M.E.P. Philippens

DNA double-strand break repair in parental chromatin of mouse zygotes, the first cell cycle as an origin of de novo mutation

In the human, the contribution of the sexes to the genetic load is dissimilar. Especially for point mutations, expanded simple tandem repeats and structural chromosome mutations, the contribution of the male germline is dominant. Far less is known about the male germ cell stage(s) that are most vulnerable to mutation contraction. For the understanding of de novo mutation induction in the germline, mechanistic insight of DNA repair in the zygote is mandatory. At the onset of embryonic development, the parental chromatin sets occupy one pronucleus (PN) each and DNA repair can be regarded as a maternal trait, depending on proteins and mRNAs provided by the oocyte. Repair of DNA double-strand breaks (DSBs) is executed by non-homologous end joining (NHEJ) and homologous recombination (HR). Differentiated somatic cells often resolve DSBs by NHEJ, whereas embryonic stem cells preferably use HR. We show NHEJ and HR to be both functional during the zygotic cell cycle. NHEJ is already active during replacement of sperm protamines by nucleosomes. The kinetics of G1 repair is influenced by DNA-PK(cs) hypomorphic activity. Both HR and NHEJ are operative in S-phase, HR being more active in the male PN. DNA-PK(cs) deficiency upregulates the HR activity. Both after sperm remodeling and at first mitosis, spontaneous levels of gammaH2AX foci (marker for DSBs) are high. All immunoflurescent indices of DNA damage and DNA repair point at greater spontaneous damage and induced repair activity in paternal chromatin in the zygote.

Validation of functional imaging with pathology for tumor delineation in the prostate

INTRODUCTION A study was performed to validate magnetic resonance (MR) based prostate tumor delineations with pathology. MATERIAL AND METHODS Five patients with biopsy proven prostate cancer underwent a T2 weighted (T2w), diffusion weighted MRI (DW-MRI) and dynamic contrast-enhanced MRI (DCE-MRI) scan before prostatectomy. Suspicious regions were delineated based on all available MR information. After prostatectomy whole-mount hematoxylin-eosin stained (H&E) sections were made. Tumor tissue was delineated on the H&E stained sections and compared with the MR based delineations. The registration accuracy between the MR images and H&E stained sections was estimated. RESULTS A tumor coverage of 44-89% was reached by the MR based tumor delineations. The application of a margin of approximately 5mm to the MR based tumor delineations yielded a tumor coverage of 85-100% in all patients. Errors created during the registration procedure were 2-3mm, which cannot completely explain the limited tumor coverage. CONCLUSIONS An accurate tissue processing and registration method was presented (registration error 2-3mm), which enables the validation of MR based tumor delineations with pathology. Reasonable tumor coverage of about 85% and larger was found when applying a margin of approximately 5 mm to the MR based tumor delineations.

A COMPARISON OF DOSE-RESPONSE MODELS FOR THE PAROTID GLAND IN A LARGE GROUP OF HEAD-AND-NECK CANCER PATIENTS

PURPOSE The dose-response relationship of the parotid gland has been described most frequently using the Lyman-Kutcher-Burman model. However, various other normal tissue complication probability (NTCP) models exist. We evaluated in a large group of patients the value of six NTCP models that describe the parotid gland dose response 1 year after radiotherapy. METHODS AND MATERIALS A total of 347 patients with head-and-neck tumors were included in this prospective parotid gland dose-response study. The patients were treated with either conventional radiotherapy or intensity-modulated radiotherapy. Dose-volume histograms for the parotid glands were derived from three-dimensional dose calculations using computed tomography scans. Stimulated salivary flow rates were measured before and 1 year after radiotherapy. A threshold of 25% of the pretreatment flow rate was used to define a complication. The evaluated models included the Lyman-Kutcher-Burman model, the mean dose model, the relative seriality model, the critical volume model, the parallel functional subunit model, and the dose-threshold model. The goodness of fit (GOF) was determined by the deviance and a Monte Carlo hypothesis test. Ranking of the models was based on Akaikes information criterion (AIC). RESULTS None of the models was rejected based on the evaluation of the GOF. The mean dose model was ranked as the best model based on the AIC. The TD(50) in these models was approximately 39 Gy. CONCLUSIONS The mean dose model was preferred for describing the dose-response relationship of the parotid gland.

Pathologic Validation of a Model Based on Diffusion-Weighted Imaging and Dynamic Contrast-Enhanced Magnetic Resonance Imaging for Tumor Delineation in the Prostate Peripheral Zone

PURPOSE For focal boost strategies in the prostate, the robustness of magnetic resonance imaging-based tumor delineations needs to be improved. To this end we developed a statistical model that predicts tumor presence on a voxel level (2.5×2.5×2.5 mm3) inside the peripheral zone. Furthermore, we show how this model can be used to derive a valuable input for radiotherapy treatment planning. METHODS AND MATERIALS The model was created on 87 radiotherapy patients. For the validation of the voxelwise performance of the model, an independent group of 12 prostatectomy patients was used. After model validation, the model was stratified to create three different risk levels for tumor presence: gross tumor volume (GTV), high-risk clinical target volume (CTV), and low-risk CTV. RESULTS The model gave an area under the receiver operating characteristic curve of 0.70 for the prediction of tumor presence in the prostatectomy group. When the registration error between magnetic resonance images and pathologic delineation was taken into account, the area under the curve further improved to 0.89. We propose that model outcome values with a high positive predictive value can be used to define the GTV. Model outcome values with a high negative predictive value can be used to define low-risk CTV regions. The intermediate outcome values can be used to define a high-risk CTV. CONCLUSIONS We developed a logistic regression with a high diagnostic performance for voxelwise prediction of tumor presence. The model output can be used to define different risk levels for tumor presence, which in turn could serve as an input for dose planning. In this way the robustness of tumor delineations for focal boost therapy can be greatly improved.

Simultaneous MRI diffusion and perfusion imaging for tumor delineation in prostate cancer patients

BACKGROUND AND PURPOSE A study was performed to investigate if we can quantify if the two imaging modalities diffusion weighted imaging (DWI) and dynamic contrast-enhanced (DCE)-MRI are consistent in what voxels they determine as being suspicious of tumor tissue. MATERIAL AND METHODS Twenty-one patients with biopsy proven prostate cancer underwent a DWI and a DCE-MRI scan. These scans were compared using a receiver operating curve (ROC) analysis, where either one of the two imaging modalities was thresholded and taken as a reference. The resulting area under the curve (AUC) reflects the consistency between target delineations based on the two imaging techniques. This analysis was performed for the complete prostate and the peripheral zone (PZ). RESULTS Consistency between DWI and DCE-MRI parameter maps varied greatly between patients. Values of the AUC up to 0.90 were found. However, on average AUC values were 0.60. The AUC values were related to the patients PSA and clinical stage. CONCLUSIONS Large variation in consistency between the two imaging modalities was found. This did not depend on the precise thresholds used. For making decisions on dose painting in the prostate, the knowledge about the inconsistency must be taken into account.

Parp1–XRCC1 and the repair of DNA double strand breaks in mouse round spermatids

The repair of DNA double strand breaks (DSBs) in male germ cells is slower and differently regulated compared to that in somatic cells. Round spermatids show DSB repair and are radioresistant to apoptosis induction. Mutation induction studies using ionizing irradiation, indicated a high frequency of chromosome aberrations (CA) in the next generation. Since they are in a G1 comparable stage of the cell cycle, haploid spermatids are expected to repair DSBs by the non-homologous end-joining pathway (NHEJ). However, immunohistochemical evidence indicates that not all components of the classical NHEJ pathway are available since the presence of DNA-PKcs cannot be shown. Here, we demonstrate that round spermatids, as well as most other types of male germ cells express both Parp1 and XRCC1. Therefore, we have determined whether the alternative Parp1/XRCC1 dependent NHEJ pathway is active in these nuclei and also have tested for classical NHEJ activity by a genetic method. To evaluate DSB repair in SCID mice, deficient for DNA-PKcs, and to study the involvement of the Parp1/XRCC1 dependent NHEJ pathway in round spermatids, the loss of gamma-H2AX foci after irradiation has been determined in nucleus spreads of round spermatids of SCID mice and in nucleus spreads and histological sections of Parp1-inhibited mice and their respective controls. Results show that around half of the breaks in randomly selected round spermatids are repaired between 1 and 8h after irradiation. The repair of 16% of the induced DSBs requires DNA-PKcs and 21% Parp1. Foci numbers in the Parp1-inhibited testes tend to be higher in spermatids of all epithelial stages reaching significance in stages I-III which indicates an active Parp1/XRCC1 pathway in round spermatids and a decreased repair capacity in later round spermatid stages. In Parp1-inhibited SCID mice only 14.5% of the breaks were repaired 8h after irradiation indicating additivity of the two NHEJ pathways in round spermatids.

Dynamic contrast enhanced MR imaging for rectal cancer response assessment after neo-adjuvant chemoradiation

Patient selection for organ sparing treatment after good response to neo‐adjuvant chemoradiation (CRT) for locally advanced rectal cancer is challenging as no optimal restaging modality is available after CRT. In this study, we assessed the value of dynamic contrast enhanced magnetic resonance imaging (DCE‐MRI) for rectal cancer pathological response prediction.

Investigating the non-linearity of the BOLD cerebrovascular reactivity response to targeted hypo/hypercapnia at 7 T

Cerebrovascular reactivity (CVR) is a mechanism responsible for maintaining stable perfusion pressure within the brain via smooth muscle mediated modulations of vascular tone. The amplitude of cerebral blood flow (CBF) change in response to a stimulus has been evaluated using Blood Oxygen Level Dependent (BOLD) MRI, however the relationship between the stimulus and the measured signal remains unclear. CVR measured invasively in animal models and using blood-velocity based measurements in humans has demonstrated a sigmoidal relationship between cerebral blood flow and CO2 partial pressure. Using an ultra-high magnetic field strength (7T) MRI scanner and a computer controlled gas delivery system, we examined the regional and voxel-wise CVR response in relation to a targeted progressively increasing hypo- to hypercapnic stimulus. The aim of this study was to assess the non-linearity/sigmoidal behavior of the CVR response at varying arterial CO2 (PaCO2) levels. We find that a sigmoidal model provides a better description of the BOLD signal response to increasing PaCO2 than a linear model. A distinct whole-brain and gray matter BOLD-CVR signal plateau was observed in both voxel-wise and regional analysis. Furthermore, we demonstrate that a progressively increasing stimulus in combination with a sigmoidal response model can be used to obtain CVR values and provides additional physiologically relevant information (such as linear and non-linear response domains, and maximum response amplitudes) that may be more difficult to obtain from blocked CVR experiments. Considering these results, we propose an alternative way in which to define CVR based on the derivative of the BOLD-CVR response curve, which can potentially be used to differentiate between healthy and diseased vascular states.

Combined T2w volumetry, DW-MRI and DCE-MRI for response assessment after neo-adjuvant chemoradiation in locally advanced rectal cancer.

Background. To assess the value of combined T2-weighted magnetic resonance imaging (MRI) (T2w) volumetry, diffusion-weighted (DW)-MRI and dynamic contrast enhanced (DCE)-MRI for pathological response prediction after neo-adjuvant chemoradiation (CRT) in locally advanced rectal cancer (LARC). Material and methods. MRI with DW-MRI and DCE-MRI sequences was performed before start of CRT and before surgery. After surgery, the tumor regression grade (TRG) was obtained based on the score by Mandard et al. Pathological complete responders (pCR, TRG 1), and pathological good responders (GR, TRG 1 + 2) were compared to non-pCR and non-GR patients, respectively. Results. In total 55 patients were analyzed, six had a pCR (10.9%) and 10 a GR (18.2%). Favorable responders had a larger decrease in tumor volume and Ktrans and a larger increase in apparent diffusion coefficient (ADC) values compared to non-responders. ADC change showed the best diagnostic accuracy for pCR. For GR, the model including ADC change and volume change showed the best diagnostic performance. However, this performance was not statistically better compared to the model with ADC change alone. Inclusion of Ktrans change did not increase the diagnostic accuracy for pathological favorable response. Conclusions. This explorative study showed that ADC change is a promising diagnostic tool for pCR and GR. Volume decrease showed potential limited additional diagnostic value for GR while Ktrans change showed no additional diagnostic value for pCR and GR.

Diffusion weighted MRI in head-and-neck cancer: geometrical accuracy.

INTRODUCTION The aim of this study is to assess the geometric accuracy of diffusion weighted (DW)-MRI by quantification of geometric distortions in the gross tumor volume (GTV) in head and neck (HN) cancer. MATERIALS & METHODS A retrospective analysis was performed on the data of 23 patients (with 24 lesions). For these patients, magnetic field maps and DW-MRI were acquired. The magnetic field maps were converted to voxel displacement maps. GTV delineations were transferred onto these voxel displacement maps and the voxel shifts in the GTV were analyzed. RESULTS The median shift was 3.2mm and the maximal posterior and anterior shifts were up to 15.0 and 26.0mm respectively. The range of shifts varied from 11.8 to 25.6mm. The percentage of GTV voxels that showed a shift of at least 6mm was found to be 23.2%. CONCLUSIONS Current DW-MRI images of HN tumors show severe distortions up to centimeters, which restrict the use of DW-MRI scans for GTV definition in RT treatment planning.