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Background To assess the feasibility of CBCT-based adaptive intensity modulated proton therapy (IMPT) using automated planning for treatment of head and neck (HN) cancers. Methods Twenty HN cancer patients treated with IMPT and had daily CBCTs were included. Initial IMPT plans were created using automated planning software for all patients. Synthetic CTs (sCT) were created by deforming planning CT (pCT) to CBCTs. rCTs were deformed to CBCT obtained on the same day to create deformed rCT (rCTdef) as gold standard. Doses recalculated on sCT and rCTdef with the were compared using gamma analysis. The dosimetric parameters were assessed based on rCTdef, sCT with deformably propagated contours or with physician-corrected contours. Weekly sCTs were then created for all patients. For non-adaptive group, dose was directly recalculated on weekly sCTs. For adaptive group, adaptive IMPT plans were created on weekly sCT using the automated planning software. The non-adapted and adapted dose-volume parameters on weekly sCTs were evaluated. The weekly doses were accumulated back to the pCT for both groups, and dosimetric parameters were compared in between. Results High dosimetric agreement was found between sCT and rCTdef with high gamma passing rate of 97.4%±1.6% using 3mm/3% criteria. With contour correction on sCT by physician, the uncertainty range of using sCT to estimate mean dose for organ at risk (OARs) can be reduced to (-2.37%, 2.19%), while for V95 of primary or secondary CTVs, the uncertainty can be controlled within (-1.09%, 0.29%). For non-adapted treatments, 15.7%, 34% and 55.7% of the fractions didn’t meet the V95 > 98% constraint for primary, secondary and tertiary CTVs respectively while all adaptive fractions met the constraint. In the accumulated dose, adaptive planning significantly reduced mean dose to constrictors (-1.42Gy ± 2.79Gy) and larynx (-2.58Gy ± 3.09Gy) compared to non-adaptive group, resulting in significantly reduction on the normal tissue complication probability (NTCP) of larynx edema by 7.52%± 13.59%. Conclusion sCT can be a powerful tool for accurate proton dose calculation. Physician contour correction is recommended to reduce the uncertainty for dose evaluation. Daily adaptation resulted in better CTV coverage, OAR sparing and lower NTCP for some OARs as compared with non-adaptive IMPT.

Background Our objective was to assess whether modifications to a customized targeted RNA sequencing (RNAseq) assay to include unique molecular identifiers (UMIs) that collapse read counts to their source mRNA counts would improve quantification of transcripts from formalin-fixed paraffin-embedded (FFPE) tumor tissue samples. The assay (SET4) includes signatures that measure hormone receptor and PI3-kinase related transcriptional activity (SETER/PR and PI3Kges), and measures expression of selected activating point mutations and key breast cancer genes. Methods Modifications included steps to introduce eight nucleotides-long UMIs during reverse transcription (RT) in bulk solution, followed by polymerase chain reaction (PCR) of labeled cDNA in droplets, with optimization of the polymerase enzyme and reaction conditions. We used Lin’s concordance correlation coefficient (CCC) to measure concordance, including precision (Rho) and accuracy (Bias), and nonparametric tests (Wilcoxon, Levene’s) to compare the modified (NEW) SET4 assay to the original (OLD) SET4 assay and to whole transcriptome RNAseq using RNA from matched fresh frozen (FF) and FFPE samples from 12 primary breast cancers. Results The modified (NEW) SET4 assay measured single transcripts (pER/PR (p=0.002) more reproducibly in technical replicates from FFPE samples. The modified SET4 assay was more precise for measuring single transcripts (Rho 0.966 vs 0.888, pER/PR (Rho 0.985 vs 0.968) or PI3Kges (Rho 0.985 vs 0.946) in FFPE, compared to FF samples. It was also more precise than wtRNAseq of FFPE for measuring transcripts (Rho 0.986 vs 0.934, pER/PR (Rho 0.993 vs 0.915, p=0.004), but not PI3Kges (Rho 0.988 vs 0.945, p=0.051). Accuracy (Bias) was comparable between protocols. Two samples carried a PIK3CA mutation, and measurements of transcribed mutant allele fraction was similar in FF and FFPE samples and appeared more precise with the modified SET4 assay. Amplification efficiency (reads per UMI) was consistent in FF and FFPE samples, and close to the theoretically expected value, when the library size exceeded 400,000 aligned reads. Conclusions Modifications to the targeted RNAseq protocol for SET4 assay significantly increased the precision of UMI-based and reads-based measurements of individual transcripts, multi-gene signatures, and mutant transcript fraction, particularly with FFPE samples.

Glioma intratumoral heterogeneity enables adaptation to challenging microenvironments and contributes to therapeutic resistance. We integrated 914 single-cell DNA methylomes, 55,284 single-cell transcriptomes and bulk multi-omic profiles across 11 adult IDH mutant or IDH wild-type gliomas to delineate sources of intratumoral heterogeneity. We showed that local DNA methylation disorder is associated with cell–cell DNA methylation differences, is elevated in more aggressive tumors, links with transcriptional disruption and is altered during the environmental stress response. Glioma cells under in vitro hypoxic and irradiation stress increased local DNA methylation disorder and shifted cell states. We identified a positive association between genetic and epigenetic instability that was supported in bulk longitudinally collected DNA methylation data. Increased DNA methylation disorder associated with accelerated disease progression and recurrently selected DNA methylation changes were enriched for environmental stress response pathways. Our work identified an epigenetically facilitated adaptive stress response process and highlights the importance of epigenetic heterogeneity in shaping therapeutic outcomes.

Glioma intratumoral heterogeneity enables adaptation to challenging microenvironments and contributes to universal therapeutic resistance. Here, we integrated 914 single-cell DNA methylomes, 55,284 single-cell transcriptomes, and bulk multi-omic profiles across 11 adult IDH-mutant or IDH-wild-type gliomas to delineate sources of intratumoral heterogeneity. We found that local DNA methylation instability, or epimutation burden, was elevated in more aggressive tumors, reflected intratumoral variability, linked with transcriptional disruption, and associated with environmental stress response. We show that the activation of cell-state specific transcription factors is impacted by epimutations and that loosened epigenetic control may facilitate cellular plasticity. Our analyses support that somatic copy number alterations (SCNAs) promote epigenetic instability and that SCNAs largely precede epigenetic and transcriptomic diversification during glioma evolution. We confirmed the link between genetic and epigenetic instability by analyzing larger cohorts of bulk longitudinally collected and spatially separated DNA methylation data. Increased DNA methylation instability was associated with accelerated disease progression, and recurrently selected DNA methylation changes were enriched for environmental stress response pathways. Our work provides an integrative framework to better understand glioma evolution and highlights the importance of epigenetic heterogeneity in shaping therapeutic response.

Critical globalization scholars argue that democracy is in crisis, negatively affecting political systems, social relations, and education. Most think education and democracy ought to play a progressive role in advancing the common good for humanity. We demonstrate that education has become a reflection of neoliberalism, replacing mutual concern and compassion with fear of uncertainty, which results in preserving global capitalism's negative processes. We propose that to achieve real democracy, education, and radical emancipatory processes that can overturn neoliberalism, we ought to make individuals aware of the role of freedom under capitalism, in warping the foundation of modern societies. To do this, education ought to be conceived as the pillar of cognitive compassion and not as a component of neoliberalism. Education must be directed at overcoming the capitalist ideology and promoting cognitive compassion. This is the path toward a good society and ultimately real democracy.

Genetic and epigenetic alterations contribute to the observed intratumoral heterogeneity in adult glioma. Current glioma classification, based on genotype (e.g., IDH1 mutations) and DNA methylation profiles (e.g., glioma CpG Island Methylator Phenotype), can provide clinically relevant tumor subgroups. However, traditional bulk sampling fails to adequately capture the full complement of epigenomic heterogeneity, and may mask deadly features present in less abundant glioma cells. To more precisely characterize the glioma epigenome, we separately profiled single-cell DNA methylation (Reduced Representation Bisulfite Sequencing, RRBS), single-cell RNA expression (10X genomics), and bulk whole genome sequencing in nine gliomas. The genomic regions profiled by scRRBS were primarily gene promoters, but adequate coverage was also reached for glioma-specific enhancer elements and binding sites of chromatin remodelers. Unsupervised clustering of single-cell DNA methylation data revealed intratumoral variability in epigenetic classification and these cell types were distinguished by regulatory element DNA methylation. We further integrated single-cell epigenetic, single-cell transcriptomic, and genomic features to better understand gene regulation and reconstruct each tumor’s lineage history. Together, our study aims to generate a glioma cellular hierarchy shaped by the epigenetic programs that drive tumor growth.

Extensive intra- and intertumoral heterogeneity in glioma contributes to therapeutic resistance and poor patient outcomes. Alterations to DNA methylation (DNAme) modulate epigenetic homeostasis, allowing tumor cells to sample alternative cell states to promote tumorigenesis. However, the epigenetic mechanisms that promote cellular plasticity and regulate cell states are still poorly understood. To characterize the epigenetic mechanisms underlying glioma heterogeneity we profiled 914 single-cell methylomes, 55,284 single-cell transcriptomes, and bulk whole genomes across 11 patient samples spanning initial and recurrent time points and 3 molecular subtypes delineated by IDH mutation status. Local DNAme disorder, defined as epimutation burden, was increased in tumor cells relative to nontumor cells, higher in IDH wild-type than in IDH mutant glioma and was positively associated with copy number alteration (CNA). Epimutation was positively associated with transcriptional variability and enriched at genes involved in cellular differentiation. Epimutation was also increased in the binding sites of transcription factors (TFs) associated with response to extracellular stimuli, suggesting that stochastic DNAme alterations enable cellular plasticity and diverse responses to microenvironmental stressors. Integrative clustering of DNAme and scRNAseq profiles defined stem-like and differentiated-like cell states which exhibited differences in TF activity. Stem-like cells were enriched for differentially methylated binding sites of TFs associated with hypoxia response. scDNAme and scRNAseq-derived copy number profiles were compared with bulk copy number profiles and inferred tumor phylogenies to assess how the timing of CNAs impact epigenetic instability, with results suggesting that early CNA events propagate both genetic and epigenetic heterogeneity. Bulk longitudinal data was used to validate the relationship of epigenetic instability with CNA burden as well as differentially methylated binding sites of cell stress response TFs. Our work suggests that local DNAme disorder promotes cellular plasticity and enables adaptive response to cellular stress such as hypoxia.

a b s t r a c t Free-living hummingbirds can learn the refill schedules of individual experimental flowers but little is known about what information they use to do this. Colour cues, in particular, may be important to hummingbirds when learning about rewarded flower properties. We investigated, therefore, whether colour cues facilitated the learning of flower refill schedules in wild, free-living rufous hummingbirds (Selasphorus rufus). In the Cued condition, we presented birds with an array of six flowers, three of one colour, each of which were refilled 10 min after being emptied by the bird and three of a different colour, which were refilled 20 min after being emptied. In the Uncued condition we presented birds with six flowers of the same colour, three of which were refilled after 10 min and three of which were refilled after 20 min as for the birds in the Cued condition. In the second part of the experiment, we moved the array 2 m and changed the shape of the array. Across both phases, birds in the Cued condition learned to discriminate between 10 and 20-min flowers more quickly than did the birds in the Uncued condition. The Cued birds were also better at discriminating between the two distinct refill intervals. Colour cues can, therefore, facilitate learning the refill schedules of experimental flowers in these birds. This article is part of a Special Issue entitled: Cognition in the wild.

Developmental, behavioural, and neurological similarities in the processing of different magnitudes (time, number, space) support the existence of a common magnitude processing system (e.g., a theory of magnitude, ATOM). It is, however, unclear whether the recruitment of wider cognitive resources (short-term memory, STM; and executive function) during magnitude processing is similar across magnitude domains or is domain specific. The current study used an individual differences approach to examine the relationship between STM, executive function, and magnitude processing. In two experiments, participants completed number, length, and duration bisection tasks to assess magnitude processing and tasks that have been shown to assess STM span and executive component processes. The results suggest that the recruitment of STM and executive resources differed for the different magnitude domains. Duration perception was associated with access, inhibition, and STM span. Length processing was associated with updating, and number processing was associated with access to semantic memory. For duration and length, greater difficulty in the magnitude judgement task resulted in more relationships to STM and executive function. It is suggested that duration perception may be more demanding of STM and executive resources because it is represented sequentially, unlike length and number which can be represented nonsequentially.

Patients with head and neck cancer are at high risk for second primary malignancies. Human papillomavirus (HPV)-driven tumors are generally high-grade oropharyngeal cancers. We analyzed the incidence of second primary malignancy of the head and neck in patients with primary squamous cell carcinoma (SCC) of the head and neck and temporal trends in the HPV era.The Surveillance, Epidemiology, and End Results (SEER) database was queried for patients with SCC of the head and neck (range, 1973-2008). Cumulative incidence rates of second primary malignancy of the head and neck were compared based on competing risk analysis.A total of 104,639 cases were included in this study, of which 4616 patients had second primary malignancy of the head and neck. Oropharyngeal cancer incidence increased over time. Estimated incidence rate/10,000 person-years (105.5, 80.6, and 50.2 for 1973-1989, 1990-1999, and 2000-2008, respectively) and cumulative incidence rates (10-year rates of 6.68%, 5.72%, and 4.59% for 1973-1989, 1990-1999, and 2000-2008, respectively) of second primary malignancies of the head and neck for patients with oropharyngeal cancer decreased over time (p .001). The second primary malignancy of the head and neck incidence rate was significantly lower in patients with high-grade oropharyngeal cancer from 2000 to 2008 (30.3 vs 65.5 and 54.6 from 1973-1989 and 1990-1999, respectively; p .001).The incidence of second primary malignancy of the head and neck in patients with head and neck cancer has decreased over time. This is driven by lower rates in patients with high-grade oropharyngeal cancer, is temporally related with increases in HPV-associated oropharyngeal cancer, and suggests that incidence rates of second primary malignancy of the head and neck may be lower for HPV-associated cancer. © 2015 Wiley Periodicals, Inc. Head Neck 38: E873-E883, 2016.

In insurance, the analyst is often faced with a large number of inter-related variables for which correlations need to be estimated. Clearly, all correlations lie in the interval [−1, 1], but the numbers cannot be assigned independently. Here, the choices left to the analyst are considered from both a geometric and a probabilistic viewpoint. In practice, the underwriter or risk manager may fix some of the correlations and this paper considers, from both these viewpoints, what effect this has on the analyst's choice of correlations between the remaining variables.

Recent psychosocial advances in colorectal surgery have greatly reduced hospital stay and anxiety and improved postsurgical recovery. The advances have put together education, preoperative care, postoperative care, pain control, mobility, and postsurgical attitude education. This chapter addresses the psychosocial aspects of colorectal surgery care to optimize the experience and healing of patients.

Incident learning systems are important tools to improve patient safety in radiation oncology, but physician participation in these systems is poor. To understand reporting practices and attitudes, a survey was sent to staff members of four large academic radiation oncology centers, all of which have in-house reporting systems.Institutional review board approval was obtained to send a survey to employees including physicians, dosimetrists, nurses, physicists, and radiation therapists. The survey evaluated barriers to reporting, perceptions of errors, and reporting practices. The responses of physicians were compared with those of other professional groups.There were 274 respondents to the survey, with a response rate of 81.3%. Physicians and other staff agreed that errors and near-misses were happening in their clinics (93.8% v 88.7%, respectively) and that they have a responsibility to report (97% overall). Physicians were significantly less likely to report minor near-misses (P = .001) and minor errors (P = .024) than other groups. Physicians were significantly more concerned about getting colleagues in trouble (P = .015), liability (P = .009), effect on departmental reputation (P = .006), and embarrassment (P.001) than their colleagues. Regression analysis identified embarrassment among physicians as a critical barrier. If not embarrassed, participants were 2.5 and 4.5 times more likely to report minor errors and major near-miss events, respectively.All members of the radiation oncology team observe errors and near-misses. Physicians, however, are significantly less likely to report events than other colleagues. There are important, specific barriers to physician reporting that need to be addressed to encourage reporting and create a fair culture around reporting.

This chapter explores the psychosocial dynamics of a preoperative patient. The theoretical foundation of the article is based on Jean Watson’s theory of human caring. Each of Watson’s Caritas Processes are presented in the context of preoperative care. Exemplars of guided imagery, creative art interventions, and personal narratives are presented. Psychosocial preparation can be profoundly empowering to patients and their families by inviting them to take the psychosocial preparation process into their own hands and become more involved in the process.

The pluripotent murine embryonal carcinoma cell line, P19, differentiates along at least three main pathways under the inductive influence of retinoic acid (RA). The events most critical to the establishment of a particular differentiation pathway must occur early since P19 cells are committed to differentiation pathways after 30 min of exposure to RA (M. W. McBurney, personal communication and our unpublished results). We have, therefore, looked for genes that are induced (or repressed) within 30 min of RA addition and find that Egr-1 is one of these genes. Egr-1 is a transcription factor of the zinc-finger class and is known to transactivate genes after binding to specific oligonucleotide sequences. We describe here the extremely rapid and transient increase of Egr-1 transcript and protein levels in P19 cells after RA addition. Stable induction of Egr-1 transcripts occurred in the presence of protein synthesis inhibitors. Simultaneous addition of RA and cycloheximide did not result in an additive effect. The mechanism of induction with either drug appears to involve relief of a block to transcriptional elongation. The response was more rapid at high RA concentrations and this suggests that the Egr-1 transcription factor could play a role in initiation of differentiation pathways of P19 EC cells.

Microarray formatted assays have become well established for the global analysis of nucleic acids, but are only beginning to be adopted for proteomic analysis. This chapter reviews the current status of protein microarray assays and highlights efforts using cells and tissues on microarrays. These ‘other’ microarrays have the potential to yield highly parallel miniaturized assays that will accelerate drug discovery efforts and facilitate understanding of biological complexity.

Cancer cachexia is a common problem among advanced cancer patients. A mixture of beta-hydroxyl beta-methyl butyrate, glutamine, and arginine (HMB/Arg/Gln) previously showed activity for increasing lean body mass (LBM) among patients with cancer cachexia. Therefore a phase III trial was implemented to confirm this activity.Four hundred seventy-two advanced cancer patients with between 2% and 10% weight loss were randomized to a mixture of beta-hydroxyl beta-methyl butyrate, glutamine, and arginine or an isonitrogenous, isocaloric control mixture taken twice a day for 8 weeks. Lean body mass was estimated by bioimpedance and skin-fold measurements. Body plethysmography was used when available. Weight, the Schwartz Fatigue Scale, and the Spitzer Quality of Life Scale were also measured.Only 37% of the patients completed protocol treatment. The majority of the patient loss was because of patient preference (45% of enrolled patients). However, loss of power was not an issue because of the planned large target sample size. Based on an intention to treat analysis, there was no statistically significant difference in the 8-week lean body mass between the two arms. The secondary endpoints were also not significantly different between the arms. Based on the results of the area under the curve (AUC) analysis, patients receiving HMB/Arg/Gln had a strong trend higher LBM throughout the study as measured by both bioimpedance (p = 0.08) and skin-fold measurements (p = 0.08). Among the subset of patients receiving concurrent chemotherapy, there were again no significant differences in the endpoints. The secondary endpoints were also not significantly different between the arms.This trial was unable to adequately test the ability of beta-hydroxy beta-methylbutyrate, glutamine, and arginine to reverse or prevent lean body mass wasting among cancer patients. Possible contributing factors beyond the efficacy of the intervention were the inability of patients to complete an 8-week course of treatment and return in a timely fashion for follow-up assessment, and because the patients may have only had weight loss possible not related to cachexia, but other causes of weight loss, such as decreased appetite. However, there was a strong trend towards an increased body mass among patients taking the Juven compound using the secondary endpoint of AUC.

Introduction To evaluate the efficacy and safety of myeloid growth factors in patients with locally advanced non-small cell lung cancer treated with combined modality therapy (CMT). Methods Patients with stage IIIA/B non-small cell lung cancer, performance status 0 to 1, and forced expiratory volume in 1 second ≥1.5, received cisplatin 75 mg/m 2 on day 1 + etoposide 80 mg/m 2 on days 1 to 3 every 3 weeks for 2 cycles concurrent with thoracic radiotherapy to 61 Gy. filgrastim 5 mcg/kg/d was administered for 10 days beginning on day 4 of each chemotherapy cycle. Patients without progression received docetaxel 75 mg/m 2 every 21 days for 3 cycles with peg-filgrastim 6 mg on day 2. The primary end point was a 50% reduction in the incidence of grade ¾ neutropenia compared with historical controls. Results A total of 26 eligible patients were enrolled. Median age was 67, 76% were men, and 58% had stage IIIA. Gr3/4 neutropenia during CMT was 19.2% and 3.8%, respectively. There were no episodes of febrile neutropenia. Gr4 thrombocytopenia was 15.4% with 2 patients requiring transfusions. Gr3 esophagitis was noted in 7.7% and Gr ¾ pneumonitis in 21.6% of patients. No patients died of treatment-related toxicities. Dose reductions/delays occurred in 3.8% of patients during CMT. Median progression-free survival and median survival were 10.7 and 27.6 months, respectively. The 1- and 2-year survival rates were 61.5% and 46.2%, respectively. Conclusions Our data suggest that the addition of filgrastim to CMT is safe and effective. The rate of grade ¾ toxicities, including febrile neutropenia, compares favorably to previous trials using a similar regimen. Dose intensity is maintained. This strategy merits further evaluation.