Target identification remains a critical challenge in inorganic drug discovery to deconvolute potential polypharmacology. Herein, we describe an improved approach to prioritize candidate protein targets based on a combination of dose‐dependent chemoproteomics and treatment effects in living cancer cells for the rhenium tricarbonyl compound TRIP. Chemoproteomics revealed 89 distinct dose‐dependent targets with concentrations of competitive saturation between 0.1 and 32 μM despite the broad proteotoxic effects of TRIP. Target‐response networks revealed two highly probable targets of which the Fe−S cluster biogenesis factor NUBP2 was competitively saturated by free TRIP at nanomolar concentrations. Importantly, TRIP treatment led to a down‐regulation of Fe−S cluster containing proteins and upregulated ferritin. Fe−S cluster depletion was further verified by assessing mitochondrial bioenergetics. Consequently, TRIP emerges as a first‐in‐class modulator of the scaffold protein NUBP2, which disturbs Fe−S cluster biogenesis at sub‐cytotoxic concentrations in ovarian cancer cells.

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During recent years, accumulating evidence suggested that metal‐based candidate drugs are promising modulators of cytoskeletal and cytoskeleton‐associated proteins. This was substantiated by the identification and validation of actin, vimentin and plectin as targets of distinct ruthenium(II)‐ and platinum(II)‐based modulators. Despite this, structural information about molecular interaction is scarcely available. Here, we compile the scattered reports about metal‐based candidate molecules that influence the cytoskeleton, its associated proteins and explore their potential to interfere in cancer‐related processes, including proliferation, invasion and the epithelial‐to‐mesenchymal transition. Advances in this field depend crucially on determining binding sites and on gaining comprehensive insight into molecular drug‐target interactions. These are key steps towards establishing yet elusive structure‐activity relationships.

The ABC transporter ABCA7 has been found to be aberrantly expressed in a variety of cancer types, including breast cancer. We searched for specific epigenetic and genetic alterations and alternative splicing variants of ABCA7 in breast cancer and investigated whether these alterations are associated with ABCA7 expression. By analyzing tumor tissues from breast cancer patients, we found CpGs at the exon 5–intron 5 boundary aberrantly methylated in a molecular subtype-specific manner. The detection of altered DNA methylation in tumor-adjacent tissues suggests epigenetic field cancerization. In breast cancer cell lines, DNA methylation levels of CpGs in promoter exon 1, intron 1, and at the exon 5–intron 5 boundary were not correlated with ABCA7 mRNA levels. By qPCR involving intron-specific and intron-flanking primers, we identified intron-containing ABCA7 mRNA transcripts. The occurrence of intron-containing transcripts was neither molecular subtype-specific nor directly correlated with DNA methylation at the respective exon–intron boundaries. Treatment of breast cancer cell lines MCF-7, BT-474, SK-BR3, and MDA-MB-231 with doxorubicin or paclitaxel for 72 h resulted in altered ABCA7 intron levels. Shotgun proteomics revealed that an increase in intron-containing transcripts was associated with significant dysregulation of splicing factors linked to alternative splicing.

Die Identifizierung von Zielmolekülen ist nach wie vor eine erhebliche Hürde in der präklinischen Entdeckungsphase metallhaltiger Wirkstoffe, und entscheidend um potenzielle polypharmakologische Aspekte zu entschlüsseln. Hier beschreiben wir einen verbesserten Ansatz zur Identifizierung von priorisierten Zielprotein-Kandidaten für die Rheniumtricarbonylverbindung TRIP, der auf einer Kombination aus dosisabhängiger Chemoproteomik und Behandlungseffekten in lebenden Krebszellen beruht. Der chemoproteomische Ansatz ergab 89 dosisabhängige Interaktoren, die zwischen 0.1-32 μM kompetitiv gesättigt wurden, und das trotz der breiten proteotoxischen Wirkung von TRIP. Durch die Erstellung von Target-Response-Netzwerken wurden weiters zwei sehr wahrscheinliche Interaktoren identifiziert, von denen der FeÀ S-Cluster-Biogenesefaktor NUBP2 durch freies TRIP bei nanomolaren Konzentrationen kompetitiv gesättigt wurde. Die Behandlung der Krebszellen mit TRIP führte zudem zu einer Herunterregulierung von FeÀ S-Cluster-haltigen Proteinen und einer Hochregulierung von Ferritin. Die Verarmung an FeÀ S-Clustern wurde durch die Analyse der mitochondrialen Bioenergetik zusätzlich verifiziert. TRIP entpuppt sich somit als erster Modulator des Gerüstproteins NUBP2 und stört die FeÀ S-Cluster-Biogenese in Eierstockkrebszellen bei subzytotoxischen Konzentrationen.