Lauss et al. (44 (link)) developed a set of 374 genes associated with breast cancer by compiling a consensus list from 44 published gene lists developed for breast cancer prognosis. This consensus list of 374 genes is also the first entry in GeneSigDB, a gene signature database created from supporting tables of research publications (45 (link)). This 374 gene list was shown to be predictive of recurrence when used as a biomarker-set to classify 1067 breast cancer patients. Since Lauss et al. (44 (link)) showed that probability of expression of these genes is associated with breast cancer recurrence, we pasted the list into the ‘Up List’ text box of the search interface of LCB, checked only the ‘Up’ checkbox, and switched the ‘Aggravate/Reverse’ slide-bar to ‘Reverse’ (Figure 1). After clicking the ‘Search’ button, the top returned experimental conditions are: Tyrphostin AG 1478 applied to A375, a skin cancer cell-line; PD0332991 applied to MDAMB231 and MCF10A breast cancer cell lines; Aminopurvalanol applied to PC3, a prostate cancer cell-line; and a benzenesulfonamide derivative also applied to PC3. Tyrphostin AG 1478 is an EGFR inhibitor, and was already reported to inhibit proliferation and progression of human breast cancer cell-lines (46 (link)). PD0332991 is a kinase inhibitor targeting CDK4/6 and was reported to suppress proliferation of ER+ human breast cancer lines (47 ). Aminopurvalanol is a CDK kinase inhibitor that exerts its effects on the canonical MAPK pathway by suppressing proliferation of several mammalian cell-lines including MCF7, a human breast cancer cell-line (48 ). The benezensulfonamide derivative is a bioactive small molecule of unknown targets and mechanisms. Among these top five matched perturbations, PD0332991 is interesting since the top matches were applied to breast cancer lines and there are three of them. Clicking on the first PD0332991 matched perturbation, the one that was applied to MDAMB231 cells at 2 µm and gene expression was measured at 24 h, we can see that the perturbation is highlighted on the experiment canvas within the average change view. The coloring of the perturbation indicates that it is not a strong perturbation relative to the other perturbations applied to the MDAMB231 cells within this same batch. The 10 µm PD0332991 perturbation, which is another top matched experiment for our query, is located left to the highlighted perturbation, suggesting that there is high similarity in gene expression changes induced signatures for these two perturbations (Figure 5A). WikiPathways (27 (link)) enrichment analysis of the down-regulated genes for this perturbation shows high enrichment for ‘DNA replication’, ‘G1 to S cell cycle control’ and ‘cell cycle’ (Figure 5B and C). This enrichment result is consistent with the knowledge that PD0332991 is a CDK inhibitor. Enrichment analysis using the MGI Mammalian Phenotype gene-set library suggests enrichment for genes that when individually knocked out in mice can cause ‘abnormal cell cycle’ and ‘abnormal cell proliferation’, confirming that PD03322991 is likely attenuating cell proliferation through inhibition of the cell cycle. For the top matched perturbation: Tyrophostin AG 1478 the perturbation appears relatively strong on the experimental canvas (Figure 5D). The WikiPathways enrichment analysis of the down genes shows enrichment for the same cell cycle related terms seen for PD0332991 as well as ‘MAPK pathway’ terms which are not present in the PD0332991 WikiPathways enrichment analysis. This observation is consistent with the notion that Tyrophostin AG 1478 inhibits EGFR. The MGI Mammalian Phenotype enrichment analysis results suggest ‘abnormal cell cycle’ and ‘abnormal tumor incidence’ as the two most highly enriched terms. This is consistent with the knowledge about the anti-tumor effects of Tyrophostin AG 1478 (Figure 5E and F).
While the analysis so far confirms known drugs and their mechanisms of action in cell-lines, LCB is useful for discovering potential mechanisms for new drugs. The benezensulfonamide derivative, the fifth drug on the list, has no known targets or mechanisms. The experimental canvas shows that this perturbation is relatively strong (Figure 5G). The WikiPathways enrichment analysis of the down genes suggest the same three cell cycle related terms shown for PD0332991 and Tyrophostin AG 1478, but also the ‘IL-3 and IL-7 pathways’. This suggests that this compound may be related to a process that inhibits the expression of genes that belong to Interleukin-related pathways. The MGI Mammalian Phenotype enrichment analysis points to tumor proliferation related terms but also contain ‘abnormal blood cell’, which adds support for a potential role for this benezensulfonamide derivative to engage with interleukin-related pathways, since interleukins are expressed in the hematopoietic system and disruption of these pathways can lead to blood cell abnormalities (Figure 5H and I).