Abstract Introduction The magnitude of intra‐ and inter‐fractional variation in the set up of breast cancer patients treated with tangential megavoltage photon beams was investigated using an electronic portal imaging device (EPID). Methods Daily cine‐EPID images were captured during delivery of the tangential fields for ten breast cancer patients treated in the supine position. Measurements collected from each image included the central lung distance (CLD), central flash distance (CFD), superior axial measurement (SAM) and the inferior axial measurement (IAM). The variation of motion within a fraction (intra‐fraction) and the variation between fractions (inter‐fraction) was analysed to quantify set up variation and motion due to respiration. Results Altogether 3775 EPID images were collected from 10 patients. The effect of respiratory motion during treatment was <0.1 cm standard deviation (SD) in the anterior–posterior (AP) direction. The inter‐fraction movement caused by variations in daily set up was larger at 0.28 cm SD in the AP direction. Superior–inferior (SI) variation was more difficult to summarise and proved unreliable as the measurements were taken to an ambiguous point on the images. It was difficult to discern true SI movement from that implicated by AP movement. Conclusion There is minimal intra‐fractional chest wall motion due to respiration during treatment. Inter‐fractional variation was larger, however, on average it remained within departmental tolerance (0.5 cm) for set up variations. This review of our current breast technique provides confidence in the feasibility of utilising advanced treatment techniques (field‐in‐field, intensity modulated radiotherapy or volumetric modulated arc therapy) following a review of the current imaging protocol.

BACKGROUND:The sap from Euphorbia peplus, commonly known as petty spurge in the U.K. or radium weed in Australia, has been used as a traditional treatment for a number of cancers. OBJECTIVE:To determine the effectiveness of E. peplus sap in a phase I/II clinical study for the topical treatment of basal cell carcinomas (BCC), squamous cell carcinomas (SCC) and intraepidermal carcinomas (IEC). METHODS:Thirty-six patients, who had refused, failed or were unsuitable for conventional treatment, were enrolled in a phase I/II clinical study. A total of 48 skin cancer lesions were treated topically with 100-300 μL of E. peplus sap once daily for 3 days. RESULTS:The complete clinical response rates at 1 month were 82% (n = 28) for BCC, 94% (n = 16) for IEC and 75% (n = 4) for SCC. After a mean follow-up of 15 months these rates were 57%, 75% and 50%, respectively. For superficial lesions < 16 mm, the response rates after follow-up were 100% for IEC (n = 10) and 78% for BCC (n = 9). CONCLUSIONS:The clinical responses for these relatively unfavourable lesions (43% had failed previous treatments, 35% were situated in the head and neck region and 30% were > 2 cm in diameter), are comparable with existing nonsurgical treatments. An active ingredient of E. peplus sap has been identified as ingenol mebutate (PEP005). This clinical study affirms community experience with E. peplus sap, and supports further clinical development of PEP005 for the treatment of BCC, SCC and IEC.

The aim was to compare a private Commonwealth-initiated regional radiation oncology facility in Toowoomba with a Queensland Health facility (QHF) in Brisbane. The comparison concentrated on staffing, case mix and operational budgets, but was not able to look at changes in access to services. Data were collected from the two facilities from January 2008 to June 2008 inclusive. A number of factors were compared, including case mix, staffing levels, delay times for treatment, research, training and treatment costs. The case mix between the two areas was similar with curative treatments making up just over half the work load in both centres and two-thirds the work being made up of cancers of breast and prostate. Staffing levels were leaner in Toowoomba, especially in the areas of nursing, administration and trial coordinators. Research activity was slightly higher in Toowoomba. The average medicare cost per treatment course was similar in both centres ($5000 per course). Total costs of an average treatment including patient, State and Commonwealth costs, showed a 30% difference in costing favouring Toowoomba. This regional radiation oncology centre has provided state-of-the-art cancer care that is close to home for patients living in the Darling Downs region. Both public and private patients have been treated with modest costs to the patient and significant savings to QH. The case mix is similar to the QHF, and there has been significant activity in clinical research. A paperless working environment is one factor that has allowed staffing levels to be reduced. Ongoing support from Governments are required if private facilities are to participate in important ongoing staff training.

Aims: To describe the toxicity and response seen in patients receiving moderate-dose radiation therapy with concurrent weekly low-dose gemcitabine in the management of locally advanced non-small cell lung cancer (NSCLC).Materials and methods: Eighteen patients with confirmed NSCLC were enrolled over a 17-month period from August 2000 until January 2002. All had localised disease but were considered unsuitable for curative therapy. Radiation therapy was given to a dose of 30 Gy in 15 fractions over 3 weeks. Gemcitabine was given weekly before and within 3 h of fractions 1, 6 and 11. The study was designed as a dose-escalation study, commencing at 100 mg/m(2) and increasing at levels of 50 mg/m(2), until the maximum tolerated dose (MTD) was reached.Results: The MTD was regarded as being 150 mg/m(2). The major acute toxicity observed was oesophagitis. Skin reactions were also reported. The overall response rate in all patients was 88%, with 44% achieving a complete response.Conclusion: The combination of gemcitabine and moderate-dose radiation therapy is feasible, and offers low toxicity and excellent response rates in patients with localised NSCLC not suitable for high-dose therapy. (c) 2005 The Royal College of Radiologists. Published by Elsevier Ltd. All rights reserved.

Ataxia-telangiectasia Mutated (ATM), mutated in the human disorder ataxia-telangiectasia, is rapidly activated by DNA double strand breaks. The mechanism of activation remains unresolved, and it is uncertain whether autophosphorylation contributes to activation. We describe an in vitro immunoprecipitation system demonstrating activation of ATM kinase from unirradiated extracts by preincubation with ATP. Activation is both time- and ATP concentration-dependent, other nucleotides fail to activate ATM, and DNA is not required. ATP activation is specific for ATM since it is not observed with kinase-dead ATM, it requires Mn2+, and it is inhibited by wortmannin. Exposure of activated ATM to phosphatase abrogates activity, and repeat cycles of ATP and phosphatase treatment reveal a requirement for autophosphorylation in the activation process. Phosphopeptide mapping revealed similarities between the patterns of autophosphorylation for irradiated and ATP-treated ATM. Caffeine inhibited ATM kinase activity for substrates but did not interfere with ATM autophosphorylation. ATP failed to activate either A-T and rad3-related protein (ATR) or DNA-dependent protein kinase under these conditions, supporting the specificity for ATM. These data demonstrate that ATP can specifically induce activation of ATM by a mechanism involving autophosphorylation. The relationship of this activation to DNA damage activation remains unclear but represents a useful model for understanding in vivo activation.

Ataxia-telangiectasia is characterized by radiosensitivity, genome instability and predisposition to cancer1,2. Heterozygous carriers of ATM, the gene defective in ataxia-telangiectasia, have a higher than normal risk of developing breast and other cancers3,4,5,6. We demonstrate here that Atm 'knock-in' (Atm-ΔSRI) heterozygous mice harboring an in-frame deletion corresponding to the human 7636del9 mutation show an increased susceptibility to developing tumors. In contrast, no tumors are observed in Atm knockout (Atm+/−) heterozygous mice. In parallel, we report the appearance of tumors in 6 humans from 12 families who are heterozygous for the 7636del9 mutation. Expression of ATM cDNA containing the 7636del9 mutation had a dominant-negative effect in control cells, inhibiting radiation-induced ATM kinase activity in vivo and in vitro. This reduces the survival of these cells after radiation exposure and enhances the level of radiation-induced chromosomal aberrations. These results show for the first time that mouse carriers of a mutated Atm that are capable of expressing Atm have a higher risk of cancer. This finding provides further support for cancer predisposition in human ataxia-telangiectasia carriers.

Mutations in ATM, the gene defective in the human genetic disorder ataxia‐telangiectasia (A‐T), have been described in A‐T patients and in a variety of tumor samples. Most of these arise due to exon skipping. We developed an RT‐PCR based protein truncation test (PTT) to screen for ATM mutations in breast cancer patients showing adverse response to radiotherapy. An additional PTT product was evident in the ATM gene in peripheral blood mononuclear cells (PBMCs) from blood samples that were collected 2 days or more prior to RNA extraction. Lymphoblastoid cell lines established from the same blood samples showed no evidence of the additional band. Cloning and sequencing of the additional RT‐PCR product revealed an exact deletion of exon 20 (2639 del 200), pointing to exon skipping. RT‐PCR analysis of RNA extracted from freshly prepared PBMCs from 3 normal individuals showed no evidence of the additional RT‐PCR product but when the blood was stored for 2‐3 days prior to RNA extraction the lower molecular weight band was evident in every sample. DNA sequencing confirmed this to be due to loss of exon 20. These data suggest that mRNA‐based mutation analysis on ATM should be carried out on unstored blood samples to avoid artificial loss of exons that give rise to apparent mutations. © 2001 Wiley‐Liss, Inc.

Purpose: The dose intensity of radiotherapy (RT) used in cancer treatment is limited in rare individuals who display severe normal tissue reactions after standard RT treatments. Novel predictive assays are required to identify these individuals prior to treatment. The mechanisms responsible for such reactions are unknown, but may involve dysfunction of genes involved in the sensing and response of cells to DNA damage. The breast cancer susceptibility genes BRCA1 and BRCA2 are implicated in DNA damage repair and the control of genome stability, The purpose of this study was to determine if clinical radiation hypersensitivity is related to mutations of the BRCA1 and BRCA2 genes. Such information is of potential use in the clinical management of BRCA mutation carriers and their families,Methods and Materials: Twenty-two cancer patients who developed severe normal tissue reactions after RT were screened for mutations of BRCA1 and BRCA2, using various methods including protein truncation testing, direct DNA sequencing, and a PCR-based BRCA1 exon 13 duplication test.Results: No mutations were detected in the 22 patients tested, despite screening for the majority of commonly described types of mutations of BRCA1 and BRCA2.Conclusion: These early results suggest that genes other than BRCA1 and BRCA2 probably account for most cases of clinical radiation hypersensitivity, and that screening for mutations of BRCA1 and BRCA2 is unlikely to be useful in predicting response to radiotherapy. However, it has not been excluded that some BRCA1 or BRCA2 heterozygotes might experience unexpected RT toxicity; further BRCA mutation screening on radiation sensitive individuals is warranted, (C) 2000 Elsevier Science Inc.

Background and purpose: To determine if clinical radiosensitivity in breast cancer patients is related to mutations of the ataxia telangiectasia gene (ATM). Materials and methods: Fifteen patients who had developed a severe late reaction to a standard radiotherapy schedule were examined for evidence of increased in vitro radiosensitivity using the MTT assay. Mutation analysis was performed using a protein truncation assay. Results: No mutations were detected in the 15 patients despite evidence of increased in vitro radiosensitivity. Conclusions: Testing for the ATM gene is unlikely to be useful for predicting clinical response to radiotherapy in breast cancer patients.

The human melanoma cell lines MM96L, A2058 and HT144 were examined for sensitivity to ionizing radiation and UVB radiation. HT144 demonstrated a significant increase in sensitivity to ionizing and UVB radiation compared with the MM96L and A2058 cells. Sensitivity to both agents was associated with susceptibility to apoptosis. Using a protein truncation assay, a mutation for the gene for ataxia telangiectasia (ATM) was identified in HT144 cells. This was confirmed to be a homozygous mutation by subsequent sequencing of the abnormal region. Protein truncation assay of the other two cell lines showed no abnormality. The results suggest that somatic mutation of the A-T gene may be important in determining tumour radiosensitivity.

Neurofibromatosis type 1 (NF 1) is associated with certain types of malignancy including childhood cancers, neuroendocrine tumours and sarcomas. An association with breast cancer has been suggested but few cases have beer reported. We report six cases of breast cancer occurring in women less than 60 years of age with NF1, most of whom hac locally advanced and poorly differentiated breast cancer on presentation. Karyotypic analysis revealed no abnormalities it these patients, including chromosome 17q where both the NF1 and BRCA1 genes are located. Since radiosensitivity has been linked to cancer predisposition in breast and some other cancer types, we examined clinical and in vitro radiosensitivity in the six patients. Three of the women showed increased in vitro radiosensitivity and two of four that received radiotherapy showed an adverse clinical response. Analysis of the ataxia-telangiectasia gene in these individuals by the proteir truncation test showed that none were likely to be mutation carriers.

The recently cloned gene (ATM) mutated in the human genetic disorder ataxia-telangiectasia (A-T) is involved in DNA damage response at different cell cycle checkpoints and also appears to have a wider role in signal transduction. Antibodies prepared against peptides from the predicted protein sequence detected a ∼ 350 kDa protein corresponding to the open reading frame, which was absent in 13/23 A-T homozygotes. Subcellular fractionation, immunoelectronmicroscopy and immunofluorescence showed that the ATM protein is present in the nucleus and cytoplasmic vesicles. This distribution did not change after irradiation. We also provide evidence that ATM protein binds to p53 and this association is defective in A-T cells compatible with the defective p53 response in these cells. These results provide further support for a role for the ATM protein as a sensor of DNA damage and in a more general role in cell signalling, compatible with the broader phenotype of the syndrome.

Exposure of human cells to gamma-radiation causes levels of the tumour-suppressor nuclear protein p53 to increase in temporal association with the decrease in replicative DNA synthesis. Cells from patients with the radiosensitive and cancer-prone disease ataxia telangiectasia (AT) exhibit radioresistant DNA synthesis and show a reduced or delayed gamma-radiation-induced increase in p53 protein levels. We have used Western immunoblotting with semiquantitative densitometry to examine the gamma-radiation-induced levels of p53 protein in 57 lymphoblastoid cell lines (LCLs) derived from patients with AT, carriers of the AT gene, breast cancer patients and normal donors. We confirm the previously reported reduced induction in AT homozygote LCLs (n = 8) compared with normal donor LCLs (n = 17, P = 0.01). We report that AT heterozygote LCLs (n = 5) also have a significantly reduced p53 induction when compared with LCLs from normal donors (n = 17, P = 0.02). The response of breast cancer patient cells was not significantly different from normal donor cells but 18% (5/27) had a p53 response in the AT heterozygote range (95% confidence interval) compared with only 6% (1/17) of the normal donor cells. We found no significant correlation between p53 induction and cellular radiosensitivity in LCLs from breast cancer patients. These methods may be useful in identifying individuals at greater risk of the DNA-damaging effects of ionising radiation.

This review was initiated to assess the outcome of treatment with radical radiotherapy with curative intent for patients diagnosed as having stage I non-small cell lung cancer. The study involved a retrospective review of 347 patients with T1 and T2N0M0 tumors treated at the Queensland Radium Institute during the period 1985 to 1992. The main reasons for not proceeding to surgery included poor performance status, old age, or refusal to submit to surgery. The median age for the group was 70 years, with the range being 34 to 90 years. Patients in this group were all treated by a standard technique involving external-beam radiotherapy to 50 Gy, minimum tumor dose, in 20 fractions over 4 weeks. The overall survival rate was 27% at 5 years with a median survival of 27.9 months. The 5-year recurrence-free survival was 23% with the median being 19.5 months. There was a strong correlation of survival to tumor size with 5-year survival rates being 32% and 21% for T1 and T2 tumors, respectively. Multivariate analysis found only T stage to be associated with overall survival (p < 0.01). In addition, the analysis showed that age younger than 70 years was a prognostic factor that approached statistical significance at the p < 0.05 level of significance. We conclude from this large series of patients with stage I non-small cell lung cancer that radical radiotherapy with curative intent may be a viable alternative to surgery in those patients who either refuse surgery or are judged to be unfit for operation.

Purpose: To investigate whether in vitro radiosensitivity of lymphocytes derived from a blood sample will predict late effects from radiotherapy in breast cancer patients.Methods and Materials: Blood samples were collect-ed from consenting patients who had received radiotherapy for breast cancer, Lymphocytes were extracted and transformed by the Epstein-Barr virus, The resulting lymphoblastoid cell lines (LCLs) were assessed for in vitro radiosensitivity using a tetrazolium-based colorimetric assay (MTT), Survival curves were generated using doses of 0.5 to 2 Gy, For each analysis an LCL derived from an individual with the radiosensitive condition ataxia-telangiectasia (AT) was run as control. Some patients also consented to give skin biopsies from which fibroblast cultures were derived, Clonogenic assays were performed to generate survical curves using doses in the range of 0.5 to 4 Gy, Comparison was made with the data obtained from LCLs. Late effects of radiotherapy were assessed using the Radiation Therapy Oncology Group (RTOG) scoring scheme and compared with the in vitro radiosensitivity data.Results: LCLs from 56 breast cancer patients were assessed for in vitro radiosensitivity, Surviving fraction to 2 Gy (SF2) generated from survical curves ranged from 0.04-0.35 with coefficient of variation for the whole group of 41%, None of the LCLs equalled the sensitivity of the AT line, but 16% showed equal or greater sensitivity to a line derived from an AT heterozygote. Comparison of LCL and fibroblast radiosensitivity showed reasonable correlation for 12 paired samples (r = 0.64), The majority of patients showed no or minimal effects after radiotherapy (Grade 0,1 effects) but seven developed a Grade 2 reaction and four a Grade 3 or 4 reaction, Patients with a Grade 2-4 reaction were found to be more sensitive in vitro than those with a Grade 0-1 reaction (p < 0.02).Conclusions: The use of the MTT assay to assess LCL radiosensitivity has demonstrated considerable heterogeneity amongst the breast cancer population, The presence of a proportion of patients showing in vitro sensitivity but normal clinical response to radiotherapy would limit the usefulness of the assay for predictive purposes.