Liposomes as Carriers of Membrane-Associated Proteins and Peptides for Mass Spectrometric Analysis.

Membrane proteins are key players of the cell. Their structure and the interactions they form with their lipid environment are required to understand their function. Here we explore liposomes as membrane mimetics for mass spectrometric analysis of peripheral membrane proteins and peptides. Liposomes are advantageous over other membrane mimetics in that they are easy to prepare, can be varied in size and composition, and are suitable for functional assays. We demonstrate that they dissociate into lipid clusters in the gas phase of a mass spectrometer while intact protein and protein-lipid complexes are retained. We exemplify this approach by employing different liposomes including proteoliposomes of two model peptides/proteins differing in size. Our results pave the way for the general application of liposomes for mass spectrometric analysis of membrane-associated proteins.

The state of survivorship care in radiation oncology: Results from a nationally distributed survey.

BACKGROUND: Survivorship care has become an increasingly critical component of oncologic care as well as a quality practice and reimbursement metric. To the authors' knowledge, the current climate of survivorship medicine in radiation oncology has not been investigated fully. METHODS: An institutional review board-approved, Internet-based survey examining practices and preparedness in survivorship care was distributed to radiation oncology practices participating in the American College of Radiology Radiation Oncology Practice Accreditation program between November 2016 and January 2017. A total of 78 surveys were completed. Among these, 2 were nonphysicians, resulting in 76 evaluable responses. RESULTS: Radiation oncologists (ROs) frequently reported that they are the primary provider in the evaluation of late toxicities and the recurrence of primary cancer. Although approximately 68% of ROs frequently discuss plans for future care with survivors, few provide a written survivorship care plan to their patients (18%) or the patients' primary care providers (24%). Patient prognosis, disease site, and reimbursement factors often influence the provision of survivorship care. Although ROs report that several platforms offer training in survivorship medicine, the quality of these resources is variable and extensive instruction is rare. Fewer than one-half of ROs believe they are expertly trained in survivorship care. CONCLUSIONS: ROs play an active role within the multidisciplinary team in the cancer-related follow-up care of survivors. Investigation of barriers to the provision of survivorship care and optimization of service delivery should be pursued further. The development of high-quality, easily accessible educational programming is needed so that ROs can participate more effectively in the care of cancer survivors. Cancer 2018;124:2653-60. © 2018 American Cancer Society.

A novel sample preparation strategy for shotgun lipidomics of phospholipids employing multilamellar vesicles.

The identification of lipids in biological samples is gaining importance. The advent of mass spectrometry-based lipidomics accelerated the field allowing nowadays for identification and quantification of complete lipidomes. However, due to solubility difficulties and varying properties of different lipid classes, sample preparation for lipidomics is still an issue. Of the many lipid classes, phospholipids are the major components of biological membranes. In solution, they spontaneously form lipid vesicles of various structures such as liposomes. They are therefore often used as membrane mimics when studying biological membranes and membrane proteins. Here, we present a novel sample preparation strategy for shotgun lipidomics employing liposomes prepared from lipid standards or lipid mixtures allowing the analysis of phospholipids directly from lipid bilayers. We validated our strategy for lipid identification by tandem mass spectrometry in positive or negative ion mode using different phospholipid species from various classes. We further tested our strategy for relative quantification by mixing different ratios of phospholipid species as well as determining the distribution of lipid species in a natural lipid extract. Graphical abstract ᅟ.

Distinct patterns of cytolytic T-cell activation by different tumour cells revealed by Ca2+ signalling and granule mobilization.

Cancer-germline genes in both humans and mice have been shown to encode antigens susceptible to targeting by cytotoxic CD8 T effector cells (CTL). We analysed the ability of CTL to kill different tumour cell lines expressing the same cancer-germline gene P1A (Trap1a). We previously demonstrated that CTL expressing a T-cell receptor specific for the P1A35-43 peptide associated with H-2Ld , although able to induce regression of P1A-expressing P815 mastocytoma cells, were much less effective against P1A-expressing melanoma cells. Here, we analysed parameters of the in vitro interaction between P1A-specific CTL and mastocytoma or melanoma cells expressing similar levels of the P1A gene and of surface H-2Ld . The mastocytoma cells were more sensitive to cytolysis than the melanoma cells in vitro. Analysis by video-microscopy of early events required for target cell killing showed that similar patterns of increase in cytoplasmic Ca2+ concentration ([Ca2+ ]i) were induced by both types of P1A-expressing tumour cells. However, the use of CTL expressing a fluorescent granzyme B (GZMB-Tom) showed a delay in the migration of cytotoxic granules to the tumour interaction site, as well as a partially deficient GZMB-Tom exocytosis in response to the melanoma cells. Among surface molecules possibly affecting tumour-CTL interactions, the mastocytoma cells were found to express intercellular adhesion molecule-1, the ligand for LFA-1, which was not detected on the melanoma cells.

Survivorship and the chronic cancer patient: Patterns in treatment-related effects, follow-up care, and use of survivorship care plans.

BACKGROUND: The survivorship needs of patients living with chronic cancer (CC) and their use of survivorship care plans (SCPs) have been overlooked and underappreciated. METHODS: A convenience sample of 39,088 SCPs completed for cancer survivors with an Internet-based SCP tool was examined; it included 5847 CC survivors (15%; CC was defined as chronic leukemia and/or recurrent/metastatic cancer of another nature). Patient-reported treatment effects and follow-up care patterns were compared between CC survivors and survivors treated with curative intent (CI). Responses from a follow-up survey regarding SCP satisfaction and use were reviewed. RESULTS: CC survivors had greater odds of experiencing multiple treatment-related effects than survivors treated with CI; these effects included fatigue, cognitive changes, dyspnea, peripheral neuropathy, lymphedema, and erectile dysfunction. Nearly half of CC survivors were managed by an oncologist alone, and they were less likely than CI patients to be comanaged by a primary care provider and an oncologist. Fewer SCPs were generated by health care providers (HCPs) for CC survivors versus CI survivors. A smaller proportion of CC users versus CI users rated their experience and satisfaction with the SCP tool as very good or excellent, and CC users were less likely to share the HCP summary with their health care team. CONCLUSIONS: A substantial number of CC survivors, often considered incurable but treatable, seek survivorship support. Tools to facilitate participation, communication, and coordination of care are valuable for these patients, and future iterations of SCPs should be designed to address the particular circumstances of living with CC. Cancer 2017;123:4268-4276. © 2017 American Cancer Society.

Survivorship after lower gastrointestinal cancer: Patient-reported outcomes and planning for care.

BACKGROUND: There is significant need for quality follow-up care to optimize long-term outcomes for the growing population of lower gastrointestinal (GI) cancer survivors. Patient-reported outcomes (PROs) provide valuable information regarding late and long-term effects (LLTEs). METHODS: A convenience sample from 1129 colon, rectal, and anal cancer survivors (n = 792; 218, and 119, respectively) who participated in an Internet-based survivorship care plan (SCP) tool between May 2010 and October 2014 was used to examine patient-reported demographics, treatment, and toxicity data. Responses from a follow-up survey were reviewed. RESULTS: The median age of diagnosis was 51 years, and 81% of survivors were Caucasian. The most commonly reported LLTEs for all survivors were neuropathy, fatigue, cognitive changes, changes in GI function, urogenital and sexual dysfunction, and dermatologic effects. The prevalence of these effects varied with time since diagnosis, treatment modality, and treatment center. Individuals who had survived anal cancer reported a high prevalence of sexual dysfunction and radiation-induced dermatologic effects. Over 87% of users reported satisfaction levels of good to excellent using the SCP tool, and 69% reported that they intend to share the SCP with their health care team. CONCLUSIONS: For lower GI cancer survivors, it is feasible to obtain PROs from an Internet-based survivorship tool. Survivors report a wide spectrum of LLTEs, and these can be used to inform counseling at the time of diagnosis and to help anticipate and respond to disease-related and treatment-related sequelae during follow-up. The authors are among the first to report on PROs in anal cancer survivors. Further investigation on the impact of SCPs on health care communication and use is needed. Cancer 2017;123:1860-1868. © 2017 American Cancer Society.

Visualization of granzyme B-expressing CD8 T cells during primary and secondary immune responses to Listeria monocytogenes.

CD8 T cells contribute to long-term protection against Listeria monocytogenes infection by differentiating into memory T cells. These rapidly respond to antigen or inflammation upon secondary infection. In this study we used CD8 T cells from OT1 mice and CD4 T cells from OT2 mice expressing a fluorescent chimeric granzyme (GZMB-Tom) protein to monitor the primary response to infection with ovalbumin-expressing L. monocytogenes (Lm-OVA). We show that, unlike poorly responding CD4 T cells, CD8 T cells readily proliferated and expressed high levels of GZMB-Tom as early as 2 days after infection. FACS analysis showed GZMB-Tom expression in undivided CD8 T cells, with its level increasing over one to four divisions. OT1 T cells were visualized in the T-cell zone by confocal microscopy. This showed GZMB-Tom-containing granules oriented towards MHCII-positive cells. Twenty hours later, most OT1 T cells had divided but their level of GZMB-Tom expression was reduced. Recently divided OT1 cells failed to express GZMB-Tom. Fourteen hours after secondary infection, GZMB-Tom was re-expressed in memory OT1 T cells responding either to Lm-OVA or L. monocytogenes. Differences in the activation phenotype and in the splenic distribution of OT1 T cells were observed, depending on the challenge. Notably, OTI T cells with polarized granules were only observed after challenge with cognate antigen. This work showed that the GZMB-Tom knock-in mice in which GZMB-Tom faithfully reproduced GZMB expression, provide useful tools to dissect mechanisms leading to the development of anti-bacterial effector and memory CD8 T cells and reactivation of the memory response to cognate antigen or inflammatory signals.

Ca2+-dependent lipid preferences shape synaptotagmin-1 C2A and C2B dynamics: Insights from experiments and simulations.

Signal transmission between neurons requires exocytosis of neurotransmitters from the lumen of synaptic vesicles into the synaptic cleft. Following an influx of Ca2+, this process is facilitated by the Ca2+ sensor synaptotagmin-1. The underlying mechanisms involve electrostatic and hydrophobic interactions tuning the lipid preferences of the two C2 domains of synaptotagmin-1; however, the details are still controversially discussed. We, therefore, follow a multidisciplinary approach and characterize lipid and membrane binding of the isolated C2A and C2B domains. We first target interactions with individual lipid species, and then study interactions with model membranes of liposomes. Finally, we perform molecular dynamics simulations to unravel differences in membrane binding. We found that both C2 domains, as a response to Ca2+, insert into the lipid membrane; however, C2A adopts a more perpendicular orientation while C2B remains parallel. These findings allow us to propose a mechanism for synaptotagmin-1 during membrane fusion.

Effect of Radiation Schedule on Transportation-Related Carbon Emissions: A Case Study in Rectal Cancer.

Purpose: The health care sector is a major contributor of worldwide greenhouse gas (GHG) emissions. Indirect emissions, including those associated with transportation, make up 82% of the US health care sector's environmental footprint. Radiation therapy (RT) treatment regimens present an opportunity for environmental health care-based stewardship owing to the high incidence of cancer diagnosis, significant utilization of RT, and myriad treatment days required for curative regimens. Because the use of short-course RT (SCRT) in the treatment of rectal cancer has demonstrated noninferior clinical outcomes compared with conventional, long-course RT (LCRT), we investigate the environmental and health equity-related outcomes. Methods and Materials: Patients treated with curative, preoperative RT for newly diagnosed rectal cancer at our institution between 2004 and 2022 and living in-state were included. Travel distance was estimated using patients' reported home address. Associated GHG emissions were calculated and reported in carbon dioxide equivalents (CO2e). Results: Of 334 patients included, the total distance traveled for the treatment course was significantly greater in patients treated with LCRT versus SCRT (median, 1417 vs 319 miles; P < .001). Total CO2e emissions for those undergoing LCRT (n = 261) and SCRT (n = 73) were 665.3 kg CO2e and 149.9 kg CO2e, respectively, per treatment course (P < .001), with a net difference of 515.4 kg CO2e. Relatively, this suggests that LCRT is associated with 4.5 times greater GHG emissions from patient transportation. Conclusions: Using treatment of rectal cancer as proof-of-principle, we advocate for the inclusion of environmental considerations in the creation of climate-resilient oncologic RT practices, especially in the context of equivocal clinical outcomes between RT fractionation schedules.

Radiation Therapy for Endometrial Cancer: An American Society for Radiation Oncology Clinical Practice Guideline.

PURPOSE: With the results of several recently published clinical trials, this guideline informs on the use of adjuvant radiation therapy (RT) and systemic therapy in the treatment of endometrial cancer. Updated evidence-based recommendations provide indications for adjuvant RT and the associated techniques, the utilization and sequencing of adjuvant systemic therapies, and the effect of surgical staging techniques and molecular tumor profiling. METHODS: The American Society for Radiation Oncology convened a multidisciplinary task force to address 6 key questions that focused on the adjuvant management of patients with endometrial cancer. The key questions emphasized the (1) indications for adjuvant RT, (2) RT techniques, target volumes, dose fractionation, and treatment planning aims, (3) indications for systemic therapy, (4) sequencing of systemic therapy with RT, (5) effect of lymph node assessment on utilization of adjuvant therapy, and (6) effect of molecular tumor profiling on utilization of adjuvant therapy. Recommendations were based on a systematic literature review and created using a predefined consensus-building methodology and system for quality of evidence grading and strength of recommendation. RESULTS: The task force recommends RT (either vaginal brachytherapy or external beam RT) be given based on the patient's clinical-pathologic risk factors to reduce risk of vaginal and/or pelvic recurrence. When external beam RT is delivered, intensity modulated RT with daily image guided RT is recommended to reduce acute and late toxicity. Chemotherapy is recommended for patients with International Federation of Gynecology and Obstetrics (FIGO) stage I to II with high-risk histologies and those with FIGO stage III to IVA with any histology. When sequencing chemotherapy and RT, there is no prospective data to support an optimal sequence. Sentinel lymph node mapping is recommended over pelvic lymphadenectomy for surgical nodal staging. Data on sentinel lymph node pathologic ultrastaging status supports that patients with isolated tumor cells be treated as node negative and adjuvant therapy based on uterine risk factors and patients with micrometastases be treated as node positive. The available data on molecular characterization of endometrial cancer are compelling and should be increasingly considered when making recommendations for adjuvant therapy. CONCLUSIONS: These recommendations guide evidence-based best clinical practices on the use of adjuvant therapy for endometrial cancer.

Universal suicide screening in college primary care.

OBJECTIVE: Implementation of a universal primary care (PC) suicide-screening program in a college student health center to heighten awareness, provide support and enhance education of staff and students served. Program feasibility, data collection, electronic medical record (EMR) adaptations and staff learning outcomes were examined. Participants: 1,607 students with PC visits during Spring 2018 semester. Methods: Annual Suicide Behaviors Questionnaire (SBQ-R), EMR template, safety alert, referral tracking and simulated staff training were employed. Chi-square tests assessed documentation, safety alerts and mental health referral changes. Paired t-tests evaluated staff learning outcomes. Results: 12.8% of students screened positively for suicide risk during implementation. Documentation consistency, EMR safety alert utilization, mental health referrals and subsequent appointments increased significantly. Staff learning outcomes yielded growth in knowledge and comfort with suicide screening and brief intervention. Conclusion: A comprehensive suicide-screening program is feasible for identifying students at-risk and promoting positive clinical changes in college PC practice settings.

Multiblock Discriminant Analysis of Integrative 18F-FDG-PET/CT Radiomics for Predicting Circulating Tumor Cells in Early-Stage Non-small Cell Lung Cancer Treated With Stereotactic Body Radiation Therapy.

PURPOSE: The main objective of the present study was to integrate 18F-FDG-PET/CT radiomics with multiblock discriminant analysis for predicting circulating tumor cells (CTCs) in early-stage non-small cell lung cancer (ES-NSCLC) treated with stereotactic body radiation therapy (SBRT). METHODS: Fifty-six patients with stage I NSCLC treated with SBRT underwent 18F-FDG-PET/CT imaging pre-SBRT and post-SBRT (median, 5 months; range, 3-10 months). CTCs were assessed via a telomerase-based assay before and within 3 months after SBRT and dichotomized at 5 and 1.3 CTCs/mL. Pre-SBRT, post-SBRT, and delta PET/CT radiomics features (n = 1548 × 3/1562 × 3) were extracted from gross tumor volume. Seven feature blocks were constructed including clinical parameters (n = 12). Multiblock data integration was performed using block sparse partial least squares-discriminant analysis (sPLS-DA) referred to as Data Integration Analysis for Biomarker Discovery Using Latent Components (DIABLO) for identifying key signatures by maximizing common information between different feature blocks while discriminating CTC levels. Optimal input blocks were identified using a pairwise combination method. DIABLO performance for predicting pre-SBRT and post-SBRT CTCs was evaluated using combined AUC (area under the curve, averaged across different blocks) analysis with 20 × 5-fold cross-validation (CV) and compared with that of concatenation-based sPLS-DA that consisted of combining all features into 1 block. CV prediction scores between 1 class versus the other were compared using the Wilcoxon rank sum test. RESULTS: For predicting pre-SBRT CTCs, DIABLO achieved the best performance with combined pre-SBRT PET radiomics and clinical feature blocks, showing CV AUC of 0.875 (P = .009). For predicting post-SBRT CTCs, DIABLO achieved the best performance with combined post-SBRT CT and delta CT radiomics feature blocks, showing CV AUCs of 0.883 (P = .001). In contrast, all single-block sPLS-DA models could not attain CV AUCs higher than 0.7. CONCLUSIONS: Multiblock integration with discriminant analysis of 18F-FDG-PET/CT radiomics has the potential for predicting pre-SBRT and post-SBRT CTCs. Radiomics and CTC analysis may complement and together help guide the subsequent management of patients with ES-NSCLC.

Circulating Tumor Cells Are Associated with Recurrent Disease in Patients with Early-Stage Non-Small Cell Lung Cancer Treated with Stereotactic Body Radiotherapy.

PURPOSE: Although stereotactic body radiotherapy (SBRT) is effective in early-stage non-small cell lung cancer (NSCLC), approximately 10%-15% of patients will fail regionally and 20%-25% distantly. We evaluate a novel circulating tumor cell (CTC) assay as a prognostic marker for increased risk of recurrence following SBRT. EXPERIMENTAL DESIGN: Ninety-two subjects (median age, 71 years) with T1a (64%), T1b (23%), or T2a (13%) stage I NSCLC treated with SBRT were prospectively enrolled. CTCs were enumerated by utilizing a GFP-expressing adenoviral probe that detects elevated telomerase activity in cancer cells. Samples were obtained before, during, and serially up to 24 months after treatment. SBRT was delivered to a median dose of 50 Gy (range, 40-60 Gy), mostly commonly in four to five fractions (92%). RESULTS: Thirty-eight of 92 subjects (41%) had a positive CTC test prior to SBRT. A cutoff of ≥5 CTCs/mL before treatment defined favorable (n = 78) and unfavorable (n = 14) prognostic groups. Increased risk of nodal (P = 0.04) and distant (P = 0.03) failure was observed in the unfavorable group. Within 3 months following SBRT, CTCs continued to be detected in 10 of 35 (29%) subjects. Persistent detection of CTCs was associated with increased risk of distant failure (P = 0.04) and trended toward increased regional (P = 0.08) and local failure (P = 0.16). CONCLUSIONS: Higher pretreatment CTCs and persistence of CTCs posttreatment is significantly associated with increased risk of recurrence outside the targeted treatment site. This suggests that CTC analysis may potentially identify patients at higher risk for regional or distant recurrences and who may benefit from either systemic therapy and/or timely locoregional salvage treatment.

Updating Photon-Based Normal Tissue Complication Probability Models for Pneumonitis in Patients With Lung Cancer Treated With Proton Beam Therapy.

PURPOSE: No validated models for predicting the risk of radiation pneumonitis (RP) with proton beam therapy (PBT) currently exist. Our goal was to externally validate and recalibrate multiple established photon-based normal tissue complication probability models for RP in a cohort with locally advanced nonsmall cell lung cancer treated with contemporary doses of chemoradiation using PBT. METHODS AND MATERIALS: The external validation cohort consisted of 99 consecutive patients with locally advanced nonsmall cell lung cancer treated with chemoradiation using PBT. RP was retrospectively scored at 3 and 6 months posttreatment. We evaluated the performance of the photon Quantitative Analyses of Normal Tissue Effects in the Clinic (QUANTEC) pneumonitis model, the QUANTEC model adjusted for clinical risk factors, and the newer Netherlands updated QUANTEC model. A closed testing procedure was performed to test the need for model updating, either by recalibration-in-the-large (re-estimation of intercept), recalibration (re-estimation of intercept/slope), or model revision (re-estimation of all coefficients). RESULTS: There were 21 events (21%) of ≥grade 2 RP. The closed testing procedure on the PBT data set did not detect major deviations between the models and the data and recommended adjustment of the intercept only for the photon-based Netherlands updated QUANTEC model (intercept update: -1.2). However, an update of the slope and revision of the model coefficients were not recommended by the closed testing procedure, as the deviations were not significant within the power of the data. CONCLUSIONS: The similarity between the dose-response relationship for PBT and photons for normal tissue complications has been an assumption until now. We demonstrate that the preexisting, widely used photon based models fit our PBT data well with minor modifications. These now-validated and updated normal tissue complication probability models can aid in individualizing selection of the most optimal treatment technique for a particular patient.

Mass spectrometry-A versatile tool for characterising the lipid environment of membrane protein assemblies.

Biological membranes are selectively permeable barriers important for cell organization and compartmentalization. Their organisation strongly depends on the lipids that constitute the lipid bilayer as well as the proteins that reside in the membrane. Unravelling the organisation of biological membranes is therefore of great importance to understand cellular function driven by integral and peripheral membrane proteins. Recent developments in mass spectrometry made it a powerful tool contributing to our present-day understanding of membrane composition and organisation. The two main deliverables of mass spectrometry are (i) the identification and quantification of the membrane components, and (ii) the analysis of their structural arrangements. In this review article, we first briefly discuss the aspects of membrane organization that are accessible through mass spectrometry. We then provide detailed insights into the various mass spectrometric strategies which help identifying lipids from membranes or membrane protein assemblies, unravelling the lipid binding modes in membrane proteins and uncovering their structural roles. We further discuss the growing interest in membrane mimetics providing membrane proteins with a native-like lipid environment for structural and functional studies and the possibilities of mass spectrometry to contribute in these experiments.

Early Detection of Recurrence in Patients With Locally Advanced Non-Small-Cell Lung Cancer via Circulating Tumor Cell Analysis.

BACKGROUND: Assays to identify circulating tumor cells (CTCs) might allow for noninvasive and sequential monitoring of lung cancer. We investigated whether serial CTC analysis could complement conventional imaging for detecting recurrences after treatment in patients with locally advanced non-small-cell lung cancer (LA-NSCLC). PATIENTS AND METHODS: Patients with LA-NSCLC (stage II-III) who definitively received concurrent chemoradiation were prospectively enrolled, with CTCs from peripheral blood samples identified using an adenoviral probe that detects elevated telomerase activity present in nearly all lung cancer cells. A "detectable" CTC level was defined as 1.3 green flourescent protein-positive cells per milliliter of collected blood. Samples were obtained before, during (at weeks 2, 4, and 6), and after treatment (post-radiation therapy [RT]; at months 1, 3, 6, 12, 18, and 24). RESULTS: Forty-eight patients were enrolled. At a median follow-up of 10.9 months, 22 (46%) patients had disease recurrence at a median time of 7.6 months post-RT (range, 1.3-32.0 months). Of the 20 of 22 patients for whom post-RT samples were obtained, 15 (75%) had an increase in CTC counts post-RT. In 10 of these 15 patients, CTCs were undetectable on initial post-RT draw but were then detected again before radiographic detection of recurrence, with a median lead time of 6.2 months and mean lead time of 6.1 months (range, 0.1-12.0 months) between CTC count increase and radiographic evidence of recurrence. One patient with an early recurrence (4.7 months) had persistently elevated detectable CTC levels during and after treatment. CONCLUSION: These results indicate that longitudinal CTC monitoring in patients with LA-NSCLC treated with chemoradiation is feasible, and that detectable CTC levels in many patients meaningfully precede radiologic evidence of disease recurrence.

Five-year Long-term Outcomes of Stereotactic Body Radiation Therapy for Operable Versus Medically Inoperable Stage I Non-small-cell Lung Cancer: Analysis by Operability, Fractionation Regimen, Tumor Size, and Tumor Location.

BACKGROUND: Stereotactic body radiation therapy (SBRT) is standard for medically inoperable stage I non-small-cell lung cancer (NSCLC) and is emerging as a surgical alternative in operable patients. However, limited long-term outcomes data exist, particularly according to operability. We hypothesized long-term local control (LC) and cancer-specific survival (CSS) would not differ by fractionation schedule, tumor size or location, or operability status, but overall survival (OS) would be higher for operable patients. PATIENTS AND METHODS: All consecutive patients with stage I (cT1-2aN0M0) NSCLC treated with SBRT from June 2009 to July 2013 were assessed. Thoracic surgeon evaluation determined operability. Local failure was defined as growth following initial tumor shrinkage or progression on consecutive scans. LC, CSS, and OS were calculated using Cox proportional hazards regression. RESULTS: A total of 186 patients (204 lesions) were analyzed. Most patients were inoperable (82%) with Eastern Cooperative Oncology Group performance status of 1 (59%) or 2 (26%). All lesions received biological effective doses ≥ 100 Gy most commonly (94%) in 3 to 5 fractions. The median follow-up was 4.0 years. LC at 2 and 5 years were 95.6% (95% confidence interval, 92%-99%) and 93.7% (95% confidence interval, 90%-98%), respectively. Compared with operable patients, inoperable patients did not have significant differences in 5-year LC (93.1% vs. 96.7%; P = .49), nodal failure (31.4% vs. 11.0%; P = .12), distant failure (12.2% vs. 10.4%; P = .98), or CSS (80.6% vs. 91.0%; P = .45) but trended towards worse OS (34.2% vs. 45.3%; P = .068). Tumor size, location, and fractionation did not significantly influence outcomes. CONCLUSIONS: SBRT has excellent, durable LC and CSS rates for early-stage NSCLC, although inoperable patients had somewhat lower OS than operable patients, likely owing to greater comorbidities.

Genetic Interactions Between BOB1 and Multiple 26S Proteasome Subunits Suggest a Role for Proteostasis in Regulating Arabidopsis Development.

Protein folding and degradation are both required for protein quality control, an essential cellular activity that underlies normal growth and development. We investigated how BOB1, an Arabidopsis thaliana small heat shock protein, maintains normal plant development. bob1 mutants exhibit organ polarity defects and have expanded domains of KNOX gene expression. Some of these phenotypes are ecotype specific suggesting that other genes function to modify them. Using a genetic approach we identified an interaction between BOB1 and FIL, a gene required for abaxial organ identity. We also performed an EMS enhancer screen using the bob1-3 allele to identify pathways that are sensitized by a loss of BOB1 function. This screen identified genetic, but not physical, interactions between BOB1 and the proteasome subunit RPT2a Two other proteasome subunits, RPN1a and RPN8a, also interact genetically with BOB1 Both BOB1 and the BOB1-interacting proteasome subunits had previously been shown to interact genetically with the transcriptional enhancers AS1 and AS2, genes known to regulate both organ polarity and KNOX gene expression. Our results suggest a model in which BOB1 mediated protein folding and proteasome mediated protein degradation form a functional proteostasis module required for ensuring normal plant development.

Patient-Reported Survivorship Care Practices and Late Effects After Treatment of Hodgkin and Non-Hodgkin Lymphoma.

PURPOSE: Multimodal treatment of Hodgkin lymphoma (HL) and non-Hodgkin lymphoma (NHL) yields excellent outcomes; however, survivors are at risk of developing myriad late and long-term effects. METHODS: From a convenience sample of 964 survivors of HL (37%) and NHL (63%) using a publicly available Internet-based survivorship care plan (SCP) tool between 2011 and 2016, we examined patient-reported cancer care, toxicities, and survivorship care data. RESULTS: Of all survivors, 67% were female and 84% were white and 88% were free of cancer. Median age of diagnosis was 28 years for survivors of HL and 49 years for NHL. Many survivors reported treatment with chemotherapy (92%), surgery (52%), and/or radiation (41%), with most radiation delivered to chest/mantle fields (81%). Survivors reported a diversity of radiation- and chemotherapy-related sequelae, including thyroid dysfunction, speaking and/or swallowing changes, pulmonary fibrosis/pneumonitis, heart disease, chronic fatigue, neurocognitive decline, neuropathy, sexual changes, and secondary breast cancers. Few reported receipt of previous survivorship information. Most reported management/comanagement by an oncology specialist after active treatment; however, a shift to management by primary care provider alone was observed as a trend over time in follow-up. Sixty-six percent of users who responded to a follow-up survey reported that they intend to share the SCP with their health care team. CONCLUSION: Survivors of lymphoma, many of whom are free of disease, report a substantial burden of late and long-term adverse effects, suboptimal delivery of survivorship information, and transitions of care in follow-up in which fragmented systems and/or poor communication may contribute to unmet survivor needs. Multiple opportunities thus exist for which SCPs may be used to improve awareness regarding survivorship and associated adverse effects in addition to communicating follow-up care plans between survivors and treatment teams.

Outcomes of invasive mediastinal nodal staging versus positron emission tomography staging alone for early-stage non-small cell lung cancer treated with stereotactic body radiation therapy.

BACKGROUND: The benefit of invasive mediastinal nodal staging (IMNS) in addition to positron emission tomography-computed tomography (PET/CT) is undefined for early stage non-small cell lung cancer (NSCLC). This multi-institutional investigation aimed to evaluate outcomes and patterns of failure in patients staged with PET/CT with or without additional IMNS. METHODS: Two academic centers assessed all consecutive patients staged with PET/CT for early-stage, primary lung NSCLC (cT1-2aN0M0) treated with SBRT. Local recurrence-free survival (LRFS), nodal recurrence-free survival (NRFS), distant metastasis-free survival (DMFS), and overall survival (OS) were calculated using Kaplan-Meier methodology. Univariate and multivariate Cox proportional hazards modeling addressed factors associated with outcomes. RESULTS: Overall, 180 patients (199 lesions) were staged with PET/CT alone and 56 patients (58 lesions) underwent additional IMNS. Among patients receiving IMNS, 52 (93%) underwent EBUS and 4 (7%) underwent mediastinoscopy. At a median follow-up of 33.5 months (range, 1.9-80.9 months), there was no significant difference in LRFS (37 vs. 47 months, p=0.309), NRFS (34 vs. 42 months p=0.370), DMFS (36 vs. 47 months, p=0.234) or OS (37 vs. 47 months, p=0.236) between patients undergoing PET/CT-only versus PET/CT+IMNS staging, respectively. Receipt of IMNS did not correlate with any outcome on either univariate or multivariate analysis (p>0.05). Patterns of failure in both groups were similar, including crude isolated nodal failure rates (8% PET/CT-only versus 14% PET+IMNS group, p=0.202). CONCLUSIONS: Patients undergoing IMNS had similar survival and patterns of recurrence as those receiving PET/CT alone. Further study, ideally prospectively, is needed to determine which subgroups might benefit from IMNS.