Therapeutic Target In Basal-like Breast Cancer Research Articles

The m6A writer RBM15 drives the growth of triple-negative breast cancer cells through the stimulation of serine and glycine metabolism

N6-adenosine methylation (m6A) is critical for controlling cancer cell growth and tumorigenesis. However, the function and detailed mechanism of how m6A methyltransferases modulate m6A levels on specific targets remain unknown. In the current study, we identified significantly elevated levels of RBM15, an m6A writer, in basal-like breast cancer (BC) patients compared to nonbasal-like BC patients and linked this increase to worse clinical outcomes. Gene expression profiling revealed correlations between RBM15 and serine and glycine metabolic genes, including PHGDH, PSAT1, PSPH, and SHMT2. RBM15 influences m6A levels and, specifically, the m6A levels of serine and glycine metabolic genes via direct binding to target RNA. The effects of RBM15 on cell growth were largely dependent on serine and glycine metabolism. Thus, RBM15 coordinates cancer cell growth through altered serine and glycine metabolism, suggesting that RBM15 is a new therapeutic target in BC.

Abstract 3087: The m6A methyltransferase RBM15 drives the growth of triple-negative breast cancer cells through the stimulation of serine and glycine metabolism

Abstract N6-adenosine methylation (m6A) is critical for controlling cancer cell growth and tumorigenesis. However, the function and detailed mechanism of how m6A methyltransferases modulate m6A levels on specific targets remains unknown. In the current study, we identified significantly elevated levels of RBM15, an m6A writer, in basal-like breast cancer (BC) patients compared to non-basal like BC and linked it to worse clinical outcome. Gene expression profiling uncovered correlations between RBM15 and serine and glycine metabolism genes including PHGDH, PSAT1, PSPH, and SHMT2. RBM15 influences overall m6A levels and, specifically, m6A of serine and glycine metabolism genes via direct binding to target RNA. Further RBM15 effects on cell growth were largely dependent on serine and glycine metabolism. Thus, RBM15 coordinates cancer cell growth through altered serine and glycine metabolism suggesting RBM15 as a new therapeutic target in BC. Citation Format: Su Hwan Park, Yun-Yong Park, Jong-Ho Lee. The m6A methyltransferase RBM15 drives the growth of triple-negative breast cancer cells through the stimulation of serine and glycine metabolism [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3087.

A feedforward circuit between KLF5 and lncRNA KPRT4 contributes to basal-like breast cancer

Basal-like breast cancer (BLBC) is the most aggressive subtype of breast cancer with a poor prognosis. Long noncoding RNAs (lncRNAs) play critical roles in human cancers. Krüppel-like Factor 5 (KLF5) is a key oncogenic transcription factor in BLBC. However, the underlying mechanism of mutual regulation between KLF5 and lncRNA remains largely unknown. Here, we demonstrate that lncRNA KPRT4 promotes BLBC cell proliferation in vitro and in vivo. Mechanistically, KLF5 directly binds to the promoter of KPRT4 to promote KPRT4 transcription. Reciprocally, KPRT4 recruits the YB-1 transcription factor to the KLF5 promoter by interacting with YB-1 at its 5′ domain and forming an RNA-DNA-DNA triplex structure at its 3′ domain, resulting in enhanced transcription of KLF5 and ultimately establishing a feedforward circuit to promote cell proliferation. Moreover, the antisense oligonucleotide (ASO)-based therapy targeting KPRT4 substantially attenuated tumor growth in vivo. Clinically, the expression levels of YB-1, KLF5 and KPRT4 are positively correlated in clinical breast specimens. Together, our data suggest that KPRT4 is a major molecule for BLBC progression and that the feedforward circuit between KLF5 and KPRT4 may represent a potential therapeutic target in BLBC.

MiR-221/222 promote epithelial-mesenchymal transition by targeting Notch3 in breast cancer cell lines

Basal-like breast cancer (BLBC) is an aggressive subtype with a strong tendency to metastasize. Due to the lack of effective chemotherapy, BLBC has a poor prognosis compared with luminal subtype breast cancer. MicroRNA-221 and -222 (miR-221/222) are overexpressed in BLBC and associate with metastasis as well as poor prognosis; however, the mechanisms by which miR-221/222 function as oncomiRs remain unknown. Here, we report that miR-221/222 expression is inversely correlated with Notch3 expression in breast cancer cell lines. Notch3 is known to be overexpressed in luminal breast cancer cells and inhibits epithelial to mesenchymal transition (EMT). We demonstrate that miR-221/222 target Notch3 by binding to its 3′ untranslated region and suppressing protein translation. Ectopic expression of miR-221/222 significantly promotes EMT, whereas overexpression of Notch3 intracellular domain attenuates the oncogenic function of miR-221/222, suggesting that miR-221/222 exerts its oncogenic role by negatively regulating Notch3. Taken together, our results elucidated that miR-221/222 promote EMT via targeting Notch3 in breast cancer cell lines suggesting that miR-221/222 can serve as a potential therapeutic target in BLBC.

ALIX Regulates Tumor-Mediated Immunosuppression by Controlling EGFR Activity and PD-L1 Presentation

SummaryThe immunosuppressive transmembrane protein PD-L1 was shown to traffic via the multivesicular body (MVB) and to be released on exosomes. A high-content siRNA screen identified the endosomal sorting complexes required for transport (ESCRT)-associated protein ALIX as a regulator of both EGFR activity and PD-L1 surface presentation in basal-like breast cancer (BLBC) cells. ALIX depletion results in prolonged and enhanced stimulation-induced EGFR activity as well as defective PD-L1 trafficking through the MVB, reduced exosomal secretion, and its redistribution to the cell surface. Increased surface PD-L1 expression confers an EGFR-dependent immunosuppressive phenotype on ALIX-depleted cells. An inverse association between ALIX and PD-L1 expression was observed in human breast cancer tissues, while an immunocompetent mouse model of breast cancer revealed that ALIX-deficient tumors are larger and show an increased immunosuppressive environment. Our data suggest that ALIX modulates immunosuppression through regulation of PD-L1 and EGFR and may, therefore, present a diagnostic and therapeutic target for BLBC.

Evaluation of FOXC1 as a therapeutic target for basal-like breast cancer.

Basal-like breast cancer (BLBC) is a malignant carcinoma with aggressive motility and rapid growth. Accounting for 15% of breast cancers, BLBC often exhibits a poor prognosis and tends to metastasize to the brain and lungs. Because most BLBC display a triple-negative phenotype (ER-, PR-, and HER2-), conventional cytotoxic chemotherapy remains the only treatment option despite poor success and high rate of relapse. The overexpression of the forkhead-box transcription factor C1 (FOXC1) was recently identified as a biomarker of BLBC. Increased expression of FOXC1 was linked to excessive mobility and growth of BLBC cell lines, suggesting FOXC1 as a therapeutic target. In this study, siRNA-mediated knockdown of FOXC1 was confirmed to decrease the proliferation rate, migration, and invasion in a model BLBC-like cell line (4T1). 4T1 and 4T1-∆FOXC1 cells lacking FOXC1 expression (generated by CRISPR/Cas9) were used to evaluate the effects of FOXC1 expression in an orthotopic murine model of BLBC. No statistically significant difference in tumor volume was observed between 4T1 and 4T1-∆FOXC1 tumors. Furthermore, tumors metastasized to the liver and lungs to a similar degree regardless of FOXC1 expression. These data suggest that, despite positive results in vitro, FOXC1 may not be a promising therapeutic target for BLBC.

Forkhead box C1 promoter upstream transcript, a novel long non-coding RNA, regulates proliferation and migration in basal-like breast cancer.

Recent studies have shown that long non‑coding RNAs (lncRNAs) have crucial regulating roles in carcinogenesis. Forkhead box C1 (FOXC1) is an important cancer‑associated gene in basal‑like breast cancer (BLBC). In the present study, a novel lncRNA, FOXC1 promoter upstream transcript (FOXCUT) was investigated in BLBC patients using polymerase chain reaction analysis. The results showed that the expression of FOXCUT and FOXC1 were positively correlated. When the expression of FOXCUT was downregulated by small interfering RNA, the expression of FOXC1 was similarly reduced. Furthermore, in MDA‑MB‑231 and MDA‑MB‑468 breast cancer cells, knockdown of FOXCUT markedly inhibited cell proliferation and migration in vitro. In conclusion, FOXCUT lncRNA may be functionally involved in the tumor progression of BLBCs through the regulation of its paired mRNA, FOXC1, demonstrating that FOXCUT may serve as a novel biomarker and therapeutic target in BLBCs.

Abstract 3742: FOXC1 is a potential prognostic marker with functional significance in basal-like breast cancer

Abstract Although basal-like breast cancer (BLBC) is a distinct molecular subtype, it has no defining gene or protein marker that can be used as the basis for therapy. Estrogen receptor (ER) and HER2 receptor guide treatment of luminal and HER2 breast cancer subtypes, respectively, but chemotherapy is still the only systemic treatment available for BLBC. In view of its poor clinical outcomes, high proliferation rates and preferential metastasis to the brain, there is an urgent need for effective targeted therapy of BLBC. In a previous study using cDNA microarray analyses and immunohistochemistry of archival breast cancer tissue specimens, we showed consistent and exclusive overexpression of FOXC1 in BLBC versus other molecular subtypes. FOXC1 overexpression was significantly associated with poor overall survival and brain metastasis. In the present study, we show that overexpression of FOXC1 in breast cancer cells with low endogenous FOXC1 levels increased cellular proliferation, migration, and invasion, and induced epithelial-mesenchymal transition. Knockdown of FOXC1 by shRNA in breast cancer cells with high endogenous FOXC1 expression had the opposite effect. Cell signaling studies showed that FOXC1 activated the NF-κB pathway by upregulating the peptidyl-prolyl isomerase Pin1 and downregulating the ubiquitin ligase SOCS-1. Pin1 binds to p65, inhibits the p65 interaction with IκBα, and thus enhances p65 nuclear localization and protein stability. SOCS-1 directly interacts with p65 and promotes its degradation. FOXC1 knockdown reduced p65 protein levels and NF-κB activity. The importance of the NF-κB pathway for FOXC1 function was confirmed by the increased sensitivity to NF-κB inhibitors in cells that overexpressed FOXC1. Because BLBC under-expresses ER, FOXC1 was inversely associated with ER, as expected. Interestingly, FOXC1 repressed ER transcription and activity via NF-κB signaling, and FOXC1 overexpression switched MCF-7 breast cancer cell growth from estrogen-dependent to estrogen-independent. This may explain why ER is not normally detected in BLBC. These results suggest that FOXC1 may be a specific diagnostic and prognostic biomarker for BLBC and may play an important role in regulating aggressive traits associated with BLBC. It might also serve as a potential molecular therapeutic target for BLBC. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3742.