High-Throughput Screening Platform for Anticancer Therapeutic Drug Cytotoxicity

Abstract

There are substances that kill cancer cells, but induce T cell proliferation, like thalidomide. To find more of these, a new anticancer drug screening strategy is vital. In this study we report the development of a differential cytotoxicity screening or evaluation platform using the CellTiter-Glo (Promega, Annandale, NSW, Australia) luminescent cell viability assay (ATP assay) and also the CellTiter 96 AQueous (Promega) one solution cell proliferation assay [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt (MTS) assay]. The results suggested the platform consisting of the combination of the ATP assay be used for quantifying peripheral blood mononuclear cells, while the more economic MTS colorimetric assay is well suited to be used detecting cell viability of cancer cells. In addition, we found paclitaxel (Taxol, MP Biomedicals Australasia Pty Ltd., Seven Hills, NSW, Australia) to be a useful control for this routine screening methodology. Taxol exhibits the desirable in vitro feature of differential cytotoxicity that spares the immunological cells, when used at a concentration that will kill the majority of the cancer cell population.

Genomic Screening for Targets Regulated by Berberine in Breast Cancer Cells

“…Tissue inhibitors of MMP (TIMP3), which is up-regulated after berberine expose in MCF-7 cells, has been recognized as potential inhibitors of angiogenesis and cancer invasion and metastasis (Ahonen et al, 2002). Interestingly, it demonstrated that berberine suppressed the activities of MMP1, MMP2, MMP9 (Sekhon et al, 2008;Kim et al, 2012), therefore, our results suggested that berberine inhibited the activities of MMPs may be via up-regulation of TIMP3 expression. SLC7A11, which mediates cystine -glutamate exchange and thereby regulates intracellular glutathione (GSH) levels, play an important role in maintaining the higher GSH and consequently in cisplatin resistance (Okuno et al, 2003).…”

“…The cell viability was determined by MTS ((Promega, Madison, WI) assay (Sekhon et al, 2008). Briefly, 5×10 3 cells per well were seeded in 96-well culture plates in triplicate and allowed to attach overnight.…”

Monitoring cellular accumulation of 3′-deoxy-3′-fluorothymidine (FLT) and its monophosphate metabolite (FLT-MP) by LC–MS/MS as a measure of cell proliferation in vitro

“…In identical parallel assays, FLT-MP formation was compared with cellular ATP generation, measured using the CellTiter-Glo ® assay. ATP generation is well-correlated with the number of viable cells in a culture cell, and this endpoint assay is widely used as a surrogate measure of cell proliferation [27]. As shown in Fig.…”

In vitroend points for the assessment of cellular immune response-modulating drugs

“…To this purpose, new non-radioactive kits are disposable for valuation of specific T-lymphocyte cytotoxicity. These methods provide a measurement of cytotoxicity by using an indirect indicator, such as the release of LDH [144] or ATP enzyme [145], or by the capacity of viable cells to incorporate alamar blue dye, an indicator of cell metabolic activity [146]. Cytolytic activity can also be assayed by flow cytometry that allows quantitative measurements further distinguishing between apoptosis and necrosis.…”