Immunologische Grundlage der abskopalen Antitumorreaktionen, induziert durch die Kombination unterschiedlicher Strahlentherapie-Fraktionierungen mit autologen Tumorimpfstoffen und Checkpoint-Inhibition

Ionizing radiation is commonly used in radiotherapy (RT) for the local treatment of solid tumors to kill cancer cells and to stop tumor progression. Meanwhile, RT is also known to have immune-stimulatory properties by causing immunogenic tumor cell death turning tumors into in situ cancer vaccines eventually leading to anti-tumor immune responses. However, due to the immunosuppressive tumor microenvironment, after RT alone the immune system often fails to completely eradicate residual tumor masses of locally irradiated lesions or distant metastases. Although therapies are continuously improving, these systemic immune responses after RT that also target distant metastases outside of the radiation field, the so called abscopal effects, are still rarely seen in the clinic. Thus, immunotherapies are needed to break immunosuppression or to amplify RT-induced anti-tumor immune responses. The latter can be addressed by autologous whole tumor cell-based vaccines which comprise tumor-associated or -specific antigens of a particular patient. Since their approval for the treatment of several tumor entities, immune checkpoint inhibitors, counteracting tolerance induction and repressed tumor cell killing by immune cells, have achieved great success. The combined treatment with RT is currently under investigation in many clinical trials but is by far not standard yet. Here we aimed to generate inactivated whole tumor cell vaccines with high hydrostatic pressure (HHP) which are safe but still immunogenic to boost RT-induced immune responses. We hypothesized that RT and such HHP-vaccines synergize to generate a tumor microenvironment fostering tumor growth retardation in locally irradiated tumors and that systemic anti-tumor immune reactions can also be triggered when anti-PD-1 immune checkpoint inhibitors are applied in addition. In pre-clinical model systems we subcutaneously (s.c.) injected mice with syngeneic B16 melanoma or CT26 colon carcinoma cells, respectively. To evaluate the efficiency of RT in combination with HHP vaccines, established tumors were locally irradiated with a hypofractionated schedule and mice were injected next to the tumor with HHP-vaccines generated with 200 MPa. In an abscopal tumor model with local irradiation of only one of two B16 tumors (one on each flank) using two differently hypofractionated schedules with 2x8 Gy or 3x8 Gy, HHP-vaccines were injected s.c. in the neck. Additionally, mice concurrently received anti-PD-1 antibodies. Tumor growth and survival of the mice was monitored and the immunological tumor microenvironment was analyzed via multi-color flow cytometry, qPCR and ELISA. Tumor cells treated with a pressure of 200 MPa were completely inactivated and lost the potential to form colonies in vitro or tumors in vivo. Combined treatment of local irradiation and therapeutic HHP-vaccination resulted in retarded tumor growth and prolonged survival of the mice in both tumor models. The number of cells per gram of tumor across several immune cell subtypes was only significantly increased in B16 tumors that were treated with both, RT and HHP-vaccines. Irradiated tumors showed higher expression of suppressive immune checkpoint ligands and most of the tumor-infiltrating T cells were positive for PD-1 expression, providing a rationale for the combination with anti-PD-1 checkpoint inhibition. Although injected distantly in the abscopal setting, HHP-vaccines also further retarded the growth of locally irradiated tumors even though in all radioimmunotherapy combinations the total number of immune cells was decreased in primary tumors and the peripheral blood. However, HHP-vaccination combined with RT failed to significantly induce abscopal anti-tumor immune responses and even partly abrogated those which were induced with RT plus anti-PD-1. In this group, the abscopal effects were accompanied by an elevated infiltration of CD8+ T cells. Adding another fraction of 8 Gy completely abrogated abscopal effects and in vitro data revealed that multiple immunosuppressive and oncogenic pathways including cell death, cytokine release, and checkpoint expression were differently modulated with 3x8 Gy when compared to 2x8 Gy. We further found that high intratumoral concentrations of IGFBP-6 correlated with abscopal responses. Surprisingly, knockdown of IGFBP-6 inhibited tumor growth in vitro and in vivo. We conclude that autologous whole tumor cell-based vaccines generated with HHP are safe and suitable to be combined with RT and work irrespective of the injection site of the vaccine if the tumor was previously irradiated. However, HHP-vaccines without any additional adjuvant do not increase abscopal anti-tumor immune responses induced with RT plus anti-PD-1 and thus need further optimization in the future.Ionisierende Strahlung wird üblicherweise in der Strahlentherapie (RT) zur lokalen Behandlung von soliden Tumoren verwendet, um Krebszellen abzutöten und das Fortschreiten des Tumors zu stoppen. Mittlerweile ist auch bekannt, dass RT immunstimulierende Eigenschaften hat, indem es immunogenen Tumorzelltod verursacht und Tumore in in-situ-Krebsimpfstoffe verwandelt, die schließlich zu anti-Tumor-Immunantworten führen. Aufgrund der immunsuppressiven Tumormikroumgebung kann das Immunsystem jedoch die verbleibenden Tumormassen lokal bestrahlter Läsionen oder Fernmetastasen außerhalb des Bestrahlungsfeldes nach RT allein oft nicht vollständig eliminieren. Obwohl sich die Therapien kontinuierlich verbessern, werden diese systemischen Immunantworten nach RT, welche auch unter dem Begriff abskopaler Effekt bekannt sind, in der Klinik immer noch selten beobachtet. Daher sind Immuntherapien erforderlich, um die Immunsuppression zu unterbrechen oder RT-induzierte anti-Tumor-Immunantworten zu verstärken. Letzteres kann durch autologe Impfstoffe auf der Basis ganzer Tumorzellen realisiert werden, die tumorassoziierte oder -spezifische Antigene eines bestimmten Patienten beinhalten. Seit ihrer Zulassung für die Behandlung mehrerer Tumorentitäten haben Immun-Checkpoint-Inhibitoren, die der Toleranzinduktion und der unterdrückten Abtötung von Tumorzellen durch Immunzellen entgegenwirken, große Erfolge erzielt. Die Behandlung in Kombination mit RT wird derzeit in vielen klinischen Studien untersucht, ist jedoch bei weitem noch nicht Standard. In dieser Arbeit war es unser Ziel inaktivierte Ganzzell-Tumorimpfstoffe mit hohem hydrostatischem Druck (HHP) zu erzeugen, die sicher und immunogen sind, um RT-induzierte Immunantworten zu verstärken. Wir stellten die Hypothese auf, dass RT und solche HHP-Impfstoffe zusammenwirken, um eine Tumormikroumgebung zu erzeugen, die die Verzögerung des Tumorwachstums bei lokal bestrahlten Tumoren fördert, und dass darüber hinaus systemische anti-Tumor-Immunreaktionen ausgelöst werden können, wenn zusätzlich anti-PD-1-Immun-Checkpoint-Inhibitoren angewendet werden. In präklinischen Modellsystemen injizierten wir Mäusen subkutan (s.c.) syngene B16-Melanom- oder CT26-Kolonkarzinomzellen. Um die Effizienz von RT in Kombination mit HHP-Impfstoffen zu untersuchen, wurden etablierte Tumoren lokal mit einem hypofraktionierten Schema bestrahlt und Mäusen wurden neben dem Tumor HHP-Impfstoffe injiziert, die mit 200 MPa erzeugt wurden. In einem abskopalen Tumormodell mit lokaler Bestrahlung von nur einem von zwei B16-Tumoren (einer an jeder Flanke) unter Verwendung von zwei unterschiedlich hypofraktionierten Zeitplänen mit 2×8 Gy oder 3×8 Gy wurden HHP-Impfstoffe s.c. im Nacken injiziert. Zusätzlich erhielten Mäuse gleichzeitig anti-PD-1-Antikörper. Das Tumorwachstum und das Überleben der Mäuse wurden überwacht und die immunologische Tumormikroumgebung wurde mittels Mehrfarben-Durchflusszytometrie, qPCR und ELISA analysiert. Mit einem Druck von 200 MPa behandelte Tumorzellen wurden vollständig inaktiviert und verloren das Potenzial zur Bildung von Kolonien in vitro oder von Tumoren in vivo. Die kombinierte Behandlung mit lokaler Bestrahlung und therapeutischer HHP-Impfung führte in beiden Tumormodellen zu einem verzögerten Tumorwachstum und einem verlängerten Überleben der Mäuse. Die Menge an Zellen pro Gramm Tumor über mehrere Immunzell-Subtypen hinweg war nur bei B16-Tumoren, die sowohl mit RT als auch mit HHP-Impfstoffen behandelt wurden, signifikant erhöht. Bestrahlte Tumoren zeigten eine höhere Expression von supprimierenden Immun-Checkpoint-Liganden und die meisten Tumor-infiltrierenden T-Zellen waren PD-1 positiv, was eine Begründung für die Kombination mit anti-PD-1-Checkpoint-Inhibition liefert. Obwohl HHP-Impfstoffe entfernt von den Tumoren verabreicht wurden, verzögerten sie trotzdem das Wachstum lokal bestrahlter Tumoren weiter, obwohl in allen Radioimmuntherapiekombinationen die Gesamtzahl der Immunzellen in Primärtumoren und im peripheren Blut verringert war. Eine mit RT kombinierte HHP-Impfung induzierte jedoch keine signifikanten abskopalen anti-Tumor-Immunantworten und hob sogar diejenigen, die mit RT plus Anti-PD-1 induziert wurden, teilweise auf. In dieser Gruppe gingen die abskopalen Effekte mit einer erhöhten Infiltration von CD8+ T-Zellen einher. Eine zusätzliche Fraktion von 8 Gy hob die abskopalen Effekte vollständig auf und in-vitro-Daten zeigten, dass mit 3×8 Gy im Vergleich zu 2x8 Gy eher immunsuppressive und onkogene Signalwege, einschließlich Zelltod, Zytokinfreisetzung und Checkpoint-Expression, moduliert wurden. Außerdem fanden wir heraus, dass hohe intratumorale Konzentrationen von IGFBP-6 mit abskopalen Reaktionen korrelierten. Überraschenderweise hemmte ein IGFBP-6 knockdown allerdings das Tumorwachstum in vitro und in vivo. Wir schließen daraus, dass mit HHP erzeugte autologe Impfstoffe auf der Basis ganzer Tumorzellen sicher und geeignet sind, mit RT kombiniert zu werden und unabhängig von der Injektionsstelle des Impfstoffs wirken, wenn der Tumor zuvor bestrahlt wurde. HHP-Impfstoffe ohne zusätzliches Adjuvans erhöhen jedoch nicht die mit RT plus Anti-PD-1 induzierten abskopalen Antitumor-Immunantworten und müssen daher in Zukunft weiter optimiert werden

Trastuzumab (Herceptin (R)): Monoclonal antibody in the treatment of HER2/neu-overexpressing breast cancer in the metastatic and (neo)adjuvant situation

Trastuzumab (Herceptin (R)) is a humanized monoclonal antibody that specifically targets HER2/neu (human epidermal growth factor receptor-2) breast cancer cells, which are overexpressed in about 25-30% of breast carcinomas. After phase I and II trials, several phase III studies of trastuzumab alone or in combination with various chemotherapies were conducted. Patients with HER2/neu overexpression levels of 3+ determined by immunohistochemical assay or gene amplification (fluorescence in situ hybridization) derive most clinical benefit from trastuzumab. Taking into consideration efficacy and side effect profile, the combination of trastuzumab and paclitaxel showed an improvement of all clinical parameters, including overall survival, for the first time in the history of palliative breast cancer therapy. The application of trastuzumab has meanwhile become an established part of systemic therapy of metastastic breast cancer, and excellent data of its application in the adjuvant setting now exist (NSABP-B31, NCCTG-N9831, HERA), with significantly better relapse-free survival in the treatment arms with trastuzumab. Ongoing trials investigate the role of trastuzumab in the neoadjuvant setting. Trastuzumab is generally well tolerated. Cardiotoxicity is the main concern, thus monitoring of cardiac function is recommended

Benchmarking of Neuromorphic Hardware Systems

Ostrau C, Klarhorst C, Thies M, Rückert U. Benchmarking of Neuromorphic Hardware Systems. In: Neuro-inspired Computational Elements Workshop (NICE ’20), March 17–20, 2020, Heidelberg, Germany. International Conference Proceeding Series (ICPS). Association for Computing Machinery (ACM); 2020.With more and more neuromorphic hardware systems for the accel- eration of spiking neural networks available in science and industry, there is a demand for platform comparison and performance esti- mation of such systems. This work describes selected benchmarks implemented in a framework with exactly this target: independent black-box benchmarking and comparison of platforms suitable for the simulation/emulation of spiking neural networks

Comparing Neuromorphic Systems by Solving Sudoku Problems

Ostrau C, Klarhorst C, Thies M, Rückert U. Comparing Neuromorphic Systems by Solving Sudoku Problems. In: Conference Proceedings: 2019 International Conference on High Performance Computing & Simulation (HPCS). Piscataway, NJ: IEEE; Accepted.In the field of neuromorphic computing several hardware accelerators for spiking neural networks have been introduced, but few studies actually compare different systems. These comparative studies reveal difficulties in porting an existing network to a specific system and in predicting its performance indicators. Finding a common network architecture that is suited for all target platforms and at the same time yields decent results is a major challenge. In this contribution, we show that a winner-takes-all inspired network structure can be employed to solve Sudoku puzzles on three diverse hardware accelerators. By exploring several network implementations, we measured the number of solved puzzles in a set of 100 assorted Sudokus, as well as time and energy to solution. Concerning the last two indicators, our measurements indicate that it can be beneficial to port a network to an analogue hardware system

Bidirectional UWB Localization: A Review on an Elastic Positioning Scheme for GNSS-deprived Zones

A bidirectional Ultra-Wideband (UWB) localization scheme is one of the three widely deployed design integration processes ordinarily destined for time-based UWB positioning systems. The key property of the bidirectional UWB localization is its ability to serve both the navigation and tracking assignments on-demand within a single localization scheme. Conventionally, the perspective of navigation and tracking in wireless localization systems is viewed distinctly as an individual system because different methodologies were required for the implementation process. The ability to flexibly or elastically combine two unique positioning perspectives (i.e., navigation and tracking) within a single scheme is a paradigm shift in the way location-based services are observed. Thus, this article addresses and pinpoints the potential of a bidirectional UWB localization scheme. Regarding this, the complete system model of the bidirectional UWB localization scheme was comprehensively described based on modular processes in this article. The demonstrative evaluation results based on two system integration processes as well as a SWOT (strengths, weaknesses, opportunities, and threats) analysis of the scheme were also discussed. Moreover, we argued that the presented bidirectional scheme can also be used as a prospective topology for the realization of precise location estimation processes in 5G/6G wireless mobile networks, as well as Wi-Fi fine-time measurement-based positioning systems in this article.Comment: 30 pages, 12 figure

Compiler-Driven Reconfiguration of Multiprocessors

Hussmann M, Thies M, Kastens U, Purnaprajna M, Porrmann M, Rückert U. Compiler-Driven Reconfiguration of Multiprocessors. In: Proceedings of the Workshop on Application Specific Processors (WASP) 2007. 2007.Multiprocessors enable parallel execution of a single large application to achieve a performance improvement. An application is split at instruction, data or task level (based on the granularity), such that the overhead of partitioning is minimal. Parallelization for multiprocessors is mostly restricted to a fixed granularity. Reconfiguration enables architectural variations to allow multiple granularities of operation within a multiprocessor. This adaptability optimizes resource utilization over a fixed organization. Here, a unified hardware-software approach to design a reconfigurable multiprocessor system called QuadroCore is presented. In our holistic methodology, compiler-driven reconfiguration selects from a fixed set of modes. Each mode relies on matching program analysis to exploit the architecture efficiently. For instance, a multiprocessor may adapt to different parallelization paradigms. The compiler can determine the best execution mode for each piece of code by analyzing the parallelism in a program. A fast, singlecycle, run-time reconfiguration between these predetermined modes is enabled by executing special instructions which switch coarse-grained components like instruction decoders, ALUs and register banks. Performance is evaluated in terms of execution cycles and achieved clock frequency. First results indicate suitability especially in audio and video processing applications

Estimating the Security of Lattice-based Cryptosystems

Encryption and signature schemes based on worst-case lattice problems are promising candidates for the post-quantum era, where classic number-theoretic assumptions are rendered false. Although there have been many important results and breakthroughs in lattice cryptography, the questions of how to systematically evaluate their security in practice and how to choose secure parameters are still open. This is mainly due to the fact that most security proofs are essentially asymptotic statements. In addition, the hardness of the underlying complexity assumption is controlled by several interdependent parameters rather than just a simple bit length as in many classic schemes. With our work, we close this gap by providing a framework that (1) distills a hardness estimate out of a given parameter set and (2) relates the complexity of practical lattice-based attacks to symmetric bit security for the first time. Our approach takes various security levels, or attacker types, into account. Moreover, we use it to predict long-term security in a similar fashion as the results that are collected on www.keylength.com. In contrast to the experiments by Gama and Nguyen (Eurocrypt 2008), our estimates are based on precisely the family of lattices that is relevant in modern lattice-based cryptography. Our framework can be applied in two ways: Firstly, to assess the hardness of the (few) proposed parameter sets so far and secondly, to propose secure parameters in the first place. Our methodology is applicable to essentially all lattice-based schemes that are based on the learning with errors problem (LWE) or the small integer solution problem (SIS) and it allows us to compare efficiency and security across different schemes and even across different types of cryptographic primitives

Safeguarding the Evidential Value of Forensic Cryptocurrency Investigations

Analyzing cryptocurrency payment flows has become a key forensic method in law enforcement and is nowadays used to investigate a wide spectrum of criminal activities. However, despite its widespread adoption, the evidential value of obtained findings in court is still largely unclear. In this paper, we focus on the key ingredients of modern cryptocurrency analytics techniques, which are clustering heuristics and attribution tags. We identify internationally accepted standards and rules for substantiating suspicions and providing evidence in court and project them onto current cryptocurrency forensics practices. By providing an empirical analysis of CoinJoin transactions, we illustrate possible sources of misinterpretation in algorithmic clustering heuristics. Eventually, we derive a set of legal key requirements and translate them into a technical data sharing framework that fosters compliance with existing legal and technical standards in the realm of cryptocurrency forensics. Integrating the proposed framework in modern cryptocurrency analytics tools could allow more efficient and effective investigations, while safeguarding the evidential value of the analysis and the fundamental rights of affected persons