Mobile zinc as a modulator of sensory perception.

Mobile zinc is an abundant transition metal ion in the central nervous system, with pools of divalent zinc accumulating in regions of the brain engaged in sensory perception and memory formation. Here, we present essential tools that we developed to interrogate the role(s) of mobile zinc in these processes. Most important are (a) fluorescent sensors that report the presence of mobile zinc and (b) fast, Zn-selective chelating agents for measuring zinc flux in animal tissue and live animals.

Aneuploidy increases resistance to chemotherapeutics by antagonizing cell division.

Aneuploidy, defined as whole chromosome gains and losses, is associated with poor patient prognosis in many cancer types. However, the condition causes cellular stress and cell cycle delays, foremost in G1 and S phase. Here, we investigate how aneuploidy causes both slow proliferation and poor disease outcome.

Enhancing chemotherapy response through augmented synthetic lethality by co-targeting nucleotide excision repair and cell-cycle checkpoints.

In response to DNA damage, a synthetic lethal relationship exists between the cell cycle checkpoint kinase MK2 and the tumor suppressor p53. Here, we describe the concept of augmented synthetic lethality (ASL): depletion of a third gene product enhances a pre-existing synthetic lethal combination. We show that loss of the DNA repair protein XPA markedly augments the synthetic lethality between MK2 and p53, enhancing anti-tumor responses alone and in combination with cisplatin chemotherapy.

A Platinum(IV) Prodrug-Perfluoroaryl Macrocyclic Peptide Conjugate Enhances Platinum Uptake in the Brain.

Effective delivery to the brain limits the development of novel glioblastoma therapies. Here, we introduce conjugation between platinum(IV) prodrugs of cisplatin and perfluoroaryl peptide macrocycles to increase brain uptake. We demonstrate that one such conjugate shows efficacy against glioma stem-like cells.

Native Zinc Catalyzes Selective and Traceless Release of Small Molecules in β-Cells.

The loss of insulin-producing β-cells is the central pathological event in type 1 and 2 diabetes, which has led to efforts to identify molecules to promote β-cell proliferation, protection, and imaging. However, the lack of β-cell specificity of these molecules jeopardizes their therapeutic potential. A general platform for selective release of small-molecule cargoes in β-cells over other islet cells or other cell-types in an organismal context will be immensely valuable in advancing diabetes research and therapeutic development.

HaloTag-Based Hybrid Targetable and Ratiometric Sensors for Intracellular Zinc.

We report a new series of small molecule-protein hybrid zinc sensors that combine genetic targetability with the spectroscopic profile of synthetic fluorophores. We functionalized the zinc sensor ZinPyr-1 (ZP1) with a chloroalkane linker (ZP1-12Cl) that reacts specifically with the engineered protein HaloTag. The resulting construct, ZP1-HaloTag, binds zinc ions with a threefold fluorescence enhancement.

Sensing intracellular calcium ions using a manganese-based MRI contrast agent.

Calcium ions are essential to signal transduction in virtually all cells, where they coordinate processes ranging from embryogenesis to neural function. Although optical probes for intracellular calcium imaging have been available for decades, the development of probes for noninvasive detection of intracellular calcium signaling in deep tissue and intact organisms remains a challenge. To address this problem, we synthesized a manganese-based paramagnetic contrast agent, ManICS1-AM, designed to permeate cells, undergo esterase cleavage, and allow intracellular calcium levels to be monitored by magnetic resonance imaging (MRI).

DNA Intercalation Facilitates Efficient DNA-Targeted Covalent Binding of Phenanthriplatin.

Phenanthriplatin, a monofunctional anticancer agent derived from cisplatin, shows significantly more rapid DNA covalent-binding activity compared to its parent complex. To understand the underlying molecular mechanism, we used single-molecule studies with optical tweezers to probe the kinetics of DNA-phenanthriplatin binding as well as DNA binding to several control complexes. The time-dependent extensions of single λ-DNA molecules were monitored at constant applied forces and compound concentrations, followed by rinsing with a compound-free solution.

A Sensitive, Nonradioactive Assay for Zn(II) Uptake into Metazoan Cells.

Sensitive measurements of cellular Zn(II) uptake currently rely on quantitating radioactive emissions from cells treated with Zn(II). Here, we describe a straightforward and reliable method employing a stable isotope to sensitively measure Zn(II) uptake by metazoan cells. First, biological medium selectively depleted of natural abundance Zn(II) using A12-resin [Richardson, C.

Superiority of SpiroZin2 Versus FluoZin-3 for monitoring vesicular Zn allows tracking of lysosomal Zn pools.

Small-molecule fluorescent probes are powerful and ubiquitous tools for measuring the concentration and distribution of analytes in living cells. However, accurate characterization of these analytes requires rigorous evaluation of cell-to-cell heterogeneity in fluorescence intensities and intracellular distribution of probes. In this study, we perform a parallel and systematic comparison of two small-molecule fluorescent vesicular Zn probes, FluoZin-3 AM and SpiroZin2, to evaluate each probe for measurement of vesicular Zn pools.

Glutathione-Scavenging Poly(disulfide amide) Nanoparticles for the Effective Delivery of Pt(IV) Prodrugs and Reversal of Cisplatin Resistance.

Despite the broad antitumor spectrum of cisplatin, its therapeutic efficacy in cancer treatment is compromised by the development of drug resistance in tumor cells and systemic side effects. A close correlation has been drawn between cisplatin resistance in tumor cells and increased levels of intracellular thiol-containing species, especially glutathione (GSH). The construction of a unique nanoparticle (NP) platform composed of poly(disulfide amide) polymers with a high disulfide density for the effective delivery of Pt(IV) prodrugs capable of reversing cisplatin resistance through the disulfide-group-based GSH-scavenging process, as described herein, is a promising route by which to overcome limitations associated with tumor resistance.

Interaction of the New Monofunctional Anticancer Agent Phenanthriplatin With Transporters for Organic Cations.

Cancer treatment with platinum compounds is an important achievement of modern chemotherapy. However, despite the beneficial effects, the clinical impact of these agents is hampered by the development of drug resistance as well as dose-limiting side effects. The efficacy but also side effects of platinum complexes can be mediated by uptake through plasma membrane transporters.

Speciation of Phenanthriplatin and Its Analogs in the Core of Tobacco Mosaic Virus.

Efficient loading of drugs in novel delivery agents has the potential to substantially improve therapy by targeting the diseased tissue while avoiding unwanted side effects. Here we report the first systematic study of the loading mechanism of phenanthriplatin and its analogs into tobacco mosaic virus (TMV), previously used by our group as an efficient carrier for anticancer drug delivery. A detailed investigation of the preferential uptake of phenanthriplatin in its aquated form (∼2000 molecules per TMV particle versus ∼1000 for the chlorido form) is provided.

The effect of geometric isomerism on the anticancer activity of the monofunctional platinum complex trans-[Pt(NH)(phenanthridine)Cl]NO.

A trans-DDP based monofunctional phenanthridine Pt(ii) complex was synthesized and characterized. Its anticancer activity was studied in vitro on a panel of human cancer cell lines and mouse intestinal cancer organoids. This complex displays significant antitumor properties, with a different spectrum of activity than that of classic bifunctional cross-linking agents like cisplatin.

Photoactivatable Sensors for Detecting Mobile Zinc.

Fluorescent sensors for mobile zinc are valuable for studying complex biological systems. Because these sensors typically bind zinc rapidly and tightly, there has been little temporal control over the activity of the probe after its application to a sample. The ability to control the activity of a zinc sensor in vivo during imaging experiments would greatly improve the time resolution of the measurement.

A Method for Selective Depletion of Zn(II) Ions from Complex Biological Media and Evaluation of Cellular Consequences of Zn(II) Deficiency.

We describe the preparation, evaluation, and application of an S100A12 protein-conjugated solid support, hereafter the "A12-resin", that can remove 99% of Zn(II) from complex biological solutions without significantly perturbing the concentrations of other metal ions. The A12-resin can be applied to selectively deplete Zn(II) from diverse tissue culture media and from other biological fluids, including human serum. To further demonstrate the utility of this approach, we investigated metabolic, transcriptomic, and metallomic responses of HEK293 cells cultured in medium depleted of Zn(II) using S100A12.

Challenges and Opportunities in Brain Bioinorganic Chemistry.

Metal ions play critical roles in neurotransmission, memory formation, and sensory perception. Understanding the molecular details of these processes is the Holy Grail of metalloneurochemistry. Here we describe five challenges for collaborative teams of chemists, biologists, and neuroscientists to help make this dream a reality.

Tuning the Diiron Core Geometry in Carboxylate-Bridged Macrocyclic Model Complexes Affects Their Redox Properties and Supports Oxidation Chemistry.

We introduce a novel platform to mimic the coordination environment of carboxylate-bridged diiron proteins by tethering a small, dangling internal carboxylate, (CH)COOH, to phenol-imine macrocyclic ligands (HPIMICn). In the presence of an external bulky carboxylic acid (RCOH), the ligands react with [Fe(Mes)] (Mes = 2,4,6-trimethylphenyl) to afford dinuclear [Fe(PIMICn)(RCO)(MeCN)] (n = 4-6) complexes. X-ray diffraction studies revealed structural similarities between these complexes and the reduced diiron active sites of proteins such as Class I ribonucleotide reductase (RNR) R2 and soluble methane monooxygenase hydroxylase.

CFH, a Hydrogen Bond Donor.

The CFH group, a potential surrogate for the OH group, can act as an unusual hydrogen bond donor, as confirmed by crystallographic, spectroscopic, and computational methods. Here, we demonstrate the bioisosterism of the OH and CFH groups and the important roles of CF-H···O hydrogen bonds in influencing intermolecular interactions and conformational preferences. Experimental evidence, corroborated by theory, reveals the distinctive nature of CFH hydrogen bonding interactions relative to their normal OH hydrogen bonding counterparts.

A subset of platinum-containing chemotherapeutic agents kills cells by inducing ribosome biogenesis stress.

Cisplatin and its platinum analogs, carboplatin and oxaliplatin, are some of the most widely used cancer chemotherapeutics. Although cisplatin and carboplatin are used primarily in germ cell, breast and lung malignancies, oxaliplatin is instead used almost exclusively to treat colorectal and other gastrointestinal cancers. Here we utilize a unique, multi-platform genetic approach to study the mechanism of action of these clinically established platinum anti-cancer agents, as well as more recently developed cisplatin analogs.

Metalloneurochemistry and the Pierian Spring: 'Shallow Draughts Intoxicate the Brain'.

Metal ions perform critical and diverse functions in nervous system physiology and pathology. The field of metalloneurochemistry aims to understand the mechanistic bases for these varied roles at the molecular level. Here, we review several areas of research that illustrate progress toward achieving this ambitious goal and identify key challenges for the future.

Repair shielding of platinum-DNA lesions in testicular germ cell tumors by high-mobility group box protein 4 imparts cisplatin hypersensitivity.

Cisplatin is the most commonly used anticancer drug for the treatment of testicular germ cell tumors (TGCTs). The hypersensitivity of TGCTs to cisplatin is a subject of widespread interest. Here, we show that high-mobility group box protein 4 (HMGB4), a protein preferentially expressed in testes, uniquely blocks excision repair of cisplatin-DNA adducts, 1,2-intrastrand cross-links, to potentiate the sensitivity of TGCTs to cisplatin therapy.

Mobile zinc increases rapidly in the retina after optic nerve injury and regulates ganglion cell survival and optic nerve regeneration.

Retinal ganglion cells (RGCs), the projection neurons of the eye, cannot regenerate their axons once the optic nerve has been injured and soon begin to die. Whereas RGC death and regenerative failure are widely viewed as being cell-autonomous or influenced by various types of glia, we report here that the dysregulation of mobile zinc (Zn) in retinal interneurons is a primary factor. Within an hour after the optic nerve is injured, Zn increases several-fold in retinal amacrine cell processes and continues to rise over the first day, then transfers slowly to RGCs via vesicular release.