PO-038 PDGFRβ as a new biomarker for metastatic triple-negative breast cancer: development of a theranostic anti-PDGFRβ aptamer for imaging and suppression of metastases

Introduction Triple-negative breast cancers (TNBCs) are a heterogeneous group of aggressive tumours lacking oestrogen and progesterone receptors and HER2 receptor, thus excluding the possibility of using targeted therapy against these proteins. Mesenchymal-like (ML) subtype, characterised by a stem-like, undifferentiated phenotype, is more invasive and metastatic than other TNBC subtypes and has a strong tendency to form vasculogenic mimicry (VM). Recently, platelet derived growth factor receptor β (PDGFRβ) has been shown to play a role in VM of TNBC. Regrettably, therapies targeting PDGFRβ with tyrosine kinase inhibitors are not effective in treating TNBCs, thus developing new strategies to target PDGFRβ in TNBC patients is crucial to improve their chances of survival. Here, we describe the characterisation of the Gint4.T anti-PDGFRβ nuclease-resistant RNA aptamer as high efficacious theranostic tool for imaging and suppression of ML TNBC metastases. Material and methods Immunohistochemical analyses on a human TNBC tissue microarray was performed to correlate PDGFRβ expression with clinical and molecular features of different subtypes. Functional assays were conducted on PDGFRβ-positive ML BT-549 and MDA-MB-231 cells to investigate the effect of Gint4.T in interfering with cell growth in 3D conditions, migration, invasion and VM formation. Gint4.T was conjugated with near-infrared (NIR) fluorescent VivoTag-S680 and its binding specificity to receptor was confirmed both in vitro (confocal microscopy and flow cytometry analyses of TNBC cells) and in vivo (fluorescence molecular tomography in mice bearing TNBC xenografts). MDA-MB-231 cells were i.v. injected in nude mice and Gint4.T-NIR was used to detect lung metastases in mice untreated or i.v. injected with Gint4.T or a scrambled aptamer. Results and discussions The expression of PDGFRβ was observed in human TNBC samples characterised by higher metastatic behaviour. Treatment of TNBC cell lines with Gint4.T aptamer blocked their invasive growth and vasculogenic properties in 3D culture conditions, and strongly reduced cell migration/invasion in vitro and metastases formation in vivo. The Gint4.T-NIR was able to specifically bind to TNBC xenografts and detect lung metastases in vivo. Therefore, the aptamer revealed a high efficacious theranostic tool for imaging and suppression of TNBC metastases. Conclusion These studies indicate PDGFRβ as a new biomarker for ML and metastatic TNBC subtype and propose a novel targeting agent for the diagnosis and treatment of metastatic TNBCs

Inhibition of Bone Marrow-Mesenchymal Stem Cell-Induced Carbonic Anhydrase IX Potentiates Chemotherapy Efficacy in Triple-Negative Breast Cancer Cells

Conventional chemotherapy represents the main systemic treatment used for triple-negative breast cancer (TNBC) patients, although many of them develop drug resistance. The hypoxic TME is the crucial driver in the onset of insensitivity to chemotherapy. In this research, we elucidated the role played by bone marrow-derived mesenchymal stem cells (BM-MSCs) in reducing cisplatin effects in TNBC. BT-549 and MDA-MB-231 cells, grown under hypoxic conditions in the presence of conditioned medium obtained from BM-MSCs (CM-MSCs), showed a strong cisplatin insensitivity and increased expression levels of carbonic anhydrase IX (CA IX). Therefore, we inhibited CM-MSC-induced CA IX by SLC-0111 to potentiate chemotherapy efficacy in TNBC cells. Our results showed that CM-MSCs under hypoxic conditions caused an increase in the ability of TNBC cells to form vascular structures, migrate and invade Matrigel. Cell treatment with cisplatin plus SLC-0111 was able to block these mechanisms, as well as the signaling pathways underlying them, such as p-AKT, p-ERK, CD44, MMP-2, vimentin, β-catenin, and N-cadherin, more effectively than treatment with single agents. In addition, a significant enhancement of apoptosis assessed by annexin V, caspase-3 expression and activity was also shown. Taken together, our results demonstrated the possibility, through CA IX inhibition, of returning TNBC cells to a more chemosensitive state

Therapeutic Potential of a Novel α_vβ₃ Antagonist to Hamper the Aggressiveness of Mesenchymal Triple Negative Breast Cancer Sub-Type

The mesenchymal sub-type of triple negative breast cancer (MES-TNBC) has a highly aggressive behavior and worse prognosis, due to its invasive and stem-like features, that correlate with metastatic dissemination and resistance to therapies. Furthermore, MES-TNBC is characterized by the expression of molecular markers related to the epithelial-to-mesenchymal transition (EMT) program and cancer stem cells (CSCs). The altered expression of αvβ3 integrin has been well established as a driver of cancer progression, stemness, and metastasis. Here, we showed that the high levels of αvβ3 are associated with MES-TNBC and therefore exploited the possibility to target this integrin to reduce the aggressiveness of this carcinoma. To this aim, MES-TNBC cells were treated with a novel peptide, named ψRGDechi, that we recently developed and characterized for its ability to selectively bind and inhibit αvβ3 integrin. Notably, ψRGDechi was able to hamper adhesion, migration, and invasion of MES-TNBC cells, as well as the capability of these cells to form vascular-like structures and mammospheres. In addition, this peptide reversed EMT program inhibits mesenchymal markers. These findings show that targeting αvβ3 integrin by ψRGDechi, it is possible to inhibit some of the malignant properties of MES-TNBC phenotype