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Are current cell-based assays truly reliable for drug screening?
Sun, December 06 2009, 12:00 AM
Posted By:
Sciclips
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Drug discovery is an expensive and a long term process. Though, the technology for drug screening is evolving everyday but only very limited number of drug candidates qualify for the final screening and get approved by FDA. We can see a drastic decline in new drug candidates in the past few years. This is mainly due to the low efficacy and toxicity of the drug on human system.
Cell-based assays are widely used for drug screening and have become powerful tools in the past decade. These cell-based assays are used in cytotoxicity, cAMP assay, ADME/Tox, GPCR, kinase assays etc. Usually, these in-vitro assays are performed in a multi-well microtiter plate. Mammalian cells are cultured in these plates and incubated with the drug for a specific period of time. The viability of the cells were then measured which correlates the toxicity level of the drug. Predictability of these assays are not at all satisfactory as in many cases the drugs fail in the human system, as the in-vitro studies are very different from the biological process inside our body.
Now the question is whether these in-vitro cell based assays are truly reliable in drug screening assays? Our major concern is the reporter driven cell based assays in drug discovery screening. In a reporter based screening assay, reporter protein will be fused with target protein/s or they may be transcriptionally controlled. The examples for reporter fusion proteins are GPCR or HERG channel proteins fused with fluorescence reporter proteins. In a transcriptionally controlled systems, reporter proteins are expressed under the control of a specific cell-signaling promoter, example is NFAT driven expression of GFP for cAMP assay.
Heart is a potential target organ to evaluate in nonclinical and clinical studies during drug development. There are many examples in the past when commercial drugs produced unexpected cardiotoxicity and have to be withdrawn from the market. Mammalian cell lines used in cardiotoxicity studies can be non-cardiac cells modified to express particular ion channels and thus do not reflect the normal physiology of a cardiomyocyte. For example, in order to develop a HERG channel assay for cardiotoxicity screening, one has to genetically fuse HERG channel protein with a marker protein (e.g. a fluorescent protein). The question is can we use non-cardiac cells for cardiotoxicity assays? The answer may be no, non- cardiac cells may not give satisfactory results. The protein content for cardiac and other cells can be different and we do not know the role of these cardiac specific proteins in any cardiac signaling pathway that might affect HERG channel activation or deactivation. Even if we use genetically modified cardiac cells in this assay, potential problems can happen. Like in any cell culture methods, we cant predict whether long-term cell culture may induce any genetic mutations which can affect the assay validation? One of the major limitations with cloned ion channel expression systems is the unstable expression of the intended ion channel protein. The over-expression of a protein, or even the transfection method itself, can also affect various signaling pathways which in turn may affect the assay results. This is also true with any other receptor binding assays for example, GPCR assays. If the receptor of interest is expressed only in cardiac or neuronal cells, how one can rely on an assay that uses a different cell line other than cardiac or neuronal cells? The genetic make-up or the signaling pathways can be different in cardiac or neuronal cells. We may not be able to replicate this is in normal cell line that over expresses a GPCR. The above mentioned points are valid to cAMP or kinase assays as well as to any transcription based cell-based assays. All these factors may contribute to the high failure rate of the drug candidate in the clinical trial.
Use of non-genetically modified (untransfected) cells or tissues (for example, cardiac tissue or cells for HERG channel assay) will be the most suitable drug screening assays. This approach will ultimately help in developing most efficient drugs, with less time and investment. More emphasis should be given, to use natural cells or tissue for developing drug screening assays using innovative approaches or technologies. This will be tough, but not impossible. There are several methods for the screening of cAMP, ADME/Tox using non-genetically modified system (non-GM cell based assays). The researchers and companies who are a heavily involved in developing cell based assays should focus on developing innovative technologies that address the potential problems associated with current cell based assays for drug screening.
Categories:
Drug Discovery
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