Abstract P2-25-01: TRPS1 Regulates angiogenesis via JAG1 and TYMP in Triple-Negative Breast Cancer

Abstract

Abstract Background: Triple-negative breast cancer (TNBC), the most difficult subtype to treat, is defined as estrogen receptor (ER), HER2, and progesterone receptor (PR) negative and comprises roughly 15-20% of all breast cancers. Patients with a TNBC diagnosis have worse outcomes and poorer survival rates when compared to women with other breast cancers, despite adjuvant chemotherapy. One of the factors contributing to its metastatic ability is the angiogenic process it undergoes. Clinical trials with anti-angiogenic therapeutics showed to be inadequate, with moderate response rates and insignificant survival gains for patients. TRPS1, a GATA and GATA-like transcription factor, has been proven to play a role in the epithelial to mesenchymal transition (EMT) which ultimately promotes tumor growth and metastasis. It has previously been shown that TRPS1 regulates angiogenesis via the expression of VEGF proteins in breast cancer. Our purpose in this study was to further investigate the functional role of TRPS1-regulated genes involved in TNBC angiogenic pathways. Methods: Sleeping-Beauty (SB) transposons help detect cancer progression genes which participate in tumorigenesis and metastasis, not easily identified with current sequencing technology. Using the SB transposon methodology, several candidate trunk drivers were elucidated using Pten mutant mice. Validation studies further showed TRPS1 as one of the eight TNBC tumor suppressor genes discovered by this technology. Then, ChIP-seq array studies were performed which showed that TRPS1 regulates expression of genes involved in the angiogenesis pathway. To corroborate the functional role of TRPS1 in angiogenesis, tube formation and sprouting assays were performed using overexpression and inactivation of TRPS1 in MDA-MD-231 and HCC70 cells, respectively. Furthermore, we performed a ChIP qPCR human angiogenesis array to determine TRPS1-regulated angiogenic genes. A luciferase reporter assay was performed to determine whether TRPS1 is a direct transcriptional regulator of these genes. Results: Inactivation of TRPS1 expression allowed tube formation and cell branching in the sprouting assays. Tumor xenografts overexpressing TRPS1 were analyzed by immunohistochemistry, showed a significant reduction of angiogenic vasculature when stained with CD31; and a substantial increase in blood vessels when TRPS1 was shRNA inactivated. Furthermore, human ChIP qPCR angiogenesis array identified 10 top candidate genes potentially regulated by TRPS1 transcription factor. Of these candidates, JAG1 and TYMP showed to have a higher fold enrichment compared to other angiogenic related genes. Finally, we validated its direct functional binding by using luciferase reporter assay, such demonstrated that TRPS1 is a direct transcriptional regulator of JAG1 and TYMP. Discussion: It has been previously established that the NOTCH ligand, JAG1 and the thymidine phosphorylase enzyme, TYMP are powerful genes participating in angiogenesis. Furthermore, elevated expression of JAG1 mRNA and protein has been associated with poor patient outcomes in breast cancer. Similarly, TYMP activity has been shown to be higher in triple-negative breast tumors, leading to poor outcomes as well. Although several anti-angiogenic drugs have been approved, tumor-acquired resistance to therapy limit their effectiveness. An increased understanding of tumor vessel mechanisms involved in tumorigenesis and metastasis is necessary to overcome the obstacles that prevent successful control of the angiogenic response in tumors. This would require as many paths as can be unraveled regarding the angiogenic mechanisms driving TNBC. Understanding the relationship between the function of TRPS1 in regulating the expression of important genes such as JAG1 and TYMP lays a better foundation for new drugs and drug targets that control and inhibit tumor angiogenesis. Citation Format: Liliana Guzman, Camila Ayerbe, Roberto Rangel, Jenny Chang, Roberto Rosato. TRPS1 Regulates angiogenesis via JAG1 and TYMP in Triple-Negative Breast Cancer [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P2-25-01.