Retinoic Acid Sensitivity of Triple-Negative Breast Cancer Cells Characterized by Constitutive Activation of the notch1 Pathway: The Role of Rarβ

Abstract

Triple-negative breast cancer (TNBC) is a heterogeneous disease that lacks effective therapeutic options. In this study, we profile eighteen TNBC cell lines for their sensitivity to the anti-proliferative action of all-trans retinoic acid (ATRA). The only three cell lines (HCC-1599, MB-157 and MDA-MB-157) endowed with ATRA-sensitivity are characterized by genetic aberrations of the NOTCH1-gene, causing constitutive activation of the NOTCH1 γ-secretase product, N1ICD. N1ICD renders HCC-1599, MB-157 and MDA-MB-157 cells sensitive not only to ATRA, but also to γ-secretase inhibitors (DAPT; PF-03084014). Combinations of ATRA and γ-secretase inhibitors produce additive/synergistic effects in vitro and in vivo. RNA-sequencing studies of HCC-1599 and MB-157 cells exposed to ATRA and DAPT and ATRA+DAPT demonstrate that the two compounds act on common gene sets, some of which belong to the NOTCH1 pathway. ATRA inhibits the growth of HCC-1599, MB-157 and MDA-MB-157 cells via RARα, which up-regulates several retinoid target-genes, including RARβ. RARβ is a key determinant of ATRA anti-proliferative activity, as its silencing suppresses the effects exerted by the retinoid. In conclusion, we demonstrate that ATRA exerts a significant anti-tumor action only in TNBC cells showing constitutive NOTCH1 activation. Our results support the design of clinical trials involving combinations between ATRA and γ-secretase inhibitors for the treatment of this TNBC subtype.

Challenges for Triple Negative Breast Cancer Treatment: Defeating Heterogeneity and Cancer Stemness

“…In addition, the M subtype highly expresses stemness-related genes and high levels of genes that are involved in developmental processes [ 14 ], thus showing dependency on retinoic acid receptors [ 19 ]. Recently, it has been proven that drug combinations comprising retinoic acid derivatives and γ-secretase inhibitors (Notch pathway inhibitors) show a synergistic effect on TNBC in vitro and in vivo models [ 19 , 32 ]. NOTCH1/2/3 mutations are frequently seen in the M and BL-1 subtypes.…”

“…Mutations involving the NOTCH PEST domain are known to be oncogenic and are key to TNBC sensitivity to γ-secretase inhibitors [ 33 ]. Therefore, targeting NOTCH could be an effective therapeutic approach for M and BL1 tumours showing NOTCH mutations [ 19 ], especially when combined with retinoic acid derivatives [ 32 ]. The M subtype also shows a negative immune signature, with the low expression of genes involved in antigen processing and presentation, interferon-gamma response, and T cell signalling.…”

Retinoic Acid Inhibits Tumor-Associated Mesenchymal Stromal Cell Transformation in Melanoma

“…However, the application of ATRA in solid tumors remains to be explored. RA can inhibit tumor cell proliferation in melanoma ( Edward and MacKie, 1989 ; Zhang and Rosdahl, 2005 ; Li and Han, 2020 ) and promote immune surveillance in breast cancer, colorectal cancer, and melanoma by influencing the metabolism of MDSCs, upregulating genes related to immune response, and supporting the survival of tumor-specific CD8 + T cells ( Guo et al, 2012 ; Paroni et al, 2020 ; Sun et al, 2020 ). Conversely, RA treatment was also reported to benefits tumor progression in sarcoma and chronic lymphocytic leukemia (CLL) by promoting the pro-tumoral differentiation of intertumoral monocytes in sarcoma and increasing CD38 expression in CLL cells ( Chen et al, 2018 ; Devalaraja et al, 2020 ).…”

A DOCK1 Gene-Derived Circular RNA Is Highly Expressed in Luminal Mammary Tumours and Is Involved in the Epithelial Differentiation, Growth, and Motility of Breast Cancer Cells

“…The growth rate of cells was determined with the sulforhodamine B assay. Single-cell motility was examined with a previously described method [ 24 ], while wound-healing assays were performed using the method of Jonkman [ 25 ].…”