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N-Alkylaminoferrocene (NAAF)-based prodrugs are activated in the presence of elevated amounts of reactive oxygen species (ROS), which corresponds to cancer specific conditions, with formation of NAAF and p-quinone methide. Both products act synergistically by increasing oxidative stress in cancer cells that causes their death. Though it has already been demonstrated that the best prodrugs of this type retain their antitumor activity in vivo, the effects were found to be substantially weaker than those observed in cell cultures. Moreover, the mechanistic studies of these compounds in vivo are missing. For clarification of these important questions, labeling of the prodrugs with radioactive moieties would be necessary. In this paper, we first observed that the representative NAAF-based prodrugs are hydrolyzed in dilute aqueous solutions to the corresponding arylboronic acids. We confirmed that these products are responsible for ROS amplification and anticancer properties of the parent prodrugs. Next, we developed the efficient synthetic protocol for radiolabeling the hydrolyzed NAAF-based prodrugs by [ 18 F]fluoroglucosylation under the conditions of the copper(I)-catalyzed azide-alkyne 1,3-dipolar cycloaddition and used this protocol to prepare one representative hydrolyzed NAAF-based prodrug radiolabeled with 18 F. Finally, we studied the stability of the 18 F-labeled compound in human serum in vitro and in rat blood in vivo and obtained preliminary data on its biodistribution in vivo in mice carrying pancreatic (AR42J) and prostate (PC3) tumors by applying PET imaging studies. The compound described in this paper will help to understand in vivo effects (e.g., pharmacokinetics, accumulation in organs, the nature of side effects) of these prodrugs that will strongly contribute to their advancement to clinical trials.

Systemic inflammation such as sepsis represents an acute life-threatening condition, to which often no timely remedy can be found. A promising strategy may be to functionalize magnetic nanoparticles with specific peptides, derived from the binding motives of agglutinating salivary proteins, that allow immobilization of pathogens. In this work, superparamagnetic iron oxide nanoparticles with stable polycondensed aminoalkylsilane layer were developed, to which the heterobifunctional linkers N-succinimidyl 3-(2-pyridyldithio)-propanoate (SDPD) and N-succinimidyl bromoacetate (SBA) were bound. These linkers were further chemoselectively reacted with the thiol group of singularly present cysteines of selected peptides. The resulting functional nanoparticles underwent a detailed physicochemical characterization. The biocompatibility of the primarily coated aminoalkylsilane particles was also investigated. To test the pathogen-binding efficacy of the particles, the lipopolysaccharide-immobilization capacity of the peptide-coated particles was compared with free peptides. Here, one particle-bound peptide species succeeded in capturing 90% of the toxin, whereas the degree of immobilization of the toxin with a system that varied in the sequence of the peptide dropped to 35%. With these promising results, we hope to develop extracorporeal magnetic clearance systems for removing pathogens from the human body in order to accelerate diagnosis and alleviate acute disease conditions such as sepsis., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The imaging of reactive oxygen species (ROS) at the molecular level with high sensitivity and specificity by positron emission tomography (PET) could be of enormous interest to increase our knowledge about ROS activity and signalling, especially in tumours. The aim of this research was to optimise the click chemistry-based radiosynthesis of an 18 F-labelled aminoferrocene glycoconjugate that was derived from an N-alkylaminoferrocene lead structure known to have anticancer activity in vitro. Applying the solvent system phosphate buffer/THF (12/5), Cu(OAc) 2 and sodium ascorbate as reducing agent at 60°C, the alkyne 1 reacted with the 18 F-labelled glycosyl azide [ 18 F]2 in the presence of carrier 3 (47μM) to obtain carrier-added [ 18 F]4 in a radiochemical yield of 85%. Interestingly, the addition of carrier was essential for sufficient radiochemical yield, because it suppressed the oxidation of no-carrier-added (n.c.a.) [ 18 F]4. Future work will include the formulation of c.a. [ 18 F]4 for studying its biodistribution in tumour-bearing mice., (© 2018 John Wiley & Sons, Ltd.)

As new generations of targeted therapies emerge and tumor genome sequencing discovers increasingly comprehensive mutation repertoires, the functional relationships of mutations to tumor phenotypes remain largely unknown. Here, we measured ex vivo sensitivity of 246 blood cancers to 63 drugs alongside genome, transcriptome, and DNA methylome analysis to understand determinants of drug response. We assembled a primary blood cancer cell encyclopedia data set that revealed disease-specific sensitivities for each cancer. Within chronic lymphocytic leukemia (CLL), responses to 62% of drugs were associated with 2 or more mutations, and linked the B cell receptor (BCR) pathway to trisomy 12, an important driver of CLL. Based on drug responses, the disease could be organized into phenotypic subgroups characterized by exploitable dependencies on BCR, mTOR, or MEK signaling and associated with mutations, gene expression, and DNA methylation. Fourteen percent of CLLs were driven by mTOR signaling in a non-BCR-dependent manner. Multivariate modeling revealed immunoglobulin heavy chain variable gene (IGHV) mutation status and trisomy 12 as the most important modulators of response to kinase inhibitors in CLL. Ex vivo drug responses were associated with outcome. This study overcomes the perception that most mutations do not influence drug response of cancer, and points to an updated approach to understanding tumor biology, with implications for biomarker discovery and cancer care.

Aminoferrocene-based prodrugs are activated in cancer cells by reactive oxygen species (ROS). They were shown to exhibit high cytotoxicity towards a variety of cancer cell lines and primary cancer cells, but remain not toxic towards non-malignant cells. However, these prodrugs have rather high lipophilicity leading to relatively low water solubility. In particular, an n-octanol/water partition coefficient for the best aminoferrocene-based prodrug (2) was found to be 4.51±0.03. Though the approaches for decreasing lipophilicity are straightforward and include the addition of polar residues to the drug structure, these modifications also lead to dramatic decrease of cell permeability and, correspondingly, lower the activity of the drug. Therefore, a delicate balance of polar and unpolar groups should be found to reduce lipophilicity without compromising the useful drug properties. In this study we optimized an N-alkyl substituent, which is a key element responsible for the stabilization of the aminoferrocene drug released in cancer cells from prodrug 2. We found that an N-propargyl residue is an optimal replacement for the N-benzyl fragment. In particular, such a substitution (prodrug 7a) leads to reduction of prodrug lipophilicity down to logP=3.78±0.05, improvement of its water solubility, decrease of its propensity towards aggregation and dramatic increase of its ROS-generating properties. Finally, we demonstrated that the optimized prodrug strongly suppresses growth of Guerin's carcinoma (T8) in vivo at the dose of 30mg/kg., (Copyright © 2017. Published by Elsevier Inc.)

Because cellular uptake of anticancer Pt II and Pt IV drugs occurs by different mechanisms, the latter ones can exhibit substantial activity towards cells, which have either intrinsic or acquired resistance towards Pt II drugs. However, this positive effect is diminished due to reductive activation of Pt IV drugs in extracellular space that can be one of the reasons why they have not yet been approved for clinical use despite over 60 clinical trials conducted worldwide. Herein, we suggest a solution to this problem by achieving highly specific intracellular versus extracellular prodrug reduction. In particular, we prepared a hybrid Pt IV prodrug containing two pro-reductants. This hybrid was uptaken by cells, the pro-reductants were activated in the cancer-specific microenvironment (high H 2 O 2 ), and reduced Pt IV by two one-electron transfers. The drug formed in this way induced cell death both in cisplatin-sensitive and resistant cell lines, but remained nontoxic to normal cells., (© 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

We developed a simple protocol for high-yielding synthesis of conjugates of a deuterated dihydro-N,N,N',N'-tetramethylrhodamine (F*) with oligodeoxyribonucleotides and a 2'-OMe RNA (a representative nuclease-resistant, chemically modified oligonucleotide) using easily accessible starting materials including NaBD4 and conjugates of oligonucleotides with N,N,N',N'-tetramethylrhodamine (F). These compounds were found to be stable in air and insensitive to light at 525, 635 and 650 nm, whereas slow activation occurs upon their exposure to 470 nm light. However, at the conditions of the templated reaction, in the presence of a target nucleic acid and a photocatalyst based on the eosin structure, the F* is oxidized forming fluorescent F. This reaction is >30-fold faster than the background reaction in the absence of the template. Moreover, the presence of a single mismatch in the target nucleic acid slows down the templated reaction by eightfold. These activatable dyes can potentially find applications as nucleic acid-specific probes for super-resolution imaging in live cells.

Peptide nucleic acid (PNA) is a non-cyclic pseudopeptide-nucleic acid structural mimic with promising applications within diagnostics and drug discovery. This review focuses on metal complex derivatives of PNA. Metal ions and their complexes display unique physical and chemical properties and offer the opportunity to introduce new labels and probes for bioanalytical and diagnostic applications of PNA, but also to modulate or to introduce new (for example catalytic) functions and biological activities.

Only two nucleic acid directed chemical reactions that are compatible with live cells have been reported to date. Neither of these processes generate toxic species from nontoxic starting materials. Reactions of the latter type could be applied as gene-specific drugs, for example, in the treatment of cancer. We report here the first example of a chemical reaction that generates a cytotoxic drug from a nontoxic prodrug in the presence of a specific endogeneous ribonucleic acid in live mammalian cells. In this case, the prodrug is triplet oxygen and the drug is singlet oxygen. The key component of this reaction is an inert molecule (InP-2'-OMe-RNA/Q-2'-OMe-RNA; P: photosensitizer; Q: quencher), which becomes an active photosensitizer (InP-2'-OMe-RNA) in the presence of single-stranded nucleic acid targets. Upon irradiation with red light, the photosensitizer produces over 6000 equivalents of toxic singlet oxygen per nucleic acid target. This reaction is highly sequence specific. To detect the generation of singlet oxygen in live cells, we prepared a membrane-permeable and water-soluble fluorescent scavenger, a derivative of 2,5-diphenylisobenzofurane. The scavenger decomposes upon reaction with singlet oxygen and this is manifested in a decrease in the fluorescence intensity. This effect can be conveniently monitored by flow cytometry.

With the improvement and completeness of genome databases, it has become possible to develop a novel fluorescence in situ hybridization (FISH) technique called COMBinatorial Oligo FISH (COMBO-FISH). In contrast to other (standard) FISH applications, COMBO-FISH makes use of a bioinformatic approach for probe set design. By means of computer genome database search, oligonucleotide stretches of typical lengths of 15-30 nucleotides are selected in such a way that they all colocalize within a given genome (gene) target. Typically, probe sets of about 20-40 stretches are designed within 50-250 kb, which is enough to get an increased fluorescence signal specifically highlighting the target from the background. Although "specific colocalization" is the only necessary condition for probe selection, i.e. the probes of different lengths can be composed of purines and pyrimidines, we additionally refined the design strategy restricting the probe sets to homopurine or homopyrimidine oligonucleotides so that depending on the probe orientation either double (requiring denaturation of the target double strand) or triple (omitting denaturation of the target strand) strand bonding of the probes is possible. The probes used for the protocols described below are DNA or PNA oligonucleotides, which can be synthesized by established automatized techniques. We describe different protocols that were successfully applied to label gene targets via double- or triple-strand bonding in fixed lymphocyte cell cultures, bone marrow smears, and formalin-fixed, paraffin-wax embedded tissue sections. In addition, we present a procedure of probe microinjection in living cells resulting in specific labeling when microscopically detected after fixation.