Citations

Mechanical ventilation is necessary to support patients with acute lung injury, but also exacerbates injury through mechanical stress–activated signaling pathways. We show that stretch applied to cultured human cells, and to mouse lungs in vivo, induces robust expression of metallothionein, a potent antioxidant and cytoprotective molecule critical for cellular zinc homeostasis. Furthermore, genetic deficiency of murine metallothionein genes exacerbated lung injury caused by high tidal volume mechanical ventilation, identifying an adaptive role for these genes in limiting lung injury. Stretch induction of metallothionein required zinc and the zinc-binding transcription factor MTF1. We further show that mouse dietary zinc deficiency potentiates ventilator-induced lung injury, and that plasma zinc levels are significantly reduced in human patients who go on to develop acute respiratory distress syndrome (ARDS) compared with healthy and non-ARDS intensive care unit (ICU) controls, as well as with other ICU patients without ARDS. Taken together, our findings identify a potentially novel adaptive response of the lung to stretch and a critical role for zinc in defining the lung’s tolerance for mechanical ventilation. These results demonstrate that failure of stretch-adaptive responses play an important role in exacerbating mechanical ventilator–induced lung injury, and identify zinc and metallothionein as targets for lung-protective interventions in patients requiring mechanical ventilation.

Background: Adjacent segment pathology is a known complication after spinal fusion, but little has been reported on junctional failure. A series of adolescent patients presented with acute distal junctional failure (DJF). We sought to determine any common features of these patients to develop a prevention strategy. Methods: A retrospective review was conducted of pediatric patients who developed DJF after instrumented spinal fusion performed at 2 institutions from 1999 to 2013. Patients with proximal junctional failure or junctional kyphosis without failure were excluded. Results: Fifteen subjects were identified with mean follow-up of 38 months. Distal failure occurred a mean of 60 days after index surgery, with history of minor trauma in 4 patients. Failures included 3-column Chance fracture (11) or instrumentation failure (4). Thirteen patients presented with back pain and/or acute kyphosis, whereas 2 asymptomatic patients presented with healed fractures. Two patients also developed new onset of severe lower extremity neurological deficit after fracture, which improved but never resolved after revision. A total of 13/15 subjects required revision surgery, typically within 1 week. Complications associated with revision surgery were encountered in 8 patients (62%). Major complications that required return to the operating room included 2 deep infections, 2 instrumentation failures, and dense lower extremity paralysis that improved after medial screw revision and decompression. At final follow-up, 10 patients are asymptomatic, 2 have persistent neurological deficit, 2 have chronic pain, and 1 has altered gait with gait aid requirement. Conclusions: This study analyzes a heterogenous cohort of spinal fusion patients who developed DJF from 3-column Chance fracture or instrumentation failure. Revision surgery is typically required, but has a high complication rate and can result in severe neurological deficit, highlighting the morbidity of this complication. It is unclear whether level of the lowest instrumented vertebra contributes to DJF. Increased awareness of junctional failure in children may prompt additional studies to further characterize risk factors and preventative strategies. Level of Evidence: Level IV—study-type case series.

NAD + can influence DNA repair by modulating protein interactions.

83 Background: Although surgery is traditionally the standard of care for esophageal cancer, esophagectomy carries significant morbidity and mortality. Alternative endoscopic therapies are needed for patients who are not candidates for conventional treatment. The objective of this study was to assess the safety, efficacy, and tolerability of spray cryotherapy of esophageal cancer. Methods: This study includes patients enrolled retrospectively and prospectively in an open-label registry and patients in a retrospective cohort from twelve academic and community practices. Endoscopic spray cryotherapy was performed until local tumor eradication was confirmed by biopsy or until treatment was halted due to progression of disease, patient withdrawal or co-morbidities. Results: One-hundred and eight patients (median age 75.5, 79.6% male, 93.5% adenocarcinoma, mean length 5.2 cm) underwent 442 treatments (mean 4.2 per patient). Tumor stages included 40 with T1a, 27 with T1b, 10 with unspecified T1, 15 with T2, and 16 with no T stage reported. One-hundred and six patients completed treatment with complete response of intraluminal disease in 54.7%, including complete response in 74.4% for T1a, 50% for T1b, 65.3% for all T1, 6.7% for T2, and 50% for those with no T stage reported. Mean follow-up was 17.3 months. There were no deaths or perforations related to spray cryotherapy. Strictures developed in 11 of 108 patients (10.2%) but were present before spray cryotherapy in 3 of 11. Conclusions: This study suggests that endoscopic spray cryotherapy is safe, well tolerated and effective for early esophageal cancer in patients who are not candidates for conventional therapy.

SummaryWhile proteasome inhibition is a validated therapeutic approach for multiple myeloma (MM), inhibition of individual constitutive proteasome (c20S) and immunoproteasome (i20S) subunits has not been fully explored owing to a lack of effective tools. We utilized the novel proteasome constitutive/immunoproteasome subunit enzyme‐linked immunosorbent (ProCISE) assay to quantify proteasome subunit occupancy in samples from five phase I/II and II trials before and after treatment with the proteasome inhibitor carfilzomib. Following the first carfilzomib dose (15–56 mg/m2), dose‐dependent inhibition of c20S and i20S chymotrypsin‐like active sites was observed [whole blood: ≥67%; peripheral blood mononuclear cells (PBMCs): ≥75%]. A similar inhibition profile was observed in bone marrow–derived CD138+ tumour cells. Carfilzomib‐induced proteasome inhibition was durable, with minimal recovery in PBMCs after 24 h but near‐complete recovery between cycles. Importantly, the ProCISE assay can be used to quantify occupancy of individual c20S and i20S subunits. We observed a relationship between MM patient response (n = 29), carfilzomib dose and occupancy of multiple i20S subunits, where greater occupancy was associated with an increased likelihood of achieving a clinical response at higher doses. ProCISE represents a new tool for measuring proteasome inhibitor activity in clinical trials and relating drug action to patient outcomes.

Transcription activator-like effectors (TALEs) are extremely effective, single-molecule DNA-targeting molecular cursors used for locus-specific genome science applications, including high-precision molecular medicine and other genome engineering applications. TALEs are used in genome engineering for locus-specific DNA editing and imaging, as artificial transcriptional activators and repressors, and for targeted epigenetic modification. TALEs as nucleases (TALENs) are effective editing tools and offer high binding specificity and fewer sequence constraints toward the targeted genome than other custom nuclease systems. One bottleneck of broader TALE use is reagent accessibility. For example, one commonly deployed method uses a multitube, 5-day assembly protocol. Here we describe FusX, a streamlined Golden Gate TALE assembly system that (1) is backward compatible with popular TALE backbones, (2) is functionalized as a single-tube 3-day TALE assembly process, (3) requires only commonly used basic molecular biology reagents, and (4) is cost-effective. More than 100 TALEN pairs have been successfully assembled using FusX, and 27 pairs were quantitatively tested in zebrafish, with each showing high somatic and germline activity. Furthermore, this assembly system is flexible and is compatible with standard molecular biology laboratory tools, but can be scaled with automated laboratory support. To demonstrate, we use a highly accessible and commercially available liquid-handling robot to rapidly and accurately assemble TALEs using the FusX TALE toolkit. Together, the FusX system accelerates TALE-based genomic science applications from basic science screening work for functional genomics testing and molecular medicine applications.

Abstract Pancreatic cancer is projected to become the second leading cause of cancer-related death in the United States by 2020. A familial aggregation of pancreatic cancer has been established, but the cause of this aggregation in most families is unknown. To determine the genetic basis of susceptibility in these families, we sequenced the germline genomes of 638 patients with familial pancreatic cancer and the tumor exomes of 39 familial pancreatic adenocarcinomas. Our analyses support the role of previously identified familial pancreatic cancer susceptibility genes such as BRCA2, CDKN2A, and ATM, and identify novel candidate genes harboring rare, deleterious germline variants for further characterization. We also show how somatic point mutations that occur during hematopoiesis can affect the interpretation of genome-wide studies of hereditary traits. Our observations have important implications for the etiology of pancreatic cancer and for the identification of susceptibility genes in other common cancer types. Significance: The genetic basis of disease susceptibility in the majority of patients with familial pancreatic cancer is unknown. We whole genome sequenced 638 patients with familial pancreatic cancer and demonstrate that the genetic underpinning of inherited pancreatic cancer is highly heterogeneous. This has significant implications for the management of patients with familial pancreatic cancer. Cancer Discov; 6(2); 166–75. ©2015 AACR. This article is highlighted in the In This Issue feature, p. 109

Background: Intraoperative computed tomography and image-guided navigation improve the accuracy of screw placement. Radiation exposure to the patient remains a primary drawback. The objective of the present study was to compare the total intraoperative radiation dose and assess the resultant image quality for O-arm-assisted pedicle screw insertion, among 3 protocols: default (manufacturer recommended), institutional (reduced dose utilized in our institution), and pediatric (new protocol with lowest dose). Methods: Thirty-seven consecutive patients under the age of 18 years underwent posterior instrumentation of the spine and underwent an intraoperative O-arm scan. Techniques (kV and mAs) for default and institutional dose settings were manually adjusted based on spinal level and body weight. Pediatric dose techniques were 80 kV/80 mAs with no adjustment for level or weight. The number of scans repeated because of inadequate imaging was assessed, and the mean estimated effective dose between the 3 protocols was compared. Results: Sixty-eight scans were performed in 37 consecutive patients with mean age of 14 years and mean weight of 55 kg. For reference, the effective radiation dose of a chest x-ray is approximately 0.10 mSv. Use of the default protocol resulted in higher mean effective dose per scan of 4.65 mSv, whereas institutional protocol resulted in 2.37 mSv. The pediatric protocol reduced the mean dose to 0.65 mSv. The total effective dose per surgery was: 1.17 mSv (pediatric), 3.83 mSv (institutional), and 12.79 mSv (default) (P<0.0001 each). All scans lead to satisfactory image quality except in 1 patient >100 kg with stainless steel implants. There were no neurological or other implant-related complications. The pediatric protocol resulted in satisfactory image quality with the lowest total radiation dose, only 1/10 of that of the default protocol. Conclusions: We successfully switched to a pediatric low-dose O-arm protocol in clinical practice, reducing the dose to <1/4 of the mean annual natural background radiation. This may allow use of intraoperative computed tomography and navigation for pedicle screw placement without excessive radiation exposure to young patients. Level of Evidence: Level III—retrospective comparative study.