Abstract C155: In-vitro multicellular 3D-spheroid model demonstrates the synergistic effect of 2-Domain soluble FMS-Like Tyrosine Kinase-1 (2d-sFlt-1) for breast cancer targeted therapies

Abstract

Using a conventional 2D-culture model, the anti-cancer drug failure rate was approximately 95%. Thus, incorporating in-vitro 3D models that better mimic in-vivo conditions became a central focus of drug discovery and regenerative medicine. Drug screening on triple-negative breast cancer (TNBC) with paclitaxel, doxorubicin alone or in combination with 2d-sFlt-1 revealed 28, 17, and 8-fold increase in the cytotoxicity effect using 2D-culture compared to 3D-spheroids. Tumor spheroids treated with 2d-sFlt-1 in combination with paclitaxel revealed a 13.5-fold higher response than paclitaxel alone. The targeted therapy was validated by a significant decrease in the formation of blood vessels either in tube formation or 3D-angiogenesis assays in endothelial-tumor cells co-cultured spheroids. The cytotoxicity effect of paclitaxel on 3D-tumor spheroids was enhanced with 2d-sFlt-1 (58% of dead cells) compared to (33% of dead cells) in paclitaxel alone. Furthermore, the antiproliferative effect of paclitaxel was significantly enhanced by combinatorial treatment with 2d-sFlt-1 (p<0.0001) compared to bevacizumab (p<0.05). Immunohistochemistry of 3D-tumor spheroids revealed inhibition in the expression of Ki-67 in combinatorial treatment. The increase in the migration rate of TNBC was significantly inhibited by combinatorial treatment as evidenced in 2D and 3D-invasion assays.  Further, immunofluorescence of 3D-spheroids showed downregulation of EMT markers (E. Cadherin, vimentin, cortactin, and paxillin) in response to combinatorial treatment compared to paclitaxel alone. Moreover, inhibition of FAK expression upon 2d-sFlt-1 treatment was exhibited either in endothelial cells, MDA-MB231, or co-cultured spheroids. Expression of vascular endothelial growth factor (VEGF) was also decreased in combinatorial treatment as evidenced by ELISA. qPCR of 3D-spheroids revealed the downregulation of EMT genes (N-cadherin, SNAIL, and TWIST) and upregulation of E-cadherin in combinatorial treatment. These findings demonstrate the antiangiogenic and anti-metastatic properties of 2d-sFlt-1 on TNBC. We conclude that multicellular 3D spheroids for bioengineering tumor models are promising in vitro animal-free models for cancer therapies and regenerative medicine.

Citation Format: Adel Zaid I. Mutahar, Bharathi Salimath. In-vitro multicellular 3D-spheroid model demonstrates the synergistic effect of 2-Domain soluble FMS-Like Tyrosine Kinase-1 (2d-sFlt-1) for breast cancer targeted therapies [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr C155.