Unbiased cell surface proteomics identifies SEMA4A as an effective immunotherapy target for myeloma.

The accessibility of cell surface proteins makes them tractable for targeting by cancer immunotherapy, but identifying suitable targets remains challenging. Here we describe plasma membrane profiling of primary human myeloma cells to identify an unprecedented number of cell surface proteins of a primary cancer. We used a novel approach to prioritize immunotherapy targets and identified a cell surface protein not previously implicated in myeloma, semaphorin-4A (SEMA4A). Using knock-down by short-hairpin RNA and CRISPR/nuclease-dead Cas9 (dCas9), we show that expression of SEMA4A is essential for normal myeloma cell growth in vitro, indicating that myeloma cells cannot downregulate the protein to avoid detection. We further show that SEMA4A would not be identified as a myeloma therapeutic target by standard CRISPR/Cas9 knockout screens because of exon skipping. Finally, we potently and selectively targeted SEMA4A with a novel antibody-drug conjugate in vitro and in vivo.

RYK-mediated filopodial pathfinding facilitates midgut elongation.

Between embryonic days 10.5 and 14.5, active proliferation drives rapid elongation of the murine midgut epithelial tube. Within this pseudostratified epithelium, nuclei synthesize DNA near the basal surface and move apically to divide. After mitosis, the majority of daughter cells extend a long, basally oriented filopodial protrusion, building a de novo path along which their nuclei can return to the basal side. WNT5A, which is secreted by surrounding mesenchymal cells, acts as a guidance cue to orchestrate this epithelial pathfinding behavior, but how this signal is received by epithelial cells is unknown. Here, we have investigated two known WNT5A receptors: ROR2 and RYK. We found that epithelial ROR2 is dispensable for midgut elongation. However, loss of Ryk phenocopies the Wnt5a-/- phenotype, perturbing post-mitotic pathfinding and leading to apoptosis. These studies reveal that the ligand-receptor pair WNT5A-RYK acts as a navigation system to instruct filopodial pathfinding, a process that is crucial for continuous cell cycling to fuel rapid midgut elongation.

Non-target profiling of bitumen-influenced waters for the identification of tracers unique to oil sands processed-affected water (OSPW) in the Athabasca watershed of Alberta, Canada.

RATIONALE: The objective of this study was to identify unique chemical tracers of oil sands process-affected water (OSPW) to enable definitive discrimination of tailings pond seepage from natural bitumen-influenced waters from the Canadian Alberta McMurray formation. METHODS: The approach involved comparing unknowns from an unprecedented sample set of OSPW (n = 4) and OSPW-affected groundwaters (n = 15) with natural bitumen-influenced groundwaters (n = 20), using high-performance liquid chromatography/electrospray ionisation high-resolution mass spectrometry (HPLC/ESI-HRMS) operated in both polarities. RESULTS: Four unknown chemical entities were identified as potential tracers of OSPW seepage and subsequently subjected to structural elucidation. One potential tracer, tentatively identified as a thiophene-containing carboxylic acid [C15 H23 O3 S]- , was only detected in OSPW and OSPW-affected samples, thereby showing the greatest diagnostic potential. The remaining three unknowns, postulated to be two thiochroman isomers [C17 H25 O3 S]+ and an ethyl-naphthalene isomer [C16 H21 ]+ , were detected in one and two background groundwaters, respectively. CONCLUSIONS: We advanced the state of knowledge for tracers of tailings seepage beyond heteroatomic classes, to identifying diagnostic substances, with structures postulated. Synthesis of the four proposed structures is recommended to enable structural confirmations. This research will guide and inform the Oil Sands Monitoring Program in its efforts to assess potential influences of oil sands development on the Athabasca River watershed.

Advances in Distinguishing Groundwater Influenced by Oil Sands Process-Affected Water (OSPW) from Natural Bitumen-Influenced Groundwaters.

The objective of this study was to advance analytical methods for detecting oil sands process-affected water (OSPW) seepage from mining containments and discriminating any such seepage from the natural bitumen background in groundwaters influenced by the Alberta McMurray formation. Improved sampling methods and quantitative analyses of two groups of monoaromatic acids were employed to analyze OSPW and bitumen-affected natural background groundwaters for source discrimination. Both groups of monoaromatic acids showed significant enrichment in OSPW, while ratios of O2/O4 containing heteroatomic ion classes of acid extractable organics (AEOs) did not exhibit diagnostic differences. Evaluating the monoaromatic acids to track a known plume of OSPW-affected groundwater confirmed their diagnostic abilities. A secondary objective was to assess anthropogenically derived artificial sweeteners and per- and polyfluoroalkyl substances (PFAS) as potential tracers for OSPW. Despite the discovery of acesulfame and PFAS in most OSPW samples, trace levels in groundwaters influenced by general anthropogenic activities preclude them as individual robust tracers. However, their inclusion with the other metrics employed in this study served to augment the tiered, weight of evidence methodology developed. This methodology was then used to confirm earlier findings of OSPW migrations into groundwater reaching the Athabasca River system adjacent to the reclaimed pond at Tar Island Dyke.

Use of mature willows (Salix nigra) for hydraulic control of landfill-impacted groundwater in a temperate climate.

Phreatophytic trees such as willows and poplars have a large capacity for extracting shallow groundwater, as evidenced by diurnal water table fluctuations corresponding to intensified transpiration during the day. As a result, they have been employed for phytoremediation of shallow contaminated groundwater. In this study, the water extraction (i.e. pumping) capacity of mature willows (Salix nigra) to capture shallow polluted groundwater in Belle Park, the site of a former landfill in Kingston, Ontario (Canada), was assessed using continuous field measurements of sap flow and water table levels associated with a single willow tree, combined with a transient numerical model (FEFLOW). On an annual basis, the sapflow averaged 2.3 m3d-1, with 70% of the cumulative sap flow occurring during the active growing season (May to September). The calibration showed a good fit (0.91 < R2<0.97) between measured groundwater levels from three shallow wells installed near the willow and the calculated water table level fluctuations, thus confirming that the water extraction rate based on sap flow data for the willow was appropriate. At stand level, additional modelling suggests that 3.4-4.7 ha of mature willows (i.e. between 7.8% and 10.6% of the Park area), could compensate for the current water volumes extracted by the municipality using a conventional pump and treat system. The results of this study indicate that willows can play a significant role in capturing contaminated groundwater underlying Belle Park, and potentially at other sites where removal of contaminants from shallow aquifers is desired.

Endobenthic Organisms Exposed to Chronically High Chloride from Groundwater Discharging along Freshwater Urban Streams and Lakeshores.

Chloride, especially from road salt, is one of the most ubiquitous contaminants of urban groundwater in temperate climates. The discharge of chloride-laden groundwater to freshwater ecosystems may pose a heightened risk to endobenthic organisms (buried in sediments), which may experience high concentrations prior to dilution from the overlying water. However, available exposure data is limited. Presented here are 22 chloride data sets from 15 urban sites across Canada, encompassing >1300 samples of shallow discharging groundwater collected principally during summer through autumn. Over half of the sites had an average chloride concentration above the long-term aquatic life guideline (120 mg/L), while 14% of each site's samples, on average, surpassed the short-term guideline (640 mg/L). Chloride concentrations frequently varied substantially (even >1000 mg/L) between adjacent locations (mostly <20 m separation), indicating patchy exposure. Chloride/bromide ratios, artificial sweeteners, and other tracers suggest a predominant contribution from road salt, with wastewater and landfill leachate important at some sites. Overall, these concentrations exceed those typically reported for urban wells and streams (even during the snowmelt period) in similar climates. These findings suggest that high chloride concentrations in shallow groundwater, largely from road salt, present a long-term threat to endobenthic organisms of urban surface waters in cold-region countries.

Endoscopic technique for closure of enterocutaneous fistulas.

BACKGROUND AND OBJECTIVES: Endoscopic over the scope clip (OTSC) closure represents a new technique for endoscopic management of enteric bleeding and tissue defects such as anastomotic leaks and enterocutaneous (EC) fistulas. We aim to describe our technical approach for OTSC closure of EC fistulas and convey our outcomes. METHODS AND PROCEDURES: This retrospective review includes ten patients who underwent OTSC application for EC fistulas by surgical endoscopists at a US tertiary care hospital from July 2015 to October 2017. Demographic data, along with type of defect, location, duration of lesion, success or failure of OTSC, and nutritional status were compiled. The ACS NSQIP surgical risk calculator was used to project the risk of mortality, complications, length of stay, and risk of readmission had our patients undergone surgical correction of their fistula. RESULTS: Overall success for EC fistula closure was 70%. Acute fistulas were closed with a success rate of 86%. Chronic fistulas were closed successfully in only 33% of cases. Of patients successfully closed, NSQIP-predicted rates of mortality, any complication, and median length of stay were 21.1%, 34.5%, and 9.5 days, respectively. With OTSC, these patients experienced 0 mortalities, 0 complications, and had a median length of stay of 4 days. CONCLUSION: OTSC is an effective adjunctive measure to improving rates of successful closure of EC fistulas and compromised anastomosis. OTSC conveys a markedly improved procedural risk profile as compared to standard surgical correction.

Transcatheter Valve-in-Valve Therapy Using a Balloon Expanding Valve for Treatment of Aortic Paravalvular Leakage.

Aortic paravalvular leakage (PVL) is a recognised complication of surgically replaced valves which is often treated using vascular plugs. Whilst transcatheter valve-in-valve therapy has been increasingly used for failed surgical bioprostheses, it is not considered as a treatment option for aortic PVL. However, the newer design of transcatheter aortic valves has a fabric skirt to create a more effective seal around the annulus. To our best knowledge, for the first time, we report successful adoption of the valve-in-valve therapy for the treatment of PVL in surgical bioprosthetic aortic valves such that the fabric skirt is placed immediately below the regurgitant orifice resulting in significant reduction in the PVL.

The biochemistry, signalling and disease relevance of RYK and other WNT-binding receptor tyrosine kinases.

The receptor tyrosine kinases (RTKs) are a well-characterized family of growth factor receptors that have central roles in human disease and are frequently therapeutically targeted. The RYK, ROR, PTK7 and MuSK subfamilies make up an understudied subset of WNT-binding RTKs. Numerous developmental, stem cell and pathological roles of WNTs, in particular WNT5A, involve signalling via these WNT receptors. The WNT-binding RTKs have highly context-dependent signalling outputs and stimulate the ß-catenin-dependent, planar cell polarity and/or WNT/Ca2+ pathways. RYK, ROR and PTK7 members have a pseudokinase domain in their intracellular regions. Alternative signalling mechanisms, including proteolytic cleavage and protein scaffolding functions, have been identified for these receptors. This review explores the structure, signalling, physiological and pathological roles of RYK, with particular attention paid to cancer and the possibility of therapeutically targeting RYK. The other WNT-binding RTKs are compared with RYK throughout to highlight the similarities and differences within this subset of WNT receptors.

Deficiency of the Wnt receptor Ryk causes multiple cardiac and outflow tract defects.

Ryk is a member of the receptor tyrosine kinase (RTK) family of proteins that control and regulate cellular processes. It is distinguished by binding Wnt ligands and having no detectable intrinsic protein tyrosine kinase activity suggesting Ryk is a pseudokinase. Here, we show an essential role for Ryk in directing morphogenetic events required for normal cardiac development through the examination of Ryk-deficient mice. We employed vascular corrosion casting, vascular perfusion with contrast dye, and immunohistochemistry to characterize cardiovascular and pharyngeal defects in Ryk-/- embryos. Ryk-/- mice exhibit a variety of malformations of the heart and outflow tract that resemble human congenital heart defects. This included stenosis and interruption of the aortic arch, ventriculoarterial malalignment, ventricular septal defects and abnormal pharyngeal arch artery remodelling. This study therefore defines a key intersection between a subset of growth factor receptors involved in planar cell polarity signalling, the Wnt family and mammalian cardiovascular development.

The "side-BASE technique": Combined side branch anchor balloon and balloon assisted sub-intimal entry to resolve ambiguous proximal cap chronic total occlusions.

Failure to penetrate the proximal cap or cross with equipment remains the most frequent cause of procedural failure in CTO PCI. In the "BASE" technique concerns were raised over the risk of proximal side branch loss. We here describe the evolution of this technique and highlight examples where the sub-intimal space was accessed proximal to the occlusion by using the side branch. This technique can be of use both in resolving proximal cap ambiguity and also in cases where there is difficulty crossing a resistant proximal cap.

Subadventitial stenting around occluded stents: A bailout technique to recanalize in-stent chronic total occlusions.

OBJECTIVES: To evaluate the outcomes of subadventitial stenting (SS) around occluded stents for recanalizing in-stent chronic total occlusions (IS-CTOs). BACKGROUND: There is little evidence on the outcomes of SS for IS-CTO. METHODS: We examined the outcomes of SS for IS-CTO PCI at 14 centers between July 2011 and June 2017, and compared them to historical controls recanalized using within-stent stenting (WSS). Target-vessel failure (TVF) on follow-up was the endpoint of this study, and was defined as a composite of cardiac death, target-vessel myocardial infarction, and target-vessel revascularization. RESULTS: During study period, 422 IS-CTO PCIs were performed, of which 32 (7.6%) were recanalized with SS, usually when conventional approaches failed. The most frequent CTO vessel was the right coronary artery (72%). Mean J-CTO score was 3.1 ± 0.9. SS was antegrade in 53%, and retrograde in 47%. Part of the occluded stent was crushed in 37%, while the whole stent was crushed in 63%. Intravascular imaging was used in 59%. One patient (3.1%) suffered tamponade. Angiographic follow-up was performed in 10/32 patients: stents were patent in six cases, one had mild neointimal hyperplasia, and three had severe restenosis at the SS site. Clinical follow-up was available for 29/32 patients for a mean of 388 ± 303 days. The 24-month incidence of TVF was 13.8%, which was similar to historical controls treated with WSS (19.5%, P = 0.49). CONCLUSIONS: SS is rarely performed, usually as last resort, to recanalize complex IS-CTOs. It is associated with favorable acute and mid-term outcomes, but given the small sample size of our study additional research is warranted.

Bivalirudin Anticoagulation Dosing Protocol for Extracorporeal Membrane Oxygenation: A Retrospective Review.

Anticoagulation with unfractionated heparin during extracorporeal membrane oxygenation (ECMO) is common, but alternative agents are being evaluated for safety and efficacy. The objective of this analysis was to assess if a comprehensive bivalirudin dosing and monitoring protocol effectively guides dose adjustments and monitoring of bivalirudin in patients during ECMO. Our analysis included 11 patients who received bivalirudin during ECMO therapy and had dosing managed using our hospital derived protocol. Patients treated over a 1-year period were included in this retrospective analysis. Clinical characteristics and changes in activated partial thromboplastin time (aPTT) were evaluated from medical records to determine the efficacy of the dosing protocol. ECMO was initiated for acute respiratory distress syndrome in eight (72.7%) patients and for cardiac arrest in three (27.3%) patients. A total of 178 protocol guided dose adjustments were made during the study. Among the dose adjustments, 56 (31.5%) attained the protocol predicted aPTT level change, 96 (53.9%) of the measured aPTT changes were less than predicted, and 26 (14.6%) of the measured aPTT changes were more than predicted. On average, patients were within their defined therapeutic aPTT target range 66.3% of the time. All patients reached their designated aPTT target range within the first 24 hours of therapy. Significant bleeding was documented in eight (72.7%) patients. No clinically evident thromboembolic events were identified in vivo while cannulated. This analysis suggests that bivalirudin can be managed using a dosing protocol to provide anticoagulation therapy to patients during ECMO and can provide foundational guidance for dose adjustment and monitoring for other institutions.

Simultaneous Transseptal Para-Ring Leak Closure and Transcatheter Mitral Valve Implantation for the Treatment of Surgical Mitral Repair Failure.

Repeat cardiac surgery in patients with a previous sternotomy is associated with significant morbidity and mortality. While transcatheter aortic valve implantation in high risk surgical patients is now well established, experience with transcatheter mitral valve replacement is still at an early stage. Although many successful reports of transcatheter mitral valve replacements now exist, the predominant approach has been via a transapical approach. It is likely that, as with the evolution of favoured access routes for transcatheter aortic therapies, future directions for transcatheter mitral valves will focus on smaller delivery systems favouring the transvenous transseptal approach where possible. We present the first reported case of combined transseptal para-ring leak closure followed by transcatheter mitral valve implantation using a 12/5mm Amplatzer III vascular plug and a 29mm SAPIEN 3 valve.

Phaeochromocytoma presenting with labile blood pressures following coronary artery bypass grafting.

Pheochromocytomas have been reported prior to and during coronary artery bypass surgery. We present a patient with an undiagnosed pheochromocytoma who presented with labile hypertension following coronary artery bypass surgery. This case calls attention to the inclusion of an undiagnosed pheochromocytoma in the differential diagnosis for all patients who develop labile hypertension in the postoperative period following cardiac surgery.

Aortic valve laceration following coronary angiography and percutaneous coronary intervention.

Valve complications following coronary angiography and percutaneous coronary interventions are rare. We report a case of an aortic valve laceration following cardiac catheterization and percutaneous coronary intervention, which required surgical valve replacement.

Elevated dissolved phosphorus in riparian groundwater along gaining urban streams.

Findings of low concentrations of dissolved phosphorus in groundwater in large surveys [e.g., United States Geological Survey's National Water-Quality Assessment (NAWQA) Program ( Dubrovsky, N. M.; et al. The Quality of Our Nation's Water: Nutrients in the Nation's Streams and Groundwater, 1992-2004 . U.S. Geological Survey Circular 1350; USGS : Reston, VA , 2010 . ); >5000 wells] support the common perception that groundwater is generally of little importance for transporting phosphorus. Here, we address whether this applies to urban riparian settings, where discharging groundwater may potentially contribute to urban stream syndrome and downstream eutrophication problems. This survey study includes 665 samples of groundwater collected along gaining stream reaches at six urban sites. Considering the combined sample set, 27% had soluble reactive phosphorus (SRP) concentrations >0.1 mg L(-1), which is more than double that determined in the NAWQA Program (12%), while for individual sites the range was 12-52%, excluding one site with consistently low SRP (0%). None of the sites showed significant correlation between SRP and the artificial sweetener acesulfame, a promising wastewater indicator, including two with known wastewater contamination (but the lowest SRP). Rather, high SRP concentrations were associated with geochemically reducing conditions. This could mean that natural aquifer or stream sediment materials were a primary contributor of the elevated SRP observed in this study.

Profiling oil sands mixtures from industrial developments and natural groundwaters for source identification.

The objective of this study was to identify chemical components that could distinguish chemical mixtures in oil sands process-affected water (OSPW) that had potentially migrated to groundwater in the oil sands development area of northern Alberta, Canada. In the first part of the study, OSPW samples from two different tailings ponds and a broad range of natural groundwater samples were assessed with historically employed techniques as Level-1 analyses, including geochemistry, total concentrations of naphthenic acids (NAs) and synchronous fluorescence spectroscopy (SFS). While these analyses did not allow for reliable source differentiation, they did identify samples containing significant concentrations of oil sands acid-extractable organics (AEOs). In applying Level-2 profiling analyses using electrospray ionization high resolution mass spectrometry (ESI-HRMS) and comprehensive multidimensional gas chromatography time-of-flight mass spectrometry (GC × GC-TOF/MS) to samples containing appreciable AEO concentrations, differentiation of natural from OSPW sources was apparent through measurements of O2:O4 ion class ratios (ESI-HRMS) and diagnostic ions for two families of suspected monoaromatic acids (GC × GC-TOF/MS). The resemblance between the AEO profiles from OSPW and from 6 groundwater samples adjacent to two tailings ponds implies a common source, supporting the use of these complimentary analyses for source identification. These samples included two of upward flowing groundwater collected <1 m beneath the Athabasca River, suggesting OSPW-affected groundwater is reaching the river system.

Spatiotemporal investigation of geochemical and hydrological controls on release of soluble reactive phosphorus from the shallow aquifer of a riparian zone.

Phosphorus (P) that accumulates in agricultural riparian zones can be released under certain hydrological and biogeochemical conditions, thereby limiting the effectiveness of these zones in reducing P loads from field to stream. The study objective was to explore factors that may be contributing to, or limiting, high soluble reactive phosphorus (SRP) concentrations in the shallow aquifer of an alluvial upland riparian zone located in a continental climate. Field investigations including porewater sampling from six vertical nests, soil sampling, and continuous soil moisture, groundwater table, and redox measurements were conducted over 19 months. Porewater SRP concentrations were generally low in the aquifer considering all sampling times (median = 14.7 µg/L; interquartile range [IQR] = 11.1 µg/L, 287 samples). The overall low SRP may be due to low reducible labile soil P (median = 21.1 µgP/g dw, IQR = 10.9 µgP/g dw, 21 samples). However, high SRP concentrations (>52 µg/L, 95% quartile) did occur intermittently in space and time with no clear spatial or temporal patterns. Analyses indicate that most high concentrations were likely not associated with factors previously reported to influence SRP release in riparian aquifers, including redox conditions, pH, and soil drying and wetting. Further, data indicate that internally released or externally supplied SRP may undergo rapid (re-) sequestration within the aquifer, limiting its vertical or horizontal transport. The study findings highlight the complexity of P behavior in riparian zones and the need for caution when assessing the effectiveness of conservation practices and in interpreting potential impacts of subsurface water quality on stream water quality when monitoring locations are distant from the stream.

Characterization of subsurface pathways contributing to freshwater salinization of urban streams using electrical and electromagnetic imaging techniques.

Salinization of inland fresh surface waters in temperate climates is a growing concern due to increasing salt inputs from sources including chloride (Cl)-containing road salt de-icers, industrial waste, and landfill leachate. Groundwater pathways play an important role in the year-round delivery of Cl to streams, but quantifying this pathway, including spatiotemporal variability and amount of Cl mass stored in the subsurface, is challenging. The objective of this study was to demonstrate, evaluate, and compare the potential applications of the geoelectrical techniques - electromagnetics (EM) and direct current (DC) resistivity - for mapping salt contamination in shallow urban groundwater and characterizing the groundwater pathways delivering Cl to urban streams. EM and DC surveys were conducted (3D mapping and 2D time-lapse) across a 20 m salt-impacted stream section and surrounding riparian zone that is located near an arterial road and parking lot. Groundwater samples and soil cores were also collected to validate the geoelectrical results. Both the EM and DC surveys detected high salt concentrations in the shallow subsurface (up to 3 m depth) near the road, parking lot, and stream; however, DC more accurately represented groundwater Cl concentrations. DC results were used to calculate the total Cl mass in the subsurface, with the spatial mass distribution used to infer the temporal variability in the subsurface salt plume. Finally, time-lapse DC showed that the highest groundwater salt concentrations existed near the stream between June and October - this is expected to contribute to the elevated salt concentrations in the stream during summer months. This study has shown that EM and DC can be useful for identifying groundwater salt concentration, storage, and transport in a non-intrusive and efficient manner, making them valuable field tools for characterizing and quantifying groundwater salt pathways to urban streams.