Platinum(II) complexes prevail as first-line treatment for many cancers but are associated with serious side effects and resistance development. Picoplatin emerged as a promising alternative to circumvent GSH-induced tumor resistance by introducing a bulky 2-picoline ligand. Although clinical studies were encouraging, picoplatin did not receive approval. Interestingly, the anticancer potential of prodrugs based on picoplatin is widely underexplored, and even less so the respective tumor-targeting approaches. We synthesized two new "hybrid" picoplatin(II) derivatives with an oxalate or cyclobutane dicarboxylate leaving group and their corresponding platinum(IV) prodrugs with an albumin-targeting maleimide moiety or a succinimide as reference. Picoplatin(II) and its derivatives indeed reacted much slower with GSH compared to the respective analogs cisplatin, carboplatin, or oxaliplatin. While PicoCarbo(IV) and PicoOxali(IV) were reduced slowly in the presence of ascorbic acid, picoplatin(IV) was extremely unstable. All three prodrugs were widely inactive in the MTT assays. The platinum(IV)-maleimide complexes rapidly bound to albumin with stable conjugates for >25 h. Albumin-binding resulted in elevated platinum plasma levels, prolonged blood circulation, and enhanced tumor accumulation of the prodrugs in mice bearing CT26 tumors. However, only maleimide-functionalized PicoCarbo(IV) and picoplatin(II) significantly inhibited tumor growth. One possible explanation is that for albumin-binding platinum(IV) prodrugs, the bulky 2-picoline moiety prevents sufficient activation/reduction to unlock their full anticancer potential.

Maleimides are widely used in anticancer drug development for linking small-molecule drugs to macromolecules like antibodies or proteins via thiol-Michael addition reactions. Despite their widespread use, even in clinically approved therapeutics, they present significant drawbacks such as hydrolysis at physiological pH and instability of the formed thiosuccinimide bond. Hence, there is a growing need for more stable yet equally efficient binding units. This is particularly important for drug-delivery systems that bind to endogenous albumin in vivo, exploiting the ability of the protein to accumulate in tumor tissue. This study compares phenyloxadiazolyl methyl sulfone (PODS) and a 2,4-difluorophenyl sulfonamide (DFSA) derivative with maleimide as endogenous albumin binders. Of note, PODS and maleimide bind to Cys34, whereas DFSA targets Lys64 of albumin. The albumin binders were conjugated as axial ligands to oxaliplatin-(IV) complexes (PODS-Ox-OAc and DFSA-Ox-OAc) and studied in comparison to a maleimide-bearing reference compound (Mal-Ox-OAc). Both PODS- and DFSA-complexes showed higher hydrolytic stability at pH 7.4 than the maleimide complex. Albumin-binding was highly efficient for the PODS and maleimide complexes. However, the DFSA derivative exhibited only slow conjugation. This was also reflected in the serum pharmacokinetic and organ distribution studies using CT26 colon cancer-bearing mice. Here, the PODS complex showed the highest platinum levels in both serum and tumor tissue. Additionally, PODS-Ox-OAc induced the most significant tumor regression and prolonged overall survival in this model. Together, our data highlight PODS as a promising alternative to maleimide as an endogenous albumin binder.

Platinum(iv) prodrugs are a promising class of anticancer agents designed to overcome the limitations of conventional platinum(ii) therapeutics. In this work, we present oxaliplatin(iv)-based complexes, which upon reduction, release acetylsalicylic acid (aspirin), known for its antitumor activity against colon cancer and currently investigated in combination with oxaliplatin in a phase III clinical study. Comparison with a recently reported cisplatin analog (asplatin) revealed a massive increase in reduction stability for the oxaliplatin complex in mouse serum. This was in line with the cell culture data indicating the desired prodrug properties for the newly synthesized complex. For in vivo studies, a new derivative containing an albumin-binding maleimide unit was synthesized. Indeed, distinctly longer plasma half-life as well as higher tumor accumulation in comparison to asplatin and oxaliplatin were observed, also leading to significantly higher antitumor activity and overall survival of CT26 tumor-bearing mice.

Platinum(IV) prodrugs are highly interesting alternatives to platinum(II) anticancer therapeutics due to their increased tumor selectivity and reduced side effects. In contrast to the established theory, we recently observed that the equatorial ligand(s) of e.g. oxaliplatin(IV) complexes can be hydrolyzed with formation of [(DACH)Pt(OHeq )2 (OAcax )2 ]. In the work presented here, we investigated the reactivity and synthetic usability of this complex to be exploited as a precursor for the development of novel platinum(IV) complexes, not able to be synthesized by conventional protocols. Indeed, we could substitute the equatorial hydroxido ligand(s) e.g. by one or two monodentate biotin ligands (which would be oxidized under standard methods). The formed complexes turned out to be very stable with slow ligand release after reduction, ideal for long-circulating tumor-targeting strategies. Therefore, two platinum(IV) complexes with equatorial maleimides, capable of exploiting serum albumin as a natural nanocarrier, were synthesized as well. The complexes showed massively prolonged plasma half-life and distinctly improved anticancer activity in vivo compared to oxaliplatin. Taken together, the newly developed synthetic platform allows the simple and specific insertion of equatorial ligands into platinum(IV) complexes. This will enable the attachment of three different (bioactive) moieties generating targeted triple-action platinum(IV) prodrugs within one single platinum complex.

While platinum-based chemotherapeutic agents have established themselves as indispensable components of anticancer therapy, they are accompanied by a variety of side effects and the rapid occurrence of drug resistance. A promising strategy to address these challenges is the use of platinum(iv) prodrugs, which remain inert until they reach the tumor tissue, thereby mitigating detrimental effects on healthy cells. Typically, platinum drugs are part of combination therapy settings. Consequently, a very elegant strategy is the development of platinum(iv) prodrugs bearing a second, clinically relevant therapeutic in axial position. In the present study, we focused on gemcitabine as an approved antimetabolite, which is highly synergistic with platinum drugs. In addition, to increase plasma half-life and facilitate tumor-specific accumulation, an albumin-binding maleimide moiety was attached. Our investigations revealed that maleimide-cisplatin(iv)-gemcitabine complexes cannot carry sufficient amounts of gemcitabine to induce a significant effect in vivo. Consequently, we designed a carboplatin(iv) analog, that can be applied at much higher doses. Remarkably, this novel analog demonstrated impressive in vivo results, characterized by significant improvements in overall survival. Notably, these encouraging results could also be transferred to an in vivo xenograft model with acquired gemcitabine resistance, indicating the high potential of this approach.

AbstractPlatin(IV)‐Prodrugs sind aufgrund ihrer erhöhten Tumorselektivität und geringeren Nebenwirkungen äußerst interessante Alternativen zu Platin(II)‐Antitumortherapeutika. Im Gegensatz zur gängigen Theorie haben wir kürzlich beobachtet, dass äquatoriale Liganden von z. B. Oxaliplatin(IV)‐Komplexen unter Bildung von [(DACH)Pt(OHeq)2(OAcax)2] hydrolysiert werden können. In der hier vorgestellten Arbeit untersuchten wir die Reaktivität und synthetische Verwendbarkeit dieses Komplexes, als Vorstufe für die Entwicklung neuartiger Platin(IV)‐Komplexe, welche mit herkömmlichen Methoden nicht zugänglich sind. Tatsächlich war es möglich die äquatorialen Hydroxidoliganden z. B. durch ein oder zwei monodentate Biotin‐Liganden, die unter Standardmethoden oxidiert werden würden, zu ersetzen. Die gebildeten Komplexe erwiesen sich als sehr stabil und zeigten auch nach der Reduktion eine langsame Ligandenfreisetzung, eine ideale Eigenschaft für lang zirkulierende zielgerichtete Strategien. Daraufhin wurden zwei Platin(IV)‐Komplexe mit äquatorialen Maleimiden, für die Bindung an Serumalbumin als natürlichen Nanocarrier, synthetisiert. Die Komplexe zeigten im Vergleich zu Oxaliplatin eine stark verlängerte Plasmahalbwertszeit und eine deutlich verbesserte Antitumoraktivität in vivo. Zusammenfassend ermöglicht diese neu entwickelte Syntheseplattform den einfachen und gezielten Einbau äquatorialer Liganden in Platin(IV)‐Komplexe. Des Weiteren können verschiedene (bioaktive) Einheiten koordiniert werden, wodurch sogar zielgerichtete dreifach‐wirksame Platin(IV)‐Prodrugs mit nur einem Platinzentrum möglich wären.

Clinical efficacy of oxaliplatin is frequently limited by severe adverse effects and therapy resistance. Acquired insensitivity to oxaliplatin is, at least in part, associated with elevated levels of glutathione (GSH). In this study we report on an oxaliplatin-based platinum(IV) prodrug, which releases L -buthionine- S , R -sulfoximine (BSO), an inhibitor of glutamate-cysteine ligase, the rate-limiting enzyme in GSH biosynthesis. Two complexes bearing either acetate ( BSO-OxOAc ) or an albumin-binding maleimide ( BSO-OxMal ) as second axial ligand were synthesized and characterized. The in vitro anticancer activity of BSO-OxOAc was massively reduced in comparison to oxaliplatin, proving its prodrug nature. Nevertheless, the markedly lower intracellular oxaliplatin uptake in resistant HCT116/OxR cells was widely overcome by BSO-OxOAc resulting in distinctly reduced resistance levels. Platinum accumulation in organs of a colorectal cancer mouse model revealed higher tumor selectivity of BSO-OxMal as compared to oxaliplatin. This corresponded with increased antitumor activity, resulting in significantly enhanced overall survival. BSO-OxMal -treated tumors exhibited reduced GSH levels, proliferative activity and enhanced DNA damage (pH2AX) compared to oxaliplatin. Conversely, pH2AX staining especially in kidney cells was distinctly increased by oxaliplatin but not by BSO-OxMal . Taken together, our data provide compelling evidence for enhanced tumor specificity of the oxaliplatin(IV)/BSO prodrug.

Chemotherapy with platinum complexes is essential for clinical anticancer therapy. However, due to side effects and drug resistance, further drug improvement is urgently needed. Herein, we report on triple-action platinum(IV) prodrugs, which, in addition to tumor targeting via maleimide-mediated albumin binding, release the immunomodulatory ligand 1-methyl-d-tryptophan (1-MDT). Unexpectedly, structure-activity relationship analysis showed that the mode of 1-MDT conjugation distinctly impacts the reducibility and thus activation of the prodrugs. This in turn affected ligand release, pharmacokinetic properties, efficiency of immunomodulation, and the anticancer activity in vitro and in a mouse model in vivo. Moreover, we could demonstrate that the design of albumin-targeted multi-modal prodrugs using platinum(IV) is a promising strategy to enhance the cellular uptake of bioactive ligands with low cell permeability (1-MDT) and to improve their selective delivery into the malignant tissue. This will allow tumor-specific anticancer therapy supported by a favorably tuned immune microenvironment.

Maleimides are essential compounds for drug conjugation reactions via thiols to antibodies, peptides and other targeting units. However, one main drawback is the occurrence of thiol exchange reactions with, for example, glutathione resulting in loss of the targeting ability. A new strategy to overcome such retro-Michael exchange processes of maleimide-thiol conjugates by stabilization of the thiosuccinimide via a transcyclization reaction is presented. This reaction enables the straightforward synthesis of stable maleimide-thiol adducts essential in drug-conjugation applications.

Due to their high kinetic inertness and consequently reduced side reactions with biomolecules, PtIV complexes are considered to define the future of anticancer platinum drugs. The aqueous stability of a series of biscarboxylato PtIV complexes was studied under physiologically relevant conditions. Unexpectedly and in contrast to the current chemical understanding, especially oxaliplatin and satraplatin complexes underwent fast hydrolysis in equatorial position (even in cell culture medium and serum). Notably, the resulting hydrolysis products strongly differ in their reduction kinetics, a crucial parameter for the activation of PtIV drugs, which also changes the anticancer potential of the compounds in cell culture. The discovery that intact PtIV complexes can hydrolyze at equatorial position contradicts the dogma on the general kinetic inertness of PtIV compounds and needs to be considered in the screening and design for novel platinum-based anticancer drugs.

AbstractPtIV‐Komplexe gelten aufgrund ihrer hohen kinetischen Inertheit und der folglich reduzierten Nebenreaktionen mit Biomolekülen als die Zukunft der Platintherapeutika. In dieser Arbeit wurde unter physiologisch relevanten Bedingungen die Stabilität einer Reihe von PtIV‐Biscarboxylatokomplexen in wässriger Lösung untersucht. Entgegen des derzeitigen chemischen Kenntnisstandes zeigten (sogar in Zellkulturmedium und Serum) vor allem Oxaliplatin‐ und Satraplatinkomplexe eine schnelle äquatoriale Hydrolyse. Die gebildeten Hydrolyseprodukte unterscheiden sich besonders stark in ihrer Reduktionskinetik, einem entscheidenden Parameter für die Aktivierung von PtIV‐Medikamenten, welcher auch mit der zytotoxischen Effizienz der Verbindungen in der Zellkultur korreliert. Die Entdeckung, dass intakte PtIV‐Komplexe an äquatorialer Position hydrolysiert werden können, widerspricht dem Dogma hinsichtlich der allgemeinen kinetischen Inertheit von PtIV‐Verbindungen und muss beim Screening und der Entwicklung neuer platin‐basierter Krebsmedikamente berücksichtigt werden.

Platin(II)-Verbindungen spielen eine sehr wichtige Rolle in der Behandlung von Krebs und sind teil von ungefahr 50% aller Chemotherapien. Die drei weltweit zugelassenen Komplexe Cis-, Carbo- und Oxaliplatin werden unter anderem gegen Hoden-, Eierstock-, Darmkrebs verwendet, aber auch gegen Kopf-Hals-Karzinome. Unglucklicherweise sind sie aber auch unter anderem fur Niere, Verdauungs- und Nervensystem toxisch. Um diese Nebenwirkungen zu verringern, liegt der Fokus heutzutage auf Platin(IV)-Prodrugs. Ihr groser Vorteil besteht darin, im Vergleich zu Platin(II), kinetisch inert zu sein, was in geringerer Reaktivitat und damit weniger Nebenreaktionen mit z.B. Biomolekulen im Blutplasma mit sich bringt. Erst innerhalb des Tumors werden die Prodrugs dann zu ihrer aktiven Form reduziert. Nun gibt es aber einige Berichte aus klinischen Studien von Satraplatin, dass sogar im intakten Platin(IV)-Komplex die aquatorial Chloridoliganden hydrolysiert werden konnen. Dieses Phanomen wurde bis jetzt noch fur keine anderen Platin(IV)-Komplexe beobachtet. Ziel dieser Masterarbeit war es deshalb das Phanomen der Hydrolyse von aquatorialen Liganden an einer Auswahl an Platin(IV)-Komplexen zu untersuchen. Dafur wurden Modellverbindungen mit einer Vielfalt an aquatorialen Liganden synthetisiert (Cis-, Carbo-, Oxaliplatin als auch einige ihrer Derivate) und auf ihre Anfalligkeit gegenuber Hydrolyse in verschiedenen (auch biologischen) Milieus, wie Puffer, Medium und Serum mittels Hochleistungsflussigkeitschromatographie gekoppelt mit Massenspektrometrie (HPLC-MS) getestet. In der Tat zeigte vor allem das Oxaliplatin(IV)derivat Hydrolyse in einem signifikanten Ausmas bei physiologischen Bedingungen innerhalb von 24 h. Die Hydrolyseprodukte dieses Komplexes wurden anschliesend synthetisiert und charakterisiert. Schlussendlich wurden die physikalisch-chemischen Eigenschaften im Vergleich zum Ausgangskomplex untersucht. Zusammenfassend zeigt diese Masterarbeit, dass einige Platin(IV)-Komplexe bei biologisch relevanten Bedingungen zu einem beachtlichen Anteil aquatorial hydrolysiert werden. Dies sollte die generelle Ansicht von Platin(IV)-Komplexen und ihrer Chemie bezuglich kinetischer Inertheit andern.