Copper‐Free Click‐Chemistry Platform to Functionalize Cisplatin Prodrugs

The ability to rationally design and construct a platform technology to develop new platinum(IV) [Pt^IV] prodrugs with functionalities for installation of targeting moieties, delivery systems, fluorescent reporters from a single precursor with the ability to release biologically active cisplatin by using well‐defined chemistry is critical for discovering new platinum‐based therapeutics. With limited numbers of possibilities considering the sensitivity of Pt^IV centers, we used a strain‐promoted azide–alkyne cycloaddition approach to provide a platform, in which new functionalities can easily be installed on cisplatin prodrugs from a single Pt^IV precursor. The ability of this platform to be incorporated in nanodelivery vehicle and conjugation to fluorescent reporters were also investigated.     

A Dual Killing Strategy: Photocatalytic Generation of Singlet Oxygen with Concomitant PtIV Prodrug Activation

A ruthenium‐based mitochondrial‐targeting photosensitiser that undergoes efficient cell uptake, enables the rapid catalytic conversion of Pt^IV prodrugs into their active Pt^II counterparts, and drives the generation of singlet oxygen was designed. This dual mode of action drives two orthogonal cancer‐cell killing mechanisms with temporal and spatial control. The designed photosensitiser was shown to elicit cell death of a panel of cancer cell lines including those showing oxaliplatin‐resistance.     

Monochalcoplatin: An Actively Transported,Quickly Reducible,and Highly Potent PtIV Anticancer Prodrug

Recently, Pt^IV prodrugs have attracted much attention as the next generation of platinum‐based antineoplastic drug candidates. Here we report the discovery and evaluation of monochalcoplatin, a monocarboxylated Pt^IV prodrug that is among the most cytotoxic Pt^IV prodrugs to date. Compared with its dicarboxylated counterpart chalcoplatin, monochalcoplatin accumulates astonishingly effectively and rapidly in cancer cells, which is not ascribed to its lipophilicity. The prodrug is quickly reduced, causes DNA damage, and induces apoptosis, resulting in superior cytotoxicity with IC₅₀ values in the nanomolar range in both cisplatin‐sensitive and ‐resistant cells; these IC₅₀ values are up to 422‐fold higher than that of cisplatin. A detailed mechanistic study reveals that monochalcoplatin actively enters cells through a transporter‐mediated process. Moreover, monochalcoplatin shows significant antitumor activity in an in vivo colorectal tumor model. Our study implies a practical strategy for the design of more effective Pt^IV prodrugs to conquer drug resistance by tuning both cellular uptake pathways and activation processes.     

Monochalcoplatin: An Actively Transported,Quickly Reducible,and Highly Potent PtIV Anticancer Prodrug

Recently, Pt^IV prodrugs have attracted much attention as the next generation of platinum‐based antineoplastic drug candidates. Here we report the discovery and evaluation of monochalcoplatin, a monocarboxylated Pt^IV prodrug that is among the most cytotoxic Pt^IV prodrugs to date. Compared with its dicarboxylated counterpart chalcoplatin, monochalcoplatin accumulates astonishingly effectively and rapidly in cancer cells, which is not ascribed to its lipophilicity. The prodrug is quickly reduced, causes DNA damage, and induces apoptosis, resulting in superior cytotoxicity with IC₅₀ values in the nanomolar range in both cisplatin‐sensitive and ‐resistant cells; these IC₅₀ values are up to 422‐fold higher than that of cisplatin. A detailed mechanistic study reveals that monochalcoplatin actively enters cells through a transporter‐mediated process. Moreover, monochalcoplatin shows significant antitumor activity in an in vivo colorectal tumor model. Our study implies a practical strategy for the design of more effective Pt^IV prodrugs to conquer drug resistance by tuning both cellular uptake pathways and activation processes.     

Activation of Platinum(IV) Prodrugs By Motexafin Gadolinium as a Redox Mediator

Water‐soluble platinum(IV) prodrugs, which proved kinetically stable to reduction in the presence of physiological concentration of ascorbate, were quickly reduced to their active form, oxaliplatin, when co‐incubated with a macrocycle metallotexaphyrin (i.e., Motexafin Gadolinium (MGd)). The reduction of Pt^IV to Pt^II promoted by MGd occurs in cell culture as well, leading to an increase in the antiproliferative activity of the Pt^IV species in question. The mediated effect is proportional to the concentration of MGd and gives rise to an enhancement when the prodrug is relatively hydrophilic. MGd is known to localize/accumulate preferentially in tumor tissues. Thus, the present “activation by reduction” approach may allow for the cancer‐selective enhancement in the cytotoxicity of Pt^IV prodrugs.     

Multitargeted Platinum(IV) Anticancer Complexes Bearing Pyridinyl Ligands as Axial Leaving Groups

Although multitargeted Pt^IV anticancer prodrugs have shown significant activities in reducing drug resistance, the types of bioactive ligands and drugs that can be conjugated to the Pt center remain limited to O-donors. Herein, we report the synthesis of Pt^IV complexes bearing axial pyridines via ligand exchange reactions. Unexpectedly, the axial pyridines are quickly released after reduction, indicating their potential to be utilized as axial leaving groups. We further expand our synthetic approach to obtaining two multitargeted Pt^IV prodrugs containing bioactive pyridinyl ligands: a PARP inhibitor and an EGFR tyrosine kinase inhibitor; these conjugates exhibit great potential for overcoming drug resistance, and the latter conjugate inhibits the growth of Pt-resistant tumor in vivo. This research adds to the array of synthetic methods for accessing Pt^IV prodrugs and significantly increases the types of bioactive axial ligands that can be conjugated to a Pt^IV center.     

Expanding the Arsenal of PtIV Anticancer Agents: Multi‐action PtIV Anticancer Agents with Bioactive Ligands Possessing a Hydroxy Functional Group

Most multi‐action Pt^IV prodrugs have bioactive ligands containing carboxylates. This is probably due to the ease of carboxylating the OH axial ligands and because following reduction, the active drug is released. A major challenge is to expand the arsenal of bioactive ligands to include those without carboxylates. We describe a general approach for synthesis of Pt^IV prodrugs that release drugs with OH groups. We linked the OH groups of gemcitabine (Gem), paclitaxel (Tax), and estramustine (EM) to the Pt^IV derivative of cisplatin by a carbonate bridge. Following reduction, the axial ligands lost CO₂, rapidly generating the active drugs. In contrast, succinate‐linked drugs did not readily release the free drugs. The carbonate‐bridged ctc‐[Pt(NH₃)₂(PhB)(Gem‐Carb)Cl₂] was significantly more cytotoxic than the succinate‐bridged ctc‐[Pt(NH₃)₂(PhB)(Gem‐Suc)Cl₂], and more potent and less toxic than gemcitabine, cisplatin, and co‐administration of cisplatin and gemcitabine.     

A Multi‐action and Multi‐target RuII–PtIV Conjugate Combining Cancer‐Activated Chemotherapy and Photodynamic Therapy to Overcome Drug Resistant Cancers

Pt^II complexes are commonly used to treat cancer. To reduce their side effects and improve their pharmacological properties, Pt^IV complexes are being developed as prodrug candidates that are activated by reduction in cancer cells. Concomitantly, Ru^II polypyridine complexes have gained much attention as photosensitizers for use in photodynamic therapy due to their attractive characteristics. In this article, a novel Pt^IV–Ru^II conjugate, which combines cancer activated chemotherapy with PDT, is presented. Upon entering the cancer cell, the Pt^IV centre is reduced to Pt^II and the axial ligands including the Ru^II complex and phenylbutyrate are released. As each component has its individual targets, the conjugate exerts a multi‐target and multi‐action effect with (photo‐)cytotoxicity values upon irradiation up to 595 nm in the low nanomolar range in various (drug resistant) 2D monolayer cancer cells and 3D multicellular tumour spheroids.     

Platinum (IV) Derivatives with Cinnamate Axial Ligands as Potent Agents Against Both Differentiated and Tumorigenic Cancer Stem Rhabdomyosarcoma Cells

To design an anticancer drug capable of inhibiting not only the proliferation of the differentiated tumor cells but also reducing the tumorigenic capability of cancer stem cells (CSCs), the new Pt^IV prodrugs with axial cinnamate ligands were synthesized. We demonstrate their superior antiproliferative activity in monolayer and 3D spheroid antiproliferative activity tests using panel of cancer cell lines. An outstanding activity was found against rhabdomyosarcoma cells, one of the most problematic and poorly treatable pediatric tumors. The results also suggest that the released Pt^II compound inhibits antiproliferative activity of cancer cells by DNA‐damage mediated mechanism; the released cinnamic acid can trigger processes leading to differentiation, making the CSCs more sensitive to killing by the platinum part of the complex. Pt^IV complex with axial cinnamate ligands is the first Pt^IV prodrug capable of overcoming CSCs resistance and induce death in both CSCs and bulk cancer.     

Human Serum Albumin Conjugated Nanoparticles for pH and Redox‐Responsive Delivery of a Prodrug of Cisplatin

Platinum anticancer drugs are particularly in need of controlled drug delivery because of their severe side effects. Platinum(IV) agents are designed as prodrugs to reduce the side effects of platinum(II) drugs; however, premature reduction could limit the effect as a prodrug. In this work, a highly biocompatible, pH and redox dual‐responsive delivery system is prepared by using hybrid nanoparticles of human serum albumin (HSA) and calcium phosphate (CaP) for the Pt^IV prodrug of cisplatin. This conjugate is very stable under extracellular conditions, so that it protects the platinum(IV) prodrug in HSA. Upon reaching the acidic and hypoxic environment, the platinum drug is released in its active form and is able to bind to the target DNA. The Pt–HSA/CaP hybrid inhibits the proliferation of various cancer cells more efficiently than cisplatin. Different cell cycle arrests suggest different cellular responses of the Pt^IV prodrug in the CaP nanocarrier. Interestingly, this delivery system demonstrates enhanced cytotoxicity to tumor cells, but not to normal cells.     

An Anticancer PtIV Prodrug That Acts by Mechanisms Involving DNA Damage and Different Epigenetic Effects

Dual- or multi-action Pt^IV prodrugs represent a new generation of platinum anticancer drugs. The important property of these Pt^IV prodrugs is that their antitumor action combines several different mechanisms owing to the presence of biologically active axial ligands. This work describes the synthesis and some biological properties of a “triple-action” prodrug that releases in cancer cells cisplatin and two different epigenetically acting moieties, octanoate and phenylbutyrate. It is demonstrated, with the aid of modern methods of molecular and cellular biology and pharmacology, that the presence of three different functionalities in a single molecule of the Pt^IV prodrug results in a selective and high potency in tumor cells including those resistant to cisplatin [the IC₅₀ values in the screened malignant cell lines ranged from as low as 9 nm (HCT-116) to 74 nm (MDA-MB-231)]. It is also demonstrated that cellular activation of the Pt^IV prodrug results in covalent modification of DNA through the release of the platinum moiety accompanied by inhibition of the activity of histone deacetylases caused by phenylbutyrate and by global hypermethylation of DNA by octanoate. Thus, the Pt^IV prodrug introduced in this study acts as a true “multi-action” prodrug, which is over two orders of magnitude more active than clinically used cisplatin, in both 2D monolayer culture and 3D spheroid cancer cells.     

Oral Anticancer Heterobimetallic PtIV−AuI Complexes Show High In Vivo Activity and Low Toxicity

Au^I-carbene and Pt^IV−Au^I-carbene prodrugs display low to sub-μM activity against several cancer cell lines and overcome cisplatin (cisPt) resistance. Linking a cisPt-derived Pt^IV(phenylbutyrate) complex to a Au^I-phenylimidazolylidene complex 2 , yielded the most potent prodrug. While in vivo tests against Lewis Lung Carcinoma showed that the prodrug Pt^IV(phenylbutyrate)-Au^I-carbene ( 7 ) and the 1 : 1 : 1 co-administration of cisPt: phenylbutyrate: 2 efficiently inhibited tumor growth (≈95 %), much better than 2 (75 %) or cisPt (84 %), 7 exhibited only 5 % body weight loss compared to 14 % for 2 , 20 % for cisPt and >30 % for the co-administration. 7 was much more efficient than 2 at inhibiting TrxR activity in the isolated enzyme, in cells and in the tumor, even though it was much less efficient than 2 at binding to selenocysteine peptides modeling the active site of TrxR. Organ distribution and laser-ablation (LA)-ICP-TOFMS imaging suggest that 7 arrives intact at the tumor and is activated there.     

Photoinduced Reduction of PtIV within an Anti‐Proliferative PtIV–Texaphyrin Conjugate

In an effort to increase the stability and control the platinum reactivity of platinum–texaphyrin conjugates, two Pt^IV conjugates were designed, synthesized, and studied for their ability to form DNA adducts. They were also tested for their anti‐proliferative effects using wild‐type and platinum‐resistant human ovarian cancer cell lines (A2780 and 2780CP, respectively). In comparison to an analogous first‐generation Pt^II chimera, one of the new conjugates provided increased stability in aqueous environments. Using a combination of ¹H NMR spectroscopy and FAAS (flameless atomic‐absorption spectrometry), it was found that the Pt^IV center within this conjugate undergoes photoinduced reduction to Pt^II upon exposure to glass‐filtered daylight, resulting in an entity that binds DNA in a controlled manner. Under conditions in which the Pt^IV complex is reduced to the corresponding Pt^II species, these new conjugates demonstrated potent anti‐proliferative activity in both test ovarian cancer cell lines.     

Development of a Pre‐assembled Through‐Bond Energy Transfer (TBET) Fluorescent Probe for Ratiometric Sensing of Anticancer Platinum(ll) Complexes

Fluorescence microscopy has emerged as an attractive technique to probe the intracellular processing of Pt‐based anticancer compounds. Herein, we reported the first through‐bond energy transfer (TBET) fluorescent probe NPR1 designed for sensitive detection and quantitation of Pt^II complexes. The novel TBET probe was successfully applied for ratiometric fluorescence imaging of anticancer Pt^II complexes such as cisplatin and JM118 in cells. Capitalizing on the ability of the probe to discriminate between Pt^II complexes and their Pt^IV derivatives, the probe was further applied to study the activation of Pt^IV prodrug complexes that are known to release active Pt^II species after intracellular reduction.     

Photolysis and Thermolysis of Platinum(IV) 2,2′‐Bipyridine Complexes Lead to Identical Platinum(II)–DNA Adducts

Two Pt^IV and two Pt^II complexes containing a 2,2′‐bipyridine ligand were treated with a short DNA oligonucleotide under light irradiation at 37 °C or in the dark at 37 and 50 °C. Photolysis and thermolysis of the Pt^IV complexes led to spontaneous reduction of the Pt^IV to the corresponding Pt^II complexes and to binding of Pt^II 2,2′‐bipyridine complexes to N7 of guanine. When the reduction product was [Pt(bpy)Cl₂], formation of bis‐oligonucleotide adducts was observed, whereas [Pt(bpy)(MeNH₂)Cl]⁺ gave monoadducts, with chloride ligands substituted in both cases. Neither in the dark nor under light irradiation was the reductive elimination process of these Pt^IV complexes accompanied by oxidative DNA damage. This work raises the question of the stability of photoactivatable Pt^IV complexes toward moderate heating conditions.     

A one‐dimensional iodine‐bridged PtII/PtIV mixed‐valence complex,catena‐poly[[[bis­(ethyl­ene­diamine)­platinum(II)]‐μ‐iodo‐[bis­(ethyl­ene­di­amine)platinum(IV)]‐μ‐iodo] hydrogenphosphate dihydrogenphosphate iodide trihydrate]

The title compound, {[Pt^IIPt^IVI₂(C₂H₈N₂)₄](HPO₄)(H₂PO₄)I·3H₂O}_n, has a chain structure composed of square‐planar [Pt(en)₂]²⁺ and elongated octa­hedral trans‐[PtI₂(en)₂]²⁺ cations (en is ethyl­ene­diamine) stacked alternately along the c axis and bridged by the I atoms; a three‐dimensionally valence‐ordered system exists with respect to the Pt sites. The title compound also has a unique cyclic tetra­mer structure composed of two hydrogenphosphate and two dihydrogenphosphate ions connected by strong hydrogen bonds [O⋯O = 2.522 (10), 2.567 (10) and 2.569 (11) Å]. The Pt and I atoms form a zigzag ⋯I—Pt^IV—I⋯Pt^II⋯ chain, with Pt^IV—I bond distances of 2.6997 (7) and 2.6921 (7) Å, inter­atomic Pt^II⋯I distances of 3.3239 (8) and 3.2902 (7) Å, and Pt^IV—I⋯Pt^II angles of 154.52 (3) and 163.64 (3)°. The structural parameters indicating the mixed‐valence state of platinum, expressed by δ = (Pt^IV—I)/(Pt^II—I), are 0.812 and 0.818 for the two independent I atoms.     

A one‐dimensional chloride‐bridged PtII/PtIV mixed‐valence complex with a 4‐[(4‐hydroxyphenyl)diazenyl]benzenesulfonate counter‐ion

The title compound, catena‐poly[[[bis(ethylenediamine‐κ²N,N′)platinum(II)]‐ μ‐chlorido‐[bis(ethylenediamine)platinum(IV)]‐μ‐chlorido] tetrakis{4‐[(4‐hydroxyphenyl)diazenyl]benzenesulfonate} dihydrate], {[Pt^IIPt^IVCl₂(C₂H₈N₂)₄](HOC₆H₄N=NC₆H₄SO₃)₄·2H₂O}_n, has a linear chain structure composed of square‐planar [Pt(en)₂]²⁺ (en is ethylenediamine) and elongated octahedral trans‐[PtCl₂(en)₂]²⁺ cations stacked alternately, bridged by Cl atoms, along the b axis. The Pt atoms are located on an inversion centre, while the Cl atoms are disordered over two sites and form a zigzag ...Cl—Pt^IV—Cl...Pt^II... chain, with a Pt^IV—Cl bond length of 2.3140 (14) Å, an interatomic Pt^II...Cl distance of 3.5969 (15) Å and a Pt^IV—Cl...Pt^II angle of 170.66 (6)°. The structural parameter indicating the mixed‐valence state of the Pt atom, expressed by δ = (Pt^IV—Cl)/(Pt^II...Cl), is 0.643.     

Reduction of Platinum(IV) Prodrug Hemoglobin Nanoparticles with Deeply Penetrating Ultrasound Radiation for Tumor-Targeted Therapeutically Enhanced Anticancer Therapy

The development of Pt^IV prodrugs that are reduced into the therapeutically active Pt^II species within the tumor microenvironment has received much research interest. In order to provide spatial and temporal control over the treatment, there is a high demand for the development of compounds that could be selectively activated upon irradiation. Despite recent progress, the majority of Pt^IV complexes are excited with ultraviolet or blue light, limiting the use of such compounds to superficial application. To overcome this limitation, herein, the first example of Pt^IV prodrug nanoparticles that could be reduced with deeply penetrating ultrasound radiation is reported, enabling the treatment of deep-seated or large tumors. The nanoparticles were found to selectively accumulate inside a mouse colon carcinoma tumor upon intravenous injection and were able to eradicate the tumor upon exposure to ultrasound radiation.     

Can an Elusive Platinum(III) Oxidation State be Exposed in an Isolated Complex?

Platinum(IV) complexes are extensively studied for their activity against cancer cells as potential substitutes for the widely used platinum(II) drugs. Pt^IV complexes are kinetically inert and need to be reduced to Pt^II species to play their pharmacological action, thus acting as prodrugs. The mechanism of the reduction step inside the cell is however still largely unknown. Gas‐phase activation of deprotonated platinum(IV) prodrugs was found to generate products in which platinum has a formal +3 oxidation state. IR multiple photon dissociation spectroscopy is thus used to obtain structural information helping to define the nature of both the platinum atom and the ligands. In particular, comparison of calculations at DFT, MP2 and CCSD levels with experimental results demonstrates that the localization of the radical is about equally shared between the d_xz orbital of platinum and the p_z of nitrogen on the amino group, the latter acting as a non‐innocent ligand.     

Bioorthogonal Catalytic Activation of Platinum and Ruthenium Anticancer Complexes by FAD and Flavoproteins

Recent advances in bioorthogonal catalysis promise to deliver new chemical tools for performing chemoselective transformations in complex biological environments. Herein, we report how FAD (flavin adenine dinucleotide), FMN (flavin mononucleotide), and four flavoproteins act as unconventional photocatalysts capable of converting Pt^IV and Ru^II complexes into potentially toxic Pt^II or Ru^II?OH₂ species. In the presence of electron donors and low doses of visible light, the flavoproteins mini singlet oxygen generator (miniSOG) and NADH oxidase (NOX) catalytically activate Pt^IV prodrugs with bioorthogonal selectivity. In the presence of NADH, NOX catalyzes Pt^IV activation in the dark as well, indicating for the first time that flavoenzymes may contribute to initiating the activity of Pt^IV chemotherapeutic agents.