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EDITORIALEvolution of Anticancer Drug Discovery and the Role of

Cell-Based Screening

Frank M. Balis

The approach to the discovery of new anticancer drugs has

recently evolved from a reliance on empiric cell-based screening

for antiproliferative effects to a more mechanistically based ap-

proach that targets the specific molecular lesions thought to be

responsible for the development and maintenance of the malig-

nant phenotype in various forms of cancer. The ultimate goal of

the development of molecularly targeted drugs is to improve the

efficacy and selectivity of cancer treatment by exploiting the

differences between cancer cells and normal cells. The success

of recently developed molecularly targeted agents, such as treti-

noin (all-trans-retinoic acid) for acute promyelocytic leukemia

(1,2) and imatinib (STI-571) for chronic myelogenous leukemia

(CML) (3,4) and gastrointestinal stromal tumors (5), providesearly clinical validation for the molecularly targeted approach to

drug discovery.

Most of the commonly used cytotoxic anticancer drugs were

discovered through random high-throughput screening of syn-

thetic compounds and natural products in cell-based cytotoxicity

assays. Despite the number and chemical diversity of these

agents, the mechanisms of action are limited (Table 1), and most

compounds are DNA-damaging agents with a low therapeutic

index. With this screening approach, mechanism of action is not

a primary determinant in selecting agents for further develop-

ment, and, as a result, none of the current drugs directly targets

the molecular lesions responsible for malignant transformation.

The initial National Cancer Institute (NCI) high-throughputscreen used the highly chemosensitive P388 leukemia cell line,

but this screen failed to identify drugs that were active against

the common adult solid tumors. In the mid-1980s, the NCI

implemented a new in vitro disease-oriented screen consisting

of 60 human tumor cell lines representing nine common forms

of cancer (6). It remains to be determined whether selective

activity in vitro against cell lines representing a particular his-

tologic form of cancer will be predictive for antitumor activity

in vivo (7).

As the molecular bases of specific forms of cancer are elu-

cidated, a variety of new potential therapeutic targets are being

identified. In the most common forms of cancers, the accumu-

lation of mutations in multiple genes is required for tumorigen-esis (8,9). These mutations, such as the ras-activating mutations

and mutations that inactivate p53, allow cancer cells to circum-

vent intrinsic and extrinsic controls that tightly regulate the cell

cycle and cell division and apoptosis in normal cells. After iden-

tification of the genetic alterations in cancer cells, the critical

transforming mutations must be discerned from genetic alter-

ations that are the result of the inherent genetic instability in

cancer cells but that do not play a direct role in tumorigenesis

(9). This target validation is performed in preclinical cancer

models. Identification of valid molecular targets has led to ra-

tional target-based drug discovery at the protein level, as illus-

trated by the development of imatinib, which was discovered by

screening compound libraries for inhibitors of the protein kinase

activity in vitro (10). Many of the proteins involved in cell cycle

regulation, signal transduction, and the regulation of apoptosis

are enzymes or receptors and are, therefore, potentially ame-

nable to inhibition by small molecules (1114). Other examples

of target-based treatments undergoing clinical evaluation in-

clude farnesyltranferase inhibitors, which block post-transla-

tional prenylation of ras, cyclin-dependent kinase inhibitors,

protein kinase C inhibitors, and epidermal growth factor receptor

kinase inhibitors (15).

However, target-based screening for discovery of new mo-

lecularly targeted cancer treatments has shortcomings. Unlike

the Bcr-Abl fusion protein in CML and ras oncogenes that rep-resent mutations leading to a gain in function, most mutations in

cancer cells result in a loss or inactivation of a protein (e.g., p53

mutations), and screening a group of drugs by use of the protein

product of mutated tumor suppressor genes is unlikely to dis-

cover a small molecule that restores protein function. In addi-

tion, compounds that are active in a target-based screening assay

may not be specific for the protein used in the screen, and, as a

result, the pharmacologic effect of the compound at a cellular or

organism level may be more closely related to its effect on other

unrelated and potentially higher affinity targets. Finally, most

molecular targets for new cancer treatments interact with other

proteins within pathways or networks in the cell, and the phar-

macologic effect resulting from the inhibition of a specific targetmay be influenced by the expression or relative levels of these

interacting proteins. Therefore, target-based screening assays

may not be predictive of drug effect within the context of the

whole cell (16).

As demonstrated by Dunstan et al. (17) in this issue of the

Journal, cell-based screening assays will continue to play an

important role in drug discovery as we move into the molecular-

targeting era. Their three-stage cell-based procedure using yeast

cells that can be genetically manipulated is adaptable to high-

throughput screening to identify agents that have a selective

effect against cells with specific mutations (gene deletions). Un-

like target-based assays, this cell-based assay is not a mechanis-

tic screen, but determining the mechanism of action of selec-tively toxic agents from this screen may identify new molecular

targets (e.g., downstream effectors in a pathway that is dere-

pressed by the deletion of a tumor suppressor gene) for subse-

quent target-based screening. From a pharmacologic perspec-

tive, this cell-based screen detects collateral sensitivity. The

Affiliation of author: Pediatric Oncology Branch, Center for Cancer Research,

National Cancer Institute, Bethesda, MD.

Correspondence to: Frank M. Balis, M.D., National Institutes of Health, Bldg.

10, Rm. 13C103, 10 Center Dr., Bethesda, MD 208921920 (e-mail: balisf@

nih.gov).

78 EDITORIAL Journal of the National Cancer Institute, Vol. 94, No. 2, January 16, 2002

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genetic mutation enhances the cells sensitivity to drugs that

affect related pathways within the cell. The assay is also a direct

application of the concept of synthetic lethality, which has been

described previously in yeast (9,18). Two genes are synthetically

lethal if mutations in either one is survivable, but mutations in

both genes are lethal. For this cell-based assay, disruption of the

function of the genes product by a drug is only lethal in cells

that have a mutation in a second related gene.

Cell-based assays are also used to confirm the activity of

agents discovered in target-based screening assays and to assess

the drugs pharmacologic effects at the cellular level. Unex-pected effects in cellular systems may suggest other targets for

the agent or interactions of the primary molecular target with

other vital proteins within the cell. The NCIs panel of 60 human

tumor cell lines has also been adapted to provide information

about the mechanism of action or molecular target of new agents

that are tested based on the drugs profile of activity in the screen

(19). As new targets are identified, their expression in each of

the 60 cell lines in the panel can be characterized and correlated

with the activity profile of the 70 000 compounds screened pre-

viously without having to retest each agent. The identification of

the topoisomerases as targets for drugs that were selectively

active in yeast cells deficient in DNA double-strand break-repair

proteins was apparently based on their activity profile in the

human tumor cell line panel.

Target-based and cell-based screening for new anticancer

drugs in the molecular targeting era are complementary methods

of identifying more selective anticancer drugs. They represent a

dramatic shift in the drug discovery process that is likely to have

an impact not only on the pharmacologic properties of new

anticancer agents reaching the clinic but also on our approach to

clinical drug development and the treatment of cancer.

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Table 1. Mechanism of action of the commonly used natural product

anticancer drugs*

Drug class Mechanism of action

Anthracyclines Topoisomerase II inhibitorsEpipodophyllotoxins Topoisomerase II inhibitorsDactinomycin Topoisomerase II inhibitorsCamptothecins Topoisomerase I inhibitorsTaxanes Tubulin-binding agentsVinca alkaloids Tubulin-binding agents

*The source and chemical structures of these agents are diverse, but themolecular targets of these agents are limited to the topoisomerases and tubulin.

Journal of the National Cancer Institute, Vol. 94, No. 2, January 16, 2002 EDITORIAL 79