Invasive lobular carcinoma of the breast (ILC) is strongly estrogen-driven and represents a unique context for estrogen receptor (ER) signaling. In ILC, ER controls the expression of the Wnt ligand WNT4, which is critical for endocrine response and anti-estrogen resistance. However, signaling mediated by WNT4 is cell type- and tissue-specific, and has not been explored in ILC. We utilized reverse phase protein array (RPPA) to characterize ER and WNT4-driven signaling in ILC cells and identified that WNT4 mediates downstream mTOR signaling via phosphorylation of S6 Kinase. Additionally, ER and WNT4 control levels of MCL-1, which is associated with regulation of mitochondrial function. In this context, WNT4 knockdown led to decreased ATP production and increased mitochondrial fragmentation. WNT4 regulation of both mTOR signaling and MCL-1 were also observed in anti-estrogen resistant models of ILC. We identified that high WNT4 expression is associated with similar mTOR pathway activation in ILC and serous ovarian cancer tumors, suggesting that WNT4 signaling is active in multiple tumor types. The identified downstream pathways offer insight into WNT4 signaling and represent potential targets to overcome anti-estrogen resistance for patients with ILC.
Invasive lobular carcinoma (ILC) is the most common special histologic subtype of breast cancer, and nearly all ILC tumors express estrogen receptor alpha (ER). However, clinical and laboratory data suggest ILC are strongly estrogen-driven but not equally antiestrogen-sensitive. We hypothesized ILC-specific ER coregulators mediate ER functions and antiestrogen resistance in ILC, and profiled ER-associated proteins by mass spectrometry. Three ER+ ILC cell lines (MDA MB 134VI, SUM44PE, and BCK4) were compared with ER+ invasive ductal carcinoma (IDC) line data, and we examined whether siRNA of identified proteins suppressed ER-driven proliferation in ILC cells. This identified mediator of DNA damage checkpoint 1 (MDC1), a tumor suppressor in DNA damage response (DDR), as a novel ER coregulator in ILC. We confirmed ER:MDC1 interaction was specific to ILC versus IDC cells, and found MDC1 knockdown suppressed ILC cell proliferation and tamoxifen resistance. Using RNA-sequencing, we found in ILC cells MDC1 knockdown broadly dysregulates the ER transcriptome, with ER:MDC1 target genes enriched for promoter hormone response elements. Importantly, our data are inconsistent with MDC1 tumor suppressor functions in DDR, but suggest a novel oncogenic role for MDC1 as an ER coregulator. Supporting this, in breast tumor tissue microarrays, MDC1 protein was frequently low or absent in IDC, but MDC1 loss was rare in ER+ ILC. ER:MDC1 interaction and MDC1 coregulator functions may underlie ER function in ILC and serve as targets to overcome antiestrogen resistance in ILC.
Implications:
MDC1 has novel ER coregulator activity in ILC, which may underlie ILC-specific ER functions, estrogen response, and antiestrogen resistance.
Invasive lobular carcinoma (ILC) is the most common histological subtype of breast cancer, and nearly all ILC tumors express estrogen receptor alpha (ER). However, clinical and laboratory data suggest ILC are strongly estrogen-driven but not equally sensitive to anti-estrogen therapies. We hypothesized that ILC-specific ER transcriptional co-regulators mediate ER functions in ILC and anti-estrogen resistance, and profiled ER-associated proteins by mass spectrometry. Three ER+ ILC cell lines, MDA MB 134VI, SUM44PE, and BCK4, were compared to published data from ER+ invasive ductal carcinoma (IDC) cell lines, and we examined whether siRNA knockdown of identified proteins suppressed ER-driven proliferation in ILC cells. This approach found mediator of DNA damage checkpoint 1 (MDC1), a key tumor suppressor in DNA damage response (DDR), as a putative novel ER co-regulator in ILC. We confirmed ER:MDC1 interaction was specific to ILC cell lines versus IDC cells, and found MDC1 knockdown suppressed ILC cell proliferation and suppressed tamoxifen resistance in MDA MB 134VI. Using RNA-sequencing, we found that in ILC cells, MDC1 knockdown broadly dysregulates the estrogen-driven ER transcriptome, with ER:MDC1 target genes enriched for hormone-response-elements in their promoter regions. Importantly, our data are inconsistent with MDC1 regulating ER via MDC1 DDR and tumor suppressor functions, but instead suggest a novel oncogenic role for MDC1 in mediating ER transcriptional activity as a co-regulator. Supporting this, in breast tumor tissue microarrays MDC1 protein was frequently low or absent in IDC or ER-ILC, but MDC1 loss is rare in ER+ ILC. ER:MDC1 interaction and MDC1 co-regulator functions may underlie cell type-specific ER functions in ILC, and serve as important biomarkers and therapeutic targets to overcome anti-estrogen resistance in ILC.
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