## 4 Discussion
### Selectivity Index of S. triloba for U2 OS and SKOV3 Cells
Additionally, stabilization of p53 by inhibiting MDM2 decreased RUNX2 protein expression [110]. RUNX2 activation during osteoblast differentiation was also dependent on p53 inhibition by MDM2 [111]. Both _RUNX2_ and _MDM2_ were downregulated in SKOV3 cells after _S. triloba_ treatment. Incontrast, _MDM2_ expression was upregulated in U2 OS cells despite _RUNX2_ downregulation. Our Transfac database [44] analysis predicted the presence of several RUNX2 transcription factor binding sites within the MDM2 promoter region. The predicted consensus sequence most likely acts as either positive or negative regulatory elements in different cellular contexts. When bound to the consensus AML site (TG[T/C]GGT), both RUNX1 and RUNX2 negatively regulated promoter activity of the human CLC gene [112; 113]. However, [PERSON] and colleagues noted that different RUNX2 isoforms can either up- or downregulate their target genes [114] when bound to the consensus RUNX2 site (5\({}^{\prime}\)-AACCAC-3\({}^{\prime}\)) [115]. Moreover, _S. triloba_ did not significantly alter _p53_ expression in both SKOV3 and U2 OS cells. [PERSON] and colleagues reported that _S. triloba_ did not affect p53 protein levels in breast cancer cell lines [24]. Hence, it is likely that RUNX2 affects MDM2 activity independent of p53. On the other hand, no RUNX2 transcription factor binding sites were predicted for the SETD7 gene. _SETD7_ upregulation after _S. triloba_ treatment suggests possible p53 stabilization [90] in U2 OS cells. Nevertheless, _SETD7_ expression in SKOV3 cells declined after treatment with _S. triloba_. This disparity in _SETD7_ expression implies that the regulatory pathways modulating _SETD7_ differ from one cellular context to another. Besides, SETD7 was unessential for p53 activation in mouse models [90]. The expression of some genes involved with the RUNX2 and p53 pathways was also assessed after _S. triloba_ treatment. _Vimentin_ and _N-cadherin_ expression was significantly decreased in U2 OS cells after treatment with _S. triloba_. Conversely, although _N-cadherin_ expression in SKOV3 cells declined, the result was not statistically significant. Vimentin and N-cadherin have been implicated as markers of epithelial-mesenchymal transition (EMT) by modulating cell migration and invasion [48; 116]. Generally, N-cadherin promotes both cell migration and invasion in cancer [117], and its expression is correlated with poor survival in ovarian cancer [118]. A few studies however reported that N-cadherin displayed anti-oncogenic properties in both human osteosarcoma tumors and mouse osteosarcoma cell lines where it was downregulated [119]. The downregulation of _N-cadherin_ in U2 OS cells by _S. triloba_ suggests an amelioration of osteosarcoma pathology; nevertheless, the exact role of N-cadherin in osteosarcoma progression remains to be elucidated. Similarly, although the exact role of vimentin in cancer progression and EMT remains unclear, its overexpression is correlated with increased tumor progression, metastasis, invasion and poor prognosis [120]. Finally, the reduced expression of _BAX_ in SKOV3 cells infers that the observed decrease in cell viability after _S. triloba_ treatment was independent of BAX mediated apoptosis [121; 122]. There is growing evidence that both RUNX2 and the PI3K/AKT pathway interact either directly or indirectly to drive tumor progression [123]. On one hand, AKT activity most likely promotes RUNX2 mRNA upregulation, protein stabilization, and transcription activity; while on the other hand, RUNX2 activity stimulates the transcription of _PI3K_ and _AKT_, along with components of the mTOR complex [123]. Since _S. triloba_ treatment significantly reduced the expression of both _RUNX2_ and _PI3K_ in U2 OS cells, it is possible that _S. triloba_ contains bioactive compounds such as phenolic acids, flavonoids, and terpenes [21; 22], that either mutually or independently target both RUNX2 and PI3K/AKT signaling.