THERAPEUTICSTRATEGIES

DRUG DISCOVERY

TODAY

Drug Discovery Today: Therapeutic Strategies Vol. 1, No. 4 2004

Editors-in-Chief

Raymond Baker – formerly University of Southampton, UK and Merck Sharp & Dohme, UK

Eliot Ohlstein – GlaxoSmithKline, USA

Infectious diseases

Finding the gems using genomicdiscovery: antibacterial drug discoverystrategies – the successes and thechallengesPan F. Chan, David J. Holmes, David J. Payne*Department of Microbiology, Microbial, Musculoskeletal and Proliferative Diseases, Center of Excellence in Drug Discovery, Anti-infectives Research (UP1345),

GlaxoSmithKline, 1250 South Collegeville Road, Collegeville, PA 19426, USA

Since the first bacterial genome was sequenced (1995)

many companies have invested in a variety of antibiotic

discovery strategies but few novel-acting antibacterial

agents have reached human trials. This is not through

lack of trying, as from literature reports alone >125

published studies on high-throughput screens of >60

different novel targets (from 34 companies) have been

identified. Post-genomics should be a new golden era of

antibiotics but there are substantial challenges of deli-

vering much needed, novel-acting antibacterials that

will be discussed.

*Corresponding author. (D.J. Payne) david_j_payne@gsk.com

1740-6773/$ � 2004 Elsevier Ltd. All rights reserved. DOI: 10.1016/j.ddstr.2004.11.003

Section Editor:Gary Woodnutt – Diversa Corp., San Diego, CA, USA

The explosion in the bacterial genomics area in the past decade has

been remarkable, but there appears to have been little reward for all of

this effort as judged by the relative lack of new antibacterial agentsentering development. Chan et al. review these developments and

suggest that, as far as target identification is concerned, the use ofgenomics has provided us with a wealth of potentially attractive

opportunities, and failure to consolidate on these reflect on otherdeficits/difficulties in the antibacterial drug discovery process.

Furthermore, the knowledge gained during this period has allowedconsiderable advances in genetic tools that can assist in tracking

structure–activity relationships in the whole cell, and should enable thedissection of compound mode of action. In addition, these advances

have been pivotal in understanding why certain promising candidatesare less viable. Thus, the bacterial genomics era is far from being over

and might actually have just begun to be applied appropriately.

Introduction

In the past 20 years, only two completely novel classes of

antibiotics have reached the market to combat the clinical

threat of multidrug-resistant bacteria (Infectious Disease

Society of America, http://www.idsociety.org/Template.

cfm?Section=AntimicrobialsTemplate=/ContentManagement/

ContentDisplay.cfmContentID=9718). Furthermore, there is

apparently only one novel-acting, systemic agent in the

antibiotic development pipeline [1]. Despite the medical

need for new antibiotics [2] there has been a trend for some

large pharmaceutical companies to downsize or abandon

their antibacterial drug discovery efforts [3]. Reasons for this

withdrawal are complex and multi-factorial but one very

significant factor is that in retrospect, it is significantly more

challenging to deliver novel classes of antibiotics than was

predicted nine years ago when the first bacterial genome was

completed. In this review, we discuss three strategies for

finding new antibiotics – (i) target-based screening (ii) anti-

bacterial whole-cell screening and (iii) structural genomics –

and examine how genomics enable or facilitate each of these

strategies. In addition, we discuss the significant challenges

that compromise the success of each approach.

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Drug Discovery Today: Therapeutic Strategies | Infectious diseases Vol. 1, No. 4 2004

Glossary

Chemical diversity: different pharmacophores and scaffolds of mole-

cules in a compound-collection. Typically, a compound library bank that is

used in HTS for inhibitors contain a large, chemically diverse range of

compounds, synthesized in combinatorial libraries or gathered from

natural-sources. Lipinski has applied computational and experimental

physical-property measurements to propose the ‘‘drug-like properties’’

of small-molecule inhibitors [66].

High-throughput screens (HTS): automated and rapid method for

identifying compounds that inhibits enzyme activity or bacterial growth.

Assay usually performed in a 96-, 384- or 1536- well-plate format and

screened against a large compound-collection.

Mode-of-action (MOA): mechanism by which a drug exerts its killing

or inhibitory effect on bacteria.

Targets: proteins that are inhibited by antibacterial agents. Microbial

drug targets are typically essential for bacteria to survive, hence, its

inhibition results in bacterial death or reduced ability to cause an

infection.

Target-based screening

To illustrate the impact of genomics on antibiotic drug dis-

covery, a literature search was conducted to identify the

number of published, antibacterial, HIGH-THROUGHPUT SCREENS

(HTS; see Glossary) reported since the release of the first

microbial genome. Fig. 1 summarizes our findings. This graph

shows there has been a clear increase in the number and

diversity of HTS of antibacterial TARGETS (see Glossary) devel-

oped for finding novel-acting antibiotics between 1995 and

2004 that probably reflect the industry’s investment in a

genomics-driven strategy. During this period, 127 antimicro-

bial screens were reportedly run against almost 70 different

bacterial targets, of which about 60 can be considered as

novel targets. However, this survey is clearly a significant

underestimate of the number of antibacterial screens run by

the pharmaceutical industry since much of that information

is proprietary.

Genomics has delivered a large number of diverse and well-

validated, novel drug targets for target-based screening for

novel-acting inhibitors [4–6] (Figs. 1, 2). Although lead anti-

bacterial compounds have been reported to several novel

classes of targets such as those to two-component signal

transduction systems (TCSTS) [4,7], coenzyme metabolism

[8], cell division [9], protein secretion [10] and novel DNA

replication proteins [11,12], only inhibitors to polypeptide

deformylase (PDF) have reached Phase I trials thus far.

Furthermore, to our knowledge, new antibacterial series iden-

tified via this strategy with in vivo efficacy are limited and

have only been reported for four novel targets namely PDF

[13,14], LpxC [15], FabI [16] and aminoacyl-tRNA synthetases

[17,18] and these will be discussed in the following sections.

In addition, rapid target identification in different patho-

genic bacteria has facilitated the development of HTS to well-

validated, biosynthetic pathways that are under-exploited as

antibacterial targets [19]. The impact of genomics on path-

520 www.drugdiscoverytoday.com

way-based screens to identify novel antibacterial agents will

also be discussed.

Polypeptide deformylase (PDF) target

PDF encoded by def (GenBank accession number X77800), is a

well-validated antibacterial target for drug discovery [6,13,

14]. Vircuron Pharmaceuticals (http://www.vicuron.com/)

in collaboration with Novartis (http://www.novartis.com/)

discovered a series of potent broad-spectrum PDF inhibitors,

of which LBM-415 is currently in Phase I clinical trials for oral

treatment of community-acquired pneumoniae [14,20].

Independently, Oscient Pharmaceuticals (http://www.os-

cient.com/) are working on another series of PDF inhibitors

that were originally identified by HTS of specific chemical

libraries targeted against metalloproteases [13]. Following

structural optimization, BB-83698 was the first compound

of its class to enter clinical trials [21]. Recent reports, how-

ever, suggest that second generation compounds with

improved Haemophilus influenzae activity may be developed

[22]. Numerous pharmaceutical companies have published

studies on PDF as an antimicrobial target [23-28]. Genomics

has also helped address the issue of development of in vitro

resistance to anti-PDF compounds. In the genome sequences

of some pathogens, the formyltransferase (fmt) (GenBank

accession number X63666) gene has been found that

bypasses the formylation pathway though these resistant

mutants show reduced virulence in vivo [15,28]. Indeed,

PDF inhibitors might prove to be the first novel class of

antibiotic to reach the market as a result of target-screening

approaches in the post-genomics era.

LpxC target

Although inhibitors with in vivo efficacy have been described

that target LpxC (GenBank accession number AAC73207), an

essential metalloenzyme involved in lipid A biosyntheis in

Gram-negative pathogens, these compounds have not pro-

gressed beyond pre-clinical stage [15].

Fatty acid biosyntheis (FAB) targets

Comparative genomics has identified all the enzymes

involved in FAB pathway in bacteria and some of the targets

have been validated with antibacterial compounds [29].

Enoyl-ACP reductase encoded by fabI (GenBank accession

number AF197058) has probably yielded the most promising

leads to date with this target screened by several companies

[16,29,30]. Leads originating from HTS have been described

by GlaxoSmithKline (http://www.gsk.com/) with efficacy in

animal infection models and exquisite antibacterial activity

against multi-resistant strains of Staphylococcus aureus (MIC90

of <0.06 mg/ml) [16]. Furthermore, Genome Therapeutic

Corporation (http://www.oscient.com/) describe a new series

of FabI inhibitors with IC50 of 4 mM and MIC of 2 mg/ml

against S. aureus, adding a new chemical scaffold for devel-

Vol. 1, No. 4 2004 Drug Discovery Today: Therapeutic Strategies | Infectious diseases

Figure 1. Literature survey of published antibacterial HTS that use genomics for finding novel-acting antibiotics since 1995. A survey of the literature

identified 127 antibacterial screening programs published by companies exemplifying the genomic strategies discussed in this review. The number of published

antibacterial HTS covers the period since the public release of the first bacterial genome in 1995 to present. Antibacterial screens have been classified by the

year of their journal publications and sub-classified by the target class or function where applicable. Further analysis of the 127 antibacterial HTS indicate that

69 different bacterial targets were screened by 34 different pharmaceutical or biotechnology companies, of which 61 of these targets are novel. Published

reports of novel targets where no HTS or chemistry efforts are mentioned have not been included in this survey. Clearly this graph is an under-representation

of the total number of targets screened industry-wide over this period. For example, GlaxoSmithKline alone have disclosed running more than 50 antibacterial

HTS [67]. * In 2000, both Cubist (http://www.cubist.com/) and GlaxoSmithKline (http://www.gsk.com/) report HTS to nearly all the 19 aminoacyl-tRNA

synthetase targets from S. aureus [32,33]. Abbreviations: HTS, high-throughout screen; TCSTS, two-component signal transduction systems.

oping anti-FabI compounds [30]. Genomics can also help to

illustrate the limitations of novel targets, for example, in

Streptococcus pneumoniae, the enoyl-ACP reductase function

is performed by FabK (GenBank accession number

AAK99183) which is structurally different to FabI [31], and

thus FabI leads are unlikely to have robust antibacterial

activity against S. pneumoniae. Other Fab enzymes, which

are conserved in bacteria, can be developed into target-based

screens for identifying potential broad-spectrum inhibitors,

although only inhibitors to FabH (GenBank accession num-

ber BAB57145) have been reported [29].

Aminoacyl-tRNA synthetase targets

Aminoacyl-tRNA synthetases are a clinically validated class of

enzymes for antimicrobial development [5]. GlaxoSmithK-

line and Cubist Pharmaceuticals (http://www.cubist.com/)

report HTS to all the 19 aminoacyl-tRNA synthetases from

S. aureus [32,33] and finding inhibitors to histidyl- [34],

isoleucyl- [35,36], methionyl- [17,37], phenylalanyl- [38],

seryl- [39], tryptophanyl- [40] and tyrosyl-tRNA synthetases

[41]. To our knowledge, leads to phenylalanyl-tRNA synthe-

tase (FRS) (GenBank accession number P07395) and methio-

nyl-tRNA synthetase (MetS or MRS) (GenBank accession

number BAB41678) are the most advanced. The FRS inhibi-

tors discovered by Bayer (http://www.bayer.com/) achieve

broad-spectrum activity (MIC < 1 mg/ml), potent IC50s in

the nM range, and in vivo efficacy [18]. The MRS leads

identified by GlaxoSmithKline have excellent antibacterial

activity against drug-resistant staphylococci and enterococci

(MIC < 0.06 mg/ml), potent IC50s (�10 nM) and in vivo effi-

cacy demonstrated in a S. aureus infection model [17]. How-

ever, these leads lacked antimicrobial activity against >40%

of clinical isolates of S. pneumoniae that was later accounted

for by the presence of a second functional copy of MetS

encoded by the gene metS2 (GenBank accession number

AY198311) [42]. MetS2 protein is distantly related to MetS

and this thus creates an unexpected complication to securing

good activity against S. pneumoniae with these leads. Further-

more, MetS2 is absent from the two completed genomes of S.

pneumoniae hence exemplifying the importance of having

multiple genome sequences from each pathogen [42].

Pathway-based screens

Genomic approaches have supported the identification and

validation of multiple enzymes of essential biosynthetic

pathways for target-based screening (Fig. 2). Pathway-based

screens have the advantage of targeting more than one

essential protein in a biosynthetic pathway with the potential

to identify inhibitors to more than one target. Generating the

complex substrates for each of the six enzymes in the Mur

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Drug Discovery Today: Therapeutic Strategies | Infectious diseases Vol. 1, No. 4 2004

Figure 2. Integrated strategies for antibiotic drug discovery using genomics. Antibacterial drug discovery strategies that use genomics for finding novel-acting

antibiotics can be divided into three areas: target-based screening, antibacterial whole-cell screening and structural genomics/screening. Once a hit molecule is

identified from screening, the target of the molecule is determined (whole-cell screen) or confirmed (target-based screen) using several MOA assays/tools. A

chemistry lead-optimization program is initiated to synthesise chemical derivatives of the hit compound with the aim of incorporating improved antibacterial

activity and other desirable clinical properties to the molecule. This flowchart only shows genomic tools used for MOA/SAR tracking, and does not include

biochemical and genetic assays reviewed elsewhere [5].

Abbreviations: HT, high-throughput; HTS, high-throughout screen; MOA, mode-of-action; OE, overexpression; ORF, open reading frame; SAR, structure–

activity relationship.

pathway involved in cell wall biosynthesis (MurA-F) (Gen-

Bank accession numbers AAC76221, AAC76950, AAC73202,

AAC73199, AAC73196, AAC73197) has likely led to the

underexploitation of the Mur enzymes as targets, while a

pathway assay circumvents the need to generate these sub-

strates [19]. Similarly, high-throughput pathway screens for

inhibitors of multiple membrane-associated proteins of pep-

tidoglycan biosynthesis that include MraY (GenBank acces-

sion number AAC73198), MurG (GenBank accession number

AAC73201) and penicillin-binding proteins (PBPs) have been

reported [43–45].

Another example of a pathway-based antibacterial screen is

an assay that targets the shikimate (Aro) pathway. The Aro

pathway is a multi-enzyme, essential process in the biosynth-

esis of chorismate, ubiquinone and folate in diverse numbers

of bacteria. An optimized, homogeneous assay of all four

essential enzymes, AroB, AroD, AroE and AroK (GenBank

accession numbers AAL00037, AAL00039, AAL00038,

AAL00032) has been developed for HTS (Goryanin I. et al,

Glaxo SmithKline, submitted for publication).

Although the essential proteins of the peptidoglycan bio-

synthesis pathway have been widely screened in pathway

assays by several groups, to date there have been no reports of

522 www.drugdiscoverytoday.com

sustainable leads from these types of screens progressing to

clinical development.

In summary, a large number and variety of novel geno-

mics-derived targets have been screened by many companies

for finding new antibiotics but the approach has delivered

relatively few new chemical series with in vivo efficacy. It is

likely that the main causes of this are the lower than expected

hit rate at HTS and that the chemical optimization of HTS hits

is far more challenging than making new derivatives of

established classes of antibiotics. In addition, progressing

entirely novel antibacterial targets can be further compli-

cated by ‘‘novel target surprises’’, such as the discovery of

a second MRS in S. pneumoniae.

Antibacterial whole-cell screening

A second strategy for identifying potential novel-acting anti-

bacterials is the HTS of large combinatorial chemical libraries

and new natural products for their ability to inhibit growth of

whole bacterial cells. Most currently marketed antibiotics

were originally discovered by random screening of com-

pounds from natural sources. Once hits have been identified,

there follows the complex task of identifying the molecular

target of the antibacterial hit as well as ensuring that the

Vol. 1, No. 4 2004 Drug Discovery Today: Therapeutic Strategies | Infectious diseases

activity is selective using biochemical and genetic tools [5].

Recent developments in expression technologies that have

facilitated MODE-OF-ACTION MOA; see Glossary) studies of com-

pounds of unknown mechanism are discussed below (Fig. 2).

Regulated strains as MOA tools

Overexpression-gene and antisense-RNA libraries have been

constructed that cover almost every single open reading

frame (ORF) in the S. aureus genome [46,47]. In theory, these

libraries enable hundreds of targets to be rapidly screened as

the potential molecular targets of whole-cell assay hits [47].

This approach has been validated for established antibiotics

of known inhibitory mechanisms but there are no reports

where this approach has identified the MOA of an unchar-

acterized antibacterial.

Proteome- and genome-profiling as MOA tools

Expression profiling can also be potentially powerful tools for

antibacterial MOA studies [48]. One successful application of

gene arrays has been the confirmation of a compound, ori-

ginally identified by Abbott (http://www.abbott.com/) from a

coupled transcription–translation HTS, as a ribosomal inhi-

bitor by comparative profiling [49]. Recently, Bayer has used

expression profiling to elucidate the MOA of two whole-cell

hits as acetyl-CoA carboxylase, AccA (GenBank accession

number BAB57862) and peptidyltransfer inhibitors, respec-

tively. [50,51]. These are some of the firsts examples of

proteome- and genome-profiling being used to identify the

MOA of an antimicrobial agent of unknown mechanism [48].

Whole-cell screening strategies targeting known MOAs

Whole-cell screens specific for targets have also been devel-

oped [52]. An advantage of this strategy is that a particular

enzyme is screened in situ so that the characteristics of an

antibacterial (for example, cell penetration) are already a

prerequisite to obtaining a hit. Screening of an Escherichia

coli strain with downregulated MurA (GenBank accession

number AAC76221) successfully identified a series of MurA

inhibitors that exemplifies the potential of the approach [53].

Another method exploits whole-cell pathway reporter strains

containing antibiotic-responsive promoters often of stress-

inducible genes, fused to a sensitive reporter gene [54,55]. A

Bacillus subtilis strain carrying a FabH reporter screened

against a chemical bank, successfully identified hits by induc-

tion of the reporter that showed weak antibacterial activity

against S. aureus, and specific inhibition of fatty acid bio-

synthesis [54].

Overall, whole-cell screening has some key advantages over

enzyme-based screening for finding novel antibacterial

agents (Table 1). However, success at defining MOA of hits

to enable rational SAR programs has been low, which might

be a function of the diversity of the compound-collection

screened. Using a whole-cell screening approach, GlaxoS-

mithKline have discovered a class of benzylidenethiazoli-

diones with Gram-positive activity though the molecular

target has yet to be reported [56], while Pharmacia (http://

www.pfizer.com/) identified a translation elongation factor-

Tu inhibitor whose MOA was elucidated using ligand-affinity

fishing and macromolecular synthesis [57]. Bayer has demon-

strated the use of expression analysis to identify novel inhi-

bitors of acetyl-CoA carboxylase and peptidyltransferase

activities [48,50,51]. Many genomic tools are now available

to study MOA of novel agents. In addition to identifying

antibacterial targets, these MOA tools are also valuable for

confirming the activity of target-based screening hits and

allowing rapid monitoring of SAR to ensure that the target

does not shift during a lead-optimization program (Fig. 2).

Structural genomics

To find novel-acting antibacterial agents, structure-based

drug design is a key strategy widely employed to assist che-

mical optimization of leads identified from antibacterial

target-based screens (Table 1). Genomics has accelerated

the identification of targets and enabled high-throughput

cloning, expression and purification of novel target proteins

for crystallisation studies. 3D structures can be exploited by

virtual screening of compound databases using docking tech-

niques that provide hits in addition to those predicted by in

silico modelling approaches [58]. Finally, structural genomics

can facilitate studies to identify the function of essential

proteins that have no significant homology with previously

characterized proteins and termed ‘‘proteins of unknown

function’’ (Fig. 2).

Structural genomics of novel targets

Extensive genomic and structural information on novel tar-

gets help characterize the active sites of targets across differ-

ent pathogens, and enable robust and rational, lead-

optimization programs to improve the inhibitory potency,

and potentially expand the antibacterial spectrum of lead-

chemical series. PDF is a good example to illustrate the impact

of structural genomics on a lead-optimization program. Sev-

eral co-crystal structures of the PDF target and anti-PDF lead

compounds have been described that have helped guide

design of potent inhibitors [13,14,24,25,28]. Furthermore,

at the 44th Interscience Conference on Antimicrobial Agents

and Chemotherapy (ICAAC) a new class of broad-spectrum

inhibitor that dual targets topoisomerase IV ParE (GenBank

accession number AAC76066) and DNA gyrase GyrB (Gen-

Bank accession number AAT48201) subunits was reported by

Vertex Pharmaceuticals (http://www.vrtx.com/) that also

achieves in vivo efficacy in animal models [59].

Structural genomics of targets of unknown function

Structural genomics is particularly useful for identifying the

function of novel protein of unknown function [60]. At

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Table 1. Comparison of strategies for antibiotic drug discovery using genomics

Strategies Pros Cons Latest developments Who is working on

this strategy?a,b

Refs

Target-based

screening

� Novel drug-target with no

pre-existing clinical resistance.

� Target validation tools.

� Known MOA.

� Multiple-enzymes might be targeted

in a single HTS run and identify

inhibitors of >1 target.

� Pathway assays enable screening

of targets not amenable to

standard single-target HTS

(do not need to generate

complex substrates).

� Enzyme hits (do not need

to penetrate bacterial cell).

� SAR assay available for tracking MOA.

� Low conservation of

target can result in narrow-

spectrum activity.

� To date, 165 whole microbial

genomes completed and 495

bacterial genome sequencing

projects ongoing.

ActivBiotics, Arrow Therapeutics,

AstraZeneca, Aventis, Basiliea,

Bayer, Bristol-Myers Squibb,

British Biotech, Cubist, Eli Lilly,

Genome Therapeutics,

GlaxoSmithKline, Johnson &

Johnson, Merck, Millenium,

Novartis, Oscient, Pfizer,

Proctor & Gamble, Replidyne,

Roche, Schering-Plough,

Vicuron,Wyeth

[9–21,27–30,36,43–45,

68-70]

� Target might be absent in

genomes of some strains. � Genomes of multiple strains.

� Target might not be amenable

to assay development.

� Many novel, validated targets.

� Deconvolution of pathway-hits

to determine the target.

� FabI, MRS, FRS, PDF inhibitors

illustrate proof of concept.

� Difficult to engineer in antibacterial

activity to hit molecule.

� Many target HTS failed

to produce sustainable leads.

� Dependent on hits from HTS,

usually low success.

Antibacterial

whole-cell

screening

� Most marketed antibiotics

discovered this way.

� Hits possess antibacterial-

activity and can penetrate

bacterial cell.

� Can inhibit more than

one target.

� Unknown MOA.

� Labor-intensive elucidation of

MOA often gives

inconclusive results.

� Challenging to differentiate

non-specific membrane effects

vs. target effects.

� Many hit known targets.

� Dependent on hits from

HTS, usually low success.

� Examples of proteome-and genome-

profiling elucidating

MOAs of antibacterial compounds of

unknown mechanism.

Abbott, Bayer, Bristol-Myers

Squibb; GlaxoSmithKline,

Pharmacia, Wyeth

[49–54,56,57]

� Panels/libraries of whole-cell

target/reporter strains developed

for MOA and screening.

� Hits are often non-sustainable leads.

Structural

genomics

screening

� Provides rational, target-based,

lead-optimization

strategies.

� Crystals can be difficult and

unpredictable to isolate.

� Dual-targeting ParE/GyrB

inhibitors achieving in vivo

efficacy reported.

Abbott, Affinium, Anadys, Aventis,

British Biotech, GlaxoSmithKline,

Merck, Morphochem, Novartis,

PanTherix, Paratek, Pharmacia,

Quorex, RiboTarget, Rib-X, Roche,

Vertex, Vicuron, Wyeth

[8,9,13,14,23–25,28,41,

58,59,61,62,71]

� Advances in HT-technologies

for structure-determination

and molecular-docking.

� Difficult to engineer in

antibacterial activity to

hit molecule.

� HT target-expression, -purification,

-crystallization and compound

co-crystallization programs.

� Might predict biochemical

function of target of unknown function.

� Predicted biochemical function of several

essential proteins of unknown function.

Abbreviations: FabI, enoyl-ACP reductase; FRS, phenylalanyl-tRNA synthetase; GyrB, DNA gyrase subunit B; HT, high-throughput; HTS, high-throughout screen; ParE, topoisomerase IV subunit B; PDF, polypeptide deformylase; MOA, mode-of-

action; MRS, methionyl-tRNA synthetase; SAR, structure–activity relationship.a Survey based on web-sites and publications, includes past and present antibacterial research from 1995 to 2004.b Abbott, http://www.abbott.com/; ActivBiotics, http://www.activbiotics.com/; Affinium, http://www.affiniumpharmaceuticals.com/; Anadys, http://www.anadyspharma.com/; Arrow Therapeutics, http://www.arrowt.co.uk/; AstraZeneca, http://

www.astrazeneca.com/; Aventis, http://www.sanofi-aventis.com/; Basiliea, http://www.basilea.com/; Bayer, http://www.pharma.bayer.com/; Bristol-Myers Squibb, http://www.bms.com/; British Biotech, http://www.vernalis.com/; Cubist, http://

www.cubist.com/; Eli Lilly, http://www.lilly.com/; Genome Therapeutic Corporation, http://www.oscient.com/; GlaxoSmithKline, http://www.gsk.com/; Johnson & Johnson, http://www.jnj.com/; Merck, http://www.merck.com/; Millenium

Pharmaceuticals, http://www.mlnm.com/; Morphochem, http://www.morphochem.com/; Novartis, http://www.novartis.com/; Oscient, http://www.oscient.com/; PanTherix, http://www.pantherix.co.uk/; Paratek, http://www.paratekpharm.com/;

Pfizer, http://www.pfizer.com/; Pharmacia, http://www.pfizer.com/; Proctor & Gamble, http://www.pg.com/; Quorex, http://www.quorex.com/profile/; Replidyne, http://www.replidyne.com/; RiboTarget, http://www.ribotargets.com/; Rib-X,

http://www.rib-x.com/; Roche, http://www.roche.com/; Schering-Plough, http://www.scheringplough.com/; Vertex, http://www.vrtx.com/; Vicuron, http://www.vicuron.com/; Wyeth, http://www.wyeth.com/.

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Links

� Infectious Diseases Society of America (IDSA) 2004 report on ‘Bad

Bugs, No Drugs’, http://www.idsociety.org/

� IDSA/PhRma/FDA Working Group Meeting, 19th November

2002 on drug development for resistant-pathogens sponsored by

the Center for Drug Evaluation and Research, Food and Drug

Administration (FDA), http://www.fda.gov/cder/present/idsaphrma/

default.htm

� The Institute for Genomic Research (TIGR) Pathogen Functional

Genomics Resource sponsored by the National Institute of Allergy

and Infectious Diseases (NIAID), http://pfgrc.tigr.org/

� Genomes Online Database. An updated, comprehensive resource of

completed and ongoing microbial genomes projects, http://www.

genomesonline.org/

� Mycobacterium Tuberculosis Structural Consortium, http://www.doe-

mbi.ucla.edu/TB/

� Structure 2 Function Project is a structural genomics project that

aims to solve structures of poorly characterised Haemophilus influ-

enzae proteins by using X-ray crystallography and protein NMR

techniques, http://s2f.carb.nist.gov/.

� The ExPASy (Expert Protein Analysis System) server of the Swiss

Institute of Bioinformatics dedicated to the analysis of protein

sequences and structures, http://us.expasy.org/

Abbott, crystallization programs have solved the structure

and determined the functions of several essential genes of

unknown function from S. pneumoniae [61]. The approach by

Aventis (http://www.sanofi-aventis.com/) involves the high-

throughput crystallization and X-ray elucidation of over 200

conserved microbial targets of unknown function that so far

has inferred the function of seven of these proteins [62]. In

addition, several consortiums are applying structural geno-

mics to systematically solve the structures of proteins of

uncharacterised function in pathogens H. influenzae and

Mycobacterium tuberculosis [63,64]. Assignments of the bio-

chemical function of these proteins by these approaches

certainly help our understanding of the genome. However,

progression of such targets as well as essential, broad-spec-

trum targets of unknown functions [65], are dependent on

identifying assayable functions that can be employed in HTS,

which adds an additional complexity to exploitation of the

unknowns.

Conclusion

The application of genomics has led to an unprecedented

number of novel, validated targets for screening of com-

pound collections to identify new classes of antibiotics. A

survey of the literature has identified more than 125 pub-

lished studies of antibacterial HTS in the nine years since the

completion of the first bacterial genome. These HTS have

targeted more than 60 different novel protein targets

screened by 34 different companies (Fig. 1). However, tar-

get-based screening has been significantly less successful in

finding novel-acting antibiotics than expected. From our

experience and others, antibacterial targets appear to have

very low hit rate and hit-to-lead success rate which compro-

mise the approach at a very early and fundamental step.

Furthermore, this strategy is frustrated by the fact that fre-

quently hits lack whole-cell antibacterial activity and no

rational SAR processes exist to improve this property. Never-

theless, there are successes such as FabI, FRS and MRS inhi-

bitors achieving promising in vivo efficacy [16–18]. However,

MRS illustrates one of the limitations of the novel target-

based strategy with the presence of a second MRS severely

compromising the broader clinical utility of these leads [42].

Target screens for PDF inhibitors by Vircuron/Novartis and

Oscient have also led to promising lead molecules, two

progressing to Phase I trials [20,21]. In contrast, there are

many more examples of enzyme-based screens of antibacter-

ial targets in the industry that have failed to deliver high-

quality antibacterial leads.

Antibacterial whole-cell screening for novel antibiotics has

the major benefit that antibacterial activity has already been

incorporated into the hit molecule. Genomics has facilitated

this approach in providing new technologies for MOA hunt-

ing studies. However, despite the availability of these power-

ful tools such studies are challenging and rarely definitive.

However, there are at least four literature examples of leads

from this approach [50,51,56,57].

Structural genomics plays a significant role in facilitating

the other antibacterial discovery strategies and there is a

promising report of a novel, dual-target inhibitor to GyrB

and ParE, predominately derived from a structural genomics

strategy that achieve broad-spectrum and in vivo activities

[59].

At present, the investment in genomics for antibacterial

research has resulted in no new genomics-derived drugs on

the market from either target-based or whole-cell screens.

However, genomics has provided numerous new antibacter-

ial target strategies and provided tools to track and identify

the MOA of antibacterial hits. One of the reasons for the low

success rate of antibacterial target screening probably lies

with the CHEMICAL DIVERSITY (see Glossary) of the compounds

being screened against these targets. There is some evidence

that chemical diversity that favors antibiotic screening can be

unique and different from the ‘‘drug-like’’ diversity defined

by Lipinski [66] that is more suited to other therapeutic areas

[4]. Consequently, one approach to increase success at anti-

bacterial HTS would be to generate new ‘‘antibacterial tar-

geted’’ diversity and instigate broader screening of natural

products.

Many companies have invested in genomics-based anti-

bacterial discovery strategies to find new classes of antibio-

tics. However, the output from this investment in terms of

new antibacterial agents in development has been disap-

pointingly low. This does not appear to be through a lack

of trying but more of a function of the unexpected complex-

ities and challenges of delivering new antibiotics targeting

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Drug Discovery Today: Therapeutic Strategies | Infectious diseases Vol. 1, No. 4 2004

novel targets. This factor might have played a contributing

role to the withdrawal of some companies from antibacterial

research just when the clinical need for new antibiotics is

beginning to peak. Although the tools of antibacterial

research have now reached a level of sophistication unim-

aginable several years ago, it is clear that new research stra-

tegies and continued investment to overcome the challenges

in antibacterial research are required.

Outstanding issues

� When will the antibacterial development pipeline give us confidence

that we have the drugs to effectively treat bacterial infections in the

10–20 year time frame?

� How can the success rate of target- and whole-cell antibacterial

screening approaches be increased?

� Do the chemical libraries assayed in HTS by pharmaceutical compa-

nies contain adequate diversity to include compounds with the

complex requirements of an antibacterial agent? If not, can effective

‘‘antibacterial-targeted’’ diversity be created?

� Will compound penetration into bacteria ever be understood to the

extent that rational SAR processes will exist to improve antibacterial

activity?

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