A two-mutation model of radiation-induced acute myeloid leukemia using historical mouse data.

From studies of the atomic bomb survivors, it is well known that ionizing radiation causes several forms of leukemia. However, since the specific mechanism behind this process remains largely unknown, it is difficult to extrapolate carcinogenic effects at acute high-dose exposures to risk estimates for the chronic low-dose exposures that are important for radiation protection purposes. Recently, it has become clear that the induction of acute myeloid leukemia (AML) in CBA/H mice takes place through two key steps, both involving the Sfpi1 gene. A similar mechanism may play a role in human radiation-induced AML. In the present paper, a two-mutation carcinogenesis model is applied to model AML in several data sets of X-ray- and neutron-exposed CBA/H mice. The models obtained provide good fits to the data. A comparison between the predictions for neutron-induced and X-ray-induced AML yields an RBE for neutrons of approximately 3. The model used is considered to be a first step toward a model for human radiation-induced AML, which could be used to estimate risks of exposure to low doses.

Biodistribution of (10)B for Boron Neutron Capture Therapy (BNCT) in a mouse model after injection of sodium mercaptoundecahydro-closo-dodecaborate and l-para-boronophenylalanine.

In boron neutron capture therapy, the absorbed dose from the (10)B(n,alpha)(7)Li reaction depends on the (10)B concentration and (10)B distribution in the irradiated volume. Thus compounds used in BNCT should have tumor-specific uptake and low accumulation in normal tissues. This study compares in a mouse model the (10)B uptake in different organs as delivered by l-para-boronophenylalanine (BPA, 700 mg/kg body weight, i.p.) and/or sodium mercaptoundecahydro-closo-dodecaborate (BSH, 200 mg/kg body weight, i.p). After BSH injection, the (10)B concentration was high in kidneys (20 +/- 12 microg/g) and liver (20 +/- 12 microg/g) but was low in brain (1.0 +/- 0.8 microg/g) and muscle (1.9 +/- 1.2 microg/g). After BPA injection, the (10)B concentration was high in kidneys (38 +/- 25 microg/g) and spleen (17 +/- 8 microg/g) but low in brain (5 +/- 3 microg/g). After combined BPA and BSH injection, the effect on the absolute (10)B concentration was additive in all organs. The ratio of the (10)B concentrations in tissues and blood differed significantly for the two compounds depending on the compound combination, which implies a different uptake profile for normal organs.

Uptake of two 10B-compounds in liver metastases of colorectal adenocarcinoma for extracorporeal irradiation with boron neutron capture therapy (EORTC Trial 11001).

Disseminated metastases of colorectal cancer in liver are incurable. The trial EORTC 11001 investigates whether autotransplantation after extracorporeal irradiation of the liver by boron neutron capture therapy (BNCT) might become a curative treatment option because of selective uptake of the compounds sodium mercaptoundecahydro-closo-dodecaborate (BSH) or L-para-boronophenylalanine (BPA). BSH (50 mg/kg bw) or BPA (100 mg/kg bw) were infused into patients who subsequently underwent resection of hepatic metastases. Blood and tissue samples were analyzed forthe (10)B-concentration with prompt gamma ray spectroscopy (PGRS). Three patients received BSH and 3 received BPA. Adverse effects from the boron carriers did not occur. For BSH, the highest (10)B-concentration was observed in liver (31.5 +/- 2.7 microg/g) followed by blood (24.8 +/- 4.7 microg/g) and tumor (23.2 +/- 2.1 microg/g) with a mean (10)B-concentration ratio metastasis/liver of 0.72 +/- 0.07. For BPA, the highest (10)B-concentration was measured in metastases (12.1 +/- 2.2 microg/g) followed by liver (8.5 +/- 0.5 microg/g) and blood (5.8 +/- 0.8 microg/g). As BPA is transported actively into cells, viable, metabolically active cells accumulate exclusively this compound. Consequently, a model is proposed to adjust the values measured by PGRS for the proportion of viable cells to express the relevant (10)B-concentration in the tumor cells, revealing a (10)B-concentration ratio metastasis/liver of 6.8 +/- 1.7. In conclusion, BSH is not suitable as (10)B-carrier in liver metastases as the (10)B-concentration in liver was higher compared to metastasis. BPA accumulates in hepatic metastases to an extent that allows for extracorporeal irradiation of the liver with BNCT.

Boron nanoparticles inhibit tumour growth by boron neutron capture therapy in the murine B16-OVA model.

BACKGROUND: Boron neutron capture therapy usually relies on soluble, rather than particulate, boron compounds. This study evaluated the use of a novel boron nanoparticle for boron neutron capture therapy. MATERIALS AND METHODS: Two hundred and fifty thousand B16-OVA tumour cells, pre-incubated with boron nanoparticles for 12 hours, were injected subcutaneously into C57BL/6J mice. The tumour sites were exposed to different doses of neutron radiation one, four, or eight days after tumour cell inoculation. RESULTS: When the tumour site was irradiated with thermal neutrons one day after injection, tumour growth was delayed and the treated mice survived longer than untreated controls (median survival time 20 days (N = 8) compared with 10 days (N = 7) for untreated mice). CONCLUSION: Boron nanoparticles significantly delay the growth of an aggressive B16-OVA tumour in vivo by boron neutron capture therapy.

Mutations of the PU.1 Ets domain are specifically associated with murine radiation-induced, but not human therapy-related, acute myeloid leukaemia.

Murine radiation-induced acute myeloid leukaemia (AML) is characterized by loss of one copy of chromosome 2. Previously, we positioned the critical haematopoietic-specific transcription factor PU.1 within a minimally deleted region. We now report a high frequency (>65%) of missense mutation at codon 235 in the DNA-binding Ets domain of PU.1 in murine AML. Earlier studies, outside the context of malignancy, determined that conversion of arginine 235 (R235) to any other amino-acid residue leads to ablation of DNA-binding function and loss of expression of downstream targets. We show that mutation of R235 does not lead to protein loss, and occurs specifically in those AMLs showing loss of one copy of PU.1 (P=0.001, Fisher's exact test). PU.1 mutations were not found in the coding region, UTRs or promoter of human therapy-related AMLs. Potentially regulatory elements upstream of PU.1 were located but no mutations found. In conclusion, we have identified the cause of murine radiation-induced AML and have shown that loss of one copy of PU.1, as a consequence of flanking radiation-sensitive fragile domains on chromosome 2, and subsequent R235 conversion are highly specific to this mouse model. Such a mechanism does not operate, or is extremely rare, in human AML.

Pre-exposure to low doses: modulation of X-ray-induced dna damage and repair?

The adaptive response to ionizing radiation may be mediated by the induction of antioxidant defense mechanisms, accelerated repair or altered cell cycle progression after the conditioning dose. To gain new insight into the mechanism of the adaptive response, nondividing lymphocytes and fibroblasts were used to eliminate possible contributions of cell cycle effects. The effect of conditioning doses of 0.05 or 0.1 Gy followed by challenging doses up to 8 Gy (with a 4-h interval between exposures) on induction and repair of DNA damage was determined by single-cell gel electrophoresis (comet assay), premature chromosome condensation, and immunofluorescence labeling for gamma-H2AX. The conditioning dose reduced the induction of DNA strand breaks, but the kinetics of strand break rejoining was not influenced by the conditioning dose in nondividing cells of either cell type. We conclude that adaptation in nondividing cells is not mediated by enhanced strand break rejoining and that protection against the induction of DNA damage is rather small. Therefore, the adaptive response is most likely a reflection of perturbation of cell cycle progression.

Dose planning with comparison to in vivo dosimetry for epithermal neutron irradiation of the dog brain.

Boron neutron capture therapy (BNCT) is an experimental type of radiotherapy, presently being used to treat glioblastoma and melanoma. To improve patient safety and to determine the radiobiological characteristics of the epithermal neutron beam of Finnish BNCT facility (FiR 1) dose-response studies were carried on the brain of dogs before starting the clinical trials. A dose planning procedure was developed and uncertainties of the epithermal neutron-induced doses were estimated. The accuracy of the method of computing physical doses was assessed by comparing with in vivo dosimetry. Individual radiation dose plans were computed using magnetic resonance images of the heads of 15 Beagle dogs and the computational model of the FiR 1 epithermal neutron beam. For in vivo dosimetry, the thermal neutron fluences were measured using Mn activation foils and the gamma-ray doses with MCP-7s type thermoluminescent detectors placed both on the skin surface of the head and in the oral cavity. The degree of uncertainty of the reference doses at the thermal neutron maximum was estimated using a dose-planning program. The estimated uncertainty (+/-1 standard deviation) in the total physical reference dose was +/-8.9%. The calculated and the measured dose values agreed within the uncertainties at the point of beam entry. The conclusion is that the dose delivery to the tissue can be verified in a practical and reliable fashion by placing an activation dosimeter and a TL detector at the beam entry point on the skin surface with homogeneous tissues below. However, the point doses cannot be calculated correctly in the inhomogeneous area near air cavities of the head model with this type of dose-planning program. This calls for attention in dose planning in human clinical trials in the corresponding areas.

Intestinal tumours induced in Apc(Min/+) mice by X-rays and neutrons.

PURPOSE: To compare the development of intestinal adenomas following neutron and X-ray exposure of Apc(Min/+) mice (Apc - adenomatous polyposis coli; Min - multiple intestinal neoplasia). MATERIALS AND METHODS: Adult mice were exposed to acute doses of X-rays or fission neutrons. Tumour counting was undertaken 200 days later and samples were taken for Loss of Heterozygosity (LOH) analysis. RESULTS: Tumour numbers (adenomas and microadenomas) increased by 1.4-fold, 1.7-fold, 2.7-fold and 9-fold, after 0.5, 1, 2 and 5 Gy X-rays, respectively, and by 2.4-fold and 5.7-fold, after 0.5 and 1 Gy fission neutrons, respectively. LOH analysis of tumours from neutron-exposed mice showed that 63% had lost Apc and 90% (cf. 53% in controls) had lost D18mit84, a marker for Epb4.1l4a/NBL4 (erythrocyte protein band 4.1-like 4a/novel band 4.1-like 4), known to be involved in the Wnt (wingless-related mouse mammary tumour virus integration site) pathway. Some tumours from neutron-exposed mice appeared to have homozygous loss of some chromosomal markers. CONCLUSIONS: X-ray or fission neutron irradiation results in strongly enhanced tumour multiplicities. Comparison of tumour yields indicated a low Relative Biological Effectiveness of around 2-8 for fission neutrons compared with X-rays. LOH in intestinal tumours from neutron-exposed mice appeared to be more complex than previously reported for tumours from X-irradiated mice.

Microsatellite instability in radiation-induced murine tumours; influence of tumour type and radiation quality.

PURPOSE: To investigate microsatellite instability (MSI) in radiation-induced murine tumours, its dependence on tissue (haemopoietic, intestinal, mammary, brain and skin) and radiation type. MATERIALS AND METHODS: DNA from spontaneous, X-ray or neutron-induced mouse tumours were used in Polymerase Chain Reactions (PCR) with mono- or di-nucleotide repeat markers. Deviations from expected allele size caused by insertion/deletion events were assessed by capillary electrophoresis. RESULTS: Tumours showing MSI increased from 16% in spontaneously arising tumours to 23% (P = 0.014) in X-ray-induced tumours and rising again to 83% (P << 0.001) in neutron-induced tumours. X-ray-induced Acute Myeloid Leukaemias (AML) had a higher level of mono-nucleotide instability (45%) than di-nucleotide instability (37%). Fifty percent of neutron-induced tumours were classified as MSI-high for mono-nucleotide markers and 10% for di-nucleotide markers. Distribution of MSI varied in the different tumour types and did not appear random. CONCLUSIONS: Exposure to ionising radiation, especially neutrons, promotes the development of MSI in mouse tumours. MSI may therefore play a role in mouse radiation tumourigenesis, particularly following high Linear Energy Transfer (LET) exposures. MSI events, for a comparable panel of genome-wide markers in different tissue types, were not randomly distributed throughout the genome.

Tissue uptake of BSH in patients with glioblastoma in the EORTC 11961 phase I BNCT trial.

PURPOSE: The uptake of the boron compound Na2B12H10-SH (BSH) in tumor and normal tissues was investigated in the frame of the EORTC phase I trial 'Postoperative treatment of glioblastoma with BNCT at the Petten Irradiation Facility' (protocol 11961). METHODS AND MATERIALS: The boron concentration in blood, tumor, normal brain, dura, muscle, skin and bone was detected using inductively coupled plasma-atomic emission spectroscopy in 13 evaluable patients. In a first group of 10 patients 100 mg BSH/kg bodyweight (BW) were administered; a second group of 3 patients received 22.9 mg BSH/kg BW. The toxicity due to BSH was evaluated. RESULTS: The average boron concentration in the tumor was 19.9 +/- 9.1 ppm (1 standard deviation (SD)) in the high dose group and 9.8 +/- 3.3 ppm in the low dose group, the tumor/blood ratios were 0.6 +/- 0.2 and 0.9 +/- 0.2, respectively. The highest boron uptake has been detected in the dura, very low uptake was found in the bone, the cerebro-spinal fluid and especially in the brain (brain/blood ratio 0.2 +/- 0.02 and 0.4 +/- 0.2). No toxicity was detected except flush-like symptoms in 2 cases during a BSH infusion at a much higher speed than prescribed. CONCLUSION: BSH proved to be safe for clinical application at a dose of 100 mg BSH/kg infused and at a dose rate of 1 mg/kg/min. The study underlines the importance of a further investigation of BSH uptake in order to obtain enough data for significant statistical analysis. The boron concentration in blood seems to be a quite reliable parameter to predict the boron concentration in other tissues.