We used a complementary gene expression profiling using the DNA microarray technology to monitor cellular changes in gene expression and discover the molecular targets upon RUNX2 suppression in EOC cells. To our knowledge, the present work represents the first effort to define global changes in gene expression upon modulation of RUNX2 expression in cancer cells. We analyzed both functionally related genes that were commonly differentially expressed in SKOV3 EOC cells upon RUNX2 knockdown. The gene expression data and consecutive network and pathway analyses were quite confirmatory of the data obtained by the RUNX2 functional assays. Indeed, microarray data sustained RUNX1 correlation with EOC cell proliferation, migration and invasion, since RUNX2 knockdown resulted in reduced expression of genes associated with metabolism, cellular growth & proliferation and cellular movement, while a number of genes linked to cell death were induced. IPA network analysis was indicative for some important gene nodes linked to RUNX2 suppression in EOC cells, as most of these substantiate and/or complement the functional data obtained. Thus, RUNX2 knockdown resulted in (+)-JQ1 upregulation of gene nodes/genes known to be implicated in apoptosis induction or displaying TSG functions. Notably the UBC interaction network and its members were shown to decrease in anchorage-independent cell growth and increase apoptosis, suggesting UBC may act as a negative regulator of skin carcinogenesis; CRYAB has been reported as a potential TSG, while increased expression of BCL-XS protein in tumors was associated with decreased proliferation and induction of apoptosis. Similarly, CTGF upregulation was found to be associated with apoptosis and decrease of tumor cell invasion; PPM1A expression could induce cell cycle arrest and apoptosis via activation of the p53 pathway, and NF2 has been characterized as a TSG in different cancers. Given the similar roles of RUNX1 and RUNX2 in EOC progression and the fact that all three RUNX proteins recognize common DNA sequence motifs, we analyzed the extent of overlap in differentially expressed genes/functional pathways following RUNX1 and RUNX2 knockdown in the SKOV3 ovarian adenocarcinoma cell line. Both the Venn diagram comparisons, as well as gene clustering and IPA functional analyses were indicative for distinct molecular mechanisms and functional pathways associated with RUNX1 or RUNX2 
implication in EOC progression, although both genes could potentially modulate the expression of some common genes involved in EOC disease advancement and metastasis. In conclusion, we have shown that the RUNX2 transcription factor is significantly overexpressed in serous EOC tumors, including LMP tumors, compared to normal ovarian tissue. BSP validation of the RUNX2 methylation status in primary EOC tumors and omental metastasis were indicative for no implication of LY2835219 epigenetics mechanisms in RUNX2 overexpression in metastatic tissues. Further functional analyses of RUNX2 in EOC cells pointed towards its association with EOC cell proliferation, migration and invasion. Gene expression profiling and consecutive network and pathway analyses confirmed these findings, as various genes and pathways known previously to be implicated in ovarian tumorigenesis, including EOC tumor invasion and metastasis, were found to be suppressed upon RUNX2 knockdown, while a number of pro-apoptotic genes and some EOC TSGs were found to be induced. Our data suggest that RUNX2 is possibly implicated in EOC tumor and cancer cell growth and invasion and could represent a potential EOC therapeutic target. The present study also reveals that RUNX1 and RUNX2 employ distinct molecular mechanisms in EOC tumorigenesis despite evident similarities of their action on EOC cell phenotype and behavior.