Can the Self-Assembling of Dicarboxylate Pt(IV) Prodrugs Influence Their Cell Uptake?

The possibility of spontaneous self-assembly of dicarboxylato Pt(IV) prodrugs and the consequences on their uptake in cancer cells have been evaluated in different aqueous solutions. Four Pt(IV) complexes, namely, (OC-6-33)-diacetatodiamminedichloridoplatinum(IV), Ace, (OC-6-33)-diamminedibutanoatodichloridoplatinum(IV), But, (OC-6-33)-diamminedichloridodihexanoatoplatinum(IV), Hex, and (OC-6-33)-diamminedichloridodioctanoatoplatinum(IV), Oct, have been dispersed in i) milliQ water, ii) phosphate buffered saline, and iii) complete cell culture media (RPMI 1640 or DMEM) containing fetal bovine serum (FBS). The samples have been analyzed by dynamic light scattering (DLS) to measure the size and distribution of the nanoparticles possibly present. The zeta potential offered an indication of the stability of the resulting aggregates. In the case of the most lipophilic compounds of the series, namely, Oct and to a lesser extent Hex, the formation of nanosized aggregates has been observed, in particular at the highest concentration tested (10 µM). The cell culture media had the effect to disaggregate these nanoparticles, mainly by virtue of their albumin content, able to interact with the organic chains via noncovalent (hydrophobic) interactions. For Oct, at the highest concentration employed for the uptake tests (10 µM), the combination between passive diffusion and endocytosis of the self-assembled nanoparticles makes the cellular uptake higher than in the presence of passive diffusion only. During the study of cellular uptake on A2780 ovarian cancer cells pretreated with cytochalasin D, a statistically significant inhibition of endocytosis was observed for Oct. In these experimental conditions, the relationship between uptake and lipophilicity becomes almost linear instead of exponential. Since Oct anticancer prodrug is active at nanomolar concentrations, where the aggregation in culture media is almost abolished, this phenomenon should not significantly impact its antiproliferative activity.

Pt(iv) complexes based on cyclohexanediamines and the histone deacetylase inhibitor 2-(2-propynyl)octanoic acid: synthesis, characterization, cell penetration properties and antitumor activity.

The Pt(iv) complexes based on (SP-4-2)-dichlorido(cyclohexane-1,4-diamine)platinum(ii) (kiteplatin) and the histone deacetylase inhibitor 2-(2-propynyl)octanoic acid (POA) were investigated. Since POA contains a chiral carbon, all the possible Pt(iv) isomers were prepared and characterized, and their antiproliferative activity on six cancer cell lines was compared with that of the corresponding Pt(iv) complexes containing the cyclohexane-1R,2R-diamine equatorial ligand. To justify the very good antiproliferative activity (nanomolar IC50), the polarity, lipophilicity, permeability, and cell accumulation of the complexes were studied. Overall, the two series of Pt(iv) complexes showed similar cell penetration properties, being significantly better than that of the Pt(ii) reference compounds. Finally, a representative compound of the whole set of complexes (i.e., that based on cyclohexane-1R,2R-diamine and racemic POA) was tested in vivo on mice bearing Lewis lung carcinoma, showing good tumor growth inhibition with negligible body weight loss.

Cis,cis,trans-[PtIVCl2(NH3)2(perillato)2], a dual-action prodrug with excellent cytotoxic and antimetastatic activity.

Two Pt(iv) conjugates containing one or two molecules of perillic acid (4-isopropenylcyclohexene-1-carboxylic acid), an active metabolite of limonene, were synthesized both with traditional and microwave-assisted methods and characterized. Their antiproliferative activity was tested on a panel of human tumor cell lines. In particular, cis,cis,trans-[PtIVCl2(NH3)2(perillato)2] exhibited excellent antiproliferative and antimetastatic activity on A-549 lung tumor cells at nanomolar concentrations. A number of in vitro biological tests were performed to decipher some aspects of its mechanism of action, including transwell migration and invasion as well as wound healing assay.

New Platinum-Based Prodrug Pt(IV)Ac-POA: Antitumour Effects in Rat C6 Glioblastoma Cells.

Gliomas are the most frequent primary tumours of the nervous system, characterised by high degree of malignancy, widespread invasion and high-rate proliferation. Cisplatin and analogue are currently employed in clinical trials as active chemotherapeutic agents for the systemic treatment of this type of malignancy. Despite therapy benefits, clinical use of these agents is hampered by severe side effects including neurotoxicity. Therefore, the aim of the present study was to analyse the effect of a new compound of platinum(IV) conjugate, named Pt(IV)Ac-POA, which can generate a synergistic antineoplastic action when released along with cisplatin, after a specific reduction reaction within tumour cells. To assess the effects of the novel compound on rat C6 glioma cells, cell cycle and cell death activation analyses were carried out using flow cytometry. Morphological changes and activation of different cell death pathways were evaluated by both transmission electron microscopy and immunofluorescence microscopy. Protein expression was investigated by western blotting analysis. The novel compound Pt(IV)Ac-POA, bearing as axial ligand (2-propynyl)octanoic acid (POA), which is a histone deacetylase inhibitor (HDACi), acts as a prodrug in tumour cells, inducing cell death through different pathways at a concentration lower than those tested for other platinum analogues. The current results showed that Pt(IV)Ac-POA could represent a promising improvement of Pt-based chemotherapy against gliomas, either inducing a chemosensitisation and reducing chemoresistance.

Antiproliferative Activity of Pt(IV) Conjugates Containing the Non-Steroidal Anti-Inflammatory Drugs (NSAIDs) Ketoprofen and Naproxen †.

Cisplatin and several non-steroidal anti-inflammatory drugs (NSAIDs) have been proven to act synergistically or at least additively on several tumor cell lines. Dual-action cisplatin-based Pt(IV) combos containing ketoprofen and naproxen offer good antiproliferative performance on a panel of human tumor cell lines, including a malignant pleural mesothelioma (MPM) one, a very chemoresistant tumor. The main reason of the increased activity relies on the enhanced lipophilicity of these Pt(IV) conjugates that in turn promotes increased cellular accumulation. A quick Pt(IV)→Pt(II) reduction generates the active cisplatin metabolite. The NSAID adjuvant action seems to be almost independent from cyclooxygenase-2 (COX-2) expression in the tumor cells under investigation (lung A-549, colon HT-29, HCT 116, SW480, ovarian A2780, and biphasic MPM MSTO-211H), but it seems to rely (at least in part) on the activation of the NSAID activated gene, NAG-1 (a member of the transforming growth factor beta, TGF-ß, superfamily), which has been suggested to be involved in NSAID antiproliferative activity.

Pt(IV) Bifunctional Prodrug Containing 2-(2-Propynyl)octanoato Axial Ligand: Induction of Immunogenic Cell Death on Colon Cancer.

The synthesis, characterization, and in vitro activity of a cyclohexane-1 R,2 R-diamine-based Pt(IV) derivative containing the histone deacetylase inhibitor rac-2-(2-propynyl)octanoato, namely, ( OC-6-44)-acetatodichlorido(cyclohexane-1 R,2 R-diamine)( rac-2-(2-propynyl)octanoato)platinum(IV), are reported together with those of its isomers containing enantiomerically enriched axial ligands. These Pt(IV) complexes showed comparable activity, of 2 orders of magnitude higher than reference drug oxaliplatin on three human (HCT 116, SW480, and HT-29) and one mouse (CT26) colon cancer cell lines. In vivo experiments were carried out on immunocompetent BALB/c mice bearing the same syngeneic tumor. The complex ( OC-6-44)-acetatodichlorido(cyclohexane-1 R,2 R-diamine)( rac-2-(2-propynyl)octanoato)platinum(IV) showed higher tumor mass Pt accumulation than oxaliplatin, due to its higher lipophilicity, with negligible nephro- and hepatotoxicities when administered intravenously. A remarkable tumor mass invasion by cytotoxic CD8+ T lymphocytes, following the Pt(IV) treatment, indicated a strong induction of immunogenic cell death.

A new platinum-based prodrug candidate: Its anticancer effects in B50 neuroblastoma rat cells.

AIMS: Neuroblastoma is a rare cancer that affects children, mostly under the age of 5. This type of cancer starts in very early forms of immature nerve cells or developing cells found in embryo or fetus. To date cisplatin represents one of the most potent antitumor agent known, however, the onset of systemic side effects and the induction of drug resistance limit its use in the clinic for long-term treatment. In the present study we have analysed the effects of a new compound of platinum(IV) conjugates, named Pt(IV)Ac-POA, which is able to generate a synergistic antineoplastic action when released along with cisplatin upon intracellular Pt(IV) → Pt(II) reduction. MAIN METHODS: To assess the growth inhibition of the compounds under investigation, a cell viability test, i.e. the resazurin reduction assay was used on the B50 neuroblastoma rat cells. Further analysis on the cell cycle and metabolic alterations were carried out through flow cytometry. Morphological changes and activation of different cell death pathways after treatment, were observed at transmission electron microscope and by immunocytochemistry at fluorescence microscopy. Protein expression was examined by western blot analysis. KEY FINDINGS: This compound bearing bioactive axial ligand, such as the active histone deacetylase inhibitor (HDACi) (2-propynyl)octanoic acid (POA), induced cell death through different pathways at a concentration ten times lower than cisplatin. SIGNIFICANCE: The results showed that Pt(IV)Ac-POA could represent a promising improvement of Pt-based chemotherapy against neuroblastoma.

An unsymmetric cisplatin-based Pt(iv) derivative containing 2-(2-propynyl)octanoate: a very efficient multi-action antitumor prodrug candidate.

The design, synthesis, characterization and biological properties of a Pt(iv) complex containing the very active inhibitor of histone deacetylase (2-propynyl)octanoic acid, POA, as an axial ligand are reported here. The title complex, namely (OC-6-44)-acetatodiamminedichlorido(2-(2-propynyl)octanoato)platinum(iv), 1, containing POA in racemic or in enantiomeric forms, was one/two orders of magnitude more active than cisplatin, depending on the chemo-sensitivity of the cancer cell lines. Moreover, 1 exhibited similar or even better antiproliferative activity than (OC-6-33)-diamminedichloridobis(2-propylpentanoato)platinum(iv), 2, containing two molecules of the well-known histone deacetylase inhibitor 2-propylpentanoic (valproic) acid. The high potency of 1 is likely due to its high cellular accumulation and to the synergism between the DNA-damaging cisplatin and the histone deacetylase inhibitor POA, both released upon the intracellular reduction of 1. Prodrug 1, after oral administration, caused an impressive reduction of the tumor mass (94%) in a model of solid tumor (murine Lewis lung carcinoma), compared to that of the control, whereas (intraperitoneal) cisplatin induced a tumor regression of 75% only. A good accumulation of 1 was observed in the tumor mass. The time course of the body weight attested that cisplatin induced elevated anorexia, whereas treatment with 1 did not induce significant body weight loss throughout the therapeutic experiment.