Improving platinum(II)-based anticancer drug delivery using cucurbit[n]urils

Despite the synthesis of hundreds of new platinum(II) and platinum(IV)-based complexes each year as potential anticancer drugs, only three have received world-wide approval: cisplatin, carboplatin and oxaliplatin. The next big advance in platinum-based chemotherapy is not likely to come from the development of new drugs, but from the controlled and targeted delivery of already approved drugs or those in late stage clinical trials. Encapsulation of platinum drugs inside macromolecules has already demonstrated promise, and encapsulation within cucurbit[n]urils has shown particular potential. Partial or full encapsulation within cucurbit[n]urils provides steric hindrance to drug degradation by peptides and proteins, and the use of different sized cucurbit[n]urils allows for the tuning of drug release rates, cytotoxicity and toxicity.     

Ensembles of carboxymethyl cyclodextrins on cationic liposomes as highly efficient nanocontainers for the delivery of hydrophobic compounds

The increase of payload is one of the key tasks in creation of nanocontainers for the delivery of bioactive substances (BAS). In this work the adsorption of anionic carboxymethyl cyclodextrins (CMCDs) on the surface of cationic liposomes was studied as mechanism of formation of capacious nanocontainers for the encapsulation and delivery of hydrophobic BAS. The formation and physico-chemical characteristics of complexes were studied by means of laser microelectrophoresis, dynamic light-scattering, conductometry and atomic force microscopy (AFM). As a model, bioactive molecule hydrophobic curcumin was chosen for the investigation. The encapsulation of curcumin was controlled by UV–Vis spectrometry. Interaction of CMCDs/liposomes complexes with model cell membranes was visualized by fluorescent microscopy. Finally, cytotoxicity of nanocontainers was studied by MTT-test. It was estimated that colloid stable complexes with net positive charge could contain up to 2.5÷5 CMCD molecules per one cationic lipid. Incorporation of curcumin in CMCDs does not change the character of interaction of oligosaccharides with liposomal membranes of individual liposome. CMCDs/liposomes complexes adsorb on model cell membranes without significant loss of CMCD molecules. This fact in addition to low cytotoxicity of cationic CMCDs/liposomes complexes demonstrates potential of their application as nanovehicles for the delivery of BAS.     

Nanocapsules: a novel lipid formulation platform for platinum-based anti-cancer drugs

Platinum-based anti-cancer agents have been used for many years to treat many different types of cancer. However, the efficacy of these drugs is limited by serious side effects. One of the strategies to reduce the side effects is encapsulation of the drug in a lipid formulation. Recently, we discovered a novel method for the efficient encapsulation of cisplatin in a lipid formulation. The method is unique in that it does not generate conventional liposomes but nanocapsules: small aggregates of solid cisplatin covered by a lipid bilayer. Also carboplatin, a cisplatin-derived anti-cancer drug with different chemical properties, can be efficiently encapsulated by a similar method. The encapsulation in nanocapsules dramatically improves the in vitro cytotoxicity of the platinum drugs. Our results hold the promise that the nanocapsule technology could prove successful in the efficient encapsulation of many other (platinum-based) drugs, and thereby improve their therapeutic index and profile in vivo.     

Nanocapsules: A Novel Lipid Formulation Platform for Platinum-based Anti-cancer Drugs

Platinum-based anti-cancer agents have been used for many years to treat many different types of cancer. However, the efficacy of these drugs is limited by serious side effects. One of the strategies to reduce the side effects is encapsulation of the drug in a lipid formulation. Recently, we discovered a novel method for the efficient encapsulation of cisplatin in a lipid formulation. The method is unique in that it does not generate conventional liposomes but nanocapsules: small aggregates of solid cisplatin covered by a lipid bilayer. Also carboplatin, a cisplatin-derived anti-cancer drug with different chemical properties, can be efficiently encapsulated by a similar method. The encapsulation in nanocapsules dramatically improves the in vitro cytotoxicity of the platinum drugs. Our results hold the promise that the nanocapsule technology could prove successful in the efficient encapsulation of many other (platinum-based) drugs, and thereby improve their therapeutic index and profile in vivo.     

Reactivity of platinum-based antitumor drugs towards a Met- and His-rich 20mer peptide corresponding to the N-terminal domain of human copper transporter 1

Cellular uptake of platinum-based antitumor drugs is a critical step in the mechanism of the drug action and associated resistance, and deeper understanding of this step may inspire development of novel methods for new drugs with reduced resistance. Human copper transporter 1 (hCtr1), a copper influx protein, was recently found to facilitate the cellular entry of several platinum drugs. In the work reported here, we constructed a Met- and His-rich 20mer peptide (hCtr1-N20) corresponding to the N-terminal domain of hCtr1, which is the essential domain of hCtr1 for transporting platinum drugs. The interactions of the peptide with cisplatin and its analogues, including transplatin, carboplatin, oxaliplatin, and [Pt(l-Met)Cl₂], were explored at the molecular level. Electrospray ionization (ESI) mass spectrometry (MS) data revealed that all of the platinum(II) complexes used in present study can bind to hCtr1-N20 in 1:1 and 2:1 stoichiometry. Four Met residues should be involved in binding to cis-platinum complexes on the basis of the tandem MS spectrometry and previously reported data. Time-dependent 2D [¹H,¹⁵N] heteronuclear single quantum coherence NMR spectra indicate the reaction of cisplatin with hCtr1-N20 is a stepwise process. The intermediate, however, is transient, which is consistent with the ESI-MS results. Time-dependent ESI-MS data revealed that the geometry and the properties of both the leaving and the nonleaving groups of platinum(II) complexes play essential roles in controlling the reactivity and formation of the final products with hCtr1-N20.     

肠道病原菌侵袭素在药物纳米载体中的应用

口服给药是药物递送系统中的优选途径。然而,在通过胃肠道时,肠细胞的低渗透性经常会阻碍药物的有效递送。包囊药物能够解决这一问题的关键,取决于其中的细胞侵袭性靶向基团包裹的纳米颗粒系统。这种药物递送系统的侵入特性是由细菌侵袭素的关键成分提供,这些成分具有快速调节药物穿越肠细胞的作用,从而促进宿主细胞对药物的有效吸收。此综述重点阐述细菌侵袭系统,对合适的侵袭素分别从功能和分子结构、作为靶向药物的相对价值以及在使用过程中可能存在的误区依次进行探讨。此外,对口服给药方法的改进和未来前景也进行了讨论。     

2-Deoxyglucose conjugated platinum (II) complexes for targeted therapy: design,synthesis, and antitumor activity

Malignant neoplasms exhibit an elevated rate of glycolysis over normal cells. To target the Warburg effect, we designed a new series of 2-deoxyglucose (2-DG) conjugated platinum (II) complexes for glucose transporter 1 (GLUT1)-mediated anticancer drug delivery. The potential GLUT1 transportability of the complexes was investigated through a comparative molecular docking analysis utilizing the latest GLUT1 protein crystal structure. The key binding site for 2-DG as GLUT1’s substrate was identified with molecular dynamics simulation, and the docking study demonstrated that the 2-DG conjugated platinum (II) complexes can be recognized by the same binding site as potential GLUT1 substrate. The conjugates were synthesized and evaluated for in vitro cytotoxicity study with seven human cancer cell lines. The results of this study revealed that 2-DG conjugated platinum (II) complexes are GLUT1 transportable substrates and exhibit improved cytotoxicities in cancer cell lines that over express GLUT1 when compared to the clinical drug, Oxaliplatin. The correlation between GLUT1 expression and antitumor effects are also confirmed. The study provides fundamental information supporting the potential of the 2-DG conjugated platinum (II) complexes as lead compounds for further pharmaceutical R&D.     

Novel cyclopalladated and coordination palladium and platinum complexes derived from alpha-diphenyl ethanedione bis(thiosemicarbazones): structural studies and cytotoxic activity against human A2780 and A2780cisR carcinoma cell lines

The preparation of new palladium(II) and platinum(II) complexes derived from alpha-diphenyl ethanedione bis(thiosemicarbazone), 1, and alpha-diphenyl ethanedione bis(4-ethylthiosemicarbazone), 2, is described. The palladium complexes 3 and 4 and platinum complexes 5 and 6 have been characterized by elemental analyses, fast atom bombardment mass spectrometry (FAB(+)) and spectroscopic studies (IR, (1)HNMR). The crystal and molecular structures of the dimeric cyclopalladated compound 4 and the mononuclear platinum complex 6 have been determined by single crystal X-ray diffraction. The cytotoxic activity of the free ligands and palladium and platinum complexes against human A2780 and A2780cisR (acquired resistance to cisplatin) epithelial ovarian carcinoma cells lines is also reported. The IC(50) values for compounds 1, 5 and 6 were found to be higher than that of cisplatin but the maximum antiproliferative activity was similar. Furthermore, the compounds largely retain their activity in the A2780cisR cell line, having a much better resistance factor than cisplatin in the pair of cell lines tested.     

Cis-platinum (II) complexes of carboxymethyl-dextran as potential antitumor agents. II. In vitro and in vivo activity

Cis-diamminedichloro platinum (II) (cis-DDP) and cis-diamminediaquo platinum (II) nitrate (cis-aq) were complexed to a macromolecular carrier carboxymethyl dextran (CM-dex). Two carriers were used in this study, one derived from dex-T-10 (Mr-10000) and the other from dex-T-40 (Mr-40000). The two platinum (II) drugs formed soluble complexes with both carriers. Uncomplexed and complexed drugs were tested and found to be cytotoxic in vitro against 5 murine and 2 human derived tumor cell lines. The two free platinum (II) drugs were cytotoxic against these cells to a similar extent. In comparison to the free drugs the complexes were somewhat less active, up to 3 fold, against murine 38C-13, L1210, EL-4 and RDM-4 leukemias, as well as against human HeLa and osteogenic sarcoma, and as active as the free drugs against murine F9 embryonal carcinoma. There were no major differences in the in vitro cytotoxic activity between CM-dex T-10 and CM-dex T-40 complexes. Differences due to the molecular size of the carrier were observed in vivo: The CM-dex T-10 complexes were significantly less toxic than the free drugs, whereas the reduction of toxicity by complexing to CM-dex T-40 was less profound. As for the efficacy, when tested in vivo against a cis-DDP sensitive tumor (F9) the T-40 complexes were equally or even more effective than the respective free drugs. The T-10 complexes were less effective than the free drugs at equal drug doses but their effectivity increased at increasing drug levels. These complexes were, however, very effective in inhibiting tumor growth upon repeated injections, leading to 100% survival.     

Quantitative measurement of the reduction of platinum(IV) complexes using X-ray absorption near-edge spectroscopy (XANES)

The platinum(II) drugs cisplatin, carboplatin and oxaliplatin are usefully employed against a range of malignancies, but toxicities and resistance have spurred the search for improved analogs. This has included investigation of the platinum(IV) oxidation state, which provides greater kinetic inertness. It is generally accepted that Pt(IV) complexes must be reduced to Pt(II) for activation. As such, the ability to monitor reduction of Pt(IV) complexes is critical to guiding the design of candidates, and providing mechanistic understanding. Here we report in full that the white line height of X-ray absorption near-edge spectra (XANES) of Pt complexes, normalized to the post-edge minima, can be used to quantitatively determine the proportion of each oxidation state in a mixture. A series of Pt(IV) complexes based on the Pt(II) complexes cisplatin and transplatin were prepared with chlorido, acetato or hydroxido axial ligands, and studies into their reduction potential and cytotoxicity against A2780 human ovarian cancer cells were performed, demonstrating the relationship between reduction potential and cytotoxicity. Analysis of white line height demonstrated a clear and consistent difference between Pt(II) (1.52 ± 0.05) and Pt(IV) (2.43 ± 0.19) complexes. Reduction of Pt(IV) complexes over time in cell growth media and A2780 cells was observed by XANES, and shown to correspond with their reduction potentials and cytotoxicities. We propose that this method is useful for monitoring reduction of metal-based drug candidates in complex biological systems.     

Meeting report on 8th International Symposium on Platinum and Other Metal Coordination Compounds in Cancer Chemotherapy

The platinum-based drugs, cisplatin and carboplatin, represent major agents in the chemotherapeutic treatment of a variety of types of cancer. Novel, "third-generation" agents aimed at broadening the clinical activity of this class of drug are currently undergoing clinical evaluation. These include oxaliplatin, ZD0473 and BBR3464. Clinical trials and preclinical studies are also being conducted with liposomal (SPI-077 and L-NDDP) and polymeric platinum complexes (linked to HPMA or albumin). Combination studies of cisplatin/carboplatin with other anticancer drugs such as gemcitabine and UCN-01 (7-hydroxystaurosporine) and agents designed to reduce platinum drug toxicities (e.g., BNP-7787, DIMESNA) are ongoing. Preclinically, there is interest in trans platinum complexes, terpyridine platinum(II) complexes and other metal-containing agents (ruthenium and gold).     

Synthesis and antitumour activity of platinum(II) and platinum(IV) complexes containing ethylenediamine-derived ligands having alcohol, carboxylic acid and acetate substituents. Crystal and molecular structure of [PtL4CL2].H20 where L4 is ethylenediamine-N,N'-diacetate

Several cisplatin analogues of ethylenediamine-derived ligands containing alcohol, carboxylic acid and acetate substituents have been prepared and characterised. Oxidation of some of these square planar platinum(II) complexes using aqueous hydrogen peroxide gave octahedral platinum(IV) complexes, containing trans hydroxo ligands. Acetylation of the hydroxo ligands was achieved by reaction with acetic anhydride, giving complexes which are analogues of the antitumour drug, JM-216. Oxidation of the complex [Pt(H2L4)Cl2], where H2L4 is ethylenediamine-N,N'-diacetic acid, with H2O2 gave the platinum(IV) complex [PtL4Cl2].H2O in which L4 is tetradentate as shown by a crystal and molecular structure. This complex was previously reported to be [Pt(HL4)(OH)Cl2] in which HL4 is tridentate. Several of the complexes were tested for antitumour activity against five human ovarian carcinoma cell lines. IC50 values range from 4.0 microM for cis,trans-PtCl2(OH)2(NH2CH2CH2NHCH2CH2OH) against the CH1 cell line to >25 microM indicating moderate to low activity relative to other platinum complexes.     

Mini-review: discovery and development of platinum complexes designed to circumvent cisplatin resistance

The discovery and development of new platinum-containing anticancer drugs have represented an integral part of anticancer drug development at the Institute of Cancer Research, Sutton, over almost 20 years. As part of a collaboration with chemists at Johnson Matthey, later AnorMED, four major new classes of platinum drug have been discovered, three of which have entered clinical trial. Earlier studies led to the clinical development of the less toxic analogue carboplatin and JM216, the first orally administerable platinum drug. In recent years, the focus has been on two lead complexes designed to overcome the major mechanisms of tumour resistance to cisplatin: JM335 (trans-ammine (cyclohexylaminedichlorodihydroxo) platinum(IV)), an active trans platinum complex; and ZD0473 (cis-amminedichloro(2-methylpyridine) platinum(II)), a sterically hindered complex shown to be less reactive towards thiol-containing molecules than cisplatin. JM335 shows some circumvention of acquired cisplatin resistance in vitro and exhibits unique cellular pharmacological properties in comparison to cisplatin or its cis-isomer in terms gene-specific repair of adducts on DNA and the rate of induction of apoptosis. ZD0473 is now in phase I clinical trial. Myelosuppression is the dose-limiting toxicity at a dose of 130 mg/m2 given i.v. every 3 weeks and there has been evidence of antitumour activity. ZD0473-resistant human ovarian carcinoma cell lines have been established in vitro. Some mechanisms of resistance common to those described for cisplatin (decreased drug uptake, increased glutathione) have been observed plus, in one cell line, increased BCL2 levels and loss of the DNA mismatch repair protein MLH1.     

Tumour-inhibiting platinum complexes—state of the art and future perspectives

Thirty years after the onset of the first clinical studies with cisplatin, the development of antineoplastic platinum drugs continues to be a productive field of research. This article reviews the current preclinical and clinical status, including a discussion of the molecular basis for the activity of the parent drug cisplatin and platinum drugs of the second and third generation, in particular their interaction with DNA. Further emphasis is laid on the development of third generation platinum drugs with activity in cisplatin-resistant tumours, particularly on chelates containing 1,2-diaminocyclohexane (DACH) and on the promising and more recently evolving field of non-classic (trans- and multinuclear) platinum complexes. The development of oral platinum drugs and drug targeting strategies using liposomes, polymers or low-molecular-weight carriers in order to improve the therapeutic index of platinum chemotherapy are also covered. Electronic Publication     

Enhancing lipophilicity as a strategy to overcome resistance against platinum complexes?

Decreased influx represents one of the major resistance mechanisms of platinum complexes. In order to address the question if this mechanism of resistance can be overcome by enhancing the lipophilicity of platinum complexes, we investigated the influence of lipophilicity on cellular accumulation and cytotoxicity in a panel of oxaliplatin analogues with different carrier ligands. Cellular accumulation, DNA platination and cytotoxicity were measured in a cisplatin-sensitive and -resistant ovarian carcinoma (A2780/A2780cis) and in an oxaliplatin-sensitive and -resistant ileocecal colorectal adenocarcinoma (HCT-8/HCT-8ox) cell line pair. Platinum concentrations were determined by flameless atomic absorption spectrometry or adsorptive stripping voltammetry. Passive diffusion represented the main influx mechanism of oxaliplatin analogues during the first minutes of incubation as indicated by a correlation between lipophilicity and early influx rate. Afterwards, the predominant influx mechanism was lipophilicity-independent. More lipophilic complexes showed a reduced cytotoxic activity, although the early influx rate was increased. The resistance profiles of the two cell line pairs were found to be different: HCT-8ox cells were less resistant against more lipophilic complexes, whereas A2780cis cells exhibited a comparable degree of resistance against all investigated compounds. However, the reduction in resistance factor of HCT-8ox cells cannot be explained by increased influx suggesting that other resistance mechanisms are circumvented upon exposure to more lipophilic compounds. Though resistance against more lipophilic platinum complexes analogues is lower we conclude that enhancing lipophilicity is not a successful strategy to overcome platinum resistance as higher lipophilicity is also associated with lower cytotoxic activity.     

The influence of temperature on antiproliferative effects, cellular uptake and DNA platination of the clinically employed Pt(II)-drugs

Cellular uptake of a drug is one of the most important factors influencing its pharmacodynamics and pharmacokinetics. Our laboratory has previously studied platinum uptake following cisplatin, carboplatin and oxaliplatin treatment at sub-lethal doses of selected tumour cell lines. Here we report on the influence of temperature on dose-dependent antiproliferative effects, cellular uptake and DNA platination of these platinum-based drugs tested on MCF-7 human mammary carcinoma cell line. Inductively coupled plasma-mass spectrometry (ICP-MS) technique has been chosen to perform Pt determinations on cells treated with drug concentrations similar with those usually found in vivo in human plasma. The high sensitivity and analytical rapidity of this technique made possible to carry out a very large amount of Pt determinations (about 300) necessary for this study. Hyperthermia (43 degrees C) proved a synergistic effect with cisplatin on cell growth inhibition, while only an additive effect was demonstrated for carboplatin and oxaliplatin. This behaviour might be explained by the higher DNA platination ratio between data at 43 and 37 degrees C of cisplatin with respect to those of carboplatin and oxaliplatin.     

Synthesis and evaluation of bi-functional 7-hydroxycoumarin platinum(IV) complexes as antitumor agents

A series of bi-functional 7-hydroxycoumarin platinum(IV) complexes were synthesized, characterized, and evaluated for antitumor activities. The 7-hydroxycoumarin platinum(IV) complexes display moderate to effective antitumor activities toward the tested cell lines and show much potential in overcoming drug resistance of platinum(II) drugs. In reducing microenvironment, the title compounds could be reduced to platinum(II) complex accompanied with two equivalents of coumarin units. By a unique mechanism, the 7-hydroxycoumarin platinum(IV) complex attacks DNA via the released platinum(II) compound, meanwhile it also inhibits the activities of cyclooxygenase by coumarin fragment. This action mechanism might be of much benefit for reducing tumor-related inflammation in the progress of inhibiting tumor proliferation and overcoming cisplatin resistance. The incorporation of 7-hydroxycoumarin leads to significantly enhanced platinum accumulation in both whole tumor cells and DNA. The HSA interaction investigation reveals that the tested coumarin platinum(IV) compound could effectively combine with HSA via van der Waals force and hydrogen bond.     

Intracellular protein binding patterns of the anticancer ruthenium drugs KP1019 and KP1339

The ruthenium compound KP1019 has demonstrated promising anticancer activity in a pilot clinical trial. This study aims to evaluate the intracellular uptake/binding patterns of KP1019 and its sodium salt KP1339, which is currently in a phase I–IIa study. Although KP1339 tended to be moderately less cytotoxic than KP1019, IC₅₀ values in several cancer cell models revealed significant correlation of the cytotoxicity profiles, suggesting similar targets for the two drugs. Accordingly, both drugs activated apoptosis, indicated by caspase activation via comparable pathways. Drug uptake determined by inductively coupled plasma mass spectrometry (ICP-MS) was completed after 1 h, corresponding to full cytotoxicity as early as after 3 h of drug exposure. Surprisingly, the total cellular drug uptake did not correlate with cytotoxicity. However, distinct differences in intracellular distribution patterns suggested that the major targets for the two ruthenium drugs are cytosolic rather than nuclear. Consequently, drug–protein binding in cytosolic fractions of drug-treated cells was analyzed by native size-exclusion chromatography (SEC) coupled online with ICP-MS. Ruthenium–protein binding of KP1019- and KP1339-treated cells distinctly differed from the platinum binding pattern observed after cisplatin treatment. An adapted SEC-SEC-ICP-MS system identified large protein complexes/aggregates above 700 kDa as initial major binding partners in the cytosol, followed by ruthenium redistribution to the soluble protein weight fraction below 40 kDa. Taken together, our data indicate that KP1019 and KP1339 rapidly enter tumor cells, followed by binding to larger protein complexes/organelles. The different protein binding patterns as compared with those for cisplatin suggest specific protein targets and consequently a unique mode of action for the ruthenium drugs investigated.     

In vitro cytotoxicity study on platinum (II) complexes with epoxysuccinates as leaving groups

A series of novel cisplatin-type platinum complexes were designed, characteristic of epoxysuccinates as leaving groups. The pertinent compounds were prepared and characterized by IR, (1)H NMR, and ESI-MS spectra with elementary analyses. The in vitro cytotoxic activities of compounds toward SPC-A1 human lung adenocarcinoma cell line and BGC823 human stomach adenocarcinoma cell line were determined. Biological tests have confirmed that complexes containing 4R,5R-DMID [abbreviation of (4R,5R)-4,5-bis (aminomethyl)-2-isopropyl-1,3-dioxolane] as carrier ligands have greater cytotoxicity toward tumor cells than the corresponding compounds with other carrier ligands. Most platinum complexes with trans-epoxysuccinates usually have higher cytotoxicity than those with cis-epoxysuccinates. Complex 4a shows the most effective among those tested platinum complexes in both cell lines, and its cytotoxicity approached that of cisplatin.     

Lipospheres and pro-nano lipospheres for delivery of poorly water soluble compounds

Lipospheres are a drug encapsulation system composed of water dispersible solid microparticles of particle size between 0.01 and 100 μm in diameter with a solid hydrophobic lipid core stabilized by a layer of phospholipid molecules embedded in their surface. The bioactive compound is dissolved or dispersed in the solid lipid matrix of the internal core. Since lipospheres were introduced in the beginning of the 1990s, they have been used for the delivery of multiple types of drugs by various routes of administration. Later, a self-assembling pro-nano lipospheres (PNL) encapsulation system was developed for oral drug delivery. Lipospheres have several advantages over other delivery systems, such as better physical stability, low cost of ingredients, ease of preparation and scale-up, high dispersibility in an aqueous medium, high entrapment of hydrophobic drugs, controlled particle size, and extended release of entrapped drug after administration, from a few hours to several days. This review article focuses on updated information on several aspects of lipospheres and PNL, including preparation techniques, physicochemical properties and in vitro evaluation methods. Additionally, it covers lipospheres and PNL utilization for oral, ocular, and parenteral delivery, with special attention to unique considerations and aspects for each route of administration.