Purpose Progress toward developing a novel radiocontrast agent for determining pO(2) in tumors in a clinical setting is described. The imaging agent is designed for use with electron paramagnetic resonance imaging (EPRI), in which the collision of a paramagnetic probe molecule with molecular oxygen causes a spectroscopic change which can be calibrated to give the real oxygen concentration in the tumor tissue.Procedures The imaging agent is based on a nanoscaffold of aluminum hydroxide (boehmite) with sizes from 100 to 200 nm, paramagnetic probe molecule, and encapsulation with a gas permeable, thin (10-20 nm) polymer layer to separate the imaging agent and body environment while still allowing O-2 to interact with the paramagnetic probe. A specially designed deuterated Finland trityl (dFT) is covalently attached on the surface of the nanoparticle through 1,3-dipolar addition of the alkyne on the dFT with an azide on the surface of the nanoscaffold. This click-chemistry reaction affords 100% efficiency of the trityl attachment as followed by the complete disappearance of the azide peak in the infrared spectrum. The fully encapsulated, dFT-functionalized nanoparticle is referred to as RADI-Sense.Results Side-by-side in vivo imaging comparisons made in a mouse model made between RADI-Sense and free paramagnetic probe (OX-071) showed oxygen sensitivity is retained and RADI-Sense can create 3D pO(2) maps of solid tumorsConclusions A novel encapsulated nanoparticle EPR imaging agent has been described which could be used in the future to bring EPR imaging for guidance of radiotherapy into clinical reality.

<p>PDF file, 40K, Dose given to MCa4 tumors vs HF10 before the radiation. MCa4 tumors of approximately the same size were treated with a range of 60-80 Gy, which is a narrow range around TCD50 value of 69 Gy consistent with the literature (Stone, 1975).</p>

[This corrects the article DOI: 10.3389/fmed.2023.1269689.].

To identify the optimal threshold in 18F-fluoromisonidazole (FMISO) PET images to accurately locate tumor hypoxia by using electron paramagnetic resonance imaging (pO2 EPRI) as ground truth for hypoxia, defined by pO2 $$\le$$ 10 mmHg. Tumor hypoxia images in mouse models of SCCVII squamous cell carcinoma (n = 16) were acquired in a hybrid PET/EPRI imaging system 2 h post-injection of FMISO. T2-weighted MRI was used to delineate tumor and muscle tissue. Dynamic contrast enhanced (DCE) MRI parametric images of Ktrans and ve were generated to model tumor vascular properties. Images from PET/EPR/MRI were co-registered and resampled to isotropic 0.5 mm voxel resolution for analysis. PET images were converted to standardized uptake value (SUV) and tumor-to-muscle ratio (TMR) units. FMISO uptake thresholds were evaluated using receiver operating characteristic (ROC) curve analysis to find the optimal FMISO threshold and unit with maximum overall hypoxia similarity (OHS) with pO2 EPRI, where OHS = 1 shows perfect overlap and OHS = 0 shows no overlap. The means of dice similarity coefficient, normalized Hausdorff distance, and accuracy were used to define the OHS. Monotonic relationships between EPRI/PET/DCE-MRI were evaluated with the Spearman correlation coefficient ( $$\rho$$ ) to quantify association of vasculature on hypoxia imaged with both FMISO PET and pO2 EPRI. FMISO PET thresholds to define hypoxia with maximum OHS (both OHS = 0.728 $$\pm$$ 0.2) were SUV $$\ge$$ 1.4 $$\times$$ SUVmean and SUV $$\ge$$ 0.6 $$\times$$ SUVmax. Weak-to-moderate correlations (| $$\rho$$ |< 0.70) were observed between PET/EPRI hypoxia images with vascular permeability (Ktrans) or fractional extracellular-extravascular space (ve) from DCE-MRI. This is the first in vivo comparison of FMISO uptake with pO2 EPRI to identify the optimal FMISO threshold to define tumor hypoxia, which may successfully direct hypoxic tumor boosts in patients, thereby enhancing tumor control.

Purpose To enhance the spatial accuracy of fluorine 18 (18F) misonidazole (MISO) PET imaging of hypoxia by using dynamic contrast-enhanced (DCE) MR images as a basis for modifying PET images and by using electron paramagnetic resonance (EPR) partial oxygen pressure (pO2) as the reference standard. Materials and Methods Mice (n = 10) with leg-borne MCa4 mammary carcinomas underwent EPR imaging, T2-weighted and DCE MRI, and 18F-MISO PET/CT. Images were registered to the same space for analysis. The thresholds of hypoxia for PET and EPR images were tumor-to-muscle ratios greater than or equal to 2.2 mm Hg and less than or equal to 14 mm Hg, respectively. The Dice similarity coefficient (DSC) and Hausdorff distance (d H ) were used to quantify the three-dimensional overlap of hypoxia between pO2 EPR and 18F-MISO PET images. A training subset (n = 6) was used to calculate optimal DCE MRI weighting coefficients to relate EPR to the PET signal; the group average weights were then applied to all tumors (from six training mice and four test mice). The DSC and d H were calculated before and after DCE MRI-corrected PET images were obtained to quantify the improvement in overlap with EPR pO2 images for measuring tumor hypoxia. Results The means and standard deviations of the DSC and d H between hypoxic regions in original PET and EPR images were 0.35 mm ± 0.23 and 5.70 mm ± 1.7, respectively, for images of all 10 mice. After implementing a preliminary DCE MRI correction to PET data, the DSC increased to 0.86 mm ± 0.18 and the d H decreased to 2.29 mm ± 0.70, showing significant improvement (P < .001) for images of all 10 mice. Specifically, for images of the four independent test mice, the DSC improved with correction from 0.19 ± 0.28 to 0.80 ± 0.29 (P = .02), and the d H improved from 6.40 mm ± 2.5 to 1.95 mm ± 0.63 (P = .01). Conclusion Using EPR information as a reference standard, DCE MRI information can be used to correct 18F-MISO PET information to more accurately reflect areas of hypoxia.Keywords: Animal Studies, Molecular Imaging, Molecular Imaging-Cancer, PET/CT, MR-Dynamic Contrast Enhanced, MR-Imaging, PET/MR, Breast, Oncology, Tumor Mircoenvironment, Electron Paramagnetic ResonanceSupplemental material is available for this article.© RSNA, 2021.

Precise quantitative delineation of tumor hypoxia is essential in radiation therapy treatment planning to improve the treatment efficacy by targeting hypoxic sub-volumes. We developed a combined imaging system of positron emission tomography (PET) and electron para-magnetic resonance imaging (EPRI) of molecular oxygen to investigate the accuracy of PET imaging in assessing tumor hypoxia. The PET/EPRI combined imaging system aims to use EPRI to precisely measure the oxygen partial pressure in tissues. This will evaluate the validity of PET hypoxic tumor imaging by (near) simultaneously acquired EPRI as ground truth. The combined imaging system was constructed by integrating a small animal PET scanner (inner ring diameter 62 mm and axial field of view 25.6 mm) and an EPRI subsystem (field strength 25 mT and resonant frequency 700 MHz). The compatibility between the PET and EPRI subsystems were tested with both phantom and animal imaging. Hypoxic imaging on a tumor mouse model using $^{18}$F-fluoromisonidazole radio-tracer was conducted with the developed PET/EPRI system. We report the development and initial imaging results obtained from the PET/EPRI combined imaging system.

12 Objectives: Tumor hypoxia is associated with resistance to therapy and tumor progression, and correlates negatively with patient survival [1, 2, 3]. 18F-Misonidazole (FMISO) is frequently used in clinical PET trials to measure and treat tumor hypoxia [4], but there is no universally accepted threshold to define tumor hypoxia with FMISO. This study uses electron paramagnetic resonance (EPR) pO2 images as true hypoxia (pO2 < 10 mmHg) [5] to calculate the optimal corresponding FMISO PET threshold for identifying hypoxic tumors in SCC7 tumor murine models of squamous cell carcinoma. Methods: Imaging: Using SCC7 squamous cell carcinoma murine models (n=14), the tumor-bearing leg was immobilized in the plastic bed in a polysiloxane dental mold cast (GC America, Alsip, IL) with embedded fiducials to allow for co-registration between modalities. FMISO PET and EPR images were acquired in a hybrid PET/EPR system for simultaneous imaging [6], which gave the advantage of identical physiological conditions of the mouse. A tail-vein cannula was used to administer an oxygen-sensitive spin probe solution for EPR imaging. A bolus injection of ~230 uCi of FMISO (produced at the in-house cyclotron facility) was used for PET imaging; images were acquired 2-hours post-injection. T2-weighted images were acquired in a 9.4 Tesla small animal imager (Bruker, Erlangen, Germany) for registration and tumor/muscle contouring. Image analysis: Following MRI/EPR/PET registration in MATLAB, images were resampled to the PET image’s isotropic voxel resolution of [0.5 mm]3. The T2 MRI-based tumor and muscle contour were transformed to the PET and EPR images in units of tumor-to-muscle ratio (TMR) and pO2, respectively. Using a custom-written script in MATLAB, ROC curves were generated for each tumor across all thresholds of PET TMR > 0 to 5.6 in increments of 0.2, using EPR pO2 < 10 mmHg as true hypoxia. The accuracy (ACC) (fraction of true negatives and positives over all true/false negatives/positives), Dice Similarity Coefficient (DSC), and Hausdorff Distance (dH) were used to quantify overlap between hypoxic regions as defined by EPR and PET. Because maximum ACC and DSC are both between 0 and 1, with 1 corresponding to highest overlap, dH was normalized and subtracted from 1 (1 - || dH ||) so that the highest value would also show maximum overlap. The peak mean of ACC, DSC, and 1 - ||dH|| averaged over all tumors was used to determine the optimal PET threshold. The hypoxic fractions of tumor voxels based on resulting thresholds was also calculated to compare between modalities. Results: For all tumors, the area under the ROC curve using pO2 2.4, and peak DSC = 0.485 (SE = 0.05) corresponding to a threshold TMR 2.0. At its minimum, mean dH= 3.40 (SE = 0.2) mm at TMR > 2.4. The average value of ACC, DSC, and 1-|| dH || showed a peak at TMR > 2.2 and pO2 < 10 mmHg. The mean hypoxic fraction of EPR images was 0.20 (SE = 0.05), and of PET images was 0.19 (SE = 0.03), which was not significant based on the two-sample t-test (p = 0.51). Conclusions: Based on this dataset of SCC7 squamous cell carcinoma murine models, the PET threshold of TMR > 2.2 has the highest ACC and DSC, and the lowest dH, when compared to hypoxic tumor regions defined by EPR pO2 < 10 mmHg. These results might help improve patient prognosis for more accurate hypoxia-based dose-painting treatment plans based on PET imaging. References: [1] Hockel, M et al. (1996). Cancer Res, 56(19): p. 4509-15.[2] Hockel, M and Vaupel, P (2001). J Natl Cancer Inst, 93(4): p. 266-76.[3] Brizel, DM et al. (1997). Int J Radiat Oncol Biol Phys, 38(2): p. 285-9.[4] Lopci, E et al. (2014). Am J Nucl Med Mol Imaging, 4: p. 365-384.[5] Epel, B et al. (2019). Int J Radiat Oncol Biol Phys, 103(4): p. 977-984.[6] Kim H et al. (2020). Nuclear Inst. and Methods in Physics Research Section, A, 959.

Abstract Introduction: In tumor mouse models, electron paramagnetic resonance (EPR) imaging has been used to quantify pO2 in-vivo for oxygen-guided radiation therapy. Irradiating the hypoxic tumor sub-regions with a boost of radiation significantly increased survival probability when compared to the boosted radiation of normoxic tumor sub-regions. These results indicate 1) the importance of accurate oxygen imaging in-vivo and 2) an advantage to dose-painting hypoxic tumor sub-regions to improve radiation therapy outcomes of hypoxic tumor treatment. The purpose of this study is to evaluate the accuracy of hypoxia imaging using 18F-misonidazole (FMISO) with PET imaging while using EPR imaging as the reference standard for true hypoxia. We include T2-weighted MRI to define tumor anatomy and dynamic contrast enhanced (DCE)-MRI imaging to model vasculature properties to make FMISO PET more accurate in its measurement of hypoxia. Methods: We used six SCC7 tumor mouse models in C3H mice, grown in the leg in the range of 200-500 mm3. Under minimal anesthesia, each mouse leg was set in a soft vinylpolysiloxane cast with embedded fiducials for registered images. A custom-built PET insert was inserted into a 720 MHz EPR imager for near-simultaneous imaging with PET and EPR, acquired two hours post-injection of FMISO to allow the radiotracer to bind to hypoxic tumor cells. Then, T2-weighted and gadodiamide DCE-MRI images were acquired and Ktrans and ve parameters were fitted to model tumor vasculature properties using the Tofts model. Data from all modalities were registered using fiducials, and resampled to isotropic (0.5 mm)3 voxels. The thresholds for hypoxia were defined as tumor to muscle ratio (TMR) ≥ 2 and pO2 ≤ 10 torr for PET and EPR, respectively. To correct the PET image using DCE-MRI parameters, we first modeled FMISO retention as a logistic function of pO2 to map the EPR image to PET TMR image, so that its sigmoidal point of inflection was at the threshold of retention. Then we estimated optimal weighting coefficients of DCE-MRI parameters to add or subtract voxel by voxel to the PET data so that its definition of hypoxia is more similar to the EPR image's hypoxia definition. The quality of overlap between hypoxic tumor regions as defined by PET and EPR was assessed using the Dice Similarity Coefficient (DSC) and the Hausdorff Distance (HD), before and after applying a correction to the PET data. Results: The DSC between hypoxic regions as defined by PET and EPR, before and after applying a correction to PET data, was 0.53 ± 0.2 and 0.80 ± 0.2, respectively. The HD was respectively 3.8 ± 0.5 and 2.1 ± 0.5 mm. Conclusion: These results indicate that there is poor agreement between FMISO PET hypoxia measurements when compared to hypoxia as defined by EPR pO2 images. Applying our correction method significantly improved the overlap between PET and EPR hypoxic tumor regions. Citation Format: Inna Gertsenshteyn, Boris Epel, Lara Leoni, Xiaobing Fan, Richard Friefelder, Eugene Barth, Heejong Kim, Marta Zamora, Erica Markiewicz, Darwin Bodero, Mohammed Bhuiyan, Anna Kucharski, Hsiu-Ming Tsai, Mellissa Grana, Subramanian V. Sundramoorthy, Gregory S. Karczmar, Chien-Min Kao, Chin-Tu Chen, Howard Halpern. Multimodal imaging of tumor hypoxia with 18F-misonidazole PET, EPR, and MRI [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1648.

Abstract Introduction: It has been known for over a century that all living systems develop radiation resistance under hypoxic conditions - low pO2. But giving extra radiation - dose painting - or boosting hypoxic tumors regions has not shown enhanced control in mammalian tumors. Pulse EPR quantitative pO2 values can be tomographically imaged as a magnetic resonance measurement with fixed stepped gradients. If there are distinct hypoxic regions, their images can then be extra radiation dose boosts. An isocentric animal irradiation system allowed conformal boosts to tiny resistant tumor subvolumes with 3D printed tungsten loaded plastic blocks. A randomized trial compared radiation boosts to hypoxic tumor vs well oxygenated tumor to determine hypoxic boost control advantage. Methods: Carcinomas (MCa4) and fibrosarcomas (FSa) in mouse legs with mean volume of 350 μl were treated with an XRAD225Cx animal irradiator to a dose that controlled 15% of tumors (TCD15), determined in separate experiments. Pulse electron paramagnetic resonance (EPR) using Spin Lattice Relaxation (SLR) and a soluble trityl spin probe has been shown to quantitatively image solution and tumor pO2. This and boost treatments were given after EPR pO2 imaging, reconstruction, registration with tumor defining T2 MRI, determination of all tumor hypoxic regions (pO2 ≤10 torr) and 3D printed tungsten loaded plastic aperture fabrication to radiate either well oxygenated or hypoxic tumor regions. Mice were followed for either 180 days (MCa4) or 90 days (FSa) sufficient for 99% of local failures to occur. Treatment of 3D hypoxic volumes was also analyzed for control dependence on surface to volume ratio (SVR). Results: Tumor control was doubled by radiating hypoxic tumor relative to well oxygenated tumor in 48 MCa4 tumors (p=0.013) and 54 FSa tumors (p=0.04). This was the first demonstration in mammalian tumors of an advantage to targeting hypoxic regions in tumors. It provides the first evidence of image based dose painting within the volume of the tumor. For MCa4 tumors, SVR values below median had significant increased survival (p=0.003) while for FSa tumors below median SVR tumors trended to increased survival (p=0.3). Clinical Relevance: Modern radiation delivery used intensity modulated radiation therapy (IMRT) for dose gradients outside the tumor volume to reduce doses to critical structures. Dose within the tumor have strict inhomogeneity limits. Allowance of higher dose variation within the tumor volume allows “hot spots” to resistant tumor volumes, sharper gradients outside the volume to protect critical structures. This will enhance the therapeutic ratio. Citation Format: Howard Halpern, Matthew Maggio, Eugene Barth, Darwin Bodero, Richard C. Miller, Charles A. Pelizzari, Martyna Krzykawska-Serda, Subramanian V. Sundramoorthy, Bulent Aydogan, Ralph R. Weichselbaum, Victor M. Tormyshev, Inna Gertsenshteyn, Boris Epel. Increased tumor control boosting hypoxic regions identified with EPR pO2 imaging in 2 tumor types [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2773.

Previously we reported the development of a positron emission tomography (PET) and electron paramagnetic resonance (EPR) combined imaging system. The combined imaging system aimed to investigate the potential of PET hypoxia imaging by using EPR oxygen imaging as a reference. Simultaneous PET/EPR data acquisition is important to make sure of recording the same biological changes in both imaging modalities as well as to shorten imaging time. Our current system does not have magnetic or RF shielding. During the initial simultaneous PET/EPR operation, we observed that the EPR RF pulsing induced spurious noise events in the PET, and significantly affected the PET detector performance. We developed a method to reject the EPR RF noise events in off-line data processing. The method is to exploit the waveform sampling capable PET data acquisition implemented in a multi-voltage-threshold (MVT) board, and to use the pulse shape difference found in the MVT waveforms between scintillation and RF noise events. Experiments were conducted to evaluate the effectiveness of the proposed method in rejecting RF noise events. Preliminary results indicate that the rejection method works effectively to enable simultaneous data acquisition of the PET/EPR system.

Purpose: Tumor oxygenation is a critical parameter influencing the efficacy of cancer therapy. Low levels of oxygen in solid tumor have been recognized as an indicator of malignant progression and metastasis, as well as poor response to chemo- and radiation therapy. Being able to measure oxygenation for an individual's tumor would provide doctors with a valuable way of identifying optimal treatments for patients. Methods: Electron paramagnetic resonance imaging (EPRI) in combination with an oxygen-measuring paramagnetic probe was performed to measure tumor oxygenation in vivo. Triarylmethyl (trityl) radical exhibits high specificity, sensitivity, and resolution for quantitative measurement of O2 concentration. However, its in vivo applications in previous studies have been limited by the required high dosage, its short half-life, and poor intracellular permeability. To address these limitations, we developed high-capacity nanoformulated radicals that employed fluorescein isothiocyanate-labeled mesoporous silica nanoparticles (FMSNs) as trityl radical carriers. The high surface area nanostructure and easy surface modification of physiochemical properties of FMSNs enable efficient targeted delivery of highly concentrated, nonself-quenched trityl radicals, protected from environmental degradation and dilution. Results: We successfully designed and synthesized a tumor-targeted nanoplatform as a carrier for trityl. In addition, the nanoformulated trityl does not affect oxygen-sensing capacity by a self-relaxation or broadening effect. The FMSN-trityl exhibited high sensitivity/response to oxygen in the partial oxygen pressure range from 0 to 155 mmHg. Furthermore, MSN-trityl displayed outstanding intracellular oxygen mapping in both in vitro and in vivo animal studies. Conclusion: The highly sensitive nanoformulated trityl spin probe can profile intracellular oxygen distributions of tumor in a real-time and quantitative manner using in vivo EPRI.

347 Objectives: Hypoxic cells are more resistant than normoxic cells to radiation treatment, so the identification of hypoxic regions within the tumor is necessary to predict or improve radiotherapeutic outcome. Preclinical studies in image-guided radiotherapy with electron paramagnetic resonance (EPR) pO2 images have shown that targeting hypoxic regions of tumors, defined as < 10 torr, significantly suppresses tumor regrowth. Though EPR is considered to be the gold standard for measuring pO2, this imaging modality is not readily available for clinical practice; both positron emission tomography (PET) and magnetic resonance imaging (MRI) are more readily available and used in the clinic routinely. In addition to EPR, 18F-fluoro-misonidazole (FMISO) PET has been used to target hypoxia in tumors. The purpose of this study is to measure hypoxia in a mouse tumor using EPR and PET to determine the efficacy of the FMISO tracer in accurately identifying hypoxia, and to consider applying a modification to the PET data using dynamic contrast enhanced (DCE)-MRI to better predict true hypoxia as identified by EPR. Methods: MCa4 tumors were grown in the leg of mice. Once the tumor grew in the range of 250-400 mm3, the mouse was anesthetized, and its leg was set in a custom-made bed and cast with soft vinylpolysiloxane. Under minimal anesthesia for a total of five hours, the mouse was imaged in EPR, followed by a T2-weighted and DCE MRI, a two-hour dynamic PET scan, and finally a CT scan. Data from all modalities were registered using fiducials and anatomic landmarks. The tumor to muscle ratio (TMR) above 1.4 defined hypoxia in the PET image in the last frame, and pO2 values below 10 torr defined hypoxia in the EPR image. Based on the Tofts model, ktransand vemaps were obtained from DCE-MRI, as well as the relative signal increase (RSI) of contrast from time of injection. The Dice similarity coefficient was used to evaluate the overlap between the PET and EPR data. In addition, linear regression, ridge regression, least absolute shrinkage and selection (LASSO) regression, and generalized additive models were used to predict EPR at each voxel in terms of PET, ktrans, ve, and RSI. Results: Axial slices of the mouse shown in Fig. 1 qualitatively show that the region of uptake in FMISO PET more closely resembles increased activity in DCE-MRI measurements of ktransand RSI, rather than the pO2 data from EPR, which we consider to be the gold standard of measuring hypoxia. Among the mice analyzed, the highest Dice coefficient between hypoxic regions in PET and EPR images was 0.23 with hypoxic fractions (HF) of 0.083 and 0.51, respectively. This follows the general trend of a higher HF in FMISO PET images than EPR images based on the set thresholds, with the exception of one mouse. Another mouse showed an HF of 0 in the EPR image, and an HF of 0.44 in the PET image, which suggests that FMISO does not target only hypoxia. The adjusted R2 of the regression models ranged from 0.20 to 0.24. Conclusions: Based on this preliminary dataset and low Dice coefficient values between PET and EPR images, we conclude that FMISO does not solely target hypoxia, and if it is to be used as a method of image-guided radiotherapy, it must be corrected by some other relevant physiological parameters, including but not limited to DCE-MRI imaging. Future analysis of this data includes adjusting hypoxia thresholds in PET data by tumor to lung ratio and by SUVmax, and developing spatial regression models for EPR in terms of PET, ktrans, ve, and RSI with better prediction properties.

We present a combined imaging system of positron emission tomography (PET) and electron paramagnetic resonance imaging (EPRI) developed for assessing tumor hypoxia. Tumor hypoxia has been known to be resistant for cancer treatment such as radiation therapy. Therefore, precise quantitative measurement of tumor hypoxia is important for effective cancer treatment planning in radiation therapy. In clinic, PET imaging by using 18 F-fluoromisonidazole (F-MISO) have been performed to delineate tumor hypoxia. However, the effectiveness of PET imaging based hypoxia targeting in radiation therapy was not yet firmly established. EPRI is a non-invasive imaging method capable of measuring the partial pressure of oxygen (pO 2 ) in tissue precisely. However, EPRI is not available for clinical use currently. The combined imaging system is to use EPRI as a gold standard to measure the pO 2 in hypoxic tumor in animal, and aims to evaluate the validity of PET imaging in tumor hypoxia by simultaneously acquired EPRI. The PET component of the combined system, which was originally developed as an insert for MRI, consists of total 14 detector modules. Each PET detector module uses of 8×4 LYSO arrays (3×3×10 mm 3 ) coupled to Hamamatsu S13361 MPPC arrays (3.2 mm pitch). The detector modules are encased in a cylindrical plastic holder with 60 mm inner diameter and 115 mm outer diameter in a ring configuration. The axial field of view of the PET is 25.6 mm. The EPRI component of the system is built with two permanent magnets (25 mT), three gradient field coils for spatial encoding (the maximum 15 mT/m) and a RF resonator. The PET is installed to fit into the two EPR magnets (12 cm distance apart) so that the PET detector center is aligned to the magnet center. The compatibility of two imaging modalities were tested by using a 22 Na point source and imaging phantoms. Animal imaging with F-MISO and FDG were also performed by running the PET/EPR simultaneously. Initial imaging results are reported.

Abstract Living tissue hypoxic resistance to radiation, has been known for over a century. Our research has validated the hypothesis that radiation dose boosts focused specifically on hypoxic regions of tumors would improve tumor curability in a fibrosarcoma mouse model. We used Electron Paramagnetic Resonance (EPR) imaging of absolute pO2 in volume elements of murine tumors with 1 torr pO2 resolution and 0.7 mm spatial resolution in FSa fibrosarcomas in the legs of C3H mice. Hypoxia was defined as subvolumes or voxels with pO2 less than 10 torr. This showed EPR pO2 images as a reliable identifier and locator of relevant radiobiologically hypoxia. For the first time in mammalian tumors pO2 based dose painting was shown to improves tumor cure. This required gantry based x-ray treatment with an XRAD225Cx system to deliver dose to mouse tumors accurately registered with EPR pO2 images. Tumors were pretreated with a dose of radiation sufficient to cure 15% of tumors, a TCD15. For hypoxic boosts or hypoxia avoidance of the same volume, 3D printing Tungsten loaded, highly conformal plastic blocks were used. This allowed comparison of treating 100% of hypoxic tumor voxels with hypoxia avoidance boosts. This showed significant (p=0.02) tumor control differences between hypoxic boosts and hypoxia avoiding boosts. This is the first validation of the curative effectiveness of focusing hypoxia based dose painting in mammalian tumors. This was motivated by a conclusive failure to demonstrate a difference between treating ~ 85% of hypoxic voxels with a hypoxic boost compared with radiation avoiding hypoxia which proved tenets of radiation biology: a few surviving hypoxic clonogens cause radiation treatment failure. To further validate this approach, we have applied the same techniques to a second mouse mammary carcinoma, the MCa4. We have determined the dose response of 325 μl IM tumors in C3H mouse legs with a 50% control at 180 days with 52 Gy radiation (the TCD50). We have begun the study of tumor control using a whole tumor TCD15 of 49 Gy and boosts of 11 Gy to hypoxic and well oxygenated tumor of the same volume. With a median follow up of35 days of 10 animals, none of the hypoxic boost cohort have failed. All failure has been in the well oxygenated tumor boost cohort. Log-Rank significance is ~0.2. Our past experience has been that these early results portend future significance. Support: R01 CA098575, P41 EB002035, R50 CA211408 Citation Format: Howard J. Halpern, Boris Epel, Matthew C. Maggio, Martyna Krzykawska-Serda, Gage H. Redler, Richard C. Miller, Eugene D. Barth, Ralph R. Weichselbaum, Victor M. Tormyshev. Second tumor type showing hypoxia targeting with radiation improves tumor control in a mammal tumor model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4157.

Purpose: It has been known for over 100 years that tumor hypoxia, a near-universal characteristic of solid tumors, decreases the curative effectiveness of radiation therapy. However, to date, there are no reports that demonstrate an improvement in radiation effectiveness in a mammalian tumor on the basis of tumor hypoxia localization and local hypoxia treatment. Methods and Materials: For radiation targeting of hypoxic subregions in mouse fibrosarcoma, we used oxygen images obtained using pulse electron paramagnetic resonance pO(2) imaging combined with 3D-printed radiation blocks. This achieved conformal radiation delivery to all hypoxic areas in FSa fibrosarcomas in mice. Results: We demonstrate that treatment delivering a radiation boost to hypoxic volumes has a significant (P = .04) doubling of tumor control relative to boosts to well-oxygenated volumes. Additional dose to well-oxygenated tumor regions minimally increases tumor control beyond the 15% control dose to the entire tumor. If we can identify portions of the tumor that are more resistant to radiation, it might be possible to reduce the dose to more sensitive tumor volumes without significant compromise in tumor control. Conclusions: This work demonstrates in a single, intact mammalian tumor type that tumor hypoxia is a local tumor phenomenon whose treatment can be enhanced by local radiation. Despite enormous clinical effort to overcome hypoxic radiation resistance, to our knowledge this is the first such demonstration, even in preclinical models, of targeting additional radiation to hypoxic tumor to improve the therapeutic ratio. (C) 2018 Elsevier Inc. All rights reserved.

Abstract Stereotactic MCa4 and FSa tumor tissue biopsies registered with electron paramagnetic resonance (EPR) O2 imaging were used to identify sampling sites suitable to measure protein levels of three hypoxic cell biomarkers, hypoxia-induced factor 1-alpha (HIF-1α), carbonic anhydrase IX (CA9), and vascular endothelial growth factor (VEGF). EPR imaging provided quantitative localization of pO2 levels in tumors grown in the legs of C3H mice. Since hypoxic cells (pO2 <10 torr) are up to three times more resistant than normoxic cells to radiation lethality, identification of hypoxic populations to predict radiotherapeutic outcome is important. The oxygen broadening of narrow EPR spectral lines, or, equivalently, the increase in relaxation rates of electron magnetization, displays pO2 with 1-2 torr resolution in image voxels less than 1 mm3. The pO2 reporter molecule OX063 was used to image hypoxic areas in tumors. The molecule is selectively retained in tumors with a half-life of ~30 minutes. We used inversion recovery electron spin echo (IRESE) to measure the T1 rate of the trityl inside the tumor-bearing leg of mice. Voxel maps with O2 levels less than 10 torr (HF10) were used to guide biopsy cannulas to obtain tumor cell samples. Each of these biopsy cores were further subdivided into samples from which protein amounts could be determined via ELISA. Results from MCa4 and FSa tumor cells indicate a strong correlation between EPR pO2-identified hypoxic areas (<10 torr) and HIF-1α, CA9, and VEGF protein. The activation of various genes, including stem-cell activation, angiogenesis, vasodilation, glucose metabolism, reduced apoptotic activity, and cell cycle changes, contributes to poor radiotherapeutic outcome. EPR pO2 imaging may be used to precisely identify hypoxic areas harboring cells that protect tumor tissue from the toxicity of conventional radiation leading to radiotherapeutic failure. Grant Support: This work was supported by grants from the NIH, including R01 CA98575 and P41 EB002034. Citation Format: Richard Miller, Boris Epel, Martyna Elas, Martyna Krzykawska-Serda, Matthew Maggio, Eugene Barth, Mihai Giurcanu, Howard Halpern. Electron paramagnetic resonance (EPR) pO2 image-guided tumor biopsies to analyze hypoxia-induced proteins [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 668.