High-throughput screen in vitro identifies dasatinib as a candidate for combinatorial treatment with HER2-targeting drugs in breast cancer.

Human epidermal growth factor receptor 2-positive (HER2+) breast cancer is an aggressive subtype of this disease. Targeted treatment has improved outcome, but there is still a need for new therapeutic strategies as some patients respond poorly to treatment. Our aim was to identify compounds that substantially affect viability in HER2+ breast cancer cells in response to combinatorial treatment. We performed a high-throughput drug screen of 278 compounds in combination with trastuzumab and lapatinib using two HER2+ breast cancer cell lines (KPL4 and SUM190PT). The most promising drugs were validated in vitro and in vivo, and downstream molecular changes of the treatments were analyzed. The screen revealed multiple drugs that could be used in combination with lapatinib and/or trastuzumab. The Src-inhibitor dasatinib showed the largest combinatorial effect together with lapatinib in the KPL4 cell line compared to treatment with dasatinib alone (p < 0.01). In vivo, only lapatinib significantly reduced tumor growth (p < 0.05), whereas dasatinib alone, or in combination with lapatinib, did not show significant effects. Protein analyses of the treated xenografts showed significant alterations in protein levels compared to untreated controls, suggesting that all drugs reached the tumor and exerted a measurable effect. In silico analyses suggested activation of apoptosis and reduced activity of survival pathways by all treatments, but the opposite pattern was observed for the combinatorial treatment compared to lapatinib alone.

Serglycin Is Involved in TGF-ß Induced Epithelial-Mesenchymal Transition and Is Highly Expressed by Immune Cells in Breast Cancer Tissue.

Serglycin is a proteoglycan highly expressed by immune cells, in which its functions are linked to storage, secretion, transport, and protection of chemokines, proteases, histamine, growth factors, and other bioactive molecules. In recent years, it has been demonstrated that serglycin is also expressed by several other cell types, such as endothelial cells, muscle cells, and multiple types of cancer cells. Here, we show that serglycin expression is upregulated in transforming growth factor beta (TGF-ß) induced epithelial-mesenchymal transition (EMT). Functional studies provide evidence that serglycin plays an important role in the regulation of the transition between the epithelial and mesenchymal phenotypes, and it is a significant EMT marker gene. We further find that serglycin is more expressed by breast cancer cell lines with a mesenchymal phenotype as well as the basal-like subtype of breast cancers. By examining immune staining and single cell sequencing data of breast cancer tissue, we show that serglycin is highly expressed by infiltrating immune cells in breast tumor tissue.

miR-101-5p Acts as a Tumor Suppressor in HER2-Positive Breast Cancer Cells and Improves Targeted Therapy.

Purpose: Human epidermal growth factor receptor 2 positive (HER2+) breast cancers responding poorly to targeted therapy need improved treatment options. miR-101-5p has shown tumor-suppressive properties in multiple cancer forms, and we assessed the effect and mechanism of action of this miRNA in HER2+ breast cancer. Methods: Expression levels of miR-101-5p in two clinical datasets, TCGA and METABRIC, were compared between tumor and normal adjacent samples, and across molecular subtypes and HER2 status. The ability of miR-101-5p to sensitize for treatment with lapatinib, tucatinib and trastuzumab was explored in HER2+ breast cancer cells responding poorly to such targeted drugs. Proliferation and apoptosis assays and downstream protein analysis were performed. Results: Expression levels of miR-101-5p were significantly lower in tumor compared to normal adjacent tissue (p < 0.001), and particularly low in HER2+ tumors, both the HER2-enriched subtype (p ≤ 0.037) and clinical HER2-status (p < 0.001). In a HER2+ cell line (KPL4) responding poorly to targeted drugs, miR-101-5p overexpression inhibited proliferation (p < 0.001), and combinatorial treatment with lapatinib and trastuzumab significantly further decreased this inhibition (p = 0.004). Proteomic data and in silico analyses revealed PI3K/Akt- and HER2-pathways among the predicted regulated pathways. miR-101-5p alone (p = 0.018) and in combination with lapatinib and trastuzumab (p < 0.001) induced apoptosis, while the targeted drugs alone did not exert any significant effect neither on proliferation nor apoptosis. Conclusion: miR-101-5p acts as a tumor suppressor by inducing apoptosis in HER2+ breast cancer and sensitizes cells with initially poor response to lapatinib and trastuzumab.

MicroRNA in combination with HER2-targeting drugs reduces breast cancer cell viability in vitro.

HER2-positive (HER2 +) breast cancer patients that do not respond to targeted treatment have a poor prognosis. The effects of targeted treatment on endogenous microRNA (miRNA) expression levels are unclear. We report that responsive HER2 + breast cancer cell lines had a higher number of miRNAs with altered expression after treatment with trastuzumab and lapatinib compared to poorly responsive cell lines. To evaluate whether miRNAs can sensitize HER2 + cells to treatment, we performed a high-throughput screen of 1626 miRNA mimics and inhibitors in combination with trastuzumab and lapatinib in HER2 + breast cancer cells. We identified eight miRNA mimics sensitizing cells to targeted treatment, miR-101-5p, mir-518a-5p, miR-19b-2-5p, miR-1237-3p, miR-29a-3p, miR-29c-3p, miR-106a-5p, and miR-744-3p. A higher expression of miR-101-5p predicted better prognosis in patients with HER2 + breast cancer (OS: p = 0.039; BCSS: p = 0.012), supporting the tumor-suppressing role of this miRNA. In conclusion, we have identified miRNAs that sensitize HER2 + breast cancer cells to targeted therapy. This indicates the potential of combining targeted drugs with miRNAs to improve current treatments for HER2 + breast cancers.