Releasable Luciferin−Transporter Conjugates: Tools for the Real-Time Analysis of Cellular Uptake and Release

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Interest in algae has significantly accelerated with the increasing recognition of their potentially unique role in medical, materials, energy, bioremediation, and synthetic biological research. However, the introduction of tools to study, control, or expand the inner-workings of algae has lagged behind. Here we describe a general molecular method based on guanidinium-rich molecular transporters (GR-MoTrs) for bringing small and large cargos into algal cells. Significantly, this method is shown to work in wild-type algae that have an intact cell wall. Developed using Chlamydomonas reinhardtii , this method is also successful with less studied algae including Neochloris oleoabundans and Scenedesmus dimorphus thus providing a new and versatile tool for algal research.

Section: Resultsmentioning

confidence: 99%

A molecular method for the delivery of small molecules and proteins across the cell wall of algae using molecular transporters

Hyman

Geihe

Trantow

et al. 2012

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“…As a prelude to animal studies with Taxol conjugates, a third small-molecule system, luciferin, the substrate for firefly luciferase, was used as a drug surrogate to compare the biodistribution and pharmacokinetics of the r8 drug surrogate conjugate and the surrogate alone. Luciferin and luciferin conjugated to r8 (7) (38,39), at concentrations equimolar to those used in the studies with Taxol (5 mg/kg), were delivered via i.p. injection into transgenic mice ubiquitously expressing firefly luciferase (Fig.…”

Section: Biodistribution and Pharmacokinetics Of An Octaarginine-lucimentioning

confidence: 99%

Overcoming multidrug resistance of small-molecule therapeutics through conjugation with releasable octaarginine transporters

Dubikovskaya¹,

Thorne

Pillow³

et al. 2008

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224

213

Many cancer therapeutic agents elicit resistance that renders them ineffective and often produces cross-resistance to other drugs. One of the most common mechanisms of resistance involves P-glycoprotein (Pgp)-mediated drug efflux. To address this problem, new agents have been sought that are less prone to inducing resistance and less likely to serve as substrates for Pgp efflux. An alternative to this approach is to deliver established agents as molecular transporter conjugates into cells through a mechanism that circumvents Pgp-mediated efflux and allows for release of free drug only after cell entry. Here we report that the widely used chemotherapeutic agent Taxol, ineffective against Taxol-resistant human ovarian cancer cell lines, can be incorporated into a releasable octaarginine conjugate that is effective against the same Taxolresistant cell lines. It is significant that the ability of the Taxol conjugates to overcome Taxol resistance is observed both in cell culture and in animal models of ovarian cancer. The generality and mechanistic basis for this effect were also explored with coelenterazine, a Pgp substrate. Although coelenterazine itself does not enter cells because of Pgp efflux, its octaarginine conjugate does so readily. This approach shows generality for overcoming the multidrug resistance elicited by small-molecule cancer chemotherapeutics and could improve the prognosis for many patients with cancer and fundamentally alter search strategies for novel therapeutic agents that are effective against resistant disease.ovarian cancer ͉ Pgp ͉ Taxol ͉ imaging ͉ luciferase

“…Work from our laboratory and others has demonstrated the utility of caged luciferins in vivo (28)(29)(30) for measuring transient small molecules (31)(32)(33)(34), enzyme and transporter activities (34)(35)(36)(37)(38)(39)(40)(41)(42)(43)(44)(45)(46), protein-protein and cell-cell interactions (42,47,48), and copper (49). Indeed, previous work from our laboratory utilized a Cu-dependent oxidation reaction to uncage luciferin for in vivo copper imaging (50), a first demonstration of a general activitybased sensing (ABS) strategy which we envisioned expanding to other essential metals in biology by changing the reaction trigger.…”

mentioning

confidence: 99%

In vivo bioluminescence imaging of labile iron accumulation in a murine model of Acinetobacter baumannii infection

Aron

Heffern

Lonergan

et al. 2017

118

Iron is an essential metal for all organisms, yet disruption of its homeostasis, particularly in labile forms that can contribute to oxidative stress, is connected to diseases ranging from infection to cancer to neurodegeneration. Iron deficiency is also among the most common nutritional deficiencies worldwide. To advance studies of iron in healthy and disease states, we now report the synthesis and characterization of iron-caged luciferin-1 (ICL-1), a bioluminescent probe that enables longitudinal monitoring of labile iron pools (LIPs) in living animals. ICL-1 utilizes a bioinspired endoperoxide trigger to release D-aminoluciferin for selective reactivity-based detection of Fe 2+ with metal and oxidation state specificity. The probe can detect physiological changes in labile Fe 2+ levels in live cells and mice experiencing iron deficiency or overload. Application of ICL-1 in a model of systemic bacterial infection reveals increased iron accumulation in infected tissues that accompany transcriptional changes consistent with elevations in both iron acquisition and retention. The ability to assess iron status in living animals provides a powerful technology for studying the contributions of iron metabolism to physiology and pathology.labile iron | molecular imaging | luciferin | metal homeostasis | infectious disease I ron is an essential mineral for nearly every form of life, owing in large part to its ability to cycle between different oxidation states for processes such as nucleotide synthesis, oxygen transport, and respiration (1, 2). At the same time, the potent redox activity of iron is potentially toxic, particularly in unregulated labile forms that can trigger aberrant production of reactive oxygen species via Fenton chemistry (3). Indeed, iron deficiency remains one of the most common nutritional deficiencies in the world (4), and aberrant iron levels have been linked to various ailments, including cancer (5-7), cardiovascular (8), and neurodegenerative (9) disorders, as well as aging (10). The situation is especially complex in infectious diseases, where the requirement for iron by both host organism and invading pathogen leads to an intricate chemical tug-of-war for this metal nutrient during various stages of the immune response (11,12).The foregoing examples provide motivation for developing technologies to monitor biological iron status, with particular interest in methods to achieve in vivo iron imaging in live animal models that go beyond current state-of-the-art assays that are limited primarily to cell culture specimens. In this regard, detection of iron with both metal and oxidation state specificity is of central importance, because while iron is stored primarily in the ferric oxidation state, a ferrous iron pool loosely bound to cellular ligands, defined as the labile iron pool (LIP), exists at the center of highly regulated networks that control iron acquisition, trafficking, and excretion. Indeed, as a weak binder on the Irving-Williams stability series (13), Fe 2+ provides a challenge for d...