Qualitative Assessment of Titanium versus Carbon Fiber/Polyetheretherketone Pedicle Screw-Related Artifacts: A Cadaveric Study.

BACKGROUND: Dorsal instrumentation and decompression are the mainstays of spinal tumor treatment. Replacing titanium screws with carbon fiber-reinforced polyetheretherketone (CFRP) screws can reduce imaging artifacts on neural structures and perturbations of radiation dose. Further reduction of metal content in such screws might enhance the benefit. The aim of this study was to assess the artifacts produced by all-titanium screws (Ti-Ti), CFRP thread-titanium screw heads (C-Ti), and all-CFRP screws (C-C). METHODS: A cadaveric spine was used to place Ti-Ti, C-Ti, and C-C consecutively from T2 to S1. Computed tomography and 1.5T and 3T magnetic resonance imaging were performed for each screw system. Axial T1- and T2-weighted sequences of representative thoracic and lumbar regions were assessed for artifacts. The artifacts were classified as not relevant, considerable, or severe. RESULTS: We evaluated 92 screws and made 178 artifact assessments. The artifacts were clearly visible in computed tomography scans but did not influence the visualization of intraspinal structures. Severe magnetic resonance imaging artifacts were found in 28% (17/60, mostly in the thoracic spine) of Ti-Ti, 2% (1/60, all T1 sequences) of C-Ti, and 0% of C-C, and considerable artifacts were found in 47% (28/60) of Ti-Ti, 10% (6/60, only 1 T2 sequence) of C-Ti, and 0% of C-C screws (P < 0.001). CONCLUSIONS: CFRP pedicle screws reduced the artifact intensity in spinal structures compared with titanium screws, and may be beneficial for planning radiotherapy and for follow-up imaging. C-C demonstrated an enhanced effect on dorsal structures.

Quality of life in lung cancer survivors treated with tyrosine-kinase inhibitors (TKI): results from the multi-centre cross-sectional German study LARIS.

PURPOSE: We aimed at exploring the quality of life (QOL) of lung cancer survivors with proven tyrosine-kinase receptor (RTK) genetic alterations and targeted tyrosine-kinase inhibitors (TKI) therapy, compared to lung cancer survivors with no-RTK alterations and no-TKI therapy. METHODS: Data were collected in a cross-sectional multi-centre study. Primary lung cancer survivors were asked about their socio-demographic and clinical information, QOL, symptom burden, and distress. QOL and symptom burden were assessed using the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire Core 30 (EORTC QLQ-C30), and distress with the Patient Health Questionnaire-4 (PHQ-4). Demographic and clinical characteristics were reported in absolute and relative frequencies, QOL, and symptom burden using mean scores. Differences in mean scores with relative 95% confidence intervals were used for comparison. RESULTS: Three groups of survivors were defined: group A with proven RTK alterations, TKI therapy at any time during therapy, and stage IV lung cancer at diagnosis (n = 49); group B: non-TKI therapy and stage IV lung cancer (n = 121); group C: non-TKI therapy and stage I-III lung cancer (n = 495). Survivors in group A reported lower QOL (mean score difference = -11.7 vs. group B) and symptom burden for dyspnoea (difference = -11.5 vs. group C), and higher symptom burden for appetite loss (difference = + 11.4 vs. group C), diarrhoea and rash (differences = + 25.6, + 19.6 and + 13.2, + 13.0, respectively, vs. both groups). CONCLUSIONS: Our results suggest that the specific side effects of TKI therapy can impair QOL among lung cancer survivors. Therefore, specific focus towards the optimal management of these side effects should be considered.

Heart atlas for retrospective cardiac dosimetry: a multi-institutional study on interobserver contouring variations and their dosimetric impact.

PURPOSE: Cardiac effects after breast cancer radiation therapy potentially affect more patients as survival improves. The heart's heterogeneous radiation exposure and composition of functional structures call for establishing individual relationships between structure dose and specific late effects. However, valid dosimetry requires reliable contouring which is challenging for small volumes based on older, lower-quality computed tomography imaging. We developed a heart atlas for robust heart contouring in retrospective epidemiologic studies. METHODS AND MATERIALS: The atlas defined the complete heart and geometric surrogate volumes for six cardiac structures: aortic valve, pulmonary valve, all deeper structures combined, myocardium, left anterior myocardium, and right anterior myocardium. We collected treatment planning records from 16 patients from 4 hospitals including dose calculations for 3D conformal tangential field radiation therapy for left-sided breast cancer. Six observers each contoured all patients. We assessed spatial contouring agreement and corresponding dosimetric variability. RESULTS: Contouring agreement for the complete heart was high with a mean Jaccard similarity coefficient (JSC) of 89%, a volume coefficient of variation (CV) of 5.2%, and a mean dose CV of 4.2%. The left (right) anterior myocardium had acceptable agreement with 63% (58%) JSC, 9.8% (11.5%) volume CV, and 11.9% (8.0%) mean dose CV. Dosimetric agreement for the deep structures and aortic valve was good despite higher spatial variation. Low spatial agreement for the pulmonary valve translated to poor dosimetric agreement. CONCLUSIONS: For the purpose of retrospective dosimetry based on older imaging, geometric surrogate volumes for cardiac organs at risk can yield better contouring agreement than anatomical definitions, but retain limitations for small structures like the pulmonary valve.

Impact of radiotherapy protocol adherence in NSCLC patients treated with concurrent chemoradiation: RTQA results of the PET-Plan trial.

INTRODUCTION: The success of intensification and personalisation of the curative treatment of non-small cell lung cancer (NSCLC) is strongly associated with the precision in radiotherapy. Here, we evaluate the impact of radiotherapy protocol adherence in a prospective multicentre trial. METHODS: In the open-label, randomised, controlled PET-Plan trial, patients with inoperable NSCLC were randomized at a 1:1 ratio regarding the target volume delineation informed by 1F-FDG PET and CT plus elective nodal irradiation (arm A) or target volumes informed by PET alone (arm B) and received iso-toxically dose-escalated concurrent chemoradiation. The prospectively organised quality assurance program (RTQA) included individual case review by predefined criteria. For evaluation, protocol adherence was scored as per protocol (pP), with minor (miD), intermediate (inD) and major (maD) deviations. In order to exclude biases through patients who discontinued treatment, patients who received ≥60 Gy were additionally analysed. RESULTS: Between 05/2009-11/2016, 205 patients were randomized, 204 patients started treatment according to protocol of which 31 (15%) patients had maD. Patients with maD had an inferior overall survival (OS) (HR 2.9, 95% CI 1.8-4.4, p < 0.0001) and a higher risk of loco-regional progression (HR 5.7, 95% CI 2.7-11.1, p < 0.0001). These results were significant also in the subgroup of patients receiving ≥ 60 Gy. Patients with maD concerning normal tissue delineation and/or dose constraints had a worse OS (p = 0.006) although no higher incidence of grade ≥ 3 toxicities. CONCLUSIONS: Non-adherence to the radiotherapy protocol was associated with an inferior OS and loco-regional control. These results underline the importance of RTQA.

Efficacy and Toxicity of Different Chemotherapy Protocols for Concurrent Chemoradiation in Non-Small Cell Lung Cancer-A Secondary Analysis of the PET Plan Trial.

(1) Background: The optimal chemotherapy (CHT) regimen for concurrent chemoradiation (cCRT) is not well defined. In this secondary analysis of the international randomized PET-Plan trial, we evaluate the efficacy of different CHT. (2) Methods: Patients with inoperable NSCLC were randomized at a 1:1 ratio regarding the target volume definition and received isotoxically dose-escalated cCRT using cisplatin 80 mg/m2 (day 1, 22) and vinorelbin 15 mg/m2 (day 1, 8, 22, 29) (P1) or cisplatin 20 mg/m2 (day 1-5, 29-33) and vinorelbin 12.5 mg/m2 (day 1, 8, 15, 29, 36, 43) (P2) or carboplatin AUC1 (day 1-5, 29-33) and vinorelbin 12.5 mg/m2 (day 1, 8, 15, 29, 36, 43) (P3) or other CHT at the treating physician's discretion. (3) Results: Between 05/2009 and 11/2016, 205 patients were randomized and 172 included in the per-protocol analysis. Patients treated in P1 or P2 had a better overall survival (OS) compared to P3 (p = 0.015, p = 0.01, respectively). Patients treated with carboplatin had a worse OS compared to cisplatin (HR 1.78, p = 0.03), but the difference did not remain significant after adjusting for age, ECOG, cardiac function creatinine and completeness of CHT. (4) Conclusions: Carboplatin doublets show no significant difference compared to cisplatin, after adjusting for possibly relevant factors, probably due to existing selection bias.

Pediatric craniospinal irradiation with a short partial-arc VMAT technique for medulloblastoma tumors in dosimetric comparison.

BACKGROUND: This study aimed to contrast four different irradiation methods for pediatric medulloblastoma tumors in a dosimetric comparison regarding planning target volume (PTV) coverage and sparing of organs at risk (OARs). METHODS: In sum 24 treatment plans for 6 pediatric patients were realized. Besides the clinical standard of a 3D-conformal radiotherapy (3D-CRT) treatment plan taken as a reference, volumetric modulated arc therapy (VMAT) treatment plans ("VMAT_AVD" vs. "noAVD" vs. "FullArc") were optimized and calculated for each patient. For the thoracic and abdominal region, the short partial-arc VMAT_AVD technique uses an arc setup with reduced arc-length by 100°, using posterior and lateral beam entries. The noAVD uses a half 180° (posterior to lateral directions) and the FullArc uses a full 360° arc setup arrangement. The prescription dose was set to 35.2 Gy. RESULTS: We identified a more conformal dose coverage for PTVs and a better sparing of OARs with used VMAT methods. For VMAT_AVD mean dose reductions in organs at risk can be realized, from 16 to 6.6 Gy, from 27.1 to 8.7 Gy and from 8.0 to 1.9 Gy for the heart, the thyroid and the gonads respectively, compared to the 3D-CRT treatment method. In addition we have found out a superiority of VMAT_AVD compared to the noAVD and FullArc trials with lower exposure to low-dose radiation to the lungs and breasts. CONCLUSIONS: With the short partial-arc VMAT_AVD technique, dose exposures to radiosensitive OARS like the heart, the thyroid or the gonads can be reduced and therefore, maybe the occurrence of late sequelae is less likely. Furthermore the PTV conformity is increased. The advantages of the VMAT_AVD have to be weighed against the potentially risks induced by an increased low dose exposure compared to the 3D-CRT method.

Imaging-based target volume reduction in chemoradiotherapy for locally advanced non-small-cell lung cancer (PET-Plan): a multicentre, open-label, randomised, controlled trial.

BACKGROUND: With increasingly precise radiotherapy and advanced medical imaging, the concept of radiotherapy target volume planning might be redefined with the aim of improving outcomes. We aimed to investigate whether target volume reduction is feasible and effective compared with conventional planning in the context of radical chemoradiotherapy for patients with locally advanced non-small-cell lung cancer. METHODS: We did a multicentre, open-label, randomised, controlled trial (PET-Plan; ARO-2009-09) in 24 centres in Austria, Germany, and Switzerland. Previously untreated patients (aged older than 18 years) with inoperable locally advanced non-small-cell lung cancer suitable for chemoradiotherapy and an Eastern Cooperative Oncology Group performance status of less than 3 were included. Undergoing 18F-fluorodeoxyglucose (18F-FDG) PET and CT for treatment planning, patients were randomly assigned (1:1) using a random number generator and block sizes between four and six to target volume delineation informed by 18F-FDG PET and CT plus elective nodal irradiation (conventional target group) or target volumes informed by PET alone (18F-FDG PET-based target group). Randomisation was stratified by centre and Union for International Cancer Control stage. In both groups, dose-escalated radiotherapy (60-74 Gy, 2 Gy per fraction) was planned to the respective target volumes and applied with concurrent platinum-based chemotherapy. The primary endpoint was time to locoregional progression from randomisation with the objective to test non-inferiority of 18F-FDG PET-based planning with a prespecified hazard ratio (HR) margin of 1·25. The per-protocol set was included in the primary analysis. The safety set included all patients receiving any study-specific treatment. Patients and study staff were not masked to treatment assignment. This study is registered with ClinicalTrials.gov, NCT00697333. FINDINGS: From May 13, 2009, to Dec 5, 2016, 205 of 311 recruited patients were randomly assigned to the conventional target group (n=99) or the 18F-FDG PET-based target group (n=106; the intention-to-treat set), and 172 patients were treated per protocol (84 patients in the conventional target group and 88 in the 18F-FDG PET-based target group). At a median follow-up of 29 months (IQR 9-54), the risk of locoregional progression in the 18F-FDG PET-based target group was non-inferior to, and in fact lower than, that in the conventional target group in the per-protocol set (14% [95% CI 5-21] vs 29% [17-38] at 1 year; HR 0·57 [95% CI 0·30-1·06]). The risk of locoregional progression in the 18F-FDG PET-based target group was also non-inferior to that in the conventional target group in the intention-to-treat set (17% [95% CI 9-24] vs 30% [20-39] at 1 year; HR 0·64 [95% CI 0·37-1·10]). The most common acute grade 3 or worse toxicity was oesophagitis or dysphagia (16 [16%] of 99 patients in the conventional target group vs 17 [16%] of 105 patients in the 18F-FDG PET-based target group); the most common late toxicities were lung-related (12 [12%] vs 11 [10%]). 20 deaths potentially related to study treatment were reported (seven vs 13). INTERPRETATION: 18F-FDG PET-based planning could potentially improve local control and does not seem to increase toxicity in patients with chemoradiotherapy-treated locally advanced non-small-cell lung cancer. Imaging-based target volume reduction in this setting is, therefore, feasible, and could potentially be considered standard of care. The procedures established might also support imaging-based target volume reduction concepts for other tumours. FUNDING: German Cancer Aid (Deutsche Krebshilfe).

[Monocentric experiences of stereotactic body radiation therapy (SBRT) for advanced hepatocellular carcinomas (BCLC-C)]. / Monozentrische Erfahrungen mit stereotaktischer Bestrahlung (SBRT) beim fortgeschrittenen hepatozellulären Karzinom im Stadium BCLC-C.

Hepatocellular carcinomas (HCCs) are highly malignant primary liver cancers with poor prognosis and limited treatment options in advanced stages of disease. Treatment of HCC requires interdisciplinary discussion and multimodal therapy approaches. Beside established loco-regional and systemic therapies, stereotactic body radiation therapy (SBRT) gained increasing importance over recent years. First results of early clinical studies indicate high rates of local control with a good safety profile. In the present work, we evaluated our single center experiences with SBRT in patients with advanced HCCs.Ten patients with 16 SBRTs were included and retrospectively analyzed in this case collection. All patients presented in advanced tumor stages with vascular invasion and/or metastases, but preserved liver function. Two patients were treated only with SBRT, two after TACE and six patients received SBRT in addition to systemic therapy. In most of the cases SBRT were applied to intrahepatic lesions. Large tumor thromboses, lymph nodes as well as bone metastases were irradiated in one, three and five fractions with a median overall dose of 38 Gy. We observed a good local tumor control with a good safety profile in all cases. No severe complications occurred in combination to sequential as well as additive approach to loco-regional or systemic treatments.In conclusion, our experiences confirm results of early clinical studies indicating safe use and good local control rates also in advanced stages of HCC with preserved liver function.

Radiation-induced vascular changes in the intracranial irradiation field in medulloblastoma survivors: An MRI study.

BACKGROUND AND PURPOSE: While survival times after treatment of medulloblastoma are increasing, little is known about radiochemotherapy (RCT)-induced cerebrovascular changes. High resolution vessel wall imaging (VWI) sequences are an emerging tool for the evaluation of cerebrovascular diseases. We performed VWI in medulloblastoma long-term survivors to screen for late sequelae of RCT. MATERIAL AND METHODS: Twenty-two pediatric medulloblastoma survivors (mean age 25.8 years (10-53 years); 16.3 years (mean) post primary RCT (range 1-45 years)) underwent 2D VWI-MRI. Vessel wall thickening, contrast enhancement and luminal narrowing were analyzed. The findings were correlated with the patients' radiation protocols. RESULTS: Vessel wall changes were observed the intracranial internal carotid artery (ICA) and the vertebrobasilar circulation (VBC) in 14 of 22 patients (63.6%). In multivariate analysis, time after RCT (OR = 1.38, p < 0.05) was strongest independent predictor for development of vessel wall alterations. The dose of radiation was not a relevant predictor. CONCLUSIONS: With longer follow-up time intracranial vessel wall changes are observed more frequently in medulloblastoma survivors. Thus VWI is a useful tool to monitor vessel wall alterations of cranially irradiated patients, creating the prerequisite for further treatment of late sequelae.

Susceptibility-weighted magnetic resonance imaging of cerebrovascular sequelae after radiotherapy for pediatric brain tumors.

BACKGROUND AND PURPOSE: Due to sensitive neuroimaging techniques, cerebrovascular complications such as cerebral microbleeds (CMB) and cerebral cavernous malformations (CCM) are increasingly recognized as considerable late effects after treatment for pediatric brain tumor. The aim of this study was to analyze CMB in a cohort of patients after cranial irradiation therapy for medulloblastoma or other pediatric brain tumors using susceptibility-weighted magnetic resonance imaging (SWI). MATERIALS AND METHODS: Forty former pediatric brain tumor patients were enrolled in this prospective cross-sectional study and examined by cranial MRI including SWI sequences. Cerebral microbleeds, clinical symptoms and disability were evaluated. RESULTS: Thirty-six (90%) of the examined individuals (mean follow-up age 22.2 y; mean follow-up time 13.5 y) were affected by CMB. Longer follow-up time and higher craniospinal irradiation doses correlated with higher total lesion count (p < 0.01). Thirteen patients (32.5%) presented with clinical symptoms. Individuals with CMB were more severely disabled than patients without CMB (p < 0.05). CONCLUSIONS: Cerebrovascular sequelae occur frequently after treatment for pediatric brain tumor. In this study, a remarkable part of pediatric brain tumor patients presents with CMB. As a sign of vascular damage, they can cause clinical symptoms and may correspond to neurocognitive decline. Further studies are needed to standardize MRI protocols and to improve quality of long-term follow-up.

Predicting Heart Dose in Breast Cancer Patients Who Received 3D Conformal Radiation Therapy.

Cardiac late effects are a major health concern for long-term survivors after radiotherapy for breast cancer. Large cohort studies to better understand the exact dose-response relationship require individual estimates of radiation dose to the heart. To predict individual cardiac dose from information that is typically available for all members of a retrospective epidemiological cohort study, 774 female breast cancer patients treated with megavoltage tangential field radiotherapy in 1998-2008 were examined. All dose distributions were calculated using Eclipse with the anisotropic analytical algorithm (AAA) for photon fields and the electron Monte Carlo algorithm for electron boost fields. Based on individual dose volume histograms, the authors calculated absorbed dose in the complete heart as well as in six functional substructures. Statistical models were developed to predict absorbed dose using only covariate information from patients' clinical records on tumor location, patient anatomy and radiotherapy prescription. The out-of-sample prediction error for mean heart dose was 54% (coefficient of variation). The prediction error in functional substructures ranged from 49-68% for mean dose and from 52-86% for extreme dose. The authors conclude that based on a patient sample with exact heart dosimetry, it is possible to use clinical information alone to predict absorbed heart dose in the remaining cohort with a quantified error suitable for dose-response analyses of cardiac late effects.

Radiation dose distribution in functional heart regions from tangential breast cancer radiotherapy.

BACKGROUND AND PURPOSE: To analyze the distribution of individually-determined radiation dose to the heart and its functional sub-structures after radiotherapy in breast cancer patients treated in Germany during 1998-2008. MATERIAL AND METHODS: We obtained electronic treatment planning records for 769 female breast cancer patients treated with megavoltage tangential field radiotherapy. All dose distributions were re-calculated using Eclipse with the anisotropic analytical algorithm (AAA) for photon fields, and the electron Monte Carlo algorithm for electron boost fields. Based on individual dose volume histograms for the complete heart and several functional sub-structures, we estimated various dose measures in patient groups. RESULTS: Mean heart dose spanned a range of 0.9-19.1Gy for left-sided radiotherapy and 0.3-11.6Gy for right-sided radiotherapy. Average (median) mean heart dose was 4.6Gy (3.7Gy) for left-sided radiotherapy, and 1.7Gy (1.4Gy) for right-sided RT. With left-sided radiotherapy, 66% of the patients had 2cm(3) of the complete heart exposed to at least 40Gy. Younger age, higher body mass index, tumor location in a medial quadrant, and presence of a parasternal field were also associated with higher heart dose. CONCLUSION: Tumor location and treatment choices influence cardiac dose with complex interactions. There is considerable variability in heart dose, with dose metrics of different cardiac sub-structures showing different patterns in their dependency on external influences. Dose-response analysis of late cardiac effects after radiotherapy requires detailed individual dosimetry.

Adjuvant temozolomide-based chemoradiotherapy versus radiotherapy alone in patients with WHO III astrocytoma: The Mainz experience.

BACKGROUND: It is currently unclear whether adjuvant therapy for WHO grade III anaplastic astrocytomas (AA) should be carried out as combined chemoradiotherapy with temozolomide (TMZ)--analogous to the approach for glioblastoma multiforme--or as radiotherapy (RT) alone. PATIENTS AND METHODS: A retrospective analysis of data from 90 patients with AA, who were treated between November 1997 and February 2014. Assessment of overall (OS) and progression-free survival (PFS) was performed according to treatment categories: (1) 50%, RT + TMZ according to protocol, (2) 11%, RT + TMZ with dose reduction, (3) 26%, RT alone, and (4) 13%, individualized, primarily palliative therapy. No dose reduction was necessary in the RT alone group. RESULTS: Median OS was 85, 69, and 43 months for treatment categories 1/2, 3, and 4, respectively. These differences were not statistically significant. PFS was 35, 29, 48, and 33 months for categories 1, 2, 3, and 4, respectively; again without significant differences between categories. In a subgroup of 39 patients with known IDH1 R132H status, the presence of this mutation correlated with significantly longer OS (p = 0.01) and PFS (p = 0.002). Complete or partial tumor resection and younger age also correlated with a significantly better prognosis, and this influence persisted in multivariate analysis. In the IDH1 R132H subgroup analysis, only this marker retained an independent prognostic value. DISCUSSION AND CONCLUSION: A general superiority of combined chemoradiotherapy compared to RT alone could not be demonstrated. Biomarkers for predicting the benefits of combination therapy using RT and TMZ are needed for patients with AA.

Strong adverse prognostic impact of hyperglycemic episodes during adjuvant chemoradiotherapy of glioblastoma multiforme.

BACKGROUND: In comparison to normal brain tissue, glioblastomas exhibit significantly increased glucose uptake. Brain edema is a common complication during adjuvant chemoradiotherapy, leading to a requirement for glucocorticoid treatment. Glucocorticoid treatment frequently causes considerable deregulation of blood glucose levels. Therefore, episodes of hyperglycemia may contribute to radio- and/or chemoresistance. PATIENTS AND METHODS: This study comprises a retrospective analysis of the influence of hyperglycemic episodes (HEs) during adjuvant therapy on the overall survival of 106 glioblastoma multiforme patients. RESULTS: The occurrence of one or more deregulated blood glucose value(s) > 10 mM is associated with a reduction in median overall survival from 16.7 to 8.8 months. A significantly poorer overall survival of patients with hyperglycemia could also be detected in subgroup analyses of patients with complete tumor resection and complete treatment according to the EORTC 22891/26891 trial protocol, as well as in a multivariate Cox proportional hazards analysis. A history of diabetes mellitus had no influence on prognosis. DISCUSSION: Our data suggest that the observed negative impact of elevated blood glucose levels on overall survival may not solely be explained by the patients' poorer general condition; the elevated blood glucose concentration itself may play a pathogenetic role. This could be due to increased activity of antioxidant systems, elevated expression of DNA damage response proteins and protection of hypoxic tumor cells against apoptosis combined with hypoxia-mediated radioresistance. CONCLUSION: A possible prognostic impact of elevated blood glucose levels during the period of adjuvant (chemo-) radiotherapy of glioblastoma should be evaluated in a prospective clinical trial.

ΔNp63 expression in four carcinoma cell lines and the effect on radioresistance--a siRNA knockdown model.

OBJECTIVES: This study investigated the expression of ΔNp63α in carcinoma cell lines of the upper aerodigestive tract and their potential influence on radioresistance using a small interfering RNA (siRNA) knockdown approach. MATERIALS AND METHODS: Four carcinoma cell lines were investigated for the expression of the ΔNp63 isoform by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) (0, 24, 48 h) with and without single dose irradiation of 6 Gy. Furthermore, all cell lines were transfected with siRNA against the ΔNp63α isoform over 24 h. Knockdown effectiveness was controlled by qRT-PCR and Western blot. Apoptotic events were evaluated by terminal transferase dUTP nick end labeling (TUNEL) assay and cross-checked by a test for cell viability (WST-1, Roche) over 48 h. RESULTS: All cell lines presented varying expression of the ΔNp63α isoform with and without irradiation. A sufficient knockdown rate was established by siRNA transfection. Knockdown of the ΔNp63 isoform showed an effect on radiation sensitivity proven by an increase of apoptotic events detectable by immunofluorescence (TUNEL assay) and likewise a significant reduction of formazan production (WST-1 test) in three cell lines. CONCLUSIONS: We found overexpression of ΔNp63α with and without irradiation in three cell lines, and the knockdown of ΔNp63α led to increased apoptotic events and fewer viable cells. Thus, the overexpression of ΔNp63α might protect carcinoma cells against irradiation effects. CLINICAL RELEVANCE: The present work supports the hypothesis that protein 63 might serve as a negative predictor for irradiation response and survival in a clinical setting and may be a target for future therapeutic strategies.

Overexpression of p63 is associated with radiation resistance and prognosis in oral squamous cell carcinoma.

BACKGROUND: The tumor suppressor homologue p63 is expressed in basal and parabasal layers of intraoral mucosa. Full length transcripts with transactivational domain (TA forms) present with homology to p53 and implicate functions governing cell proliferation, differentiation and apoptosis control. To date studies show an increase of p63 expression in oral dysplasia and additionally high expression levels correlated with poor prognosis for patients with OSCC, whereas a possible link to radiation resistance of tumors has not been investigated yet. In the present study we tested the hypothesis for p63 being a marker of radioresistance and overall survival in OSCC. METHODS: p63 Expression was labeled by immunohistochemistry in pre-treatment biopsies collected from 33 patients with OSCC. Quantification of the labeling index (Li) based on the relation of p63 positive cells to overall tumor cell count. Histological examination of the resection specimen allowed categorization of the radiation response. Statistical analyses of the association between Li and radiation response were performed. Survival analysis utilized Kaplan-Meier estimates and additionally a Cox regression model was built for p63 (Li), T stage, N-stage and chemotherapy and presented as hazard ratios. RESULTS: All tumors had enhanced p63 expression. The median Li was 63.1% (range 36-87%). Tumors with a p63 positive cell count>63.1% showed increased resistance to radiation (p=0.027). Overall survival was higher (p=0.001) for patients with low Li (median value) and multivariate Cox regression analysis confirmed the significance of p63 as a prognostic marker of survival. CONCLUSIONS: The results of this analysis advocate p63 expression in pre-treatment tumor tissue to be a marker of radiation resistance in OSCC, with high expression levels being associated with poor radiation response and shorter survival. The promising results of this biomarker should now be confirmed by a study with larger patient counts.

Correcting false gene expression measurements from degraded RNA using RTQ-PCR.

This paper describes a method allowing correcting false gene expression measured on highly degraded RNA using real-time quantitative reverse transcription-polymerase chain reaction (RTQ-PCR). RNA was isolated from different models (in vitro cell lines, in vivo models of human and dog) and different tissue types. In vitro RNA degradation and modeling of in vivo degradation were applied on intact and degraded total RNA. Gene expression (eg, Bcl-2, GAPDH, PGK, PSME3, RAB2, BAX) was measured using RTQ-PCR. 18S rRNA proved to be the most constant house-keeping gene. Less than 10-fold degraded RNA can be quantified correctly when using 18S rRNA for normalization purposes. Higher-fold degraded RNA can be quantified correctly up to a precision that is comparable to RTQ-PCR measurements on intact RNA when simulating the RNA-species and tissue-specific degradation kinetic.

Overexpression of GLUT-1 is associated with resistance to radiotherapy and adverse prognosis in squamous cell carcinoma of the oral cavity.

This study tested the hypothesis that GLUT-1 is a marker of radioresistance in oral squamous cell carcinomas (OSCC). A GLUT-1 labeling index (LI) was determined by immunohistochemistry in 40 pretreatment OSCC biopsies. Radiation responses were categorized by histopathology of the resection specimens. Associations between the LI and radiation response, Kaplan-Meier survival estimates and Cox regression analysis for the variables GLUT-1, T-stage, N-stage and chemotherapy were examined. The median LI was 64.2% (range 14-100%). Tumors with >65% of GLUT-1+cells were more resistant to radiation (p=0.023). Overall survival was higher (p=0.044) for subjects with low LI (median value). The Cox regression analysis confirmed the prognostic significance of GLUT-1. Our results indicate that pre-treatment GLUT-1 expression in the tumor is a marker of radioresistance in OSCC, with high expression being associated with poor radiation response and shorter survival.