Curr. Med. Chem.

- Anti-Cancer Agents, 2005, 5, 363-372 363

Etoposide, Topoisomerase II and Cancer

E.L. Baldwin1 and N. Osheroff1,2,*

Departments of 1Biochemistry and 2Medicine (Hematology/Oncology), Vanderbilt University School of Medicine,

Nashville TN 37232-0146, USA

Abstract: Etoposide is an important chemotherapeutic agent that is used to treat a wide spectrum of human cancers. It has
been in clinical use for more than two decades and remains one of the most highly prescribed anticancer drugs in the
world. The primary cytotoxic target for etoposide is topoisomerase II. This ubiquitous enzyme regulates DNA under- and
overwinding, and removes knots and tangles from the genome by generating transient double-stranded breaks in the
double helix. Etoposide kills cells by stabilizing a covalent enzyme-cleaved DNA complex (known as the cleavage
complex) that is a transient intermediate in the catalytic cycle of topoisomerase II. The accumulation of cleavage
complexes in treated cells leads to the generation of permanent DNA strand breaks, which trigger recombination/repair
pathways, mutagenesis, and chromosomal translocations. If these breaks overwhelm the cell, they can initiate death
pathways. Thus, etoposide converts topoisomerase II from an essential enzyme to a potent cellular toxin that fragments
the genome. Although the topoisomerase II-DNA cleavage complex is an important target for cancer chemotherapy, there
also is evidence that topoisomerase II-mediated DNA strand breaks induced by etoposide and other agents can trigger
chromosomal translocations that lead to specific types of leukemia. Given the central role of topoisomerase II in both the
cure and initiation of human cancers, it is imperative to further understand the mechanism by which the enzyme cleaves
and rejoins the double helix and the process by which etoposide and other anticancer drugs alter topoisomerase II
function.
Key Words: Etoposide, topoisomerase II, anticancer drugs, DNA cleavage, DNA ligation, DNA cleavage complex, cancer
chemotherapy, leukemia.

A. INTRODUCTION It also will discuss therapy-related leukemias that are

Etoposide is one of the most widely prescribed anticancer
drugs in the world. It is used to treat a variety of cancers,
including small cell lung cancer, germ-line malignancies,
sarcomas, leukemias, and lymphomas. The primary cellular
target for etoposide is topoisomerase II. This essential
enzyme removes knots and tangles from the genome by
introducing transient double-stranded breaks in the DNA
backbone.
Etoposide kills cells in an insidious fashion. Rather than
robbing the cell of the critical functions of topoisomerase II,
the drug stabilizes a covalent enzyme-cleaved DNA complex
(known as the cleavage complex) that is a requisite
intermediate in the catalytic cycle of topoisomerase II. The
accumulation of cleavage complexes in treated cells leads to
the generation of permanent breaks in the genetic material,
which ultimately trigger cell death pathways. Thus, etopo-
side converts this essential enzyme into a potent cellular
toxin that fragments the genome.
This review will focus on etoposide, its interactions with
topoisomerase II, and the cellular pathways that are involved
in processing the DNA breaks that are induced by the drug.
*Address correspondence to this author at the Department of Biochemistry,
654 Robinson Research Building, Vanderbilt University School of
Medicine, Nashville, TN 37232-0146, USA; Tel: (615)322-4338; Fax:
(615)343-1166; E-mail: neil.osheroff@vanderbilt.edu
associated with the use of etoposide.
B. HISTORY OF ETOPOSIDE
Etoposide is derived from podophyllotoxin (Fig. (1)) [1,
2]. This natural product is found in the plant Podophyllum
peltatum, more commonly known as mandrake or May apple.
Podophyllotoxin has been used as a folk remedy for a variety
of illnesses for over a thousand years [2]. It has well estab-
lished antimitotic properties, which are related to its ability
to inhibit tubulin polymerization. Although podophyllotoxin
demonstrated potential as an antineoplastic agent, clinical
development was prevented by its high toxicity [2-4].
In the 1950’s, investigators at Sandoz Pharmaceuticals
initiated a program in the hopes of overcoming the prohibi-
tive toxicity of the parent compound [1, 2]. As a result, a
series of semi-synthetic podophyllotoxin derivatives was
generated. Ultimately, two analogs with increased antineo-
plastic activity and decreased toxicity were identified in the
mid-1960’s, etoposide and teniposide (Fig. (1)) [1, 2].
Following pre-clinical development and a series of clinical
trials, the United States Food and Drug Administration
approved the use of etoposide for cancer chemotherapy in
1983. Remarkably, at the time of this approval, the mech-
anism of action of the drug was unknown. In contrast to
podophyllotoxin, etoposide displayed no ability to inhibit
tubulin polymerization [5]. It was not until the mid-1980’s
that it was determined that the primary cellular target of
etoposide was topoisomerase II [6-8].

1568-0118/05 $50.00+.00 © 2005 Bentham Science Publishers Ltd.

364 Curr. Med. Chem. – Anti-Cancer Agents, 2005, Vol. 5, No. 4 Baldwin and Osheroff

g8
OH g7
H3 C
g6
O O g
4
O
O g5
O
g2
O HO g3
O
O g1
OH
11
O 5 4
6 3
A B C D O12
7 8 2
CH3O OCH3 1 13
O
OCH3 O
1' 6'
2'
3' E
Podophyllotoxin Etoposide 4' 5'
H3CO OCH3
OH

H3 C CH3
H N
O
O
S O
HO
H3 C N
OH
O

O O
O O
O O
O O

CH3O OCH3 CH3O OCH3

OH OH

Teniposide TOP-53

Fig. (1). Structures of podophyllotoxin, etoposide, teniposide and TOP-53. Etoposide can be separated into three distinct structural domains:
a glycosidic group substituted at the C-4 position, a polycyclic core (A-D rings), and a pendant ring (E-ring). Teniposide contains of a
thiophene moiety in place of the methyl group at the g-8 position, whereas TOP-53 contains an amino-alkyl side chain at the C-4 position.

C. TOPOISOMERASE II Topoisomerase IIα is the isoform originally described in

1. Isoforms
Topoisomerase II is a ubiquitous enzyme that regulates
DNA topology by passing an intact double helix through a
transient double-stranded break that it generates in the back-
bone of a separate DNA segment [9-15]. In contrast to lower
eukaryotes such as yeast and Drosophila that encode a single
type II enzyme [16, 17], vertebrates have two isoforms of
topoisomerase II, α and β [18, 19]. These two isoforms share
~70% amino acid sequence identity and display similar
enzymatic activities. However, they are encoded by separate
genes, and differ in their protomer molecular masses (~170
kDa and ~180 kDa, respectively), physiological regulation,
and cellular functions [18-25]. Topoisomerase IIα levels
increase dramatically during periods of cell growth, and this
isoform appears to be primarily responsible for the required
roles of the enzyme during DNA replication and mitosis [26-
31]. Topoisomerase IIα is an essential enzyme [23, 32, 33].
Vertebrate cells can survive in the absence of topoisomerase
IIβ, but this isoform is required for proper neurological
development in mouse embryos [34].
vertebrate species and in general, represents the enzyme that
was characterized as “topoisomerase II” in early studies in
these organisms. Since the enzymological properties of all
eukaryotic type II enzymes appear to be similar and distinc-
tions between isoforms are not always apparent, unless
specifically stated otherwise, the term topoisomerase II will
be used to refer to all members of this enzyme family.
2. Catalytic Cycle
One round of catalysis mediated by topoisomerase II is
depicted in Fig. (2) (see Refs. [10-13, 15] for comprehensive
reviews of the topoisomerase II catalytic cycle). A brief
description follows: 1) Topoisomerase II binds two segments
of DNA to initiate a round of catalysis [35, 36]. 2) In the
presence of a divalent cation [37], the two subunits of the
enzyme each cleave one strand of the double helix [38-40].
The two scissile bonds are located four base pairs apart
directly across the major groove from one another. Thus,
cleavage generates DNA molecules with 4-base single-
stranded cohesive ends on their 5'-termini [38, 39]. It is
Etoposide, Topoisomerase II and Cancer
Fig. (2). Catalytic cycle of topoisomerase II. One round of catalysis is shown as a series of six discrete steps. 1) Topoisomerase II-DNA
binding; 2) pre-strand passage DNA cleavage/religation equilibrium; 3) ATP binding and DNA strand passage; 4) post-strand passage DNA
cleavage-religation equilibrium; 5) ATP hydrolysis and gate opening; 6) DNA release and enzyme turnover. Adapted from [13].
notable that this reaction is reversible and that the enzyme
establishes a DNA cleavage-religation equilibrium. 3)
Topoisomerase II binds two ATP molecules and undergoes a
conformational change that causes the passage of the intact
DNA segment through the cleaved double helix [41, 42].
DNA strand passage appears to be faster if one of the two
ATP molecules is hydrolyzed [43]. The transported DNA is
passed into a central cavity in topoisomerase II through an
N-terminal gate [44, 45]. The gate is closed and the enzyme
forms a protein clamp on the DNA [44, 45]. 4) Following
DNA translocation, topoisomerase II reseals the original
nucleic acid break, and the enzyme once again establishes a
DNA cleavage-religation equilibrium [44]. 5) Topoiso-
merase II hydrolyzes the second ATP molecule, which opens
a C-terminal gate on the protein and allows the transported
DNA segment to be released [41, 44, 45]. 6) Finally, the
enzyme returns to its original conformation and regains the
ability to begin a new round of catalysis [41, 44].
As a result of its catalytic mechanism, topoisomerase II
can relax under- or overwound (i.e., negatively or positively
supercoiled) DNA molecules and can remove knots and
tangles from the genome [10-13, 15]. It is these latter func-
tions that make topoisomerase II essential for cell survival.
In the absence of the type II enzyme, cells cannot segregate
daughter chromosomes and die as a result of mitotic failure.
3. Topoisomerase II-DNA Cleavage Complexes
In order to maintain the integrity of the genetic material
throughout the double-stranded DNA passage reaction,
topoisomerase II forms covalent bonds between active site
tyrosyl residues (one per each subunit of the homodimeric
enzyme) and the newly generated termini of the cleaved
DNA [38-40, 46-48]. This covalent enzyme-cleaved DNA
complex, which is referred to as the cleavage complex, is a
hallmark of all type II enzymes. Beyond the essential role of
the cleavage complex in the catalytic cycle of topoisomerase
Curr. Med. Chem. – Anti-Cancer Agents, 2005, Vol. 5, No. 4 365
II, it is the primary link between the enzyme and cancer. The
topoisomerase II-DNA cleavage complex is the cytotoxic
target for etoposide and other anticancer drugs and also
appears to play a central role in the initiation of leukemic
chromosomal translocations.
Normally, cleavage complexes are fleeting intermediates
in the catalytic cycle of topoisomerase II. Consequently, the
enzyme-linked DNA breaks are present at low steady-state
levels and are tolerated by cells [10, 13, 15, 49]. However,
increases in the physiological concentration or lifetime of
cleavage complexes can initiate a number of mutagenic
events (Fig. (3)) [10, 13, 15, 50-53]. For example, when
DNA tracking systems such as replication forks collide with
a topoisomerase II-DNA cleavage complex, the transient
enzyme-associated DNA break is converted to a permanent
double-stranded break in the genome [54-57]. Once the
DNA termini are no longer bridged by topoisomerase II, they
become targets for recombination and repair pathways.
Depending on the pathway employed, repair of topoisomer-
ase II-mediated DNA strand breaks can generate chromo-
somal insertions, deletions, translocations, or other aberra-
tions [10, 50, 52, 58]. If these strand breaks are present in
sufficient numbers, they initiate programmed cell death
pathways (Fig. (3)) [51].
Agents such as etoposide that increase the concentration
of cleavage complexes are known as topoisomerase II
poisons because they convert this essential enzyme to a
potent cellular toxin [59]. This is in contrast to catalytic
inhibitors, which block the overall catalytic activity of the
enzyme. Due to the mechanism of action of topoisomerase II
poisons, the higher the cellular concentration of the type II
enzyme, the more potent these drugs become [10, 13, 15,
60]. The high levels of topoisomerase II often expressed in
cancer cells provide part of the therapeutic window for
etoposide and other chemotherapeutic agents that target this
enzyme.
366 Curr. Med. Chem. – Anti-Cancer Agents, 2005, Vol. 5, No. 4
Fig. (3). Effects of topoisomerase II-DNA cleavage complexes in the cell. Topoisomerase II-DNA cleavage complexes normally are
transient intermediates in the catalytic cycle of the enzyme and are present at low cellular concentrations. However, when levels of cleavage
complexes rise, they can be converted to permanent DNA strand breaks that trigger DNA recombination/repair pathways or cell death
pathways. Alternatively, chromosomal translocations can be generated that lead to specific types of leukemia.
D. ACTIONS OF ETOPOSIDE AGAINST TOPOISO-
MERASE II
1. Cellular Target
As discussed earlier, etoposide kills cells by increasing
levels of topoisomerase II-DNA cleavage complexes [2, 10,
13, 15, 60, 61]. This drug was among the earliest anticancer
agents that were identified as targeting the type II enzyme.
Several lines of evidence led to this identification.
By the early 1980’s, it was well established that treat-
ment of mammalian cells with etoposide induced the forma-
tion of protein-associated single-and double-stranded DNA
breaks [62-64]. It was not until the mid-1980’s, however,
that the role of topoisomerase II in mediating the toxicity of
the drug became apparent. The first direct evidence that the
enzyme was the target of etoposide came from in vitro
studies in the Liu laboratory [6, 7]. Inclusion of etoposide in
topoisomerase II-DNA cleavage assays significantly raised
levels of enzyme-mediated strand breaks. Subsequent studies
demonstrated that DNA breaks generated following treat-
ment of cells with the drug were attached to topoisomerase
II. The initial work utilized SV40 as a model system [8], but
later experiments extended this result to chromosomal DNA
[65-67].
Mutagenesis studies also played an important role in the
identification of topoisomerase II as the primary cellular
target of etoposide. Mammalian cell lines selected for
resistance to etoposide or teniposide were demonstrated to
express a type II enzyme that displayed resistance to these
drugs in vitro [68-71].
Baldwin and Osheroff
The final conclusive evidence that etoposide killed cells
by acting as a topoisomerase II poison came from studies
that used the budding yeast, Saccharomyces cerevisiae. This
organism allowed, for the first time, the cellular effects of
etoposide to be studied in a series of isogenic strains. The
initial work of Nitiss and colleagues determined that overex-
pression of topoisomerase II rendered yeast cells hypersen-
sitive to etoposide [72]. Subsequent experiments demon-
strated that yeast expressing a temperature-sensitive mutant
of the enzyme were refractory to drug treatment at semiper-
missive temperatures (at which topoisomerase II activity is
reduced to less than 10% that of the wild-type enzyme) [73].
These two studies were seminal to our understanding of
etoposide action. Together they provided the first direct
evidence that topoisomerase II is the cytotoxic target of
etoposide and that the drug kills cells by converting the
enzyme into a physiological toxin. As further evidence that
topoisomerase II is the primary cellular target of etoposide,
yeast strains that expressed mutant type II enzymes that were
either resistant or hypersensitive to etoposide in vitro
displayed the corresponding drug phenotype in vivo [74-78].
The relative contributions of topoisomerase IIα and β to
the actions of etoposide in mammalian cells are still under
debate. The drug has similar effects on DNA cleavage
mediated by the two isoforms in vitro [19, 67, 79] and
appears to target both enzymes in cultured human cells [65,
67]. Thus, it is likely that topoisomerase IIα and β both play
a role in mediating the cellular efficacy of etoposide, and that
the relative contributions of the two isoforms are dependent
on the tumor and the individual patient.
Etoposide, Topoisomerase II and Cancer
2. Topoisomerase II-DNA-Etoposide Ternary Complex
Formation
The topoisomerase II-DNA-etoposide ternary complex
can form by one of three different pathways: etoposide can
enter the complex through interactions with the enzyme,
with DNA, or with the binary topoisomerase II-DNA com-
plex [10, 13, 80]. Originally, it was believed that anticancer
drugs entered the complex through interactions with either
the DNA or the enzyme-DNA binary complex. However, all
currently available evidence suggests that etoposide enters
the complex through interactions with topoisomerase II.
First, in contrast to most other topoisomerase II poisons,
etoposide displays little ability to bind free DNA [81].
Second, the drug binds to yeast topoisomerase II and human
topoisomerase IIα in a stoichiometric fashion in the absence
of DNA [82, 83]. Furthermore, the concentrations of etopo-
side required for binding approximate the levels required to
increase levels of topoisomerase II-DNA cleavage com-
plexes [82, 83]. Third, the binding affinity of etoposide for a
mutant drug-resistant enzyme is decreased as compared to
wild-type topoisomerase II [77, 82]. Finally, rates of topo-
isomerase II-mediated DNA cleavage are higher when etopo-
side is incubated with the enzyme prior to the addition of
DNA, as compared to those generated when the drug is incu-
bated with the DNA prior to the addition of topoisomerase II
[84].
The above notwithstanding, it is likely that etoposide
contacts DNA in the ternary complex. For example, the drug
alters DNA cleavage site utilization by topoisomerase II and
preferentially induces scission at sites that have a cytosine
one base 5’ to the scissile bonds [85-89].
3. Mechanism of Action
Topoisomerase II poisons increase levels of cleavage
complexes by two non-mutually exclusive mechanisms [10,
13, 15]. Some drugs act by inhibiting the ability of topoiso-
merase II to ligate cleaved DNA. Agents in this class not
only raise the concentration of cleavage complexes, but
increase their lifetime as well. Other drugs have little effect
on the ligation activity of topoisomerase II and are presumed
to act primarily by increasing the forward rate of enzyme-
catalyzed DNA cleavage.
On the basis of assays that monitor the ability of topoiso-
merase II to religate DNA that it has cleaved or to ligate two
separate fragments of DNA, etoposide acts primarily by
inhibiting the ligation activity of the type II enzyme [90-92].
Although the mechanism of drug action against topoiso-
merase II had long been debated, etoposide was the first
compound to be analyzed experimentally [90].
Unfortunately, none of the available crystal structures for
the DNA cleavage-ligation domain of topoisomerase II
include either DNA or anticancer drugs [93, 94]. Therefore,
it is not known precisely how etoposide impairs the ability of
topoisomerase II to seal DNA strand breaks. The drug may
interfere with noncovalent enzyme-DNA interactions. Alter-
natively, it may alter the position of the DNA termini to be
ligated or insert between the DNA termini and act as a
physical block to ligation [15, 95].
Curr. Med. Chem. – Anti-Cancer Agents, 2005, Vol. 5, No. 4 367
Irrespective of the precise mechanism, evidence suggests
that etoposide acts in close proximity to the scissile bonds
[85-88, 92]. Furthermore, it appears that the interaction of
etoposide with one scissile bond is independent from its
interaction with the other, and that each drug molecule
stabilizes only a single-stranded break [92]. Thus, two drug
molecules are required in order to increase levels of topo-
isomerase II-mediated double-stranded DNA breaks [92].
This finding explains the fact that etoposide produces high
levels of enzyme-associated single-stranded DNA breaks at
physiologically relevant drug concentrations [62-64, 92]. In
addition, it implies that the cytotoxic effects of etoposide
may be more closely associated with the generation of topo-
isomerase II-mediated single-, rather than double-stranded
DNA breaks. These single-stranded DNA breaks subse-
quently are converted to permanent double-stranded DNA
breaks by the actions of replication forks. Consistent with
this suggestion, etoposide activates the ATR- rather than
ATM-dependent DNA damage checkpoint in Xenopus egg
extracts which were not undergoing active DNA replication
[96].
4. Structure-Activity Relationships
Etoposide can be separated into three distinct structural
domains: a glycosidic group (at the C-4 position), a
polycyclic core (A-D rings), and a pendant ring (E-ring) (see
Fig. (1)) [97]. The glycosidic group contributes little to the
activity of the drug against topoisomerase II [98]. However,
it plays important roles in the physiology of the compound.
If the glycosidic group is removed, the remaining demethyl-
epipodophyllotoxin displays strong interactions with tubulin
[1, 5, 99]. Furthermore, exchanging the g-8 methyl group of
etoposide for a thiophene moiety in teniposide increases the
physiological potency of the drug by ~10–fold [1, 100-102].
If the glycosidic group at the C-4 position is replaced by an
amino-alkyl side chain, such as that found in TOP-53 (Fig.
(1)), activity against topoisomerase II rises significantly [67].
Other modifications at the C-4 position also alter the activity
of the drug against topoisomerase II both in cells and in vitro
[103-105]. Thus, while the C-4 glycosidic group in etoposide
does not appear to promote drug interactions with topoiso-
merase II, modifications at this position can significantly
affect drug activity.
The polycyclic core of etoposide plays an important role
in promoting drug-enzyme interactions [98]. There is little
evidence that etoposide binds free nucleic acids [81].
However, it is possible that portions of the polycyclic core
interact with DNA in the active site of topoisomerase II [15,
95].
Substituents on the pendant E-ring appear to be essential
for the activity of etoposide. The 4’-OH group is very
important for increasing levels of topoisomerase II-DNA
cleavage complexes. If this group is removed or converted to
a methoxy group, the activity of etoposide against topoiso-
merase II is reduced significantly [62, 98-100, 106]. Removal
of both methoxy groups (at the 3’- and 5’-positions) decre-
ases drug activity, however, the loss of only one of these
groups appears to have little effect [98, 107]. In addition,
replacement of the 3’- and 5’-methoxy groups by hydroxyl
groups increases drug activity [102].
368 Curr. Med. Chem. – Anti-Cancer Agents, 2005, Vol. 5, No. 4
Finally, one of the methoxy groups on the E-ring can be
converted to a hydroxyl group in vivo by the actions of
cytochrome P450 3A4 (CYP 3A4) [108-111]. The resulting
etoposide catechol, which has been found in patients that
were treated with etoposide [112, 113], can be further
oxidized to form etoposide quinone [108-111]. Both meta-
bolites display higher activity against topoisomerase II than
the parent compound [89].
E. CELLULAR REPAIR OF ETOPOSIDE-INDUCED
DNA DAMAGE
Double-stranded breaks in the genetic material are
repaired primarily by DNA recombination pathways. S.
cerevisiae has been used as a model system to investigate the
pathways by which etoposide-induced DNA damage is
repaired. The most common pathways used by budding yeast
are depicted in Fig. (4) [114-116]. The initial processing of
double-stranded DNA breaks utilizes the Rad50p/Mre11p/
Xrs2p complex to generate single-stranded ends at the site of
the break [114-117]. Following this processing, the DNA can
be shuttled into three distinct recombination pathways. The
break can be repaired by the single-strand invasion pathway
of homologous recombination [114-116]. This pathway,
which utilizes Rad51p/52p/54p/55p/57p as well as the
replication machinery, is capable of repairing the initial
double-stranded DNA break in an error-free manner. Alter-
natively, the break can be repaired by the single-strand
annealing pathway of homologous recombination [114-116].
Fig. (4). Pathways that repair double-stranded DNA breaks in S. cerevisiae. Components of the pathways that play integral roles in
homologous recombination (single-strand invasion or single-strand annealing) and nonhomologous end-joining are shown. Adapted from
[120].
Baldwin and Osheroff
This pathway is dependent on the presence of direct repeats
(or closely related sequences) proximal to and flanking the
initial break site. It relies on Rad52p and the Rad1p/Rad10p
endonuclease. Single-strand annealing is not an error-free
pathway and deletes one of the repeated sequences, as well
as the genetic information that is located between them.
Finally, the break can be rejoined by the nonhomologous
end-joining pathway [114, 115, 118, 119]. This pathway
utilizes Ku70p/Ku80p and Lig4p and results in the loss of
sequences proximal to the original DNA break. If multiple
breaks are present in the genome, nonhomologous end-join-
ing can lead to the formation of chromosomal rearrange-
ments or translocations. In general, homologous recombina-
tion pathways are considerably more active than nonho-
mologous end-joining in S. cerevisiae [114-119].
Two lines of evidence strongly suggest that the single-
strand invasion pathway of homologous recombination is the
primary pathway used to repair topoisomerase II-associated
DNA breaks that are induced by etoposide. First, deletion of
RAD54 significantly decreases cell survival and the induc-
tion of recombination/repair in the presence of etoposide. In
contrast, deletion of RAD1 or KU70 does not sensitize yeast
cells to the drug [120]. Second, a genome-wide screen for
hypersensitivity to etoposide identified every member of the
single-strand invasion pathway, but did not identify genes
that were exclusive to either the single-strand annealing
pathway or nonhomologous end-joining [121]. In addition to
the single-strand invasion pathway, recent evidence suggests
Etoposide, Topoisomerase II and Cancer
an important role for MMS22 in the repair of etoposide-
induced DNA damage in yeast [121]. This gene is not part of
the single-strand invasion pathway and is thought to be
involved in the resolution of aberrant DNA structures that
accumulate at replication forks in response to genomic
damage [121, 122].
Studies using the fission yeast, Schizosaccharomyces
pombe, also suggest that homologous recombination path-
ways play a major role in repairing topoisomerase II-
mediated DNA breaks that are induced by the etoposide
derivative TOP-53 [123]. Furthermore, they implicate
nucleotide excision repair proteins in the processing of these
DNA breaks [123].
Nonhomologous end-joining appears to play a more
significant role in the repair of double-stranded DNA breaks
in mammalian species [124]. A number of studies indicate
that homologous recombination and nonhomologous end-
joining pathways both are involved in the repair of topoiso-
merase II-mediated DNA damage induced by etoposide.
Evidence for the importance of homologous recombination
comes largely from studies on the Rad51 protein (which is
homologous to S. cerevisiae Rad51p) [125-129]. There is a
strong correlation between levels of this protein and
sensitivity to etoposide in human small cell lung cancer cells.
The lower the level of Rad51 protein, the more sensitive
cells are to the drug and the greater the physiological
concentration of DNA breaks [129]. In addition, treatment of
mammalian cell lines with etoposide induces the formation
of Rad51 nuclear foci [125, 126].
Evidence for the importance of nonhomologous end-
joining comes from studies on the Ku70/80 and Lig4
proteins. In a variety of mammalian cell lines, deficiencies in
any of these proteins result in hypersensitivity to etoposide
[130]. Deletion of DNA-PKcs (the catalytic subunit, which
together with the Ku70/80 proteins, comprises the DNA-PK
complex [131]) shows a less severe phenotype [130]. In this
regard, it is notable that the Ku70 protein does not recognize
the DNA in a topoisomerase II cleavage complex unless the
enzyme is first proteolyzed [132]. Similarly, the enzyme has
to be processed from the DNA termini in order for DNA-PK
to be activated [132].
Finally, studies using mutant chicken cell lines suggest
that nonhomologous end-joining plays a more important role
than homologous recombination in the repair of etoposide-
induced topoisomerase II-mediated DNA breaks [133, 134].
Whereas RAD54 null cells are 10- to 100-fold hypersensitive
to etoposide, KU70 or LIG4 null cells are 1,000- to 10,000-
fold hypersensitive the drug [133].
F. INITIATION OF LEUKEMIA BY ETOPOSIDE
Despite the importance of etoposide in the chemothera-
peutic treatment of human cancers, considerable evidence
suggests that the drug also triggers chromosomal trans-
locations that lead to specific types of leukemia (see Fig. (3))
[52, 58, 135-141]. It is estimated that 2-3% of patients who
receive drug regimens that include etoposide eventually
develop treatment-related leukemias [140]. The majority of
etoposide-related leukemias display translocations with
breakpoints that cluster within an 8.3 kb region between
Curr. Med. Chem. – Anti-Cancer Agents, 2005, Vol. 5, No. 4 369
exons 5-11 of the mixed lineage leukemia (MLL) gene at
chromosome band 11q23 [52, 58, 135, 142-147]. Transloca-
tions involving chromosomal band 11q23 also are observed
in primary human CD34+ cells following treatment with
etoposide [148].
MLL translocations involve partner genes on a variety of
different chromosomes [52, 58, 144, 145, 147, 149, 150].
However, the der(11) chromosomes in all of these trans-
locations express a chimeric protein consisting of the N-
terminus of MLL and the C-terminus of the partner protein.
The mechanism through which the chimeric fusion protein
leads to leukemia is not known. It has been suggested that
the fusion gene encodes an inactive MLL protein that acts as
a dominant negative to block the actions of the normal
protein [151]. It also has been proposed that the translocation
partner protein provides an active functional role [152].
Several lines of evidence point to a direct role for
topoisomerase II-mediated DNA cleavage in the initiation of
at least some MLL translocations. Treatment-related leuke-
mias with 11q23 chromosomal rearrangements are observed
primarily in patients that have been treated with regimens
that include etoposide [52, 58, 141]. Similar chromosomal
rearrangements also are seen in patients that receive other
topoisomerase II-targeted drugs as part of their therapy [52,
58, 141]. In addition, all of the DNA breakpoints mapped in
patient samples are located in close proximity to topoiso-
merase II-DNA cleavage sites [89, 153-157]. Furthermore,
these chromosomal translocations often involve precise or
near-precise rearrangements between topoisomerase II
cleavage sites on chromosome 11 and its partners [157, 158].
Finally, as discussed above, etoposide is converted to its
more active catechol (and subsequent quinone) metabolite by
CYP3A4 [108-111]. Individuals who carry a polymorphism
in the promoter region of the CYP3A4 gene display a decre-
ased probability of developing treatment-related leukemias
[159].
It should be noted that recent studies with human cell
lines indicate that DNA breaks within the MLL gene and
tranlocations involving this locus are observed in cells that
survive apoptotic events [160-162]. Therefore, it is possible
that the etoposide-induced breakpoints observed in deriva-
tive chromosomes do not arise directly from the drug-
stabilized DNA breaks mediated by topoisomerase II, but
rather from DNA breaks generated by apoptotic processes. In
this latter case, increased cellular levels of cleavage com-
plexes would serve primarily to initiate apoptosis. However,
since topoisomerase II has been implicated in excising DNA
loop domains during apoptosis [163, 164], topoisomerase II
could still be generating the apoptotic DNA breaks that
eventually recombine to form the chromosomal transloca-
tions.
In addition to treatment-related leukemias, translocations
that involve the MLL gene are associated with infant acute
myeloid leukemias (AMLs) [147, 165-167]. The chromo-
somal translocations associated with these cancers are
observed in utero [167-169]. Epidemiological studies indi-
cate that maternal consumption of foods during pregnancy
that are high in naturally occurring topoisomerase II poisons
increase the risk of developing infant AMLs ~10-fold [170,
171].
370 Curr. Med. Chem. – Anti-Cancer Agents, 2005, Vol. 5, No. 4 Baldwin and Osheroff

The ability of etoposide and other topoisomerase II [20] Tsai-Pflugfelder, M.; Liu, L. F.; Liu, A. A.; Tewey, K. M.; Whang-
poisons to cure rather than cause cancer may be related to Peng, J.; Knutsen, T.; Huebner, K.; Croce, C. M.; Wang, J. C.
Proc. Natl. Acad. Sci. USA, 1988, 85, 7177-7181.
cellular levels of topoisomerase II-mediated DNA cleavage [21] Jenkins, J. R.; Ayton, P.; Jones, T.; Davies, S. L.; Simmons, D. L.;
and the activity of cell death pathways. If the concentration Harris, A. L.; Sheer, D.; Hickson, I. D. Nucleic Acids Res., 1992,
of enzyme-associated DNA breaks is too low to overwhelm 20, 5587-5592.
recombination/repair pathways or to trigger cell death, the [22] Tan, K. B.; Dorman, T. E.; Falls, K. M.; Chung, T. D.; Mirabelli,
C. K.; Crooke, S. T.; Mao, J. Cancer Res., 1992, 52, 231-234.
pathways that promote cell survival can lead to the formation [23] Austin, C. A.; Marsh, K. L. BioEssays, 1998, 20, 215-226.
of the chromosomal translocations that ultimately lead to [24] Nitiss, J. L. Biochim. Biophys. Acta, 1998, 1400, 63-81.
cancerous growth. [25] Wang, J. C. Nat. Rev. Mol. Cell. Biol., 2002, 3, 430-440.
[26] Woessner, R. D.; Mattern, M. R.; Mirabelli, C. K.; Johnson, R. K.;
G. CONCLUSIONS AND PERSPECTIVES Drake, F. H. Cell Growth Differ., 1991, 2, 209-214.
[27] Sullivan, D. M.; Latham, M. D.; Ross, W. E. Cancer Res. , 1987,
Etoposide is an important chemotherapeutic agent that is 47, 3973-3979.
[28] Heck, M. M.; Hittelman, W. N.; Earnshaw, W. C. Proc. Natl. Acad.
used to treat a wide spectrum of human cancers. However, Sci. USA, 1988, 85, 1086-1090.
there is also evidence that the drug has the capacity to trigger [29] Hsiang, Y. H.; Wu, H. Y.; Liu, L. F. Cancer Res., 1988, 48, 3230-
distinct leukemias that are associated with chromosomal 3235.
translocations involving the MLL gene. Both of these conse- [30] Isaacs, R. J.; Davies, S. L.; Sandri, M. I.; Redwood, C.; Wells, N.
quences of etoposide treatment appear arise from the ability J.; Hickson, I. D. Biochim. Biophys. Acta, 1998, 1400, 121-137.
[31] Carpenter, A. J.; Porter, A. C. Mol. Biol. Cell, 2004, 15, 5700-
of the drug to stabilize topoisomerase II-DNA cleavage 5711.
complexes. Given the clinical prevalence of etoposide, it is [32] Dereuddre, S.; Delaporte, C.; Jacquemin-Sablon, A. Cancer Res.,
critical to fully understand the mechanism by which this 1997, 57, 4301-4308.
drug affects the catalytic functions of topoisomerase II and [33] Grue, P.; Grasser, A.; Sehested, M.; Jensen, P. B.; Uhse, A.;
Straub, T.; Ness, W.; Boege, F. J. Biol. Chem., 1998, 273, 33660-
the cellular processes that repair topoisomerase II-mediated 33666.
DNA breaks. With this knowledge, it may be possible to [34] Yang, X.; Li, W.; Prescott, E. D.; Burden, S. J.; Wang, J. C.
improve the anticancer properties of etoposide while Science, 2000, 287, 131-134.
decreasing the risk of treatment-related leukemias. [35] Zechiedrich, E. L.; Osheroff, N. EMBO J., 1990, 9, 4555-4562.
[36] Roca, J.; Berger, J. M.; Wang, J. C. J. Biol. Chem., 1993, 268,
14250-14255.
ACKNOWLEDGEMENTS [37] Osheroff, N. Biochemistry, 1987, 26, 6402-6406.
[38] Liu, L. F.; Rowe, T. C.; Yang, L.; Tewey, K. M.; Chen, G. L. J.
The authors are grateful to Jennifer S. Dickey and Renier Biol. Chem., 1983, 258, 15365-15370.
Vélez-Cruz for the critical reading of this manuscript. The [39] Sander, M.; Hsieh, T. J. Biol. Chem., 1983, 258, 8421-8428.
senior author’s laboratory is supported by National Institutes [40] Zechiedrich, E. L.; Christiansen, K.; Andersen, A. H.; Westergaard,
of Health research grants GM33944 and GM53960. E.L.B. O.; Osheroff, N. Biochemistry, 1989, 28, 6229-6236.
was a trainee under National Institutes of Health grant 5 T32 [41] Osheroff, N.; Shelton, E. R.; Brutlag, D. L. J. Biol. Chem., 1983,
258, 9536-9543.
CA09582. [42] Lindsley, J. E.; Wang, J. C. Proc. Natl. Acad. Sci. USA, 1991, 88,
10485-10489.
REFERENCES [43] Baird, C. L.; Harkins, T. T.; Morris, S. K.; Lindsley, J. E. Proc.
Natl. Acad. Sci. USA, 1999, 96, 13685-13690.
[1] Stahelin, H. F.; von Wartburg, A. Cancer Res., 1991, 51, 5-15. [44] Osheroff, N. J. Biol. Chem., 1986, 261, 9944-9950.
[2] Hande, K. R. Eur. J. Cancer, 1998, 34, 1514-1521. [45] Roca, J.; Wang, J. C. Cell, 1992, 71, 833-840.
[3] Vogelzang, N. J.; Raghavan, D.; Kennedy, B. J. Am. J. Med., 1982, [46] Rowe, T. C.; Chen, G. L.; Hsiang, Y. H.; Liu, L. F. Cancer Res.,
72, 136-144. 1986, 46, 2021-2026.
[4] Sinkule, J. A. Pharmacotherapy, 1984, 4, 61-73. [47] Horowitz, D. S.; Wang, J. C. J. Biol. Chem., 1987, 262, 5339-5344.
[5] Loike, J. D.; Horwitz, S. B. Biochemistry, 1976, 15, 5435-5443. [48] Worland, S. T.; Wang, J. C. J. Biol. Chem., 1989, 264, 4412-4416.
[6] Ross, W.; Rowe, T.; Glisson, B.; Yalowich, J.; Liu, L. Cancer Res., [49] Wang, J. C. Annu. Rev. Biochem., 1996, 65, 635-692.
1984, 44, 5857-5860. [50] Baguley, B. C.; Ferguson, L. R. Biochim. Biophys. Acta, 1998,
[7] Chen, G. L.; Yang, L.; Rowe, T. C.; Halligan, B. D.; Tewey, K. M.; 1400, 213-222.
Liu, L. F. J. Biol. Chem., 1984, 259, 13560-13566. [51] Kaufmann, S. H. Biochim. Biophys. Acta, 1998, 1400, 195-211.
[8] Yang, L.; Rowe, T. C.; Liu, L. F. Cancer Res., 1985, 45, 5872- [52] Felix, C. A. Biochim. Biophys. Acta, 1998, 1400, 233-255.
5876. [53] Holden, J. A. Curr. Med. Chem. Anti-Canc. Agents, 2001, 1, 1-25.
[9] Wang, J. C. Quart. Rev. Biophys., 1998, 31, 107-144. [54] Holm, C.; Covey, J. M.; Kerrigan, D.; Pommier, Y. Cancer Res. ,
[10] Burden, D. A.; Osheroff, N. Biochim. Biophys. Acta, 1998, 1400, 1989, 49, 6365-6368.
139-154. [55] D'Arpa, P.; Beardmore, C.; Liu, L. F. Cancer Res., 1990, 50, 6919-
[11] Berger, J. M. Biochim. Biophys. Acta, 1998, 1400, 3-18. 6924.
[12] Berger, J. M. Curr. Opin. Struct. Biol., 1998, 8, 26-32. [56] Corbett, A. H.; Osheroff, N. Chem. Res. Toxicol., 1993, 6, 585-597.
[13] Fortune, J. M.; Osheroff, N. Prog. Nucleic Acid Res. Mol. Biol., [57] Howard, M. T.; Neece, S. H.; Matson, S. W.; Kreuzer, K. N. Proc.
2000, 64, 221-253. Natl. Acad. Sci. USA, 1994, 91, 12031-12035.
[14] Champoux, J. J. Annu. Rev. Biochem., 2001, 70, 369-413. [58] Felix, C. A. Med. Pediatr. Oncol., 2001, 36, 525-535.
[15] Wilstermann, A. M.; Osheroff, N. Curr. Top. Med. Chem., 2003, 3, [59] Kreuzer, K. N.; Cozzarelli, N. R. J. Bacteriol., 1979, 140, 424-435.
321-338. [60] Walker, J. V.; Nitiss, J. L. Cancer Invest., 2002, 20, 570-589.
[16] Wyckoff, E.; Natalie, D.; Nolan, J. M.; Lee, M.; Hsieh, T. J. Mol. [61] Hande, K. R. Biochim. Biophys. Acta, 1998, 1400, 173-184.
Biol., 1989, 205, 1-13. [62] Loike, J. D.; Horwitz, S. B. Biochemistry, 1976, 15, 5443-5448.
[17] Goto, T.; Wang, J. C. Cell, 1984, 36, 1073-1080. [63] Wozniak, A. J.; Ross, W. E. Cancer Res., 1983, 43, 120-124.
[18] Drake, F. H.; Zimmerman, J. P.; McCabe, F. L.; Bartus, H. F.; Per, [64] Glisson, B. S.; Smallwood, S. E.; Ross, W. E. Biochim. Biophys.
S. R.; Sullivan, D. M.; Ross, W. E.; Mattern, M. R.; Johnson, R. Acta, 1984, 783, 74-79.
K.; Crooke, S. T. J. Biol. Chem., 1987, 262, 16739-16747. [65] Willmore, E.; Frank, A. J.; Padget, K.; Tilby, M. J.; Austin, C. A.
[19] Drake, F. H.; Hofmann, G. A.; Bartus, H. F.; Mattern, M. R.; Mol. Pharmacol., 1998, 54, 78-85.
Crooke, S. T.; Mirabelli, C. K. Biochemistry, 1989, 28, 8154-8160. [66] Hsiang, Y. H.; Liu, L. F. J. Biol. Chem., 1989, 264, 9713-9715.
Etoposide, Topoisomerase II and Cancer Curr. Med. Chem. – Anti-Cancer Agents, 2005, Vol. 5, No. 4 371

[67] Byl, J. A.; Cline, S. D.; Utsugi, T.; Kobunai, T.; Yamada, Y.; [104] Arimondo, P.; Boukarim, C.; Bailly, C.; Dauzonne, D.; Monneret,
Osheroff, N. Biochemistry, 2001, 40, 712-718. C. Anticancer Drug Des., 2000, 15, 413-421.
[68] Glisson, B. S.; Sullivan, D. M.; Gupta, R.; Ross, W. E. Natl. [105] Guianvarc'h, D.; Duca, M.; Boukarim, C.; Kraus-Berthier, L.;
Cancer Inst. Monogr., 1987, 4, 89-93. Leonce, S.; Pierre, A.; Pfeiffer, B.; Renard, P.; Arimondo, P. B.;
[69] Danks, M. K.; Yalowich, J. C.; Beck, W. T. Cancer Res., 1987, 47, Monneret, C.; Dauzonne, D. J. Med. Chem., 2004, 47, 2365-2374.
1297-1301. [106] Sinha, B. K.; Politi, P. M.; Eliot, H. M.; Kerrigan, D.; Pommier, Y.
[70] Danks, M. K.; Schmidt, C. A.; Cirtain, M. C.; Suttle, D. P.; Beck, Eur. J. Cancer, 1990, 26, 590-593.
W. T. Biochemistry, 1988, 27, 8861-8869. [107] Saulnier, M. G.; Vyas, D. M.; Langley, D. R.; Doyle, T. W.; Rose,
[71] Sullivan, D. M.; Latham, M. D.; Rowe, T. C.; Ross, W. E. W. C.; Crosswell, A. R.; Long, B. H. J. Med. Chem., 1989, 32,
Biochemistry, 1989, 28, 5680-5687. 1418-1420.
[72] Nitiss, J. L.; Liu, Y. X.; Harbury, P.; Jannatipour, M.; Wasserman, [108] van Maanen, J. M.; Retel, J.; de Vries, J.; Pinedo, H. M. J. Natl.
R.; Wang, J. C. Cancer Res., 1992, 52, 4467-4472. Cancer Inst., 1988, 80, 1526-1533.
[73] Nitiss, J. L.; Liu, Y. X.; Hsiung, Y. Cancer Res., 1993, 53, 89-93. [109] Mans, D. R.; Retel, J.; van Maanen, J. M.; Lafleur, M. V.; van
[74] Jannatipour, M.; Liu, Y. X.; Nitiss, J. L. J. Biol. Chem., 1993, 268, Schaik, M. A.; Pinedo, H. M.; Lankelma, J. Br. J. Cancer, 1990,
18586-18592. 62, 54-60.
[75] Liu, Y.-X.; Hsiung, Y.; Jannatipour, M.; Yeh, Y.; Nitiss, J. L. [110] Relling, M. V.; Nemec, J.; Schuetz, E. G.; Schuetz, J. D.;
Cancer Res., 1994, 54, 2943-2951. Gonzalez, F. J.; Korzekwa, K. R. Mol. Pharmacol., 1994, 45, 352-
[76] Nitiss, J. L.; Vilalta, P. M.; Wu, H.; McMahon, J. Mol. Pharmacol., 358.
1994, 46, 773-777. [111] Zhuo, X.; Zheng, N.; Felix, C. A.; Blair, I. A. Drug Metab. Dispos.,
[77] Elsea, S. H.; Hsiung, Y.; Nitiss, J. L.; Osheroff, N. J. Biol. Chem. , 2004, 32, 993-1000.
1995, 270, 1913-1920. [112] Stremetzne, S.; Jaehde, U.; Kasper, R.; Beyer, J.; Siegert, W.;
[78] Hsiung, Y.; Elsea, S. H.; Osheroff, N.; Nitiss, J. L. J. Biol. Chem. , Schunack, W. Eur. J. Cancer, 1997, 33, 978-979.
1995, 270, 20359-20364. [113] Relling, M. V.; Yanishevski, Y.; Nemec, J.; Evans, W. E.; Boyett,
[79] Cornarotti, M.; Tinelli, S.; Willmore, E.; Zunino, F.; Fisher, L. M.; J. M.; Behm, F. G.; Pui, C. H. Leukemia, 1998, 12, 346-352.
Austin, C. A.; Capranico, G. Mol. Pharmacol., 1996, 50, 1463- [114] Paques, F.; Haber, J. E. Microbiol. Mol. Biol. Rev., 1999, 63, 349-
1471. 404.
[80] Froelich-Ammon, S. J.; Osheroff, N. J. Biol. Chem., 1995, 270, [115] Haber, J. E. Trends Genet., 2000, 16, 259-264.
21429-21432. [116] Dudas, A.; Chovanec, M. Mutat. Res., 2004, 566, 131-167.
[81] Chow, K. C.; Macdonald, T. L.; Ross, W. E. Mol. Pharmacol., [117] Hopfner, K. P.; Putnam, C. D.; Tainer, J. A. Curr. Opin. Struct.
1988, 34, 467-473. Biol., 2002, 12, 115-122.
[82] Kingma, P. S.; Burden, D. A.; Osheroff, N. Biochemistry, 1999, 38, [118] Moore, J. K.; Haber, J. E. Mol. Cell. Biol., 1996, 16, 2164-2173.
3457-3461. [119] Lewis, L. K.; Resnick, M. A. Mutat. Res., 2000, 451, 71-89.
[83] Leroy, D.; Kajava, A. V.; Frei, C.; Gasser, S. M. Biochemistry, [120] Sabourin, M.; Nitiss, J. L.; Nitiss, K. C.; Tatebayashi, K.; Ikeda,
2001, 40, 1624-1634. H.; Osheroff, N. Nucleic Acids Res., 2003, 31, 4373-4384.
[84] Burden, D. A.; Kingma, P. S.; Froelich-Ammon, S. J.; Bjornsti, M.- [121] Baldwin, E. L.; Berger, A. C.; Corbett, A. H.; Osheroff, N. Nucleic
A.; Patchan, M. W.; Thompson, R. B.; Osheroff, N. J. Biol. Chem., Acids Res., 2005, 33, 1021-1030.
1996, 271, 29238-29244. [122] Araki, Y.; Kawasaki, Y.; Sasanuma, H.; Tye, B. K.; Sugino, A.
[85] Pommier, Y.; Capranico, G.; Orr, A.; Kohn, K. W. Nucleic Acids Genes Cells, 2003, 8, 465-480.
Res., 1991, 19, 5973-5980. [123] Malik, M.; Nitiss, J. L. Eukaryot. Cell, 2004, 3, 82-90.
[86] Capranico, G.; Zunino, F. Eur. J. Cancer, 1992, 12, 2055-2060. [124] Valerie, K.; Povirk, L. F. Oncogene, 2003, 22, 5792-5812.
[87] Pommier, Y.; Fesen, M. R.; Goldwasser, F. In Cancer [125] Haaf, T.; Raderschall, E.; Reddy, G.; Ward, D. C.; Radding, C. M.;
Chemotherapy and Biotherapy: Principles and Practice., B. A. Golub, E. I. J. Cell Biol., 1999, 144, 11-20.
Chabner; D. L. Longo, eds.; Lippincott-Raven Publishers: [126] Raderschall, E.; Golub, E. I.; Haaf, T. Proc. Natl. Acad. Sci. USA,
Philadelphia, 1996, pp. 435-461. 1999, 96, 1921-1926.
[88] Capranico, G.; Binaschi, M. Biochim. Biophys. Acta, 1998, 1400, [127] Raderschall, E.; Bazarov, A.; Cao, J.; Lurz, R.; Smith, A.; Mann,
185-194. W.; Ropers, H. H.; Sedivy, J. M.; Golub, E. I.; Fritz, E.; Haaf, T. J.
[89] Lovett, B. D.; Strumberg, D.; Blair, I. A.; Pang, S.; Burden, D. A.; Cell Sci., 2002, 115, 153-164.
Megonigal, M. D.; Rappaport, E. F.; Rebbeck, T. R.; Osheroff, N.; [128] Lundin, C.; Schultz, N.; Arnaudeau, C.; Mohindra, A.; Hansen, L.
Pommier, Y. G.; Felix, C. A. Biochemistry, 2001, 40, 1159-1170. T.; Helleday, T. J. Mol. Biol., 2003, 328, 521-535.
[90] Osheroff, N. Biochemistry, 1989, 28, 6157-6160. [129] Hansen, L. T.; Lundin, C.; Spang-Thomsen, M.; Petersen, L. N.;
[91] Robinson, M. J.; Osheroff, N. Biochemistry, 1991, 30, 1807-1813. Helleday, T. Int. J. Cancer, 2003, 105, 472-479.
[92] Bromberg, K. D.; Burgin, A. B.; Osheroff, N. J. Biol. Chem., 2003, [130] Jin, S.; Inoue, S.; Weaver, D. T. Carcinogenesis, 1998, 19, 965-
278, 7406-7412. 971.
[93] Berger, J. M.; Gamblin, S. J.; Harrison, S. C.; Wang, J. C. Nature, [131] Jackson, S. P. Int. J. Biochem. Cell. Biol., 1997, 29, 935-938.
1996, 379, 225-232. [132] Martensson, S.; Nygren, J.; Osheroff, N.; Hammarsten, O. Radiat.
[94] Fass, D.; Bogden, C. E.; Berger, J. M. Nat. Struct. Biol., 1999, 6, Res., 2003, 160, 291-301.
322-326. [133] Adachi, N.; Suzuki, H.; Iiizumi, S.; Koyama, H. J. Biol. Chem.,
[95] Wilstermann, A. M.; Osheroff, N. J. Biol. Chem., 2001, 276, 2003, 278, 35897-35902.
17727-17731. [134] Adachi, N.; Iiizumi, S.; So, S.; Koyama, H. Biochem. Biophys. Res.
[96] Costanzo, V.; Shechter, D.; Lupardus, P. J.; Cimprich, K. A.; Commun., 2004, 318, 856-861.
Gottesman, M.; Gautier, J. Mol. Cell, 2003, 11, 203-213. [135] DeVore, R.; Whitlock, J.; Hainsworth, T.; Johnson, D. Ann. Intern.
[97] Macdonald, T. L.; Legnert, E. K.; Loper, J. T.; Chow, K.-C.; Ross, Med., 1989, 110, 740-742.
W. E. In DNA Topoisomerases in Cancer, M. Potmesil; K.W. [136] Ratain, M. J.; Kaminer, L. S.; Bitran, J. D.; Lawson, R. A.; Le
Kohn, eds.; Oxford University Press: New York, 1991, pp. 199- Beau, M. M.; Skosey, C.; Purl, S.; Hoffman, P. C.; Wade, J.;
214. Vardiman, J. W.; Daly, K.; Rowley, J. D.; Golomb, H. M. Blood,
[98] Wilstermann, A. M. G., M.; Anklin, C.; Berkowitz, D. B.; Choi, S.; 1987, 70, 1412-1417.
Osheroff, N.; Graves, D. E. Proc. Am. Assoc. Cancer Res., 2003, [137] Pui, C.-H.; Ribeiro, R. C.; Hancock, M. L.; Rivera, G. K.; Evans,
135. W. E.; Raimondi, S. C.; Head, D. R.; Behm, F. G.; Mahmoud, M.
[99] Loike, J. D. Cancer Chemother. Pharmacol., 1982, 7, 103-111. H.; Sandlund, J. T.; Crist, W. M. N. Engl. J. Med., 1991, 325,
[100] Long, B. H.; Musial, S. T.; Brattain, M. G. Biochemistry, 1984, 23, 1682-1687.
1183-1188. [138] Winick, N. J.; McKenna, R. W.; Shuster, J. J.; Schneider, N. R.;
[101] Long, B. H. Sem. Oncol., 1992, 19, 3-19. Borowitz, M. J.; Bowman, W. P.; Jacaruso, D.; Kamen, B. A.;
[102] Long, B. H.; Casazza, A. M. Cancer Chemother. Pharmacol., Buchanan, G. R. J. Clin. Oncol., 1993, 11, 209-217.
1994, 34, S26-S31. [139] Sandoval, C.; Pui, C. H.; Bowman, L. C.; Heaton, D.; Hurwitz, C.
[103] Lee, K. H. J. Biomed. Sci., 1999, 6, 236-250. A.; Raimondi, S. C.; Behm, F. G.; Head, D. R. J. Clin. Oncol.,
1993, 11, 1039-1045.
372 Curr. Med. Chem. – Anti-Cancer Agents, 2005, Vol. 5, No. 4
[140] Smith, M. A.; Rubinstein, L.; Anderson, J. R.; Arthur, D.;
Catalano, P. J.; Freidlin, B.; Heyn, R.; Khayat, A.; Krailo, M.;
Land, V. J.; Miser, J.; Shuster, J.; Vena, D. J. Clin. Oncol., 1999,
17, 569-577.
[141] Andersen, M. K.; Christiansen, D. H.; Jensen, B. A.; Ernst, P.;
Hauge, G.; Pedersen-Bjergaard, J. Br. J. Haematol., 2001, 114,
539-543.
[142] Pedersen-Bjergaard, J.; Philip, P. Blood, 1991, 78, 1147-1148.
[143] Pedersen-Bjergaard, J. Leuk. Res., 1992, 16, 733-735.
[144] Felix, C. A.; Winick, N. J.; Negrini, M.; Bowman, W. P.; Croce, C.
M.; Lange, B. J. Cancer Res., 1993, 53, 2954-2956.
[145] Felix, C. A.; Hosler, M. R.; Winick, N. J.; Masterson, M.; Wilson,
A. E.; Lange, B. J. Blood, 1995, 85, 3250-3256.
[146] Broeker, P. L.; Super, H. G.; Thirman, M. J.; Pomykala, H.;
Yonebayashi, Y.; Tanabe, S.; Zeleznik-Le, N.; Rowley, J. D.
Blood, 1996, 87, 1912-1922.
[147] Rowley, J. D. Annu. Rev. Gen., 1998, 32, 495-519.
[148] Libura, J.; Slater, D. J.; Felix, C. A.; Richardson, C. Blood, 2004.
[149] Super, H. J.; McCabe, N. R.; Thirman, M. J.; Larson, R. A.; Le
Beau, M. M.; Pedersen-Bjergaard, J.; Philip, P.; Diaz, M. O.;
Rowley, J. D. Blood, 1993, 82, 3705-3711.
[150] Domer, P. H.; Head, D. R.; Renganathan, N.; Raimondi, S. C.;
Yang, E.; Atlas, M. Leukemia, 1995, 9, 1305-1312.
[151] Schichman, S. A.; Canaani, E.; Croce, C. M. JAMA, 1995, 273,
571-576.
[152] Biondi, A.; Cimino, G.; Pieters, R.; Pui, C. H. Blood, 2000, 96, 24-
33.
[153] Negrini, M.; Felix, C. A.; Martin, C.; Lange, B. J.; Nakamura, T.;
Canaani, E.; Croce, C. M. Cancer Res., 1993, 53, 4489-4492.
[154] Felix, C. A.; Lange, B. J.; Hosler, M. R.; Fertala, J.; Bjornsti, M.-A.
Cancer Res., 1995, 55, 4287-4292.
[155] Aplan, P. D.; Chervinsky, D. S.; Stanulla, M.; Burhans, W. C.
Blood, 1996, 87, 2649-2658.
Baldwin and Osheroff
[156] Strissel, P. L.; Strick, R.; Rowley, J. D.; Zeleznik-Le, N. J. Blood,
1998, 92, 3793-3803.
[157] Lovett, B. D.; Lo Nigro, L.; Rappaport, E. F.; Blair, I. A.; Osheroff,
N.; Zheng, N.; Megonigal, M. D.; Williams, W. R.; Nowell, P. C.;
Felix, C. A. Proc. Natl. Acad. Sci. USA, 2001, 98, 9802-9807.
[158] Whitmarsh, R. J.; Saginario, C.; Zhuo, Y.; Hilgenfeld, E.;
Rappaport, E. F.; Megonigal, M. D.; Carroll, M.; Liu, M.;
Osheroff, N.; Cheung, N. K.; Slater, D. J.; Ried, T.; Knutsen, T.;
Blair, I. A.; Felix, C. A. Oncogene, 2003, 22, 8448-8459.
[159] Felix, C. A.; Walker, A. H.; Lange, B. J.; Williams, T. M.; Winick,
N. J.; Cheung, N. K.; Lovett, B. D.; Nowell, P. C.; Blair, I. A.;
Rebbeck, T. R. Proc. Natl. Acad. Sci. USA, 1998, 95, 13176-13181.
[160] Stanulla, M.; Wang, J.; Chervinsky, D. S.; Thandla, S.; Aplan, P.
D. Mol. Cell. Biol., 1997, 17, 4070-4079.
[161] Betti, C. J.; Villalobos, M. J.; Diaz, M. O.; Vaughan, A. T. Cancer
Res., 2001, 61, 4550-4555.
[162] Betti, C. J.; Villalobos, M. J.; Diaz, M. O.; Vaughan, A. T. Cancer
Res., 2003, 63, 1377-1381.
[163] Li, T. K.; Chen, A. Y.; Yu, C.; Mao, Y.; Wang, H.; Liu, L. F.
Genes Dev., 1999, 13, 1553-1560.
[164] Solovyan, V. T.; Bezvenyuk, Z. A.; Salminen, A.; Austin, C. A.;
Courtney, M. J. J. Biol. Chem., 2002, 277, 21458-21467.
[165] Felix, C. A.; Hosler, M. R.; Slater, D. J.; Parker, R. I.; Masterson,
M.; Whitlock, J. A.; Rebbeck, T. R.; Nowell, P. C.; Lange, B. J. J.
Pediatr. Hematol. Oncol., 1998, 20, 299-308.
[166] Felix, C. A.; Lange, B. J. Oncologist, 1999, 4, 225-240.
[167] Strick, R.; Strissel, P. L.; Borgers, S.; Smith, S. L.; Rowley, J. D.
Proc. Natl. Acad. Sci. USA, 2000, 97, 4790-4795.
[168] Ford, A. M.; Ridge, S. A.; Cabrera, M. E.; Mahmoud, H.; Steel, C.
M.; Chan, L. C.; Greaves, M. Nature, 1993, 363, 358-360.
[169] Ross, J. A. Proc. Natl. Acad. Sci. USA, 2000, 97, 4411-4413.
[170] Ross, J. A.; Potter, J. D.; Reaman, G. H.; Pendergrass, T. W.;
Robison, L. L. Cancer Causes Control, 1996, 7, 581-590.
[171] Ross, J. A. Int. J. Cancer Suppl., 1998, 11, 26-28.