Minimum reporting standards should be expected for preclinical radiobiology irradiators and dosimetry in the published literature.

The cornerstones of science advancement are rigor in performing scientific research, reproducibility of research findings and unbiased reporting of design and results of the experiments. For radiation research, this requires rigor in describing experimental details as well as the irradiation protocols for accurate, precise and reproducible dosimetry. Most institutions conducting radiation biology research in in vitro or animal models do not have describe experimental irradiation protocols in sufficient details to allow for balanced review of their publication nor for other investigators to replicate published experiments. The need to increase and improve dosimetry standards, traceability to National Institute of Standards and Technology (NIST) standard beamlines, and to provide dosimetry harmonization within the radiation biology community has been noted for over a decade both within the United States and France. To address this requirement subject matter experts have outlined minimum reporting standards that should be included in published literature for preclinical irradiators and dosimetry.

Dichotomic Potency of IFNγ Licensed Allogeneic Mesenchymal Stromal Cells in Animal Models of Acute Radiation Syndrome and Graft Versus Host Disease.

Mesenchymal stromal cells (MSCs) are being tested as a cell therapy in clinical trials for dozens of inflammatory disorders, with varying levels of efficacy reported. Suitable and robust preclinical animal models for testing the safety and efficacy of different types of MSC products before use in clinical trials are rare. We here introduce two highly robust animal models of immune pathology: 1) acute radiation syndrome (ARS) and 2) graft versus host disease (GvHD), in conjunction with studying the immunomodulatory effect of well-characterized Interferon gamma (IFNγ) primed bone marrow derived MSCs. The animal model of ARS is based on clinical grade dosimetry precision and bioluminescence imaging. We found that allogeneic MSCs exhibit lower persistence in naïve compared to irradiated animals, and that intraperitoneal infusion of IFNγ prelicensed allogeneic MSCs protected animals from radiation induced lethality by day 30. In direct comparison, we also investigated the effect of IFNγ prelicensed allogeneic MSCs in modulating acute GvHD in an animal model of MHC major mismatched bone marrow transplantation. Infusion of IFNγ prelicensed allogeneic MSCs failed to mitigate acute GvHD. Altogether our results demonstrate that infused IFNγ prelicensed allogeneic MSCs protect against lethality from ARS, but not GvHD, thus providing important insights on the dichotomy of IFNγ prelicensed allogenic MSCs in well characterized and robust animal models of acute tissue injury.

Radiation Biology Irradiator Dose Verification Survey.

Interest in standardized dosimetry for radiobiological irradiators has expanded over the last decade. At a symposium held at NIST, "The Importance of Standardization of Dosimetry in Radiobiology", a set of 12 criteria necessary for adequate irradiation was developed by the authors. Here we report on our review of dosimetry methods from various peer-reviewed publications and found that none of them satisfied all 12 criteria set forth by the authors of the NIAD/NCI/NIST proceedings. The inadequate reporting of dosimetry methods in the literature raises questions regarding the accuracy of the dose delivered to animal test subjects and the resulting experimental results. For this reason, we investigated the level of accuracy of dose delivery in radiation biology studies. We performed an irradiator output verification study of 12 radiation biology laboratories (7 gamma and 5 X-ray units) using polymethyl methacrylate (PMMA) mouse phantoms and thermoluminescent dosimeters (TLDs) readouts at the University of Wisconsin Medical Radiation Research Center (UWMRRC). The laboratories housing each of these irradiators were asked to deliver specific doses to individual mouse phantoms. Simultaneously, mouse phantoms at the UWMRRC were irradiated with NIST-traceable reference beams representative of the subject laboratories' beam energies. The irradiated mouse phantoms were returned from the various institutions to the UWMRRC and the TLDs were processed, with their measured dose response compared to the known dose response of the calibration phantom TLDs. Of the five facilities using X-ray irradiators, only one delivered an output within 5% of the target dose. The dose differences for the other four X-ray irradiators ranged from 12 to 42%. These results indicate the potential need for standardization of dose determination and additional oversight of radiobiology investigations.

Determination of exit skin dose for 192Ir intracavitary accelerated partial breast irradiation with thermoluminescent dosimeters.

PURPOSE: Intracavitary accelerated partial breast irradiation (APBI) has become a popular treatment for early stage breast cancer in recent years due to its shortened course of treatment and simplified treatment planning compared to traditional external beam breast conservation therapy. However, the exit dose to the skin is a major concern and can be a limiting factor for these treatments. Most treatment planning systems (TPSs) currently used for high dose-rate (HDR) 192Ir brachytherapy overestimate the exit skin dose because they assume a homogeneous water medium and do not account for finite patient dimensions. The purpose of this work was to quantify the TPS overestimation of the exit skin dose for a group of patients and several phantom configurations. METHODS: The TPS calculated skin dose for 59 HDR 192Ir APBI patients was compared to the skin dose measured with LiF:Mg,Ti thermoluminescent dosimeters (TLDs). Additionally, the TPS calculated dose was compared to the TLD measured dose and the Monte Carlo (MC) calculated dose for eight phantom configurations. Four of the phantom configurations simulated treatment conditions with no scattering material beyond the point of measurement and the other four configurations simulated the homogeneous scattering conditions assumed by the TPS. Since the calibration TLDs for this work were irradiated with 137Cs and the experimental irradiations were performed with 192Ir, experiments were performed to determine the intrinsic energy dependence of the TLDs. Correction factors that relate the dose at the point of measurement (center of TLD) to the dose at the point of interest (basal skin layer) were also determined and applied for each irradiation geometry. RESULTS: The TLD intrinsic energy dependence for 192Ir relative to 137Cs was 1.041 +/- 1.78%. The TPS overestimated the exit skin dose by an average of 16% for the group of 59 patients studied, and by 9%-15% for the four phantom setups simulating treatment conditions. For the four phantom setups simulating the conditions assumed by the TPS, the TPS calculated dose agreed well with the TLD and MC results (within 3% and 1%, respectively). The inverse square geometry correction factor ranged from 1.023 to 1.042, and an additional correction factor of 0.978 was applied to account for the lack of charged particle equilibrium in the TLD and basal skin layer. CONCLUSIONS: TPS calculations that assume a homogeneous water medium overestimate the exit skin dose for intracavitary APBI treatments. It is important to determine the actual skin dose received during intracavitary APBI to determine the skin dose-response relationship and establish dose limits for optimal skin sparing. This study has demonstrated that TLDs can measure the skin dose with an expanded uncertainty (k = 2) of 5.6% when the proper corrections are applied.

Continuous 28-day iododeoxyuridine infusion and hyperfractionated accelerated radiotherapy for malignant glioma: a phase I clinical study.

PURPOSE: To investigate the maximal tolerated dose of a continuous 28-day iododeoxyuridine (IUdr) infusion combined with hyperfractionated accelerated radiotherapy (HART); to analyze the percentage of IUdr-thymidine replacement in peripheral granulocytes as a surrogate marker for IUdr incorporation into tumor cells; to measure the steady-state serum IUdr levels; and to assess the feasibility of continuous IUdr infusion and HART in the management of malignant glioma. METHODS AND MATERIALS: Patients were required to have biopsy-proven malignant glioma. Patients received 100 (n = 4), 200 (n = 3), 300 (n = 3), 400 (n = 6), 500 (n = 4), 625 (n = 5), or 781 (n = 6) mg/m(2)/d of IUdr by continuous infusion for 28 days. HART was started 7 days after IUdr initiation. The total dose was 70 Gy (1.2 Gy b.i.d. for 25 days with a 10-Gy boost [2.0 Gy for 5 Saturdays]). Weekly assays were performed to determine the percentage of IUdr-DNA replacement in granulocytes and serum IUdr levels using standard high performance liquid chromatography methods. Standard Phase I toxicity methods were used. RESULTS: Between June 1994 and August 1999, 31 patients were enrolled. No patient had Grade 3 or worse HART toxicity. Grade 3 or greater IUdr toxicity predominantly included neutropenia (n = 3), thrombocytopenia (n = 3), and elevated liver function studies (n = 3). The maximal tolerated dose was 625 mg/m(2)/d. Thymidine replacement in the peripheral granulocytes peaked at 3 weeks and increased with the dose (maximal thymidine replacement 4.9%). The steady-state plasma IUdr level increased with the dose (maximum, 1.5 microM). CONCLUSION: In our study, continuous long-term IUdr i.v. infusion had a maximal tolerated dose of 625 mg/m(2)/d. Granulocyte incorporation data verified the concept that prolonged IUdr infusion results in IUdr-DNA replacement that corresponds to a high degree of cell labeling. IUdr steady-state plasma levels increased with increasing dose and attained levels needed for clinical radiosensitization. Continuous IUdr infusion and HART were both feasible and well tolerated.

Overexpression of the R2 subunit of ribonucleotide reductase in human nasopharyngeal cancer cells reduces radiosensitivity.

PURPOSE: Ribonucleotide reductase is the rate-limiting enzyme in the de novo synthesis of deoxyribonucleotide triphosphates, which are utilized in both DNA synthesis and DNA repair. We reported previously that RR enzyme activity and R2 (catalytic subunit of RR) protein levels were increased after exposure to ionizing radiation (IR) in growth-arrested human tumor cells, suggesting that R2 protein expression regulates RR activity to allow for IR damage repair. Using isogenic human nasopharyngeal carcinoma cells in this study, we examine the relationship of overexpression of either the R1 regulatory subunit or the R2 catalytic subunit of RR to the cellular response of IR damage. MATERIALS AND METHODS: We used three isogenic human nasopharyngeal cancer cell lines previously derived by Zhou et al, including KB, the parental tumor cell line; KB/M1, an R1 protein-overexpressing clone stably transfected with human R1 complementary DNA; and KB/M2, a R2 protein-overexpressing clone stably transfected with human R2 complementary DNA. We initially characterized these isogenic human tumor cell lines in exponential growth for R2 protein expression, RR enzyme activity, and R2 protein changes during the cell cycle by flow cytometry. Subsequently, the IR response in these cell lines was determined by clonogenic survival, cell cycle changes occurring after IR, and an analysis of IR DNA damage determined by pulsed field gel electrophoresis. The effect of combining IR and hydroxyurea, a RR (R2) inhibitor, was also studied in KB and KB/M2 cells. RESULTS: KB/M2 cells were found to have 4.5-fold higher R2 protein expression and a threefold higher RR enzyme activity in exponential growth than KB and KB/M1. Although R2 protein levels increased at the G1/S transition in all cell lines, KB/M2 cells also demonstrated consistently higher R2 protein levels throughout the cell cycle. Using a linear-quadratic analysis of IR clonogenic survival data, KB/M2 cells were more radioresistant than KB and KB/M1 cells, including both decreased alpha and decreased beta values, a finding that correlates with increased reparable IR damage. KB/M2 cells also show a reduced G2 cell cycle arrest and fewer DNA double strand breaks 18 hours after IR (6 Gy). Exposure of KB/M2 cells to hydroxyurea (300 microM) after exposure to IR restored in vitro radiosensitivity in a manner similar to that found in KB and KB/M1 cells. DISCUSSION: An increase in R2 protein levels and RR activity in KB/M2 cells results in IR resistance, which appears mediated by enhanced IR damage repair during G2. R1 protein overexpression in these isogenic human tumor cells (KB/M1) did not affect RR activity or IR response.