TRIP12 as a mediator of human papillomavirus/p16-related radiation enhancement effects.

Patients with human papillomavirus (HPV)-positive head and neck squamous cell carcinoma (HNSCC) have better responses to radiotherapy and higher overall survival rates than do patients with HPV-negative HNSCC, but the mechanisms underlying this phenomenon are unknown. p16 is used as a surrogate marker for HPV infection. Our goal was to examine the role of p16 in HPV-related favorable treatment outcomes and to investigate the mechanisms by which p16 may regulate radiosensitivity. HNSCC cells and xenografts (HPV/p16-positive and -negative) were used. p16-overexpressing and small hairpin RNA-knockdown cells were generated, and the effect of p16 on radiosensitivity was determined by clonogenic cell survival and tumor growth delay assays. DNA double-strand breaks (DSBs) were assessed by immunofluorescence analysis of 53BP1 foci; DSB levels were determined by neutral comet assay; western blotting was used to evaluate protein changes; changes in protein half-life were tested with a cycloheximide assay; gene expression was examined by real-time polymerase chain reaction; and data from The Cancer Genome Atlas HNSCC project were analyzed. p16 overexpression led to downregulation of TRIP12, which in turn led to increased RNF168 levels, repressed DNA damage repair (DDR), increased 53BP1 foci and enhanced radioresponsiveness. Inhibition of TRIP12 expression further led to radiosensitization, and overexpression of TRIP12 was associated with poor survival in patients with HPV-positive HNSCC. These findings reveal that p16 participates in radiosensitization through influencing DDR and support the rationale of blocking TRIP12 to improve radiotherapy outcomes.

Inflammation-related genetic variants predict toxicity following definitive radiotherapy for lung cancer.

Definitive radiotherapy improves locoregional control and survival in inoperable non-small cell lung cancer patients. However, radiation-induced toxicities (pneumonitis/esophagitis) are common dose-limiting inflammatory conditions. We therefore conducted a pathway-based analysis to identify inflammation-related single-nucleotide polymorphisms associated with radiation-induced pneumonitis or esophagitis. A total of 11,930 single-nucleotide polymorphisms were genotyped in 201 stage I-III non-small cell lung cancer patients treated with definitive radiotherapy. Validation was performed in an additional 220 non-small cell lung cancer cases. After validation, 19 single-nucleotide polymorphisms remained significant. A polygenic risk score was generated to summarize the effect from validated single-nucleotide polymorphisms. Significant improvements in discriminative ability were observed when the polygenic risk score was added into the clinical/epidemiological variable-based model. We then used 277 lymphoblastoid cell lines to assess radiation sensitivity and expression quantitative trait loci (eQTL) relationships of the identified single-nucleotide polymorphisms. Three genes (PRKCE, DDX58, and TNFSF7) were associated with radiation sensitivity. We concluded that inflammation-related genetic variants could contribute to the development of radiation-induced toxicities.

ALDH-1 expression levels predict response or resistance to preoperative chemoradiation in resectable esophageal cancer patients.

PURPOSE: Operable thoracic esophageal/gastroesophageal junction carcinoma (EC) is often treated with chemoradiation and surgery but tumor responses are unpredictable and heterogeneous. We hypothesized that aldehyde dehydrogenase-1 (ALDH-1) could be associated with response. METHODS: The labeling indices (LIs) of ALDH-1 by immunohistochemistry in untreated tumor specimens were established in EC patients who had chemoradiation and surgery. Univariate logistic regression and 3-fold cross validation were carried out for the training (67% of patients) and validation (33%) sets. Non-clinical experiments in EC cells were performed to generate complimentary data. RESULTS: Of 167 EC patients analyzed, 40 (24%) had a pathologic complete response (pathCR) and 27 (16%) had an extremely resistant (exCRTR) cancer. The median ALDH-1 LI was 0.2 (range, 0.01-0.85). There was a significant association between pathCR and low ALDH-1 LI (p ≤ 0.001; odds-ratio [OR] = 0.432). The 3-fold cross validation led to a concordance index (C-index) of 0.798 for the fitted model. There was a significant association between exCRTR and high ALDH-1 LI (p ≤ 0.001; OR = 3.782). The 3-fold cross validation led to the C-index of 0.960 for the fitted model. In several cell lines, higher ALDH-1 LIs correlated with resistant/aggressive phenotype. Cells with induced chemotherapy resistance upregulated ALDH-1 and resistance conferring genes (SOX9 and YAP1). Sorted ALDH-1+ cells were more resistant and had an aggressive phenotype in tumor spheres than ALDH-1- cells. CONCLUSIONS: Our clinical and non-clinical data demonstrate that ALDH-1 LIs are predictive of response to therapy and further research could lead to individualized therapeutic strategies and novel therapeutic targets for EC patients.

Improving imaging diagnosis of persistent nodal metastases after definitive therapy for oropharyngeal carcinoma: specific signs for CT and best performance of combined criteria.

BACKGROUND AND PURPOSE: Criteria for detection of persistent nodal metastases in treated oropharyngeal tumors are sensitive but nonspecific, leading to unnecessary nodal dissections. Developing specific imaging criteria for persistent nodal metastases could improve diagnosis while decreasing patient morbidity. MATERIALS AND METHODS: Patients with oropharyngeal squamous cell carcinoma with nodal metastases treated by definitive radiation therapy and subsequent nodal dissection were retrospectively evaluated. One hundred thirty-eight patients had pre- and posttherapy contrast-enhanced CTs evaluated by radiologists blinded to the status of pathologically proved hemineck persistent nodal metastases. Composite scoring criteria for CT, combined from individual parameters, were compared with radiologists' opinions, previous multiparameter criteria, and outcome data. RESULTS: New low-attenuation areas and a lack of size change (<20% cross sectional area) were both highly specific for persistent nodal metastases (99%; P = .0004). Extranodal disease on pretherapy imaging was moderately specific (86%; P = .001). The CSC correctly placed 29 patients in a low-risk category compared with 14 by previously reported criteria and radiologist reports. With good second-rater reliability, the CSC cutoff values stratified patients at highest risk of persistent nodal metastases, thereby improving specificity while maintaining sensitivity. CONCLUSIONS: Comparing pre- and posttherapy examinations improves specificity by discriminating focal findings and size change compared with a single time point. The CSC can categorize the risk of persistent nodal metastases more accurately than previous CT methods. This finding has the potential to improve resource use and reduce surgical morbidity.

Prenatal stress influences 8-OH-DPAT modulated startle responding and [3H]-8-OH-DPAT binding in rats.

The present study was to investigate some aspects of the 5-HT1A receptor system in adult-aged rats (50-60 days) that were either exposed to prenatal stress (PS) or not exposed to prenatal stress (CON). In the first series of experiments, rats were pretreated with vehicle, the 5-HT1A agonist 8-OH-DPAT or the 5-HT1A antagonist, WAY-100635 and exposed to 120 acoustic startle stimuli (95 dB) using a 30 s inter-trial interval. 8-OH-DPAT produced a dose-dependent increase in acoustic startle responding in CON and PS rats, with the PS rats exhibiting greater responding than CON rats. WAY-100635 depressed startle amplitudes only in the CON group. Finally, radioligand binding studies using [3H]-8-OH-DPAT indicated a significant decrease in receptor density in hippocampal homogenates from PS rats but no difference in [3H]-8-OH-DPAT binding from homogenates of the amygdala. Our results are consistent with earlier reports indicating that prenatal stress alters the serotonergic system. Specifically, our results indicate that gestational exposure to chronic mild stress enhances startle amplitudes following 8-OH-DPAT administration, prevents the depression in startle amplitudes following WAY-100635 administration and reduces [3H]-8-OH-DPAT binding in hippocampal preparations.

Mild prenatal stress in rats is associated with enhanced conditioned fear.

We tested the hypothesis that prenatal stress would enhance conditioned fear in adult rats. Pregnant Sprague-Dawley rats were stressed by exposure to a novel environment and subcutaneous injection of saline (0.1 ml 0.9% NaCl) at random times daily from Days 14 to 21 of pregnancy. When compared to adult control (CON) male rats from unmanipulated pregnancies, adult prenatally stressed (PS) male rats showed increased freezing behavior in response to acute footshock as well as increased freezing behavior the next day in the same context, without shock delivery. In another experiment, the gestational stressor was examined for elevations in corticosterone and ACTH. At gestational days (G)15, G17, G19 and G21, maternal and fetal plasma was collected. Analysis showed elevations in corticosterone and ACTH in the PS dams when compared to the CON dams. Additionally, increased corticosterone was found in the PS fetuses when compared to the CON fetuses. Finally, some CON and PS litters were examined for alterations in length of gestation, number of pups born, bodyweight on postnatal day (P)1 and anogenital distance on P1 and differences were not found. In conclusion, our data demonstrate that a mild stressor during gestation, sufficient to raise plasma corticosterone and ACTH, is associated with enhanced conditioned fear during adulthood.