tricolor-transgenic-murine-model-for-studying-growth-plate-injury

Tricolor Transgenic Murine Model for Studying Growth Plate Injury

The cartilage growth plates in the bones of children enable limb lengthening but are weak relative to the bone, making them prone to fracturing when bones ...

University of Connecticut Health Center

Intravenously (IV)-injected gold nanoparticles (AuNPs) powerfully enhance the efficacy of X-ray therapy of tumors including advanced gliomas. However, pharmacokinetic issues, such as slow tissue clearance and skin discoloration, may impede clinical translation. The direct infusion of AuNPs into the tumor might be an alternative mode of delivery.

Intravenously-injected gold nanoparticles (AuNPs) access intracerebral F98 rat gliomas better than AuNPs infused directly into the tumor site by convection enhanced delivery.

Standard clinical X-ray contrast agents are small iodine-containing molecules that are rapidly cleared by the kidneys and provide robust imaging for only a few seconds, thereby limiting more extensive vascular and tissue biodistribution imaging as well as optimal tumor uptake. They are also not generally useful for preclinical microCT imaging where longer scan times are required for high resolution image acquisition. We here describe a new iodine nanoparticle contrast agent that has a unique combination of properties: 20 nm hydrodynamic diameter, covalent PEG coating, 40 hour blood half-life, 50% liver clearance after six months, accumulation in tumors, and well-tolerated to at least 4 g iodine/kg body weight after intravenous administration in mice. These characteristics are unique among the other iodine nanoparticles that have been previously reported and provide extended-time high contrast vascular imaging and tumor loading. As such, it is useful for preclinical MicroCT animal studies. Potential human applications might include X-ray radiation dose enhancement for cancer therapy and vascular imaging for life-threatening situations where high levels of contrast are needed for extended periods of time.

Small, Long Blood Half-Life Iodine Nanoparticle for Vascular and Tumor Imaging.

Gliomas and other brain tumors have evaded durable therapies, ultimately causing about 20% of all cancer deaths. Tumors are widespread in the brain at time of diagnosis, limiting surgery and radiotherapy effectiveness. Drugs are also poorly effective. Radiotherapy (RT) is limited by dose to normal tissue. However, high-atomic-number elements absorb X-rays and deposit the absorbed dose locally, even doubling (or more) the local dose. Previously we showed that gold nanoparticles (AuNPs) with RT could eradicate some brain tumors in mice and many other preclinical studies confirmed AuNPs as outstanding radioenhancers. However, impediments to clinical translation of AuNPs have been poor clearance, skin discoloration, and cost. We therefore developed iodine nanoparticles (INPs) that are almost colorless, non-toxic, lower cost, and have reasonable clearance, thus overcoming major drawbacks of AuNPs. Here we report the use of iodine nanoparticle radiotherapy (INRT) in treating advanced human gliomas (U87) grown orthotopically in nude mice resulting in a more than a doubling of median life extension compared to RT alone. Significantly, INRT also enhanced the efficacy of chemotherapy when it was combined with the chemotherapeutic agent Doxil, resulting in some longer-term survivors. While ongoing optimization studies should further improve INRT, clinical translation appears promising.

Iodine nanoparticles enhance radiotherapy of intracerebral human glioma in mice and increase efficacy of chemotherapy.

About 30% of breast cancers metastasize to the brain; those widely disseminated are fatal typically in 3-4 months, even with the best available treatments, including surgery, drugs, and radiotherapy. To address this dire situation, we have developed iodine nanoparticles (INPs) that target brain tumors after intravenous (IV) injection. The iodine then absorbs X-rays during radiotherapy (RT), creating free radicals and local tumor damage, effectively boosting the local RT dose at the tumor. Efficacy was tested using the very aggressive human triple negative breast cancer (TNBC, MDA-MB-231 cells) growing in the brains of athymic nude mice. With a well-tolerated non-toxic IV dose of the INPs (7 g iodine/kg body weight), tumors showed a heavily iodinated rim surrounding the tumor having an average uptake of 2.9% iodine by weight, with uptake peaks at 4.5%. This is calculated to provide a dose enhancement factor of approximately 5.5 (peaks at 8.0), the highest ever reported for any radiation-enhancing agents. With RT alone (15 Gy, single dose), all animals died by 72 days; INP pretreatment resulted in longer-term remissions with 40% of mice surviving 150 days and 30% surviving > 280 days.

Iodine nanoparticle radiotherapy of human breast cancer growing in the brains of athymic mice.

 To analyze the localization, distribution and effect of iodine nanoparticles (INPs) on radiation therapy (RT) in advanced intracerebral gliomas over time after intravenous injection.  Luciferase/td-tomato expressing U87 human glioma cells were implanted into mice which were injected intravenously with INPs. Mice with gliomas were followed for tumor progression and survival. Immune-stained mouse brain sections were examined and quantified by confocal fluorescence microscopy.  INPs injected intravenously 3 days prior to RT, compared with 1 day, showed greater association with CD31-staining structures, accumulated inside tumor cells more, covered more of the tumor cell surface and trended toward increased median survival.  INP persistence and redistribution in tumors over time may enable greater RT enhancement and clinically relevant hypo-fractionated-RT and may enhance INP efficacy.

Distributions of intravenous injected iodine nanoparticles in orthotopic u87 human glioma xenografts over time and tumor therapy.

Triple negative breast cancer (TNBC), ~ 10-20% of diagnosed breast cancers, metastasizes to brain, lungs, liver. Iodine nanoparticle (INP) radioenhancers specifically localize to human TNBC MDA-MB-231 tumors growing in mouse brains after iv injection, significantly extending survival of mice after radiation therapy (RT). A prominent rim of INP contrast (MicroCT) previously seen in subcutaneous tumors but not intracerebral gliomas, provide calculated X-ray dose-enhancements up to > eightfold. Here, MDA-MB-231-cells, INPs, CD31 were examined by fluorescence confocal microscopy. Most INP staining co-localized with CD31 in the tumor center and periphery. Greatest INP/CD31 staining was in the tumor periphery, the region of increased MicroCT contrast. Tumor cells are seen to line irregularly-shaped spaces (ISS) with INP, CD31 staining very close to or on the tumor cell surface and PAS stain on their boundary and may represent a unique form of CD31-expressing vascular mimicry in intracerebral 231-tumors. INP/CD31 co-staining is also seen around ISS formed around tumor cells migrating on CD31 blood-vessels. The significant radiation dose enhancement to the prolific collagen I containing, INP-binding ISS found throughout the tumor but concentrated in the tumor rim, may contribute significantly to the life extensions observed after INP-RT; VM could represent a new drug/NP, particularly INP, tumor-homing target.

Novel Iodine nanoparticles target vascular mimicry in intracerebral triple negative human MDA-MB-231 breast tumors.

Cell-cell communications are central to a variety of physiological and pathological processes in multicellular organisms. Cells often rely on cellular protrusions to communicate with one another, which enable highly selective and efficient signaling within complex tissues. Owing to significant improvements in imaging techniques, identification of signaling protrusions has increased in recent years. These protrusions are structurally specialized for signaling and facilitate interactions between cells. Therefore, physical regulation of these structures must be key for the appropriate strength and pattern of signaling outcomes. However, the typical approaches for understanding signaling regulation tend to focus solely on changes in signaling molecules, such as gene expression, protein-protein interaction, and degradation. In this short review, we summarize the studies proposing the removal of different types of signaling protrusions-including cilia, neurites, MT (microtubule based)-nanotubes and microvilli-and discuss their mechanisms and significance in signaling regulation.

Removal of cellular protrusions.

Effective and durable treatment of glioblastoma is an urgent unmet medical need. In this article, we summarize a novel approach of a physical method that enhances the effectiveness of radiotherapy. High atomic number nanoparticles that target brain tumors are intravenously administered. Upon irradiation, the nanoparticles absorb X-rays creating free radicals, increasing the tumor dose several fold. Radiotherapy of mice with orthotopic human gliomas and human triple negative breast cancers growing in the brain showed significant life extensions when the nanoparticles were included. An extensive study of the properties of the iodine-containing nanoparticle (Niodx) by the Nanotechnology Characterization Laboratory, including sterility, physicochemical characterization, in vitro cytotoxicity, in vivo immunological characterization, and in vivo toxicology, is presented. In summary, the iodine nanoparticle Niodx appears safe and effective for translational studies toward human use.

Iodine Nanoparticles (Niodx) for Radiotherapy Enhancement of Glioblastoma and Other Cancers: An NCI Nanotechnology Characterization Laboratory Study.

Radiation therapy (RT) treats approximately half of all cancers and most brain cancers. RT is variably effective at inducing a dormant tumor state i.e. the time between RT and clinical recurrence of tumor growth. Interventions that significantly lengthen tumor dormancy would improve long-term outcomes. Inflammation can promote the escape of experimental tumors from metastatic dormancy in the lung. Previously we showed intracerebral B16F10 melanoma dormancy varied with RT dose; 20.5 Gy induced dormancy lasted ~ 2 to 4 weeks-sufficient time to study escape from dormancy. Tumors were followed over time using bioluminescence. Surprisingly, some tumors in endotoxin-treated mice exited from dormancy slower; a large fraction of the mice survived more than 1-year. A cohort of mice also experienced an accelerated exit from dormancy and increased mortality indicating there might be variation within the tumor or inflammatory microenvironment that leads to both an early deleterious effect and a longer-term protective effect of inflammation. Some of the melanin containing cells at the site of the original tumor were positive for senescent markers p16, p21 and βGal. Changes in some cytokine/chemokine levels in blood were also detected. Follow-up studies are needed to identify cytokines/chemokines or other mechanisms that promote long-term dormancy after RT.

Radiation-induced dormancy of intracerebral melanoma: endotoxin inflammation leads to both shortened tumor dormancy and long-term survival with localized senescence.

Drosophila male germline stem cells (GSCs) reside at the tip of the testis and surround a cluster of niche cells. Decapentaplegic (Dpp) is one of the well-established ligands and has a major role in maintaining stem cells located in close proximity. However, the existence and the role of the diffusible fraction of Dpp outside of the niche have been unclear. Here, using genetically-encoded nanobodies called Morphotraps, we physically block Dpp diffusion without interfering with niche-stem cell signaling and find that a diffusible fraction of Dpp is required to ensure differentiation of GSC daughter cells, opposite of its role in maintenance of GSC in the niche. Our work provides an example in which a soluble niche ligand induces opposed cellular responses in stem cells versus in differentiating descendants to ensure spatial control of the niche. This may be a common mechanism to regulate tissue homeostasis.

Sharif M. Ridwan

University of Connecticut Health Center

Tricolor Transgenic Murine Model for Studying Growth Plate Injury