The discovery of a new potential anticancer drug: A case history

Article abstract of DOI:10.1016/S0014-827X(03)00014-4

DNA minor groove binders (MGB) represent a class of anticancer agents whose DNA sequence specificity was hypothesized to lead to high selectivity of action. Tallimustine (TAM), a benzoyl nitrogen mustard derivative of distamycin A (DST), showed excellent antitumor activity in preclinical tests, but also a severe myelotoxicity. Novel nitrogen mustard, nitrogen half-mustard and sulfur mustard derivatives of DST showing excellent activity were recently identified and SAR reported. In particular nitrogen half-mustard and sulfur mustard derivatives, as one-arm alkylating agents, represent interesting structural novelties. A further new class of cytotoxic anticancer agents is that of α-halogenoacrylamido derivatives of DST-like oligopeptides, which show an activity profile substantially improved in comparison to TAM. In particular brostallicin (PNU-166196), α-bromo-acrylamido tetra-pyrrole derivative ending with a guanidino moiety, showed high cytotoxic potency and myelotoxicity dramatically reduced in comparison to TAM and other MGB. Brostallicin binds to the minor groove but appears unreactive in classical in vitro DNA alkylation assays. About the apparent lack of DNA alkylation we speculated that an intracellular nucleophile, e.g. glutathione (GSH), could activate the reactivity of the compound leading to alkylation of DNA in vivo. Evidence of both covalent interaction of brostallicin with plasmidic DNA in the presence of GSH and of enhanced cytotoxicity in cancer cells characterized by high levels of GSH were obtained. Brostallicin was selected for clinical development and is now undergoing Phase II studies.

Full text of DOI:10.1016/S0014-827X(03)00014-4

Il Farmaco 58 (2003) 213Á220
/
www.elsevier.com/locate/farmac
The discovery of a new potential anticancer drug: a case history
Paolo Cozzi
Department of Chemistry, Pharmacia, Global Chemistry, Discovery Research Oncology, Viale Pasteur 10, 20014 Nerviano, Milan, Italy
Accepted 24 July 2002
Abstract
DNA minor groove binders (MGB) represent a class of anticancer agents whose DNA sequence specificity was hypothesized to
lead to high selectivity of action. Tallimustine (TAM), a benzoyl nitrogen mustard derivative of distamycin A (DST), showed
excellent antitumor activity in preclinical tests, but also a severe myelotoxicity. Novel nitrogen mustard, nitrogen half-mustard and
sulfur mustard derivatives of DST showing excellent activity were recently identified and SAR reported. In particular nitrogen half-
mustard and sulfur mustard derivatives, as one-arm alkylating agents, represent interesting structural novelties. A further new class
of cytotoxic anticancer agents is that of a-halogenoacrylamido derivatives of DST-like oligopeptides, which show an activity profile
substantially improved in comparison to TAM. In particular brostallicin (PNU-166196), a-bromo-acrylamido tetra-pyrrole
derivative ending with a guanidino moiety, showed high cytotoxic potency and myelotoxicity dramatically reduced in comparison to
TAM and other MGB. Brostallicin binds to the minor groove but appears unreactive in classical in vitro DNA alkylation assays.
About the apparent lack of DNA alkylation we speculated that an intracellular nucleophile, e.g. glutathione (GSH), could activate
the reactivity of the compound leading to alkylation of DNA in vivo. Evidence of both covalent interaction of brostallicin with
plasmidic DNA in the presence of GSH and of enhanced cytotoxicity in cancer cells characterized by high levels of GSH were
obtained. Brostallicin was selected for clinical development and is now undergoing Phase II studies.
´
# 2003 Editions scientifiques et me´dicales Elsevier SAS. All rights reserved.
Keywords: Minor groove; Distamycin; Brostallicin; a-Halogenoacrylamides; Glutathione
1. Introduction
32 versus for prostate; 24 versus 19 for urinary tract; 14
versus 12 for ovary [1,4]. The classical 5-year survival
rate for the average of the more common cancers is
about 60%, but is as low as 14% for lung cancer. In
general the 5-year survival rate is increasing, however,
this reflects more the great progress in early diagnosis,
as the large scale practices of mammography, PSA and
Pap test, than the real capability of therapeutic control
[5].
Cancer is an heterogeneous group of diseases, char-
acterized by uncontrolled growth of the malignant cell
population. The estimated worldwide incidence of
different types of cancers is around 10 millions, roughly
half of which in developed countries. The incidence,
prevalence and mortality in the USA and in Italy are,
respectively, about 1.2, 8 millions, 550 thousands and
A dramatic decrease of mortality occurred, at least in
developed countries, only for those cancers in which
both early diagnosis and full organ surgery are possible,
as in the case of uterus and cervix or for which a
substantial drop of incidence occurred, as in the case of
the stomach. In the USA, the cancer mortality 1995
versus 1950 dropped by 67% for uterus, 76% for cervix,
80% for stomach, while e.g. both mortality and in-
cidence increased by about 250% for lung cancer [5].
In spite of the large number of available chemother-
apeutic agents the medical need is still largely unmet.
The main reasons are: the lack of selectivity of conven-
about 250 thousands, 1 million, 80 thousands [1Á3].
/
In spite of impressive progress in diagnosis, surgery
and therapy, occurred since the Sixties, the overall
cancer mortality, even somewhat declined in late Nine-
ties in some countries, is still very high, with some
exceptions for some specific tumor types. For instance in
the USA the cancer deaths 2000 versus 1985 in
thousands were, respectively, 160 versus 120 for lung;
110 versus 60 for colon-rectum; 41 versus 38 for breast;
E-mail address: paolo.cozzi@cu.pnu.com (P. Cozzi).
´
0014-827X/03/$ - see front matter # 2003 Editions scientifiques et me´dicales Elsevier SAS. All rights reserved.
doi:10.1016/S0014-827X(03)00014-4

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P. Cozzi / Il Farmaco 58 (2003) 213Á220
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tional drugs, leading to toxicity; the metastatic spread-
ing, implying early tumor implantation in organs other
than primary site; the heterogeneity of the disease,
comprising about 100 types of cancer; the intrinsic or
acquired resistance to chemotherapy developed after few
therapeutic cycles (multi-drug resistance).
A number of innovative strategies are in development,
aimed to target the malignant abnormalities of tumor
cells arising from the activation of tumor promoter
genes or inhibition of tumor suppressor genes. Some
focused molecular targets are e.g. oncogene encoded
proteins acting as signal transducer like RAS, cell cycle
targets as cyclin-dependent kinases, telomerase. In
alternative, angiogenesis inhibition may prevent the
formation of tumor blood vessels, thus limiting the
tumor ability to grow and metastasize. Focused mole-
cular targets of angiogenesis cascade are e.g. the
proteases degrading cellular matrix, endothelial tyrosine
kinases, avb5 transmembrane integrins. However, it will
likely take some years to fully define the role of
innovative agents in preventing disease progression.
Therefore, cytotoxic drugs will continue to represent a
chief part of the therapy in the near future, and this
implies a need for new agents with better activity and
safety profile.
Fig. 1. Distamycin A and its benzoyl nitrogen mustard tallimustine.
IC50 against L1210 murine leukemia. Distamycin A: 10.0 mM,
Tallimustine: IC50
ꢀ/68.5 nM.
TAM showed cytotoxicity against L1210 murine
leukemia more than two orders of magnitude higher
then DST and more than one order of magnitude higher
then classical nitrogen mustard melphalan [11]. It may
be hypothesized that distamycin-driven DNA binding
increases the TAM concentration near the DNA target,
avoiding as much as possible a specific alkylation of
biological nucleophiles, as e.g. glutathione (GSH),
which is a major drawback of classical mustards [12].
TAM showed excellent antitumor activity in the pre-
clinical tests, however, its clinical evaluation showed a
severe myelotoxicity [13] and its development was
discontinued. Nevertheless TAM has represented an
important model for the design of new minor groove
alkylating agents having proved the possibility of
obtaining potent antitumor activity by tethering a
moiety of mild chemical reactivity, such as that repre-
sented by benzoic acid nitrogen mustard, to a DNA
selective binding-frame, thus leading to DNA alkylation
with highT4GA sequence specificity.
2. Distamycin-derived nitrogen mustards
The putative mode of action of many antitumor
agents involves a DNA damage, either by direct binding
of the drug to DNA or to DNA-binding proteins such as
topoisomerases. Most of DNA-interacting agents have
only a limited degree of sequence specificity, which
imply that they may hit most cellular genes. The minor
groove represents a vulnerable site of attack normally
unoccupied, while most of DNA interactive proteins
bind to the major groove. However, the minor groove is
the site of non-covalent, reversible, highly specific
interaction with thymine-adenine (TA)-rich sequences
for several molecules of MW less than 1 kDa endowed
with antibacterial and antiviral activities. One of these
molecules is distamycin A (DST) [6], which show a
highly selective reversible interaction with TA sequences
containing at least four T4GA base pairs [7].
DNA minor groove binders (MGB) able to damage
irreversibly DNA, e.g. to alkylate DNA, may represent
a new class of cytotoxics, whose increased gene specifi-
city may provide significant improvement in potency
and selectivity [8]. The main representatives of this class
that were clinically tested in the recent past are
tallimustine (TAM) [9], a benzoic acid nitrogen mustard
derivative of DST, Fig. 1, and the compounds structu-
rally derived from antibiotic CC-1065, i.e. adozelesin,
carzelesin, bizelesin [10].
Derivatives of DST called a particular interest also
because of their oligopeptidic nature, which suggested
the possibility of modulating the DNA binding cap-
ability by varying the number and the characteristics of
the azole units [14Á20]. Moreover the role of the
/
chemical reactivity of the phenyl mustard and that of
basicity of the amidino moiety were specifically investi-
gated by us [21,22]. The medicinal chemistry and the
SAR of distamycin-derived nitrogen mustards, has been
reviewed by several authors including us [23,24] and will
not be further discussed here.
However, a specific finding arising from our studies
seems worthwhile of a particular mention: the potent
activity of one-arm mustards derived form DST. This is
well exemplified by two cinnamic derivatives, the nitro-
gen ethyl half-mustard PNU-160366 and the sulfur
mustard PNU-193821 [21,25]. Fig. 2. Both compounds

P. Cozzi / Il Farmaco 58 (2003) 213Á
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215
Fig. 3. PNU-151807: the lead of distamycin-derived a-halogenoacry-
lamides. IC50 against L1210 murine leukemia.
The cytotoxicity of the compounds of this series
depends upon the chemical reactivity of a-halogen-
oacrylic moiety, that we studied with simple model
compounds, i.e. 4-(a-haloacrylamido)-pyrrole-2-carbox-
yanilides. The structures of derivatives obtained by the
reaction of these compounds with amines or the
imidazole ring are in agreement with a first-step Michael
nucleophilic attack, followed by a further reaction
leading to a second nucleophilic substitution or to b-
elimination [24] Fig. 4.
The dramatically different reactivity toward nucleo-
philic attack of a-bromoacrylamido and of a-fluoroa-
crylamido moieties corresponds to the dramatically
different cytotoxicity of a-bromo and a-fluoro-acryla-
mido DST derivatives, the former very potent, the latter
inactive. Also acrylamido DST derivative is devoid of
activity confirming the requirement of a reactive halo-
gen [29].
The relevant activity of PNU-151807, its cytotoxicity/
myelotoxicity ratio significantly better than that of
TAM [30] and its unusual mechanistic features,
prompted us to the synthesis of new halogenoacrylic
derivatives of DST and congeners. Therefore, we
modified the length of the DST frame, replaced the
amidino moiety with moieties of different features and
replaced one or more pyrrole units with other imidazole
or pyrazole rings.
Fig. 2. Cinnamic nitrogen half-mustard and sulfur mustard derivatives
of distamycin. IC50 against L1210 murine leukemia.
show the possibility of achieving potent activity by
tethering a reactive one-arm mustard to a minor groove
binding ligand, while classical nitrogen half-mustards
and sulfur mustards are substantially non cytotoxic [26],
due to the impossibility of crosslinking the DNA strands
[27]. Sulfur mustard PNU-193821 was the most potent
DST-derived cytotoxic found in our laboratories (IC50
0.9 nM against L1210 leukemia). The corresponding
sulfoxide derivative shows a substantial lack of cyto-
toxicity, suggesting that the formation of a thiiranium
cation, which is no more possible in the case of the
sulfoxide, may play a key role for the mechanism
leading to cytotoxicity.
The three pyrrole unit DST derivatives, both bromo
and chloro, are about one order of magnitude less
cytotoxic than the corresponding four pyrrole conge-
ners. The same decrease of activity occurs with a-bromo
derivatives with two and one-pyrrole units, which are,
therefore, devoid of significant activity (Table 1). This
feature was classically considered to arise from a tighter
DNA binding depending on the increased multiplicity of
interaction between the pyrrolecarboxamide units and
minor groove TA-rich sequences [31]. However, recent
permeability data obtained using a Caco-2 cells line
(Table 2) show that in the series of a-bromoacrylic
polypyrrole derivatives there is a progressive strong
increase of permeability from one to four pyrrole
compounds, which goes together with their increased
3. Distamycin-derived a-halogenoacrylamides
A new class of cytotoxic MGB showing a-bromo or
chloro-acrylamido moiety linked to DST or DST-like
frames was identified in our laboratory [11]. The first
lead of this class, PNU-151807, Fig. 3, the a-bromoa-
crylamido derivative of four-pyrrole DST homologue,
showed high cytotoxicity and in vivo activity. Moreover
PNU-151807 was found to bind to DNA minor groove
but unable to alkylate minor groove AT-rich sequences
in classical in vitro experiments [28] at variance with
TAM.

216
P. Cozzi / Il Farmaco 58 (2003) 213Á220
/
Fig. 4. Reactivity of a-bromoacrylamido moiety (a-bromoacrylamino-pyrrolecarboxyanilide). a-Fluoroacrylamino-pyrrolecarboxyanilide does not
react in the same or harder conditions.
Table 1
The effect of the length of distamycin-like frame on in vitro and in vivo
activity (L1210 leukemia)
Table 2
Caco-2 cells permeability for a series of a-bromoacrylamido deriva-
tives
a
b
n
Pappꢁ
/
10ꢂ6 (cm/s)
ClogD, pH 7.4
IC50 (nM)
X
n
In vitro IC50 (nM)
In vivo OD (mg/kg)
T/C (%)
4
3
2
1
38.20
15.21
5.58
ꢂ
/
6.63
6.3
Br
Br
Br
Br
Br
Cl
Cl
5
4
3
2
1
4
3
23.0 a9
6.39
98.89
/
5.0
0.78
1.56
12.5
n.d.
n.d.
1.56
12.5
167
200
100
n.d.
n.d.
133
117
ꢂ
/
6.0
98.8
1300
/
1.3
ꢂ
/
5.38
/
24.2
0.69
ꢂ
/
4.75
ꢀ/3000
ꢀ/1300
ꢀ/3000
a
Apparent permeability, method of P. Artusson.
ACD calculated LogD at pH 7.4.
3.89
/
1.4
b
96.89
/
24.2
more pyrrole units and suggests the existence of an
active transport mechanism for these compounds, whose
permeability cannot by related to a passive transport,
due to high MW, full ionization, high hydrophilicity and
high hydrogen bonding capability.
IC50
response curves of at least two experiments.
ꢀ
/
50% inhibitory concentration as the mean9
/
SE from dose-
a
Drug sensitivity determined after 4 h of continuous exposure
against L1210 cells, after 48 h for other compounds; for in vivo studies
cells were injected i.v. and mice were treated i.v. the day after
tumorinjection; O.D., optimal (non toxic) dose
B/
LD10; T/Cꢀ
/
median survival time of treated versus untreated miceꢁ
/
100.
As far as the role of strongly basic amidino moiety,
typical of DST, is concerned, the activity of the parent
amidino lead PNU-151807 is fully maintained not only
by basic amidino-like compounds, but also by non-basic
cytotoxicity. This may represent an additional explana-
tion for high cytotoxicity of derivatives with three or

P. Cozzi / Il Farmaco 58 (2003) 213Á
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217
amidino derivatives such as amidoxime and cyanoami-
dine and by carboxyamide derivative [32]. The amidino
moiety, due to its strong basic nature, implies the total
protonation at any biological condition, which may play
a key role for DNA binding. The presence of non-basic
moieties in highly active DST derivatives contrasts with
the common opinion that electrostatic interaction be-
tween the cationic moiety and the negatively charged
DNA phosphate residues may represent a main con-
tribution to molecular recognition of DST and DST-like
derivatives.
Several a-bromoacrylic derivatives of DST-like
frames in which one or more pyrrole units were replaced
by other pentaatomic heteroaromatic rings, mainly
imidazole or pyrazole, were synthesized. Some of these
derivatives appear significantly more potent than TAM,
however, even best compounds of this series do not
show a profile of activity better than that of the
corresponding pyrrole analogs.
Extensive antitumor investigation among the whole
series of oligopeptidic a-bromoacrylic derivatives led to
the selection for the clinical development of PNU-
166196, INN brostallicin, an a-bromoacrylamido-tetra-
pyrrole derivative ending with a guanidino moiety (Fig.
5), which is presently undergoing Phase II clinical trials.
Brostallicin shows a broad spectrum of activity,
circumvents resistance to alkylating agents and camp-
tothecins, results more active than TAM in inducing
apoptosis and shows an outstanding favorable cytotoxi-
city/myelotoxicity ratio. In fact its mean IC50 against a
series of tumor cell lines is about eighty times lower than
its IC50 on human CFU-GM hematopoietic progenitors
cells [33]. The dramatic improvement of the cytotoxicity/
myelotoxicity ratio of brostallicin in comparison with
TAM and other MGB represents a determinant feature
to make possible the reaching of effective therapeutic
doses that was not possible in the case of TAM.
presence of GSH, which is the most abundant intracel-
lular thiol, present in the millimolar range in mamma-
lian cells [34].
Our hypothesis was supported by experiments of
interaction of brostallicin and parent PNU-151807
with plasmidic DNA, in the presence or in the absence
of GSH. Agarose gel electrophoresis showed that both
compounds induced the change of plasmid DNA, from
the supercoiled form to the circular form (nicking), only
in the presence of GSH, while in the absence no change
occurred in the plasmid topology, at variance with
TAM. Inactive fluoroacrylic analog of brostallicin was
unable to relax plasmidic supercoiled DNA in the
presence of GSH [35].
Moreover the cytotoxicity of brostallicin and parent
PNU-151807, against melphalan-resistant leukemia
cells, characterized by high levels of GSH, showed a
three-fold increase in comparison to wild L1210 cells,
while the cytotoxicity of TAM was comparable on the
two cell lines. The role of GSH on brostallicin cytotoxi-
city was further confirmed by the fact that inhibition of
GSH formation, with buthionine sulfoximine (BSO), led
to a significant decrease of both cytotoxic and apoptotic
effects on A2780 human ovarian carcinoma cells [35].
Evidence for the role of GSH on brostallicin efficacy has
been obtained also in vivo. Clones expressing high levels
of GSH-S-transferase-p (GST-p), obtained from A2780
cell line after transfection with human GST-p gene,
showed a three-fold increased sensitivity to brostallicin
in comparison to transfected ones with low GST-p
expression. In tumors developed in nude mice after
implantation of GST-p-transfected clones brostallicin
showed greater antitumor activity. Finally recent experi-
ments proved that while the reaction of brostallicin and
GSH alone is negligible within few minutes, the presence
of GSTs, particularly of GST-p significantly enhance the
reaction [36]. These findings suggest that GSH may
affect both the mechanism of brostallicinÁDNA inter-
/
This compound, as the parent PNU-151807, appears
unreactive in DNA in vitro alkylation assays [33]. About
the apparent lack of DNA alkylation we speculated that
an intracellular reactive nucleophilic species, e.g. glu-
tathione (GSH), could perform a first-step Michael-type
attack, which may be followed by a further reaction,
leading to alkylation of DNA nucleophilic functions.
Fig. 6. Thus brostallicin might alkylate DNA only in the
action and antitumor activity, with a potential value in
cancer treatment. In fact high levels of GSH and of
GSTs, the enzymes which catalyze the nucleophilic GSH
reactivity [37], have been reported to play a role in the
resistance of tumor cells to different anticancer drugs,
such as classical mustards and cis-platinum [38]. There
is the evidence moreover that a number of human
tumors display increased levels of GSH and GST-p
isozyme in respect to normal tissues [39].
4. The synthesis of brostallicin and congeners
The syntheses of nitrogen mustards, sulfur mustards
and a-halogenoacrylic DST and DST-like derivatives
cited in this paper were already reported
[11,21,22,25,31]. However, some synthetic pathways
are outlined as representative examples of the syntheses
Fig. 5. Brostallicin (PNU-166196) undergoing Phase II studies.

218
P. Cozzi / Il Farmaco 58 (2003) 213Á220
/
Fig. 6. Putative role of GSH in the interaction of a-bromoacrylic derivatives and DNA.
of DST derivatives. In the case of tri-pyrrole mustards,
which represent the majority of tested mustards, the
synthesis was based upon the coupling of the appro-
priate mustard acid with desformyldistamycin, in turn
easily obtained by acidic hydrolysis of distamycin A.
Most of the mustard derivatives modified at the amidino
group were obtained by the direct reaction of the parent
amidine mustard with the appropriate amine derivative,
Fig. 7. The occurrence of this reaction on amidine
moiety, in good yields, appears noteworthy and under-
lines the mild electrophilic reactivity of our nitrogen
mustard moieties.
of the distamycin A with the appropriate amine
derivative and subsequent hydrolysis. Fig. 7. This path-
way was applied also to the synthesis of analogs in
which the pyrrole ring near the a-halogenoacrylic
moiety was replaced by other azoles. The original
synthesis of brostallicin required, however, an inter-
mediate desformyldistamycin analog bearing a guanidi-
no moiety which was obtained by a step-by-step total
synthesis from guanidinoethylamine, by iterative acyla-
tion with 4-nitro-pyrrolecarboxylic acid chloride and
catalytic reduction of the nitro group Fig. 9.
In the case of tetra-pyrrole a-halogenoacrylic deriva-
tives, which represent the majority of a-halogenoacrylic
tested compounds, the synthesis was based upon the
coupling of the appropriate 4-(a-haloacrylamido)-pyr-
role-2-carboxylic acid with desformyldistamycin or
amidino-modified N-desformyldistamycin analogs, Fig.
8. The latter were obtained in turn by the direct reaction
5. Conclusions
Our studies led to the identification of a new class of
cytotoxic MGB, a-bromoacrylamido DST-like deriva-
tives, endowed with potent antitumor activity and
reduced myelotoxicity. Compounds of this class interact
Fig. 7. A general method of synthesis of distamycin and tallimustine analogs modified at the amidino group. Replacement of the amidine by:
amidoxime, amidrazone, cyanamidine, methylamidines, imidazoline, imidazole.

P. Cozzi / Il Farmaco 58 (2003) 213Á
/220
219
Fig. 8. Synthesis of PNU 151807 as representative example of a-halogenoacrylic derivatives.
with TA-rich sequence of DNA, but at variance with
Acknowledgements
other MGB, appear unreactive in classical in vitro DNA
alkylation assays. There are evidences, however, that
these compounds may be activated in vivo by GSH,
leading to irreversible DNA interaction. Brostallicin
(PNU-166196), now undergoing Phase II clinical trials,
shows potent antitumor activity, very favorable cyto-
toxicity/myelotoxicity ratio, is well tolerated in patients
(Phase I) and, due to its interaction with GSH, may have
a specific role for the treatment of tumors characterized
by constitutive or therapy-induced overexpression of
GSH-GST levels.
I wish to underline the particular contribution to
design, synthesis and mechanism studies of: Nicola
Mongelli, Italo Beria, Marina Caldarelli (Pharmacia,
Global Chemistry), Cristina Geroni (Pharmacia, Exter-
nal Research), PierGiovanni Baraldi, Romeo Romag-
noli (Department of Pharmaceutical Chemistry,
University of Ferrara), Enzio Ragg, Stefania Mazzini
(D.I.S.M.A., University of Milan), Maurizio D’Incalci,
Massimo Broggini (Department of Oncology, Mario
Negri Institute, Milan).
Fig. 9. Synthesis of brostallicin.

220
P. Cozzi / Il Farmaco 58 (2003) 213Á220
/
[20] P.G. Baraldi, R. Romagnoli, G. Spalluto, P. Cozzi, N. Mongelli,
N. Bianchi, R. Gambari, DNA sequence-recognizing properties of
minor groove alkylating agents: effects of the replacement of N-
methylpyrrole by an N-methylimidazole on tallimustine and its
References
[1] R.T. Greenlee, T. Murray, S. Bolden, P.A. Wingo, Cancer
Statistics 2000, CA, Cancer J. Clin. 50 (2000) 7Á
[2] A.P. Polednak, Estimating the prevalence of cancer in the United
States, Cancer 80 (1997) 136Á141.
/
33.
own homologue, Arzneim. -Forsch. 50 (2000) 309Á315.
/
[21] P. Cozzi, I. Beria, G. Biasoli, M. Caldarelli, L. Capolongo, R.
D’Alessio, C. Geroni, S. Mazzini, E. Ragg, C. Rossi, N. Mongelli,
Novel phenyl nitrogen mustard and half-mustard derivatives of
/
[3] E. Negri, C. La Vecchia, A. Decarli, Cancer mortality in Italy,
1997: quantifying the fall in rates in women and men, Tumori 87
distamycin A, Biorg. Med. Chem. Lett. 7 (1997) 2985Á2990.
/
(2001) 290Á
[4] E. Silverberg, Cancer Statistics 2000, CA, Cancer J. Clin. 35
(1985) 19Á35.
/
298.
[22] P. Cozzi, I. Beria, G. Biasoli, M. Caldarelli, C. Capolongo, C.
Geroni, N. Mongelli, Novel phenyl nitrogen mustard and half-
mustard derivatives of amidino-modified distamycin, Bioorg.
/
[5] H.G. Welch, L.M. Schwartz, S. Woloshin, Are increasing 5-year
Med. Chem. Lett. 7 (1997) 2979Á
[23] P. Cozzi, N. Mongelli, Cytotoxics derived from distamycin and
congeners, Curr. Pharm. Des. 4 (1998) 181Á201.
[24] P. Cozzi, Distamycin derivatives as potential anticancer agents,
Idrugs 4 (2001) 573Á581.
/
2984.
survival rates evidence of success against cancer, J. Am. Med.
Assoc. 284 (2000) 2053Á2055.
[6] A.M. Casazza, A. Fioretti, M. Ghione, M. Soldati, M. Verini, A.
/
/
Distamycin, A new antiviral antibiotic, Antimic. Agents Che-
/
mother. 5 (1965) 593Á
[7] B.C. Baguley, Non-intercalative DNA-binding antitumor com-
pounds, Mol. Cell. Biochem. 43 (1982) 167Á181.
/
598.
[25] P. Cozzi, I. Beria, M. Caldarelli, L. Capolongo, C. Geroni, S.
Mazzini, E. Ragg, Phenyl sulfur mustard derivatives of distamy-
/
cin A, Biorg. Med. Chem. Lett. 10 (2000) 1653Á1656.
/
[8] M. Broggini, M. D’Incalci, Modulation of transcription factor-
[26] M. Brendel, A. Ruhland, Relationships between functionality and
genetic toxicology of selected DNA-damaging agents, Mutat. Res.
DNA interaction by anticancer drugs, Anti-cancer Drug Des. 9
(1994) 373Á387.
/
133 (1984) 51Á85.
/
[9] G. Pezzoni, M. Grandi, G. Biasoli, L. Capolongo, D. Ballinari,
F.C. Giuliani, B. Barbieri, A. Pastori, E. Pesenti, N. Mongelli, F.
Spreafico, Biological profile of FCE-24517, a novel benzoyl
mustard analogue of distamycin A, Br. J. Cancer 64 (1991)
[27] A. Sunters, C.J. Springer, K.D. Bagshawe, R.L. Souhami, J.A.
Hartley, The cytotoxicity, DNA crosslinking ability and DNA
sequence selectivity of the aniline mustards melphalan, chloram-
bucil and 4-[bis(2-chloroethyl)amino]benzoic acid, Biochem.
1047Á1050.
/
Pharmacol. 44 (1992) 59Á64.
/
[10] M. D’Incalci, C. Sessa, DNA minor groove binding ligands: a new
[28] S. Marchini, M. Ciro`, F. Gallinari, C. Geroni, P. Cozzi, M.
D’Incalci, M. Broggini, a-Bromo-acryloyl derivative of distamy-
cin A (PNU 15): a new non-covalent minor groove DNA binder
class of anticancer agents, Expert. Opin. Invest. Drugs 6 (1997)
875Á884.
/
[11] R. D’Alessio, C. Geroni, G. Biasoli, E. Pesenti, M. Grandi, N.
Mongelli, Structure-activity relationship of novel distamycin A
derivatives: synthesis and antitumor activity, Bioorg. Med. Chem.
with antineoplastic activity, Br. J. Cancer 80 (1999) (1807) 991Á
/
997.
[29] P. Cozzi, I. Beria, M. Caldarelli, L. Capolongo, C. Geroni, N.
Mongelli, Cytotoxic a-halogenoacrylic derivatives of distamycin
Lett. 4 (1994) 1467Á
[12] W.A. Denny, DNA minor groove alkylating agents, Curr. Med.
Chem. 8 (2001) 533Á544.
/1472.
A, Bioorg. Med. Chem. Lett. 10 (2000) 1269Á1272.
/
/
[30] M. Ghielmini, G. Bosshard, L. Capolongo, C. Geroni, E. Pesenti,
V. Torri, M. D’Incalci, F. Cavalli, C. Sessa, Estimation of the
haematological toxicity of minor groove alkylators using tests on
[13] M. Ghielmini, G. Bosshard, L. Capolongo, C. Geroni, E. Pesenti,
V. Torri, M. D’Incalci, F. Cavalli, C. Sessa, Estimation of the
haematological toxicity of minor groove alkylators using tests on
human cord blood cells, Br. J. Cancer 75 (1997) 878Á
[31] P.B. Dervan, Design of sequence-specific DNA-binding mole-
cules, Science 232 (1986) 464Á471.
/883.
human cord blood cells, Brit. J. Cancer 75 (1997) 878Á883.
/
[14] H.H. Lee, M. Boyd, G.L. Gravatt, W.A. Denny, Pyrazole
analogues of the bispyrrolecarboxamide anti-tumor antibiotics:
synthesis, DNA binding and antitumor properties, Anti-Cancer
/
[32] P. Cozzi, I. Beria, M. Caldarelli, C. Geroni, N. Mongelli, G.
Pennella, Cytotoxic a-bromoacrylic derivatives of distamycin
analogues modified at the amidino moiety, Bioorg. Med. Chem.
Drug Des. 6 (1991) 501Á517.
/
[15] M. Lee, A.L. Rhodes, M.D. Wyatt, S. Forrow, J.A. Hartley,
Design, synthesis and biological evaluation of DNA sequence and
minor groove selective alkylating agents, Anti-Cancer Drug Des.
Lett. 10 (2000) 1273Á1276.
/
[33] C. Geroni, G. Pennella, C. Capolongo, D. Moneta, R. Rossi, M.
Farao, S. Marchini, P. Cozzi, Antitumor activity of PNU 166196,
a novel DNA minor groove binder selected for clinical develop-
ment, Proc. AACR Ann. Meeting 41 (2000) 425 (Abstract 2708).
[34] A. Meister, M.E. Anderson, Glutathione, Ann. Rev. Biochem. 52
8 (1993) 173Á192.
/
[16] M.D. Wyatt, B.J. Garbiras, M.K. Haskel, M. Lee, R.L. Souhami,
J.A. Hartley, Structure-activity relationship of a series of nitrogen
mustard- and pyrrole-containing minor groove binding agents
(1983) 711Á760.
/
related to distamycin, Anti-Cancer Drug Des. 9 (1994) 511Á525.
/
[35] P. Cozzi, A new class of cytotoxic DNA minor groove binders: a-
halogenoacrylic derivatives of pyrrolecarbamoyl oligomers,
[17] P.G. Baraldi, I. Beria, B. Cacciari, L. Capolongo, P. Cozzi, N.
Mongelli, R. Romagnoli, G. Spalluto, Structure-activity relation-
ship of novel tallimustine derivatives: synthesis and antitumor
Farmaco 56 (2001) 57Á65.
/
[36] C. Geroni, S. Marchini, P. Cozzi, E. Galliera, E. Ragg, T.
Colombo, R. Battaglia, M. Howard, M. D’Incalci, M. Broggini,
Brostallicin, a novel anticancer agent whose activity is enhanced
activity, Bioorg. Med. Chem. Lett. 6 (1996) 1247Á1252.
/
[18] P.G. Baraldi, I. Beria, B. Cacciari, L. Capolongo, P. Cozzi, C.
Franzetti, N. Mongelli, R. Romagnoli, G. Spalluto, Synthesis and
antitumor activity of novel distamycin derivatives, Bioorg. Med.
upon binding to glutathione, Cancer Res. 62 (2002) 2332Á
[37] A.E. Salinas, M.G. Wong, Glutathione S-transferases: a review,
Curr. Med. Chem. 6 (1999) 279Á309.
[38] K.D. Tew, Glutathione-associated enzymes in anticancer drug
resistance, Cancer Res. 54 (1994) 4313Á4320.
[39] S. Tsuchida, K. Sato, Glutathione transferases and cancer, Crit.
Rev. Biochem. Mol. Biol. 27 (1992) 337Á384.
/
2336.
Chem. Lett. 6 (1996) 1241Á1246.
/
/
[19] P.G. Baraldi, P. Cozzi, C. Geroni, N. Mongelli, R. Romagnoli, G.
Spalluto, Novel benzoyl nitrogen mustard derivatives of pyrazole
analogues of distamycin A: synthesis and antileukemic activity,
/
Bioorg. Med. Chem. 7 (1999) 251Á
/262.
/