Platinum Anticancer Agents Research Articles

Solution Structures of a DNA Dodecamer Duplex with and without a Cisplatin 1,2-d(GG) Intrastrand Cross-Link: Comparison with the Same DNA Duplex Containing an Oxaliplatin 1,2-d(GG) Intrastrand Cross-Link,

Proteins that discriminate between cisplatin-DNA adducts and oxaliplatin-DNA adducts are thought to be responsible for the differences in tumor range, toxicity, and mutagenicity of these two important chemotherapeutic agents. However, the structural basis for differential protein recognition of these adducts has not been determined and could be important for the design of more effective platinum anticancer agents. We have determined high-resolution NMR structures for cisplatin-GG and undamaged DNA dodecamers in the AGGC sequence context and have compared these structures with the oxaliplatin-GG structure in the same sequence context determined previously in our laboratory. This structural study allows the first direct comparison of cisplatin-GG DNA and oxaliplatin-GG DNA solution structures referenced to undamaged DNA in the same sequence context. Non-hydrogen atom rmsds of 0.81 and 1.21 were determined for the 15 lowest-energy structures for cisplatin-GG DNA and undamaged DNA, respectively, indicating good structural convergence. The theoretical NOESY spectra obtained by back-calculation from the final average structures showed excellent agreement with the experimental data, indicating that the final structures are consistent with the NMR data. Several significant conformational differences were observed between the cisplatin-GG adduct and the oxaliplatin-GG adduct, including buckle at the 5' G6.C19 base pair, opening at the 3' G7.C18 base pair, twist at the A5G6.T20C19 base pair step, slide, twist, and roll at the G6G7.C19C18 base pair step, slide at the G7C8.C18G17 base pair step, G6G7 dihedral angle, and overall bend angle. We hypothesize that these conformational differences may be related to the ability of various DNA repair proteins, DNA binding proteins, and DNA polymerases to discriminate between cisplatin-GG and oxaliplatin-GG adducts.

Understanding the Effect of Carbonate Ion on Cisplatin Binding to DNA

The role of carbonate in the binding of cis-diamminedichloroplatinum(II) to DNA was investigated in order to understand the potential involvement of carbonato-cisplatin species in the mechanism of action of platinum anticancer agents. Cisplatin was allowed to react with both double- and single-stranded DNA in carbonate, phosphate, and HEPES buffers, and the products were analyzed by agarose gel electrophoresis and enzymatic digestion/mass spectrometry, respectively. The data from these experiments demonstrate (1) that carbonate, like other biological nucleophiles, forms relatively inert complexes with platinum that inactivate cisplatin and (2) that the major cisplatin−DNA adduct formed is a bifunctional cross-link. These results are in accord with previous studies of cisplatin−DNA binding and reveal that the presence of carbonate has no consequence on the nature of the resulting adducts.

Pre-association of polynuclear platinum anticancer agents on a protein, human serum albumin. Implications for drug design.

The interactions of polynuclear platinum complexes with human serum albumin were studied. The compounds examined were the "non-covalent" analogs of the trinuclear BBR3464 as well as the dinuclear spermidine-bridged compounds differing in only the presence or absence of a central -NH(2)-(+) (BBR3571 and analogs). Thus, closely-related compounds could be compared. Evidence for pre-association, presumably through electrostatic and hydrogen-bonding, was obtained from fluorescence and circular dichroism spectroscopy and Electrospray Ionization Mass Spectrometry (ESI-MS). In the case of those compounds containing Pt-Cl bonds, further reaction took place presumably through displacement by sulfur nucleophiles. The implications for protein pre-association and plasma stability of polynuclear platinum compounds are discussed.

Structural and mechanistic aspects of platinum anticancer agents

After the discovery of the anticancer activity of cisplatin many studies have focused on elucidating its mechanism of action. The antitumor effects of platinum complexes originate from their interaction with DNA, which causes interference with normal transcription or DNA replication. Pt–DNA adducts produced by cisplatin and many of its analogues are almost identical, and would explain their similar patterns of tumor sensitivity and susceptibility to resistance. However, platinum compounds bearing trans-amine (ammine) ligands, and those of multinuclear Pt complexes give rise to radically different DNA–Pt adducts. Platinum–sulfur interactions are associated with undesired phenomena such as resistance and toxicity. Modern multinuclear n.m.r. approaches are very powerful for the investigation of thermodynamics and kinetics of the reactions of metal compounds with biomolecules, and it is possible to study the coordination chemistry of platinum drugs under physiological relevant conditions. In this review biocoordination chemistry of platinum anticancer drugs and various mechanistic aspects related to the antitumor effects of platinum complexes have been explained.

Differential nephrotoxicity of cisplatin and a novel series of traditional Chinese medicine–platinum anticancer agents correlates with their chemical reactivity towards sulfur-containing nucleophiles

A series of novel traditional Chinese medicine-platinum compounds has been found to be active against a number of murine and human cancers both in vitro and in vivo. Their high potency and the lack of cisplatin cross-resistance are believed to be due to the inclusion of the protein phosphatase 2A-inhibiting demethylcantharidin in the novel structures. A simple reversed-phase high-performance liquid chromatographic method was developed and validated as a stability-indicating assay for the platinum compounds. Using cisplatin and carboplatin as reference compounds, the stability study agrees well with the literature-reported findings. The novel traditional Chinese medicine-platinum compounds were more stable than cisplatin in water and dextrose, but became unstable in normal saline, a characteristic similar to that of carboplatin. The developed assay was further applied to study the chemical reactivity of the novel platinum compounds towards physiologically important nucleophiles such as glutathione and cysteine. The novel compounds were considerably less reactive to the sulfur-containing nucleophiles than cisplatin. In-vitro cytotoxicity assay was performed in a porcine kidney LLC-PK1 cell line model to investigate the nephrotoxicity potential of the platinum compounds. The lower rate of hydrolysis and the decreased reactivity of the novel traditional Chinese medicine-platinum compounds towards sulfur-containing bionucleophiles appear to have reduced their toxicity when compared with cisplatin, yet the antitumor activities of the novel compounds have not been compromised.

The fate of platinum(II) and platinum(IV) anti-cancer agents in cancer cells and tumours

SRIXE mapping has been used to gain insight into the fate of platinum(II) and platinum(IV) complexes in cells and tumours treated with anticancer active complexes to facilitate the development of improved drugs. SRIXE maps were collected of thin sections of human ovarian (A2780) cancer cells treated with bromine containing platinum complexes, cis-[PtCl 2(3-Brpyr)(NH 3)] (3-Brpyr = 3-bromopyridine) and cis,trans,cis-[PtCl 2(OAcBr) 2(NH 3) 2] (OAcBr = bromoacetate), or a platinum complex with an intercalator attached cis-[PtCl 2(2-[(3-aminopropyl)amino]-9,10-anthracenedione)(NH 3)]. After 24 h the complexes appear to be localised in the cell nucleus with a lower concentration in the surrounding cytoplasm. In cells treated with cis-[PtCl 2(3-Brpyr)(NH 3)] the concentration of bromine was substantially higher than in control cells and the bromine was co-localised with the platinum consistent with the 3-bromopyridine ligand remaining bound to the platinum. The cells treated with cis,trans,cis-[PtCl 2(OAcBr) 2(NH 3) 2] also showed an increased level of bromine, but to a much lesser extent than for those treated with cis-[PtCl 2(3-Brpyr)(NH 3)] suggestive of substantial reduction of the platinum(IV) complex. Maps were also collected from thin sections of a 4T1.2 neo 1 mammary tumour xenograft removed from a mouse 3 h after treatment with cis,trans,cis-[PtCl 2(OH) 2(NH 3) 2] and revealed selective uptake of platinum by one cell.

Mechanism of the Membrane Interaction of Polynuclear Platinum Anticancer Agents. Implications for Cellular Uptake

The interaction between phospholipids and polynuclear platinum drugs was studied as a mechanism model for cellular uptake of anticancer drugs. The interaction was studied by differential scanning calorimetry (DSC), 31P nuclear magnetic resonance spectroscopy (NMR), inductively coupled plasma optical emission spectroscopy (ICP-OES), and electrospray ionization mass spectrometry (ESI-MS). The transition temperature, enthalpy, and entropy of negatively charged phospholipids DPPS, DPPA, and DPPG were changed upon reaction with the trinuclear platinum complex [{trans-PtCl(NH3)2}2mu-Pt(NH3)2{H2N(CH2)6NH2}2](NO3)4 (I, BBR3464) and the dinuclear analogue [{trans-PtCl(NH3)2}mu-{(NH2)(CH2)3NH2(CH2)4(NH2)}Cl3 (II, BBR3571). This suggests that these platinum complexes interacted not only with the phosphate headgroup but also with the region of the fatty acid tail of liposomes and finally changed the fluidity of the membrane. Both noncovalent (presumably electrostatic and hydrogen bonding) and covalent interactions were involved in the reactions of the negatively charged phospholipids DPPA, DPPS, and DPPG with the highly positively charged platinum complexes. In contrast, few differences were seen for the zwitterionic phospholipids DPPC and DPPE. The binding ratio of BBR3464 to DPPA liposomes was higher than the ratio of BBR3464 to DPPS liposomes, and similar differences were seen for BBR3571. The binding ratios of the platinum complexes to negatively charged phospholipids DPPA, DPPS, and DPPG were slightly lower in a 100 mM chloride solution than in a chloride-free solution. The binding of BBR3464 and BBR3571 with the liposomes was significantly stronger than that with cis-[PtCl2(NH3)2], cisplatin. ESI-MS confirmed that the products of the incubation of BBR3464 with DPPA and DPPS correspond to chloride displacement and formation of [Pt3(NH3)6{NH2(CH2)6NH2}2(DPPA)2]2+ (1) and [Pt3(NH3)6{NH2(CH2)6NH2}2(DPPS)2]2+ (2), respectively. Similar observations were made for BBR3571. 31P NMR spectra confirmed that the site of binding for DPPA was the phosphate oxygen, whereas for DPPS, a binding site of the nitrogen of the serine side chain is indicated. Noncovalent interactions were also confirmed by use of the analogue [{Pt(NH3)3}2mu-Pt(NH3)2{H2N(CH2)6NH2}2](NO3)6 (III, 0,0,0/t,t,t). The implications of these results for the mechanism of cellular uptake of polynuclear platinum complexes are discussed.

Platinum Complexes as Anticancer Agents

The application of inorganic chemistry to medicine is a rapidly developing field, and novel therapeutic and diagnostic metal complexes are now having an impact on medical practice. Advances in biocoordination chemistry are crucial for improving the design of compounds to reduce toxic side effects and understand their mechanisms of action. Cisplatin, as one of the leading metal-based drugs, is widely used in the treatment of cancer. Significant side effects and drug resistance, however, have limited its clinical applications. Biological carriers conjugated to cisplatin analogs have improved specificity for tumor tissue, thereby reducing side effects and drug resistance. Platinum complexes with distinctively different DNA binding modes from that of cisplatin also exhibit promising pharmacological properties. This review focuses on recent advances in developing platinum anticancer agents with an emphasis on platinum coordination complexes.

The β-Carboline Analog Mana-Hox Causes Mitotic Aberration by Interacting with DNA

Mana-Hox, an analog of beta-carbolines with anticancer activity, induces aberrant mitosis and delays mitotic exit. However, the cellular target is not known. In this study, we visualized the intracellular localization of Mana-Hox. Mana-Hox rapidly penetrated into cells (within 1 min) and concentrated on disorganized metaphase chromosomes after 13 hr of exposure. We demonstrated that Mana-Hox is a noncovalent DNA binder that can interact with DNA through intercalation and/or through minor groove binding. Furthermore, Mana-Hox also inhibits topoisomerase II relaxation activity in vitro, suggesting that Mana-Hox could perturb mitotic chromosome decatenation. Overall, Mana-Hox binding to DNA plays a critical role in the induction of aberrant mitosis and contributes to its anticancer activity.

In vitro and in vivo suppression of growth of hepatocellular carcinoma cells by novel traditional Chinese medicine–platinum anti-cancer agents

Protein phosphatase 2A (PP2A) is a new target for platinum (Pt)-based cancer chemotherapeutic agents. A series of novel Pt complexes containing demethylcantharidin, a modified component of a traditional Chinese medicine (TCM), [Pt(C8H8O5)(NH2R)2] 1-5 have been shown to inhibit PP2A both in its purified form and in cell homogenates. In this study, the potential efficacy of compounds 1-5 in suppressing the growth of PP2A-highly expressed liver cancer was evaluated. The in vitro anti-proliferative activity of compounds 1-5 was investigated in human hepatocellular carcinoma (HCC) cell lines using the MTT assay. Compounds 1-5 were about 2-20 and 20-200 times more potent than cisplatin and carboplatin, respectively, in SK-Hep1 and HepG2 cells. The in vivo anti-tumor efficacies of 1-5 were evaluated in a s.c. inoculated SK-Hep1 xenograft model in nude mice. Compounds 1-5 demonstrated definite in vivo activity (giving rise to an optimal %T/C as low as 14.5%) without inducing undue toxicity, contrasting the lack of activity of cisplatin and carboplatin. In a cisplatin-resistant model established in vivo in human HCC, compounds 1-5 could still elicit the same level of tumor growth suppression as in the control tumors, demonstrating the circumvention of cisplatin cross-resistance. An acute toxicity study in ICR mice showed that compounds 1-5 are not nephrotoxic at LD10. The high potency of the novel TCM-Pt compounds against liver cancer and the minimal toxicity suggest that they have significant potential to be developed into useful Pt-based anti-tumor drugs.

Novel Concepts in the Development of Platinum Antitumour Drugs: An Update

The limitations of cisplatin as an anticancer drug have stimulated the search for other antitumour-active platinum complexes with improved pharmacological properties. The two main goals in the search for new platinum anticancer agents are the reduction of the dose-limiting toxicities of cisplatin and the circumvention of cisplatin resistance. However, it should be noted that this has proven to be a difficult task. In fact, less than 1% of the thousand of platinum complexes tested for pre-clinical antitumour activity have entered clinical trials in the past 30 years. Nonetheless, at present, several new platinum complexes are in clinical trials; a proof of the continued belief that platinum complexes may still fulfil the needs for novel antitumour drugs. This review will focus on the three main innovative bioinorganic approaches found in the platinum anticancer-field, namely, (1) compounds with a slowed down kinetic of activation, (2) compounds with carrier ligands, and (3) compounds that bind differently to DNA as compared to cisplatin. In the latter class, special attention is paid to dinuclear and polynuclear platinum complexes. Keywords: platinum antitumor drugs, dna binding, decreased reactivity, carrier ligands

Multiple States of Stalled T7 RNA Polymerase at DNA Lesions Generated by Platinum Anticancer Agents

Transcription inhibition by DNA adducts of cisplatin is considered to be one of the major routes by which this anticancer drug kills cancer cells. Stalled RNA polymerases at platinum-DNA lesions evoke various cellular responses such as nucleotide excision repair, polymerase degradation, and apoptosis. T7 RNA polymerase and site-specifically platinated DNA templates immobilized on a solid support were used to study stalled transcription elongation complexes. In vitro transcription studies were performed in both a promoter-dependent and -independent manner. An elongation complex is strongly blocked by cisplatin 1,2-intrastrand d(GpG) and 1,3-intrastrand d(GpTpG) cross-links located on the template strand. Polymerase action is inhibited at multiple sites in the vicinity of the platinum lesion, the nature of which can be altered by the choice and concentration of NTPs. The [(1R,2R-diaminocyclohexane)Pt]2+ DNA adducts formed by oxaliplatin, which carries a stereochemically more demanding spectator ligand than the ammine groups in cisplatin, also strongly block the polymerase with measurable differences compared with cis-[(NH3)2Pt]2+ lesions. Elongation complexes stopped at sites of platinum damage were isolated and characterized. The stalled polymerase can be dissociated from the DNA by subsequent polymerases initiated from the same template. We also discovered that a polymerase stalled at the platinum-DNA lesion can resume transcription after the platinum adduct is chemically removed from the template.

An examination of the aquation kinetics and equilibria of dinuclear platinum anticancer agents

An examination of the aquation kinetics and equilibria of dinuclear platinum anticancer agents

XANES determination of the platinum oxidation state distribution in cancer cells treated with platinum(IV) anticancer agents.

Here we describe the use of X-ray absorption near edge spectroscopy (XANES) to provide information about the relative proportions of platinum(II) and platinum(IV) complexes by analyzing the XANES edge height. The intracellular reduction of platinum(IV) complexes in cancer cells has been observed directly, and the proportion of reduction after 2 h was found to correlate with the reduction potentials of the complexes.

New cytotoxic and water-soluble bis(2-phenylazopyridine)ruthenium(II) complexes.

New water-soluble bis(2-phenylazopyridine)ruthenium(II) complexes, all derivatives of the highly cytotoxic alpha-[Ru(azpy)(2)Cl(2)] (alpha denoting the coordinating pairs Cl, N(py), and N(azo) as cis, trans, cis, respectively) have been developed. The compounds 1,1-cyclobutanedicarboxylatobis(2-phenylazopyridine)ruthenium(II), alpha-[Ru(azpy)(2)(cbdca-O,O')] (1), oxalatobis(2-phenylazopyridine)ruthenium(II), alpha-[Ru(azpy)(2)(ox)] (2), and malonatobis(2-phenylazopyridine)ruthenium(II), alpha-[Ru(azpy)(2)(mal)] (3), have been synthesized and fully characterized. X-ray analyses of 1 and 2 are reported, and compound 1 is the first example in which the cbdca ligand is coordinated to a ruthenium center. The cytotoxicity of this series of water-soluble bis(2-phenylazopyridine) complexes has been determined in A2780 human ovarian carcinoma and A2780cisR, the corresponding cisplatin-resistant cell line. For comparison reasons, the cytotoxicity of the complexes alpha-[Ru(azpy)(2)Cl(2)], alpha-[Ru(azpy)(2)(NO(3))(2)], beta-[Ru(azpy)(2)Cl(2)] (beta indicating the coordinating pairs Cl, N(py), and N(azo) as cis, cis, cis, respectively), and beta-[Ru(azpy)(2)(NO(3))(2)] have been determined in this cell line. All the bis(2-phenylazopyridine)ruthenium(II) compounds display a promising cytotoxicity in the A2780 cell line (IC(50) = 0.9-10 microM), with an activity comparable to that of cisplatin and even higher than the activity of carboplatin. Interestingly, the IC(50) values of this series of ruthenium compounds (except the beta isomeric compounds) are similar in the cisplatin-resistant A2780cisR cell line compared to the normal cell line A2780, suggesting that the activity of these compounds might not be influenced by the multifactorial resistance mechanism that affect platinum anticancer agents.

Pharmaceutical Development of a Parenteral Lyophilized Formulation of the Investigational Polymer-Conjugated Platinum Anticancer Agent AP 5280

AP 5280 is a novel polymer-conjugated platinum anticancer agent showing promising in vitro and in vivo activity against solid tumors. The aim of this study was to develop a parenteral pharmaceutical dosage form for phase I clinical trials. AP 5280 drug substance was characterized by using a wide range of analytical techniques and showed excellent solubility in water. However, as aqueous solutions of AP 5280 proved to be labile upon sterilization by moist heat, it was decided to develop a lyophilized dosage form. Initially, glass vials were used as primary packaging, but this led to a high breakage rate, which could be completely prevented by the use of CZ® resin vials. Stability studies to date show that the lyophilized product in glass vials is stable for at least 12 months when stored at 2–8°C in the dark and the lyophilized product in CZ resin vials is stable for at least 6 months under these conditions. Photostability testing revealed photolability of AP 5280 drug substance and lyophilized product in both types of primary container, necessitating storage in the dark. The first clinical experiences indicate that the proposed formulation is fully applicable for use in the clinical setting.

The nature of the DNA template (single- versus double-stranded) affects the rate of aquation of a dinuclear Pt anticancer drugElectronic supplementary information (ESI) available: experimental conditions for the NMR reactions, the models used for the kinetic fits and [1H,15N] HSQC NMR spectra of the final products from reactions of 1 with the single strand (I) (before and after addition of

The rate of aquation of a dinuclear platinum anticancer agent is altered in the presence of template DNA with enhancement of hydrolysis in the presence of single-stranded over double-stranded DNA, emphasising how the alteration of chemical properties of small molecules in the presence of large host interactions is also dependent on the conformation and nature of that host.

Stability and compatibility of the investigational polymer-conjugated platinum anticancer agent AP 5280 in infusion systems and its hemolytic potential.

AP 5280 is a novel polymer-conjugated platinum anticancer agent currently undergoing phase I clinical trials. It is pharmaceutically formulated as a lyophilized product containing 200 mg platinum per dosage unit. The aim of this study was to determine the reconstitution and dilution fluid of choice, and to investigate the stability and compatibility of AP 5280 in solution under different storage conditions and with several container materials. Furthermore, the hemolytic potential of AP 5280 infusion solution was investigated. AP 5280 slowly released small platinum species in all solutions, although this release was enhanced in normal saline. Accordingly, 5% dextrose in water (D W) was selected for reconstitution and dilution of AP 5280. Container material [glass or polyvinylchloride (PVC)] did not influence the stability of AP 5280 in solution. Storage at refrigerated temperature (2-8 degrees C) marginally decreased the release rate of liberated platinum. The infusion solutions are compatible with the PVC infusion system and do not cause hemolysis. In conclusion, AP 5280 lyophilized product should be reconstituted and diluted to infusion concentration with D W, and administered within 8 h after preparation to ensure that less than 1.0% of the total platinum concentration is present as liberated platinum.

Novel Concepts in the Development of Platinum Antitumor Drugs

The limitations of cisplatin as an anticancer drug have stimulated the search for other antitumor-active platinum complexes with improved pharmacological properties. The two main goals in the search for new platinum anti-cancer agents are the reduction of the dose-limiting toxicities of cisplatin and the circumvention of cisplatin resistance. However, it should be pointed out that this has proven to be a difficult task. In fact, less than 1% of the thousand of platinum complexes tested for pre-clinical antitumor activity have entered clinical trials in the past 30 years. Nonetheless, right now, several new platinum complexes are in clinical trials, a proof of the continued belief that platinum complexes may still fulfil the needs for novel antitumor drugs. This review will focus on the three main innovative approaches found in the platinum anticancer-field, namely, (1) compounds with decreased reactivity against nucleophiles, (2) compounds with carrier ligands, and (3) compounds which bind differently to DNA as compared to cisplatin. In the latter class, special attention is paid to dinuclear and polinuclear platinum complexes.

Mechanism of action of non-cisplatin type DNA-targeted platinum anticancer agents: DNA interactions of novel acridinylthioureas and their platinum conjugates

The DNA binding of two novel acridinylthioureas, ACR-NH-(CH 2) 2-C(S)-NHCH 3 ( 1) and ACR-N(CH 3)-C(S)-NHCH 3 ( 3), and their platinum conjugates 4 and 5—derived from [PtCl 2(en)]—was studied in cell-free model systems using various physico-chemical and biophysical methods. These included: spectrophotometric drug–DNA titrations, ethidium−DNA fluorescence quenching, competitive drug displacement, high-resolution NMR spectroscopy, and unwinding of plasmid DNA monitored by agarose gel electrophoresis. The acridinium cation of 1 showed strong binding to native DNA with K i =1.5×10 6 M −1 and an excluded site size ( n) of 2 bp (McGhee–von Hippel fits of absorbance data). Compound 3 showed no measurable association with DNA. Binding of 1 was an order of magnitude stronger than that of simple 9-methylaminoacridine ( 2). In alternating copolymers, 1 exhibited slight AT preference. In poly(dA-dT) 2, enhanced association was accompanied by an increased binding site (approximately 3 bp), while parameters in poly(dG-dC) 2 were consistent with classical intercalation. Displacement of 1 by distamycin from calf thymus DNA was suggestive of non-intercalating thiourea in 1 being located in the minor groove of the duplex. 1 H NMR data of d(GGAGCTCC) 2 modified with 1 indicated intercalative binding of planar acridine, based on upfield shifts of aromatic proton signals relative to those in unbound 1 (Δδ≈−0.5 to −1 ppm). Finally, 4 and 5 were found to unwind negatively supercoiled pUC19 plasmid by 21° and 7° per adduct, respectively (electrophoretic gel mobility assays). The difference in DNA binding modes of 4 and 5 is discussed as the ultimate source of the distinctly different biological activities of the conjugates.