Tools for target identification and validation

doi:10.1016/j.cbpa.2004.06.001

Current Opinion in Chemical Biology

Volume 8, Issue 4, August 2004, Pages 371-377

https://doi.org/10.1016/j.cbpa.2004.06.001 Get rights and content

Abstract

Reliable technologies for addressing target identification and validation are the foundation of successful drug development. Microarrays have been well utilized in genomics/proteomics approaches for gene/protein expression profiling and tissue/cell-scale target validation. Besides being used as an essential step in analyzing high-throughput experiments such as those involving microarrays, bioinformatics can also contribute to the processes of target identification and validation by providing functional information about target candidates and positioning information to biological networks. Antisense technologies (including RNA interference technology, which is recently very ‘hot’) enable sequence-based gene knockdown at the RNA level. Zinc finger proteins are a DNA transcription-targeting version of knockdown. Chemical genomics and proteomics are emerging tools for generating phenotype changes, thus leading to target and hit identifications. NMR-based screening, as well as activity-based protein profiling, are trying to meet the requirement of high-throughput target identification.

Introduction

Target identification and validation are the first key stages in the drug discovery pipeline. Therefore, researchers are necessarily concerned with this initial aspect of the drug discovery process.

In the past, researchers had a tendency to work on a handful of favored genes, often identified in the literature by academic groups, amenable to low-throughput analysis. Thus, a majority of successful drug discovery projects have targeted the relatively small numbers of protein classes that have proved amenable to pharmaceutical development. For example, around 40% of marketed small-molecule therapeutics target G-protein coupled receptors [1]. Other favored protein classes include ion channels, proteases, kinases and nuclear receptors 2., 3., 4..

With the publication of the human genome sequence 5., 6., the newly revealed potential target pool shows promising prospect for drug development. Various tools and technologies have been used in different approaches to accelerate target identification and validation. In this review, we explore the recently developed, cutting-edge technologies and their potentials in this field (Figure 1).

Section snippets

Microarrays

Target identification seeks to identify new targets, normally proteins (or DNA/RNA), whose modulation might inhibit or reverse disease progression. Current technologies enable researchers to attempt to correlate changes in gene (genomics) and protein (proteomics) expression with human disease, in the hope of finding new targets. Microarrays are a well-utilized tool in both academic and industrial research laboratories. They can be used to assess gene and protein expression (via nucleic acid or

Bioinformatics

Besides being used as an essential step in analyzing high-throughput experiments such as DNA microarrays, bioinformatics can contribute to the processes of target identification and validation by providing functional information of target candidates and positioning information on the biological networks. Numerous public biological databases are warehousing and providing a great amount of functional information for genes or proteins, and many useful bioinformatics tools are continuously

Antisense technology

A key strategy in target validation is to determine what happens, with respect to phenotype and/or the expression of other genes in cells or model organisms, if a gene of interest is either deleted or its activity is inhibited. Gene knockout mimics the activity of a drug that completely inhibits the normal function of the gene’s product. A temporary knockout, a so called knockdown, is another popular alternative for real-time analysis of the gene function. Several important strategies for gene

Zinc finger proteins

Zinc finger proteins (ZFPs) have remarkable versatility for recognizing different sequences of DNA, and variations in the amino acid sequence of the C2H2 domains allow them to be targeted to different locations in the genome. Each zinc finger is a short stretch of 30 amino acids, containing two conserved cysteines and two conserved histidines. These proteins have been used as the DNA-binding domains of novel transcription factors (ZFP TFs). ZFP TFs can be applied to potential new drug target

Haplotype analysis

Recently, haplotype analysis has attracted more and more interest. The phenomenon of haploinsufficiency, in which loss of function of just one gene copy leads to an abnormal phenotype, has led to the assumption that, regardless of their frequency, haploinsufficient loci define a set of genes whose dosage and function are critical to the organism. Building on this idea, lowering the dosage of a single gene may result in a heterozygote that is sensitized to any drug that acts on the product of

Chemical-driven random mutagenesis

Much of the above discussion concerned experiments in silico and in vitro, rather than whole-organism physiology. These studies still cannot predict the integrated response of a potential drug as accurately as work in living systems, thus in vivo testing is a somewhat more reliable method for target identification and validation [33]. Random mutagenesis of the mouse, driven by N-ethyl-N-nitrosourea, is important for large-scale drug target identification and validation. The technique is used in

Chemical genomics and proteomics

Rather than finding drugs for targets in the conventional pharmaceutical approach, forward chemical genomics, in a sense, finds targets for known drugs. Its goal is to discover the specific molecular targets and pathways that are modulated by particular chemical molecules (i.e. study the biochemistry underling the phenotype changes induced by chemicals).

Activity-based protein profiling

The activity of proteins is regulated by widespread post-translational regulation in vivo, thus protein abundance may not directly correlate with protein activity. Accordingly, methods for activity-based protein profiling (ABPP) may serve as a valuable complement to conventional abundance variations based approaches [38^••]. Since it was hypothesized that chemical probes capable of directly reporting on the integrity of enzyme active sites in complex proteomes might serve as effective

Conclusions

Novel therapeutic target identification and validation is a highly complex and resource-intensive process, which requires an integral use of various tools, approaches and information. It will be largely affected by the genomic and proteomic tools in the post-genome era, which may speed up the whole biological/chemical/medical community, and lead to the high-throughput, low cost, and the means to save time and energy. Currently, microarray technology continues to rapidly identify novel

References and recommended reading

Papers of particular interest, published within the annual period of review, have been highlighted as:

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of special interest
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of outstanding interest

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Citation Excerpt :
Consistently, In vivo results showed that kaempferol (145 μg/kg) or ginkgolide A (205 μg/kg) significantly reduced the pulmonary resistance in mouse model (P < 0.05 vs. methacholine, Fig. 3B). Target identification and validation are the first key stages of the drug development (Stitziel and Kathiresan, 2017; Wang et al., 2004). The three basic strategies, which remain dominant for classical drug target identification, are the physiological, mechanism-driven and gene-driven approaches (Knowles and Gromo, 2003; Lindsay, 2005).
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Tools for target identification and validation

Abstract

Introduction

Section snippets

Microarrays

Bioinformatics

Antisense technology

Zinc finger proteins

Haplotype analysis

Chemical-driven random mutagenesis

Chemical genomics and proteomics

Activity-based protein profiling

Conclusions

References and recommended reading

Drug Discov Today

Drug Discov Today

J Biol Chem

Curr Opin Chem Biol

Nat Rev Drug Discov

Mol Pharmacol

J Am Chem Soc

Trends Biochem Sci

Drug discovery: a historical perspective

Science

Collecting and harvesting biological data: the GPCRDB and NucleaRDB information systems

Nucleic Acids Res

LGICdb: the ligand-gated ion channel database

Nucleic Acids Res

Protein kinase inhibitors: emerging pharmacophores 1997–2000

Expert Opin Ther Pat

The sequence of the human genome

Science

A perspective on protein microarrays

Nat Biotechnol

Identification of novel proteins associated with hepatocellular carcinomas using protein microarrays

J Pathol

Developing a strategy for activity-based detection of enzymes in a protein microarray

ChemBioChem

Identification and characterization of human SNAIL3 (SNAI3) gene in silico

Int J Mol Med

Diffuse large B-cell lymphoma outcome prediction by gene-expression profiling and supervised machine learning

Nat Med

In silico model-driven assessment of the effects of single nucleotide polymorphisms (SNPs) on human red blood cell metabolism

Genome Res