Image-guided Small Animal Irradiator Research Articles

Technical note: Multi-MATE, a high-throughput platform for automated image-guidedsmall-animal irradiation.

Small animal irradiation is essential to study the radiation response of new interventions before or parallel to human therapy. Image-guided radiotherapy (IGRT) and intensity-modulated radiotherapy (IMRT) are recently adopted in small animal irradiation to more closely mimic human treatments. However, sophisticated techniques require exceedingly high time, resources, and expertize that are often impractical. We propose a high throughput and high precision platform named Multiple Mouse Automated Treatment Environment (Multi-MATE) to streamline image-guided small animal irradiation. Multi-MATE consists of six parallel and hexagonally arranged channels, each equipped with a transfer railing, a 3D-printed immobilization pod, and an electromagnetic control unit, computer-controlled via an Arduino interface. The mouse immobilization pods are transferred along the railings between the home position outside the radiation field and the imaging/irradiation position at the irradiator isocenter. All six immobilization pods are transferred to the isocenter in the proposed workflow for parallel CBCT scans and treatment planning. The immobilization pods are then sequentially transported to the imaging/therapy position for dose delivery. The positioning reproducibility of Multi-MATE are evaluated using CBCT and radiochromic films. While parallelizing and automating the image-guided small animal radiation delivery, Multi-MATE achieved the average pod position reproducibility of 0.17 ± 0.04 mm in the superior-inferior direction, 0.20 ± 0.04 mm in the left-right direction, and 0.12 ± 0.02mm in the anterior-posterior direction in repeated CBCT tests. Additionally, in image-guided dose delivery tasks, Multi-MATE demonstrated the positioning reproducibility of 0.17 ± 0.06 mm in the superior-inferior direction, 0.19 ± 0.06 mm in the left-right direction. We designed, fabricated, and tested a novel automated irradiation platform, Multi-MATE to accelerate and automate image-guided small animal irradiation. The automated platform minimizes human operation and achieves high setup reproducibility and image-guided dose delivery accuracy. Multi-MATE thus removes a major barrier to implementing high-precision preclinical radiation research.

Integrated Small Animal PET/CT/RT with Onboard PET/CT Image Guidance for Preclinical Radiation Oncology Research.

We have integrated a compact and lightweight PET with an existing CT image-guided small animal irradiator to enable practical onboard PET/CT image-guided preclinical radiation therapy (RT) research. The PET with a stationary and full-ring detectors has ~1.1 mm uniform spatial resolution over its imaging field-of-view of 8.0 cm diameter and 3.5 cm axial length and was mechanically installed inside the irradiator in a tandem configuration with CT and radiation unit. A common animal bed was used for acquiring sequential dual functional and anatomical images with independent PET and CT control and acquisition systems. The reconstructed dual images were co-registered based on standard multi-modality image calibration and registration processes. Phantom studies were conducted to evaluate the integrated system and dual imaging performance. The measured mean PET/CT image registration error was ~0.3 mm. With one-bed and three-bed acquisitions, initial tumor focused and whole-body [18F]FDG animal images were acquired to test the capability of onboard PET/CT image guidance for preclinical RT research. Overall, the results have shown that integrated PET/CT/RT can provide advantageous and practical onboard PET/CT image to significantly enhance the accuracy of tumor delineation and radiation targeting that should enhance the existing and enable new and potentially breakthrough preclinical RT research and applications.

A comprehensive and efficient quality assurance program for an image-guided small animal irradiation system

In the field of preclinical radiotherapy, many new developments were driven by technical innovations. To make research of different groups comparable in that context and reliable, high quality has to be maintained. Therefore, standardized protocols and programs should be used. Here we present a guideline for a comprehensive and efficient quality assurance program for an image-guided small animal irradiation system, which is meant to test all the involved subsystems (imaging, treatment planning, and the irradiation system in terms of geometric accuracy and dosimetric aspects) as well as the complete procedure (end-to-end test) in a time efficient way. The suggestions are developed on a Small Animal Radiation Research Platform (SARRP) from Xstrahl (Xstrahl Ltd., Camberley, UK) and are presented together with proposed frequencies (from monthly to yearly) and experiences on the duration of each test. All output and energy related measurements showed stable results within small variation. Also, the motorized parts (couch, gantry) and other geometrical alignments were very stable. For the checks of the imaging system, the results are highly dependent on the chosen protocol and differ according to the settings. We received nevertheless stable and comparably good results for our mainly used protocol. All investigated aspects of treatment planning were exactly fulfilled and also the end-to-end test showed satisfying values. The mean overall time we needed for our checks to have a well monitored machine is less than two hours per month.

Evaluation of a micro ionization chamber for dosimetric measurements in image-guided preclinical irradiation platforms

Image-guided small animal irradiation platforms deliver small radiation fields in the medium energy x-ray range. Commissioning of such platforms, followed by dosimetric verification of treatment planning, are mostly performed with radiochromic film. There is a need for independent measurement methods, traceable to primary standards, with the added advantage of immediacy in obtaining results. This investigation characterizes a small volume ionization chamber in medium energy x-rays for reference dosimetry in preclinical irradiation research platforms. The detector was exposed to a set of reference x-ray beams (0.5–4 mm Cu HVL). Leakage, reproducibility, linearity, response to detector’s orientation, dose rate, and energy dependence were determined for a 3D PinPoint ionization chamber (PTW 31022). Polarity and ion recombination were also studied. Absorbed doses at 2 cm depth were compared, derived either by applying the experimentally determined cross-calibration coefficient at a typical small animal radiation platform ‘user’s’ quality (0.84 mm Cu HVL) or by interpolation from air kerma calibration coefficients in a set of reference beam qualities. In the range of reference x-ray beams, correction for ion recombination was less than 0.1%. The largest polarity correction was 1.4% (for 4 mm Cu HVL). Calibration and correction factors were experimentally determined. Measurements of absorbed dose with the PTW 31022, in conditions different from reference were successfully compared to measurements with a secondary standard ionization chamber. The implementation of an End-to-End test for delivery of image-targeted small field plans resulted in differences smaller than 3% between measured and treatment planning calculated doses. The investigation of the properties and response of a PTW 31022 small volume ionization chamber in medium energy x-rays and small fields can contribute to improve measurement uncertainties evaluation for reference and relative dosimetry of small fields delivered by preclinical irradiators while maintaining the traceability chain to primary standards.

Development of an Orthotopic Murine Model of Rectal Cancer in Conjunction With Targeted Short-Course Radiation Therapy

PurposeOrthotopic tumors more closely recapitulate human cancers than do ectopic models; however, precision targeting of such internal tumors for radiation therapy (RT) without inducing systemic toxicity remains a barrier. We developed an innovative murine orthotopic rectal tumor model where the insertion of clinical grade titanium fiducial clips on opposing sides of the rectal tumor allowed for targeted administration of short-course radiation therapy (SCRT). With this novel approach, clinically relevant RT regimens can be administered to orthotopic tumors to explore the biology and efficacy of radiation alone or as a combination therapy in a murine model that closely recapitulates human disease. Methods and MaterialsMurine Colon 38-luciferase tumor cells were injected into the rectal wall of syngeneic mice, and fiducial clips were applied to demarcate the tumor. An SCRT regimen consisting of 5 consecutive daily doses of 5 Gy delivered by an image-guided conformal small animal irradiator was administered 9 days after implantation. Tumor burden and survival were monitored along with histological and flow cytometric analyses on irradiated versus untreated tumors at various time points. ResultsSCRT administered to orthotopic rectal tumors resulted in a reduction in tumor burden and enhanced overall survival with no apparent signs of systemic toxicity. This treatment paradigm resulted in significant reductions in tumor cellularity and increases in fibrosis and hyaluronic acid production, recapitulating the SCRT-induced effects observed in human cancers. ConclusionsWe have established a means to target murine orthotopic rectal tumors using fiducial markers with a fractionated and clinically relevant SCRT schedule that results in an RT response similar to what is observed in human rectal cancer. We also validated our model through examining various parameters associated with human cancer that are influenced by irradiation. This model can be used to further explore RT doses and scheduling, and to test combinatorial therapies.

SPOP Deregulation Improves the Radiation Response of Prostate Cancer Models by Impairing DNA Damage Repair.

Speckle-type POZ (pox virus and zinc finger protein) protein (SPOP) is the most commonly mutated gene in prostate cancer (PCa). Recent evidence reports a role of SPOP in DNA damage response (DDR), indicating a possible impact of SPOP deregulation on PCa radiosensitivity. This study aimed to define the role of SPOP deregulation (by gene mutation or knockdown) as a radiosensitizing factor in PCa preclinical models. To express WT or mutant (Y87N, K129E and F133V) SPOP, DU145 and PC-3 cells were transfected with pMCV6 vectors. Sensitivity profiles were assessed using clonogenic assay and immunofluorescent staining of γH2AX and RAD51 foci. SCID xenografts were treated with 5 Gy single dose irradiation using an image-guided small animal irradiator. siRNA and miRNA mimics were used to silence SPOP or express the SPOP negative regulator miR-145, respectively. SPOP deregulation, by either gene mutation or knockdown, consistently enhanced the radiation response of PCa models by impairing DDR, as indicated by transcriptome analysis and functionally confirmed by decreased RAD51 foci. SPOP silencing also resulted in a significant downregulation of RAD51 and CHK1 expression, consistent with the impairment of homologous recombination. Our results indicate that SPOP deregulation plays a radiosensitizing role in PCa by impairing DDR via downregulation of RAD51 and CHK1.

A failure modes and effects analysis quality management framework for image-guided small animal irradiators: A change in paradigm for radiation biology.

Image-guided small animal irradiators (IGSAI) are increasingly being adopted in radiation biology research. These animal irradiators, designed to deliver radiation with submillimeter accuracy, exhibit complexity similar to that of clinical radiation delivery systems, including image guidance, robotic stage motion, and treatment planning systems. However, physics expertise and resources are scarcer in radiation biology, which makes implementation of conventional prescriptive QA infeasible. In this study, we apply the failure modes and effect analysis (FMEA) popularized by the AAPM task group 100 (TG-100) report to IGSAI and radiation biological research. Radiation biological research requires a change in paradigm where small errors to large populations of animals are more severe than grievous errors that only affect individuals. To this end, we created a new adverse effects severity table adapted to radiation biology research based on the original AAPM TG-100 severity table. We also produced a process tree which outlines the main components of radiation biology studies performed on an IGSAI, adapted from the original clinical IMRT process tree from TG-100. Using this process tree, we created and distributed a preliminary survey to eight expert IGSAI operators in four institutions. Operators rated proposed failure modes for occurrence, severity, and lack of detectability, and were invited to share their own experienced failure modes. Risk probability numbers (RPN) were calculated and used to identify the failure modes which most urgently require intervention. Surveyed operators indicated a number of high (RPN >125) failure modes specific to small animal irradiators. Errors due to equipment breakdown, such as loss of anesthesia or thermal control, received relatively low RPN (12-48) while errors related to the delivery of radiation dose received relatively high RPN (72-360). Errors identified could either be improved by manufacturer intervention (e.g., electronic interlocks for filter/collimator) or physics oversight (errors related to tube calibration or treatment planning system commissioning). Operators identified a number of failure modes including collision between the collimator and the stage, misalignment between imaging and treatment isocenter, inaccurate robotic stage homing/translation, and incorrect SSD applied to hand calculations. These were all relatively highly rated (90-192), indicating a possible bias in operators towards reporting high RPN failure modes. The first FMEA specific to radiation biology research was applied to image-guided small animal irradiators following the TG-100 methodology. A new adverse effects severity table and a process tree recognizing the need for a new paradigm were produced, which will be of great use to future investigators wishing to pursue FMEA in radiation biology research. Future work will focus on expanding scope of user surveys to users of all commercial IGSAI and collaborating with manufacturers to increase the breadth of surveyed expert operators.

Preclinical Models of Craniospinal Irradiation for Medulloblastoma.

Medulloblastoma is an embryonal tumor that shows a predilection for distant metastatic spread and leptomeningeal seeding. For most patients, optimal management of medulloblastoma includes maximum safe resection followed by adjuvant craniospinal irradiation (CSI) and chemotherapy. Although CSI is crucial in treating medulloblastoma, the realization that medulloblastoma is a heterogeneous disease comprising four distinct molecular subgroups (wingless [WNT], sonic hedgehog [SHH], Group 3 [G3], and Group 4 [G4]) with distinct clinical characteristics and prognoses has refocused efforts to better define the optimal role of CSI within and across disease subgroups. The ability to deliver clinically relevant CSI to preclinical models of medulloblastoma offers the potential to study radiation dose and volume effects on tumor control and toxicity in these subgroups and to identify subgroup-specific combination adjuvant therapies. Recent efforts have employed commercial image-guided small animal irradiation systems as well as custom approaches to deliver accurate and reproducible fractionated CSI in various preclinical models of medulloblastoma. Here, we provide an overview of the current clinical indications for, and technical aspects of, irradiation of pediatric medulloblastoma. We then review the current literature on preclinical modeling of and treatment interventions for medulloblastoma and conclude with a summary of challenges in the field of preclinical modeling of CSI for the treatment of leptomeningeal seeding tumors.

Effects of Brain Irradiation in Immune-Competent and Immune-Compromised Mouse Models.

Patient-derived orthotopic xenografts (PDOXs) closely recapitulate primary human glioblastoma (GBM) tumors in terms of histology and genotype. Compared to other mouse strains, NOD-scid IL2Rgammanull (NSG) mice show excellent tumor take rates, which makes them an ideal host for PDOXs. However, NSG mice harbor a mutation in the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs), which renders them relatively radiosensitive. This has been a frequently voiced concern in studies involving ionizing radiation. In this study, we assessed brain toxicity in NSG mice compared to three other different mouse strains frequently used in radiation studies at radiation doses commonly used in experimental combination therapy studies. C3H/Sed/Kam, C57Bl/6, nude and NOD-scid IL2Rgammanull mice received a single dose of 4 Gy to the right brain hemispheres using an image-guided small animal irradiator. Brains were stained using H&E, luxol fast blue, and antibodies against IBA1 and GFAP one, two, four or six months postirradiation. Additional animals of all four strains were exposed to five daily fractions of 2 Gy (5 × 2 Gy), and tissue sections were stained 72 h later against gH2AX, NeuN, GFAP and IBA1. None of the mouse strains displayed radiation-induced toxicity at any of the time points tested. Radiation doses relevant for testing combination therapies can be safely applied to the brains of NSG mice without the occurrence of radiation-induced normal tissue toxicity.

P1.06-15 Safety of Irradiation Combined with Intracavitary Cisplatin-Fibrin After Lung-Sparing Surgery in a Rat Model of Mesothelioma

To investigate feasibility and toxicity of new localized therapeutic treatment combinations for malignant pleural mesothelioma (MPM), we performed lung-sparing surgery followed by cisplatin-fibrin application and hemithoracic irradiation in an orthotopic immunocompetent rat model of MPM. Male F344 rats (n=9) were implanted sub-pleurally (parietal pleura) with 1 million rat mesothelioma cells (IL45-luciferase). Formed tumor nodules confirmed by IVIS bioluminescence imaging (BLI) were resected on day 9 after implantation. Following resection, animals were treated with local-intracavitary cisplatin-fibrin or placebo (NaCl-fibrin). Three days later, CT guided local irradiation of the former tumor region, resembling IMRT in human patients, was performed. Irradiation was given in a single high dose application using the image-guided stereotactic small animal irradiation X-RAD SmART (small animal radiotherapy) and image guided biological irradiator PXi (precision X-Ray) with precise localization. Treatment schemes after tumor resection were as followed: i) Intracavitary cisplatin-fibrin application (n=2) ii) Irradiation with 10 Gy (n=2) iii) Irradiation with 20 Gy (n=1) iv) Intracavitary cisplatin-fibrin plus 10 Gy radiotherapy (n=2) v) Intracavitary cisplatin-fibrin plus 20 Gy radiotherapy (n=2) Wellbeing of the animals was monitored daily until the predefined termination criteria were reached. Particular attention was given to possible irradiation toxicity related pulmonary side effects and weight loss. We successfully treated 1-2 animals per group according to the methods above. The irradiation was performed after visualization of the tumor with BLI- and CT-imaging to ensure an individual treatment plan. None of the animals, whether with radiotherapy alone or in combination with cisplatin-fibrin application, showed any signs of pulmonary side effects. In addition, none had reduced pulmonary functions, measured by increased breathing or the appearance of blue or white colored ear/extremities/eyes assuming desaturation. Furthermore, neither significant body weight loss of ≥15%, deterioration of body conditioning score nor of the activity score were observed in the immediate post-interventional phase. In all animals, termination endpoint was reached because of tumor relapse. In this pilot study, we have shown that irradiation alone and in combination with local intracavitary cisplatin-fibrin application in rats is safe and feasible up to a dosage of 20 Gy. The efficacy of the various treatment schemes and a possible radio-sensitizing effect by intracavitary cisplatin is currently being evaluated in the same animal model.

Preclinical dose verification using a 3D printed mouse phantom for radiobiology experiments.

Dose verification in preclinical radiotherapy is often challenged by a lack of standardization in the techniques and technologies commonly employed along with the inherent difficulty of dosimetry associated with small-field kilovoltage sources. As a consequence, the accuracy of dosimetry in radiobiological research has been called into question. Fortunately, the development and characterization of realistic small-animal phantoms has emerged as an effective and accessible means of improving dosimetric accuracy and precision in this context. The application of three-dimensional (3D) printing, in particular, has enabled substantial improvements in the conformity of representative phantoms with respect to the small animals they are modeled after. In this study, our goal was to evaluate a fully 3D printed mouse phantom for use in preclinical treatment verification of sophisticated therapies for various anatomical targets of therapeutic interest. An anatomically realistic mouse phantom was 3D printed based on segmented microCT data of a tumor-bearing mouse. The phantom was modified to accommodate both laser-cut EBT3 radiochromic film within the mouse thorax and a plastic scintillator dosimeter (PSD), which may be placed within the brain, abdomen, or 1-cm flank subcutaneous tumor. Various treatments were delivered on an image-guided small-animal irradiator in order to determine the doses to isocenter using a PSD and validate lateral- and depth-dose distributions using film dosimeters. On-board cone-beam CT imaging was used to localize isocenter to the film plane or PSD active element prior to irradiation. The PSD irradiations comprised a 3×3mm2 brain arc, 5×5mm2 parallel-opposed pair (POP), and 5-beam 10×10mm2 abdominal coplanar arrangement while two-dimensional (2D) film dose distributions were acquired using a 3×3mm2 arc and both 5×5 and 10×10mm2 3-beam coplanar plans. A validated Monte Carlo (MC) model of the source was used as to verify the accuracy of the film and PSD dose measurements. computer-aided design (CAD) geometries for the mouse phantom and dosimeters were imported directly into the MC code to allow for highly accurate reproduction of the physical experiment conditions. Experimental and MC-derived film data were co-registered and film dose profiles were compared for points above 90% of the dose maximum. Point dose measurements obtained with the PSD were similarly compared for each of the candidate (brain, abdomen, and tumor) treatment sites. For each treatment configuration and anatomical target, the MC-calculated and measured doses met the proposed 5% agreement goal for dose accuracy in radiobiology experiments. The 2D film and MC dose distributions were successfully registered and mean doses for lateral profiles were found to agree to within 2.3% in all cases. Isocentric point-dose measurements taken with the PSD were similarly consistent, with a maximum percentage deviation of 3.2%. Our study confirms the utility of 3D printed phantom design in providing accurate dose estimates for a variety of preclinical treatment paradigms. As a tool for pretreatment dose verification, the phantom may be of particular interest to researchers for its ability to facilitate precise dosimetry while fostering a reduction in cost for radiobiology experiments.

Development of a Compact PET for integrated PET/CT/RT to Streamline and Enhance Functional/Anatomic Image-Guided Preclinical Radiation Oncology Researches

To present latest progress in developing a compact and high-performance PET that will be inserted inside an existing CT image-guided small animal irradiator to establish an integrated PET/CT/RT for substantially expanded and improved image-guided RT research capabilities. 1) PET will consist of 12 detector panels in a multi-polygon configuration to provide ∼10 cm diameter trans-axial and ∼3.2 cm axial image field-of-view: Each panel consists of 4 detectors that closely tiled in a 2x2 matrix: Each detector consists of 15x15 array of 1x1x20 mm LYSO scintillators, with each end of the scintillator array read out by an 8x8 array of 2x2 mm Silicon Photomultiplier (SiPM) array to provide dual-scintillator-end readout for measuring depth-of-interaction (DOI); 2) PC Board of discrete components based electronics have been developed for reading/processing total 768 detector output channels (after readout channel reduction) with both positive and negative polarity signals from SiPMs’ anodes and cathodes; 3) Standard detector performance evaluation have been conducted, including energy, coincidence timing, DOI, and intrinsic detector spatial resolutions; 4) Imaging performance based on rotating detector panels is under investigation. Two full detector panels have already been assembled and placed opposite to each other on a rotating table, with a radioactive rod phantom between the panels. Coincident data will be acquired by the detector pairs at 6 different rotating angles span over 180 degrees, with 30 degrees between each consecutive detector angles. Data corrections, including attenuation, scatter and random will be applied before the image reconstruction. Expanded dual positive and negative polarity signal readout and processing electronics works well. All 900 crystals on each detector panel can be very well identified; energy, coincidence timing, and DOI are 18%-34%, ∼2.0 ns, and 2.4 – 3.7 mm, respectively. The detector intrinsic spatial resolution is < 1.0 mm. Overall, detectors have achieved expected performance. As anticipated, ongoing phantom imaging study with dual rotating detector panels, potentially along with the performance evaluation study of the assembled PET system, will be reported in the conference. We have developed PET detectors and readout/processing electronics with desired performance, and expect the ongoing system construction and imaging studies will quantify the PET capability for functional image guidance for small animal radiotherapy research.

Pre-clinical and small field dosimetry

Preclinical in vivo studies have drastically improved over the past decade with the development of cone beam computed tomography (CBCT) image-guided small animal irradiation systems. Such systems produce 220 or 225 kV x-rays with square and circular field sizes ranging from 0.5 to 10 mm. The dosimetry of such equipment involving kilovoltage small-field dosimetry has not received as much attention as the megavoltage small-field dosimetry. The dosimetry of megavoltage small fields can be challenging due to lateral charged particle disequilibrium, detector volume averaging effect, and high dose gradients. Clinically there has been a rapid increase in the use of small fields in modern radiotherapy techniques such as stereotactic radiosurgery (SRS) and stereotactic radiotherapy (SBRT). This study presents dosimetric properties of image-guided small animal irradiation systems. Both EBT Gafchromic films and 3-D PRESAGE measured beam data were presented and compared with the calculated dose distribution from a commissioned planning system. For megavoltage small-field dosimetry, EBT3 films and PRESAGE dosimeters were used to measure the dose distributions for MLC-delimited 6 x 6 to 20 x 20 mm2 square fields, and for selected IMRT and VMAT plans with small field sizes or segments of 1-4 cm. A single-beam optical CT scanner was used as the readout mechanism of the radiation-induced 3-D information in the PRESAGE phantoms. Measured data sets were compared with calculated results from Eclipse Acuros XB. The results for kilovoltage small animal irradiator showed that PDD data measured from EBT films and PRESAGE dosimeters are in agreement within 3%; profiles and 2-D dose distributions measured from PRESAGE present a larger penumbra compared with those from EBT films. Discrepancies between measured beam data and treatment planning data were identified. For megavoltage small fields, the measured data percent depth dose, beam profiles, and dose distributions were found to agree within experimental uncertainties for EBT films and PRESAGE dosimeters. Beam profiles for MLC-delimited field sizes less than 10 mm reveal some discrepancies in the penumbra region between measured data and calculated results. Dose distributions from EBT film and PRESAGE measurements demonstrate that 3-D dosimetry measurement for small-field IMRT and VMAT QA may be necessary to ensure a complete verification of dose delivery and accuracy.

A 3D printed modular phantom for quality assurance of image-guided small animal irradiators: Design, imaging experiments, and Monte Carlo simulations.

The goal of this work was to develop and test a cylindrical tissue-equivalent quality assurance (QA) phantom for micro computed tomography (microCT) image-guided small animal irradiators that overcomes deficiencies of existing phantoms due to its mouse-like dimensions and composition. The 8.6-cm-long and 2.4-cm-diameter phantom was three-dimensionally (3D) printed out of Somos NeXt plastic on a stereolithography (SLA) printer. The modular phantom consisted of four sections: (a) CT number evaluation section, (b) spatial resolution with slanted edge (for the assessment of longitudinal resolution) and targeting section, (c) spatial resolution with hole pattern (for the assessment of radial direction) section, and (d) uniformity and geometry section. A Python-based graphical user interface (GUI) was developed for automated analysis of microCT images and evaluated CT number consistency, longitudinal and radial modulation transfer function (MTF), image uniformity, noise, and geometric accuracy. The phantom was placed at the imaging isocenter and scanned with the small animal radiation research platform (SARRP) in the pancake geometry (long axis of the phantom perpendicular to the axis of rotation) with a variety of imaging protocols. Tube voltage was set to 60 and 70kV, tube current was set to 0.5 and 1.2mA, voxel size was set to 200 and 275μm, imaging times of 1, 2, and 4min were used, and frame rates of 6 and 12 frames per second (fps) were used. The phantom was also scanned in the standard (long axis of the phantom parallel to the axis of rotation) orientation. The quality of microCT images was analyzed and compared to recommendations presented in our previous work that was derived from a multi-institutional study. Additionally, a targeting accuracy test with a film placed in the phantom was performed. MicroCT imaging of the phantom was also simulated in a modified version of the EGSnrc/DOSXYZnrc code. Images of the resolution section with the hole pattern were acquired experimentally as well as simulated in both the pancake and the standard imaging geometries. The radial spatial resolution of the experimental and simulated images was evaluated and compared to experimental data. For the centered phantom images acquired in the pancake geometry, all imaging protocols passed the spatial resolution criterion in the radial direction (>1.5lp/mm @ 0.2MTF), the geometric accuracy criterion (<200μm), and the noise criterion (<55HU). Only the imaging protocol with 200-μm voxel size passed the criterion for spatial resolution in the longitudinal direction (>1.5lp/mm @ 0.2MTF). The 70-kV tube voltage dataset failed the bone CT number consistency test (<55HU). Due to cupping artifacts, none of the imaging protocols passed the uniformity test of <55HU. When the phantom was scanned in the standard imaging geometry, image uniformity and longitudinal MTF were satisfactory; however, the CT number consistency failed the recommended limit. A targeting accuracy of 282 and 251μm along the x- and z-direction was observed. Monte Carlo simulations confirmed that the radial spatial resolution for images acquired in the pancake geometry was higher than the one acquired in the standard geometry. The new 3D-printed phantom presents a useful tool for microCT image analysis as it closely mimics a mouse. In order to image mouse-sized animals with acceptable image quality, the standard protocol with a 200-μm voxel size should be chosen and cupping artifacts need to be resolved.

Small animal image-guided radiotherapy

Small animal image-guided radiotherapy

Characterization of a plastic scintillating detector for the Small Animal Radiation Research Platform (SARRP).

The purpose of this study was to characterize a small plastic scintillator developed for high resolution, real-time dosimetry of therapy and imaging x-ray beams delivered by an image-guided small animal irradiator. A 1mm diameter, 1mm long polystyrene BCF-60 scintillating fiber dosimeter was characterized with 220kVp therapy and 40, 50, 60, 70, and 80kVp imaging beams on the Small Animal Research Platform (SARRP). Scintillator output, sensitivity (charge per unit dose), linearity, and 0.2-mm resolution beam profile measurements were performed. A validated in-house Monte Carlo (MC) model of the SARRP was used to compute detailed energy spectra at locations of dosimetry, and validated scintillator measurement with MC simulations. Mass energy-absorption coefficients from the National Institute of Standards and Technology (NIST) tables convolved with MC-derived spectra were used in conjunction with Birks ionization quenching factors to correct scintillator output. An air kerma calibration method was employed to correct scintillator output for in-air beam profile measurements with open, 5×5, and 3×3mm2 square field sizes, and compared to MC simulations. Scintillator dose response showed excellent linearity (R2 ≥0.999) for all sensitivity measurements, including output as a function of tube current. Detector sensitivity was 2.41μCGy-1 for the 220kVp therapy beam, and it ranged from 1.21 to 1.32μCGy-1 for the 40-80 imaging beams. Percentage difference in sensitivity between the therapy and imaging beams before sensitivity correction and after using the Birks quenching factors were 52.3% and 10.2%, respectively. Percentage differences between the therapy and imaging beam sensitivities after using the air kerma calibration method for in-air measurements was excellent and below 0.3%. In-air beam profile measurements agreed to MC simulations within a mean difference of 2.4% for the 5×5 and 3×3mm2 field sizes, however, the scintillator showed signs of volume averaging at the penumbra edges. A small plastic scintillator was characterized for therapy and imaging energies of a small animal irradiator, with output corrected for using an in-house MC model of the irradiator. The characterization of the scintillator detector system for small fields presents steps toward implementing real-time measurements for quality assurance and small animal treatment and imaging dose verification.

The potential impact of ultrathin filter design on dosimetry and relative biological effectiveness in modern image-guided small animal irradiators.

Modern image-guided small animal irradiators like the Xstrahl Small Animal Radiation Research Platform (SARRP) are designed with ultrathin 0.15 mm Cu filters, which compared with more heavily filtrated traditional cabinet-style biological irradiators, produce X-ray spectra weighted toward lower energies, impacting the dosimetric properties and the relative biological effectiveness (RBE). This study quantifies the effect of ultrathin filter design on relative depth dose profiles, absolute dose output, and RBE using Monte Carlo techniques. The percent depth-dose and absolute dose output are calculated using kVDoseCalc and EGSnrc, respectively, while a tally based on the induction of double-strand breaks as a function of electron spectra invoked in PENELOPE is used to estimate the RBE. The RBE increases by >2.4% in the ultrathin filter design compared to a traditional irradiator. Furthermore, minute variations in filter thickness have notable effects on the dosimetric properties of the X-ray beam, increasing the percent depth dose (at 2 cm in water) by + 0.4%/0.01 mm Cu and decreasing absolute dose (at 2 cm depth in water) by -1.8%/0.01 mm Cu for the SARRP. These results show that modern image-guided irradiators are quite sensitive to small manufacturing variations in filter thickness, and show a small change in RBE compared to traditional X-ray irradiators. We quantify the consequences of ultrathin filter design in modern image-guided biological irradiators on relative and absolute dose, and RBE. Our results show these to be small, but not insignificant, suggesting laboratories transitioning between irradiators should carefully design their radiobiological experiments.

Dosimetric Assessment of Treatment Planning in an Image-guided Small Animal Irradiator

Dosimetric Assessment of Treatment Planning in an Image-guided Small Animal Irradiator

Abstract 3766: Radiation enhances anti-tumor activity of immune checkpoint blockade in syngeneic tumor models

Abstract More than 50% of cancer patients receive some sort of radiation therapy during the course of their illness. While radiation treatment is a mainstay in clinical oncology, there is limited preclinical data in syngeneic models. The advent of image-guided small animal irradiators such as the Small Animal Radiation Research Platform (SARRP; Xstrahl) allow for use of targeted focal irradiation (RT) in a broad range of models. Here we evaluated the use of RT to broaden efficacy and response duration of immunomodulatory therapies. As a first step, subcutaneous syngeneic tumor models were tested with single doses of RT. The A20 B cell lymphoma, the CT26 colon carcinoma, and the Pan02 pancreatic carcinoma all demonstrated dose response anti-tumor activity with increasing dose (5, 10 or 20Gy) of RT. In the A20 model, doses of 10 and 20Gy RT resulted in 2 tumor free survivors (TFS) each. CT26 had 1, 2 and 2 TFS each in RT groups of 5, 10 and 20Gy, respectively. In these 2 models, tumor free mice were rechallenged with appropriate tumor cell implants on the contralateral flank and, in all cases, tumors failed to develop suggesting a memory immune response was elicited. In Pan02 we found 1 and 5 TFS in the 10 and 20Gy groups, respectively. In follow-on work, we used the CT26 model to compare 5Gy RT, bi-weekly dosing of anti-PD-1 (RMP1-14; Bio X Cell) or the combination. Treatments were initiated at approximately 100mm3 tumor volumes. Anti-PD-1 resulted in a slight tumor growth delay (TGD) of 5 days compared to isotype control, whereas RT resulted in a 19 day TGD and more than doubled the time to evaluation size (TES) from 16 days in the control to 35 days. In combination, the TGD was further improved to 40 days and TES to 56 days. An additional colon model, MC38, was tested with 10Gy RT alone or in combination with anti-PD-1, or anti-PD-L1. Either antibody resulted in modest single agent activity with anti-PD-1 being slightly more efficacious than anti-PD-L1 with 15 days TGD compared to 7 days, respectively. Mice in the RT group demonstrated 20 days TGD. RT in combination with anti-PD-1 resulted in 37 days TGD, however, in combination with anti-PD-L1 TGD was not improved over RT alone. Effects of radiation and anti-PD-1 therapy on tumor infiltrating lymphocytes is being investigated in the MC38 model, and data will be shown. GL261-luc, an orthotopic model of murine GBM, was also tested. We found that anti-PD-1 improved anti-tumor activity from control, a single dose of 7.5Gy RT further improved activity against control, and the combination was 100% curative. Evaluation of tumor infiltrating lymphocytes showed increased CD8+ T cells along with increased regulatory T cells (Tregs) as the model progressed. RT reduced the number of Tregs and a further reduction was observed following combination therapy. In summary, the data presented here provides rationale for further evaluation of immunomodulatory agents in combination with RT. Citation Format: Sumithra Urs, Mary Anne Meade, Kevin Guley, Sarah Krueger, Alden Wong, Scott Wise, Maryland Franklin. Radiation enhances anti-tumor activity of immune checkpoint blockade in syngeneic tumor models [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 3766.

32 - Protection of rat prostate and erectile function from radiation-induced damage by novel Mn(III) N- substituted pyridylporphyrin and ascorbate

Prostate cancer is the most common cancer in American men, with approximately 30-50% of men developing loss of erectile function after definitive radiotherapy (RT). Currently, no treatment is available that preserves erectile function. Therefore there is a significant reduction of the quality of life of survivors which incurs considerable healthcare costs. Hence, there is an imminent need to protect normal tissues during prostate cancer RT. We have developed Mn porphyrin-based SOD mimics and demonstrated that they radioprotect normal tissues such as brain, salivary glands and mouth mucosa while radio- and chemosensitizing tumors, such as brain, breast and lymphoma. We have also demonstrated and explained the reasoning behind the significant enhancement of tumor radiosensitization by Mn porphyrins when combined with ascorbate (Asc). A lead drug, BMX-001 is now in two Phase I/II clinical trials as a normal tissue radioprotector with glioma and head&neck cancer patients. Herein we have explored the radioprotection of a lower pelvic region by a member of the new class of powerful Mn porphyrin-based SOD mimics – Mn(III) meso-tetrakis(5',5',5'-trifluoropentylpyridinium-2-yl)porphyrin, MnF3Pen. The SOD-like activity given by log kcat(O2.-) is 7.45, and the reduction potential of the Mn site, E1/2 is +288 mV vs NHE. In this study we also aimed to show that the addition of Asc would not diminish radioprotection of a normal tissue by MnF3Pen. The rat toxicity and pharmacokinetic studies of MnF3Pen/Asc system showed that MnF3Pen reached ~100 to 500 nM levels at targeted sites (penis, prostate, bladder, testes and rectum) within few hours after single subcutaneous (sc) 1 mg/kg injection. We subsequently performed the efficacy study. Single radiation dose of 20 Gy was delivered stereotactically to the prostate of Sprague-Dawley 8 week old rats using image-guided small animal irradiator. MnF3Pen was given sc at 0.5 mg/kg/day for four weeks and continued twice weekly for the next 5 weeks. Ascorbate was given intraperitoneally at 1 g/kg/day for first 3 days and then twice weekly for the rest of 9 weeks. We showed that MnF3Pen/Asc treatment prevented erectile dysfunction observed in RT-treated group. We further showed that MnF3Pen/Asc treatment greatly suppressed RT-induced damage to prostate as evaluated by H&E staining.