Objective: Evaluate a patient-reported-oral mucositis scale (PROMS) on its own and in relation to existing measures of mucositis. Methods: 50 patients with head and neck cancer receiving radiotherapy were examined before cancer treatment, twice weekly during 6-7 weeks of therapy and post-therapy. Oral mucositis (OM) signs were evaluated clinically using NCI-CTCAE v.3, OMAS criteria and Total VAS-OMAS score. OM symptoms were recorded on PROMS-VAS questionnaires. Albumin and polymorphonuclear neutrophils were measured in saline rinses. The PROMS data were subjected to Spearman rank correlations versus the other clinical and biomarker data. Results: 33 participants completed the study. Significant correlations (p<.001) were seen between PROMS scores and other clinical and biomarker indicators of OM at a group level. Significant variations were seen between individuals. Conclusion: The PROMS tool demonstrates good correlation with other clinical indicators of OM and adds novel dimensions to currently available methods of assessments used for quantification of OM.

Cervix cancer affects women of all ages and causes significant morbidity and mortality. Locally advanced disease is curable with radiotherapy (RT) in about 50% of patients, although often at the expense of serious side effects. In order to improve the therapeutic ratio of tumour control versus normal tissue toxicity, conformal intensity-modulated radiotherapy (IMRT) is being investigated. However, inter- and intra-fractional motion of cervix cancer can contribute to both geographical miss of the target and overdosing of surrounding normal tissues, particularly in the setting of conformal IMRT with steep dose gradients. Defining the target volume accurately and understanding the dose consequence of these complex intra-pelvic organ dynamics during external beam radiotherapy forms the essential foundations for future treatment optimization and adaptation. This in turn will lead to improvements in tumour control and disease-free survival while minimising treatment toxicity.

Objective: Evaluate a patient-reported-oral mucositis scale (PROMS) on its own and in relation to existing measures of mucositis. Methods: 50 patients with head and neck cancer receiving radiotherapy were examined before cancer treatment, twice weekly during 6-7 weeks of therapy and post-therapy. Oral mucositis (OM) signs were evaluated clinically using NCI-CTCAE v.3, OMAS criteria and Total VAS-OMAS score. OM symptoms were recorded on PROMS-VAS questionnaires. Albumin and polymorphonuclear neutrophils were measured in saline rinses. The PROMS data were subjected to Spearman rank correlations versus the other clinical and biomarker data. Results: 33 participants completed the study. Significant correlations (p<.001) were seen between PROMS scores and other clinical and biomarker indicators of OM at a group level. Significant variations were seen between individuals. Conclusion: The PROMS tool demonstrates good correlation with other clinical indicators of OM and adds novel dimensions to currently available methods of assessments used for quantification of OM.

Localized prostate cancer (LPC) is predominantly a tumor among older men, and few patients will get symptoms from the disease. All methods to treat LPC with a curative intent have different types and degrees of side effects. It is therefore very important to evaluate the side effects thoroughly to make sure that treatment complications will not decrease the quality of life more than the disease would have done. In search for new and better treatments, complications has to be registered and evaluated in relation to quality of life (QOL) for the patients. Few validated self-assessment questionnaires for evaluation of external radiotherapy (EBRT) induced side effects has yet been developed. The present project focus on the development of the PC-specific questionnaire, QUFW94, and evaluation of symptoms in patients treated with EBRT and un-treated (watchful waiting) patients with a LPC. In the newly developed LPC-specific questionnaire a reliability and responsiveness test was performed. Both the inter-rater test and the test-retest show high correlation coefficients (ICC), above 0.60 for all scales. The internal reliability exceeded the lower acceptable limit (Cronbach a &gt;0.70). The questionnaire was proven to be valid for the evaluations of EBRT side effects in LPC patients. Late side effects were evaluated 4 years after treatment in 181 LPC patients, treated with conventional large field EBRT, and compared with 141 age-matched PC disease free men. The most prominent urinary side effects were urgency and leakage which were doubled in the patient group. A ten fold increase was seen in comparison to controls at the most prominent intestinal problems, blood, mucus and leakage. The results support the use 3-D conformai therapy to decrease irradiation dose to the rectum and the bladder and thereby decreased side effects. A prospective additional evaluation 8 years after EBRT did not show any changes in urinary problems between 4 and 8-yr follow-up in the patients or the controls. EBRT of LPC is also accompanied by disturbances in sexual function. These problems were therefore evaluated, 4 years after EBRT, in relation to the function in PC free men. Patients treated with EBRT indicated higher levels of sexual dysfunction than age-matched controls. No erection was reported from 12% of the control subjects, 56% of the patients who had only received radiotherapy (RT) and 87% of the RT+castration (RT+A) patients. The extended evaluation 8 years after EBRT show similar sexual function in all groups. QOL and late side effects/symptoms were evaluated in the first and only randomized trial between RT and deferred treatment (DT) and compared to age-matched controls. QOL was evaluated with the general QOL formula, EORTC's QLQ-C30 (+3), and LPC specific side effects with QUFW94 in 108 randomized patients with LPC 3 years after diagnosis. Social functioning was the only QOL scale where a significant difference was found between the two patient groups and in comparison with the control group. Multivariate regression analysis showed that hematuria, incontinence, mucus, and planning of the daily activities due to intestinal problems caused this decrease in QOL in the RT group. In conclusion, the LPC specific QUFW94 questionnaire was proven to be valid for evaluation of side effects and showed increased intestinal problems in the patients treated with conventional large field EBRT in comparison to untreated LPC patients. No difference in urinary and intestinal late side effects or sexual function was seen between a 4 year vs. 8 year follow-up. / digitalisering@umu

The radiosensitivity of tumour cells depends mainly on their capacity to maintain genomic integrity. This requires efficient repair of radiation-induced DNA double strand breaks, a process governed by the cell cycle. Based on their functions in cell cycle regulation and DNA damage repair, we hypothesised that targeting of CDK2 and CDK9 modifies cancer cell response to radiotherapy. Therefore, we evaluated the significance of CDK2 and CDK9 for the cellular radiation response in a panel of human head and neck squamous cell carcinoma (HNSCC) cell lines. In order to achieve our goal, we performed a series of experiments to measure several key parameters such as clonogenic radiation survival, cell cycling, DNA damage repair and apoptosis. We found that loss of CDK2 radiosensitises mouse embryonic fibroblasts (MEFs) as well as HNSCC two dimensional (2D) cell cultures. However, under more physiological three dimensional (3D) growth conditions in laminin-rich extracellular matrix, targeting of CDK2 failed to modulate the radiosensitivity of HNSCC cells. Moreover, CDK2 attenuated the repair of radiogenic double strand breaks (DSBs) in MEFs as well as SAS and FaDu HNSCC cells indicating a possible role of CDK2 in DNA damage repair. However, we found that CDK2 is dispensable for cell cycle and checkpoint regulation in response to irradiation in SAS and FaDu cells. Taken together, our results suggest that targeting of CDK2 may not provide a therapeutic benefit to overcome HNSCC cell resistance to radiotherapy. We also showed that depletion of CDK9 clearly enhances the radiosensitivity of HNSCC cultures. In addition, the ectopic expression of CDK9 has a radioprotective effect. These findings suggest a potential role of CDK9 in the radiation response of HNSCC cells. Moreover, our study indicates a possible role of CDK9 in the DNA damage repair response and cell cycling of HNSCC cells. Conclusively, on the basis of these data, targeting of CDK9 in addition to conventional radiotherapy might be a viable strategy to overcome cancer cell resistance.

Replacement of thymidine in DNA by halopyrimidines, such as bromodeoxyuridine (BrdU) and iododeoxyuridine (IdU), has long been known to enhance DNA damage and cell death induced by ionizing/UV radiation, but the mechanism of action of halopyrimidines at the molecular level is poorly understood. We have applied advanced time-resolved femtosecond laser spectroscopy to this molecular system of biological, chemical and medical significance. We obtained the first real-time observations of the transition states of the ultrafast electron transfer (UET) reactions of halopyrimidines with the ultrashort-lived precursor to the hydrated electron, which is a general product in ionizing/UV radiation. Our results provide a mechanistic understanding of these photo-/radiosensitizing drugs at the molecular level. We found that the UET reaction of BrdU is completed within 0.2 picosecond (ps) after the electronic exciataion, leading to the formation of the transition state BrdU* with a lifetime of ~1.5 ps that then dissociates into Br and a high reactive radical dU•. We have also demonstrated that the reaction efficiency for the formation of the reactive radical dU• to cause DNA damage and cell death is in the order of IdU>>BrdU>CldU>>FdU. This is due to the availability of two precursor states of ~0.2 ps and ~ 0.54 ps lifetimes for dissociative electron attachment (DEA) to IdU, of one precursor state of ~0.2 ps lifetime for DEAs to BrdU and CldU, and no precursors for DEA to FdU. This explains why BrdU and IdU were found to be effective radio-/photosensitizers and indicates that IdU should be explored as the most effective radiosensitizer among halopyrimidines. Moreover, as a by-product of this project, these halopyrimidines have been employed as quantum-state-specific molecular probes to resolve a long-standing controversy about the nature and lifetimes of prehydrated electrons. These findings also have a broader significance as they indicated that nonequilibrium precursor electrons may play an important role in electron-initiated reactions in many biological, chemical and environmental systems. We have also demonstrated UET reactions of nucleotides with the precursor to the hydrated electrons. Our results indicate that among DNA bases, adenine is the most efficient electron trapper and an effective electron transfer promoter, while guanine is the most effective in dissociative electron attachment. These results not only primarily explain the sequence selectivity of duplex DNA containing BrdU/IdU, but imply that the DEA of guanine is an important mechanism for radiation-induced DNA damage in ionizing radiation and radiotherapy of cancer.

Purpose: To examine the dosimetric characteristics of Gafchromic EBT3 film when measuring small fields of radiation, and compare it against other common radiation detectors. Methods and Materials: EBT3 film was placed in a solid water phantom and irradiated with 6MV photons, field sizes from 10x10cm2 down to 6x6mm2. The films were scanned with a Vidar DosimetryPRO Advantage Red scanner, and analyzed with RIT113 software. The films were also scanned at different orientations and times to quantify the discrepancies associated with scanning orientation and post-exposure darkening. The same fields were measured with a PTW TN30013 farmer chamber, an Exradin T1 cylindrical ion chamber, a PTW parallel plate ion chamber, and a Sun Nuclear Edge Detector (diode). Output factors were calculated for each detector and compared for accuracy. The output factors were measured from a Varian Clinac iX, Clinac 21EX, Trilogy, and TrueBeam; as well as a Novalis Tx. The outputs from different machines at different clinics were compared. Results: The EBT3 film and Edge Detector were the only detectors that succeeded in accurately measuring the output from all field sizes; the ion chambers were too large and failed for field sizes below 4x4cm2 due to volume averaging. The dose measured with the film increased by an average of 8.8% after one week post-irradiation. The dose measured was also reduced by an average of 4.4% by scanning the film in landscape orientation, as opposed to portrait orientation. It was shown that the output factors for the smallest field of 6x6mm2--successfully measured with film and diode--varied between 0.54-0.74 for five different machines at three different clinics. Conclusions: The feasibility of using Gafchromic EBT3 film to measure very small fields of radiation is confirmed. Of the other 4 detectors used, only the diode was shown to be capable of accurately measuring small fields of radiation. The need to optimize the film dosimetry process--including the time films are scanned post-irradiation, the consistency of the scanning orientation of the calibration and subsequent films, and the measurement procedure on the computer software--is highlighted.

Abstract   Background Semi-automated segmentation using deformable registration of atlases consisting of pre-segmented patient images can facilitate the tedious task of delineating structures and organs in patients subjected to radiotherapy planning. However, a generic atlas based on a single patient may not function well enough due to the anatomical variation between patients. Fusion of segmentation proposals from multiple atlases has the potential to provide a better segmentation due to a more complete representation of the anatomical variation. Purpose The main goal of the present study was to investigate potential operator timesavings from use of atlas-based segmentation compared to manual segmentation of patients with prostate cancer. It was also anticipated that, and evaluated if, the use of semi-automated segmentation workflows would reduce the operator dependent variations in delineation. Materials and Methods A commercial atlas-based segmentation software (VelocityAI from Nucletron AB) was used with several atlases of consistently, protocol based, delineated CT images to create multiple-atlas segmentation proposals through deformable registration. The atlas that was considered most representative was selected to construct single generic atlas segmentation proposals. For fusion of the multiple-atlas segmentations an in-house developed algorithm, which includes information of local registration success was used in a MATLAB-environment[1]. The algorithm used weighted distance map calculations where weights represent probabilities of improving the segmentation results. Based on results from Sjöberg and Ahnesjö the probabilities were estimated using the cross correlation image similarity measure evaluated over a region within a certain distance from the segmentation. 10 patients were included in the study. Each patient was delineated three times, (a) manually by the radiation oncologist, (b) with a generic single-atlas segmentation and (c) with a fusion of multiple-atlas segmentations. For the methods (b) and (c) the radiation oncologist corrected the proposed segmentations blindly without using the result from method (a) as reference. The total number of atlases used for case (c) was 15. The operator time spent by the radiation oncologist was recorded separately for each method. In addition a grading was used to score how helpful the segmentation proposals were for the delineations. The Dice Similarity Coefficient, the Hausdorff distance and the segmented volumes were used to evaluate the similarity between the delineated structures and organs. Results An average time reduction of 26% was found when the radiation oncologist corrected the multiple atlas-based segmentation proposals as compared to manual segmentations. Due to more accurate segmentations and more time saved, segmentation with fused multiple-atlases (c) was superior to the generic single-atlas (b) method, which showed a time reduction of 17%. Hints of an affected intra- and inter-operator variability were seen. Conclusions Atlas-based segmentation saves time for the radiation oncologist but the segmentation proposals always need editing to be approved for dose planning. The atlases, the fusion of these and the software implementation needs to be improved for optimal results and to extend the clinically usefulness.

Background Semi-automated segmentation using deformable registration of atlases consisting of pre-segmented patient images can facilitate the tedious task of delineating structures and organs in patients subjected to radiotherapy planning. However, a generic atlas based on a single patient may not function well enough due to the anatomical variation between patients. Fusion of segmentation proposals from multiple atlases has the potential to provide a better segmentation due to a more complete representation of the anatomical variation. Purpose The main goal of the study was to investigate potential operator timesaving from editing of atlas-based segmentation compared to manual segmentation for head &amp; neck cancer. Materials and Methods A commercial atlas-based segmentation software (VelocityAI from Nucletron AB) was used together with several expert generated and protocol-based atlases of delineated CT images to create multiple atlas segmentations through deformable registration. The atlas that was considered most universal was selected to construct single atlas segmentation proposals. For fusion of the multiple atlas segmentations an in-house developed algorithm, including information of local registration success was used in a MATLAB-environment1. The algorithm uses weighted distance map calculations where weights represent probabilities of improving the segmentation results. Based on previous results1 the probabilities were estimated using the cross correlation image similarity measure evaluated over a region within a certain distance from the segmentation. Ten patients were incorporated in the study. Each patient was delineated three times, (a) manually by the radiation oncologist, (b) with a single atlas segmentation and (c) with a fusion of multiple atlas segmentations. For the methods (b) and (c) the radiation oncologist corrected the proposed segmentations blindly without using the result from method (a) as reference. For case (c) a total number of 11 atlas segmentations were used. The time spent for segmenting or editing the segmentation proposals by the radiation oncologist was recorded separately for each method and each individual ROI. In addition a grading was used to score how helpful the candidate segmentation proposals were for the structure delineations. The Dice Similarity Coefficient, the Hausdorff distance and the volume were used to evaluate the similarity between the delineated structures. Results The results show a time reduction in the order of 40% when the radiation oncologist only has to correct the multiple atlas-based segmentation proposal compared to manual segmentation. When using single atlas the corresponding figure is 21%. Conclusions Using atlas-based segmentation can reduce the time needed for delineation in the head and neck area of patients admitted for radiotherapy. 1C. Sjöberg and A. Ahnesjö, Evaluation of atlas-based segmentation using probabilistic weighted distance maps, Manuscript, Uppsala University, 2011 / Bakgrund Atlasbaserad, semiautomatisk segmentering skulle kunna användas för att underlätta den för onkologen tidskrävande uppgiften med att manuellt segmentera strukturer och organ i patienter vid behandlingsplanering inför strålbehandling. Tidigare segmenterade atlaspatienter ger med hjälp av deformeringsalgoritmer segmenteringsförslag för strukturer i den aktuella patienten. Dessa kan sedan kontrolleras och editeras av onkologen med en tidsbesparing gentemot manuell segmentering som följd. En atlas som baserats på en enstaka individ (singelatlas) kan dock ha begränsningar när det gäller att täcka de anatomiska variationer som finns mellan olika patienter. Därför har metoder med fusionering av multipla segmenteringsförslag från en databas bestående av ett antal sedan tidigare segmenterade patienter (fusionerad multipelatlas) potential att ge ett bättre segmenteringsresultat. Syfte Huvudsyftet med arbetet var att undersöka de möjliga tidsbesparingar för onkologen som kan åstadkommas när editering av atlasbaserad segmentering används vid planering inför strålbehandling i huvud- och halsområdet istället för manuell segmentering Material och metoder En kommersiell, atlasbaserad segmenteringsprogramvara (VelocityAI från Nucletron AB) användes i studien. Genom att låta en erfaren onkolog segmentera ett antal CT-studier (11 st) enligt ett vedertaget protokoll skapades en databas av atlaser som sedan, via deformerbara registreringar, kunde generera lika många segmenteringsförslag för en nytillkommen patient. Den enskilda atlas som ansågs mest representativ valdes till att framställa segmenteringsförslaget för metoden med singelatlas. Till metoden med fusionerade multipla atlaser användes en lokalt utvecklad MATLAB-algoritm baserad på viktade    distansmappar. Vikterna representerar sannolikheten för förbättrat segmenteringsresultat och baseras på tidigare resultat1 där sannolikheterna bestämts utifrån en beräkning av likheterna mellan bilderna i ett visst område från den specifika segmenteringen. Tio patienter har inkluderats i studien. Varje patient segmenterades tre gånger, (a) manuellt, (b) med singelatlas och (c) med fusionerade multipla atlaser. För metoderna (b) och (c) editerades sedan segmenteringsförslagen av onkologen utan att denne fick använda resultatet från metod (a) som referens. För fallet med fusionerade multipla atlaser, (c), användes databasen med 11 atlaser. Tiden onkologen behövde för segmentering respektive editering av segmenteringsförslaget uppmättes i varje enskilt fall för jämförelse. Onkologen fick även göra en bedömning av hur hjälpsamt segmenteringsförslaget var i samband med editeringen. För utvärdering av resultaten användes Dice’s similaritetskoefficient, Hausdorff’s distansmått samt strukturernas volym. Resultat Resultaten visar på att en tidsbesparing i storleksordningen 40 % är rimlig när onkologen editerar förslag från fusioneringen av multipla atlassegmenteringar i jämförelse med manuell segmentering. Vid användning av singelatlas är motsvarande siffra 21 %. Slutsatser Användandet av atlasbaserad segmentering kan reducera tidsåtgången för segmentering av patienter inför strålbehandling i huvud-halsområdet. 1C. Sjöberg and A. Ahnesjö, Evaluation of atlas-based segmentation using probabilistic weighted distance maps, Manuscript, Uppsala University, 2011

Replacement of thymidine in DNA by halopyrimidines, such as bromodeoxyuridine (BrdU) and iododeoxyuridine (IdU), has long been known to enhance DNA damage and cell death induced by ionizing/UV radiation, but the mechanism of action of halopyrimidines at the molecular level is poorly understood. We have applied advanced time-resolved femtosecond laser spectroscopy to this molecular system of biological, chemical and medical significance. We obtained the first real-time observations of the transition states of the ultrafast electron transfer (UET) reactions of halopyrimidines with the ultrashort-lived precursor to the hydrated electron, which is a general product in ionizing/UV radiation. Our results provide a mechanistic understanding of these photo-/radiosensitizing drugs at the molecular level. We found that the UET reaction of BrdU is completed within 0.2 picosecond (ps) after the electronic exciataion, leading to the formation of the transition state BrdU* with a lifetime of ~1.5 ps that then dissociates into Br and a high reactive radical dU•. We have also demonstrated that the reaction efficiency for the formation of the reactive radical dU• to cause DNA damage and cell death is in the order of IdU>>BrdU>CldU>>FdU. This is due to the availability of two precursor states of ~0.2 ps and ~ 0.54 ps lifetimes for dissociative electron attachment (DEA) to IdU, of one precursor state of ~0.2 ps lifetime for DEAs to BrdU and CldU, and no precursors for DEA to FdU. This explains why BrdU and IdU were found to be effective radio-/photosensitizers and indicates that IdU should be explored as the most effective radiosensitizer among halopyrimidines. Moreover, as a by-product of this project, these halopyrimidines have been employed as quantum-state-specific molecular probes to resolve a long-standing controversy about the nature and lifetimes of prehydrated electrons. These findings also have a broader significance as they indicated that nonequilibrium precursor electrons may play an important role in electron-initiated reactions in many biological, chemical and environmental systems. We have also demonstrated UET reactions of nucleotides with the precursor to the hydrated electrons. Our results indicate that among DNA bases, adenine is the most efficient electron trapper and an effective electron transfer promoter, while guanine is the most effective in dissociative electron attachment. These results not only primarily explain the sequence selectivity of duplex DNA containing BrdU/IdU, but imply that the DEA of guanine is an important mechanism for radiation-induced DNA damage in ionizing radiation and radiotherapy of cancer.