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Bronchialkarzinome sind in Deutschland die dritthäufigste Tumorerkrankung. Trotz erheblicher therapeutischer Fortschritte ist die Prognose der Lungentumoren nach wie vor ungünstig, die relative 5-Jahres- Überlebensrate nach Diagnose eines Bron- chialkarzinoms beträgt lediglich 16 % [3]. Hinsichtlich der Tumorart werden nicht- kleinzellige (NSCLC) und kleinzellige Bron- chialkarzinome unterschieden. Bei fortgeschrittenen NSCLC besteht die Standardtherapie aus einer gleichzei- tig durchgeführten Radiochemotherapie, gefolgt von einer Immuntherapie. In Metaanalysen konnte die Überlegenheit der simultanen Radiochemotherapie ge- genüber einem sequenziellen Vorgehen gezeigt werden.

In times of high‐precision radiotherapy, the accurate and precise definition of the primary tumor localization and its microscopic spread is of enormous importance. In glioblastoma, the microscopic tumor extension is uncertain and, therefore, population‐based margins for Clinical Target Volume (CTV) definition are clinically used, which could either be too small—leading to increased risk of loco‐regional recurrences r too large, thus, enhancing the probability of normal tissue toxicity. Therefore, the aim of this project is to investigate an individualized definition of the CTV in preclinical glioblastoma models based on specific biological tumor characteristics. The microscopic tumor extensions of two different orthotopic brain tumor models (U87MG_mCherry; G7_mCherry) were evaluated before and during fractionated radiotherapy and correlated with corresponding histological data. Representative tumor slices were analyzed using Matrix‐Assisted Laser Desorption/Ionization (MALDI) and stained for putative stem‐like cell markers as well as invasion markers. The edges of the tumor are clearly shown by the MALDI segmentation via unsupervised clustering of mass spectra and are consistent with the histologically defined border in H&E staining in both models. MALDI component analysis identified specific peaks as potential markers for normal brain tissue (e.g., 1339 m/z), whereas other peaks demarcated the tumors very well (e.g., 1562 m/z for U87MG_mCherry) irrespective of treatment. MMP14 staining revealed only a few positive cells, mainly in the tumor border, which could reflect the invasive front in both models. The results of this study indicate that MALDI information correlates with microscopic tumor spread in glioblastoma models. Therefore, an individualized CTV definition based on biological tumor characteristics seems possible, whereby the visualization of tumor volume and protein heterogeneity can be potentially used to define radiotherapy sensitive and resistant areas.

Purpose: The standardized uptake value (SUV) is widely used for quantitative evaluation in oncological FDG-PET but has well-known shortcomings as a measure of the tumor’s glucose consumption. The standard uptake ratio (SUR) of tumor SUV and arterial blood SUV (BSUV) possesses an increased prognostic value but requires image-based BSUV determination, typically in the aortic lumen. However, accurate manual ROI delineation requires care and imposes an additional workload which makes the SUR approach less attractive for clinical routine. The goal of the present work was the development of a fully automated method for BSUV determination in whole-body PET/CT. Methods: Automatic delineation of the aortic lumen was performed with a convolutional neural network (CNN), using the U-Net architecture. 946 FDG PET/CT scans from several sites were used for network training (N = 366) and testing (N = 580). For all scans, the aortic lumen was manually delineated, avoiding areas affected by motion-induced attenuation artifacts or potential spill-over from adjacent FDG-avid regions. Performance of the network was assessed using the fractional deviations of automatically and manually derived BSUVs in the test data. Results: The trained U-Net yields BSUVs in close agreement with those obtained from manual delineation. Comparison of manually and automatically derived BSUVs shows excellent concordance: the mean relative BSUV difference was (mean ± SD) = (-0.5± 2.2)% with a 95% confidence interval of [−5.1, 3.8]% and a total range of [-10.0, 12.0]%. For four test cases the derived ROIs were unusable (

Background The limited availability of proton beam therapy (PBT) requires individual treatment selection strategies that can be based on normal tissue complication probability (NTCP) models. We developed and externally validated NTCP models for common late side-effects following PBT in brain tumour patients to optimise patients’ quality of life. Methods Cohorts from three PBT centres (216 patients) were investigated for several physician-rated endpoints at 12 and 24 months after PBT: alopecia, dry eye syndrome, fatigue, headache, hearing and memory impairment, and optic neuropathy. Dose-volume parameters of associated normal tissues and clinical factors were used for logistic regression modelling in a development cohort. Statistically significant parameters showing high area under the receiver operating characteristic curve (AUC) values in internal cross-validation were externally validated. In addition, analyses of the pooled cohorts and of time-dependent generalised estimating equations including all patient data were performed. Results In the validation study, mild alopecia was related to high dose parameters to the skin [e.g. the dose to 2% of the volume (D2%)] at 12 and 24 months after PBT. Mild hearing impairment at 24 months after PBT was associated with the mean dose to the ipsilateral cochlea. Additionally, the pooled analyses revealed dose-response relations between memory impairment and intermediate to high doses to the remaining brain as well as D2% of the hippocampi. Mild fatigue at 24 months after PBT was associated with D2% to the brainstem as well as with concurrent chemotherapy. Moreover, in generalised estimating equations analysis, dry eye syndrome was associated with the mean dose to the ipsilateral lacrimal gland. Conclusion We developed and in part validated NTCP models for several common late side-effects following PBT in brain tumour patients. Validation studies are required for further confirmation.

Resistance of cancer stem-like and cancer tumor bulk cells to radiochemotherapy and destructive infiltration of the brain fundamentally influence the treatment efficiency to cure of patients suffering from Glioblastoma (GBM). The interplay of adhesion and stress-related signaling and activation of bypass cascades that counteract therapeutic approaches remain to be identified in GBM cells. We here show that combined inhibition of the adhesion receptor β1 integrin and the stress-mediator c-Jun N-terminal kinase (JNK) induces radiosensitization and blocks invasion in stem-like and patient-derived GBM cultures as well as in GBM cell lines. In vivo, this treatment approach not only significantly delays tumor growth but also increases median survival of orthotopic, radiochemotherapy-treated GBM mice. Both, in vitro and in vivo, effects seen with β1 integrin/JNK co-inhibition are superior to the monotherapy. Mechanistically, the in vitro radiosensitization provoked by β1 integrin/JNK targeting is caused by defective DNA repair associated with chromatin changes, enhanced ATM phosphorylation and prolonged G2/M cell cycle arrest. Our findings identify a β1 integrin/JNK co-dependent bypass signaling for GBM therapy resistance, which might be therapeutically exploitable.

Aim: The standardized uptake value (SUV) is widely used for quantitative evaluation in oncological FDG-PET but has well-known shortcomings as a measure of the tumor's glucose consumption. The standard uptake ratio (SUR) of tumor SUV and arterial blood SUV (BSUV) possesses an increased prognostic value but requires image-based BSUV determination, typically in the aortic lumen. However, accurate manual ROI delineation requires care and imposes an additional workload which makes the SUR approach less attractive for clinical routine. The goal of the present work was the development of a fully automated method for BSUV determination in whole-body PET/CT. Methods: Automatic delineation of the aortic lumen was performed with a convolutional neural network (CNN), namely U-Net. 632 FDG PET/CT scans from 4 different sites were used for network training (N=208) and testing (N=424). For all scans, the aortic lumen was manually delineated, avoiding areas affected by motion-induced attenuation artifacts or potential spill-over from adjacent FDG-avid regions. Performance of the network was assessed using the fractional deviations of automatically and manually derived BSUVs in the test data. Results: The trained U-Net yields BSUVs in close agreement with those obtained from manual delineation. Notably, using both CT and PET data as input for network training allows the trained network to derive unbiased BSUVs by detecting and excluding aorta segments affected by attenuation artifacts or spill-over. Comparison of manually (M) and automatically (A) derived BSUVs shows excellent concordance: the mean paired M-A difference in the 424 test cases is (mean +/- SD)=(0.2 +/- 3.1)% with a 95% confidence interval of [-6.6, 5.7]%. For a single test case the M-A difference exceeded 10%. Conclusion: CNNs offer a viable approach for automatic BSUV determination. Our trained network exhibits a performance comparable to an experienced human observer and might already be considered suitable for supervised clinical use.

Background Neurocognitive function of adult patients with brain tumours may deteriorate after radiotherapy. Proton beam therapy (PBT) reduces the volume of irradiated healthy brain tissue and could potentially preserve neurocognition and quality of life (QoL). As present data are still limited, the impact of clinical factors and dosimetric parameters on neurocognitive function and QoL during recurrence-free follow-up after PBT is investigated. Methods The current study includes 62 brain tumour patients treated with PBT between 2015 and 2017. Neurocognition and QoL were assessed at baseline and every 3 months after PBT using the Montreal Cognitive Assessment (MoCA) test together with EORTC-QLQ-C30 and BN20 questionnaires, respectively. Objective and self-reported measures of neurocognitive functions were correlated. During two years of follow-up, the impact of clinical co-factors as well as dosimetric parameters of several brain structures were analysed using a mixed-model approach. Results At baseline, mean MoCA total score was 24.8/30 and self-reported cognitive function was 68.9/100. Both remained stable over time. Patients with impaired neurocognition on the MoCA test reported significantly lower global health status, cognitive, physical and role function as well as more fatigue, pain, headache and communication deficits compared to normal performing patients. For most follow-up time points, the majority of MoCA subitems correlated significantly to QoL items regarding neurocognition. Slight deterioration of the MoCA score was associated with tumours located in the left hemisphere and with an increase in relative volume of the anterior cerebellum that received doses of 30 to 40 Gy(RBE). Conclusion Self-reported and objectively measured neurocognition and most other QoL domains remained largely stable over time during recurrence-free follow-up for brain tumour patients treated with PBT. The association between reduced cognitive function and irradiated volume of the anterior cerebellum requires validation in larger studies and comparison to patients treated with photon therapy.

Purpose: To investigate late physician-rated side effects and their association with dosimetric parameters of various organs at risk (OARs) as well as clinical cofactors in adult brain tumour patients following proton beam therapy (PBT). Material and methods: Adult patients with brain tumours who underwent PBT at three different institutes were included in this study (N1=57, N2=47, N3=63). The radiation-induced side effects alopecia, fatigue, headache, memory impairment, hearing impairment, optic nerve disorder, dry eye and seizure (CTCAE v4.0) at 12 and 24 months after PBT were investigated. Side effects with sufficiently high incidence were dichotomised and correlated to different dose-volume histogram (DVH) parameters of associated OARs, such as skin, remaining brain, brainstem, cerebellum, hippocampi and cochlea. Clinical parameters comprised age, gender, tumour volume, prescribed dose, concomitant chemotherapy, resection of the tumour, diagnosis and WHO grading. Normal tissue complication probability (NTCP) models were developed on a combined cohort from two institutes (N=104) using logistic regression. The area under the receiver operating characteristic curve (AUC) was used to assess the prognostic ability in external validation on the remaining cohort. Results: In all cohorts, low toxicity rates were observed at 12 and 24 months after PBT. Most common side effects were fatigue (grade≥1: 32%/27%, grade≥2: 14%/6% at 12/24 months), alopecia (grade≥1: 30%/22%) and mild memory impairment (grade≥1: 23% at 24 months). Mild headache and hearing impairment (grade≥1) occurred in 20%/19% and 8%/9% of all patients at 12/24 months, respectively. Logistic regression revealed significant correlations between the incidence of alopecia grade≥1 at both times and high dose regions of the skin (D2%, p

Magnetic resonance imaging (MRI) and immunohistochemical tissue stainings are pivotal for radiotherapeutic workflows. Yet, recent efforts herald a paradigm shift: Radiomic methods are used to extract a large number of quantitative features from image data to detect high-dimensional patterns, which are correlated with relevant clinical endpoints. Preclinical experiments help to understand underlying mechanisms, yet require the backtranslation of clinically used methods and their application to a heterogeneous patient cohort. In the present preclinical experiment, we determine the tumor phenotype from MRI and tumor microenvironment (TME) features in a patient cohort of xenograft tumor models of the head and neck. An artificial heterogeneous patient population was created by mixing two tumor models of different radiosensitivity (SAS & UT-SCC-14) in pooled cohort (N = 108) and exposure to one week of fractionated irradiation with photons and protons. After irradiation, contrast agent-enhanced T1-weighted 3D gradient-echo MRI scans were acquired, tumors were excised and characterized immunohistochemically regarding vascularity (CD31), hypoxia (Pimonidazole) and morphology (H&E). Approximately 200 quantitative features were extracted from MRI and light-microscopy image data with an automated medical image radiomics processor and trainable image segmentation, respectively. TME parameters were analyzed regarding effects of radiation with two-sided t-tests. A fully automated radiomic framework was used for feature selection, model generation using leave-one-out cross validation with the individual tumor model’s identity (i.e. its phenotype) as endpoint. Model performance was assessed through area under the curve (AUC). The used image quantification methods allowed for robust feature extraction. No effects of radiation on the TME were detected except for changes in vessel-adjacent hypoxia. Radiomic analysis was able to predict the tumor model based on TME features (AUC = 0.86), MRI features (AUC = 0.90) or combined features (AUC = 0.86). We demonstrated backtranslation of radiomic methods in a preclinical setting with multi-modal image data. Further analysis of automatically extracted MRI and TME features may allow for a more biologically informed interpretation of MRI data.

Purpose: Proton radiotherapy offers the potential to reduce normal tissue toxicity. However, clinical safety margins, range uncertainties and varying relative biological effectiveness (RBE) may result in a critical dose in tumor-surrounding normal tissue. To assess potential adverse effects in preclinical studies, we established stereotactic proton mouse brain irradiation and a cell-based analysis of radiation damage repair. Material and methods: A setup to shape a proton beam with 7 mm range in water and 3 mm in diameter was built and dosimetrically characterized. Cone-beam computed tomography (CBCT) and orthogonal X-ray imaging were used to delineate the right hippocampus (target) and to position the mice, respectively. For two mouse strains (C57BL/6 and C3H), brains were irradiated with 4 Gy or 8 Gy and excised after 30 min or 3 h. Brain sections (3 µm) were cut every 100 µm and DNA double-strand break (DSB) repair kinetics was visualized by staining for cell nuclei and H2AX. Imaged sections were analyzed with an automated and validated processing pipeline to provide a quantitative, spatially-resolved damage indicator. Results: Twenty mice underwent the treatment workflow including imaging, target delineation, positioning, and irradiation. The analyzed DNA damage pattern clearly visualized the radiation effect and could be mapped onto the measured dose distribution. For all evaluated C3H mice, the proton beam hit the right hippocampus and stopped in the brain. Damage pattern became spatially more extended and diffuse for 8 Gy and 3 h after irradiation, respectively. C57BL/6 mice showed comparable damage distributions, however, with larger spatial variation of the beam alignment relative to the hippocampus. Conclusion: We established and biologically validated stereotactic proton irradiation of mouse brains. The clinically-oriented workflow facilitates (back-) translational studies. Geometric accuracy and cell-based assessment enable a biologically and spatially resolved analysis of radiation response and RBE.

Background and purpose: Early radiation-induced esophageal toxicity (RIET) is one of the major side effects in patients with non-small cell lung cancer (NSCLC) and can be a reason for treatment interruptions. As the age of patients with NSCLC and corresponding comorbidities continue to increase, primary radiotherapy alone is a commonly used alternative treatment in these cases. The aim of the present study is to compare dosimetric and clinical parameters from the previously reported CHARTWEL trial for their ability to predict esophagitis and investigate potential differences in the accelerated and conventional fractionation arm. Material and methods: 146 patients of the Dresden cohort of the randomized phase III CHARTWEL trial were included in this post-hoc analysis. Side effects were prospectively scored weekly during the first 8 weeks from start of radiotherapy. To compare both treatment arms, recorded dose-volume parameters were adjusted for the different fractionation schedules. Logistic regression was performed to predict early RIET for the entire study group as well as for the individual treatment arms. Differentdosimetric and clinical parameters were tested. Results: Patients receiving the accelerated CHARTWEL schedule experienced earlier and more severe esophagitis (e.g. 20.5% v . 9.6% ≤ grade 2 at week 3, respectively). In contrast, the median time period for recovery of grade 1 esophagitis was significantly longer for patients with conventional fractionation compared to the CHARTWEL group (median [range]: 21 [12-49] days vs. 15 [7-84] days, p=0.028). In univariable logistic regression none of the dose-volume parameters showed a significant correlation with early RIET grade  2 in the conventional irradiation group. In contrast, for patients receiving CHARTWEL, the physical dose-volumes parameters V40 and V50; and re-scaled values VEQD2,50 and VEQD2,60 were significant predictors of early RIET grade  2. Dose-volume parameters remained different between CHARTWEL and conventional fractionation even after biological rescaling. Conclusion: Our results show a more dominant dose-volume effect in the CHARTWEL arm compared to conventional fractionation, especially for higher esophageal doses. These findings support the notion that dose-volume parameters for radiation esophagitis determined in a specific and time dependent setting of field arrangements can not be easily transferred to another setting. In clinical practice esophageal volumes receiving 40 Gy or more should be strictly limited in hyperfractionated accelerated fraction schemes.

Objective: Radiotherapy leads to inactivation of tumor cells following radiation-induced DNA damage. Compared to conventional photon-based radiotherapy, proton therapy offers the potential of normal tissue sparring due to its favorable depth-dose distribution. However, acute or long-term side effects could still occur due to clinical safety margins and uncertainties about the relative biological effectiveness (RBE). While a variable RBE has been demonstrated in in vitro studies, especially at the end of the proton range, in clinical practice, a constant RBE value of 1.1 is applied. To elucidate the RBE issue based on in vivo experiments, proton irradiation of mouse brains was realized in Dresden. Methods: Experiments were performed at the experimental beam line [1] of the University Proton Therapy Dresden. For beam characterization and dosimetry, a 2D scintillation detector, ionization chambers and radiochromic films [2] were used. A multi-modality mouse bed suitable for imaging, transportation and irradiation was developed in-house. Like clinical applications, the workflow includes computed tomography scans for treatment planning and X-ray images for refined positioning. By combining these images with proton radiographies [3] of the setup, it was possible to accurately locate the animals relative to the beam. To confirm positioning, DNA damage was visualized by immunofluorescent staining of gH2AX in the irradiated mouse brain. Results and conclusion: Proton mouse brain irradiation was successfully performed. Distribution of DNA DSB via gH2AX revealed that the proton beam stopped in the beam facing brain hemisphere. The setup enables the comparison to corresponding photon experiments with SAIGRT [4] and clinically relevant long-term experiments, such as measuring cognitive functions and anomalies in imaging, to directly relate potential photon and proton side effects in brain radiotherapy. [1] Helmbrecht et al. J Instrum 2016 [2] Beyreuther et al. IJPT 2018 (accepted) [3] Müller et al. Acta Oncologica 2017 [4] Tillner et al. Phys Med Biol 2016

The prognosis of patients with esophageal carcinoma remains dismal despite ongoing efforts to improve treatment options. For locally advanced tumors, several randomized trials have shown the benefit of neoadjuvant chemoradiation followed by surgery compared to surgery alone. The aim of this exploratory study was to evaluate the prognostic value of different baseline positron emission tomography (PET) parameters and their potentially additional prognostic impact at the end of neoadjuvant radiochemotherapy. Furthermore, the standard uptake ratio (SUR) as a new parameter for quantification of tumor metabolism was compared to the conventional PET parameters metabolic active volume (MTV), total lesion glycolysis (TLG), and standardized uptake value (SUV) taking into account known basic parameters. Methods: 18F-FDG-PET/CT was performed in 76 consecutive patients ((60±10) years, 71 males) with newly diagnosed esophageal cancer before and during the last week of neoadjuvant radiochemotherapy. MTV of the primary tumor was delineated with an adaptive threshold method. The blood SUV was determined by manually delineating the aorta in the low dose CT. SUR values were computed as scan time corrected ratio of tumor SUVmax and mean blood SUV. Univariate Cox regression and Kaplan-Meier analysis with respect to locoregional control (LRC), freedom from distant metastases (FFDM), and overall survival (OS) was performed. Additionally, independence of PET parameters from standard clinical factors was analyzed with multivariate Cox regression. Results: In multivariate analysis two parameters showed a significant correlation with all endpoints: restaging MTV and restaging SUR. Furthermore, restaging TLG was prognostic for LCR and FFDM. For all endpoints the largest effect size was found for restaging SUR. The only basic factors remaining significant in multivariate analyses were histology for OS and FFDM and age for LRC. Conclusion: PET provides independent prognostic information for OS, LRC, and FFDM in addition to standard clinical parameters in this patient cohort. Our results suggest that the prognostic value of tracer uptake can be improved when characterized by SUR rather than by SUV. Overall, our investigation revealed a higher prognostic value of restaging parameters compared to baseline PET; therapy-adjustments would still be possible at this point of time. Further investigations are required to confirm these hypothesis-generating results.

Purpose: To investigate changes in neurocognitive function and quality of life (QoL) and their association with dosimetric parameters of various brain structures as well as clinical cofactors in adult brain tumour patients following proton beam therapy (PBT). Material and methods: Sixty-nine adult patients with primary brain tumours who received conventionally fractionated PBT were included in this study. Neurocognitive function according to the Montreal Cognitive Assessment (MoCA) test and QoL according to general EORTC-QLQ-C30 and brain tumour specific QLQ-BN20 questionnaires were scored prospectively at baseline and within 3-month-intervals up to one year after PBT. Dose-volume parameters of the retrospectively contoured structures hippocampus, thalamus, frontal and temporal lobes, amygdala, entire cerebellum, anterior cerebellum, and posterior cerebellum were extracted. Clinical parameters comprised age, sex, diagnosis and WHO grading, tumour volume, prescribed dose, concomitant chemotherapy, tumour resection and administration of corticosteroids. MoCA scores and differences to baseline values at different time points were correlated with self-reported QoL items (Spearman correlation rs), clinical and dosimetric parameters (Mann-Whitney U test, logistic regression). A change of ≥3 points of the MoCA total score compared to baseline was considered clinically relevant. Unless otherwise stated, differences at 3 months after PBT compared to baseline are given. Results: The MoCA total score remained stable over time for the majority of patients: Less than 10% of the patients had clinically relevant changes at respective time points. The QLQ-C30 items did not change over time. On the QLQ-BN20 symptom scale, significant increases were observed for the items hair loss (p=0.002) and seizure (p

Objectives: Clinically relevant animal models of non-small cell lung carcinoma (NSCLC) are required for the validation of novel treatments. We compared two different orthotopic transplantation techniques as well as imaging modalities to identify suitable mouse models mimicking clinical scenarios. Methods: We used three genomically diverse NSCLC cell lines (NCI-H1703 adenosquamous cell carcinoma, NCI-H23 adenocarcinoma and A549 adenocarcinoma) for implanting tumour cells either as spheroids or cell suspension into lung parenchyma. Bioluminescence imaging (BLI) and contrast-enhanced cone beam computed tomography (CBCT) were performed twice weekly to monitor tumour growth. Tumour histological data and microenvironmental parameters were determined. Results: Tumour development after spheroid-based transplantation differs probably due to the integrity of spheroids, as H1703 developed single localized nodules, whereas H23 showed diffuse metastatic spread starting early after transplantation. A549 transplantation as cell suspension with the help of a stereotactic system was associated with initial single localized tumour growth and eventual metastatic spread. Imaging techniques were successfully applied to monitor longitudinal tumour growth: BLI revealed highly sensitive qualitative data, whereas CBCT was associated with less sensitive quantitative data. Histology revealed significant model dependent heterogeneity in proliferation, hypoxia, perfusion and necrosis. Conclusions: Our developed orthotopic NSCLC tumours have similarity with biological growth behavior similar to that seen in the clinic and could therefore be used as attractive models to study tumour biology and evaluate new therapeutic strategies. The use of human cancer cell lines facilitates testing of different genomic tumor profiles that may affect treatment outcomes. Advances in knowledge: The combination of different imaging modalities and orthotopic transplantation techniques pave the way towards representative preclinical NSCLC models for experimental testing of novel therapeutic options in future studies.

Introduction: The Small Animal Image-Guided Radiation Therapy (SAIGRT) system has been developed at OncoRay by downsizing treatment technology of human radiation oncology towards small animal experiments [1,2]. The system consists of a gantry module, covered by a radiation protection housing, and a supply module. The gantry module comprises (1) a stationary unit incorporating a 3D computerised animal stage positioner and (2) a 360° rotating arm holding a 225 kV X-ray tube as radiation source, a system of flat aperture collimators for beam shaping and a flat-panel detector for X-ray imaging. Supported by dedicated software, the SAIGRT system allows for a precise and accurate, conformal irradiation and X-ray imaging of small animals using an experimental workflow resembling the patient treatment process. However, only simplified treatment planning has been applied so far, in lack of a full 3D treatment planning system (TPS). Materials & Methods: RayStation 5 is an advanced, clinical TPS by RaySearch Laboratories AB, which works across different external beam radiotherapy devices (e.g. linear accelerators, proton therapy). Import / export options are available for various image modalities. Furthermore, it provides versatile manual and automatic tools for contouring as well as different methods for rigid and deformable image registration. Various irradiation devices can be modelled by specifying characteristics of design features and beam shaping components (e.g. material, motion, geometry) as well as dosimetric properties of the irradiation field. For plan design, the software comprises machine-dependent functions and dose calculation engines resulting in realistic dose distributions. In addition, different features for plan evaluation and comparison are accessible. However, all tools are intended for patients and thereby not suitable for the submillimetre dimensions of small animals. For this reason, µ-RayStation 5 has been developed in collaboration with RaySearch. Based on a research version of the clinical TPS, functionality has been expanded to comply with the requirements of small animal irradiators such as the SAIGRT system. Results: Tools for contouring and image registration as well as the dose grid have been modified to support dimensions down to 0.1 mm. The machine model for small animal irradiators includes characteristics such as the geometry of cone-shaped beams (e.g. distances, aperture sizes), a focal spot model (e.g. 2D Gaussian distribution) and a photon energy spectrum of an X-ray tube with several hundred kV accelerating potential. Plan design is supported for 3D conformal radiotherapy using fixed beams and static arcs. Dose calculation is performed by the VMC++ Monte-Carlo engine [6]. The number of simulated histories and the dose quantity (dose to water or medium) can be selected. Monitor units have been replaced by irradiation time. The SAIGRT system was successfully modelled using a 200 kV spectrum generated from SpekCalc [3-5] showing good agreement with verification measurements. Conclusion: µ-RayStation 5 provides comprehensive functionality of a clinical TPS for small animal studies allowing an efficient experimental workflow for experienced Raystation users. Flexibility of the software facilitates adaption to other small animal irradiators and expansion of usage for preclinical research in our institute especially for commencing small animal proton irradiations. References: [1] Tillner et al. (2014), Z Med Phys. 24(4): 335-51 [2] Tillner et al. (2016), Phys Med Biol. 61(8): 3084-108 [3] Poludniowski GG, Evans PM (2007), Med Phys. 34(6): 2164-74 [4] Poludniowski GG (2007), Med Phys. 34(6): 2175-86 [5] Poludniowski GG et al. (2009), Phys Med Biol. 54(19): 433-8 [6] Kawrakow I, Fippel M (2000), In: The Use of Computers in Radiation Therapy, Springer: 126-8

Proton therapy holds the potential to spare tumor-surrounding tissue and is therefore frequently used in brain tumor treatment. Clear indications from in vitro experiments show that the clinically applied relative biological effectiveness of 1.1 is higher towards the distal edge of the proton Bragg peak. Due to planning margins and range uncertainties the Bragg peak is typically placed in normal tissue and might increase neurological toxicities. Clinical evidence for radiation induced brain necrosis is yet scarce, nonetheless, needs critical evaluation. Sophisticated in vivo studies might be able to close this knowledge gap when designed to mirror the clinical exposure scenario. Here, the setup for in vivo experiments together with the first results of mouse brain irradiation at the experimental beam line [1] of the UPTD will be presented. Absolute dosimetry in treatment position was done with ionization chambers and EBT3 radiochromic films. The anesthetized mice were fixed in an in-house developed multi-modality bed suitable for imaging and irradiation. Following CT for target delineation and proton radiography [2] for treatment positioning, mice were irradiated with a lateral collimated proton beam of 2 mm in diameter to the proximal hemisphere of the brain. Following radiation, mouse brains were excised and analyzed for DNA and tissue damage. Matching photon experiments with SAIGRT [3] are currently underway allowing in the future a direct comparison of treatment related side effects in the brain. [1] Helmbrecht et al. J Instrum 2016 [2] Müller et al. Acta Oncologica 2017 [3] Tillner et al. Phys Med Biol 2016

Over the past decade, functional imaging by means of 18F-fluorodeoxyglucose positron emission tomography (FDG-PET/CT) has improved tumor staging and treatment planning leading to somewhat higher survival rates, in particular in NSCLC patients. This review focuses on the recent insight gained and at current challenges encountered while pursuing improved outcome in patients suffering from NSCLC or SCLC. © 2018, Italian Association of Nuclear Medicine and Molecular Imaging.

Background: The poor prognosis of patients suffering from Glioblastoma multiforme (GBM) is mainly basedon therapy resistances of GBM stem- and tumor bulk cells and their invasive growth within the brain. Neglected are therapy-induced adaptation mechanisms. Here, we blocked bypass mechanisms simultaneously to radiochemotherapy by targeting the pro-survival beta1 integrins and the stress-related c-Jun N-terminal kinases (JNK) and evaluated the effectiveness of this strategy on GBM radiochemosensitization and invasion in vitro and in vivo. Methods: An Oncomine database analysis was conducted to compare the expression of JNK, beta1 integrin and collagen type-I in GBM and brain. The clonogenic survival and the invasion of human GBM cell lines (U343-MG, T4), GBM stem-like (GS-8) and patient-derived cells (DK32, DK41) was quantified upon irradiation (0-6 Gy X-ray) in 2- and 3-dimensional collagen type-I matrix. On top of this treatment, beta1 integrins (AIIB2) and JNK (SP600125, siRNA) were inhibited in a single or dual manner. The effect of a combined beta1 integrin/JNK inhibition on tumor growth and survival was evaluated in orthotopic GBM mice treated with radiochemotherapy. Furthermore, underlying changes of cellular signaling cascades (phosphoproteome array), cell cycle (FACS), DNA damage (53BP1) and chromatin organization were evaluated upon beta1 integrin/JNK co-targeting. Results: Oncomine data showed an increased expression of beta1 integrins and collagen type-I in GBM. While neither a single inhibition of beta1 integrins nor JNK reduced cell survival, co-targeting of both molecules induced radiosensitization and blocked cell invasion in all GBM cell populations tested. This treatment effect was promoted by an increased expression of pro-survival beta1 integrin upon JNK inhibition. Moreover, in combination with radiochemotherapy, beta1 integrin/JNK co-inhibition significantly delayed tumor growth in vivo leading to a significant longer survival of orthotopic GBM mice. Mechanistically, the radiosensitization upon beta1 integrin/JNK targeting was attributed to an amplified ATM phosphorylation and G2/M cell cycle arrest, which was accompanied by an increase in 53BP1 foci and euchromatin formation. Conclusion: Our data show that a combined deactivation of beta1 integrin/JNK efficiently targets adaptation mechanisms and reduces GBM radiochemoresistance and invasion. Further understanding of therapy-induced bypass mechanisms is key for therapy optimization for GBM and other malignancies.

Glioblastoma multiforme (GBM) is the most common brain tumor in adults and characterized by poor clinical outcome due to genetic and epigenetic alterations in resistance-mediating genes and destructive infiltration into the normal brain. Upon therapy, malignant tumors show adaptation to maintain their homeostasis. Two critical determinants of this adaptation process are cell adhesion by beta1 integrins and stress signaling via c-Jun N-terminal kinases (JNK). Here, we evaluated the potential of simultaneous beta1 integrin/JNK targeting to overcome GBM adaptation controlling radiochemoresistance and invasion. Comparative Oncomine data base analysis was performed on the expression of JNK1/2/3 isoforms, beta1 integrin and its ligands in GBM with normal brain. Different human GBM cell populations (patient-derived, stem-like, established) were analyzed for sphere formation, clonogenicity, 3D collagen type-1 invasion, cell cycling, chromatin organization, DNA double strand break (DSB) repair (γH2AX foci assay), broad-spectrum phosphoproteome analysis, FACS analysis and protein expression/phosphorylation upon irradiation (0-6 Gy X-rays) and chemotherapy (Temozolomide) with and without single and simultaneous inhibition of beta1 integrin (AIIB2) and JNK (SP600125, JNKi). The radiochemosensitizing potential of AIIB2/JNKi was also investigated in an orthotopic GBM mouse model using stem-like cells. In contrast to JNK isoforms, beta1 integrin and col1 showed significant overexpression in GBM compared with normal brain. While single inhibition of beta1 integrin and JNK mediated cytotoxicity, only combined targeting resulted in radiochemosensitization. Intriguingly, double AIIB2/JNKi treatment abrogated GBM cell invasion. Importantly, dual beta1 integrin/JNK inhibition elicited a significant reduction in tumor growth and longer survival of mice concomitantly treated with radiotherapy/Temozolomide. Mechanistically, JNK blocking induced beta1 integrin expression for stimulating diverse signaling pathways controlling cell cycling, invasion and radiochemosensitivity. Radiosensitization by AIIB2/JNKi is caused by enhanced ATM phosphorylation and prolonged G2/M cell cycle arrest as well as impaired DNA double strand break repair in the context of elevated levels of euchromatin. In summary, our data reveal that dual beta1 integrin/JNK targeting efficiently impairs adhesion and stress-related adaptation mechanisms involved in radiochemoresistance and invasion. More in-depth evaluation is warranted to clarify the potential of this kind of beta1 integrin/JNK multi-targeting strategy administrated concomitantly to standard radiochemotherapy in patients suffering from GBM.

In this abstract, we present an experimental setup that allows for multi-modal, cross-platform imaging of small animals as well as image-guided proton- and photon irradiation under laboratory conditions. The setup consists of two units: A primary bedding unit which holds the animal and which is equipped with a breathing mask for inhalation anesthesia, an inlet for warm air and a breathing sensor. The primary unit was designed to meet the demands of the various imaging (magnetic resonance imaging, positron emission tomography, computed tomography, proton radiography,) and treatment modalities (photon- and proton irradiation). The bedding unit can be mounted inside a container, which is designed to maintain pathogen-free conditions outside designated animal laboratory facilities. The second, peripheral unit comprises a heating module, several sensors and read-out electronics to control and monitor temperature as well as vital signs. Moreover, it allows for remote emergency intervention (e.g. oxygen flush) during the animal’s anesthesia. The setup has currently been tested for proton irradiation in an experimental area. A method was implemented to perform on-line position verification by proton radiography. The presented setup features multiple advantages for combined, multi-modal treatment which is of special importance for the monitoring and treatment planning of experimental tumor models. In particular, orthotopic tumors which require accurate imaging – the modality of which can be chosen based upon tissue and treatment – and treatment planning. It satisfies the various modalities’ requirements, hence allowing for one combined workflow. Moreover, image analysis is strongly simplified as multi-modal images can be co-registered without sophisticated techniques.

Purpose Early side effects including oesophagitis are potential prognostic factors in patients undergoing radiochemotherapy (RCT) for locally advanced oesophageal cancer (LAEC). We assessed the prognostic value of 18F-fluorodeoxyglucose (FDG) uptake within irradiated non-tumour-affected oesophagus (NTO) during restaging positron emission tomography (PET) as a surrogate for inflammation/oesophagitis. Methods This retrospective evaluation included 64 patients with LAEC who had completed neoadjuvant RCT and had successful oncological resection. All patients underwent FDG PET/CT before and after RCT. In the restaging PET scan maximum and mean standardized uptake values (SUVmax, SUVmean) were determined in the tumour and NTO. Univariate Cox regression with respect to overall survival, local control, distant metastases and treatment failure was performed. Independence of clinically relevant parameters was tested in a multivariate Cox regression analysis. Results Increased FDG uptake, measured in terms of SUVmean in NTO during restaging was significantly associated with complete pathological remission (p = 0.002) and did not show a high correlation with FDG response of the tumour (rho < 0.3). In the univariate analysis, increased SUVmax and SUVmean in NTO was associated with improved overall survival (p = 0.011, p = 0.004), better local control (p = 0.051, p = 0.044), a lower rate of treatment failure (p < 0.001 for both) and development of distant metastases (p = 0.012, p = 0.001). In the multivariate analysis, SUVmax and SUVmean in NTO remained a significant prognostic factor for treatment failure (p < 0.001, p = 0.004) and distant metastases (p = 0.040, p = 0.011). Conclusions FDG uptake in irradiated normal tissues measured on restaging PET has significant prognostic value in patients undergoing neoadjuvant RCT for LAEC. This effect may potentially be of use in treatment personalization.

BACKGROUND: In early-stage non-small cell lung cancer (NSCLC) without affected lymph nodes detected at staging, surgical resection is still the mainstay of treatment. However, in patients with metastatic mediastinal lymph nodes (pN2) or non-radically resected primary tumors (R1/R2), postoperative radiotherapy (possibly combined with chemotherapy) is indicated. So far, investigations about time factors affecting postoperative radiotherapy have only examined the waiting time defined as interval between surgery and start of radiotherapy, but not the overall treatment time (OTT) itself. Conversely, results from trials on primary radio(chemo)therapy in NSCLC show that longer OTT correlates with significantly worse local tumor control and overall survival rates. This time factor of primary radio(chemo)therapy is thought to mainly be based on repopulation of surviving tumor cells between irradiation fractions. It remains to be elucidated if such an effect also occurs when patients with NSCLC are treated with postoperative radiotherapy after surgery (and chemotherapy). Our own retrospective data suggest an advantage of shorter OTT also for postoperative radiotherapy in this patient group. METHODS/DESIGN: This is a multicenter, prospective randomized trial investigating whether an accelerated course of postoperative radiotherapy with photons or protons (7 fractions per week, 2 Gy fractions) improves locoregional tumor control in NSCLC patients in comparison to conventional fractionation (5 fractions per week, 2 Gy fractions). Target volumes and total radiation doses will be stratified in both treatment arms based on individual risk factors. DISCUSSION: For the primary endpoint of the study we postulate an increase in local tumor control from 70% to 85% after 36 months. Secondary endpoints are overall survival of patients; local recurrence-free and distant metastases-free survival after 36 months; acute and late toxicity and quality of life for both treatment methods.

Primary radiotherapy is the treatment of choice for patients with locally advanced non-small cell lung carcinoma where surgery is unable to perform. Orthotopic tumor models, where tumor material is transplanted into the corresponding organ of origin, are used in preclinical studies to predict the clinical efficacy of newly developed treatment options. For radiooncological research, orthotopic lung tumor models should have the characteristics of growing locally and spread in a manner that resembles the growth and metastasis of a real clinical situation. In light of these respects, luciferases expressing human lung carcinoma cells (A549) were used for orthotopic transplantation. Different transplantation techniques were tested: (i) injection of small tumor pieces (< 1 mm) of subcutaneous source tumors, (ii) percutaneous injection of cell suspensions in matrigel between two ribs or (iii) stereotactic-guided injection of few microliters of cell suspensions with matrigel. Tumor development was imaged twice weekly by optical imaging using IVIS Spectrum and cone beam CT integrated in the Small-Animal Image-Guided Radiotherapy (SAIGRT) platform, developed in our institute. Tumor histology was analyzed via staining with hematoxylin and eosin and human origin of tumors were verified by a specific anti-human Ki-67 antibody. Our experiments revealed a multifocal, early metastatic spreading after percutaneous injection of cell suspensions while the transplantation of small tumor pieces lead to a defined tumor mass after quite a long period of time. In contrast, our new stereotactic approach resulted in early solitary lung lesions mimicking a clinical scenario which will allow us to start image-guided radiotherapy treatment at an appropriate time point. In the future different lung carcinoma cell lines will be orthotopically transplanted using the stereotactic technique and corresponding growth and metastatic potential will be compared in order to improve the selection of models for preclinical radiooncological experiments.

For preclinical cancer research, dedicated small animal imaging and irradiation devices are increasingly gaining importance to examine experimental tumors and normal tissue models. We designed a special small animal bed, which is intended to be used for multimodal imaging (e.g. MRI, CT, PET) and image-guided treatment with different radiation types (e.g. photons, protons). The multimodality small animal bed was constructed with CAD software and produced with a 3D printer using the fused deposition modeling procedure. The bed is made of thin acrylonitrile butadiene styrene (ABS), which is fully compatible with magnetic fields, barely influence radiation and is chemically resistant against most frequently used disinfectants. To facilitate proper positioning and alignment of the animal, the bed contains a tooth bar and ear pins for cranial fixation as well as a detachable distal foot holder for the hind leg. For inhalation anesthesia, a respective mask is integrated. To avoid hypothermia, the bed is supplied with a controllable stream of HEPA-filtered warm air. Furthermore, the respiration of the animal can be monitored during the experiment using a respiratory cushion. Suitability of the prototype was tested for photon and proton irradiation as well as CT imaging and proton radiography. In conclusion, the designed multimodality bed enables standardized positioning of small animals and provides integrated solutions for their anesthesia, warming and monitoring. Ist use for multimodal monitoring and image-guided irradiation of experimental tumors will simplify the workflow and image analyses for preclinical radiooncological investigations.