S): Short-term treatment with vitamin D did not appear to alter leiomyoma size, based on in vivo monitoring and macroscopic examination. However, long-term high-dose treatment induced a significant reduction in leiomyoma size. Cell proliferation was not decreased in the short term, whereas 1 µg/kg/d vitamin D in the long term significantly reduced proliferation compared with control. Although collagen-I and plasminogen activator inhibitor 1 were not modified by short-term treatment, they were both significantly reduced by long-term high-dose vitamin D. Similarly, long-term high-dose vitamin D significantly reduced TGF-ß3 expression. Finally, apoptosis significantly increased with both short- and long-term high-dose vitamin D treatment. CONCLUSION(S): Long-term vitamin D acts as an antiproliferative, antifibrotic, and proapoptotic therapy that provides a safe, nonsurgical therapeutic option for reducing uterine leiomyoma size without side-effects.