Technical Step Research Articles

Abstract A190: Management of germline findings revealed throughout the course of tumor-normal whole genome sequencing in oncology

Abstract Background: The Personalized OncoGenomics (POG) project at the BC Cancer Agency utilizes tumor-normal whole genome sequencing (WGS) to understand key driver pathways and guide personalized treatment decisions. Analysis of the germline data can reveal variants; these may be presumed pathogenic, presumed benign, or of unknown significance (VUS). We have developed a process for evaluating and returning presumed pathogenic variants in known cancer susceptibility genes to patients, for counseling and validation in a clinical-accredited laboratory. Methods: Patients receive germline cancer-related information as part of the consent process for participation in the POG program. A subcommittee comprising medical geneticists, bioinformaticians, pathologists, oncologists, and an ethicist review the germline results. Any variants suspicious of being an artifact undergo a technical validation step. Presumed pathogenic findings of known cancer susceptibility genes are returned to the patient by their treating oncologist and patients are referred to the Hereditary Cancer Program (HCP), for genetic counseling and clinical confirmation. Results: From June 2012-January 2017, 466 patients have consented to the project. To date, 39 cases (8.4%) had at least one variant that was deemed pathogenic, and 86 cases had at least one VUS in a known cancer susceptibility gene. 11 out of 23 cases (47.8%) with high-penetrance mutations were already known to HCP. All VUS were reviewed by the subcommittee, taking into consideration the VUS and clinical context. 8 of the subjects with pathogenic results and 3 with VUS were known to HCP before POG data were generated. A VUS in 7 cases (1.5%) was returned after review. Conclusions: The number of pathogenic variants in known cancer susceptibility genes is consistent with published oncology results. We created a process to manage clinically relevant germline findings discovered during the course of genomic research to ensure appropriate care for patients. Genetic counseling within HCP and validation of variants in the clinically accredited Cancer Genetics Laboratory enables seamless return of research-generated clinically relevant germline results to affected subjects. Citation Format: Howard J. Lim, Kasmintan A. Schrader, Sean Young, Jessica M T Nelson, Alexandra Fok, Erin Pleasance, Martin Jones, Yaoqing Shen, Linlea Armstrong, Alice Virani, Shahrad Rassekh, Rebecca Deyell, Stephen Yip, Robyn Roscoe, Aly Karsan, Marco Marra, Janessa J. Laskin. Management of germline findings revealed throughout the course of tumor-normal whole genome sequencing in oncology [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr A190.

Neutron Imaging Facilities in a Global Context

Neutron Imaging (NI) has been developed in the last decades from a film-based inspection method for non-destructive observations towards a powerful research tool with many new and competitive methods. The most important technical step forward has been the introduction and optimization of digital imaging detection systems. In this way, direct quantification of the transmission process became possible—the basis for all advanced methods like tomography, phase-contrast imaging and neutron microscopy. Neutron imaging facilities need to be installed at powerful neutron sources (reactors, spallation sources, other accelerator driven systems). High neutron intensity can be used best for either highest spatial, temporal resolution or best image quality. Since the number of such strong sources is decreasing world-wide due to the age of the reactors, the number of NI facilities is limited. There are a few installations with pioneering new concepts and versatile options on the one hand, but also relatively new sources with only limited performance thus far. It will be a challenge to couple the two parts of the community with the aim to install state-of-the-art equipment at the suitable beam ports and develop NI further towards a general research tool. In addition, sources with lower intensity should be equipped with modern installations in order to perform practical work best.

The impact of antegrade intramedullary nailing start site using the SIGN nail in proximal femoral fractures: A prospective cohort study

IntroductionIn many low and middle-income countries (LMICs) SIGN nail is commonly used for antegrade femoral intramedullary (IM) nailing, using a start site either at the tip of the greater trochanter or piriformis fossa. While a correct start site is considered an essential technical step; few studies have evaluated the impact of using an erroneous start site. This is particularly relevant in settings with limited access to fluoroscopy to aid in creating a nail entry point. The purpose of this study was to evaluate the impact of antegrade SIGN IM nailing start site on radiographic alignment and health-related quality of life. MethodsIn this prospective cohort study, adult patients with proximal femur fractures (OTA 32, subtrochanteric zone) treated with antegrade IM SIGN nail at Muhimbili Orthopaedic Institute (MOI), Dar es Salaam, Tanzania were enrolled. Start site was determined on the immediate postoperative X-ray and was graded on a continuous scale based on distance of the IM nail center from the greater trochanteric tip. The primary outcome measurement was coronal alignment on the post-operative x-ray. The secondary outcomes were reoperation rates, RUST scores and EQ5D scores at one year follow-up. ResultsSeventy-nine patients were enrolled. 50 of them (63.3%) had complete data at 1year and were included in the final data analysis. Of the fifty patients, nine (18%) had IM nails placed laterally, 26 (52%) medially and 15 (30%) directly over the tip of the greater trochanter. Compared to a start site at the tip or medial to the greater trochanter, a lateral start site was 9 times more likely to result in a varus malalignment (95% CI: 1.42–57.70, p=0.021). ConclusionsLateral start site was associated with varus malalignment. Although lateral start site was not significantly associated with reoperation, varus deformity was associated with higher reoperation rates. Surgeons should consider avoiding a start site lateral to the tip of the greater trochanter or allow the nail to rotate to avoid malalignment when using the SIGN nail for proximal femur fractures.

Erratum: A Simple Technique to Prevent Early or Late EndoButton Deployment in Anterior Cruciate Ligament Reconstructions: A Technical Note.

The use of suspensory graft fixation methods in arthroscopic reconstruction of the anterior cruciate ligament has become increasingly popular with the more frequent use of tendon grafts and anatomical techniques involving the creation of bone tunnels. An important technical step to ensure adequate fixation of the graft when using EndoButton-type implants, particularly in the femur, involves flipping the EndoButton plate at the correct length to avoid performing this maneuver before the appropriate time or leaving soft tissue between the plate and lateral cortex of the femur. In the present study, we describe a simple arthroscopic technique for indicating the correct time to flip/deploy the EndoButton plate.

Lyapunov stability analysis of a linear system coupled to a heat equation.

This paper addresses the stability analysis of a system of ordinary differential equations coupled with a classic heat equation using a Lyapunov approach. Relying on recent developments in the area of time-delay systems, a new method to study the stability of such a class of coupled finite/infinite dimensional systems is presented here. It consists in a Lyapunov analysis of the infinite dimensional state of the system using an energy functional enriched by the mean value of the heat variable. The main technical step relies on the use an efficient Bessel-like integral inequality on Hilbert space leading to tractable conditions expressed in terms of linear matrix inequalities. The results are then illustrated on academic examples and demonstrate the potential of this new approach.

Almost algebraic actions of algebraic groups and applications to algebraic representations

Let G be an algebraic group over a complete separable valued field k . We discuss the dynamics of the G -action on spaces of probability measures on algebraic G -varieties. We show that the stabilizers of measures are almost algebraic and the orbits are separated by open invariant sets. We discuss various applications, including existence results for algebraic representations of amenable ergodic actions. The latter provides an essential technical step in the recent generalization of Margulis–Zimmer super-rigidity phenomenon [2].

A technical step forward in the integration of visible-induced luminescence imaging methods for the study of ancient polychromy

Photo-induced luminescence imaging techniques, such as UV-induced visible luminescence (UVL) and the more recently developed technique of visible-induced infrared luminescence (VIL), have been invaluable for the study of ancient polychromy, allowing the detection and mapping of luminescent materials, such as varnishes, consolidants, organic binders, and crucially, traces of pigments, organic and inorganic, that are often not visible to the naked eye. In the context of works from the Hellenistic period onwards, the detection of two pigments, Egyptian blue and rose madder lake, has been particularly pivotal in advancing the field. Current conventional methodologies for the digital mapping of these two luminescent pigments rely on the separate application of two techniques (VIL and UVL), each requiring a different illumination source and acquisition set-up. In this study, a novel approach is proposed, combining the use of visible-induced infrared luminescence and visible-induced visible luminescence to locate these two pigments. As the source of illumination in both cases is the same system of LEDs, the set-up has the advantage of requiring only minor filter changes between luminescence modes. The increased portability and safety compared to the use of methodologies that employ UV radiation represent notable advantages of this integrated system. The interchangeability between highly selective excitation sources, also significantly simplifies the experimental set-up and the need to adjust the object or equipment between acquisitions, ensuring better reproducibility of the data acquired and facilitating any post-processing procedures. This results in a user-friendly methodology for both experts and non-specialists alike. Three Hellenistic period terracottas; two from Canosa di Puglia, Italy (270–200 BC) and one from Myrina, Turkey (c. 100 BC), all characterised by large well-preserved areas of decoration in Egyptian blue and red lake, were studied in order to trial the approach. Comparisons were made with the more standard techniques of VIL and UVL, and it was shown that the combined method proposed efficiently detects and maps both of these pigments with analogous results to those obtained by more established methodologies. The observations made from the multispectral images acquired were verified by analysis of small samples of the pigments, using FTIR and Raman spectroscopy and HPLC-DAD analysis.

A GENERALIZATION OF PRINCIPAL BUNDLES WITH A PARABOLIC OR LEVEL STRUCTURE

We define a parameter-dependent notion of stability for principal bundles with a certain local decoration, which generalizes both parabolic and level structures, and construct their coarse moduli space. A necessary technical step is the construction of the moduli space of tuples of vector bundles with a global and a local decoration, which we call locally decorated tumps. We introduce a notion of asymptotic stability for locally decorated tumps and show that stable locally decorated principal bundles can be described as asymptotically stable locally decorated tumps.

Management of germline findings revealed throughout the course of tumor-normal whole genome sequencing in oncology.

e13113 Background: The Personalized OncoGenomics (POG) project at the BC Cancer Agency utilizes tumor-normal whole genome sequencing (WGS) to understand key driver pathways and guide personalized treatment decisions. Analysis of the germline data can reveal variants; these may be presumed pathogenic, presumed benign or of unknown significance (VUS). We have developed a process for evaluating and returning presumed pathogenic variants in known cancer susceptibility genes to patients, for counseling and validation in a clinical-accredited laboratory. Methods: Patients receive germline cancer related information as part of the consent process for participation in the POG program. A sub-committee comprised of medical geneticists, bioinformaticians, pathologists, oncologists and an ethicist review the germline results. Any variants suspicious of being an artifact undergo a technical validation step. Presumed pathogenic findings of known cancer susceptibility genes are returned to the patient by their treating oncologist and patients are referred to the Hereditary Cancer Program (HCP), for genetic counseling and clinical confirmation. Results: From June 2012 - January 2017 – 466 patients have consented to the project. To date, 39 cases (8.4%) had at least one variant that was deemed pathogenic, 86 cases had at least one VUS in a known cancer susceptibility gene. 11 out of 23 cases (47.8%) with high penetrance mutations were already known to HCP. All VUS were reviewed by the sub-committee taking in to consideration the VUS and clinical context. 8 of the subjects with pathogenic results and 3 with VUS were known to HCP before POG data was generated. A VUS in 7 cases (1.5%) was returned after review. Conclusions: The number of pathogenic variants in known cancer susceptibility genes is consistent with published oncology results. We created a process to manage clinically relevant germline findings discovered during the course of genomic research to ensure appropriate care for patients. Genetic counseling within HCP and validation of variants in the clinically accredited Cancer Genetics Laboratory enables seamless return of research generated clinically relevant germline results to affected subjects. Clinical trial information: NCT02155621.

A Simple Technique to Prevent Early or Late EndoButton Deployment in Anterior Cruciate Ligament Reconstructions: A Technical Note.

The use of suspensory graft fixation methods in arthroscopic reconstruction of the anterior cruciate ligament has become increasingly popular with the more frequent use of tendon grafts and anatomical techniques involving the creation of bone tunnels. An important technical step to ensure adequate fixation of the graft when using EndoButton-type implants, particularly in the femur, involves flipping the EndoButton plate at the correct length to avoid performing this maneuver before the appropriate time or leaving soft tissue between the plate and lateral cortex of the femur. In the present study, we describe a simple arthroscopic technique for indicating the correct time to flip/deploy the EndoButton plate.

Modified MicroSecure Vitrification: A Safe, Simple and Highly Effective Cryopreservation Procedure for Human Blastocysts.

Clinical embryo vitrification evolved with the development of unique vitrification devices in the 21st century and with the misconception that ultra-rapid cooling in an "open" system (i.e., direct LN2 contact) was a necessity to optimize vitrification success. The dogma surrounding the importance of cooling rates led to unsafe practices subject to technical variation and to the creation of vitrification devices that disregarded important quality-control factors (e.g., ease of use, repeatability, reliability, labeling security, and storage safety). Understanding the quality-control flaws of other devices allowed for the development of a safe, secure, repeatable, and reliable µS-VTF method aimed to minimize intra- and inter-technician variation. Equally important, it combined the availability of two existing FDA-compliant devices: 1) a 0.3-mL ionomeric resin embryo straw with internalized, dual-colored, tamper-proof labeling with repeatable weld seal potential; and 2) shortened, commonly-used, 300-µm ID sterile flexipettes to directly load the embryo(s) in order to create a highly-effective global vitrification device. Like other aseptic, closed vitrification systems (e.g., High Security Vitrification (HSV), Rapid-i, and VitriSafe) effectively used in reproductive medicine, microSecure Vitrification (µS-VTF) has proven that it can achieve high post-warming survival and pregnancy outcomes with its attention to simplicity, and reduced technical variation. Although the 0.3-mL embryo straw containing an internal hydrophobic plug was commercially replaced with a standard semen straw possessing cotton-polyvinyl pyrrolidone (PVP) plugs, it maintained its ionomeric resin composition to ensure weld sealing. However, the cotton plugs can wick out the fluid-embryo contents of the flexipettes upon contact. A modified µS-VTF method was adapted to include an additional internal weld seal before the plug on the device loading side. The added technical step to the µS-VTF procedure has not affected its successful application, as high survival rates (> 95%) and pregnancy rates continue today.

Modified MicroSecure Vitrification: A Safe, Simple and Highly Effective Cryopreservation Procedure for Human Blastocysts

Clinical embryo vitrification evolved with the development of unique vitrification devices in the 21st century and with the misconception that ultra-rapid cooling in an "open" system (i.e., direct LN2 contact) was a necessity to optimize vitrification success. The dogma surrounding the importance of cooling rates led to unsafe practices subject to technical variation and to the creation of vitrification devices that disregarded important quality-control factors (e.g., ease of use, repeatability, reliability, labeling security, and storage safety). Understanding the quality-control flaws of other devices allowed for the development of a safe, secure, repeatable, and reliable µS-VTF method aimed to minimize intra- and inter-technician variation. Equally important, it combined the availability of two existing FDA-compliant devices: 1) a 0.3-mL ionomeric resin embryo straw with internalized, dual-colored, tamper-proof labeling with repeatable weld seal potential; and 2) shortened, commonly-used, 300-µm ID sterile flexipettes to directly load the embryo(s) in order to create a highly-effective global vitrification device. Like other aseptic, closed vitrification systems (e.g., High Security Vitrification (HSV), Rapid-i, and VitriSafe) effectively used in reproductive medicine, microSecure Vitrification (µS-VTF) has proven that it can achieve high post-warming survival and pregnancy outcomes with its attention to simplicity, and reduced technical variation. Although the 0.3-mL embryo straw containing an internal hydrophobic plug was commercially replaced with a standard semen straw possessing cotton-polyvinyl pyrrolidone (PVP) plugs, it maintained its ionomeric resin composition to ensure weld sealing. However, the cotton plugs can wick out the fluid-embryo contents of the flexipettes upon contact. A modified µS-VTF method was adapted to include an additional internal weld seal before the plug on the device loading side. The added technical step to the µS-VTF procedure has not affected its successful application, as high survival rates (> 95%) and pregnancy rates continue today.

Radiologists - Threatened by a veritable identity crisis?

Radiologists - Threatened by a veritable identity crisis?

The social security number: A small device underpinning big systems

AbstractThe possession and use of a personal social security number helps to structure people's daily lives. However, despite its fundamental normative importance, the social security number remains a little‐known entity. Increasingly universal and yet diverse in form, it is a legal and technical norm which is as much a mechanism for surveillance and monitoring as it is a necessary instrument for giving effect to social rights. Analysis of this constituent element of social security systems permits as assessment of some of the technical difficulties presented by the ever‐increasing movement of people and data. Overcoming these technical difficulties should permit to envisage a first technical step towards realizing a universal and global social security system.

Drill wobble - effect on femoral tunnel aperture during anterior cruciate ligament reconstruction.

BackgroundIn anterior cruciate ligament reconstruction performed using cortical button fixation on the femur, we have observed a “wobble” effect that can occur when a cannulated femoral drill is used over a guide pin that is not securely fixed in bone. Our study assessed the effect of drill “wobble” on femoral tunnel aperture in sawbones.MethodsFemoral tunnels were drilled in sawbones, which had been divided in two groups of 10 each, per drilling technique. The “wobble” technique group had the smaller cortical button drill passed before drilling the graft socket with the bigger diameter femoral drill. In contrast, in the “non-wobble” technique group, the smaller cortical button drill was passed after drilling the graft socket. The aperture dimensions: antero-posterior, proximo-distal and oblique, as well as the length of each tunnel, were measured.ResultsWhile the average dimensions of the tunnels were similar between the two techniques, there was significantly more variation in the antero-posterior measurements for the wobble technique as compared to the non-wobble technique (mean 7.3 mm, SD 0.28 mm, and mean 7.3 mm, SD 0.11 mm, respectively; Brown-Forsythe test, p 0.02).ConclusionWe conclude that using the “socket first” “non-wobble” technique is a single surgical technical step surgeons can employ to decrease variability in tunnel aperture and size.

A redox-stable direct-methane solid oxide fuel cell (SOFC) with Sr2FeNb0.2Mo0.8O6−δ double perovskite as anode material

Development of high-performing and redox-stable ceramic oxide electrode materials is a crucial technical step for direct hydrocarbon solid oxide fuel cells (SOFCs) operating at intermediate temperatures (550–700 °C). Here we report a nickel-free double perovskite, Sr2FeNb0.2Mo0.8O6−δ (SFNM20), for SOFC anode, and this anode shows outstanding performances with high resistance against carbon build-up and redox cycling in hydrocarbon fuels. At 800 °C, the SFNM20 anode shows electrical conductivity of 5.3 S cm−1 in 5% H2 and peak power densities of 520 and 380 mW cm−2 using H2 and CH4 as the fuel, respectively. The cell exhibits a very stable performance under different constant current loads in H2 and CH4 at 700 °C and high redox stability against the gas environment changes in the anode chamber. In addition, the electrode is structurally stable in various fuels, suggesting that it is a feasible material candidate for the electrode of high-performing SOFCs.

Geometry of the Aortic Arch After Initial Hybrid or Norwood Palliation

Aortic arch reconstruction is a challenging technical step in the Norwood operation or the comprehensive stage II operation. This study sought to analyze differences in aortic arch geometry and dimensions in patients undergoing Norwood or hybrid palliation. Retrospective data collection included all patients who underwent Norwood or hybrid palliation at the Hospital for Sick Children in Toronto between 2007 and 2014; 139 patients were analyzed. Lateral angiograms obtained before stage II or comprehensive stage II and Fontan completion were measured at the level of the ascending aorta, transverse arch, isthmus, and descending aorta. Reintervention rate and type were assessed. Before stage II or comprehensive stage II hybrid procedures, patients had significantly larger descending aorta z-scores. Patients undergoing Norwood operations showed a significant increase in descending aorta z-scores before Fontan completion. Comparable dimensions (absolute and z-scores) were found at all points of measurements before Fontan completion. Geometry was similar in both groups but significantly different compared with normal dimensions. Reduction in aortic arch diameter happened almost solely in the segment between the transverse arch and the isthmus. Stent inclusion in patients undergoing hybrid procedures led to similar dimensions. Reintervention rates were verylow (Norwood 9.6% vs hybrid 7.6%). Reintervention for excessive dimensions was as common as for recoarctation. Aortic arch geometry and growth is not altered by palliation type. The increase in descending aorta diameter seen in patients undergoing Norwood operations is in accord with physiologic changes and may reflect catch-up growth. Reintervention rates are low and are not related to recoarctation alone.

A galectin from Hyriopsis cumingii involved in the innate immune response against to pathogenic microorganism and its expression profiling during pearl sac formation.

Hyriopsis cumingii is the most important freshwater pearl mussel cultured in China. The operation for implantation is one necessary technical step for pearl culture. However, implantation-induced trauma results in a series of immune responses and can enable the invasion of pathogenic microbes. Lectin proteins are found widely in nature and play important roles in innate immunity. Galectins are members of the lectin superfamily and are characterized by one or several carbohydrate recognition domains (CRDs) that produce multiple sugar binding sites on the protein. Here we cloned and characterized the H.cumingii galectin gene HcGal1, which encodes a 312 amino acid galectin protein. The HcGal1 transcript was detected in all tested H.cumingii tissues and showed higher expression specifically in immune tissues. The significant upregulation of HcGal1 expression was observed after challenging the mussel with lipopolysaccharide or Gram-negative and Gram-positive bacteria. After implantation, significant downregulation of the HcGal1 transcript was noted in the mantle, hemocytes, and pearl sac in the acute-stress stage (0-24h) and the stage of wound healing and pearl-sac formation (24h-7d). In addition, significant upregulation of HcGal1 expression was observed in the liver in the stage of wound healing and pearl-sac formation. In the pearl-secretion stage (7-35d), the HcGal1 transcript levels returned to normal in all tested tissues. We also show that recombinantly expressed and purified HcGal1 can agglutinate some Gram-negative and Gram-positive bacteria. In addition, invivo experiments showed that the recombinant protein HcGal1 could promote phagocytosis by hemocytes. Our data suggest that HcGal1 plays a role in innate immune responses involved in pathogen recognition and wound healing.

An algebraic approach to virtual fundamental cycles on moduli spaces of pseudo-holomorphic curves

We develop techniques for defining and working with virtual fundamental cycles on moduli spaces of pseudo-holomorphic curves which are not necessarily cut out transversally. Such techniques have the potential for applications as foundations for invariants in symplectic topology arising from "counting" pseudo-holomorphic curves. We introduce the notion of an *implicit atlas* on a moduli space, which is (roughly) a convenient system of local finite-dimensional reductions. We present a general intrinsic strategy for constructing a canonical implicit atlas on any moduli space of pseudo-holomorphic curves. The main technical step in applying this strategy in any particular setting is to prove appropriate gluing theorems. We require only topological gluing theorems, that is, smoothness of the transition maps between gluing charts need not be addressed. Our approach to virtual fundamental cycles is algebraic rather than geometric (in particular, we do not use perturbation). Sheaf-theoretic tools play an important role in setting up our functorial algebraic "VFC package". We illustrate the methods we introduce by giving definitions of Gromov--Witten invariants and Hamiltonian Floer homology over $\mathbb Q$ for general symplectic manifolds. Our framework generalizes to the $S^1$-equivariant setting, and we use $S^1$-localization to calculate Hamiltonian Floer homology. The Arnold conjecture (as treated by Floer, Hofer--Salamon, Ono, Liu--Tian, Ruan, and Fukaya--Ono) is a well-known corollary of this calculation.

Clinical Application of an Original Vascular Anastomosis: A Clinical Multicenter Study

Vascular anastomosis is the most important technical step required for the possibility of free tissue transfer, and mismatch of the donor and recipient vessel size is the most common surgical challenge. As recent reports have described a new method (Ren anastomosis) to resolve this challenge, the goal of this study was to assess these newly described microvascular anastomosis methods. The study was conducted at 2 institutes in China in different periods. Patients were recruited from the Second Xiangya Hospital between February and May 2013 and from the Shanghai Ninth People's Hospital between March and May 2015. All patients who participated in this study needed free flaps for oral and maxillofacial defects. Patients were divided into the experimental group and the control group. In the experimental group, isometric double-notch end-in-end microvascular anastomosis (Ren anastomosis) was performed, whereas end-to-end anastomosis was performed in the control group. Statistical differences were assessed by use of χ2 and t tests. A total of 148 patients (108 men and 40 women) were treated over the course of this study. There was a statistically significant difference (P < .001) in the operation time for the microscopic arteryanastomosis between the experimental group (70 cases; 5.6 ± 1.8minutes) and the control group (78 cases; 14.6 ± 3.7minutes). In each group, 1 case of artery compromise was observed. The Ren anastomosis was time-saving, straightforward, efficient, and easy to learn, with a high patency rate.