Non-endometrioid and high-grade endometrioid endometrial cancers show DNA fragmentation factor 40 (DFF40) and B-cell lymphoma 2 protein (BCL2) underexpression, which predicts disease-free and overall survival, but not DNA fragmentation factor 45 (DFF45) underexpression

Abstract

BackgroundThe expression of DNA fragmentation factor 45 (DFF45) and B-cell lymphoma 2 (BCL2) in glands of the normal human endometrium is related to phases of the menstrual cycle and decreases after menopause, whereas the expression of DNA fragmentation factor 40 (DFF40) is stable. Moreover, DF45, BCL2 and DFF40 underexpression has been reported in numerous malignancies, including uterine leiomyosarcomas. In this study, we aimed to investigate DFF45, BCL2 and DFF40 expression in endometrioid and non-endometrioid types of endometrial cancers (ECs). We also evaluated the correlations between DFF45, BCL2 and DFF40 expression levels and clinicopathological parameters and determined the value of these three proteins as prognostic markers of disease-free survival (DFS) and overall survival (OS).MethodsImmunohistochemistry was performed to evaluate DFF45, BCL2 and DFF40 expression in 342 cases of ECs. Student’s t-test, the Mann-Whitney U-test, and the chi-squared test were used for the statistical analyses as appropriate. The Cox-Mantel test, Cox’s proportional hazard model, and relative risk analyses were used to evaluate associations between DFF40, DFF45, and BCL2 expression and clinicopathological characteristics.ResultsDFF40 and BCL2, but not DFF45, were significantly underexpressed in non-endometrioid and high-grade endometrioid ECs compared with low- and moderate-grade endometrioid ECs. Women with DFF40- and BCL2-negative tumors had higher risks of disease recurrence, lymph node involvement, lympho-vascular space infiltration, and deep myometrial invasion compared with women with DFF40- and BCL2-positive tumors. Additionally, women with DFF40- and BCL2-negative tumors had significantly lower OS and DFS than women with DFF40- and BCL2-positive tumors. A multivariable analysis of the model, including the clinicopathological characteristics and immunohistochemical results, showed that negative BCL2 expression, lymph node involvement, and high-stage and high-grade disease were independent predictors of OS, whereas negative BCL2 expression, lymph node involvement, and high-stage disease were independent predictors of DFS.ConclusionsCompared with low- and moderate-grade endometrioid ECs, non-endometrioid and high-grade endometrioid ECs showed significant DFF40 and BCL2 underexpression. The absence of DFF40 and BCL2 expression negatively affects DFS and OS. Further prospective studies are warranted to assess the potential utility of DFF40 and BCL2 as targets in the diagnosis or treatment of ECs.

Development of prediction models for lymph node metastasis in endometrioid endometrial carcinoma

“…Similar alterations in expression patterns of the individual proteins have previously been demonstrated and extensively linked to LNM by us and others. 18 , 55 – 57 These findings support that LIMMA correctly identifies proteins associated with nodal metastasis and indicate that multiple proteins are involved; our model only selects the most contributing proteins. However, functional studies are needed to build up a picture of protein networks that drives metastasis.…”

Laparoscopic versus open hysterectomy in type I endometrial cancer – a tertiary referral center experience

“…Type II endometrial carcinomas are usually characterized by p53 and PIK3CA overexpression, while type I tumors are characterized by PTEN mutations (15) . Also, type II endometrial carcinomas compared to type I showed significant DFF40 and BCL2 underexpression as reported by Banas et al (16) . Minimally invasive surgery has been widely accepted to be used as the preferred approach in type I endometrial cancers and in selected cases of type II endometrial cancer (3,7,17,18) .…”

Energetic metabolic reprogramming in Jurkat DFF40-deficient cancer cells

“…Finally, caspase-3/7 activity was assessed by flow cytometer with the Caspase-3/7 Green detection reagent (Thermo Fisher Scientific, Cat#R37111) according to the manufacturer's instructions with some modifications. Briefly, 1x10 6 Jurkat cells were treated or not with STS [1 µM] for up to 2, 4 and 6 h in RPMI medium supplemented with 0.1% FBS. Cells were then washed twice with PBS 1X and suspended in PBS 1X containing the caspase-3/7 probe (3 drops in 4 ml).…”

“…Extracellular acidification rates (ECAR) and oxygen consumption rates (OCR) were assessed using the Seahorse XF Cell Mito Stress Test Kit (Agilent Cat#103015), according to the manufacturer's instructions with some modifications. Briefly, Jurkat cells were seeded in Seahose XFe96 cell culture microplates at a density of 0.2x10 6 Briefly, Jurkat DFF40 WT and DFF40 KO cells were washed once with PBS 1X and suspended in complete RPMI medium and fixed with 4% (v/v) paraformaldehyde for 15 min at room temperature. Samples were then vortexed 5 min at 400xg and pellets were suspended in PBS 1X.…”

“…Multiple studies have reported the importance of DFF40 in cancer cell resistance, by linking DFF40 deficient tumors with poor prognosis. Under expression of DFF40 in uterine leiomyosarcoma's [5], non-endometrioid and high grade endometrioid cancers [6], glioblastoma's and astrocytoma's [7] was shown to be associated with poor prognosis in patients. Recently, our laboratory has shown that knocking out the DFF40 gene in Jurkat cells by CRISPR-cas9 conferred cell resistance to tributyltin (TBT)-induced apoptosis through a delayed activation of the extrinsic apoptotic pathway [8], raising a new hypothesis for its potential novel role in genomic stability [9].…”