Cell cycle, energy metabolism and DNA repair pathways in cancer cells are suppressed by Compound Kushen Injection.

BACKGROUND: In this report we examine candidate pathways perturbed by Compound Kushen Injection (CKI), a Traditional Chinese Medicine (TCM) that we have previously shown to alter the gene expression patterns of multiple pathways and induce apoptosis in cancer cells. METHODS: We have measured protein levels in Hep G2 and MDA-MB-231 cells for genes in the cell cycle pathway, DNA repair pathway and DNA double strand breaks (DSBs) previously shown to have altered expression by CKI. We have also examined energy metabolism by measuring [ADP]/[ATP] ratio (cell energy charge), lactate production and glucose consumption. Our results demonstrate that CKI can suppress protein levels for cell cycle regulatory proteins and DNA repair while increasing the level of DSBs. We also show that energy metabolism is reduced based on reduced glucose consumption and reduced cellular energy charge. RESULTS: Our results validate these pathways as important targets for CKI. We also examined the effect of the major alkaloid component of CKI, oxymatrine and determined that it had no effect on DSBs, a small effect on the cell cycle and increased the cell energy charge. CONCLUSIONS: Our results indicate that CKI likely acts through the effect of multiple compounds on multiple targets where the observed phenotype is the integration of these effects and synergistic interactions.

An algorithm for standardization of tumor Infiltrating lymphocyte evaluation in head and neck cancers.

PURPOSE: The prognostic and predictive significance of pathologist-read tumor infiltrating lymphocytes (TILs) in head and neck cancers have been demonstrated through multiple studies over the years. TILs have not been broadly adopted clinically, perhaps due to substantial inter-observer variability. In this study, we developed a machine-based algorithm for TIL evaluation in head and neck cancers and validated its prognostic value in independent cohorts. EXPERIMENTAL DESIGN: A network classifier called NN3-17 was trained to identify and calculate tumor cells, lymphocytes, fibroblasts and "other" cells on hematoxylin-eosin stained sections using the QuPath software. These measurements were used to construct three predefined TIL variables. A retrospective collection of 154 head and neck squamous cell cancer cases was used as the discovery set to identify optimal association of TIL variables and survival. Two independent cohorts of 234 cases were used for validation. RESULTS: We found that electronic TIL variables were associated with favorable prognosis in both the HPV-positive and -negative cases. After adjusting for clinicopathologic factors, Cox regression analysis demonstrated that electronic total TILs% (p = 0.025) in the HPV-positive and electronic stromal TILs% (p < 0.001) in the HPV-negative population were independent markers of disease specific outcomes (disease free survival). CONCLUSIONS: Neural network TIL variables demonstrated independent prognostic value in validation cohorts of HPV-positive and HPV-negative head and neck cancers. These objective variables can be calculated by an open-source software and could be considered for testing in a prospective setting to assess potential clinical implications.