GSE125743
Fractional Deletion of Compound Kushen Injection Indicates Cytokine Signaling Pathways are Critical for its Perturbation of the Cell Cycle
We have used computational and experimental biology approaches to identify candidate mechanisms of action of a traditional Chinese medicine; Compound Kushen Injection (CKI), in a breast cancer cell line in which CKI has been shown to cause apoptosis. Because CKI is a complex mixture of plant secondary metabolites, we used a high-performance liquid chromatography (HPLC) fractionation and reconstitution approach to define chemical fractions required for CKI to induce apoptosis in MDA-MB-231 cells. Our initial fractionation separated major from minor compounds, and showed that the major compounds accounted for little of the activity of CKI. By systematically perturbing the major compounds in CKI we found that removal of no single major compound could alter the effect of CKI on cell viability and apoptosis. However, simultaneous removal of two major compounds identified oxymatrine and oxysophocarpine as critical compounds with respect to CKI activity. We then used RNA sequencing and transcriptome analysis to correlate compound removal with gene expression and phenotype data. We determined that many compounds in CKI are required for its effectiveness in triggering apoptosis but that significant modulation of its activity is conferred by a small number of compounds. In conclusion, CKI may be typical of many plant based extracts that contain many compounds in that no single compound is responsible for all of the bioactivity of the mixture and that many compounds interact in a complex fashion to influence a network containing many targets. Overall design: High-depth paired-end RNA-seq from MDA-MB-231 cell line. Each sample contains 3 biological replicates. N_2(where N represents the total number of compounds in CKI, N_2 represents removal of Omt and Ospc from CKI which is N-OmtOspc), N_3 represents removal of Mac, Omt and Ospc from CKI (hence N-MacOmtOspc), OO represents two compounds only: Omt and Ospc, MOO represents three compounds only: Mac, Omt and Ospc, N_Mac represents removal of Mac from CKI (hence N-Mac), N_Nme represents removal of Nme from CKI, N_Omt represents removal of Omt from CKI, and N_Tri represents removal of Tri from CKI. See the manuscript doi: https://doi.org/10.1101/462135 for more details.