Is it Time to Change Radiotherapy: The Dawning of Chronoradiotherapy?

A minority of radiotherapy patients experience adverse reactions as a result of the inevitable irradiation of the surrounding healthy tissue. These reactions range in severity and affect the patient's quality of life, as well as being dose-limiting. If the patients most at risk of toxicity could be identified before radiotherapy, the treatment pathway, radiation dose or fractionation could be altered to reduce toxicity while maintaining efficacy. Previous research is described on how chemotherapy treatments could be improved through the delivery of drugs at specific times of the day ('chronomodulation') based on the circadian rhythm. More recently time-of-day effects have been investigated for radiotherapy, yielding complex results, but with some promise for genetic prediction of the optimal time for treatment. This would allow an almost cost-free modification to treatment that would reduce toxicity. Despite the increasing evidence for 'chronotherapy' for treating cancer, little work has looked into the potential mechanisms underlying the time-of-day effect, which potentially include differences in inflammation, cell cycle or hormones. This overview discusses the main findings from chronotherapy so far and comments on why elucidating the biological mechanisms relating radiotherapy toxicity to the circadian cycle warrants further investigation.

Genetic Variants Predict Optimal Timing of Radiotherapy to Reduce Side-effects in Breast Cancer Patients.

AIMS: Radiotherapy is an important treatment for many types of cancer, but a minority of patients suffer long-term side-effects of treatment. Multiple lines of evidence suggest a role for circadian rhythm in the development of radiotherapy late side-effects. MATERIALS AND METHODS: We carried out a study to examine the effect of radiotherapy timing in two breast cancer patient cohorts. The retrospective LeND cohort comprised 535 patients scored for late effects using the Late Effects of Normal Tissue-Subjective Objective Management Analytical (LENT-SOMA) scale. Acute effects were assessed prospectively in 343 patients from the REQUITE study using the CTCAE v4 scales. Genotyping was carried out for candidate circadian rhythm variants. RESULTS: In the LeND cohort, patients who had radiotherapy in the morning had a significantly increased incidence of late toxicity in univariate (P = 0.03) and multivariate analysis (P = 0.01). Acute effects in the REQUITE group were also significantly increased in univariate analysis after morning treatment (P = 0.03) but not on multivariate analysis. Increased late effects in the LeND group receiving morning radiotherapy were associated with carriage of the PER3 variable number tandem repeat 4/4 genotype (P = 6 × 10-3) and the NOCT rs131116075 AA genotype (P = 5 × 10-3). CONCLUSION: Our results suggest that it may be possible to reduce toxicity associated with breast cancer radiotherapy by identifying gene variants that affect circadian rhythm and scheduling for appropriate morning or afternoon radiotherapy.

The Patient Perspective on Radiogenomics Testing for Breast Radiation Toxicity.

AIMS: In the field of radiogenomics, several potential predictive genetic markers have been identified that are associated with individual susceptibility to radiation toxicity. Predictive models of radiation toxicity incorporating radiogenomics and other biomarkers are being developed as part of the ongoing multicentre REQUITE trial. The purpose of this study was to explore patient attitudes towards future predictive radiogenomics testing for breast radiation toxicity. PATIENTS AND METHODS: Twenty-one semi-structured interviews were conducted with breast cancer patients taking part in the REQUITE study at one centre. We used inductive thematic analysis to generate common themes. RESULTS: We identified three emerging themes describing attitudes and feelings towards a predictive radiogenomics test for breast radiation toxicity: theme 1 - willingness to undergo a test (subthemes - information, trusted expert); theme 2 - implications of a test (subthemes - preparation and planning, anxiety without recourse); theme 3 - impact on treatment decision-making (subthemes - prioritising cancer cure, preserving breast integrity, patient preferences). CONCLUSIONS: Results from the present study indicate that patients support and have confidence in the validity of a radiogenomics test for breast radiation toxicity, but they would prefer the result be provided to healthcare professionals. Except in cases of significant chronic symptoms and pain or significant end-organ damage, participants in this study rarely felt that advance knowledge of their personal risk of breast radiation toxicity would influence their treatment decision-making. These findings provide a number of insights that will allow us to anticipate how patients are likely to engage with predictive radiogenomics testing in the future.

Finding the genetic determinants of adverse reactions to radiotherapy.

Individual variation in radiosensitivity is thought to be at least partly determined by genetic factors. The remaining difference between individuals is caused by comorbidities, variation in treatment, body habitus and stochastic factors. Evidence for the heritability of radiosensitivity comes from rare genetic disorders and from cell-based studies. To what extent common and rare genetic variants might explain the genetic component of radiosensitivity has not been fully elucidated. If the genetic variants accounting for this heritability were to be determined, they could be incorporated into any future predictive statistical model of adverse reactions to radiotherapy. With the evolution of DNA sequencing and bioinformatics, radiogenomics has emerged as a new research field with the aim of finding the genetic determinants of adverse reactions to radiotherapy. Similar to the investigation of other complex genetic disease traits, early studies in radiogenomics involved candidate gene association studies--many plagued by false associations caused by low sample sizes and problematic experimental design. More recently, some promising genetic associations (e.g. with tumour necrosis factor) have emerged from large multi-institutional cohorts with built-in replication. At the same time, several small- to medium-sized genome-wide association studies (GWAS) have been or are about to be published. These studies will probably lead to an increasing number of genetic polymorphisms that may predict adverse reactions to radiotherapy. The future of the field is to create large patient cohorts for multiple cancer types, to validate the genetic loci and build reliable predictive models. For example, the REQUITE project involves multiple groups in Europe and North America. For further discovery studies, larger GWAS will be necessary to include rare sequence variants through next generation sequencing. Ultimately, radiogenomics seeks to predict which cancer patients will show radiosensitivity or radioresistance, so oncologists and surgeons can alter treatment accordingly to lower adverse reactions or increase the efficacy of radiotherapy.

A replicated association between polymorphisms near TNFα and risk for adverse reactions to radiotherapy.

BACKGROUND: Response to radiotherapy varies between individuals both in terms of efficacy and adverse reactions. Finding genetic determinants of radiation response would allow the tailoring of the treatment, either by altering the radiation dose or by surgery. Despite a growing number of studies in radiogenomics, there are no well-replicated genetic association results. METHODS: We carried out a candidate gene association study and replicated the result using three additional large cohorts, a total of 2036 women scored for adverse reactions to radiotherapy for breast cancer. RESULTS: Genetic variation near the tumour necrosis factor alpha gene is shown to affect several clinical endpoints including breast induration, telangiectasia and overall toxicity. In the combined analysis homozygosity for the rare allele increases overall toxicity (P=0.001) and chance of being in the upper quartile of risk with odds ratio of 2.46 (95% confidence interval 1.52-3.98). CONCLUSION: We have identified that alleles of the class III major histocompatibility complex region associate with overall radiotherapy toxicity in breast cancer patients by using internal replication through a staged design. This is the first well-replicated report of a genetic predictor for radiotherapy reactions.

Death-associated protein kinase (DAPK1) in cerebral cortex of late-onset Alzheimer's disease patients and aged controls.

AIMS: Here our objective was to detect the pro-apoptotic serine/threonine kinase death-associated protein kinase (DAPK1) in aged human cerebral cortex and to test the hypothesis that DAPK1 abundance is associated with late-onset Alzheimer's disease (AD). METHODS: Using Western analysis and immunohistochemistry we evaluated post mortem frontal cerebral cortex from patients with severe AD (mean age 76 years, range 66-91, n = 11, all male), and from control cases without serious central nervous system illness (mean age 77 years, range 61-95, n = 12, all male). We also examined brains of Tg2576 transgenic mice (males, aged 16-21 months), a model for chronic amyloid-induced brain injury. RESULTS: Immunohistochemical labelling showed DAPK1 expression in cortical neurones of human cortex and axonal tracts within subcortical white matter, both in AD and in control brains. Western analysis confirmed DAPK1 expression in all samples, although expression was very low in some control cases. DAPK1 abundance in the AD group was not significantly different from that in controls (P = 0.07, Mann-Whitney test). In brains of Tg2576 mice DAPK1 abundance was very similar to that in wild-type littermates (P = 0.96, Mann-Whitney test). CONCLUSION: We found that DAPK1 was expressed in neurones of aged human frontal cortex, both in AD and in control cases.

Can cutaneous telangiectasiae as late normal-tissue injury predict cardiovascular disease in women receiving radiotherapy for breast cancer?

BACKGROUND: Overall, approximately 5% of patients show late normal-tissue damage after radiotherapy with a smaller number having a risk of radiation-induced heart disease. Although the data are conflicting, large studies have shown increased risks of cardiovascular disease (CVD) for irradiated patients compared with non-irradiated ones, or for those treated to the left breast or chest wall compared with those treated to the right. Cutaneous telangiectasiae as late normal-tissue injury have so far only been regarded as a cosmetic burden. METHODS: The relationship between late normal-tissue radiation injury phenotypes in 149 irradiated breast cancer patients and the presence of cardiovascular disease were examined. RESULTS: A statistically significant association between the presence of skin telangiectasiae and the long-term risk of CVD was shown in these patients (P=0.017; Fisher's exact test). INTERPRETATION: This association may represent initial evidence that telangiectasiae can be used as a marker of future radiation-induced cardiac complications. It could also suggest a common biological pathway for the development of both telangiectasiae and CVD on the basis of a genetically predisposed endothelium. To our knowledge this is the first reported study looking at this association.

Mouse bone marrow and peripheral blood erythroid cell counts are regulated by different autosomal genetic loci.

Erythropoiesis is under fine control and genetic loci that affect it are likely to be important in a range of conditions. To assess the relative contributions of different genetic loci to parameters of erythropoiesis, we have measured RBC counts in the peripheral circulation and committed erythroid cells (RBC and small normoblasts) in the bone marrow in a cohort of (CBA/H x C57BL/6) F2 mice to map quantitative trait loci (QTL). Candidate genes were assessed using bioinformatics and DNA sequencing. Different autosomal loci affect bone marrow (chromosomes 5, 11 and 19) and peripheral blood (chromosome 4) erythroid cell counts but there may be a common chromosome X locus. Spleen weight QTL were found on chromosomes 3, 15 and 17. Surprisingly, erythropoietin (Epo) is the best candidate quantitative trait gene (QTG) in the chromosome 5 locus that affects bone marrow but not peripheral blood erythroid cell counts. Epo gene expression is known to be genetically regulated in mice, but our data suggest a tissue-specific role for epo in mouse erythropoiesis that is also genetically determined. The identity of the other QTG will be important both to further knowledge of the control of erythropoiesis and as potential modifier genes for haematological disorders.

A method for fine mapping quantitative trait loci in outbred animal stocks.

High-resolution mapping of quantitative trait loci (QTL) in animals has proved to be difficult because the large effect sizes detected in crosses between inbred strains are often caused by numerous linked QTLs, each of small effect. In a study of fearfulness in mice, we have shown it is possible to fine map small-effect QTLs in a genetically heterogeneous stock (HS). This strategy is a powerful general method of fine mapping QTLs, provided QTLs detected in crosses between inbred strains that formed the HS can be reliably detected in the HS. We show here that single-marker association analysis identifies only two of five QTLs expected to be segregating in the HS and apparently limits the strategy's usefulness for fine mapping. We solve this problem with a multipoint analysis that assigns the probability that an allele descends from each progenitor in the HS. The analysis does not use pedigrees but instead requires information about the HS founder haplotypes. With this method we mapped all three previously undetected loci [chromosome (Chr.) 1 logP 4.9, Chr. 10 logP 6.0, Chr. 15 logP 4.0]. We show that the reason for the failure of single-marker association to detect QTLs is its inability to distinguish opposing phenotypic effects when they occur on the same marker allele. We have developed a robust method of fine mapping QTLs in genetically heterogeneous animals and suggest it is now cost effective to undertake genomewide high-resolution analysis of complex traits in parallel on the same set of mice.

High-resolution mapping of quantitative trait loci in outbred mice.

Screening the whole genome of a cross between two inbred animal strains has proved to be a powerful method for detecting genetic loci underlying quantitative behavioural traits, but the level of resolution offered by quantitative trait loci (QTL) mapping is still too coarse to permit molecular cloning of the genetic determinants. To achieve high-resolution mapping, we used an outbred stock of mice for which the entire genealogy is known. The heterogeneous stock (HS) was established 30 years ago from an eight-way cross of C57BL/6, BALB/c, RIII, AKR, DBA/2, I, A/J and C3H inbred mouse strains. At the time of the experiment reported here, the HS mice were at generation 58, theoretically offering at least a 30-fold increase in resolution for QTL mapping compared with a backcross or an F2 intercross. Using the HS mice we have mapped a QTL influencing a psychological trait in mice to a 0.8-cM interval on chromosome 1. This method allows simultaneous fine mapping of multiple QTLs, as shown by our report of a second QTL on chromosome 12. The high resolution possible with this approach makes QTLs accessible to positional cloning.

Autosomal dominant dementia with widespread neurofibrillary tangles.

Several familial dementing conditions with atypical features have been characterized, but only rarely is the neuropathology dominated solely by neurofibrillary lesions. We present a Midwestern American pedigree spanning four generations in which 15 individuals were affected by early-onset dementia with long disease duration, with an autosomal dominant inheritance pattern, and with tau-rich neurofibrillary pathology found in the brain post mortem. The average age at presentation was 55 years with gradual onset and progression of memory loss and personality change. After 30 years' disease duration, the proband's neuropathologic examination demonstrated abundant intraneuronal neurofibrillary tangles (NFTs) involving the hippocampus, pallidum, subthalamic nucleus, substantia nigra, pons, and medulla. Only sparse neocortical tangles were present and amyloid plaques were absent. The tangles were recognized by antibodies specific for phosphorylation-independent (Tau-2, T46, 133, and Alz-50) and phosphorylation-dependent epitopes (AT8, T3P, PHF-1, 12E8, AT6, AT18, AT30) in tau proteins. Electron microscopy of NFTs in the dentate gyrus and midbrain demonstrated paired helical filaments. Although the clinical phenotype resembles Alzheimer's disease, and the neuropathologic phenotype resembles progressive supranuclear palsy, an alternative consideration is that this familial disorder may be a new or distinct disease entity.

Genetic association studies between dementia of the Alzheimer's type and three receptors for apolipoprotein E in a Caucasian population.

The epsilon 4 allele of the apolipoprotein E gene (ApoE) is associated with an increased risk for sporadic and some familial forms of Alzheimer's disease (AD) but the precise mechanism of pathogenesis is unknown. ApoE is a ligand for at least three receptors in the central nervous system, low density lipoprotein receptor (LDL-R), very low density lipoprotein receptor VLDL-R and low density lipoprotein-like receptor (LRP). We have tested for association between these receptors and dementia of the Alzheimer's type (DAT) in a clinically based sample of Caucasian cases and age-matched controls. In contrast to findings in a Japanese cohort we detected no association between DAT and a polymorphism in the VLDL-R gene. No association was detected with the LDL-R gene. We observed a possible association between the 87 allele of a polymorphism within the LRP gene and DAT which remained significant after correction for multiple testing. When the effects of known risk factors for AD such as ApoE epsilon 4 were applied, the effect of LRP no longer reached conventional levels of statistical significance. Nevertheless, LRP is a plausible candidate gene and we may be observing a minor risk factor that will require further examination in other large independent samples to assess whether it truly modifies susceptibility to DAT.

No association found between Alzheimer's disease and a mitochondrial tRNA glutamine gene variant.

We have screened a large sample of patients with sporadic late-onset dementia of the Alzheimer type (DAT) and age-matched controls for a mitochondrial tRNA(Gln) variant previously reported to be associated with increased risk of developing Alzheimer's disease (AD). The frequency of an Ava II site gain was determined by restriction analysis of a PCR-amplified mitochondrial DNA product. One of 155 DAT cases and four of 105 age-matched controls carried the variant. Both the affected and control frequencies are statistically different from those previously reported. The mitochondrial lineage of those individuals harboring the variant was determined by sequencing a short region of the hypervariable mitochondrial D-loop. The affected individual and three of the four controls carrying the Ava II variant belong to the same mitochondrial lineage previously reported to be associated with AD.

Seasonal movements in a salt glacier in iran.

The philosophy of the storage of high-grade radioactive wastes in salt seems to be based on the assumption that the salt glaciers of Iran are no longer moving. Monitoring the movements of markers painted onto one of the salt glaciers suggests that the glacier deforms elastically as a result of temperature changes most of the time but flows plastically when it is sufficiently wet during each annual rainy season.