The management of stage I Non-Small cell lung cancer (NSCLC) in Ontario: A Population-Based study of patterns of care and Stereotactic Ablative Body radiotherapy (SABR) utilization from 2010 to 2019.

BACKGROUND: Stereotactic Ablative Body Radiotherapy (SABR) is the standard of care for medically inoperable patients with Stage I NSCLC. The adoption of SABR and its association with cancer outcomes requires characterization. AIM: We described the management of biopsy-proven Stage I NSCLC with SABR, surgery, non-SABR curative radiotherapy (RT) and observation in Ontario, Canada, between 2010 and 2019. Temporal and geographic trends in practice and survival outcomes were analyzed. METHODS: This was a retrospective population-based cohort study conducted by linking electronic radiotherapy (RT) records to a population-based cancer registry. RESULTS: A total of 12,065 patients were identified, 61.7 % underwent surgery, 17.9 % received SABR, 8.6 % received non-SABR curative RT and 11.7 % were observed. Between 2010 and 2019, the utilization of surgery decreased (63.8 % to 49.9 %, p < 0.0001), while SABR use increased (7.5 % to 24.4 %, p < 0.0001), non-SABR curative RT use increased (6.7 % to 9.6 %, p < 0.0014) and patients observed decreased (14.4 % to 12.0 %, p < 0.0001). Substantial variation in practice exists across Ontario. Two- yr CSS improved for the entire cohort (81.9 % to 85.0 %, p < 0.0001). While there was improvement in 2 yr CSS for surgical patients (92.1 %% to 95.7 %, p < 0.001), survival for patients who received SABR, Non-SABR curative RT and observation remained stable. CONCLUSION: There has been an increase in SABR utilization and a reduction in surgical utilization with a corresponding increased survival of stage I patients in Ontario between 2010 and 2019. Substantial differences in practice patterns exist across health regions, suggesting the need for strategies to improve access to SABR in many jurisdictions.

Survival outcomes and predicting intracranial metastasis in stage III non-small cell lung cancer treated with definitive chemoradiation: Real-world data from a tertiary cancer center.

PURPOSE/OBJECTIVE: Around 30% of patients with non-small cell lung cancers (NSCLC) are diagnosed with stage III disease at presentation, of which about 50% are treated with definitive chemoradiation (CRT). Around 65-80% of patients will eventually develop intracranial metastases (IM), though associated risk factors are not clearly described. We report survival outcomes and risk factors for development of IM in a cohort of patients with stage III NSCLC treated with CRT at a tertiary cancer center. MATERIALS/METHODS: We identified 195 patients with stage III NSCLC treated with CRT from January 2010 to May 2021. Multivariable logistic regression was used to generate odds ratios for covariates associated with development of IM. Kaplan-Meier analysis with the Log Rank test was used for unadjusted time-to-event analyses. P-value for statistical significance was set at < 0.05 with a two-sided test. RESULTS: Out of 195 patients, 108 (55.4%) had stage IIIA disease and 103 (52.8%) had adenocarcinoma histology. The median age and follow-up (in months) was 67 (IQR 60-74) and 21 (IQR 12-43), respectively. The dose of radiation was 60 Gy in 30 fractions for148 patients (75.9%). Of the 77 patients who received treatment since immunotherapy was available and standard at our cancer center, 45 (58.4%) received at least one cycle. During follow-up, 84 patients (43.1%) developed any metastasis, and 33 (16.9%) developed IM (either alone or with extracranial metastasis). 150 patients (76.9%) experienced a treatment delay (interval between diagnosis and treatment > 4 weeks). Factors associated with developing any metastasis included higher overall stage at diagnosis (p = 0.013) and higher prescribed dose (p = 0.022). Factors associated with developing IM included higher ratio of involved over sampled lymph nodes (p = 0.001) and receipt of pre-CRT systemic or radiotherapy for any reason (p = 0.034). On multivariate logistical regression, treatment delay (OR 3.9, p = 0.036) and overall stage at diagnosis (IIIA vs. IIIB/IIIC) (OR 2.8, p = 0.02) predicted development of IM. These findings were sustained on sensitivity analysis using different delay intervals. Median OS was not reached for the overall cohort, and was 43.1 months for patients with IM and 40.3 months in those with extracranial-only metastasis (p = 0.968). In patients with any metastasis, median OS was longer (p = 0.003) for those who experienced a treatment delay (48.4 months) compared to those that did not (12.2 months), likely due to expedited diagnosis and treatment in patients with a higher symptom burden secondary to more advanced disease. CONCLUSIONS: In patients with stage III NSCLC treated with definitive CRT, the risk of IM appears to increase with overall stage at diagnosis and, importantly, may be associated with experiencing a treatment delay (> 4 weeks). Metastatic disease of any kind remains the primary life-limiting prognostic factor in these patients with advanced lung cancer. In patients with metastatic disease, treatment delay was associated with better survival. Patients who experience a treatment delay and those initially diagnosed at a more advanced overall stage may warrant more frequent surveillance for early diagnosis and treatment of IM. Healthcare system stakeholders should strive to mitigate treatment delay in patients with locally NSCLC to reduce the risk of IM. Further research is needed to better understand factors associated with survival, treatment delay, and the development of IM after CRT in the immunotherapy era.

Lung SBRT credentialing in the Canadian OCOG-LUSTRE randomized trial.

Purpose: To report on the Stereotactic Body Radiation Therapy (SBRT) credentialing experience during the Phase III Ontario Clinical Oncology Group (OCOG) LUSTRE trial for stage I non-small cell lung cancer. Methods: Three credentialing requirements were required in this process: (a) An institutional technical survey; (b) IROC (Imaging and Radiation Oncology Core) thoracic phantom end-to-end test; and (c) Contouring and completion of standardized test cases using SBRT for one central and one peripheral lung cancer, compared against the host institution as the standard. The main hypotheses were that unacceptable variation would exist particularly in OAR definition across all centres, and that institutions with limited experience in SBRT would be more likely to violate per-protocol guidelines. Results: Fifteen Canadian centres participated of which 8 were new, and 7 were previously established (≥2 years SBRT experience), and all successfully completed surveys and IROC phantom testing. Of 30 SBRT test plans, 10 required replanning due to major deviations, with no differences in violations between new and established centres (p = 0.61). Mean contouring errors were highest for brachial plexus in the central (C) case (12.55 ± 6.62 mm), and vessels in the peripheral (P) case (13.01 ± 12.55 mm), with the proximal bronchial tree (PBT) (2.82 ± 0.78 C, 3.27 ± 1.06 P) as another variable structure. Mean dice coefficients were lowest for plexus (0.37 ± 0.2 C, 0.37 ± 0.14 P), PBT (0.77 ± 0.06 C, 0.75 ± 0.09 P), vessels (0.69 ± 0.29 C, 0.64 ± 0.31 P), and esophagus (0.74 ± 0.04 C, 0.76 ± 0.04 P). All plans passed per-protocol planning target volume (PTV) coverage and maximum/volumetric organs-at-risk constraints, although variations existed in dose gradients within and outside the target. Conclusions: Clear differences exist in both contouring and planning with lung SBRT, regardless of centre experience. Such an exercise is important for studies that rely on high precision radiotherapy, and to ensure that implications on trial quality and outcomes are as optimal as possible.

Low-dose aspirin confers protection against acute cellular allograft rejection after primary liver transplantation.

This study investigated the effect of low-dose aspirin in primary adult liver transplantation (LT) on acute cellular rejection (ACR) as well as arterial patency rates. The use of low-dose aspirin after LT is practiced by many transplant centers to minimize the risk of hepatic artery thrombosis (HAT), although solid recommendations do not exist. However, aspirin also possesses potent anti-inflammatory properties and might mitigate inflammatory processes after LT, such as rejection. Therefore, we hypothesized that the use of aspirin after LT has a protective effect against ACR. This is an international, multicenter cohort study of primary adult deceased donor LT. The study included 17 high-volume LT centers and covered the 3-year period from 2013 to 2015 to allow a minimum 5-year follow-up. In this cohort of 2365 patients, prophylactic antiplatelet therapy with low-dose aspirin was administered in 1436 recipients (61%). The 1-year rejection-free survival rate was 89% in the aspirin group versus 82% in the no-aspirin group (hazard ratio [HR], 0.77; 95% confidence interval [CI], 0.63-0.94; p = 0.01). The 1-year primary arterial patency rates were 99% in the aspirin group and 96% in the no-aspirin group with an HR of 0.23 (95% CI, 0.13-0.40; p < 0.001). Low-dose aspirin was associated with a lower risk of ACR and HAT after LT, especially in the first vulnerable year after transplantation. Therefore, low-dose aspirin use after primary LT should be evaluated to protect the liver graft from ACR and to maintain arterial patency.

Outcomes of Hypofractionated Stereotactic Radiotherapy for Small and Moderate-Sized Brain Metastases: A Single-Institution Analysis.

Background: Stereotactic radiosurgery (SRS) is the standard treatment for limited intracranial metastases. With the advent of frameless treatment delivery, fractionated stereotactic radiotherapy (FSRT) has become more commonly implemented given superior control and toxicity rates for larger lesions. We reviewed our institutional experience of FSRT to brain metastases without size restriction. Methods: We performed a retrospective review of our institutional database of patients treated with FSRT for brain metastases. Clinical and dosimetric details were abstracted. All patients were treated in 3 or 5 fractions using LINAC-based FSRT, did not receive prior cranial radiotherapy, and had at least 6 months of MRI follow-up. Overall survival was estimated using the Kaplan-Meier method. Local failure and radionecrosis cumulative incidence rates were estimated using a competing risks model with death as the competing risk. Univariable and multivariable analyses using Fine and Gray's proportional subdistribution hazards regression model were performed to determine covariates predictive of local failure and radionecrosis. Results: We identified 60 patients and 133 brain metastases treated at our institution from 2016 to 2020. The most common histologies were lung (53%) and melanoma (25%). Most lesions were >1 cm in diameter (84.2%) and did not have previous surgical resection (88%). The median duration of imaging follow-up was 9.8 months. The median survival for the whole cohort was 20.5 months. The local failure at 12 months was 17.8% for all lesions, 22.1% for lesions >1 cm, and 13.7% for lesions ≤1 cm (p = 0.36). The risk of radionecrosis at 12 months was 7.1% for all lesions, 13.2% for lesions >1 cm, and 3.2% for lesions ≤1 cm (p = 0.15). Conclusions: FSRT is safe and effective in the treatment of brain metastases of any size with excellent local control and toxicity outcomes. Prospective evaluation against single-fraction SRS is warranted for all lesion sizes.

A Comparison of Hypofractionated and Twice-Daily Thoracic Irradiation in Limited-Stage Small-Cell Lung Cancer: An Overlap-Weighted Analysis.

Despite evidence for the superiority of twice-daily (BID) radiotherapy schedules, their utilization in practice remains logistically challenging. Hypofractionation (HFRT) is a commonly implemented alternative. We aim to compare the outcomes and toxicities in limited-stage small-cell lung cancer (LS-SCLC) patients treated with hypofractionated versus BID schedules. A bi-institutional retrospective cohort review was conducted of LS-SCLC patients treated with BID (45 Gy/30 fractions) or HFRT (40 Gy/15 fractions) schedules from 2007 to 2019. Overlap weighting using propensity scores was performed to balance observed covariates between the two radiotherapy schedule groups. Effect estimates of radiotherapy schedule on overall survival (OS), locoregional recurrence (LRR) risk, thoracic response, any ≥grade 3 (including lung, and esophageal) toxicity were determined using multivariable regression modelling. A total of 173 patients were included in the overlap-weighted analysis, with 110 patients having received BID treatment, and 63 treated by HFRT. The median follow-up was 20.4 months. Multivariable regression modelling did not reveal any significant differences in OS (hazard ratio [HR] 1.67, p = 0.38), LRR risk (HR 1.48, p = 0.38), thoracic response (odds ratio [OR] 0.23, p = 0.21), any ≥grade 3+ toxicity (OR 1.67, p = 0.33), ≥grade 3 pneumonitis (OR 1.14, p = 0.84), or ≥grade 3 esophagitis (OR 1.41, p = 0.62). HFRT, in comparison to BID radiotherapy schedules, does not appear to result in significantly different survival, locoregional control, or toxicity outcomes.

The Economic Burden of Postoperative Complications Predicted by the Comprehensive Complication Index® in Patients Undergoing Elective Major Hepatopancreaticobiliary Surgery for Malignancy - A Prospective Cost Analysis.

BACKGROUND/AIM: Postoperative complications have a tremendous impact on in-hospital costs. The novel Comprehensive Complication Index® (CCI®) summarizes all complications together and is more sensitive than existing morbidity endpoints. The purpose of this study was to assess the correlation of CCI® with in-hospital costs and externally validate a novel cost prediction calculator. PATIENTS AND METHODS: This was a prospective study including consecutive patients undergoing elective major hepatopancreaticobiliary (HPB) surgery for malignancy at a London tertiary referral hospital. A priori sample size and post-hoc power calculations were performed. RESULTS: Thirty patients were included in the analysis, 14 were female, and the median age was 67 [interquartile range (IQR)=54-74] years. The median Charlson Comorbidity Index was 6 (IQR=5-8). Eighteen patients underwent liver, 9 pancreatic surgery and three a palliative bypass; 11 patients had a major complication (≥grade 3a) according to the Clavien-Dindo classification. The median CCI® was 30.2 (IQR=12.18-39.5). The mean cost per case was 13,908 (SD=4,600) GBP. There was no correlation between the Charlson Comorbidity Index or age with actual cost. However, there was very good correlation of actual cost with the CCI® (r=0.77, 95% confidence interval=0.57-0.89, p<0.001) as well as with the predicted cost (Clavien Cost Prediction Calculator) (r=0.70, 95% confidence interval=0.44-0.85, p<0.001). CONCLUSION: These findings support the hypothesis that complications are the most important predictor of overall cost in the setting of elective major HPB surgery for malignancy. Furthermore, CCI® and the novel Cost Prediction Calculator can be used in this setting to accurately predict costs using no additional resources.

Signatures of Post-Glacial Genetic Isolation and Human-Driven Migration in the Dothistroma Needle Blight Pathogen in Western Canada.

Many current tree improvement programs are incorporating assisted gene flow strategies to match reforestation efforts with future climates. This is the case for the lodgepole pine (Pinus contorta var. latifolia), the most extensively planted tree in western Canada. Knowledge of the structure and origin of pathogen populations associated with this tree would help improve the breeding effort. Recent outbreaks of the Dothistroma needle blight (DNB) pathogen Dothistroma septosporum on lodgepole pine in British Columbia and its discovery in Alberta plantations raised questions about the diversity and population structure of this pathogen in western Canada. Using genotyping-by-sequencing on 119 D. septosporum isolates from 16 natural pine populations and plantations from this area, we identified four genetic lineages, all distinct from the other DNB lineages from outside of North America. Modeling of the population history indicated that these lineages diverged between 31.4 and 7.2 thousand years ago, coinciding with the last glacial maximum and the postglacial recolonization of lodgepole pine in western North America. The lineage found in the Kispiox Valley from British Columbia, where an unprecedented DNB epidemic occurred in the 1990s, was close to demographic equilibrium and displayed a high level of haplotypic diversity. Two lineages found in Alberta and Prince George (British Columbia) showed departure from random mating and contemporary gene flow, likely resulting from pine breeding activities and material exchanges in these areas. The increased movement of planting material could have some major consequences by facilitating secondary contact between genetically isolated DNB lineages, possibly resulting in new epidemics.

How to Handle Arterial Conduits in Liver Transplantation? Evidence From the First Multicenter Risk Analysis.

OBJECTIVE: The aims of the present study were to identify independent risk factors for conduit occlusion, compare outcomes of different AC placement sites, and investigate whether postoperative platelet antiaggregation is protective. BACKGROUND: Arterial conduits (AC) in liver transplantation (LT) offer an effective rescue option when regular arterial graft revascularization is not feasible. However, the role of the conduit placement site and postoperative antiaggregation is insufficiently answered in the literature. STUDY DESIGN: This is an international, multicenter cohort study of adult deceased donor LT requiring AC. The study included 14 LT centers and covered the period from January 2007 to December 2016. Primary endpoint was arterial occlusion/patency. Secondary endpoints included intra- and perioperative outcomes and graft and patient survival. RESULTS: The cohort was composed of 565 LT. Infrarenal aortic placement was performed in 77% of ACs whereas supraceliac placement in 20%. Early occlusion (≤30 days) occurred in 8% of cases. Primary patency was equivalent for supraceliac, infrarenal, and iliac conduits. Multivariate analysis identified donor age >40 years, coronary artery bypass, and no aspirin after LT as independent risk factors for early occlusion. Postoperative antiaggregation regimen differed among centers and was given in 49% of cases. Graft survival was significantly superior for patients receiving aggregation inhibitors after LT. CONCLUSION: When AC is required for rescue graft revascularization, the conduit placement site seems to be negligible and should follow the surgeon's preference. In this high-risk group, the study supports the concept of postoperative antiaggregation in LT requiring AC.

Protein-Nucleic Acid Conjugation with Sterol Linkers Using Hedgehog Autoprocessing.

Hedgehog (Hh) precursor proteins contain an autoprocessing domain called HhC whose native function is protein cleavage and C-terminal glycine sterylation. The transformation catalyzed by HhC occurs in cis from a precursor protein and exhibits wide tolerance toward both sterol and protein substrates. Here, we repurpose HhC as a 1:1 protein-nucleic acid ligase, with the sterol serving as a molecular linker. A procedure is described for preparing HhC-active sterylated DNA, called steramers, using aqueous compatible chemistry and commercial reagents. Steramers have KM values of 7-11 µM and reaction t1/2 values of ∼10 min. Modularity of the HhC/steramer method is demonstrated using four different proteins along with structured and unstructured sterylated nucleic acids. The resulting protein-DNA conjugates retain the native solution properties and biochemical function. Unlike self-tagging domains, HhC does not remain fused to the conjugate; rather, enzymatic activity is mechanistically coupled to conjugate release. That unique feature of HhC, coupled with efficient kinetics and substrate tolerance, may ease access and open new applications for these suprabiological chimeras.

Improving Timeliness of Oncology Assessment and Cancer Treatment Through Implementation of a Multidisciplinary Lung Cancer Clinic.

PURPOSE: Timely lung cancer care has been associated with improved clinical outcomes and patient satisfaction. We identified improvement opportunities in lung cancer management pathways at Kingston Health Sciences Centre. Quality improvement strategies led to the implementation of a multidisciplinary lung cancer clinic (MDC). METHODS: We set an outcome measure of decreasing the time from diagnosis to first cancer treatment by 10 days within 6 months of clinic implementation. We implemented a weekly MDC that involved respirologists, medical oncologists, and radiation oncologists at which patients with new lung cancer diagnoses were offered concurrent oncology consultation. We used Plan-Do-Study-Act cycles to guide our improvement initiatives. A total of five Plan-Do-Study-Act cycles spanned 14 months and consisted of an MDC pilot clinic, large-scale MDC launching, debriefing meetings, and clinic expansion. Pre-MDC data were analyzed retrospectively to establish baseline and prospectively for improvement. Statistical Process Control XmR(i) charts were used to report data. RESULTS: Since MDC initiation, 128 patients have been seen in 34 MDC clinics (3.8 patients per clinic). Mean days from diagnosis to first oncology assessment decreased from 12.4 days to 3.9 days, and mean days from diagnosis to first cancer treatment decreased from 39.5 to 15.0 days, both of which demonstrated special cause variation. Time to assessment and treatment improved for patients with every stage of lung cancer and for both small-cell and non-small-cell subtypes. CONCLUSION: MDC shortens the time from lung cancer diagnosis to oncology assessment and treatment. Time to treatment improved more than time to oncology assessment, which suggests the improvement is related to benefits beyond faster oncology assessment.

Therapeutic interventions in children and adolescents with patellar tendon related pain: a systematic review.

OBJECTIVE: Evaluate effectiveness and harms of interventions for patellar tendon related pain in children and adolescents. DESIGN: Systematic review and meta-analysis. DATA SOURCES: Medline via Pubmed, Embase via OVID, CINAHL via Ebsco, SportDiscus up until 24 November 2017 were searched. ELIGIBILITY CRITERIA FOR SELECTING STUDIES: Inclusion criteria were (1) controlled or randomised controlled clinical trials (RCTs), (2) participants with diagnosis of patellar tendon related disorder, (3) participants≤18 years of age at enrolment and (4) published in a peer-reviewed English or Scandinavian language journal. RESULTS: Of 530 studies identified, eight were included after screening, with three included in data synthesis. To be included in data synthesis, we required studies to have included (and have data available for) a minimum of 10 participants under 18 years. All studies were rated as being at high risk of bias. For adolescents with patellar tendinopathy, one RCT compared eccentric exercises to usual care and found no difference between groups. In adolescents with Osgood-Schlatter disease (OSD), injection of local anaesthetic with dextrose proved superior to either usual care or local anaesthetic alone (three armed RCTs). In a retrospective case controlled study in adolescents with OSD, surgery provided no benefit over conservative management in terms of persistent symptoms and had a higher complication rate. CONCLUSION: There is weak evidence to support the use of dextrose injection with local anaesthetic and no evidence to support the use of specific types of exercises to treat children/adolescents with OSD/patellar tendinopathy. Until further evidence arises, clinicians should include load modification and advise on a return to sport based on symptoms.

Increasing the Stability of Recombinant Human Green Cone Pigment.

Three types of cone cells exist in the human retina, each containing a different pigment responsible for the initial step of phototransduction. These pigments are distinguished by their specific absorbance maxima: 425 nm (blue), 530 nm (green), and 560 nm (red). Each pigment contains a common chromophore, 11-cis-retinal covalently bound to an opsin protein via a Schiff base. The 11-cis-retinal protonated Schiff base has an absorbance maxima at 440 nm in methanol. Unfortunately, the chemistry that allows the same chromophore to interact with different opsin proteins to tune the absorbance of the resulting pigments to distinct λmax values is poorly understood. Rhodopsin is the only pigment with a native structure determined at high resolution. Homology models for cone pigments have been generated, but experimentally determined structures are needed for a precise understanding of spectral tuning. The principal obstacle to solving the structures of cone pigments has been their innate instability in recombinant constructs. By inserting five different thermostabilizing proteins (BRIL, T4L, PGS, RUB, and FLAV) into the recombinant green opsin sequence, constructs were created that were up to 9-fold more stable than WT. Using cellular retinaldehyde-binding protein (CRALBP), we developed a quick means of assessing the stability of the green pigment. CRALBP testing also confirmed an additional 48-fold increase in pigment stability when varying the detergent used. These results suggest an efficient protocol for routine purification and stabilization of cone pigments that could be used for high-resolution determination of their structures, as well as for other studies.

Quality of life and swallowing with standard chemoradiotherapy versus accelerated radiotherapy and panitumumab in locoregionally advanced carcinoma of the head and neck: A phase III randomised trial from the Canadian Cancer Trials Group (HN.6).

AIM: To compare quality of life (QOL) between standard (SFX) chemoradiotherapy (arm A) and altered fractionation radiotherapy (AFX) with panitumumab (PMab; arm B). METHODS: Patients with T any N + M0 or T3-4N0M0 squamous cell head-neck carcinoma were randomised to SFX (70 Gy/35/7 wks) plus cisplatin (100 mg/m2 IV × 3) versus AFX (70 Gy/35/6 wks) plus PMab (9 mg/kg IV × 3). QOL was collected at baseline, end of radiation therapy (RT) and 2, 4, 6, 12, 24 and 36 months post-RT using the Functional Assessment of Cancer Therapy Head and Neck (FACT-H&N), MD Anderson Dysphagia Index (MDADI) and SWAL-QOL. We hypothesised a 6-point more favourable change in FACT-H&N score from baseline to 1 year in arm B over arm A. RESULTS: Among 320 patients, median follow-up was 46 (range: 0.1-64.3) months, median age 56, 84% male, Eastern Cooperative Oncology Group PS 0 (71%), 1 (29%). Primary site was oropharynx in 81% (p16+ 68%, p16- 16%, missing 16%). Baseline scores did not differ by arm (A/B): FACT-H&N 116.5/115, MDADI Global 83/77, SWAL-QOL General 67/68. At 1 year, no difference was seen between arms in FACT-H&N change from baseline: A -1.70, B -4.81, p = 0.194. Subscale change scores by arm were (A/B): last week RT, FACT-Physical (-11.6, -10, p = 0.049), MDADI Physical (-40.4, -33.9, p = 0.045), and SWAL-QOL Eating Duration (-61.2, -51.2, p = 0.02), Eating Desire (-53.3, -43.9, p = 0.031) and Mental Health (-42, -32.6, p = 0.009); 4 months, HN subscale (-7.7, -10, p = 0.014). No clinically important differences by arm were seen post-treatment. CONCLUSIONS: PMab with AFX did not durably improve QOL or swallowing as compared with SFX with cisplatin. TRIAL REGISTRATION: ClinicalTrials.gov: NCT00820248.

Canadian Phase III Randomized Trial of Stereotactic Body Radiotherapy Versus Conventionally Hypofractionated Radiotherapy for Stage I, Medically Inoperable Non-Small-Cell Lung Cancer - Rationale and Protocol Design for the Ontario Clinical Oncology Group (OCOG)-LUSTRE Trial.

We describe a Canadian phase III randomized controlled trial of stereotactic body radiotherapy (SBRT) versus conventionally hypofractionated radiotherapy (CRT) for the treatment of stage I medically inoperable non-small-cell lung cancer (OCOG-LUSTRE Trial). Eligible patients are randomized in a 2:1 fashion to either SBRT (48 Gy in 4 fractions for peripherally located lesions; 60 Gy in 8 fractions for centrally located lesions) or CRT (60 Gy in 15 fractions). The primary outcome of the study is 3-year local control, which we hypothesize will improve from 75% with CRT to 87.5% with SBRT. With 85% power to detect a difference of this magnitude (hazard ratio = 0.46), a 2-sided α = 0.05 and a 2:1 randomization, we require a sample size of 324 patients (216 SBRT, 108 CRT). Important secondary outcomes include overall survival, disease-free survival, toxicity, radiation-related treatment death, quality of life, and cost-effectiveness. A robust radiation therapy quality assurance program has been established to assure consistent and high quality SBRT and CRT delivery. Despite widespread interest and adoption of SBRT, there still remains a concern regarding long-term control and risks of toxicity (particularly in patients with centrally located lesions). The OCOG-LUSTRE study is the only randomized phase III trial testing SBRT in a medically inoperable population, and the results of this trial will attempt to prove that the benefits of SBRT outweigh the potential risks.

A Single Aspartate Coordinates Two Catalytic Steps in Hedgehog Autoprocessing.

Hedgehog (Hh) signaling is driven by the cholesterol-modified Hh ligand, generated by autoprocessing of Hh precursor protein. Two steps in Hh autoprocessing, N-S acyl shift and transesterification, must be coupled for efficient Hh cholesteroylation and downstream signal transduction. In the present study, we show that a conserved aspartate residue, D46 of the Hh autoprocessing domain, coordinates these two catalytic steps. Mutagenesis demonstrated that D46 suppresses non-native Hh precursor autoprocessing and is indispensable for transesterification with cholesterol. NMR measurements indicated that D46 has a pKa of 5.6, ∼2 units above the expected pKa of aspartate, due to a hydrogen-bond between protonated D46 and a catalytic cysteine residue. However, the deprotonated form of D46 side chain is also essential, because a D46N mutation cannot mediate cholesteroylation. On the basis of these data, we propose that the proton shuttling of D46 side chain mechanistically couples the two steps of Hh cholesteroylation.

Förster resonance energy transfer-based cholesterolysis assay identifies a novel hedgehog inhibitor.

Hedgehog (Hh) proteins function in cell/cell signaling processes linked to human embryo development and the progression of several types of cancer. Here, we describe an optical assay of hedgehog cholesterolysis, a unique autoprocessing event critical for Hh function. The assay uses a recombinant Förster resonance energy transfer (FRET)-active Hh precursor whose cholesterolysis can be monitored continuously in multi-well plates (dynamic range=4, Z'=0.7), offering advantages in throughput over conventional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) assays. Application of the optical assay in a pilot small molecule screen produced a novel cholesterolysis inhibitor (apparent IC50=5×10(-6)M) that appears to inactivate hedgehog covalently by a substitution nucleophilic aromatic (SNAr) mechanism.

Zinc inhibits Hedgehog autoprocessing: linking zinc deficiency with Hedgehog activation.

Zinc is an essential trace element with wide-ranging biological functions, whereas the Hedgehog (Hh) signaling pathway plays crucial roles in both development and disease. Here we show that there is a mechanistic link between zinc and Hh signaling. The upstream activator of Hh signaling, the Hh ligand, originates from Hh autoprocessing, which converts the Hh precursor protein to the Hh ligand. In an in vitro Hh autoprocessing assay we show that zinc inhibits Hh autoprocessing with a Ki of 2 µm. We then demonstrate that zinc inhibits Hh autoprocessing in a cellular environment with experiments in primary rat astrocyte culture. Solution NMR reveals that zinc binds the active site residues of the Hh autoprocessing domain to inhibit autoprocessing, and isothermal titration calorimetry provided the thermodynamics of the binding. In normal physiology, zinc likely acts as a negative regulator of Hh autoprocessing and inhibits the generation of Hh ligand and Hh signaling. In many diseases, zinc deficiency and elevated level of Hh ligand co-exist, including prostate cancer, lung cancer, ovarian cancer, and autism. Our data suggest a causal relationship between zinc deficiency and the overproduction of Hh ligand.