Targeted imaging and inhibition of triple-negative breast cancer metastases by a PDGFRβ aptamer

Abstract

While the overall mortality for breast cancer has recently declined, management of triple-negative breast cancer (TNBC) is still challenging because of its aggressive clinical behavior and the lack of targeted therapies. Genomic profiling studies highlighted the high level of heterogeneity of this cancer, which comprises different subtypes with unique phenotypes and response to treatment. Platelet-derived growth factor receptor β (PDGFRβ) is an established mesenchymal/stem cell-specific marker in human glioblastoma and, as recently suggested, it may uniquely mark breast cancer cells with stem-like characteristics and/or that have undergone epithelial-mesenchymal transition.Methods: Immunohistochemical analysis for PDGFRβ expression was performed on a human TNBC tissue microarray. Functional assays were conducted on mesenchymal-like TNBC cells to investigate the effect of a previously validated PDGFRβ aptamer on invasive cell growth in three-dimensional culture conditions, migration, invasion and tube formation. The aptamer was labeled with a near-infrared (NIR) dye and its binding specificity to PDGFRβ was assessed both in vitro (confocal microscopy and flow cytometry analyses) and in vivo (fluorescence molecular tomography in mice bearing TNBC xenografts). A mouse model of TNBC lung metastases formation was established and NIR-labeled PDGFRβ aptamer was used to detect lung metastases in mice untreated or intravenously injected with unlabeled aptamer.Results: Here, we present novel data showing that tumor cell expression of PDGFRβ identifies a subgroup of mesenchymal tumors with invasive and stem-like phenotype, and propose a previously unappreciated role for PDGFRβ in driving TNBC cell invasiveness and metastases formation. We show that the PDGFRβ aptamer blocked invasive growth and migration/invasion of mesenchymal TNBC cell lines and prevented TNBC lung metastases formation. Further, upon NIR-labeling, the aptamer specifically bound to TNBC xenografts and detected lung metastases.Conclusions: We propose PDGFRβ as a reliable biomarker of a subgroup of mesenchymal TNBCs with invasive and stem-like phenotype as well as the use of the PDGFRβ aptamer as a high efficacious tool for imaging and suppression of TNBC lung metastases. This study will allow for the significant expansion of the current repertoire of strategies for managing patients with more aggressive TNBC.

Aptamer targeted therapy potentiates immune checkpoint blockade in triple-negative breast cancer

“…As both PDGFRβ and PD-L1 are crucial targets for TNBC treatments [31,43,44], we tested whether their co-inhibition would enhance TNBC cells killing. To this aim, we used an aptamer and a mAb that we previously validated as high a nity binders and inhibitors of PDGFRβ [31] and PD-L1 [40,45], respectively, in different tumor types including TNBC.…”

“…As both PDGFRβ and PD-L1 are crucial targets for TNBC treatments [31,43,44], we tested whether their co-inhibition would enhance TNBC cells killing. To this aim, we used an aptamer and a mAb that we previously validated as high a nity binders and inhibitors of PDGFRβ [31] and PD-L1 [40,45], respectively, in different tumor types including TNBC. Speci cally, Gint4.T is a 2'F-Py RNA aptamer that binds to the extracellular domain of PDGFRβ and inhibits receptor activation and downstream phosphatidyl-3-kinase (PI3K)/Akt and ERK1/2 pathways [22,31,35], thus affecting growth, migration and invasion of mesenchymal TNBC cell lines [31].…”

“…RNA extraction and RT-qPCR were performed as previously described [31] on tumors from four animals per group. Primers used were: IL-2, Fwd 5′-TTGTCGTCCTTGTCAACAGC-3′, Rev 5′-CTGGGGAGTTTCAGGTTCCT-3′; IFN-γ, Fwd, 5'-AGCGGCTGACTGAACTCAGATTGTAG-3', Rev 5'-GTCACAGTTTTCAGCTGTATAGGG-3'.…”

“…Moreover, PDGFRβ expression has been shown as a unique feature of tumor cells characterized by a mesenchymal/stem and poorly differentiated phenotype, and it correlates with aggressiveness and resistance to therapy in multiple tumor types [23][24][25][26][27][28][29][30]. Recent ndings prove that PDGFRβ is expressed on the surface of tumor cells belonging to a subgroup of mesenchymal TNBC with invasive and stem-like phenotype and contributes to drive the metastatic potential [31] and vasculogenic mimicry [31,32] of these tumors.…”

“…In searching e cacious strategies to target PDGFRβ-positive TNBC in alternative to PDGFRβ tyrosine kinase inhibitors, which showed limited clinical activity in TNBC as single agents and severe side effects [33,34], we recently tested the Gint4.T nuclease-resistant RNA aptamer, which we previously validated as a high a nity ligand/inhibitor of PDGFRβ in glioblastoma (GBM) [35,36] and human bone marrowderived mesenchymal stem cells (BM-MSCs) [22]. We found that Gint4.T is a potent theranostic agent in TNBC [31], as it e ciently detected lung metastases derived from TNBC cells and suppressed their formation when intravenously administrated in a mouse model [31].…”