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Chapter 5 : Cell-Based Screening Methods for Anti-Infective Compounds

Authors: Stefano Donadio, Margherita Sosio
Content Type: Reference
Publication Year: 2010

Category: Applied and Industrial Microbiology

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Abstract:

The chapter talks about selected cell-based assays, and the selection of the corresponding target(s), amenable for the discovery of novel antibacterial and antifungal agents from microbial products, screened as complex mixtures resulting from partial processing of fermentation broths. The authors also provide a framework to view target choice and assay design in the context of the microbial diversity available for screening, highlighting the importance of designing an overall strategy for an effective drug discovery program. In the anti-infective field, a drug discovery program based on microbial products has the ultimate goal of discovering a new, patentable chemical entity possessing desired properties, such as antimicrobial spectrum, molecular weight, solubility, and preferred route of administration. Cell-free assays directly measure the effect of a sample on the biological activity of one or more relevant targets. In general, these types of assays are more specific and sensitive than cell-based assays. The objective of the assay phase is to ensure that the assay is more sensitive than growth inhibition of the same test strain. The ultimate goal of microbial product screening is to establish the chemical identity of the hits and their biological properties. In antibacterial and antifungal screening programs, hits are characterized for their activity on different microbial strains.

Citation: Donadio S, Sosio M. 2010. Cell-Based Screening Methods for Anti-Infective Compounds, p 62-72. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch5

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Cell-Based Screening Methods for Anti-Infective Compounds
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Citation: Donadio S, Sosio M. 2010. Cell-Based Screening Methods for Anti-Infective Compounds, p 62-72. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch5
/content/10.1128/9781555816827.ch05.ch05fig01
FIGURE 1
Scheme of a screening process.
10.1128/9781555816827/f0063-01_thmb.gif
10.1128/9781555816827/f0063-01.gif
Image of FIGURE 1
FIGURE 1

Scheme of a screening process.

Citation: Donadio S, Sosio M. 2010. Cell-Based Screening Methods for Anti-Infective Compounds, p 62-72. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch5
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/content/10.1128/9781555816827.ch05.ch05fig02
FIGURE 2
From target to screens. Main conceptual passages required (top) and the experimental steps required at each stage (bottom).
10.1128/9781555816827/f0064-01_thmb.gif
10.1128/9781555816827/f0064-01.gif
Image of FIGURE 2
FIGURE 2

From target to screens. Main conceptual passages required (top) and the experimental steps required at each stage (bottom).

Citation: Donadio S, Sosio M. 2010. Cell-Based Screening Methods for Anti-Infective Compounds, p 62-72. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch5
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/content/10.1128/9781555816827.ch05.ch05fig03
FIGURE 3
Concept of reporter assays. In a bacterial cell, the antibiotics 1 and 2 act on the different pathways and , respectively (top panel), leading to pathway perturbation due to specific stress-response signals and upregulation of genes under the same stimulon, as exemplified by the promoters p and p, which responds to perturbations in pathways and , respectively (middle panel). Two reporter assays can be designed by engineering cells so that expression of the reporter gene is under the control of the inducible promoter p or p, resulting in increased activity of the reporter enzyme Enz (bottom panel).
10.1128/9781555816827/f0067-01_thmb.gif
10.1128/9781555816827/f0067-01.gif
Image of FIGURE 3
FIGURE 3

Concept of reporter assays. In a bacterial cell, the antibiotics 1 and 2 act on the different pathways and , respectively (top panel), leading to pathway perturbation due to specific stress-response signals and upregulation of genes under the same stimulon, as exemplified by the promoters p and p, which responds to perturbations in pathways and , respectively (middle panel). Two reporter assays can be designed by engineering cells so that expression of the reporter gene is under the control of the inducible promoter p or p, resulting in increased activity of the reporter enzyme Enz (bottom panel).

Citation: Donadio S, Sosio M. 2010. Cell-Based Screening Methods for Anti-Infective Compounds, p 62-72. In Baltz R, Demain A, Davies J, Bull A, Junker B, Katz L, Lynd L, Masurekar P, Reeves C, Zhao H (ed), Manual of Industrial Microbiology and Biotechnology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555816827.ch5
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