Th2 Cells Are Associated with Tumor Recurrence Following Radiation.

The curative treatment of multiple solid tumors, including head and neck squamous cell carcinoma (HNSCC), utilizes radiation. The outcomes for HPV/p16-negative HNSCC are significantly worse than HPV/p16-positive tumors, with increased radiation resistance leading to worse locoregional recurrence (LRR) and ultimately death. This study analyzed the relationship between immune function and outcomes following radiation in HPV/p16-negative tumors to identify mechanisms of radiation resistance and prognostic immune biomarkers. A discovery cohort of 94 patients with HNSCC treated uniformly with surgery and adjuvant radiation and a validation cohort of 97 similarly treated patients were utilized. Tumor immune infiltrates were derived from RNAseq gene expression. The immune cell types significantly associated with outcomes in the discovery cohort were examined in the independent validation cohort. A positive association between high Th2 infiltration and LRR was identified in the discovery cohort and validated in the validation cohort. Tumor mutations in CREBBP/EP300 and CASP8 were significantly associated with Th2 infiltration. A pathway analysis of genes correlated with Th2 cells revealed the potential repression of the antitumor immune response and the activation of BRCA1-associated DNA damage repair in multiple cohorts. The Th2 infiltrates were enriched in the HPV/p16-negative HNSCC tumors and associated with LRR and mutations in CASP8, CREBBP/EP300, and pathways previously shown to impact the response to radiation.

FAK Drives Resistance to Therapy in HPV-Negative Head and Neck Cancer in a p53-Dependent Manner.

PURPOSE: Radiation and platinum-based chemotherapy form the backbone of therapy in human papillomavirus (HPV)-negative head and neck squamous cell carcinoma (HNSCC). We have correlated focal adhesion kinase (FAK/PTK2) expression with radioresistance and worse outcomes in these patients. However, the importance of FAK in driving radioresistance and its effects on chemoresistance in these patients remains unclear. EXPERIMENTAL DESIGN: We performed an in vivo shRNA screen using targetable libraries to identify novel therapeutic sensitizers for radiation and chemotherapy. RESULTS: We identified FAK as an excellent target for both radio- and chemosensitization. Because TP53 is mutated in over 80% of HPV-negative HNSCC, we hypothesized that mutant TP53 may facilitate FAK-mediated therapy resistance. FAK inhibitor increased sensitivity to radiation, increased DNA damage, and repressed homologous recombination and nonhomologous end joining repair in mutant, but not wild-type, TP53 HPV-negative HNSCC cell lines. The mutant TP53 cisplatin-resistant cell line had increased FAK phosphorylation compared with wild-type, and FAK inhibition partially reversed cisplatin resistance. To validate these findings, we utilized an HNSCC cohort to show that FAK copy number and gene expression were associated with worse disease-free survival in mutant TP53, but not wild-type TP53, HPV-negative HNSCC tumors. CONCLUSIONS: FAK may represent a targetable therapeutic sensitizer linked to a known genomic marker of resistance.

Cs-131 prostate brachytherapy boost and effect of hydrogel rectal spacer on long-term patient-reported rectal bleeding and bowel quality of life.

PURPOSE: This study evaluates long-term patient-reported bowel quality of life (QOL), rectal bleeding, and bleeding bother in patients with prostate cancer treated with external beam radiation therapy (EBRT) and Cesium-131 LDR brachytherapy (LDR-BT) boost with and without hydrogel rectal spacer. METHODS AND MATERIALS: This is a retrospective analysis of prostate cancer patients treated between 2007 and 2022 with 45 Gy EBRT followed by 85 Gy Cs-131 LDR-BT boost with or without hydrogel rectal spacer. Expanded Prostate Cancer Index Composite (EPIC) QOL questionnaires pre-treatment and at each follow-up were collected. Patient-reported rectal bleeding occurring more than "rarely" and bother from rectal bleeding occurring more than a "very small problem" were deemed clinically significant. Fisher's exact test was used to test the association of rectal spacer use and the incidence of clinically significant rectal bleeding and bleeding bother. Paired samples t-test was used to analyze mean bowel scores at each time point. RESULTS: Three hundred and forty-one patients were included in the analysis. The rectal spacer was used in 108 patients. Overall median follow-up was 48 months (IQR, 24-72), with a median follow-up of 24 months (IQR, 12-37.5) for the hydrogel group and 60 months (IQR, 36-84) for the non-hydrogel group. EPIC questionnaire response rates at median follow-up were 33% and 37% for the hydrogel and non-hydrogel groups, respectively. A clinically significant decrease in mean bowel domain scores was seen in the bowel bother domain at 6 and 12 months for patients who did not receive a rectal spacer. At the last follow-up of 60 months, the prevalence of clinically significant rectal bleeding and bleeding bother were 2.2% and 2.2%, respectively. The cumulative incidence of clinically significant long-term rectal bleeding was 2.8% and 18.9% in the hydrogel group and non-hydrogel group, respectively (Fisher's exact test, p < 0.0001). The cumulative incidence of clinically significant long-term bowel bother was 4.6% and 19.7% in the hydrogel group and non-hydrogel group, respectively (Fisher's exact test, p < 0.001). CONCLUSIONS: Use of hydrogel rectal spacer with EBRT and Cs-131 LDR-BT boost was significantly associated with a lower incidence of patient-reported rectal bleeding and bother from rectal bleeding, and better long-term bowel QOL. Cumulative incidence was 2.8% (hydrogel group) versus 18.9% (non-hydrogel group) and 4.6% (hydrogel group) versus 19.7% (non-hydrogel group) for clinically significant long-term rectal bleeding and long-term bleeding bother, respectively.

Effect of short-term corticosteroid usage on acute urinary toxicity following Cs-131 prostate brachytherapy.

PURPOSE: To evaluate short-term patient reported urinary quality of life scores in patients with prostate cancer treated at our institution with and without perioperative prednisone following Cesium-131 (131Cs) prostate LDR brachytherapy. METHODS AND MATERIALS: We started routinely using a perioperative 7-day course of prednisone at a dose of 5 mg per day, beginning 1 day prior to 131Cs prostate LDR brachytherapy from 2013 with goal of improving acute urinary symptomatology. One hundred consecutive patients treated with prednisone were selected, with comparison to 100 consecutive patients who were not treated with prednisone. We analyzed for differences in mean change with standard deviation (SD) in EPIC and AUA scores at 0.5-1 month and 3 months with or without prednisone by Mann-Whitney U Test. Binary logistic regression was performed to assess for impact of prednisone on postoperative urinary catheter use. RESULTS: Pretreatment EPIC and AUA scores were available in 197 patients. Less reduction in EPIC US score was noted at 0.5-1.0 month in the group who received prednisone with mean change of -22.9 (SD 15.4) when compared to the group who did not receive prednisone with mean change of -31.7 (SD 19.3), p < 0.01, with significance lost at 3 months. There was no significant difference in acute urinary retention requiring postoperative urinary catheter placement with perioperative prednisone (OR 1.13, p = 0.71). CONCLUSIONS: A short course of perioperative low-dose prednisone was associated with less severe worsening in urinary symptoms by the EPIC questionnaire at the 0.5-1.0-month timepoint suggesting some improvement in acute urinary quality of life, although differences did not remain statistically significant at 3 months.

Dosimetric parameters related to occurrence of distant metastases and regional nodal relapse after SBRT for early-stage non-small cell lung cancer.

PURPOSE: Previous studies have suggested that the dose immediately outside the PTV may impact the incidence of distant metastases after stereotactic body radiation therapy (SBRT) for patients with early-stage non-small cell lung cancer (NSCLC). In particular, Diamant et al. [1,2] reported a correlation between the mean EQD2 of a 30 mm shell around the PTV and both local control and the rate of distant metastases. In this study, we assess this parameter and others in a series of patients with radiographically presumed or biopsy-proven early-stage NSCLC treated at our institution with stereotactic body radiotherapy (SBRT) between 2017 and 2019. MATERIALS/METHODS: We reviewed the dosimetry, local control, regional nodal relapse, and rate of distant metastases for 304 patients with 325 lesions treated with SBRT at our institution. Dosimetric parameters investigated include the prescribed dose, minimum and mean doses to the PTV, conformity index, and the mean EQD2 to a 30 mm shell around the PTV. Time to each event was defined from date of last fraction of SBRT to date of event, with event-free patients censored at last radiographic follow-up. Univariate (UVA) Cox regression analysis was performed on the collected parameters to assess for correlation with regional nodal relapse and rate of distant metastases. RESULTS: There was no significant correlation between the mean EQD2 dose to a 30 mm shell around the PTV and the rate of distant metastases. On UVA Cox proportional hazards analysis, positive predictors of reduced incidence of distant metastases were PTV <22 cc (vs. ≥22 cc, p = 0.01) and GTV <10 cc (vs. ≥10 cc, p < 0.01), with GTV <10 cc also being a positive predictor of reduced incidence of regional nodal relapse (p < 0.01). In the subset of patients treated with 4-5 fractions, mean EQD2 dose to the 30 mm shell around the PTV ≥21 Gy was associated with increased incidence of distant metastases (HR 2.42, 95% CI 1.06-5.53, p = 0.04), differing from prior data from Diamant et al. CONCLUSIONS: We did not observe a correlation between the rate of distant metastases and dose outside the PTV, as reported by other groups; rather, we noted an opposite trend in patients treated with 4-5 fractions. Our data show additional correlations between distant metastases and tumor size.

p16 Represses DNA Damage Repair via a Novel Ubiquitin-Dependent Signaling Cascade.

Squamous cell carcinoma driven by human papillomavirus (HPV) is more sensitive to DNA-damaging therapies than its HPV-negative counterpart. Here, we show that p16, the clinically used surrogate for HPV positivity, renders cells more sensitive to radiotherapy via a ubiquitin-dependent signaling pathway, linking high levels of this protein to increased activity of the transcription factor SP1, increased HUWE1 transcription, and degradation of ubiquitin-specific protease 7 (USP7) and TRIP12. Activation of this pathway in HPV-positive disease led to decreased homologous recombination and improved response to radiotherapy, a phenomenon that can be recapitulated in HPV-negative disease using USP7 inhibitors in clinical development. This p16-driven axis induced sensitivity to PARP inhibition and potentially leads to "BRCAness" in head and neck squamous cell carcinoma (HNSCC) cells. Thus, these findings support a functional role for p16 in HPV-positive tumors in driving response to DNA damage, which can be exploited to improve outcomes in both patients with HPV-positive and HPV-negative HNSCC. SIGNIFICANCE: In HPV-positive tumors, a previously undiscovered pathway directly links p16 to DNA damage repair and sensitivity to radiotherapy via a clinically relevant and pharmacologically targetable ubiquitin-mediated degradation pathway.

Inhibition of histone acetyltransferase function radiosensitizes CREBBP/EP300 mutants via repression of homologous recombination, potentially targeting a gain of function.

Despite radiation forming the curative backbone of over 50% of malignancies, there are no genomically-driven radiosensitizers for clinical use. Herein we perform in vivo shRNA screening to identify targets generally associated with radiation response as well as those exhibiting a genomic dependency. This identifies the histone acetyltransferases CREBBP/EP300 as a target for radiosensitization in combination with radiation in cognate mutant tumors. Further in vitro and in vivo studies confirm this phenomenon to be due to repression of homologous recombination following DNA damage and reproducible using chemical inhibition of histone acetyltransferase (HAT), but not bromodomain function. Selected mutations in CREBBP lead to a hyperacetylated state that increases CBP and BRCA1 acetylation, representing a gain of function targeted by HAT inhibition. Additionally, mutations in CREBBP/EP300 are associated with recurrence following radiation in squamous cell carcinoma cohorts. These findings provide both a mechanism of resistance and the potential for genomically-driven treatment.

The influence of breast cancer subtype on survival after palliative radiation for osseous metastases.

BACKGROUND: Among patients with osseous metastases, breast cancer (BC) patients typically have the best prognosis. In the palliative setting, BC is often considered a single disease, but based on receptor status there are four distinct subtypes: luminal A (LA), luminal B (LB), triple negative (TN), and HER2-enriched (HER2). We hypothesize that survival and palliative outcomes following palliative RT for osseous metastases correlate with breast cancer subtype (BCS). METHODS: We identified 3,895 BC patients with known receptor status who received palliative RT for osseous metastases from 2004-2013 in the National Cancer Database. Kaplan-Meier method with log-rank testing and univariate/multivariate Cox-regression was used to identify survival factors. Incomplete radiation courses, 30-day mortality rate, and percentage remaining life spent receiving RT (PRLSRT) were calculated. RESULTS: Subtypes were 54% LA, 33% LB, 8% TN, and 5% HER2 with median survival of 34.1, 28.2, 5.3, and 15.7 months, respectively (p < 0.001). Overall 82% of patients received ≥10 fractions. Although BCS had limited effect on radiation regimens, TN received nearly twice as many single or hypofractionated (≤5 fractions) treatments, but the overall rate of these fraction schemes was low at 3.7 and 13.7%, respectively. Compared to LA and LB, TN and HER2 patients had worse palliative outcomes; higher rates of incomplete courses at 18.8% and 18.3% versus 12.7%-14.4%; higher 30-day mortality post-radiotherapy at 21.5% and 16.0% versus 6.3%-7.9%, and higher median PRLSRT of 7.7% and 3.7% versus 2.2%-2.4% for LA and LB. On multivariate analysis, BCS was associated with overall survival with TN (HR 3.7), HER2 (HR 1.75), and LB (HR 1.28) fairing worse than LA (p < 0.001). CONCLUSIONS: BCS correlated with survival and palliative outcome following radiation to osseous metastases. BCS should be considered by physicians when planning palliative RT to maximize quality-of-life, avoid unnecessary treatment, and ensure palliative benefits.

Enzyme and Cancer Cell Selectivity of Nanoparticles: Inhibition of 3D Metastatic Phenotype and Experimental Melanoma by Zinc Oxide.

Biomedical applications for metal and metal oxide nanoparticles are rapidly increasing. Here their functional impact on two well-characterized model enzymes, Luciferase (Luc) or ß-galactosidase (ß-Gal) was quantitatively compared. Nickel oxide nanoparticle (NiO-NP) activated ß-Gal (>400% control) and boron carbide nanoparticle (B4C-NP) inhibited Luc(<10% control), whereas zinc oxide (ZnO-NP) and cobalt oxide (Co3O4-NP) activated ß-Gal to a lesser extent and magnesium oxide (MgO) moderately inhibited both enzymes. Melanoma specific killing was in the order; ZnO > B4C ≥ Cu > MgO > Co3O4 > Fe2O3 > NiO, ZnO-NP inhibiting B16F10 and A375 cells as well as ERK enzyme (>90%) and several other cancer-associated kinases (AKT, CREB, p70S6K). ZnO-NP or nanobelt (NB) serve as photoluminescence (PL) cell labels and inhibit 3-D multi-cellular tumor spheroid (MCTS) growth and were tested in a mouse melanoma model. These results demonstrate nanoparticle and enzyme specific biochemical activity and suggest their utility as new tools to explore the important model metastatic foci 3-D environment and their chemotherapeutic potential.

Optimizing otoscopy competency in audiology students through supplementary otoscopy training.

BACKGROUND: Scope of practice in audiology encompasses proficiency in visual inspection of ear canal and tympanic membrane (TM) as well as otoscopy interpretation skills to determine normal versus abnormal conditions of outer and middle ear. Audiology students can develop skills in otoscopy through education and supervised training. Studies have shown that additional otoscopy training increased skills in medical students and general practitioners. However, educational and supervised practices targeting otoscopy competency during audiology graduate coursework are lacking. Also, no studies have attempted to determine otoscopy skills among audiology students. PURPOSE: To determine the effectiveness of the otoscopy training model on clinical competency and confidence level of audiology students in performing and interpreting otoscopy. RESEARCH DESIGN: A combination of experimental treatment design with random assignment of treatment and control groups and delayed treatment for control group. STUDY SAMPLE: Thirty-two first- and second-year audiology graduate students who were enrolled in a pediatric audiology class participated in this study. Students were randomly assigned to the control (n = 16, 14 females) or experimental (n = 16, 14 females) group. INTERVENTION: Participants in the experimental group received supplementary otoscopy training including didactic otoscopy lectures as well as clinical training using manikin ears. The control group received the same pretest and posttest and then completed a third assessment (posttest 2) after receiving the same training. DATA COLLECTION AND ANALYSIS: An evaluation of knowledge and skills regarding otoscopy between groups and time was conducted at three times: (a) pretraining, (b) upon completion of training for the experimental group, (c) upon completion of training by the control group. The evaluation consisted of a written exam, a clinical exam, and a self-perception rating of confidence. Both written exam scores and clinical exam scores (otoscopy manikin) were analyzed via two-way analyses of variance (ANOVAs), whereas chi-square (χ²) statistic was conducted to evaluate the effects of training on the confidence level of students of both groups. RESULTS: Experimental and control groups demonstrated significant increased overall competency in otoscopy following the otoscopy training model with didactic and laboratory components. Posttest confidence ratings showed increases in all groups, and there were no significant differences between groups. CONCLUSIONS: The need for supplementary otoscopy training was warranted by low knowledge and clinical competency in otoscopy skills of audiology students as measured by pretest mean scores. After completing the training, both experimental and control groups showed significant improvement in knowledge and competency. Results also suggest that perceived confidence ratings may be misleading in determining students' clinical otoscopy skills.