Metastatic inhibitory effect of gallic acid combined with cisplatin on hela cells proliferation

Due to the multiple adverse effects of chemotherapy treatment, more researches with natural product discovery has been conducted to treat various types of cancer. A polyphenol derivative, gallic acid, was previously reported to combat the growth of cancer cells. Hence, this study aims to assess the metastatic inhibitory effect of gallic acid combined with cisplatin against HeLa cells. Cell migration, cell invasion and cell proliferation were measured by scratch-wound healing assay, Transwell invasion assay and colony formation assay, respectively. After 24-hour of scratch-wound healing assay, the combined treatment showed significant decrease in wound closure rate where 43.08 ± 6.21% of the wound was filled by the HeLa cells in comparison to control wound width. Combination of gallic acid and cisplatin also exerted a significant decrease in the number of invaded cells at 44.55 ± 6.04% as compared to the control group by Transwell invasion assay. Gallic acid and cisplatin were able to significantly reduce the proliferation HeLa cells by colony formation assay. However, the effect that was produced by their combination was greater than that of each agent alone. Hence further study either in vitro and in vivo is required to strengthen the current findings such as research regarding the involvement of PI3K/AKT/mTOR signaling pathway in decreasing migration and invasion capability of HeLa cells

Cancer Cell-Derived PDGFB Stimulates mTORC1 Activation in Renal Carcinoma

Clear cell renal cell carcinoma (ccRCC) is a hypervascular tumor that is characterized by bi-allelic inactivation of the VHL tumor suppressor gene and mTOR signalling pathway hyperactivation. The pro-angiogenic factor PDGFB, a transcriptional target of super enhancer-driven KLF6, can activate the mTORC1 signalling pathway in ccRCC. However, the detailed mechanisms of PDGFB-mediated mTORC1 activation in ccRCC have remained elusive. Here, we investigated whether ccRCC cells are able to secrete PDGFB into the extracellular milieu and stimulate mTORC1 signalling activity. We found that ccRCC cells secreted PDGFB extracellularly, and by utilizing KLF6- and PDGFB-engineered ccRCC cells, we showed that the level of PDGFB secretion was positively correlated with the expression of intracellular KLF6 and PDGFB. Moreover, the reintroduction of either KLF6 or PDGFB was able to sustain mTORC1 signalling activity in KLF6-targeted ccRCC cells. We further demonstrated that conditioned media of PDGFB-overexpressing ccRCC cells was able to re-activate mTORC1 activity in KLF6-targeted cells. In conclusion, cancer cell-derived PDGFB can mediate mTORC1 signalling pathway activation in ccRCC, further consolidating the link between the KLF6-PDGFB axis and the mTORC1 signalling pathway activity in ccRCC

Cancer Cell-Derived PDGFB Stimulates mTORC1 Activation in Renal Carcinoma

Clear cell renal cell carcinoma (ccRCC) is a hypervascular tumor that is characterized by bi-allelic inactivation of the VHL tumor suppressor gene and mTOR signalling pathway hyperactivation. The pro-angiogenic factor PDGFB, a transcriptional target of super enhancer-driven KLF6, can activate the mTORC1 signalling pathway in ccRCC. However, the detailed mechanisms of PDGFB-mediated mTORC1 activation in ccRCC have remained elusive. Here, we investigated whether ccRCC cells are able to secrete PDGFB into the extracellular milieu and stimulate mTORC1 signalling activity. We found that ccRCC cells secreted PDGFB extracellularly, and by utilizing KLF6- and PDGFB-engineered ccRCC cells, we showed that the level of PDGFB secretion was positively correlated with the expression of intracellular KLF6 and PDGFB. Moreover, the reintroduction of either KLF6 or PDGFB was able to sustain mTORC1 signalling activity in KLF6-targeted ccRCC cells. We further demonstrated that conditioned media of PDGFB-overexpressing ccRCC cells was able to re-activate mTORC1 activity in KLF6-targeted cells. In conclusion, cancer cell-derived PDGFB can mediate mTORC1 signalling pathway activation in ccRCC, further consolidating the link between the KLF6-PDGFB axis and the mTORC1 signalling pathway activity in ccRCC

Reverting <i>TP53</i> Mutation in Breast Cancer Cells: Prime Editing Workflow and Technical Considerations

Breast cancer is the leading cause of cancer-related deaths in women. The aggressive breast cancer subtype is commonly linked to the genetic alterations in the TP53 tumor suppressor gene, predominantly the missense mutations. Robust experimental models are needed to gain better insights into these mutations’ molecular properties and implications in tumorigenesis. The generation of such models harboring the alterations is feasible with the CRISPR-based gene editing technology. Moreover, the development of new CRISPR applications, particularly DNA base and prime editing, has considerably improved the precision and versatility of gene editing. Here, we employed the prime editing tool to revert a TP53 missense C > T mutation (L194F) in a T47D luminal A breast cancer cell line. In parallel, this prime editing tool was also utilized to introduce the L194F mutation in HEK293T cells. To assess the prime editing efficiency in both cell lines, we first performed Sanger sequencing in the prime-edited cells pool and single cell-derived clones. However, the Sanger sequencing approach did not detect any base substitution in these cell lines. Next, by employing the more sensitive amplicon target sequencing, we managed to identify the expected substitution in these T47D and HEK293T cells, albeit the editing efficiency was low. In light of these findings, we discussed the technical aspects and provided suggestions for improve the prime editing workflow and efficiency for future experiments