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Cancer is a major public health problem worldwide and is the second leading cause of death in the United States. About two-thirds of cancer patients are treated with radiation, either alone or in combination with immunotherapy, chemotherapy, or surgery. It is known that hypoxia or low partial oxygen pressure (pO₂) causes resistance to radiation therapy, immunotherapy, and chemotherapy.

Solid tumors treated with radiotherapy often experience decreased oxygen delivery.  This early tumor response to radiotherapy can be monitored by measuring the pO₂ in the tumor microenvironment.  These in vivo pO₂ measurements can be accomplished using the JIVA-25™ Electron Paramagnetic Resonance Oxygen Imaging (EPROI) instrument from O2M Technologies.  During EPROI studies, a gas anesthetic is often used to immobilize the mouse.  Unfortunately, a gas anesthetic that uses medical grade air (21% O₂) or 100% O₂ breathing gas can alter the measured pO₂ in the tumor, potentially complicating the interpretation of the imaging results.

Figure 1.  Oxygen Enhanced EPROI.  The workflow acquires anatomical MR images, two pO₂ maps with 21% O₂ breathing gas, and two pO₂ maps of 100% O₂ breathing gas.  The test-retest shows outstanding precision of 3.1 Torr with both breathing gases.

Prof. Marty Pagel and graduate student Tianzhe Li at the MD Anderson Cancer Center have exploited this potential pitfall to develop a new biomarker for assessing early tumor response to radiotherapy.  They have measured ΔpO₂, which is the difference in pO₂ measured with 21% O₂ and 100% O₂ breathing gases.  As an analogy, a car with a ¼-full gas tank has an available capacity of ¾-empty tank.  ΔpO₂ represents the available capacity of a tumor to take more O₂.  To demonstrate the value of this new biomarker, Li and Pagel have developed an Oxygen Enhanced (OE)-EPROI protocol that interperitoneally administers OX071 agent, acquires MR images for anatomical reference, and then obtains pO­₂ maps with 21% O2 and 100% O₂ breathing gases (Figure 1).  They have applied their protocol to study the early response of a Colo357 model treated with 10 Gy radiotherapy (Figure 2).

Figure 2.  Oxygen Enhanced (OE)-EPROI showing a significant decrease in ΔpO_2.

Their results have shown that radiotherapy caused a large and significant decrease in ΔpO₂ (Table 1). For comparison, radiotherapy caused a small, statistically insignificant decrease in tumor pO₂ with 21% O₂ breathing gas.  Similarly, radiotherapy caused a larger yet still statistically insignificant decrease in tumor pO₂ with 100% O₂ breathing gas.  Furthermore, ΔpO₂ showed the greatest effect size, known as the change in a biomarker relative to its variance, demonstrating that ΔpO₂ was most effective in evaluating an early tumor response to radiotherapy.  This OE-EPROI protocol can provide a new diagnostic method for evaluating early response to radiotherapy.

Table 1. OE-EPROI of the Colo357 model undergoing radiotherapy (n=9)

1. differences determined for each mouse, and not based on the averages of the groups.
2. p < 0.01 is considered to be statistically significant, from a two-tailed Student’s T-Test assuming equal variances
3. Effect size: |{(average post-pre) / (average standard deviation of post and pre)}|

Prof. Marty Pagel leads the Contrast Agent Molecular Engineering Laboratory (CAMEL) at the MD Anderson Cancer Center, Houston TX, USA.  CAMEL evaluates hypoxia, acidosis, enzyme activity and vascular perfusion in the tumor microenvironment, using a variety of molecular imaging methods.  More specifically, CAMEL uses EPROI to evaluate chemotherapy, immunotherapy, radiotherapy and radiosensitizers in small animal models of cancer and models of wound healing.

Learn more about EPR Oxygen Imaging

Magnetic resonance imaging (MRI) and electron paramagnetic resonance oxygen imaging (eMRI or EPROI) are fundamentally based upon the same principles, however a key difference is the resonant species. MRI aligns water protons, that are abundant in the body, in a magnetic field to produce exquisite anatomical images whereas eMRI relies on aligning electron spins. Unlike the ubiquity of protons, unpaired electrons sensitive to oxygen are rarely found in living organisms. A nontoxic contrast agents, with high specificity and sensitivity to detect molecular oxygen, is a necessity for EPR-based oxygen imaging; and now, it is being synthesized by O2M Technologies. Introducing incredible molecule: OX071!

Tetrathiatriarylmethyl or “trityl” radicals are not unfamiliar in MRI applications. They have been used as a polarizing agent. Previous versions of trityls, such as the largely used Finland trityl (FT), were unsuitable for in vivo experimentation. Its lipophilic core has a propensity to aggregate with plasma proteins, resulting in some amount of toxicity. In the next iteration, trityl OX063, 12 methyl groups in FT are replaced with hydroxyethyl groups, which greatly increased it’s hydrophilicity. The result is a completely non-toxic, injectable probe. OX063 is used a polarizing agent in 13C MRI.

The latest evolution of trityl is OX071, a partially deuterated analogue of OX063. Deuterium is EPR-silent, which eliminates a source of hyperfine coupling with trityl’s radical center. OX071 retains all the benefits of it’s predecessor while also showing a higher oxygen sensitivity, a higher signal-to-noise ratio, and under physiological conditions, a single-line EPR spectrum with ultra-narrow linewidth. OX071 is especially sensitive in low oxygen conditions. Even a small amount of OX071 will generate signal. The probe can be delivered intravenously, intraperitonially or intratumorally. OX071 is cleared from the animal in 10-30 minutes virtually unchanged.

All these properties make OX071 the probe of choice for a wide array of in vivo EPR oximetry experiments. OX071 has already been published in applications of solid tumor oximetry, viability studies in highly oxygen dependent cell populations like transplanted islets of Langerhans, bioscaffold device development, and much more. Reach out to O2M for OX063 or OX071 in your experiments!

Tissue engineered grafts and cell encapsulation devices have significant potential to improve human health. The site of transplantation is a factor that has profound effect on graft survival. The ideal chosen site should be easy to access for implantation and retrieval, while providing adequate diffusion of nutrients and oxygen for support of cell survival until vascularization is established. The two most commonly used sites for implantation of tissue grafts and cell encapsulation devices are subcutaneous (SC) and intraperitoneal (IP). Evaluating the native oxygenation in these sites is valuable information towards understanding the physiological milieu of implants in vivo.

O2M’s recent peer-reviewed publication in Tissue Engineering Part C: Methods presents the first ever solid probe oxygen imaging, reporting pO₂ values, of subcutaneous and intraperitoneal spaces in mice. We observe that solid probe oxygen imaging is a technology that offers researchers oxygenation data in vivo without needing further invasive measures or sacrifice. This study was funded by Juvenile Diabetes Research Foundation (JDRF) and will help scientists develop better therapies for type I diabetes as well as other health conditions.

Visit O2M’s Publication Page to learn about latest research in oxygen imaging.

O2M is at BioFAB and TERMIS!

Come see us in Manchester, New Hampshire!

“In Vitro and In Vivo Oxygen Imaging of Islet Encapsulation Devices: Lessons Learned and Path Forward”

Come See Us in Toronto!

Are you at TERMIS 2022? Make sure your plans include a visit to our booth in the Exhibition Hall. We hope to see you there!

Upcoming O2M Webinar

Moderator: Dr. Martyna Elas, Jagiellonian University, Krakow, Poland

Abstract: Previous studies with EPR imaging have shown that measurements of tumor pO₂ can indicate the status of hypoxia in the tumor microenvironment, which can be used to predict response to radiation therapy.  To build on these previous studies, we are developing Oxygen Enhanced (OE) EPR imaging that challenges a pre-clinical tumor model with 21% O₂ (medical grade air) and 100% O₂ in the anesthetic carrier gas, which can be a useful biomarker for evaluating early response to radiation therapy.  In addition, we are developing Dynamic Contrast Enhanced (DCE) EPR imaging that measures tumor vascular perfusion, as a complimentary biomarker for evaluating early response to cancer treatment.  This presentation will also discuss our ongoing research studies that show how tumor hypoxia causes resistance to immunotherapy, and how reducing hypoxia can improve tumor control with immune checkpoint blockade.

Biomaterials that host live cells must overcome the crucial hurdle of sustaining sufficient oxygenation for cell survival. Several approaches for improving oxygenation within the biomaterials have been developed and investigated for improved cell survival in artificial tissue constructs. Regardless of the method of addressing the oxygen needs of the cells, all three-dimensional constructs require in situ pO₂ assessment in vitro and in vivo to gauge the success of the approach. In addition, in vivo models can have very divergent results because of inherent variability between the animals, which demands a greater understanding for the breadth of variation in normal and diseased states.  O2M’s oxygen imaging technology can improve the outcome of cell and tissue engineering by providing real time in vitro and in vivo oxygen maps for improved therapy outcome.

O2M is at SFB!