The role of stimulating bystander cells in bispecific antibody-mediated T cell activation

Recently, bispecific monoclonal antibodies (BsMAb) yielded promising results regarding the treatment of various malignant diseases. However, the serum levels that can be safely achieved in humans remain suboptimal in terms of saturation of the tumor-associated antigen (TAA), as the application of higher doses results in systemic cytokine release and transient leukopenia. Off-target activation of circulating immune cells, stimulated by the BsMAb, could explain most of the side effects observed after in-vivo application of BsMAb. In this work, immunoassays were conducted, which showed that BsMAb are capable of inducing T cell proliferation in the absence of target cells. Furthermore, it was demonstrated that human umbilical vein endothelial cells (HUVECs) as well as various lymphoid cell lines markedly amplify this effect. They are acting as stimulating bystander cells (SBCs). Several lymphoid and non-lymphoid cell lines did not exhibit this property, thus ruling out allogeneic effects. Functional assays were conducted to characterize blocking antibodies directed against costimulatory and adhesion molecules on both stimulating bystander cells and immune cells, that are capable of reducing off-target activation. Antibodies against CD54, CD2, and TNFα reduced off-target activation markedly, whereas antibodies directed against CD18 as well as prednisolone blocked it completely. Further experiments addressed the effect of blocking reagents on the induction of on- target activation and subsequent killing of tumor cells by T cells after stimulation with BsMAb. Prednisolone and aCD2 antibodies impaired tumor cell killing markedly, while only minor reduction was observed with aCD18 and aTNFα antibodies. Using an adhesion assay, it was shown that PBMCs adhere to a HUVEC layer to a higher extent when stimulated with BsMAb compared to unstimulated PBMCs. Antibodies directed against CD18 could greatly reduce this phenomenon. In summary, aCD18 and aTNFα antbodies appear to be promising agents for the prevention of cytokine release after application of BsMAb. Eventually, they could replace prednisolone as first line drug, since prednisolone exerted a pronounced inhibition of BsMAb-induced tumor cell killing. However, further in-vivo assays need to be conducted to further characterize the blocking properties of aCD18 and aTNFα blocking antibodies and possible side effects, such as a higher susceptibility to infection.In den letzten Jahren erzielten bispezifische monoklonale Antikörper (BsMAb) bemerkenswerte Erfolge in der Behandlung von malignen Erkrankungen. Die in Menschen erreichbaren Serumkonzentrationen dieser Antikörper sind jedoch nicht ausreichend, um die anvisierten Tumorantigene (TAA) zu sättigen. Der Grund dafür ist das Auftreten eines Zytokinfreisetzungssyndroms und einer transienten Leukopenie. Die Aktivierung von zirkulierenden Immunzellen durch BsMAb abseits von Tumorzellen würde die meisten Nebeneffekte erklären, die bisher beobachtet wurden. In dieser Arbeit wurden Immunassays durchgeführt, die zeigten, dass BsMAb T-Zell-Proliferation in der Abwesenheit von Zielzellen induzieren können. Außerdem konnte gezeigt werden, dass humane Nabelschnurvenenendothelzellen (HUVECs) sowie verschiedene lymphatische Zelllinien diesen Effekt verstärken. Deswegen wurden sie im weiteren als stimulierende Bystanderzellen (SBCs) bezeichnet. Andere lymphatische und nicht-lymphatische Zellen verstärkten die unspezifische Aktivierung nicht, wodurch ein rein allogener Effekt ausgeschlossen werden konnte. Blockierende Antikörper, die gegen bestimmte Zytokine, kostimulierende und Adhäsionsmoleküle auf Immunzellen und stimulierenden Nebenzellen gerichtet waren, konnten die unspezifische Aktivierung reduzieren. So konnten Antikörper gegen CD54, CD2 oder TNFα den Effekt deutlich eindämmen. Antikörper gegen CD18 sowie das Steroid Prednisolon konnten die unspezifische Aktivierung vollständig unterbinden. Weitere Experimente wurden durchgeführt, um den Einfluss der blockierenden Antikörper auf die BsMAb-induzierte T-Zell-Aktivierung, sowie das nachfolgende Killing von Tumorzellen zu untersuchen. So blockierten Prednisolon und Antikörper gegen CD2 die Aktivierung und Tumorzelllyse vollständig, während Anti-CD18-Antikörper und Anti- TNFα-Antikörper kaum negative Effekte hatten. Mittels eines Adhäsionsassays konnte gezeigt werden, dass PBMCs vermehrt an Endothelzellen (HUVECs) haften, wenn sie vorher mit BsMAb stimuliert wurden. Dieser Effekt konnte mit Antikörpern gegen CD18 unterbrochen werden. Im Endeffekt stellen Antikörper gegen CD18 und TNFα mögliche Medikamente zur Verhinderung von Zytokinfreisetzungssyndromen dar. Nach eingehender Charakterisierung könnten beide Prednisolon als Therapeutikum der Wahl für diese Indikation ersetzen. Letzteres hatte die Anti-Tumor-Aktivität der T-Zellen in den hier durchgeführten Experimenten deutlich eingeschränkt. Nichtsdestotrotz sollten weitere in-vivo Assays durchgeführt werden, um Antikörper gegen CD18 und TNFα weiter zu charakterisieren und mögliche Nebenwirkungen zu erkennen, zum Beispiel eine höhere Anfälligkeit für Infektionen

The IceCube Neutrino Observatory: Instrumentation and Online Systems

The IceCube Neutrino Observatory is a cubic-kilometer-scale high-energy neutrino detector built into the ice at the South Pole. Construction of IceCube, the largest neutrino detector built to date, was completed in 2011 and enabled the discovery of high-energy astrophysical neutrinos. We describe here the design, production, and calibration of the IceCube digital optical module (DOM), the cable systems, computing hardware, and our methodology for drilling and deployment. We also describe the online triggering and data filtering systems that select candidate neutrino and cosmic ray events for analysis. Due to a rigorous pre-deployment protocol, 98.4% of the DOMs in the deep ice are operating and collecting data. IceCube routinely achieves a detector uptime of 99% by emphasizing software stability and monitoring. Detector operations have been stable since construction was completed, and the detector is expected to operate at least until the end of the next decade.Comment: 83 pages, 50 figures; updated with minor changes from journal review and proofin

Mechanical design of the optical modules intended for IceCube-Gen2

IceCube-Gen2 is an expansion of the IceCube neutrino observatory at the South Pole that aims to increase the sensitivity to high-energy neutrinos by an order of magnitude. To this end, about 10,000 new optical modules will be installed, instrumenting a fiducial volume of about 8 km3. Two newly developed optical module types increase IceCube’s current sensitivity per module by a factor of three by integrating 16 and 18 newly developed four-inch PMTs in specially designed 12.5-inch diameter pressure vessels. Both designs use conical silicone gel pads to optically couple the PMTs to the pressure vessel to increase photon collection efficiency. The outside portion of gel pads are pre-cast onto each PMT prior to integration, while the interiors are filled and cast after the PMT assemblies are installed in the pressure vessel via a pushing mechanism. This paper presents both the mechanical design, as well as the performance of prototype modules at high pressure (70 MPa) and low temperature (−40∘C), characteristic of the environment inside the South Pole ice

Cross Correlation of IceCube Neutrinos with Tracers of Large Scale Structure

The origin of most astrophysical neutrinos is unknown, but extragalactic neutrino sources may follow the spatial distribution of the large-scale structure of the universe. Galaxies also follow the same large scale distribution, so establishing a correlation between galaxies and IceCube neutrinos could help identify the origins of the diffuse neutrinos observed by IceCube. Following a preliminary study based on the WISE and 2MASS catalogs, we will investigate an updated galaxy catalog with improved redshift measurements and reduced stellar contamination. Our IceCube data sample consists of track-like muon neutrinos selected from the Northern sky. The excellent angular resolution of track-like events and low contamination with atmospheric muons is necessary for the sensitivity of the analysis. Unlike a point source stacking analysis, the calculation of the cross correlation does not scale with the number of entries in the catalog, making the work tractable for catalogs with millions of objects. We present the development and performance of a two-point cross correlation of IceCube neutrinos with a tracer of the large scale structure

The next generation neutrino telescope: IceCube-Gen2

The IceCube Neutrino Observatory, a cubic-kilometer-scale neutrino detector at the geographic South Pole, has reached a number of milestones in the field of neutrino astrophysics: the discovery of a high-energy astrophysical neutrino flux, the temporal and directional correlation of neutrinos with a flaring blazar, and a steady emission of neutrinos from the direction of an active galaxy of a Seyfert II type and the Milky Way. The next generation neutrino telescope, IceCube-Gen2, currently under development, will consist of three essential components: an array of about 10,000 optical sensors, embedded within approximately 8 cubic kilometers of ice, for detecting neutrinos with energies of TeV and above, with a sensitivity five times greater than that of IceCube; a surface array with scintillation panels and radio antennas targeting air showers; and buried radio antennas distributed over an area of more than 400 square kilometers to significantly enhance the sensitivity of detecting neutrino sources beyond EeV. This contribution describes the design and status of IceCube-Gen2 and discusses the expected sensitivity from the simulations of the optical, surface, and radio components

Sensitivity of IceCube-Gen2 to measure flavor composition of Astrophysical neutrinos

The observation of an astrophysical neutrino flux in IceCube and its detection capability to separate between the different neutrino flavors has led IceCube to constraint the flavor content of this flux. IceCube-Gen2 is the planned extension of the current IceCube detector, which will be about 8 times larger than the current instrumented volume. In this work, we study the sensitivity of IceCube-Gen2 to the astrophysical neutrino flavor composition and investigate its tau neutrino identification capabilities. We apply the IceCube analysis on a simulated IceCube-Gen2 dataset that mimics the High Energy Starting Event (HESE) classification. Reconstructions are performed using sensors that have 3 times higher quantum efficiency and isotropic angular acceptance compared to the current IceCube optical modules. We present the projected sensitivity for 10 years of data on constraining the flavor ratio of the astrophysical neutrino flux at Earth by IceCube-Gen2

Estimating the coincidence rate between the optical and radio array of IceCube-Gen2

The IceCube-Gen2 Neutrino Observatory is proposed to extend the all-flavour energy range of IceCube beyond PeV energies. It will comprise two key components: I) An enlarged 8km3 in-ice optical Cherenkov array to measure the continuation of the IceCube astrophysical neutrino flux and improve IceCube\u27s point source sensitivity above ∼100TeV; and II) A very large in-ice radio array with a surface area of about 500km2. Radio waves propagate through ice with a kilometer-long attenuation length, hence a sparse radio array allows us to instrument a huge volume of ice to achieve a sufficient sensitivity to detect neutrinos with energies above tens of PeV. The different signal topologies for neutrino-induced events measured by the optical and in-ice radio detector - the radio detector is mostly sensitive to the cascades produced in the neutrino interaction, while the optical detector can detect long-ranging muon and tau leptons with high accuracy - yield highly complementary information. When detected in coincidence, these signals will allow us to reconstruct the neutrino energy and arrival direction with high fidelity. Furthermore, if events are detected in coincidence with a sufficient rate, they resemble the unique opportunity to study systematic uncertainties and to cross-calibrate both detector components

Direction reconstruction performance for IceCube-Gen2 Radio

The IceCube-Gen2 facility will extend the energy range of IceCube to ultra-high energies. The key component to detect neutrinos with energies above 10 PeV is a large array of in-ice radio detectors. In previous work, direction reconstruction algorithms using the forward-folding technique have been developed for both shallow (≲20 m) and deep in-ice detectors, and have also been successfully used to reconstruct cosmic rays with ARIANNA. Here, we focus on the reconstruction algorithm for the deep in-ice detector, which was recently introduced in the context of the Radio Neutrino Observatory in Greenland (RNO-G)

Sensitivity of the IceCube-Gen2 Surface Array for Cosmic-Ray Anisotropy Studies

The energy of the transition from Galactic to extra-galactic origin of cosmic rays is one of the major unresolved issues of cosmic-ray physics. However, strong constraints can be obtained from studying the anisotropy in the arrival directions of cosmic rays. The sensitivity to cosmic-ray anisotropy is, in particular, a matter of statistics. Recently, the cosmic ray anisotropy measurements in the TeV to PeV energy range were updated from IceCube using 11 years of data. The IceCube-Gen2 surface array will cover an area about 8 times larger than the existing IceTop surface array with a corresponding increase in statistics and capability to investigate cosmic-ray anisotropy with higher sensitivity. In this contribution, we present details on the performed simulation studies and sensitivity to the cosmic-ray anisotropy signal for the IceCube-Gen2 surface array

Deep Learning Based Event Reconstruction for the IceCube-Gen2 Radio Detector

The planned in-ice radio array of IceCube-Gen2 at the South Pole will provide unprecedented sensitivity to ultra-high-energy (UHE) neutrinos in the EeV range. The ability of the detector to measure the neutrino’s energy and direction is of crucial importance. This contribution presents an end-to-end reconstruction of both of these quantities for both detector components of the hybrid radio array (\u27shallow\u27 and \u27deep\u27) using deep neural networks (DNNs). We are able to predict the neutrino\u27s direction and energy precisely for all event topologies, including the electron neutrino charged-current (νe-CC) interactions, which are more complex due to the LPM effect. This highlights the advantages of DNNs for modeling the complex correlations in radio detector data, thereby enabling a measurement of the neutrino energy and direction. We discuss how we can use normalizing flows to predict the PDF for each individual event which allows modeling the complex non-Gaussian uncertainty contours of the reconstructed neutrino direction. Finally, we discuss how this work can be used to further optimize the detector layout to improve its reconstruction performance