Electron Spin Resonance Probe Incorporation into Bioinks Permits Longitudinal Oxygen Imaging of Bioprinted Constructs.

Purpose: Bioprinting is an additive manufacturing technology analogous to 3D printing. Instead of plastic or resin, cell-laden hydrogels are used to produce a construct of the intended biological structure. Over time, cells transform this construct into a functioning tissue or organ.

Dose-Specific Intratumoral GM-CSF Modulates Breast Tumor Oxygenation and Antitumor Immunity.

GM-CSF has been employed as an adjuvant to cancer immunotherapy with mixed results based on dosage. We previously showed that GM-CSF regulated tumor angiogenesis by stimulating soluble vascular endothelial growth factor (VEGF) receptor-1 from monocytes/macrophages in a dose-dependent manner that neutralized free VEGF, and intratumoral injections of high-dose GM-CSF ablated blood vessels and worsened hypoxia in orthotopic polyoma middle T Ag (PyMT) triple-negative breast cancer (TNBC). In this study, we assessed both immunoregulatory and oxygen-regulatory components of low-dose versus high-dose GM-CSF to compare effects on tumor oxygen, vasculature, and antitumor immunity.

In Vivo Electron Paramagnetic Resonance Molecular Profiling of Tumor Microenvironment upon Tumor Progression to Malignancy in an Animal Model of Breast Cancer.

Purpose: Hypoxia and acidosis are recognized tumor microenvironment (TME) biomarkers of cancer progression. Alterations in cancer redox status and metabolism are also associated with elevated levels of intracellular glutathione (GSH) and interstitial inorganic phosphate (Pi). This study aims to evaluate the capability of these biomarkers to discriminate between stages and inform on a switch to malignancy.

Improving combination therapies: targeting A2B-adenosine receptor to modulate metabolic tumor microenvironment and immunosuppression.

Background: We investigated the role of A2B-adenosine receptor in regulating immunosuppressive metabolic stress in the tumor microenvironment. Novel A2B-adenosine receptor antagonist PBF-1129 was tested for antitumor activity in mice and evaluated for safety and immunologic efficacy in a phase I clinical trial of patients with non-small cell lung cancer.

Methods: The antitumor efficacy of A2B-adenosine receptor antagonists and their impact on the metabolic and immune tumor microenvironment were evaluated in lung, melanoma, colon, breast, and epidermal growth factor receptor-inducible transgenic cancer models.

Rapid scan EPR: Automated digital resonator control for low-latency data acquisition.

Automation has become an essential component of modern scientific instruments which often capture large amounts of complex dynamic data. Algorithms are developed to read multiple sensors in parallel with data acquisition and to adjust instrumental parameters on the fly. Decisions are made on a time scale unattainable to the human operator.

Rapid Scan EPR Oxygen Imaging in Photoactivated Resin Used for Stereolithographic 3D Printing.

Oxygen plays a critical role in the photopolymerization process resulting in the formation of solid structures from liquid resins during three-dimensional (3D) printing: it acts as a polymerization inhibitor. Upon exposure to light, oxygen is depleted. As a result, the polymerization process becomes activated.

Rapid Scan EPR imaging as a Tool for Magnetic Field Mapping.

Functional four-dimensional spectral-spatial electron paramagnetic imaging (EPRI) is routinely used in biomedical research. Positions and widths of EPR lines in the spectral dimension report oxygen partial pressure, pH, and other important parameters of the tissue microenvironment. Images are measured in the homogeneous external magnetic field.

Heterogeneity of radiation response in mesenchymal subtype glioblastoma: molecular profiling and reactive oxygen species generation.

Background: Radiotherapy-induced tumor death remains critical in the successful first-line management of glioblastoma, whereas resistance to radiation serves as a major factor in disease progression. Mesenchymal shift has been identified as a driver in GBM recurrence, with gene expression associated with enhanced repair of macromolecular damage caused by radiation.

Methods: Using distinct mesenchymal subtype GBM cells lines, radiation response was assessed by clonogenic assay and orthotopic mouse tumor model.

Tuning the size and composition of manganese oxide nanoparticles through varying temperature ramp and aging time.

Manganese oxide (MnO) nanoparticles (NPs) can serve as robust pH-sensitive contrast agents for magnetic resonance imaging (MRI) due to Mn2+ release at low pH, which generates a ~30 fold change in T1 relaxivity. Strategies to control NP size, composition, and Mn2+ dissolution rates are essential to improve diagnostic performance of pH-responsive MnO NPs. We are the first to demonstrate that MnO NP size and composition can be tuned by the temperature ramping rate and aging time used during thermal decomposition of manganese(II) acetylacetonate.

Hypoxia-Inducible Factor α Subunits Regulate Tie2-Expressing Macrophages That Influence Tumor Oxygen and Perfusion in Murine Breast Cancer.

Tie2-expressing monocytes/macrophages (TEMs) are a distinct subset of proangiogenic monocytes selectively recruited to tumors in breast cancer. Because of the hypoxic nature of solid tumors, we investigated if oxygen, via hypoxia-inducible transcription factors HIF-1α and HIF-2α, regulates TEM function in the hypoxic tumor microenvironment. We orthotopically implanted PyMT breast tumor cells into the mammary fat pads of syngeneic LysMcre, HIF-1α /LysMcre, or HIF-2α /LysMcre mice and evaluated the tumor TEM population.

Chemical exchange induced Hahn echo modulation in pulsed electron paramagnetic resonance experiment.

Triarylmethyl (TAM) radicals are probes well suited to study microenvironment parameters using pulse electron paramagnetic resonance (EPR) due to their long relaxation times. Here, we are reporting the study of relaxation properties of monophosphonated TAM radicals in the presence of chemical exchange processes. The dependence of Hahn and inversion recovery echo on the solution pH and the chemical exchange rate are discussed.

Development of multifunctional Overhauser-enhanced magnetic resonance imaging for concurrent in vivo mapping of tumor interstitial oxygenation, acidosis and inorganic phosphate concentration.

Tumor oxygenation (pO), acidosis (pH) and interstitial inorganic phosphate concentration (Pi) are important parameters of the malignant behavior of cancer. A noninvasive procedure that enables visualization of these parameters may provide unique information about mechanisms of tumor pathophysiology and provide clues to new treatment targets. In this research, we present a multiparametric imaging method allowing for concurrent mapping of pH, spin probe concentration, pO, and Pi using a single contrast agent and Overhauser-enhanced magnetic resonance imaging technique.

Modular imaging system: Rapid scan EPR at 800 MHz.

An electron paramagnetic resonance (EPR) imaging system has been custom built for use in pre-clinical and, potentially, clinical studies. Commercial standalone modules have been used in the design that are MATLAB-controlled. The imaging system combines digital and analog technologies.

Oxygen-induced leakage of spin polarization in Overhauser-enhanced magnetic resonance imaging: Application for oximetry in tumors.

Overhauser-enhanced Magnetic Resonance Imaging (OMRI) is a double resonance technique applied for oxygen imaging in aqueous samples and biological tissues. In this report, we present an improved OMRI approach of oxygen measurement using the single line "Finland" trityl spin probe. Compared to a traditional approach, we introduced an additional mechanism of leakage of spin polarization due to an interaction of a spin system with oxygen.

A combined positron emission tomography (PET)-electron paramagnetic resonance imaging (EPRI) system: initial evaluation of a prototype scanner.

The advent of hybrid scanners, combining complementary modalities, has revolutionized the application of advanced imaging technology to clinical practice and biomedical research. In this project, we investigated the melding of two complementary, functional imaging methods: positron emission tomography (PET) and electron paramagnetic resonance imaging (EPRI). PET radiotracers can provide important information about cellular parameters, such as glucose metabolism.

In Vivo EPR Assessment of pH, pO2, Redox Status, and Concentrations of Phosphate and Glutathione in the Tumor Microenvironment.

This protocol demonstrates the capability of low-field electron paramagnetic resonance (EPR)-based techniques in combination with functional paramagnetic probes to provide quantitative information on the chemical tumor microenvironment (TME), including pO2, pH, redox status, concentrations of interstitial inorganic phosphate (Pi), and intracellular glutathione (GSH). In particular, an application of a recently developed soluble multifunctional trityl probe provides unsurpassed opportunity for in vivo concurrent measurements of pH, pO2 and Pi in Extracellular space (HOPE probe). The measurements of three parameters using a single probe allow for their correlation analyses independent of probe distribution and time of the measurements.

LPA signaling is regulated through the primary cilium: a novel target in glioblastoma.

The primary cilium is a ubiquitous organelle presented on most human cells. It is a crucial signaling hub for multiple pathways including growth factor and G-protein coupled receptors. Loss of primary cilia, observed in various cancers, has been shown to affect cell proliferation.

Triarylmethyl-based biradical as a superoxide probe.

Superoxide radical represents one of the most biologically relevant reactive oxygen species involved in numerous physiological and pathophysiological processes. Superoxide measurement through the decay of an electron paramagnetic resonance (EPR) signal of a triarylmethyl (TAM) radical possesses the advantage of a high selectivity and relatively high rate constant of TAM reaction with the superoxide. Hereby we report a straightforward synthesis and characterization of a TAM-TAM biradical showing a high reactivity with superoxide (second-order rate constant, (6.

Exchange Phenomena in the Electron Paramagnetic Resonance Spectra of the Nitroxyl and Trityl Radicals: Multifunctional Spectroscopy and Imaging of Local Chemical Microenvironment.

This Feature overviews the basic principles of using stable organic radicals involved in reversible exchange processes as functional paramagnetic probes. We demonstrate that these probes in combination with electron paramagnetic resonance (EPR)-based spectroscopy and imaging techniques provide analytical tools for quantitative mapping of critical parameters of local chemical microenvironment. The Feature is written to be understandable to people who are laymen to the EPR field in anticipation of future progress and broad application of these tools in biological systems, especially in vivo, over the next years.

Interstitial Inorganic Phosphate as a Tumor Microenvironment Marker for Tumor Progression.

Noninvasive in vivo assessment of chemical tumor microenvironment (TME) parameters such as oxygen (pO), extracellular acidosis (pH), and concentration of interstitial inorganic phosphate (Pi) may provide unique insights into biological processes in solid tumors. In this work, we employ a recently developed multifunctional trityl paramagnetic probe and electron paramagnetic resonance (EPR) technique for in vivo concurrent assessment of these TME parameters in various mouse models of cancer. While the data support the existence of hypoxic and acidic regions in TME, the most dramatic differences, about 2-fold higher concentrations in tumors vs.

Nitro-Triarylmethyl Radical as Dual Oxygen and Superoxide Probe.

Superoxide radical is involved in numerous physiological and pathophysiological processes. Tetrathiatriarylmethyl (TAM) radicals are known to react with superoxide allowing measurement of superoxide production in biological media. We report the synthesis of a Nitro conjugated TAM radical showing a rate constant of 7 × 10 M s which is two order of magnitude higher than other TAMs, allowing high sensitivity measurement of superoxide.

Concurrent Longitudinal EPR Monitoring of Tissue Oxygenation, Acidosis, and Reducing Capacity in Mouse Xenograft Tumor Models.

Tissue oxygenation, extracellular acidity, and tissue reducing capacity are among crucial parameters of tumor microenvironment (TME) of significant importance for tumor pathophysiology. In this paper, we demonstrate the complementary application of particulate lithium octa-n-butoxy-naphthalocyanine and soluble nitroxide paramagnetic probes for monitoring of these TME parameters using electron paramagnetic resonance (EPR) technique. Two different types of therapeutic interventions were studied: hypothermia and systemic administration of metabolically active drug.

Poly-arginine conjugated triarylmethyl radical as intracellular spin label.

Stable triarylmethyl radicals are ideal spin labels used for biomedical electron paramagnetic resonance applications. Previously reported structures exhibit polar charged functions for water solubilization preventing them from crossing the cell membrane. We report the synthesis of a triarylmethyl radical conjugated to poly-arginine peptide allowing intracellular delivery of the paramagnetic label.

Effect of Sterical Shielding on the Redox Properties of Imidazoline and Imidazolidine Nitroxides.

The oxidant properties of the series of 2,2,5,5-tetraalkyl imidazoline and imidazolidine nitroxides were investigated. An increase in the number of bulky alkyl substituents leads to a decrease in the rate of reduction with ascorbate, which makes the electrochemical reduction potential more negative and shifts the equilibrium in the mixture of nitroxide and reference hydroxylamine (3-carboxy-1-hydroxy-2,2,5,5-tetramethylpyrrolidine-1-oxyl-1-(15)N) toward the starting compounds. The effect of structural factors on these reactions was analyzed by means of multiple regression using the Fujita steric constant Es and the inductive Hammett constant σI.