Biochemical Mechanisms of Cisplatin Cytotoxicity

Anti-Cancer Agents in Medicinal Chemistry, 2007, 7, 3-18 3

1871-5206/07 $50.00+.00 © 2007 Bentham Science Publishers Ltd.

Biochemical Mechanisms of Cisplatin Cytotoxicity

Victoria Cepeda1, Miguel A. Fuertes1, Josefina Castilla2, Carlos Alonso1, Celia Quevedo3 andJose M. Pérez3,*

1Centro de Biología Molecular “Severo Ochoa”(CSIC-UAM), Facultad de Ciencias, Universidad Autónoma de Madrid,28049-Madrid, Spain. 2Farmacia Castilla, Hermanos García Noblejas 174, 28037-Madrid, Spain and 3Departamentode Química Inorgánica, Facultad de Ciencias, Universidad Autónoma de Madrid, 28049-Madrid, Spain

Abstract: Since the discovery by Rosenberg and collaborators of the antitumor activity of cisplatin 35 years ago, threeplatinum antitumor drugs (cisplatin, carboplatin and oxaliplatin) have enjoyed a huge clinical and commercial hit. Eversince the initial discovery of the anticancer activity of cisplatin, major efforts have been devoted to elucidate the bio-chemical mechanisms of antitumor activity of cisplatin in order to be able to rationally design novel platinum based drugswith superior pharmacological profiles. In this report we attempt to provide a current picture of the known facts pertainingto the mechanism of action of the drug, including those involved in drug uptake, DNA damage signals transduction, andcell death through apoptosis or necrosis. A deep knowledge of the biochemical mechanisms, which are triggered in thetumor cell in response to cisplatin injury not only may lead to the design of more efficient platinum antitumor drugs butalso may provide new therapeutic strategies based on the biochemical modulation of cisplatin activity.

Key Words: Cisplatin, cancer, DNA, apoptosis, necrosis.

1. INTRODUCTION

The great impact in the treatment of cancer of the plati-num coordination complex cisplatin, [cis-diamminedichloroplatinum(II)] or cis-DDP, is a paradigm within the use ofmetals in medicine. In fact, cisplatin and its analogue car-boplatin, [cis-diammine-1,1-cyclobutanedicarboxylate plati-num(II)] are among the most commonly used antitumordrugs today.

The biological activity of cisplatin, was discovered ser-endipitously about 125 years after the initial report of itssynthesis and characterization. Although the synthesis andcharacterization of cisplatin was first reported by Peyrone in1845 [1], its anticancer properties remained unnoticed untilthe mid-1960s, when Rosenberg and co-workers studied theeffects of electric fields on Escherichia coli growth [2]. Theplatinum electrodes released by redox reactions some plati-num complexes which provoked complete stop of cell divi-sion in the bacterial rods. Amongst the platinum complexesformed, cisplatin was identified as the main antiproliferativeagent [3]. In the next years cisplatin developed into one ofthe most widely used drugs in cancer chemotherapy.

Cisplatin is highly effective in the treatment of testicularand ovarian cancers and is also widely employed for treatingbladder, cervical, head and neck, oesophageal and small celllung cancer [4]. Despite its success, cisplatin has severaldisadvantages, which include severe side effects includingnephrotoxicity, neurotoxicity, ototoxicity, nausea and vom-iting [4]. These toxic effects limit the dose that can be ap-plied to patients. Cisplatin also has limited aqueous solubil-ity and is administered intravenously, a major inconveniencefor outpatient treatment [5]. Although cisplatin is extensively

*Address correspondence to this author at the Departmento de QuímicaInorgánica, Facultad de Ciencias, Universidad Autónoma de Madrid, 28949-Spain; Tel: +34914978 454; Fax: 34913274457;E-mail: [email protected]

used in clinic, its applicability is still limited to a relativelynarrow range of tumour types. Some tumours such as colo-rectal and non-small cell lung cancers have intrinsic resis-tance to cisplatin, while others such as ovarian or small celllung cancer develop acquired resistance after the initialtreatment [6].

Since the introduction of cisplatin in standard clinicalprotocols, numerous laboratory studies have provided con-siderable information on the molecular mechanisms of anti-tumor action of the drug [6]. It has emerged from thesestudies, that the biochemical mechanisms of cisplatin cyto-toxicity involve the binding of the drug to DNA and non-DNA targets and further induction of cell death throughapoptosis, necrosis or both within the heterogeneous popula-tion of cells that forms a tumoral mass [7].

Tumor resistance to cis-DDP coupled with cisplatin tox-icity have stimulated the search for other antitumor-activeplatinum complexes with improved pharmacological proper-ties. The only registered platinum drug that has consistentlydemonstrated antitumor activity against cisplatin resistanttumors such as colorectal cancers is oxaliplatin, [trans-L-1,2-diaminocyclohexaneoxalatoplatinum(II)] [7]. Fig. (1) depictsthe structures of the three platinum antitumor drugs currentlyapproved for clinical use by both the American Food andDrug administration (FDA) and the European Agency for theEvaluation of Medicinal Products (EMEA) as well as thestructure of the inactive isomer of cisplatin, transplatin(trans-DDP).

Although the large number of biochemical studies carriedout on cis-DDP activity have not clearly established yet themolecular bases of tumor resistance to this drug in any typeof cell, at least they have identified several mechanisms thatcan contribute to this process. As shown in Fig. (2), resis-tance to cisplatin is generally considered a multifactorialphenomenon, which is mainly due to (i) reduced drug accu-mulation, (ii) inactivation by thiol containing species, (iii)

4 Anti-Cancer Agents in Medicinal Chemistry, 2007, Vol. 7, No. 1 Cepeda et al.

increased repair of platinum-DNA adducts, and (iv) increaseof cis-DDP adducts tolerance and failure of cell death path-ways. These resistance mechanisms are cell line-dependentso that a particular tumor may exhibit one, two or even allthe above-mentioned mechanisms [8, 9].

An alternative way to circumvent cisplatin resistance andto enhance its anticancer activity is the use of biochemicalmodulation strategies. This new approach involves the com-bination of cisplatin with drugs that interfere with specificcisplatin-resistance pathways. So far, the biochemical

Fig. (1). The three FDA and EMEA approved platinum anticancer drugs: (A) cisplatin, (B) carboplatin, (C) oxaliplatin and (D) biologicallyinactive transplatin.

Fig. (2). Schematic drawing of the main biochemical mechanisms of resistance to cis-DDP. Resistance mechanisms may act prior of aftercisplatin binding to DNA. MMR = mismatch repair, NER = nucleotide excision repair, GSH = glutathione.

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Biochemical Mechanisms of Cisplatin Cytotoxicity Anti-Cancer Agents in Medicinal Chemistry, 2007, Vol. 7, No. 1 5

mechanisms of modulation of cisplatin-resistance by an in-creasing number of drugs have been identified [6].

2. BIOCHEMICAL MECHANISMS OF ACTION OFCISPLATIN

It is generally accepted that binding of cis-DDP to ge-nomic DNA (gDNA) in the cell nucleus is the main eventresponsible for its antitumor properties [10]. Thus, the dam-age induced upon binding of cisplatin to gDNA may inhibittranscription, and/or DNA replication mechanisms. Subse-quently, these alterations in DNA processing would triggercytotoxic processes that lead to cancer cell death. Fig. (3)shows the main molecular events in which cisplatin is in-volved before reaching its ultimate target gDNA.

2.1. Drug Accumulation

Once cis-DDP has been intravenously administered to thepatient, it rapidly diffuses into tissues and is highly bound toplasma proteins. Binding of cis-DDP to plasma proteins is aresult of the strong reactivity of platinum against sulphur ofthiol groups of amino acids such as cysteine. Hence, near 90% of the platinum in the blood is bound to albumin and otherplasma proteins leading to inactivation of a great amount ofcisplatin molecules [11]. Loss of the chloride groups from

the cisplatin molecule is required before it binds to gDNA.Outside the cell, chloride concentration is around 100 mM.However, within the cell, chloride concentrations range be-tween 2 and 30 mM and cisplatin aquation occurs. Conse-quently, water molecules replace one or both chloride leav-ing groups. The result is the formation of the [Pt(H2O)Cl(NH3)2]

+ and [Pt(H2O)2(NH3)2]2+ cations. These mono and

diaquo species of cisplatin are very reactive towards nucleo-phile centres of biomolecules because H2O is a much betterleaving group than Cl- [12, 13].

The biochemical mechanism by which cisplatin crossesthe cell membrane still remains unclear. Early studies re-ported that cis-DDP uptake was not inhibited by their struc-tural analogues and this entry into the cell did not seem to bedependent on an optimum pH [14-16]. In addition, cis-DDPaccumulation in the cell proceeded linearly with time over 60minutes and did not reach a plateau up to a drug concentra-tion of 1 mM. So, it was suggested that passive diffusion wasthe main mechanism by which cisplatin enters the cell. How-ever, it was found later that a certain degree of cis-DDP up-take seemed to be energy dependent and could be modulatedby pharmacological agents such as the Na+/K+-ATPase in-hibitor ouabain and the membrane-interactive agents am-photericin B and digitonin [17]. Moreover, recent observa-tions point towards a direct connection between the cellular

Fig. (3). Schematic depiction of cisplatin uptake and efflux and binding to nuclear DNA. In addition to passive diffusion, cisplatin is alsoactively uptaked through the copper transporter CTR1. Copper-transporting P-type adenosine triphosphate (ATP7B) is involved in cisplatinefflux. Both copper and cis-DDP show the ability to decrease the import of the other and can induce the degradation and delocalization ofCTR1. Once cis-DDP is in the cytoplasm, it may be inactivated by gluthathione (GSH) or may bind to its main cellular target, nuclear DNA.


concentrations of copper and platinum, which leads to pro-pose an active transport for cisplatin in addition to passivediffusion. Thus, experiments with the yeast protein Ctr1 (ahigh-affinity copper transporter) and cis-DDP, revealed thatmutation or deletion of the CTR1 gene leads to stronger cis-platin resistance and to intracellular reduction of platinumlevels in yeast and in mouse cells [18]. Interestingly, cis-DDP accumulation in cells became increased when humanCTR1 gene was overexpressed, although the ability of thedrug to access its cytotoxic targets was unaltered [19]. Onthe other hand, it has been found that copper-transporting P-type adenosine triphosphate (ATP7B), which regulates cop-per homeostasis in the cell, has a role in cisplatin efflux andis associated with cisplatin resistance in vitro [20], and invarious cancers [21-24]. Hence, expression of ATP7B in hu-man carcinoma cells modulates sensitivity to cis-DDP andcopper through a more efficient efflux of the two agents [19].Another exporter protein implicated in cisplatin resistancethrough drug efflux is the ATP-binding cassette, sub-familyC 2 (ABCC2 also known as MRP2 or cMOAT) [25-27].Altogether these data indicate that there is a connected trans-port for copper and cisplatin because both copper and cis-platin (i) can reduce the uptake of each other, (i) trigger thedegradation and delocalization of CTR1 17, and (iii) alsoshow bidirectional cross-resistance [28-30].

2.2. Binding to Non-DNA Targets

When cis-DDP passes trough the cell membrane, the cis-Pt(II) center may coordinate to some constituents of thelipidic bilayer, which contain nitrogen and sulphur atomsincluding phospholipids and phosphatidylserine [31]. In thecytoplasm many cellular components that have soft nucleo-philic sites such as cytoskeletal microfilaments, thiol-con-taining peptides and proteins and RNA, may react with cis-platin [32]. Hence, of interest is the observation that only 5-10% of covalently bound cell-associated cisplatin is found inthe gDNA fraction, whereas 75-85% of the drug binds to

proteins and other cellular constituents [33, 34]. The mostimportant non-DNA target of cisplatin is probably thetripeptide glutathione (GSH) which is present in cells at highconcentrations (0.5-10 mM). GSH and other thiol-containingbiomolecules such as metalothioneins (MT) bind quickly toplatinum [6]. Cis-DDP binding to GSH and MT has primar-ily been associated with negative pharmacological proper-ties, including the development of resistance and toxicity. Onthe other hand, cis-DDP may alter the activity of enzymes,receptors, and other proteins through coordination to sulfuratoms of cysteine and/or methionine residues and to nitrogenatoms of histidine residues. In fact, binding of cis-DDP tomethionine 1 (met1) and/or histidine 68 (his68) of ubiquitinmay inhibit the ubiquitin-proteasome pathway of selectivedegradation of cellular proteins, which ends up in cytotoxicevents [35]. In addition, it has been found that cis-DDP, be-sides inhibiting in vitro the activity of heat shock protein 90(Hsp90), an ATP-binding chaperone, efficiently and specifi-cally blocks its C-terminal ATP binding site. The C-terminalof Hsp90 via its ATP hydrolytic function is involved in thecorrect folding of proteins which play a role in signal trans-duction and cell cycle regulation [36].

2.3. Binding to DNA

Experimental data accumulated over the last 35 yearsindicates that gDNA is the main cellular target of cis-DDP[12]. The N7 atoms of guanine and adenine located in themajor groove of the double helix are the most accessible andreactive nucleophilic sites for platinum coordination to DNAdue to both their high nucleophilicity and accessibility. Asshown in Fig. (4), cis-DDP may form on DNA variousstructurally different adducts. The binding kinetics of cis-DDP to DNA begins with the formation of monofunctionalcis-[Pt(NH3)2(H2O)]:DNA adducts, but most cis-Pt(II) cen-ters react further to produce cis-[Pt(NH3)2]:DNA bifunc-tional adducts called interstrand or intrastrand cross-links,which then block replication and/or prevent transcription

Fig. (4). Main adducts formed after binding of cis-DDP to DNA. (A) 1,2-intrastrand cross-link, (B) interstrand cross-link, (C) monofunc-tional adduct, and (D) protein-DNA cross-link. The main site of attack of cis-DDP to DNA (N7 of guanine) is shown in the central panel.


[37]. Hence, 60-65% of the adducts formed by cis-DDP are1,2-d(GpG) intrastrand cross-links and 20-25% d(ApG) in-trastrand cross-links. It has been recently shown that the mi-nor 1,3-d(GpTpG) intrastrand cross-link could be also im-portant in the mechanism of action of cis-DDP. In fact, theminor 1,3-d(GpTpG) intrastrand cross-link adduct induces innucleosomes an asymmetric arrangement of the DNA in re-lation to the histone octamer, which affects the translationalpositioning of the DNA.

These phasing phenomenon may be crucial to the recog-nition and processing of platinum-DNA adducts in cancercells [38]. DNA-protein cross-links are also formed by cis-DDP and other cisplatin analogues [6]. On the other hand, ofparticulat interest to medicinal chemists and biochemicalpharmacologists was the initial observation that the trans iso-mer of cisplatin, trans-diamminedichloroplatinum(II) or trans-DDP, see Fig.(1), shows low in vitro cytotoxic activity andlacks in vivo antitumor activity [39]. However, both cis- andtrans-DDP bind to DNA almost exclusively by coordination tothe N7 nitrogen atom of purine bases, with a higher affinity forguanine over adenine [40, 41]. Therefore, from a biochemicalpoint of view the lack of pharmacological activity of trans-DDP or transplatin must be related to the different type ofDNA adducts formed by this trans isomer relative to cis-DDP.Thus, due to steric reasons, trans-DDP is unable to form 1,2-intrastrand adducts between two adjacent purines in the sameDNA strand. In fact, transplatin mainly forms 1,3-intrastrandand interstrand cross-links [42].

It is generally assumed that the 1,2-intrastrand DNA ad-duct, which bends the double helix towards the major grooveis responsible for cisplatin cytotoxicity, although this is not aproven fact (see “(Fig.5)”). This hypothesis is based on thediscovery that some HMG (high mobility group) proteinsspecifically recognize this type of DNA adduct and therefore

may regulate the processing of 1,2-intrastrand cross-linksformed by cisplatin [43]. Further support to this assumptioncame from the finding that nucleotide excision repair (NER),an important DNA-repair system, repairs 1,3-intrastrand ad-ducts with higher efficiency than the 1,2-intrastrand adducts[44]. However, it should be pointed out that other adducts,such as DNA interstrand cross-links and DNA-protein cross-links might also contribute to the mechanism of cytotoxicactivity of cisplatin [10].

3. MOLECULAR PHARMACOLOGY OF CISPLATINCYTOTOXICITY

The specific mechanisms that trigger cytotoxic pathwaysin response to cis-DDP injury mainly include ways to detectas well as to determine whether cisplatin-induced DNAdamage is strong enough to be lethal. In certain cases, how-ever, direct damage of cisplatin to proteins or other bio-molecules might trigger cytotoxicity mechanisms.

Research in this field has been focused on DNA damagerecognition proteins that recognize the DNA bending in-duced by specific cisplatin adducts [10, 12]. So, these pro-teins recognize the kink in DNA induced by cisplatin 1,2-intrastrand adduct and include the XPA-RPA complex, theTATA-box binding protein TBP, hMSH2, and nonhistoneHMG1 and HMG2. Hence, it has been hypothesized that therecognition of 1,2-intrastrand adduct by these proteins maybe the first step towards the initiation of apoptosis or pro-grammed cell death (PCD). However, as mentioned before,most cisplatin molecules bind to proteins rather than DNA sothat there is experimental evidence that the former type ofcisplatin damage also plays a role in triggering PCD path-ways [10, 45]. Moreover, necrosis or accidental cell deathdue to general cell machinery failure is also reported as acell-killing mechanism for cisplatin [10, 46].

Fig. (5). A depiction of the effect of the major 1,2-d(GpG) intrastrand cross-link in a double-stranded oligonucleotide. The bending of theDNA backbone towards the major grove is clearly perceptible.


3.1. Repair of Cisplatin-DNA Adducts

Cisplatin-DNA adducts may be repaired by several typesof proteins. The most important ones are (i) nucleotide exci-sion repair (NER) proteins, (ii) mismatch repair (MMR)proteins and (iii) DNA-dependent protein kinase (DNA-PK)protein.

3.1.1. NER Proteins

Nucleotide excision repair (NER) appears to be a majormechanism of cisplatin resistance. NER is an ATP-depen-dent multiprotein complex that recognizes the bending in-duced on DNA by 1,2-intrastrand cross-links, and subse-quently excises the part of the DNA that includes the kink as27- to 29-base-pair oligonucleotides. The gap that remains isthen filled by DNA polymerase [47]. An schematic represen-tation of removal of 1,2-intrastrand cross-links by NER isshown in Fig. (6).

It has been reported that at least 16 genes are essential forthe DNA damage recognition and excision function of theintrastrand adducts formed by cis-DDP between to adjacentguanines [48].

3.1.2. MMR Proteins

Mistmatch repair is a post-replication repair system thatcorrects unpaired or mispared nucleotides. The mismatchrepair system involves at least five proteins (MLH1, MSH2,MSH3, MSH6 and PMS2) and functions as a repair mecha-nism that needs ATP [49]. The relationship between DNAdamage recognition by MMR proteins and cytotoxicity re-mains undefined yet. However, it has been proposed that theMMR proteins would try to insert the “correct” nucleotide onthe non-damaged strand opposite to the intrastrand adductbetween to adjacent guanines and that this “futile” repaircycle might then induce cell death pathways [12]. It is inter-esting to point out that, at least in ovarian and colon cancers,MMR is a comparative small contributor to the cis-DDPresistance phenotype in comparison with NER. In fact inthese two types of tumors an intact MMR system appears tobe essential for the linkage of DNA damage/repair with theinitiation of apoptosis [50].

3.1.3. DNA-PK Protein

The DNA-PK protein mainly repairs DNA double-strandbreaks (DSB) induced by cell exposure to ionizing radiation.

However, it has been described that the DNA-PK protein canalso interact with cisplatin-DNA lesions [51]. DNA bindingof the Ku subunits of DNA-PK in vitro is essential to acti-vate the kinase activity of DNA-PK to phosphorilate itself orother transcription factors. Interestingly, it has been reportedthat in ovarian cancer cells the presence of cisplatin-DNAadducts serves to inhibit the ability of the Ku subunits ofDNA-PK to translocate on a duplex DNA substrate so thatkinase activity is inhibited [52].

3.2. Mechanisms of Cytotoxicity Associated with HMGProteins

Several HMG-domain proteins, which belong to thehigh-mobility group of proteins recognize and bind to cis-platin-modified DNA. The greatest attention has been con-centrated on the studies of recognition of platinated DNA byHMGB1 and HMGB2 proteins, which belong to architec-tural chromatin proteins. In contrast to NER proteins,HMGB1 is able to inhibit the repair of the major 1,2-d(GpG)intrastrand cisplatin-DNA adduct by human excision nucle-ase in vitro, but not the adducts formed by transplatin rein-forcing the hypothesis that 1,2-intrastrand cross-links are themajor DNA adducts involved in the mechanism of action ofcisplatin [12]. Several mechanisms have been proposed forhow HMG domain proteins might modulate the sensitivity ofcells to cis-DDP but two of them seem to be the most feasibleones. The “repair shielding model” postulates that HMG pro-teins could protect cisplatin-DNA adducts from recognition byDNA repair enzymes [53]. The second one, the so-called “hi-jacking model” establishes that HMG proteins could modulatecell cycle events subsequent to DNA damage and trigger celldeath. Thus, this latter model postulates that the recognition byHMG cellular factors of cisplatin-DNA lesions would deviatethem from their natural binding sites resulting in inhibition ofvital cellular functions [54].

As shown in Fig. (7), HGMB1 recognizes the major 1,2-d(GpG) intrastrand cisplatin-DNA adduct and may avoid thatNER can repair this DNA lesion. In addition, HMGB1 hasbeen linked to other DNA-dependent pathways. Hence,HMGB1 (i) may activate cleavage of recombination activa-tion genes 1 and 2 (RAG1 and RAG2), (ii) may stimulate thebinding of sequence-specific transcription factors, (iii) mayinteract with the MMR protein MutSα then having a possiblerole in mismatch repair, (iv) binds to the tumor suppressor

Fig. (6). Scheme of the steps involved in the repair of the major cisplatin 1,2-intrastrand crosslink by the nucleotide excision repair system(NER).


protein p53 in vitro therefore increasing p53 DNA-bindingactivity, and (v) HMGB1 affinity for platinated DNA is sig-nificantly improved in the presence of p53 [55]. However, itshould be pointed out that all these biochemical functionsattributed to HMGB1 in response to cisplatin damage toDNA are dependent on the cell type. In fact, it has been re-ported that when both the growth inhibition by cisplatin andcisplatin-induced cell death were analyzed in HMGB1knockout mouse embryonic fibroblasts and its wild-typecounterpart no significant differences were observed in theresponses of these cells to cisplatin [56].

3.3. Mechanisms of Cytotoxicity Associated with Non-HMG Proteins

Besides HMG-domain family of proteins, many otherimportant nuclear proteins such as the TATA-binding protein(TBP) which is involved in transcription initiation also rec-ognize the major 1,2-d(GpG) intrastrand cisplatin-DNA ad-duct [57]. Most of the transcribed genes in mammalian cellsuse a TATA box within the promotor region for initiation oftranscription. Therefore, TBP could be efficiently removedfrom its natural binding site by 1,2 intrastrand adducts sur-rounding the TATA box, resulting in additional pathways by

which cis-DDP induces cell death [58]. Such a process isknown as “the transcription factor hijacking hypothesis”.

It has also been reported that histone linker protein H1binds to DNA platinated by cis-DDP but not by trans-DDP[59]. H1 histone is an important nuclear factor that wrapsand maintains the integrity of DNA-linking nucleosome coreparticles. Consequently, binding of cisplatin to H1 histonemight initiate cytotoxic processes and eventually lead to celldeath.

3-methyladenine DNA glycosylase (AAG) mammalianrepair proteins are also able to recognize cisplatin-DNA 1,2-intrastrand adducts [60]. Binding of AAG is favored by tyro-sine intercalation, an interaction that the enzyme exhibitswith other substrates. Hence, by analogy to “transcriptionfactor hijacking mechanisms”, cis-DDP may also be in-volved in the so-called “repair factor hijacking pathways”,preventing binding of AAG to its natural substrates and,therefore, leaving other DNA lesions to initiate cell death.This hypothesis has been proposed to explain the synergisticeffect shown by cis-DDP when is administered to patients incombination with the alkylating agent 1,3-bis-(2-chloroethyl)-1-nitrosourea (BCNU) [61].

Fig. (7). Recognition of the major cisplatin 1,2-d(GpG) intrastrand cross-link by HMGB1 (high mobility group box 1) protein and its possi-ble role in adduct processing and interaction with cellular regulatory proteins. HMGB1 recognizes the DNA bending induced by cisplatin 1,2d(GpG) adduct and may regulate the efficiency of nucleotide excision repair (NER) in vitro. In addition, HMGB1 may interact with RAG1/2,p53 and MutSα. HMGB1 has been associated to mismatch repair (MMR), V(D)J recombination, p53 and MAPK pathways. HMGB1 may bealso involved in nucleosome remodelling and may function as a cytokine, being secreted by necrotic and immune cells. ACF, ATP-usingchromatin assembly and remodelling factor; MAPK, mitogen-activated protein kinase; NF-κB, nuclear factor-κB; RAG, recombination acti-vating genes.


3.4. Signals Transduction from Cisplatin-DNA Damage

3.4.1. AKT Pathway

AKT (a member of the family of phosphatidylinositol 3-OH-kinase-regulated serine/threonine kinases) protects cellsfrom apoptotic death induced by several stimuli, includingcis-DDP damage to DNA, and promotes cell survivalthrough the modulation of several downstream target pro-teins. AKT stabilizes the X-linked inhibitor of apoptosis(XIAP) by inhibiting its auto- and cisplatin-induced- ubiq-uitination activities. It has been reported that low cellularlevels of XIAP and/or AKT expression or function are asso-ciated with circumvention of cis-DDP chemoresistance inovarian cancer cells that express either endogenous or recon-stituted wild-type p53 [62-64], whereas high levels of XIAPare associated with decreased cisplatin-induced apoptosis.On the other hand, AKT may also activate nuclear factor-_B(NF-_B), which has a key role in cell survival. Thus, highNF-κB activity indicates decreased apoptosis so that inhibi-tion of NF-κB enhances the effect of cis-DDP [30, 65, 66].

3.4.2. c-ABL Pathway

c-ABL is a member of the SRC family of non-receptortyrosine kinases. Cisplatin and other DNA-damaging agentscan stimulate c-ABL so that it may transmit the DNA-damage signal from the nucleus to the cytoplasm [67]. It hasbeen found that this protein has a role in cisplatin-activatedapoptosis because overexpression of c-ABL tyrosine kinasein SaOS-2 cells and NIH3T3 cells activates apoptosis [68,69], whereas c-ABL-deficient mouse fibroblasts are resistantto cisplatin-induced apoptosis and reintroduction of c-ABLfunction in these cells restores their sensitivity to the drug[70]. In addition, retinoblastoma tumour-suppressor protein(RB) is able to bind to c-ABL, blocking its tyrosine kinaseactivity and avoiding its activation by DNA-damage signals.The RB–c-ABL interaction can be disrupted by the phos-phorylation of RB so that c-ABL is released in the nucleus.On the other hand, c-ABL may phosphorilate downstreamtranscription factors p73 and p53, although it is not essentialfor the induction and activation of p53 by cisplatin damageto DNA [55]. In wild-type fibroblasts treated with cis-DDP,c-ABL phosphorilates downstream MAPK/ERK kinasekinase 1 (MEKK1) and the stress-activated kinases JNK andp38 MAPK [71-73]. In addition, c-ABL deficient cells fail toactivate p38 MAPK and JNK after cis-DDP administration,but not after exposure to ultraviolet irradiation. These cellsrecover p38 MAPK and JNK activation ability when resto-ration of c-ABL is produced, which shows both that p38MAPK and JNK c-ABL-dependent activation is specific forcis-DDP and that p38 MAPK and JNK are regulated in adifferent way in response to diverse classes of DNA-damag-ing agents [74]. Interestingly, c-ABL contains three HMG-like boxes that bind preferably to A/T duplexes and distortedDNA structures. This structural feature suggests that c-ABLcould recognize and interact with cisplatin–DNA adductsjust the same way as HMGB1 [75, 76]. However, it has beenalso reported that c-ABL has unique DNA binding propertiesas compared with HMGB1 [30]. So, whether c-ABL couldrecognize and interact with the major 1,2-d(GpG) intrastrandcisplatin-DNA adduct like HMGB1 is at present an openquestion.

3.4.3. p53 Protein

The tumor suppressor gene p53 facilitates DNA repairbefore DNA replication. As shown in Fig. (8), p53 protein isa transcription factor that is considered a “guardian of thegenome” because it activates a host of other genes (p21/Waf1, MDM2, GADD45 and so on) that lead to cell cyclearrest and DNA repair [77].

Interaction between p53 and HMGB1 after DNA platina-tion might provide a molecular link between cisplatin DNAdamage and p53-mediated apoptosis because in this eventp53 could not activate DNA repair. The above-mentionedmechanisms of p53 response to cisplatin-induced DNA dam-age are not necessarily exclusive and could work in concert.In particular, cis-DDP cytotoxicity might be determined by adynamic competition among proteins that repair DNA andproteins that interfere with DNA repair and trigger cell death[55].

p53 protein is mutated in more than 50% of all humantumors and, therefore, plays a central role in chemotherapy-induced apoptosis. Several studies have shown that sensitiv-ity to cis-DDP usually correlates with the presence of wildtype p53, while the lack of functional p53 is related to cis-DDP resistance [78]. However, a clear relationship betweenp53 cellular status and cisplatin-induced cytotoxicity has notbeen found yet. In fact, the cell type and cellular contextexert a strong influence in p53-mediated responses to cis-platin DNA damage [79].

3.4.4. MAPK/JNK/ERK Pathways

MAP kinases protein family (MAPK) includes threemain mammalian subfamilies: ERK, JNK (also known asstress-activated protein kinase) and p38 MAPK. These pro-teins play key roles in signal transduction pathways in re-sponse to different extracellular stimuli. JNK and p38 arestrongly activated by several stress signals such as tumournecrosis factor, ionizing and short-wavelength ultravioletirradiation, but they are only weakly activated by growthfactors [30, 80, 81]. In contrast, the ERK pathway is highlyinduced in response to growth factors and cytokines, and isalso activated by oxidative stress. As shown in Fig. (9), cis-DDP is also able to activate these MAPK pathways in tu-mour cells or transformed cell lines.

3.4.5. p38 MAPK Pathway

It has been recently reported that cis-DDP triggers theactivation of p38 MAPK for 8–12 hours in sensitive cellsand for 1–3 hours in resistant cells [82]. Furthermore, a re-sistant phenotype was observed in human cells lacking p38MAPK activation or function [81, 82]. So, there is accumu-lating evidence of a key role for p38 MAPK pathway activa-tion in the cellular response to cisplatin [74, 83-85]. On theother hand, MAP kinases may also phosphorylate severalsubstrates, including transcription factors, chromatin proteinconstituents and downstream serine/threonine effectorkinases [86, 87], which indicates a putative role for theseproteins in the regulation of immediate-early gene expression[88]. Moreover, p38 MAPK pathway is also involved in invitro cisplatin-induced phosphorylation of histone H3 atSer10, together with the downstream kinase mitogen- andstress-activated protein kinase 1 (MSK1) [89].


3.4.6. ERK Pathway

ERK pathway can be activated by growth factors, cytoki-nes, oxidative stress and also cis-DDP. So, it has been re-ported that in vitro treatment with cisplatin triggers time-dependent activation of ERK1/2 in cells [90]. However, thelevel of ERK activity following cis-DDP treatment divergesbetween different cell lines as some cells show weak activa-tion of ERK or no activation at all [91]. There are also somestudies indicating that ERK activation might be associatedwith enhanced survival of cisplatin-treated cells [92, 93].However, the role of ERK in the survival of cisplatin-treatedcells remains still unclear. Cisplatin-induced ERK activityhas been blocked using some inhibitors of upstream proteinssuch as MEK1 inhibitor PD98059. So, it has been reportedthat PD98059 sensitizes the human melanoma cell lineC8161 to cisplatin-induced cell demise. In contrast,PD98059 did not enhance cisplatin-induced apoptosis inthree other human melanoma cell lines. Moreover, PD98059seems to protect against cisplatin-induced cell death in oneof these cell lines, apparently through the improvement ofcisplatin induction of NF-_B activity [94, 95]. All these dif-ferences indicate a direct dependence on the individual na-ture or context of specific cell lines for the relationship be-tween the activity of ERK proteins and the cellular responseto cisplatin [55].

3.4.7. JNK Pathway

There are several studies about JNK activation in re-sponse to cis-DDP treatment. However, at present, the rolefor this pathway in cell survival is still ambiguous. Somestudies show that JNK pathway participates in cisplatin-induced apoptosis [82, 91, 96, 97]. In contrast, it has alsobeen reported that JNK activation in response to cis-DDPpromotes cell survival [92, 98-100]. The inconsistent find-ings observed in these studies may be related to the differentnature of the cell lines. On the other hand, there is a studyshowing that the key factor in deciding between cell prolif-eration and apoptosis might be the time length of JNK in-duction [101]. Interestingly, cis-DDP induces JNK1 in a per-sistent way whereas trans-DDP activates JNK in a transientmanner. This fact might be due to their differential effectover MKP1/CL100 (MAP kinase phosphatase 1), which in-hibits JNK and p38 activation after cis-DDP treatment.

3.4.6. MKP1/CL100

MAP kinase phosphatase 1 (MKP1/CL100) is able toinhibit JNK and p38 induction after cis-DDP treatment,which increases cell survival. However, cisplatin weaklyinduces MKP1/CL100 in a continuous way, whereas trans-DDP is able to activate MKP1/CL100 powerfully but tran-siently, and its expression is connected with JNK inactiva-

Fig. (8). Transduction of cisplatin-DNA damage signal by p53 protein. After DNA damage, p53 is activated and it subsequently activatesdifferent sets of downstream target genes, which in turn induce diverse cellular responses. ATM, ataxia telangiectasia mutated; ATR, ataxiatelangiectasia and Rad3 related; BRCA1, breast cancer 1, early onset; CSB, cockayne syndrome B; GADD45, growth arrest and DNA dam-age 45; MDM2, mouse double minute 2 homologue; PCNA, proliferating cell nuclear antigen; RPA, replication A; TFIIH, transcription fac-tor IIH; XPC, xeroderma pigmentosum complementation group C.


tion [102]. In addition, it has been reported that mutant 293cells without ectopic MKP1/CL100 catalytic activity showlonger JNK induction by trans-DDP, and increased cytotox-icity of both cisplatin and trans-DDP [102, 103]. Therefore,these data indicate that cis-DDP treatment might induce per-sistent activation of JNK in the cell because of its inability toinduce the expression of MKP1/CL100. Consequently, theinhibition of MKP1/CL100 activity in tumour cells could bean interesting approach to improve the response to cisplatinin cancer cells [55].

3.4.7. Cell Cycle Arrest

There are two main checkpoints (G1/S and G2/M)through the cell cycle that screen the proper evolution of theprocess in order to avoid the acquisition and accumulation ofmutations [104]. In the event of DNA damage, the cell cyclewill be arrested to help the function of the repair machinery.The G1/S checkpoint allows DNA restoration before repli-cation, and G2/M facilitates the reparation of DNA damagedduring S or G2 steps to prevent its segregation into daughtercells. Treatment with cis-DDP does not usually induce G1arrest. In fact, even though cisplatin may stimulate p53 andlead to S-phase arrest, it fails to induce G1 arrest in synchro-nized wild-type MEF cells [105]. In contrast, G2 arrestseems to be essential to trigger cisplatin-induced cell death[106]. In the event of DNA damage, ATM and its relatedATR (ataxia-telangiectasia and Rad3-related) kinase activatethrough phosphorilation checkpoint kinases CHK1 andCHK2, which subsequently phosphorilate cell divison cycle

25C (CDC25C). Then, activated CDC25C binds to 14-3-3adaptor proteins and is thereby split off from CDC2 bytranslocation of CDC25C to the cytoplasm [55]. As a conse-quence, CDC2 phophorylation is enhanced and provokes cellarrest in G2 [12].

3.4.8. Telomere Shortening

It is now well-established that the limitless replicativepotential of tumor cells is related to their ability to maintain aconstant telomere length. Telomeres are tandem repeatedDNA sequences, comprising a G-rich strand and a comple-mentary C-rich strand, which are located at the end of thechromosomes. Telomeres protect the chromosomes fromDNA degradation, end-to-end fusions, rearrangements, andchromosome loss and maintain nuclear structure [107]. Insomatic cells, telomeres are shortened in every cell divisionand when they are critically shortened cells stop dividing andfinally die [108]. In tumor cells, telomere length is main-tained, mainly through activation of the reverse transcriptaseenzyme, telomerase [109]. Telomerase is a ribonucleoproteinthat uses its RNA component as a template to synthesize the5’-TTAGGG-3’ repeats at the ends of the chromosomes.Therefore, to maintain telomere length, cancer cells have toreplicate the telomeric motif several times yielding a(TTAGGG)n telomeric sequence [110]. Interestingly, it hasbeen found that exposure of HeLa human cervix carcinomacells to low concentrations of cis-DDP produces shorteningand degradation of telomeres, which leads to cell death. Apotential explanation to this phenomenon could be that te-

Fig. (9). Role of MAPK pathways in signal transduction of cisplatin damage to DNA. Cisplatin triggers the activation of the three majormammalian MAPK cascades (ERK, JNK and p38 MAPK) in tumor cells and transformed cell lines. ERK, extracellular signal-regulatedkinase; JNK, c-Jun NH2-terminal kinase; MAPK, mitogen-activated protein kinase.


lomeres are not transcribed so that they will not be repairedby the NER system. So, even low levels of telomere damagemay be enough to induce cell death by cisplatin [111]. Onthe other hand, it has also been reported that cis-DDP mayaffect telomerase activity by direct interaction with this en-zyme. However, other DNA-binding drugs including trans-DDP seems to have no effect on telomerase activity [36].

3.4.9. Ubiquitination

Ubiquitin (Ub) is a small and tightly folded protein,which is implicated through the ubiquitin-proteasome path-way in the selective degradation of short-lived cellular regu-latory proteins [112]. In view of the crucial importance of Ubin cellular metabolism, it has been hypothesized that a directinteraction of cis-DDP with Ub may trigger cytotoxic proc-esses leading to cell death [113]. In fact, it is known that cis-DDP has two binding sites in Ub, one N-terminal methionine(Met1) and one histidine (His68) and that the drug forms atleast 4 types of adducts on the protein [35]. Of interest is theobservation that inhibition of the ubiquitin-proteasomepathway by several proteasome inhibitors is being exploredas a new target for cancer chemotherapy.

DNA transcription can also be inhibited by DNA lesionsprovoked by cis-DDP. In fact, cisplatin-induced cell cyclearrest in the G2 phase suggests that some proteins necessaryfor mitosis are not being synthesized, which indicates thattranscription has been halted. Transcription failure might bea result of RNA polymerase blocking at cisplatin-DNA ad-ducts or cisplatin-DNA-protein ternary complexes. Inhibitionof RNA pol II transcription is provoked by ultraviolet radia-tion or cisplatin-induced damage leading to phosphorilationof its COO-terminal, which ends up in ubiquitination anddegradation in proteasomes [114]. Altogether all these ob-servations suggest that one feasible mechanism of cisplatincytotoxicity may be the inhibition of transcription and sub-sequent degradation of RNA pol II as a result of ubiquitina-tion. RNA pol II levels would not be restored becauseshielding of cisplatin-DNA adducts by repair proteins pro-tects cisplatin lesions from nucleotide excision repair. There-fore, there would be a constant decrease in RNA pol II levelsand transcriptional activity that might initiate cell deathpathways. This is an interesting hypothesis for a possiblemechanism of cisplatin cytotoxicity, which combines tran-scription inhibition with the “repair shielding model” of cis-platin-DNA intrastrand cross-links [115].

4. CISPLATIN INDUCED CELL DEATH PATHWAYS:INTERCONNECTIONS BETWEEN APOPTOTIC ANDNECROTIC PATHWAYS.

It is commonly accepted that futile attempts to repaircisplatin-DNA adducts induce cytotoxic processes that fi-nally end up in the triggering of apoptosis [10]. Apoptosisalso called “programmed cell death (PCD), is considered acontrolled pathway that requires ATP (adenosine triphosphate)and de novo protein synthesis. However, as mentioned before,most cis-DDP molecules bind to proteins rather than DNA,and there is experimental evidence that the former type ofdamage also plays a role in triggering PCD pathways [10,45]. It has been reported that some types of cancer cells whenare exposed to cis-DDP show internucleosomal DNA degrada-tion in approximately 180 base pair fragments, blebbing of the

cell surface, and cell shrinkage, all of which is consistent withapoptosis as a mode of cell death [52]. Besides, it has also beenreported that, in other cell lines, particularly those with resis-tance to the drug, cisplatin produces characteristic features ofnecrosis, which is considered a mode of cell death due to gen-eral cellular machinery failure [116-118]. Moreover, it has beenfound that necrotic and apoptotic cell death may take placetogether in the same population of cisplatin-treated cells [119].

PCD and necrosis have been traditionally considered astwo separate modes of cell demise, not only morphologicallybut also mechanistically [120]. In the 1980s, necrosis wasconsidered the mode of cell-death induced by DNA-damag-ing anticancer agents because of the activity poly(ADP-ribose) polymerase-1 (PARP-1) [121]. PARP-1 is activatedby the DNA strands breaks caused by anticancer agents andcleaves the glycolytic coenzyme NAD+, provoking the for-mation of poly(ADP-ribose) moieties. Depletion of NAD+

inhibits glycolytic production of ATP with subsequent ATPdepletion, which leads to necrotic cell death [6]. By the1990s, it was thought that most clinically effective anticanceragents that bind to DNA kill cancer cells by apoptosis. Anoutstanding contribution to the study of the biochemicalmechanisms of cell death was the discovery by the end of the1990s that intracellular ATP levels dictate whether antitumordrugs including cis-DDP as well as other chemical andphysical agents induce cell death by necrosis or apoptosisand that both processes of cell death are linked [122-124].Fig. (10) shows an outline of the interconnections betweenapoptotic and necrotic pathways within the cell in responseto cisplatin assault. It may be seen that activation ofcaspases, before PARP-1 cleavage of NAD+ induces deple-tion of ATP to levels causing cell death by necrosis, leads toexecution of apoptosis. In fact, the destruction of PARP-1activity by cleavage due to caspase-3-6-7 allows caspaseactivity to complete apoptosis. Caspase blocking by inhibi-tors of apoptosis (IAPs) plus continued activity by PARP-1,plus the ATP decrease from the loss of electron transport inbroken mitochondria, lead the cell to necrosis because ofcontinuation of PARP-induced ATP depletion [125]. Sur-prisingly, however, it has been also reported that failure ofPARP-1 cleavage enhances apoptosis [126]. A possible ex-planation for this paradox might be related with the fact thatcontinued activity of PARP -1 induces changes in the pyri-dine nucleotide pool (NADH/NAD+ and NADPH/NADP+)and nucleotide pool of ATP/ADP that regulate MPT [127].All of the above observations indicate that, rather than afunctional opposition between necrosis and apoptosis, thereis a functional cooperativity. This hypothesis would explainthe data that suggests that some cells might die as a result ofan unfinished apoptotic program. For instance, in L1210murine leukemic cells, cisplatin-induced cell death might bethe consequence of a defective apoptotic program that lackssome morphological and biochemical properties attributed to“classic” apoptosis [128].

6. DEVELOPMENT OF NEW PLATINUM ANTI-TUMOR DRUGS

Understanding the molecular mechanisms by which tu-mor cells react to cis-DDP damage may lead to the design ofmore pharmacologically efficient platinum antitumor drugs.Platinum anti-cancer research has been going on for three


decades now, and the increased understanding of the bio-chemical mechanism of action of cisplatin and other clini-cally-used platinum drugs, has allowed a change from puretrial-and-error testing to a more rational approach in the de-sign of new platinum agents. Development of platinum com-pounds capable of circumventing cis-DDP resistance isthought to be a feasible strategy. However, this approach hasproven to be a difficult task. In fact, most of the compoundssynthesized and biologically tested are simple analogues ofthe parent compounds cisplatin, carboplatin or oxaliplatin.Therefore, little success has been achieved so far in devel-oping better platinum drugs by simple substitution of theammine ligands or chloride leaving groups of cisplatin. Cur-rent knowledge about the molecular bases of platinum resis-tance has led to the design of cis-ammine(2-methylpyridine)dichloroplatinum(II) (ZD0473), which shows improved invitro activity compared with cis-DDP [5, 129]. The rationale

for the synthesis of this compound was to delay the action ofglutathione and other cellular thiols that inactivate platinumdrugs before they can reach target DNA. An interesting strat-egy for building potential novel platinum anticancer drugcandidates might be the addition to the platinum center of amolecular fragment that can interfere with a specific cellularpathway. So, in these drugs it might be expected a synergyeffect between the platinum atom, which carries into the tu-mor cell the DNA-binding bullet and the molecular frag-ment, which can inhibit a cellular protein involved in a spe-cific transduction pathway. Almost any molecular fragmentcan be attached to cis-DDP, carboplatin or oxaliplatin byoxidation to form the trans-dihydroxoplatinum (IV) pro-druganalogue. Once inside the tumour cell, the Pt(IV) complex israpidly reduced to its Pt(II) precursor, recovering its potencywhile liberating the pathway-modulating moiety [130, 131].These complexes have several advantages in addition to their

Fig. (10). Biochemical cell death pathways induced by cis-DDP showing the interconnections between apoptosis and necrosis (discontinuousarrows) depending on the metabolic condition of the tumor cell. DNA strand breaks induced by cis-DDP activates PARP-1, which cleavesNAD+ into poly(ADP-ribose) and nicotinamide moieties. The result is a decrease in NAD+ level with a concomitant fall of ATP levels. IfATP levels in the tumor cell are low, then, it dies through necrosis. In contrast, if ATP levels are enough to sustain cell survival, the caspase-3-6-7 complex cleaves PARP-1 so that necrosis is blocked and the tumor cell dies through apoptosis. Apaf-1, apoptotic protease-activatingfactor-1; Bid, a type of proapoptotic protein; FAAD, Fas-associated death domain; Fas, cell surface membrane receptor; TNRF-1, tumornecrosis factor receptor; IAP, inhibitor of apoptosis; PARP-1, poly(ADP-ribose) polymerase-1; ROS, reactive oxygen species; cyto C, cyto-chrome C; Mito, mitochondrial; MPT, mitochondrial permeability transition.


fast activation within the cell; they can also be chemicallymodified at three locations: the axial, the ammine function-alities, and the leaving groups, which offer a wide range ofpossibilities. The moieties that can be attached to the plati-num core could be, as an example, an AKT inhibitor; a sti-mulator of c-ABL tyrosine kinase; a molecule that promotesp53 activity; or an inhibitor of RB, BRCA1 or p38 MAPK.Moreover, the complexes could be orally delivered as previ-ously assessed in clinical trials with the Pt(IV) complexJM216 [132]. A more advanced option would be transportingthe platinum complex to tumor cells with specifics receptorsfor the attached fragment [130, 133]. This approach offers anunlimited number of exciting possibilities; however, furtherclinical testing will be needed.

7. BIOCHEMICAL MODULATION OF CISPLATINCHEMOSENSITIVITY

Biochemical modulation of key cellular pathways di-rected to circumvent cisplatin resistance may be a comple-mentary way to the development of novel platinum antitu-mor drugs [6]. Hence, understanding of the major mecha-nisms of cisplatin resistance in cancer cells has also openednew directions for pharmacological exploitation of cis-DDPchemotherapy.

An interesting approach is the combination of cis-DDPwith two or more known non-platinum antitumor drugs. Thisstrategy is markedly improving cellular survival in severalcancers. Nevertheless, the mechanisms of such synergisticinteractions remain poorly understood. Cisplatin, carboplatinand oxaliplatin have been evaluated in combination withother chemotherapeutic agents, such as etoposide, mitomycinC, vinblastine, paclitaxel, docetaxel, vinorelbine, gemcit-abine, cyclophosphamide, doxorubicin, epirubicin, metho-trexate, lonidamine and 5-fluorouracil (5-FU) [5, 134]. Pa-tients with metastatic breast cancer showed good clinicalresponses when treated with cis-DDP in combination withetoposide, a topoisomerase II inhibitor that inhibits cell-cycleprogression before mitosis. Combination with 5-FU alsoimproved response rates in head and neck cancer [135, 136].Carboplatin, combined with paclitaxel, a microtubule disas-sembly inhibitor, has been also used in ovarian cancer, non-small-cell lung cancer, and head and neck cancer [137],whereas oxaliplatin with 5-FU improved survival in coloncancer [135]. However, deeper studies of the molecularmechanisms of these synergistic interactions are needed.

An alternative strategy is the development of rationalcombination therapies which interfere with known platinumprocessing pathways, specially those processes that takeplace closest in time to the formation of DNA adducts. Forinstance, the pharmacological inhibition of NER DNA repairsystem is attracting much attention recently. In addition, theuse of topoisomerase II inhibitors or DNA polymerase in-hibitors may help to overcome acquired cis-DDP resistance.In fact, as mentioned above, breast cancer patients achievedexcellent clinical responses rates when treated with cis-DDPin combination with topo II inhibitor etoposide, which haltscell-cycle progression before mitosis. [5]. On the other hand,a relationship between cisplatin killing efficacy and somehistone deacetylase inhibitors has been recently reported.Thus, pretreatment with trichostatin A (TSA) or suberoy-

lamilide hydroxamic acid (SAHA), two histone deacetylaseinhibitors, enhanced the damaging effect of cis-DDP. Theresults of this study show that the relaxation of the chromatinstructure induced by histone acetylation increases the effectsof the drug, maybe by influencing the adduct formation[138]. Therefore, the pharmacological management of chro-matin organization could be a complementary way to en-hance cisplatin cytotoxicity.

As mentioned already, PARP-1 enzyme is involved in thecellular response to DNA damage so that in the event of ir-reparable DNA damage overactivation of PARP-1 leads tonecrotic cell death [125]. As shown in Fig. (11), inhibitors ofPARP-1 activity in combination with cis-DDP may consti-tute a suitable strategy in cancer chemotherapy [139]. WhenDNA is moderately damaged, PARP-1 participates in theDNA repair process and then the cell survives. However, inthe case of extensive DNA damage PARP-1 overactivationinduces a decrease of NAD+ and ATP levels leading to celldysfunction or even to necrotic cell death. Therefore, due toPARP-1 involvement in cell death, pharmacological inhibi-tion of PARP-1 activity by PARP-1 inhibitors may constitutea suitable target to enhance the activity of cis-DDP throughinhibition of necrosis and activation of apoptosis. PARP-1inhibitors such as 3-aminobenzamide, 1,5-dihydroxyisoqui-nolinone and the recently patented tryciclic benzimidazoleshave shown potent inhibitory effects of PARP-1 activity intumor cells [125].

8. CONCLUDING REMARKS AND FUTURE TRENDS

Basic research on platinum antitumor drugs has beengoing on for more than three decades now, and the increasedunderstanding of the biochemical mechanism of action ofcis-DDP has allowed a change from pure trial-and-errortesting to a more rational approach in the design of newplatinum agents. Cisplatin is one of the most potent com-pounds currently used in cancer chemotherapy and one of thevery few therapeutic drugs available for cancer patients.However, improvement of its potency and avoidance of un-desired side effects are still important goals for the future.

It is interesting to point out that apoptosis and necrosishas been frequently observed in the same population of tu-mor cells treated with cisplatin [119]. It is known that mostcisplatin molecules that enter the cell are bound to proteinsrather than DNA and there is experimental evidence showingthat the former type of damage also plays a role in the initia-tion of apoptotic pathways [35, 113]. Tumor cell populationscontain cancer cells of variable sensitivity to cisplatin. Hence,either apoptotic or necrotic cell death has been observed inindividual cells within the same tumor cell population [140].Therefore, it can be hypothesized that apoptosis and necrosisare the two extremes of a continuum of possible modes of celldeath so that apoptotic and necrotic pathways of cell death areinterconnected [122, 123]. This hypothesis establishes that in-dividual cell death after cisplatin attack might be decided byfactors such as the availability of energy and the metaboliccondition of the cell. Moreover, it is likely that even some cellsthat initiate the apoptotic program in response to cisplatin dam-age could not finish it dying then by necrosis.

Biochemical modulation of cisplatin mechanisms of re-sistance may offer multiple opportunities for future applica-


tions in clinical cancer research. Thus, Modulation of cellu-lar sensitivity to cisplatin by the use of inhibitors, activatorsor combinations of drugs that interfere with certain pathwayswould improve therapeutic response and may constitute acomplementary strategy to the discovery of novel platinumcomplexes.

ACKNOWLEDGEMENTS

This work was supported by Spanish Ministerio de Edu-cación y Ciencia (SAF 2004-03111). We also thank sponsor-ship by European COST Action D20/0003/00. An institu-tional grant from Fundación Ramón Areces is also acknowl-edged. VC and CQ are the recipients of predoctoral (FPI)and postdoctoral (Juan de la Cierva) fellowships from theSpanish Ministerio de Educación y Ciencia, respectively.

REFERENCES

[1] Peyrone, M. Ann. Chem. Pharm., 1845, 51, 1.[2] Rosenberg, B.;VanCamp, L.; Krigas, T. Nature, 1965, 205, 698.[3] Rosenberg,B; VanCamp, L.; Trosko, J.E.; Mansour, V.H. Nature,

1969, 222, 385.[4] Giaccone, G. Drugs, 2000, 59, U4.[5] Wong, E; Giandomenico, C.M. Chem. Rev., 1999, 99, 2451.

[6] Fuertes, M.A.; Alonso, C.; and Pérez, J.M. Chem. Rev, 2003, 103,645.

[7] Cvitkovic, E. Semin. Oncol., 1998, 25, 1.[8] Chu, G. J. Biol. Chem., 1994, 269, 787.[9] Kelland, L.R. Drugs, 2000, 59, 1.[10] González, V.M.; Fuertes, M.A.; Alonso, C.; Pérez, J.M. Mol.

Pharmacol., 2001, 59, 657.[11] Judson, I.; Kelland, L.R. Drugs, 2000, 59, 29.[12] Jamieson, E.R.; Lippard, S.J. Chem. Rev., 1999, 99, 2467.[13] Miller, S.E.; House, D.A. Inorg. Chim. Acta, 1991, 187, 125.[14] Binks, S.P.; Dobrota, M. Biochem. Pharmacol., 1990, 40, 1329.[15] Hromas, R.A.; North, J.A.; Burns, C.P. Cancer Lett., 1987, 36, 197.[16] Mann, S.C.; Andrews, P.A.; Howell, S.B. Int. J. Cancer , 1991, 48,

866.[17] Gately, D.P.; Howell, S.B. Brit. J. Cancer, 1993, 67, 1171.[18] Ishida, S.; Lee, J.; Thiele, D.J.; Herskowitz, I. Proc. Natl. Acad.Sci.

USA, 2002, 99, 14298.[19] Holzer, A.K.; Sanini, G.; Katano, K.; Naerdeman, W., Lin, X.;

Safaei, R.; Howell, S.B. Mol. Pharmacol., 2004, 66, 817.[20] Komatsu, M.; Sumizawa, T.; Mutoh, M.; Chen, Z.S.; Terada, K.;

Furukawa, T.; Yang, X.L.; Gao, H.; Miura, N.; Sugiyama, T.; Aki-yama, S. Cancer Res., 2000, 60, 1312.

[21] Miyashita, H.; Nitta, Y.; Mori, S.; Kanzaki, A.; Nakayama, K.;Terada, K.; Sugiyama, T.; Kawamura, H.; Suto A.; Morikawa, H.;Motegi, K., Takebayashi, Y. Oral Oncol., 2003, 39, 157.

[22] Nakayama, K.; Kanzaki, A.; Ogawa, K.; Miyazaki, K.; Neamati,N.; Takebayashi, Y. Int. J. Cancer, 2002, 101, 488.

Fig. (11). Biochemical modulation of cis-DDP cytotoxicity by PARP-1 inhibitors. PARP-1 overactivation by cisplatin leads to necrotic celldeath so that PARP-1 inhibition by compounds such as 3-aminobenzamide (3-AB) ends up in cisplatin-induced apoptosis.


[23] Nakayama, K.; Kanzaki, A; Terada, K.; Mutoh, M.; Ogawa, K.;Sugiyama, T.; Takenoshita, S.; Itoh, K.; Yaegashi, N.; Miyazaki,K.; Neamati, N.; Takebayashi, Y. Clin. Cancer Res., 2004, 10,2804.

[24] Ohbu, M.; Ogawa, K.; Konno, S.; Kanzaki, A.; Terada, K.; Sugi-yama, T.; Takebayashi, Y. Cancer Lett., 2003, 189, 33.

[25] Cui, Y.; Konig, J.; Bucholz, J.K.; Spring, H.; Leier, I.; Keppler, A .Mol. Pharmacol., 1999, 55, 929.

[26] Koike, K.; Kawabe, T.; Tanaka, T., Toh, S.; Uchiumi, T.; Wada,M.; Akiyama, S., Ono, M.; Kuwano, M. Cancer Res., 1997, 57,5475.

[27] Kool, M.; de Haas, M.; Scheffer, G.L.; Scheper, R.J.; van Eijk,M.J.; Juijn, J.A.; Baas, F., Borst, P. Cancer Res., 1997, 57, 3537.

[28] Katano, K.; Kondo, A.; Safei, R.; Holzer, A.; Samimi, G.;Mishima, M.; Kuo, Y.M. Cancer Res., 2002, 62, 6559.

[29] Safaei, R.; Howell, S.B. Crit. Rev. Oncol. Hematol., 2005, 53, 13.[30] Andrews, P.A. In Platinum-Based Drugs in Cancer Therapy; Kel-

land, L.R., Farrell, N.P., Eds.; Totowa: Humana Press, 2000, pp.89-113.

[31] Speelmans, G.; Staffhorst, R.W.; Versluis, K.; Reedijk, J.; deKruijff, B. Biochemistry, 1997, 36, 10545.

[32] Jordan, P.; Carmo-Fonseca, M. Cell. Mol. Life Sci., 2000, 57, 1229.[33] Akaboshi, M.; Kawai, K.; Maki, H.; Akuta, K.;Ujeno, Y.; Miya-

hara, T. Jap. J. Cancer Res., 1992, 83, 522.[34] Akaboshi, M.; Kawai, K.; Ujeno, Y.; Takada, S.; Miyahara, T. Jap.

J. Cancer Res., 1994, 85, 106.[35] Peleg-Shulman, T.; Gibson, D. J. Am.Chem. Soc., 2001, 123, 3171.[36] Soti, C.; Racz, A.; Csermely, P. J. Biol.Chem., 2002, 277, 7066.[37] Payet, D.G.; Sip, F.; Leng, M. Nucl. Acids Res., 1993, 21, 5846.[38] Danford, A.J.; Wang, D.; Wang Q.; Tullius, T.D.; Lippard, S.J.

Proc. Natl. Acad. Sci. USA, 2005, 102, 12311.[39] Pérez, J.M.; Fuertes, M.A.; Alonso, C.; Navarro-Ranninger, C.

Crit. Rev. Oncol. Hematol., 2000, 35, 109.[40] Leng, M.; Locker, D.; Giraud-Panis, M.J.; Schwartz, A.; Intini,

F.P.; Natile, G.; Pisano, C.; Boccarelli, A.; Giordano, D.; Coluccia,M. Mol. Pharm., 2000, 58, 1525.

[41] Yang, X-L.; Wang, A.H-J. Pharmacol. Therapeut., 1999, 83, 181.[42] Fuertes, M.A.; Castilla, J.; Alonso, C.; Pérez, J.M. Curr. Med.

Chem. Anticancer Agents, 2002, 2, 539.[43] Pil, P.M.; Lippard, S.J. Science, 1992, 256, 234.[44] Zamble, D.B.; Mu, D; Reardon, J. T.; Sancar, A.; Lippard, S.J.

Biochemistry, 1996, 35, 10004.[45] Kruidering, M.; van de Water, B.; Zhan, Y.; Baelde, J.J.; Heer, E.;

Mulder, G.J., Stevens, J.L.; Nagelkerke, J.F. Cell Death Diff., 1998,5, 601.

[46] Lau, A.H. Kidney Int., 1999, 56, 1295.[47] Chaney, S.G.; and Sancar, A. J. Natl Cancer Inst., 1996, 88, 1346.[48] Bruhn, S.L.; Pil, P.M.; Essigmann, J.M.; Housman, D.E.; Lippard,

S.J. Proc. Natl. Acad. Sci. USA, 1992, 89, 2307.[49] Fishel, R. Cancer Res., 2001, 61, 7369.[50] Reed, E. In Cancer Chemotherapy and Biological Response Modi-

fiers. Pinedo H.M., Longo, D.L., Chabner, B.A., Eds.; Elsevier Sci-ence BV: Amsterdan, 1999, pp. 144-151.

[51] Turchi, J.J.; Henkels, K. J. Biol. Chem., 1996, 271, 13861.[52] Henkels, K.M.; Turchi, J.J. Cancer Res., 1997; 57, 4488.[53] Brown, S.J.; Kellett, P.J.; Lippard, S.J. Science, 1993, 261, 603.[54] Orphanides, G.; Wu, WH.; Lane W.S.; Hampsey, M.; Reinberg, D.

Nature, 1999, 400, 284.[55] Wang, D.; Lippard, S.J. Nat. Rev. Drug Discover, 2005, 4, 307.[56] Wei, M.; Burenkova, O.; Lippard, S.J. J. Biol. Chem., 2003, 278,

1769.[57] Juo, Z.S.; Chiu, T.K.; Leiberman, P.M.; Baikalov, I.; Berk, A.J.;

Dickerson, R.E. J. Mol. Biol., 1996, 261, 239.[58] Wei, M.; Cohen, S.M.; Silverman, A.P.; Lippard, S.J. J. Biol.

Chem., 2001, 276, 38774.[59] Yaneva, J.; Leuba, S.H.; van Holde K.; Zlatanova, J. Proc. Natl.

Acad. Sci. USA, 1997, 94, 13448.[60] Kartalou, M.; Samson, L.D.; Essigmann, J.M. Biochemistry, 2000,

39, 8032.[61] Blackburn, E.H. Nature, 1991, 350, 569.[62] Asselin, E.; Mills, G.B.; Tsang, B.K. Cancer Res., 2001, 61, 1862.[63] Dan, H.C.; Sun M.; Kameko, S.; Feldman, R.I.; Nicosia, S.V.;

Wang, H.G.; Tsang, B.K.; Chen, J.Q. J. Biol.Chem., 2004, 279,5405.

[64] Fraser, M.; Leung, B.M.; Yan, X.; Dan H.C.; Cheng, J.Q.; Tsang,B.K. Cancer Res., 2003, 63, 7081.

[65] Mabuchi, S.; Ohmichi M.; Nishio Y.; Hayasaka, T.; Kimura, A.;Ohta, T.; Saito, M.; Kawagoe, J.; Takahashi, K.; Yada-Hashimoto,N.; Sakata, M.; Motoyama, T.; Kurachi, H.; Tasaka, K.; Murata Y.J. Biol. Chem., 2004, 279, 23477.

[66] Pommier, Y.; Sordet, O.; Antony, S.; Hayward, R.L.; Kohn, K.W.Oncogene, 2004, 23, 2934.

[67] Shaul, Y. Cell Death Diff., 2000, 7, 10.[68] Cong, F.; Goff, S.P. Proc. Natl. Acad. Sci. USA, 1999, 96, 13819.[69] Theis, S.; Roemer, K. Oncogene, 1998, 17, 557.[70] Gong, J.G.; Constanzo, A.; Yan H.Q.; Melino, G.; Kaelin, W.G.

Jr.; Levrero, M.; Wang, J.Y. Nature, 1999, 399, 806.[71] Kharbanda, S.; Pandey, P.; Yamauchi, T.; Kumar, S.; Kaneki, M.;

Kumar, V.; Barthi, A.; Yuan, Z.M.; Ghanem, L.; Rana, A.; Weich-selbaum, R.; Johnson, G.; Kufe, D. Mol. Cell. Biol., 2000, 20,4979.

[72] Kharbanda, S.; Ren, R.; Pandey, P.; Shafman, T.D.; Feller, S.M.,Weichselbaum, R.R.; Kufe, D.W. Nature, 1995, 376, 785.

[73] Sánchez-Prieto, R.; Sánchez-Arevalo, V.J.; Servitja, J.M.; Gutkind,J.S. Oncogene, 2002, 21, 974.

[74] Pandey, P.; Raingeaud, J.; Kaneki, M.; Weichselbaum, R.; Davis,R.J.; Kufe, D.; Kharbanda, S. J. Biol. Chem., 1996, 271, 23775.

[75] David-Cordonnier, M.H.; Payet, D.; D'Halluin, J.C.; Waring, M.J.;Travers, A.A.; Bailly, C. Nucleic Acids Res., 1999, 27, 2265.

[76] Miao, Y.J.; Wang, J.Y.J. J. Biol. Chem., 1996, 271, 22823.[77] Fisher, D.E. Cell, 1994, 78, 539.[78] Dempke, W.; Voigt, W.; Grothey, A.; Schmoll, H.J. Anticancer

Drugs, 2000, 11, 225.[79] Weller, M. Cell Tissue Res., 1998, 292, 435.[80] Olson, J.M.; Hallahan, A.R. Trends in Mol. Med., 2004, 10, 125.[81] Wada, T.; Penninger, J.M. Oncogene, 2004, 23, 2838.[82] Mansouri, A., Ridway, L.D.; Korapati, A.L.; Zhang, Q.; Tian, L.;

Wang, Y.; Siddik, Z.H.; Mills, G.B.; Claret, F.X. J. Biol. Chem.,2003, 278, 19245.

[83] Losa, J.H.; Parada Cobo, C.; Viniegra, J.G.; Sánchez-ArevaloLobo, V.J; Ramón y Cajal, S.; Sánchez-Prieto, R. Oncogene, 2003,22, 3998.

[84] Kumar, S.; Boehm, J.; Lee, J.C. Nat. Rev. Drug Discover ., 2003, 2,717.

[85] Pillaire, M.J.; Nebreda, A.R.; Darbon, J.M. Biochem. Biophys. Res.Comm., 2000, 278, 724.

[86] Soloaga, A.; Thomson, S.; Wiggin, G.R.; Rampersaud, N.; Dyson,M.H.; Hazzalin, C.A.; Mahadevan, L.C.; Arthur, J.S. EMBO J.,2003, 22, 2788.

[87] Thomson, S; Clayton, A.L.; Hazzalin, C.A.; Rose, S.; Barrat, M.J.;Mahadevan, L.C. EMBO J., 1999, 18, 4779.

[88] Yang, S.H.; Sharrocks, A.D.; Whitmarsh, A.J. Gene, 2003, 320; 3.[89] Wang, D.; Lippard, S.J. J. Biol. Chem., 2004, 279, 20622.[90] Persons, D.L.; Yazlovitskaya, E.M.; Pelling, J.C. J. Biol. Chem.,

2000, 275, 35778.[91] Sánchez-Párez, I.; Murguia, J.R.; Perona, R. Oncogene, 1998, 16,

533.[92] Hayakawa, J.; Ohmichi, M.; Kurachi, H.; Ikegami, H.; Kimura, A.;

Matsuoka, T.; Jikihara, H.; Mercola, D.; Murata, Y. J. Biol. Chem .,1999, 274, 31648.

[93] Persons, D.L.; Yazlovitskaya, E.M.; Cui, W.; Pelling, J.C. Clin.Cancer Res., 1999, 5, 1007.

[94] Mandic, A.; Viktorsson, K.; Molin, M.; Akusjarvi, G.; Eguchi, H.;Hayashi, S.I.; Toi, M.; Hansson, J.; Linder, S.; Shosshan, M.C.Melanoma Res., 2001, 11, 11.

[95] Yeh, P.Y.; Chuang, S.E.; Yeh, K.H.; Song, Y.C.; Ea, C.K.; Cheng,A.L. Biochem. Pharmacol., 2002, 63, 1423.

[96] Sánchez-Pérez, I.; Perona, R. FEBS Lett., 1999, 453, 151.[97] Zanke, B.W.; Boudreau, K.; Rubie, E.; Winnett, E.; Tibbles, L.A.;

Zon, L.; Kyriakis, J.; Liu, F.F. Curr. Biol., 1996, 6, 606.[98] Davis, R.J. Cell, 2000, 103, 239.[99] Levresse, V.; Marek, L.; Blumberg, D.; Heasley, L.E. Mol. Phar-

macol., 2002, 62, 689.[100] Potapova, O.; Haghighi, A.; Bost, F.; Liu, C.; Birrer, M.J.; Gjerset,

R.; Mercola, D. J. Biol. Chem., 1997, 272, 14041.[101] Chen, Y.R.; Wang, X.; Templeton, D.; Davis, R.J.; Tan, T.H. J.

Biol. Chem., 1996, 271, 31929.[102] Sánchez-Pérez, I.; Martínez-Gomariz, M.; Williams, D.; Keyse,

S.M.; Perona, R. Oncogene, 2000, 19, 5142.[103] Franklin, C.C.; Srikanth, S.; Kraft, A.S. Proc. Natl. Acad. Sci. USA,

1998, 95, 3014.


[104] Dasika, G.K.; Lin, S.C.; Zhao, S.; Sung, P.; Tomkinson, A.; Lee,E.Y. Oncogene, 1999, 18, 7883.

[105] Attardi, L.D.; de Vries, A.; Jacks, T. Oncogene, 2004, 23, 973.[106] Sorenson, C.M.; Eastman, A. Cancer Res., 1988, 48, 6703.[107] Blackburn, E.H. Cell, 2001, 106, 661.[108] Harley, C.B.; Futcher, A.B.; Greider, C.W. Nature, 1990, 345, 458.[109] Neidle, S.; Kelland, L.R. Anticancer Drug Des., 1999, 14, 341.[110] Shay, J.W.; Bacchetti, S. Eur. J. Cancer, 1997, 33, 787.[111] Ishibashi, T.; Lippard, S.J. Proc. Natl. Acad. Sci. USA, 1998, 95,

4219.[112] Daino, H.; Matsumura, I.; Takada, K.; Odajima, J.; Tamaka, H.;

Ueda, S.; Shibayama, H.; Ikeda, H., Hibi, M.; Machii, T.; Hirano,T.; Kanakura, Y. Blood, 2000, 95, 2577.

[113] Nguewa, P.A.; Fuertes, M.A.; Cepeda, V.; Iborra, S.; Carrión, J.;Valladares, B.; Alonso, C.; Pérez, J.M. Chem. Biodiver ., 2005, 2,1387.

[114] Hanahan, D; Weinberg, R.A. Cell, 2000, 100, 57.[115] Cohen, S.M.; Lippard, S.J. Prog. Nucleic Acid Res. Mol. Biol.,

2001, 67, 93.[116] Guchelaar, H.J.; Vermes, I.; Koopmans, R.P.; Reutenlingsperger,

C.P.; Haanen, C. Cancer Chem. Pharmacol., 1998, 42, 77.[117] Matsumoto, M.; Tsuchida, T.; Kawamoto, K. Int. J. Oncol., 1997,

11, 1209.[118] Pérez, J.M.; Quiroga, A.G.; Montero, E.I.; Alonso, C.; Navarro-

Ranninger, C. J. Inorg. Biochem., 1999, 73, 235.[119] Montero, E.I.; Pérez, J.M.; Schwartz, A.; Fuertes, M.A.; Malinge,

J.M.; Alonso, C.; Leng, M., Navarro-Ranninger, C. Chembiochem,2002, 3, 61.

[120] Wyllie, A.H. J. Pathol., 1987, 153, 313.[121] Tanizawa, A.; Kubota, M.; Hashimoto, H.; Shimizu, T.; Takimoto,

T.; Kitoth, T.; Akiyama, Y.; Mikama, H. Exp. Cell Res., 1989, 185,237.

[122] Eguchi, Y.; Shimizu, S.; Tsujimoto, Y. Cancer Res., 1997, 57,1835.

[123] Leist, M.; Single, B.; Castoldi, A.F.; Kühnle, S.; Nicotera, P. J.Exp. Med., 1997, 185, 1481.

[124] Zhou, R.; Vander Heiden, M.G.; Rudin, C.M. Cancer Res ., 2002,62, 3515.

[125] Fuertes, M.A.; Castilla, J.; Nguewa, P.; Alonso, C., Pérez, J.M.Med. Chem. Rev. Online, 2004, 1, 187.

[126] Herceg, Z.; Wang, Z.Q. Mol. Cell. Biol., 1999, 19, 5124.[127] Hirsch, T.; Marchetti, P.; Susin, S.; Dellaporta, B.; Zamzani, M.;

Marzo, I.; Geuskens, N.; Kroemer, G. Oncogene, 1997, 15, 1573.[128] Segal-Bendirdjian, E.; Jacquemin-Sablon, A. Exp. Cell. Res., 1995,

218, 201.[129] Ho, Y.P.; Au-Yeung, S.C.F.; To, K.K.W. Med. Res. Rev., 2003, 23,

633.[130] Barnes, K.R.; Kutikov, A.; Lippard, S.J. Chem. Biol., 2004, 11,

557.[131] Pérez, J.M.; Kelland, L.R.; Montero, E.I.; Boxall, F.E.; Fuertes,

M.A.; Alonso, C.; Navarro-Ranninger, C. Mol. Pharm ., 2003, 63,933.

[132] Nguewa, P.; Cepeda, V.; Fuertes, M.A.; Castilla, J.; Alonso, C.;Pérez, J.M. In Metal Compounds in Cancer Chemotherapy, J.M.Pérez, M.A. Fuertes, C. Alonso, Eds., Research Signpost: Trivan-drum, Kerala, India, 2005, pp. 1-29.

[133] Jackson, A.; Davis, J.; Pither, R.J.; Rodgers, A.; Hannon, M.J.Inorg. Chem., 2001, 40, 3964.

[134] Bunn, P.A. J. Clin.Oncol., 2002, 20, 23S.[135] Decatris, M.P.; Sundar, S.; O'Byrne, K.J. Cancer Treat. Rev., 2004,

30, 53.[136] Shelley, M.D.; Burgon, K.; Mason, M.D. Cancer Treat. Rev., 2002,

28, 237.[137] Lebwohl, D.; Canetta, R. Eur. J. Cancer, 1998, 34, 1522.[138] Kim, M.S.; Blake, M.; Baek, J.M.; Kohlhagen, G.; Pommier, Y.;

Carrier, F. Cancer Res., 2003, 63, 7291.[139] Cepeda, V.; Fuertes, M.A.; Castilla, J.; Alonso, C.; Quevedo, C.;

Soto, M.; Pérez, J.M. Rec. Pat. Anticancer Drug Discover., 2006,1, (in press).

[140] Martin, D.S.; Bertino, J.R.; Koutcher, J.A. Cancer Res ., 2000, 60,6776.

Received: November 3, 2005 Revised: December 21, 2005 Accepted: April 10, 2006

Biochemical Mechanisms of Cisplatin Cytotoxicity

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