Type 1 diabetes (T1D) is an autoimmune disease that leads to the loss of insulin-producing beta cells. Bioartificial pancreas (BAP) or beta cell replacement strategies have shown promise in curing T1D and providing long-term insulin independence. Hypoxia (low oxygen concentration) that may occur in the BAP devices due to cell oxygen consumption at the early stages after implantation damages the cells, in addition to imposing limitations to device dimensions when translating promising results from rodents to humans. Finding ways to provide cells with sufficient oxygenation remains the major challenge in realizing BAP devices' full potential. Therefore, in vitro oxygen imaging assessment of BAP devices is crucial for predicting the devices' in vivo efficiency. Electron paramagnetic resonance oxygen imaging (EPROI, also known as electron MRI or eMRI) is a unique imaging technique that delivers absolute partial pressure of oxygen (pO2) maps and has been used for cancer hypoxia research for decades. However, its applicability for assessing BAP devices has not been explored. EPROI utilizes low magnetic fields in the mT range, static gradients, and the linear relationship between the spin-lattice relaxation rate (R1) of oxygen-sensitive spin probes such as trityl OX071 and pO2 to generate oxygen maps in tissues. With the support of the Juvenile Diabetes Research Foundation (JDRF), an academic-industry partnership consortium, the "Oxygen Measurement Core" was established at O2M to perform oxygen imaging assessment of BAP devices originated from core members' laboratories. This article aims to establish the protocols and demonstrate a few examples of in vitro oxygen imaging of BAP devices using EPROI. All pO2 measurements were performed using a recently introduced 720 MHz/25 mT preclinical oxygen imager instrument, JIVA-25™. We began by performing pO2 calibration of the biomaterials used in BAPs at 25 mT magnetic field since no such data exist. We compared the EPROI pO2 measurement with a single-point probe for a few selected materials. We also performed trityl OX071 toxicity studies with fibroblasts, as well as insulin-producing cells (beta TC6, MIN6, and human islet cells). Finally, we performed proof-of-concept in vitro pO2 imaging of five BAP devices that varied in size, shape, and biomaterials. We demonstrated that EPROI is compatible with commonly used biomaterials and that trityl OX071 is nontoxic to cells. A comparison of the EPROI with a fluorescent-based point oxygen probe in selected biomaterials showed higher accuracy of EPROI. The imaging of typically heterogenous BAP devices demonstrated the utility of obtaining oxygen maps over single-point measurements. In summary, we present EPROI as a quality control tool for developing efficient cell transplantation devices and artificial tissue grafts. Although the focus of this work is encapsulation systems for diabetes, the techniques developed in this project are easily transferable to other biomaterials, tissue grafts, and cell therapy devices used in the field of tissue engineering and regenerative medicine (TERM). In summary, EPROI is a unique noninvasive tool to experimentally study oxygen distribution in cell transplantation devices and artificial tissues, which can revolutionize the treatment of degenerative diseases like T1D.
© 2023. Springer Nature Limited.

In this study, we focus on investigating the therapeutic effects of camptothesome on treating metastatic triple-negative breast cancer (TNBC). We elucidate that camptothesome elicited stronger immunogenic cell death (ICD) compared to free camptothecin (CPT) and Onivyde in 4T1 TNBC cells. In addition, camptothesome is mainly internalized by the 4T1 and MDA-MB-231 cells through clathrin-mediated endocytosis based on the results of flow cytometry. Through real-time Lago optical imaging, camptothesome shows excellent tumor-targeting efficiency in orthotopic TNBC tumors. We demonstrate that camptothesome can upregulate programmed death-ligand 1 (PD-L1) in 4T1 tumors in an interferon gamma (IFN-γ)-dependent manner. Furthermore, the anti-TNBC efficacy studies reveal that camptothesome is superior to Onivyde and markedly potentiates PD-L1 immune checkpoint blockade therapy with complete lung metastasis remission in an orthotopic 4T1-Luc2 tumor model. This combination therapy eliciting robust cytotoxic T lymphocytes (CTL) response via boosting tumor-infiltrating cluster of differentiation 8 (CD8), calreticulin (CRT), high mobility group box 1 protein (HMGB-1), low-density lipoprotein receptor-related protein 1 (LRP1), IFN-γ, and granzyme B. Our work corroborates the promise of camptothesome in favorably modulating tumor immune microenvironment via inducing ICD to fortify the PD-L1 checkpoint blockade therapy for improved treatment of intractable TNBC.

Camptothesome is an innovative nanovesicle therapeutic comprising the sphingomyelin-derived camptothecin (CPT) lipid bilayer. In this work, we deciphered that Camptothesome was taken up by colorectal cancer (CRC) cells through primarily the clathrin-mediated endocytotic pathway and displayed the potential of eliciting robust immunogenic cancer cell death (ICD) via upregulating calreticulin, high mobility group box 1 protein (HMGB-1), and adenosine triphosphate (ATP), three hallmarks involved in the induction of ICD. In addition, use of dying MC38 tumor cells treated with Camptothesome as vaccine prevented tumor growth in 60% mice that received subsequent injection of live MC38 cells on the contralateral flank, validating Camptothesome was a legitimate ICD inducer in vivo. Camptothesome markedly reduced the acute bone marrow toxicity and gastrointestinal mucositis associated with free CPT and beat free CPT and Onivyde on anti-CRC efficacy and immune responses in a partially interferon gamma (IFN-γ)-dependent manner. Furthermore, Camptothesome enhanced the efficacy of immune checkpoint inhibitors to shrink late-stage orthotopic MC38 CRC tumors with diminished tumor metastasis and markedly prolonged mice survival.
Copyright © 2022 Elsevier B.V. All rights reserved.

Radiodynamic therapy (RDT) holds the potential to overcome the shallow tissue penetration issue associated with conventional photodynamic therapy (PDT). To this end, complex and sometimes toxic scintillator-photosensitizer nanoconjugates are often used, posing barriers for large-scale manufacturing and regulatory approval.
Herein, we report a streamlined RDT strategy based on CsI(Na)@MgO nanoparticles and 5-aminolevulinic acid (5-ALA). 5-ALA is a clinically approved photosensitizer, converted to protoporphyrin IX (PpIX) in cancer cells' mitochondria. CsI(Na)@MgO nanoparticles produce strong ~ 410 nm X-ray luminescence, which matches the Soret band of PpIX. We hypothesize that the CsI(Na)@MgO-and-5-ALA combination can mediate RDT wherein mitochondria-targeted PDT synergizes with DNA-targeted irradiation for efficient cancer cell killing. Because scintillator nanoparticles and photosensitizer are administered separately, the approach forgoes issues such as self-quenching or uncontrolled release of photosensitizers.
When tested in vitro with 4T1 cells, the CsI(Na)@MgO and 5-ALA combination elevated radiation-induced reactive oxygen species (ROS), enhancing damages to mitochondria, DNA, and lipids, eventually reducing cell proliferation and clonogenicity. When tested in vivo in 4T1 models, RDT with the CsI(Na)@MgO and 5-ALA combination significantly improved tumor suppression and animal survival relative to radiation therapy (RT) alone. After treatment, the scintillator nanoparticles, made of low-toxic alkali and halide elements, were efficiently excreted, causing no detectable harm to the hosts.
Our studies show that separately administering CsI(Na)@MgO nanoparticles and 5-ALA represents a safe and streamlined RDT approach with potential in clinical translation.
© 2022. The Author(s).

Restoring the control of food intake is the key to obesity management and prevention. The arcuate nucleus (ARC) of the hypothalamus is extensively being studied as a potential anti-obesity target. Animal studies showed that neuropeptide FF (NPFF) reduces food intake by its action in neuropeptide Y (NPY) neurons of the hypothalamic ARC, but the detailed mode of action observed in human neurons is missing, due to the lack of a human-neuron-based model for pharmacology testing. Here, we validated and utilized a human-neural-stem-cell-based (hNSC) model of ARC to test the effects of NPFF on cellular pathways and neuronal activity. We found that in the human neurons, decreased cAMP levels by NPFF resulted in a reduced rate of cytoplasmic calcium oscillations, indicating an inhibition of ARC NPY neurons. This suggests the therapeutic potential of NPFFR2 in obesity. In addition, we demonstrate the use of human-stem-cell-derived neurons in pharmacological applications and the potential of this model to address functional aspects of human hypothalamic neurons.