The completion of the first draft of the human genome has provided an
unprecedented opportunity to understand the genetic and molecular basis of
disease. Parallel developments of new biological technologies, such as transcript
profiling, allow scientists to examine almost any biological system in high
molecular resolution. Contemporary drug discovery research is now focusing
on the identification and validation of pharmaceutical targets in the molecular
pathways/systems embedded in this information. Novel therapeutic interventions
are being developed and evaluated as a result of this research which will
be the basis of innovative pharmaceuticals of the future.
Despite many advances in medicine, disease burdens remain significant
in both developed and emerging countries.1
Effective drugs for treatment and prevention are needed for many disease areas,
including cardiovascular disease, cancer, neurological disorders, infectious
diseases, endocrinology, and inflammatory and chronic degenerative diseases.
Therefore, there is excitement about the potential biological revolution that
will emerge with understanding the human genome.2
The various genome initiatives have provided drafts of the chromosomal
sequences of humans and other species. The enabling technologies for this
accomplishment, such as transcript profiling, now provide new tools to examine
complex biological systems at the level of essentially all expressed messenger
RNA (mRNA) and corresponding proteins.
This explosion of biologic information about the proteins and pathways
relevant to cellular physiology and disease has stimulated biotechnology and
pharmaceutical researchers to assign top priority to identification and validation
of key targets (known or novel) to develop therapies for the many remaining
diseases. Hypothesis-based biological research is now supplemented with multidisciplinary
approaches to systems and circuit-based biology that integrate bioinformatics,
genomic databases, and cellular and molecular biology with the traditional
drug discovery disciplines of physiological biochemistry, pharmacology, and
medicinal chemistry. As a consequence, interventional strategies now include
recombinant proteins, monoclonal antibodies, peptides, and small organic molecules
as drug candidates. The goal is to expedite the testing of novel therapeutic
hypotheses in humans and to develop strategies to identify optimal therapy
for individual patients.
Perspectives on Pharmaceutical Research
Successful drug treatments of today and in the past involve fewer than
500 targets or growth factors as of 1996,3
whereas the human genome contains 35 000 to 120 000 genes.4,5 At least 5000 of these genes should
be important targets or produce therapeutic proteins, suggesting that only
10% of potential therapeutic strategies have been identified and exploited
to date.6 This avalanche of genetic information,
largely coming from genomic sequences and expressed sequence tag–based
sequencing of complementary DNA (cDNA) libraries, initially represents a complete
catalogue of component parts of the cell and tissues but does not yet provide
insight into how all of the protein products of these genes interact or function
within the cell.
The challenge for pharmaceutical research is to unravel the pathophysiology
of human diseases and thus, make it possible to identify targets accessible
to drug intervention. The new systems or circuit view of biology that has
evolved from gene research must be considered in successful drug discovery.7,8 This perspective requires integration
of various new technologies into the traditional toolbox of pharmaceutical
research. This article focuses on this new genomically influenced multidisciplinary
approach to contemporary drug discovery by examining genomic information for
hypothesis and target generation and the technologies being developed to validate
targets for new therapies.
Target Identification and Hypothesis Generation
Biochemistry, pharmacology, and medicinal chemistry will continue to
play an essential role in identification of pharmacological targets in the
posthuman genome era. Expertise in preclinical models and human biology is
necessary to incorporate genomic information into a molecular systems approach
to physiology. The traditional drug discovery process will now be supplemented
with additional sources of genomic information at the level of chromosomal
DNA, disease gene associations, mRNA transcript profiling of tissues, human
genetic variance data, and animal and developmental models relevant to disease
For example, if the disease target involves the human endothelium, genomic
information on the target organ system can be mined with bioinformatics for
the discovery process.9,10 DNA
sequencing of a cDNA library of human endothelial cells or the use of transcriptional
arrays can provide a view of gene transcription in normal human endothelium.11,12 Likewise, proteomics (or the systematic
study of expressed proteins) can provide insight into the functional proteins
in specific cells.13,14
Additional uses of these technologies include the ability to compare
normal and disease paradigms of human endothelium by forward and backward
cDNA library subtraction technologies and differential display techniques
to concentrate on transcripts associated with the disease hypothesis.15,16 These approaches will provide a high
resolution view of the endothelial cell system with thousands of components
catalogued as mRNAs of the corresponding proteins.
In the future, many of these component parts will still be proteins
of unknown functions. Identification of the functions of these proteins will
be part of biological research for years to come. The discovery of new pathways
and new molecular interactions could form the basis of new pharmacological
strategies. Protein-protein interaction maps are being constructed using strategies
such as the yeast 2-hybrid system, which eventually will provide detailed
insight into the pathways and networks operational in biological systems.17 Until all these interactions and pathways are mapped
out, a pragmatic approach is to view the data in the context of drug target
A biological target is pharmacologically accessible (or "druggable")
when an organic molecule (peptide, protein, or monoclonal antibody) can modulate
the target's function. The predominant target of currently available drugs
is the G protein coupled–receptor family of molecules.18
Other cell surface receptors and molecules, proteases, protein kinases, and
phosphatases are also "druggable."19-21
In addition, secreted hormones, growth factors, chemokines, soluble receptors,
and decoys may serve either as a drug substance or as targets for other biological
The sequence databases described for the example of human endothelial
cells could be scanned with sequence searching algorithms programmed to recognize
molecular signatures of these druggable families. This "filtered" genomic
information from bioinformatics may identify pharmacologically accessible
targets in the absence of clues to their biological relevance to the disease.
Target identification could be routine with these technologies, whereas, at
present, selection of the correct target is the key strategic challenge.
Target Validation and Pharmaceutical Intervention
Ultimate biological validation of a pharmaceutical target comes with
a successful phase 3 clinical trial and broad application of the therapy to
large populations of patients. Intermediate stages of target validation are
summarized in Figure 2. The initial
stages involve relating the target hypothesis to the disease under study.
The hypothesis may originate from a relevant animal model, such as the discovery
that the molecular defect in ob/ob mice involved the hormone leptin.25 This observation was linked to human obesity by identification
of the human homolog and study of its expression in human obesity.
Mining druggable genomic targets as transcripts from various tissue
libraries, prompts additional questions of whether the protein is expressed
in vivo and whether the target protein is expressed in various physiological
states. Human and animal tissue banks are an invaluable resource for genomic
target validation. In situ and immunohistology studies make it possible to
correlate mRNA and target protein levels with genetics, transcript profiling,
and proteomic or biochemical pharmacology studies. Such analyses of target
expression in all human tissues is critical to understanding the potential
toxicology of the drug strategy under development.
Molecular and cellular biology strategies are frequently used to ablate,
overexpress, or modulate the expression of an in vitro or in vivo target.
Such studies can help provide evidence for the function of the target as well
as models for drug screening and further pharmacological research when drug
candidates have been identified.26,27
Species such as Drosophila melanogaster and Caenorhabditis elegans are assuming increasing roles for
At this point or earlier in the process, drug discovery often involves
developing screening strategies to discover drug candidates that modulate
the targets to test the therapeutic hypotheses. These agents essentially become
validation tools in the process. Various analytical technologies have been
used to develop highly leads for potential targets.29,30
The types of drug candidates screened are originally derived from synthetic
organic chemistry, combinatorial chemistry, which reflects parallel synthesis
on chemical templates; natural product chemistry; and use of chemical libraries.31-33 Additional technologies
include development of monoclonal antibodies for the target (restricted to
cell surface or soluble extracellular molecules), which may act as antagonists,
agonists, or other modulating functions of the proposed target.34
Candidate drugs can then be evaluated in preclinical pharmacological investigations.
For example, in vivo models can be tested with the drug candidate to evaluate
efficacy. If the results are promising, further optimization and evaluations
of potency, efficacy, bioavailability, and toxicology can be conducted prior
to testing in humans.
In the interim, the strategies used to find the target initially, such
as transcript profiling, can be used to examine the effects of the drug in
the biological system under study. An example of this approach involves recombinant
human activated protein C(rhaPC), an anticoagulant protein that functions
as a serine protease to regulate the activity of factors V and VII in the
coagulation cascade (Figure 3).35 Preclinical studies in models of sepsis suggested
that rhaPC also has anti-inflammatory properties. A proinflammatory response
was induced in human endothelium by exposing human umbilical cord endothelial
cells to tumor necrosis factor, and rhaPC downregulated expression of several
proinflammatory genes and up-regulated antiapoptotic transcripts. The effects
are potentially relevant to the utility of the drug in acute inflammatory
disease (D. Joyce, MD, unpublished data, 2001). While these studies were performed
after preclinical pharmacology, novel pharmacological actions of rhaPC were
uncovered. These strategies appear broadly applicable to investigation of
disease and drug mechanisms, structure activity relationships, and toxicogenomics,
and they should enhance the potential success of drug candidates in the future.
In addition, the use of singular transcript profiling strategies in preclinical
models may identify compensatory systems or circuits in biology that can be
targets of a future pharmacological intervention.
Future Prospects for Novel Therapeutics
These strategies will continue to be enhanced with the development of
new biotechnologies and knowledge about the biological circuits of life. Although
new validated targets will emerge in unprecedented numbers, several key questions
also arise. For instance, which targets and interventions should receive high
priority for clinical investigation? What drug modality can be used to rapidly
evaluate the hypothesis in humans? When will the first new therapies from
these new strategies emerge?
It is likely that the initial impact of genetic and genomic technologies
for new therapies will involve therapeutic proteins and monoclonal antibodies.36-38 Numerous examples
exist of novel therapeutic protein candidates derived from genomic databases
and some of these agents are currently in clinical development.36-38
Innovative developments in monoclonal antibody technology such as engineered
mice with human immunoglobulin repertoires have also provided researchers
with the tools to rapidly identify and evaluate human monoclonal antibodies
as drug candidates. Taken together, therapeutic proteins and monoclonal antibodies
represent one of the largest classes of drugs in development, with estimates
of more than 350 molecules under investigation in early 2000.39
Most of these agents are directed toward cancer or inflammation targets for
which biological drugs continue to provide significant innovation.40,41 These approaches accelerate human
clinical trials, but some diseases are not amenable to protein therapies because
of lack of accessibility of the target (eg, intracellular enzyme) or because
of physiological constraints (eg, blood brain barrier).
However, a parallel explosion of many drug candidates of small molecular
weight will occur in the next decade as research intensifies into the regulatory
circuits of disease coupled with parallel drug discovery on every new validated
target. It is likely that polygenic diseases will provide multiple targets
for therapy, each of which will need clinical investigation. An example of
a disease platform for which genetic and genomic technologies have had an
impact is in obesity research, for which several intervention options have
been identified recently.42 An alternate strategy
may involve the in-depth understanding of a family of potential targets that
are implicated in key biological activities. For instance, this strategy is
apparent in developments in the pharmacology of excitatory amino acid receptors,
which have potential therapeutic applications in several diseases.43
The chemical "hit/lead" database will be combined with bioinformatics
and structural genomics to expedite the development of new leads in the future.44,45 It seems inevitable that these various
therapeutic intervention strategies would identify targets that today are
not presently accessible to pharmacology. Gene therapy may also provide alternative
pharmaceutical strategies as may cellular therapies, particularly with human
embryonic and adult stem cells.46,47
The genetic and biological revolution undoubtedly will change clinical
trials and clinical practice in the future. The effect of human genetic variance
on responses to therapy will influence drug-development clinical trials and
the use of products in clinical practice.48,49
The "resequencing" of the human genome will establish the frequency
of single nucleotide polymorphisms in the genome and other types of genetic
variance. DNA, which is now routinely obtained in many clinical trials, will
be examined retrospectively for the association of genetic variance or mutations
with outcomes. Smaller clinical studies prospectively biased toward the target,
the drug, and the appropriate patient will be designed. These approaches and
the accompanying data could make it possible to devise optimal strategies
for future therapies. Biomarker/surrogate marker panels for human diseases
will provide additional strategies to diagnose, monitor, and predict outcomes
of various therapies.
The potential of genetics, genomic information, and genomic technologies
for new treatments will only be realized if these new disciplines are integrated
into the drug discovery process at every stage—from hypothesis generation
to clinical evaluation. Translation of biological information to disease knowledge,
validated target mechanisms, and new therapies will indeed make the coming
century an era of biomedical revolution.
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