ALBERT

Description: Approximately 25% of childhood acute lymphoblastic leukemias carry the ETV6/RUNX1 fusion gene. Despite their excellent initial treatment response, up to 20% of patients relapse. To gain insight into the relapse mechanisms, we analyzed single nucleotide polymorphism arrays for DNA copy number aberrations (CNAs) in 18 matched diagnosis and relapse leukemias. CNAs were more abundant at relapse than at diagnosis (mean 12.5 vs 7.5 per case; P = .01) with 5.3 shared on average. Their patterns revealed a direct clonal relationship with exclusively new aberrations at relapse in only 21.4%, whereas 78.6% shared a common ancestor and subsequently acquired distinct CNA. Moreover, we identified recurrent, mainly nonoverlapping deletions associated with glucocorticoid-mediated apoptosis targeting the Bcl2 modifying factor (BMF) (n = 3), glucocorticoid receptor NR3C1 (n = 4), and components of the mismatch repair pathways (n = 3). Fluorescence in situ hybridization screening of additional 24 relapsed and 72 nonrelapsed ETV6/RUNX1-positive cases demonstrated that BMF deletions were significantly more common in relapse cases (16.6% vs 2.8%; P = .02). Unlike BMF deletions, which were always already present at diagnosis, NR3C1 and mismatch repair aberrations prevailed at relapse. They were all associated with leukemias, which poorly responded to treatment. These findings implicate glucocorticoid-associated drug resistance in ETV6/RUNX1-positive relapse pathogenesis and therefore might help to guide future therapies.

Description: Abstract 1214 Immune thrombocytopenia (ITP) is the most common acquired thrombocytopenia in children. Typically, external triggers as infections or vaccinations cause the rise of antibodies that crossreact with antigens expressed on the platelet surface. These anti-platelet antibodies are mostly directed against glycoprotein complexes GPIIb/IIIa or GPIb/IX/V, resulting in an increased turnover of antibody-decorated platelets which are then sequestered by the reticuloendothelial system. Recently, it has been suggested that thrombocytopenia might also be due to an insufficient platelet production as serum of some patients with ITP can impair the maturation of CD34+ hematopoietic stem cells to bone marrow megakaryocytes (MKs) in vitro or abrogate the formation of proplatelets in an in vitro culture system. The accelerated platelet turnover demands the generation of platelets de novo. Bone marrow smears often reveal normal or slightly increased MKs, although they seem to be smaller and of altered morphology. However, very little is known about the consequences of anti-platelet antibodies on bone marrow MKs in vivo and in situ. Here, we took advantage of a simple animal model of passive ITP by single or multiple intraperitoneal injections of an anti-GPIb antibody into mice. MKs were evaluated by multi-color immunofluorescence histology on whole femur sections in a modified staining procedure that bypasses decalcification. MK numbers on day 3 were doubled in response to a single injection and tripled on day 8 when mice were injected additionally on day 3 and 7. In these mice platelet counts were up to 2000/nL on day 10, indicating the power to produce platelets. MK area per section was transiently upregulated on day 3 in single injected mice and quadrupled after multiple injections on day 8 before shrinking below norm on day 14. Staining with an anti-rat IgG antibody showed that the antibody was present on MKs within the bone marrow several hours to days after injection. The signal was present for 5 days and no antibody was detected on day 7. MKs had an overall normal morphology and showed no signs of apoptosis or DNA blebbing. All MKs analyzed were negative for TdT in a classical TUNEL assay, indicating that there were no single strand breaks. As platelet counts rose markedly while the antibody was still present on the MK surface, we sought to identify whether the pool of MKs is expanded or formed de novo. To address this, mice where fed with nucleotide analogue EdU for up to 12 days and femur sections stained with Click-It-647 reagent to stain for newly incorporated DNA while mice were treated with anti-platelet antibody or isotype control. We found EdU-positive MKs after 12 days in control isotype-injected mice indicating the de novo formation from hematopoietic stem cells. In antibody-injected mice, newly formed MKs were negative or stained weakly for EdU on day 12, suggesting that they arise partially from an existing pool of progenitors. Finally, we analyzed platelet formation in vivo by imaging of the cranial bone marrow of GPIIb-eYFP-heterozygous mice. The depletion antibody was labeled with Atto-590-fluorophore and injected hours before imaging. Vasculature was counterstained by Quantum dots. We found that MKs residing at the bone marrow were decorated with the antibody and released pre- and proplatelets into the vasculature, indicating that platelet biogenesis can occur in the presence of anti-platelet antibodies on MKs. Our data thus provide novel insight into the pathomechanism of platelet production in patients with ITP. Disclosures: No relevant conflicts of interest to declare.

Description: Acute lymphoblastic leukemia (ALL) is the most common malignancy in childhood. While improved multi-agent chemotherapy regimens with individualized risk stratification have led to increased survival rates of approximately 80 percent, 20 percent of patients respond poorly to therapy or relapse. Therefore, novel therapeutic avenues are urgently needed to improve treatment outcome, overcome resistance and reduce side effects. Failure to undergo cell death represents a key survival mechanism of cancer cells and results in drug resistance and clonal escape. Since inhibitor of apoptosis proteins (IAPs) are often overexpressed in malignant cells and their overexpression correlates with inferior survival rates, they provide an attractive molecular target for therapeutic intervention. Small molecule inhibitors have been developed that act as SMAC mimetics (SMs) to counteract the cell death inhibitory function of IAPs. SMs can activate and/or modulate cell death pathways, and are currently being evaluated in clinical trials. Their successful therapeutic implementation requires identification of patients who could benefit from a SM-based treatment regimen ideally before start of therapy. Here, we analyzed the intrinsic activity of two monovalent (AT406 and LCL161) and two bivalent (Birinapant or BV6) SMs on 29 unselected patient-derived pediatric precursor B-cell (BCP)-ALL samples and identified a subset of BCP-ALL primografts to be sensitive to SM treatment (n=8). When we compared gene expression of SM-sensitive (n=8) and SM-insensitive (n=6) patient-derived BCP-ALL samples, we identified a characteristic gene expression signature with 127 differentially regulated genes, amongst them upregulation of TNFRSF1A (TNFR1) in the SM-sensitive subset. In line with previous reports, we confirmed a critical role of the TNF/TNFR1-axis for SM-induced cell death in BCP-ALL by functional analysis. Expression of TNFRSF1A alone, however, did not correlate with sensitivity to SM-induced cell death indicating that TNFR1 is not the only factor regulating cell fate decisions in response to SM treatment. To identify potential biomarker genes for prediction of patient response to SM monotherapy in BCP-ALL, we compared differentially regulated genes of SM responders and non-responders from our cohort with data from a published cohort. Interestingly, we found 4 genes to overlap between these two cohorts. Of these 4 genes TSPAN7, FAM69C, and TNFRSF1A were upregulated whereas MTX2 was downregulated in SM-sensitive samples. The signature identified may reflect a particular TNF network. Analysis of expression levels of these 4 genes in BCP-ALL cell lines (Nalm6, Reh, UoCB6 and RS4;11) revealed that Reh cells, sensitive to SM-induced cell death, exhibited the biomarker profile of primograft sensitivity, i.e. upregulation of TSPAN7, FAM69C, TNFRSF1A and downregulation of MTX2. Nalm6 cells resembled the expression pattern of SM-insensitive samples with a downregulation of TSPAN7, FAM69C, TNFRSF1A and an upregulation of MTX2 and were resistant to SM-induced cell death. RS4;11 and UoCB6 cells showed no pattern. Based on these findings we hypothesized that the respective expression patterns of TSPAN7, FAM69C, TNFRSF1A and MTX2 could predict sensitivity to SMs. An extended screen of additional primary BCP-ALL samples for their expression levels of TSPAN7, FAM69C, TNFRSF1A and MTX2 and response to SMs substantiated this hypothesis. In summary, the subset of primary BCP-ALL samples with sensitivity to SMs is characterized by a gene signature with MTX2 low and TSPAN7, FAM69C and TNFRSF1A high. By using this expression profile, sensitivity to SMs in BCP-ALL could be identified in cell lines and additional primografts. Based on these results, we suggest the identified gene expression pattern as a biomarker for selecting patients to be treated by SM monotherapy in clinical trials. Disclosures No relevant conflicts of interest to declare.

Description: Eastern boundary upwelling systems (EBUS) are among the most productive marine ecosystems on Earth. The production of organic material is fueled by upwelling of nutrient-rich deep waters and high incident light at the sea surface. However, biotic and abiotic factors can modify surface production and related biogeochemical processes. Determining these factors is important because EBUS are considered hotspots of climate change, and reliable predictions of their future functioning requires understanding of the mechanisms driving the biogeochemical cycles therein. In this field experiment, we used in situ mesocosms as tools to improve our mechanistic understanding of processes controlling organic matter cycling in the coastal Peruvian upwelling system. Eight mesocosms, each with a volume of ∼55 m3, were deployed for 50 d ∼6 km off Callao (12∘ S) during austral summer 2017, coinciding with a coastal El Niño phase. After mesocosm deployment, we collected subsurface waters at two different locations in the regional oxygen minimum zone (OMZ) and injected these into four mesocosms (mixing ratio ≈1.5 : 1 mesocosm: OMZ water). The focus of this paper is on temporal developments of organic matter production, export, and stoichiometry in the individual mesocosms. The mesocosm phytoplankton communities were initially dominated by diatoms but shifted towards a pronounced dominance of the mixotrophic dinoflagellate (Akashiwo sanguinea) when inorganic nitrogen was exhausted in surface layers. The community shift coincided with a short-term increase in production during the A. sanguinea bloom, which left a pronounced imprint on organic matter C : N : P stoichiometry. However, C, N, and P export fluxes did not increase because A. sanguinea persisted in the water column and did not sink out during the experiment. Accordingly, export fluxes during the study were decoupled from surface production and sustained by the remaining plankton community. Overall, biogeochemical pools and fluxes were surprisingly constant for most of the experiment. We explain this constancy by light limitation through self-shading by phytoplankton and by inorganic nitrogen limitation which constrained phytoplankton growth. Thus, gain and loss processes remained balanced and there were few opportunities for blooms, which represents an event where the system becomes unbalanced. Overall, our mesocosm study revealed some key links between ecological and biogeochemical processes for one of the most economically important regions in the oceans.

Description: In this paper we present a method to detect airflow through ice caves and to quantify the corresponding airflow speeds by the use of temperature loggers. The time series of temperature observations at different loggers are cross-correlated. The time shift of best correlation corresponds to the travel time of the air and is used to derive the airflow speed between the loggers. We apply the method to test data observed inside Schellenberger Eishöhle (ice cave). The successful determination of airflow speeds depends on the existence of distinct temperature variations during the time span of interest. Moreover the airflow speed is assumed to be constant during the period used for the correlation analysis. Both requirements limit the applicability of the correlation analysis to determine instantaneous airflow speeds. Nevertheless the method is very helpful to characterize the general patterns of air movement and their slow temporal variations. The correlation analysis assumes a linear dependency between the correlated data. The good correlation we found for our test data confirms this assumption. We therefore in a second step estimate temperature biases and scale factors for the observed temperature variations by a least squares adjustment. The observed phenomena, a warming and a damping of temperature variations depending on the distance the air traveled inside the cave, are explained by a mixing of the inflowing air with the air inside the cave. Furthermore we test the significance of the determined parameters by a standard F test and study the sensitivity of the procedure to common manipulations of the original observations like smoothing. In the end we will give an outlook on possible applications and further development of this method.

Description: In this paper we present a method to detect airflow through ice caves and to quantify the corresponding airflow speeds by the use of temperature loggers. The time series of temperature observations at different loggers are cross-correlated. The time shift of best correlation corresponds to the travel time of the air and is used to derive the airflow speed between the loggers. We apply the method to test data observed inside Schellenberger Eishöhle (ice cave). The successful determination of airflow speeds depends on the existence of distinct temperature variations during the time span of interest. Moreover the airflow speed is assumed to be constant during the period used for the correlation analysis. Both requirements limit the applicability of the correlation analysis to determine instantaneous airflow speeds. Nevertheless the method is very helpful to characterize the general patterns of air movement and their slow temporal variations. The correlation analysis assumes a linear dependency between the correlated data. The good correlation we found for our test data confirms this assumption. We therefore in a second step estimate temperature biases and scale factors for the observed temperature variations by a least-squares adjustment. The observed phenomena, a warming and an attenuation of temperature variations, depending on the distance the air traveled inside the cave, are explained by a mixing of the inflowing air with the air inside the cave. Furthermore we test the significance of the determined parameters by a standard F test and study the sensitivity of the procedure to common manipulations of the original observations like smoothing. In the end we will give an outlook on possible applications and further development of this method.

Description: In 2003 sediment core Lz1024 was drilled at Lake El'gygytgyn, far east Russian Arctic, in an area of the Northern Hemisphere which has not been glaciated for the last 3.6 Ma. Biogenic silica was used for analysing the oxygen isotope composition (δ18Odiatom) in the upper 13 m long section dating back about 250 ka with samples dominated by one taxa in the

Description: In 2003 sediment core Lz1024 was drilled at Lake El'gygytgyn, far east Russian Arctic, in an area of the Northern Hemisphere which has not been glaciated for the last 3.6 Ma. Biogenic silica was used for analysing the oxygen isotope composition (δ18Odiatom) in the upper 13 m long section dating back about 250 ka with samples dominated by one taxa in the

Description: More than 300 non-dispersive infrared (NDIR) CO2 low-cost sensors labelled as LP8 were integrated into sensor units and evaluated for the purpose of long-term operation in the Carbosense CO2 sensor network in Switzerland. Prior to deployment, all sensors were calibrated in a pressure and climate chamber and in ambient conditions co-located with a reference instrument. To investigate their long-term performance and to test different data processing strategies, 18 sensors were deployed at five locations equipped with a reference instrument after calibration. Their accuracy during 19 to 25 months deployment was between 8 and 12 ppm. This level of accuracy requires careful sensor calibration prior to deployment, continuous monitoring of the sensors, efficient data filtering, and a procedure to correct drifts and jumps in the sensor signal during operation. High relative humidity (〉 ∼85 %) impairs the LP8 measurements, and corresponding data filtering results in a significant loss during humid conditions. The LP8 sensors are not suitable for the detection of small regional gradients and long-term trends. However, with careful data processing, the sensors are able to resolve CO2 changes and differences with a magnitude larger than about 30 ppm. Thereby, the sensor can resolve the site-specific CO2 signal at most locations in Switzerland. A low-power network (LPN) using LoRaWAN allowed for reliable data transmission with low energy consumption and proved to be a key element of the Carbosense low-cost sensor network.

Description: Snow densification stores water in alpine regions and transforms snow into ice on the surface of glaciers. Despite its importance in determining snow-water equivalent and glacier-induced sea level rise, we still lack a complete understanding of the physical mechanisms underlying snow compaction. In essence, compaction is a rheological process, where the rheology evolves with depth due to variation in temperature, pressure, humidity, and meltwater. The rheology of snow compaction can be determined in a few ways, for example, through empirical investigations (e.g., Herron and Langway, 1980), by microstructural considerations (e.g., Alley, 1987), or by measuring the rheology directly, which is the approach we take here. Using a French-press or cafetière-à-piston compression stage, Wang and Baker (2013) compressed numerous snow samples of different densities. Here we derive a mixture theory for compaction and airflow through the porous snow to compare against these experimental data. We find that a plastic compaction law explains experimental results. Taking standard forms for the permeability and effective pressure as functions of the porosity, we show that this compaction mode persists for a range of densities and overburden loads. These findings suggest that measuring compaction in the lab is a promising direction for determining the rheology of snow through its many stages of densification.