A COVID-19 peptide vaccine for the induction of SARS-CoV-2 T cell immunity.

T cell immunity is central for the control of viral infections. CoVac-1 is a peptide-based vaccine candidate, composed of SARS-CoV-2 T cell epitopes derived from various viral proteins1,2, combined with the Toll-like receptor 1/2 agonist XS15 emulsified in Montanide ISA51 VG, aiming to induce profound SARS-CoV-2 T cell immunity to combat COVID-19. Here we conducted a phase I open-label trial, recruiting 36 participants aged 18-80 years, who received a single subcutaneous CoVac-1 vaccination. The primary end point was safety analysed until day 56. Immunogenicity in terms of CoVac-1-induced T cell response was analysed as the main secondary end point until day 28 and in the follow-up until month 3. No serious adverse events and no grade 4 adverse events were observed. Expected local granuloma formation was observed in all study participants, whereas systemic reactogenicity was absent or mild. SARS-CoV-2-specific T cell responses targeting multiple vaccine peptides were induced in all study participants, mediated by multifunctional T helper 1 CD4+ and CD8+ T cells. CoVac-1-induced IFNγ T cell responses persisted in the follow-up analyses and surpassed those detected after SARS-CoV-2 infection as well as after vaccination with approved vaccines. Furthermore, vaccine-induced T cell responses were unaffected by current SARS-CoV-2 variants of concern. Together, CoVac-1 showed a favourable safety profile and induced broad, potent and variant of concern-independent T cell responses, supporting the presently ongoing evaluation in a phase II trial for patients with B cell or antibody deficiency.

IRF2BP2 counteracts the ATF7/JDP2 AP-1 heterodimer to prevent inflammatory overactivation in acute myeloid leukemia (AML) cells.

Acute myeloid leukemia (AML) is a hematological malignancy characterized by abnormal proliferation and accumulation of immature myeloid cells in the bone marrow. Inflammation plays a crucial role in AML progression, but excessive activation of cell-intrinsic inflammatory pathways can also trigger cell death. IRF2BP2 is a chromatin regulator implicated in AML pathogenesis, although its precise role in this disease is not fully understood. In this study, we demonstrate that IRF2BP2 interacts with the AP-1 heterodimer ATF7/JDP2, which is involved in activating inflammatory pathways in AML cells. We show that IRF2BP2 is recruited by the ATF7/JDP2 dimer to chromatin and counteracts its gene-activating function. Loss of IRF2BP2 leads to overactivation of inflammatory pathways, resulting in strongly reduced proliferation. Our research indicates that a precise equilibrium between activating and repressive transcriptional mechanisms creates a pro-oncogenic inflammatory environment in AML cells. The ATF7/JDP2-IRF2BP2 regulatory axis is likely a key regulator of this process and may, therefore, represent a promising therapeutic vulnerability for AML. Thus, our study provides new insights into the molecular mechanisms underlying AML pathogenesis and identifies a potential therapeutic target for AML treatment.

The histone acetyltransferase KAT6A is recruited to unmethylated CpG islands via a DNA binding winged helix domain.

The lysine acetyltransferase KAT6A (MOZ, MYST3) belongs to the MYST family of chromatin regulators, facilitating histone acetylation. Dysregulation of KAT6A has been implicated in developmental syndromes and the onset of acute myeloid leukemia (AML). Previous work suggests that KAT6A is recruited to its genomic targets by a combinatorial function of histone binding PHD fingers, transcription factors and chromatin binding interaction partners. Here, we demonstrate that a winged helix (WH) domain at the very N-terminus of KAT6A specifically interacts with unmethylated CpG motifs. This DNA binding function leads to the association of KAT6A with unmethylated CpG islands (CGIs) genome-wide. Mutation of the essential amino acids for DNA binding completely abrogates the enrichment of KAT6A at CGIs. In contrast, deletion of a second WH domain or the histone tail binding PHD fingers only subtly influences the binding of KAT6A to CGIs. Overexpression of a KAT6A WH1 mutant has a dominant negative effect on H3K9 histone acetylation, which is comparable to the effects upon overexpression of a KAT6A HAT domain mutant. Taken together, our work revealed a previously unrecognized chromatin recruitment mechanism of KAT6A, offering a new perspective on the role of KAT6A in gene regulation and human diseases.

Potent drug delivery enhancement of betulinic acid and NVX-207 into equine skin in vitro - a comparison between a novel oxygen flow-assisted transdermal application device and microemulsion gels.

BACKGROUND: Gray horses are predisposed to equine malignant melanoma (EMM) with advancing age. Depending on the tumor's location and size, they can cause severe problems (e.g., defaecation, urination, feeding). A feasible therapy for EMM has not yet been established and surgical excision can be difficult depending on the location of the melanoma. Thus, an effective and safe therapy is needed. Naturally occurring betulinic acid (BA), a pentacyclic triterpene and its synthetic derivate, NVX-207 (3-acetyl-betulinic acid-2-amino-3-hydroxy-2-hydroxymethyl-propanoate) are known for their cytotoxic properties against melanomas and other tumors and have already shown good safety and tolerability in vivo. In this study, BA and NVX-207 were tested for their permeation potential into equine skin in vitro in Franz-type diffusion cell (FDC) experiments after incubation of 5 min, 30 min and 24 h, aiming to use these formulations for prospective in vivo studies as a treatment for early melanoma stages. Potent permeation was defined as reaching or exceeding the half maximal inhibitory concentrations (IC50) of BA or NVX-207 for equine melanoma cells in equine skin samples. The active ingredients were either dissolved in a microemulsion (ME) or in a microemulsion gel (MEG). All of the formulations were transdermally applied but the oil-in-water microemulsion was administered with a novel oxygen flow-assisted (OFA) applicator (DERMADROP TDA). RESULTS: All tested formulations exceeded the IC50 values for equine melanoma cells for BA and NVX-207 in equine skin samples, independently of the incubation time NVX-207 applied with the OFA applicator showed a significant time-dependent accumulation and depot-effect in the skin after 30 min and 24 h (P < 0.05). CONCLUSIONS: All tested substances showed promising results. Additionally, OFA administration showed a significant accumulation of NVX-207 after 30 min and 24 h of incubation. Further in vivo trials with OFA application are recommended.

Non-equilibrium structural dynamics of supercoiled DNA plasmids exhibits asymmetrical relaxation.

Many cellular processes occur out of equilibrium. This includes site-specific unwinding in supercoiled DNA, which may play an important role in gene regulation. Here, we use the Convex Lens-induced Confinement (CLiC) single-molecule microscopy platform to study these processes with high-throughput and without artificial constraints on molecular structures or interactions. We use two model DNA plasmid systems, pFLIP-FUSE and pUC19, to study the dynamics of supercoiling-induced secondary structural transitions after perturbations away from equilibrium. We find that structural transitions can be slow, leading to long-lived structural states whose kinetics depend on the duration and direction of perturbation. Our findings highlight the importance of out-of-equilibrium studies when characterizing the complex structural dynamics of DNA and understanding the mechanisms of gene regulation.

Knife wound or nosebleed-where does the blood at the crime scene come from?

Secretion analysis is a useful tool in forensic genetics, since it establishes the (cellular) origin of the DNA prior in addition to the identification of the DNA donor. This information can be crucial for the construction of the crime sequence or verification of statements of people involved in the crime. For some secretions, rapid/pretests already exist (blood, semen, urine, and saliva) or can be determined via published methylation analyses or expression analyses (blood, saliva vaginal secretions, menstrual blood, and semen). To discriminate nasal secretion/blood from other secretions (like oral mucosa/saliva, blood, vaginal secretion, menstrual blood, and seminal fluid), assays based on specific methylation patterns at several CpGs were set up in this study. Out of an initial 54 different CpG markers tested, two markers showed a specific methylation value for nasal samples: N21 and N27 with a methylation mean value of 64.4% ± 17.6% and 33.2% ± 8.7%, respectively. Although identification or discrimination was not possible for all nasal samples (due to partial overlap in methylation values to other secretions), 63% and 26% of the nasal samples could be unambiguously identified and distinguished from the other secretions using the CpG marker N21 and N27, respectively. In combination with a blood pretest/rapid test, a third marker (N10) was able to detect nasal cells in 53% of samples. Moreover, the employment of this pretest increases the proportion of identifiable or discriminable nasal secretion samples using marker N27 to 68%. In summary, our CpG assays proved to be promising tools in forensic analysis for the detection of nasal cells in samples from a crime scene.

Medication errors in cancer therapy: Reports from German hospital pharmacists between 2008 and 2019.

OBJECTIVE: Since medication errors can have severe consequences, the development of methods to improve patient safety is becoming increasingly important. The aim of this evaluation was to identify frequent medication errors in oncology as well as characteristic correlations in the various error patterns. In addition, the implementation rate of the proposed pharmaceutical intervention was determined in order to assess the benefit of a clinical pharmacist in the field of oncology. METHODS: The evaluation was based on a data-set from a national documentation system for medication errors and interventions (DokuPIK) used by hospital pharmacists in the field of oncology from 2008 to 2019, namely 6684 reported cases in oncology, representing about 5% of all reports in DokuPIK. RESULTS: The most frequently reported errors were incorrect doses (22% of reported errors), followed by interactions (14%); in 10% of errors the prescription/documentation was incomplete/incorrect. The intervention suggested by the pharmacist was implemented in 97% of the cases. Based on the respective Anatomical Therapeutical Chemical Classification (ATC codes), drugs (or groups of drugs) were identified that were reported frequently in connection with medication errors, namely carboplatin and cyclophosphamide as anticancer drugs pantoprazole as non-anticancer drug. CONCLUSION: Frequently occurring medication errors in the field of oncology were identified, facilitating the development of specific recommendations for action and prevention strategies. The implementation of an electronic prescription software is particularly recommended for the avoidance of dosage errors in chemotherapy.

Safety of a Novel Dialyzer Containing a Fluorinated Polyurethane Surface-Modifying Macromolecule in Patients with End-Stage Kidney Disease.

BACKGROUND: By inhibiting the adsorption of protein and platelets, surface-modifying macromolecules (SMMs) may improve the hemocompatibility of hemodialyzers. This trial aims to assess the performance and safety of a novel dialyzer with a fluorinated polyurethane SMM, Endexo™. METHODS: This prospective, sequential, multicenter, open-label study (NCT03536663) was designed to meet regulatory requirements for clinical testing of new hemodialyzers, including assessment of the in vivo ultrafiltration coefficient (Kuf). Adults prescribed thrice-weekly hemodialysis were eligible for enrollment. After completing 12 hemodialysis sessions with an Optiflux® F160NR dialyzer, patients received 38 sessions with the dialyzer with Endexo. Evaluated parameters included the in vivo Kuf of the dialyzer with Endexo extent of removal of urea, albumin, and ß2-microglobulin (ß2M), as well as complement activation. RESULTS: Twenty-three patients received 268 hemodialysis treatments during the Optiflux period, and 18 patients received 664 hemodialysis treatments during the Endexo period. Three serious adverse events were reported, and none of them were considered device related. No overt complement activation was observed with either dialyzer. Both dialyzers were associated with comparable mean increases in serum albumin levels from pre- to posthemodialysis (Optiflux: 7.9%; Endexo: 8.0%). These increases can be viewed in the context of a mean increase in hemoglobin of approximately 5% and a mean ultrafiltration volume removed of approximately 2.2 L. The corrected mean ß2M removal rate was 47% higher during the Endexo period (67.73%). Mean treatment times (208 vs. 205 min), blood flow rates (447.7 vs. 447.5 mL/min), dialysate flow rates (698.5 vs. 698.0 mL/min), urea reduction ratio (82 vs. 81%), and spKt/V (2.1 vs. 1.9) were comparable for the Endexo and Optiflux periods, respectively. The mean (SD) Kuf was 15.85 (10.33) mL/h/mm Hg during the first use of the dialyzer with Endexo (primary endpoint) and 16.36 (9.92) mL/h/mm Hg across the Endexo period. CONCLUSIONS: The safety of the novel dialyzer with Endexo was generally comparable to the Optiflux dialyzer, while exhibiting a higher ß2M removal rate.

The effects of examiner fatigue on the diagnostic accuracy of dental radiographs.

OBJECTIVES: The aim of the study was to find out whether and to what extent the performance of dentists regarding diagnostic evaluation of dental radiographs is influenced by symptoms of fatigue. MATERIALS AND METHODS: Over a period of 40 minutes, 21 dentists evaluated a database of 96 randomly selected, infinitely repeating intraoral dental radiographs for the presence of periapical radiolucencies. Both before and after, participants were asked to assess their subjective fatigue using the Swedish Occupational Fatigue Inventory (SOFI), Visual Analogue Scale (VAS), and Numerical Rating Scale (NRS). Diagnostic accuracy was analyzed using the Receiver Operating Characteristics (ROC) method. Furthermore, the correlation between diagnostic accuracy and radiographic experience, image viewing time, and level of training was also evaluated. RESULTS: The study showed that despite increasing fatigue, the diagnostic accuracy of the examiners remained consistent with an average AUC value of 0.768 ± 0.091. Within the 40-min reporting period, no statistically significant fluctuations were found. The diagnostic accuracy varied depending on the radiographic experience: with many years of radiographic experience, the diagnostic accuracy increased. At the same time, the older study participants with greater radiographic experience became less tired compared to younger study participants during the examination. CONCLUSIONS: Although an increase in fatigue was observed during the 40-min examination, the diagnostic accuracy of the doctors remained constant. CLINICAL RELEVANCE: Due to the high workload which needs to be handled in a limited time, medical and dental professionals have reached a certain level of exposition to stress that can lead to physical fatigue. However, the increasing fatigue should not negatively influence the work of the doctors. The study shows that the radiodiagnostic accuracy remained the same.

Concentration profiles and safety of topically applied betulinic acid and NVX-207 in eight healthy horses-A randomized, blinded, placebo-controlled, crossover pilot study.

The naturally occurring betulinic acid (BA) and its derivative NVX-207 show anticancer effects against equine malignant melanoma (EMM) cells and a potent permeation in isolated equine skin in vitro. The aim of the study was to determine the in vivo concentration profiles of BA and NVX-207 in equine skin and assess the compounds' local and systemic tolerability with the intent of developing a topical therapy against EMM. Eight horses were treated percutaneously in a crossover design with 1% BA, 1% NVX-207 or a placebo in a respective vehicle twice a day for seven consecutive days with a seven-day washout period between each formulation. Horses were treated at the neck and underneath the tail. Concentration profiles of the compounds were assessed by high-performance liquid chromatography in the cervical skin. Clinical and histopathological examinations and blood analyses were performed. Higher concentrations of NVX-207 were found in the skin compared to BA. Good systemic tolerability and only mild local adverse effects were observed in all three groups. This study substantiates the topical application of BA and NVX-207 in further clinical trials with horses suffering from EMM; however, penetration and permeation of the compounds may be altered in skin affected by tumors.

Betulinic acid shows anticancer activity against equine melanoma cells and permeates isolated equine skin in vitro.

BACKGROUND: Equine malignant melanoma (EMM) is a frequently occurring dermoepidermal tumor in grey horses. Currently available therapies are either challenging or inefficient. Betulinic acid (BA), a naturally occurring triterpenoid, is a promising compound for cancer treatment. To evaluate the potential of BA as a topical therapy for EMM, its anticancer effects on primary equine melanoma cells and dermal fibroblasts and its percutaneous permeation through isolated equine skin were assessed in vitro. RESULTS: BA showed antiproliferative and cytotoxic effects on both primary equine melanoma cells and fibroblasts in a time- and dose-dependent manner. The lowest half-maximal inhibitory concentrations were obtained 96 h after the beginning of drug exposure (12.7 µmol/L and 23.6 µmol/L for melanoma cells eRGO1 and MelDuWi, respectively, in cytotoxicity assay). High concentrations of the compound were reached in the required skin layers in vitro. CONCLUSION: BA is a promising substance for topical EMM treatment. Further clinical studies in horses are necessary to assess safety and antitumoral effects in vivo.

[Blessing or curse? The World Wide Web as information source for autism and Asperger Syndrome]. / Segen oder Fluch? Das Internet als Informationsquelle über Autismus und Asperger-Syndrom.

Blessing or curse? The World Wide Web as information source for autism and Asperger Syndrome Abstract. Objective: The World Wide Web is today one of the most common methods used for obtaining health-related information, though the quality of the information is sometimes questionable. The present study addresses the quality of the information source internet and the resulting implications and discusses examples related to autism spectrum disorder. Method: We systematically evaluated 96 German websites, with the aim of estimating specific characteristic features, reliability of publications, presentation of information as well as overall website quality. We also analyzed the clinical implications of the presentations. Results: Only 39 % of the websites provided references to scientifically well-founded information, whereas advertisements were found on 53 % of websites. The greatest percentage of false information (17 %) was disseminated concerning therapy. 75 % of the websites provided incomplete information. Only 10 % of websites discussed the impairment or familial burden. The quality of information was insufficient on 30 %, poor on 41 %, and good on only 6 % of the websites. Conclusions: Similar to results available for English-language websites, the quality of German websites providing health-related information can be considered low. Implications concerning confirmation bias, stigma, overidentification, in-group/outgroup, contrast and snowball effects are discussed.

Identification of gene regulation models from single-cell data.

In quantitative analyses of biological processes, one may use many different scales of models (e.g. spatial or non-spatial, deterministic or stochastic, time-varying or at steady-state) or many different approaches to match models to experimental data (e.g. model fitting or parameter uncertainty/sloppiness quantification with different experiment designs). These different analyses can lead to surprisingly different results, even when applied to the same data and the same model. We use a simplified gene regulation model to illustrate many of these concerns, especially for ODE analyses of deterministic processes, chemical master equation and finite state projection analyses of heterogeneous processes, and stochastic simulations. For each analysis, we employ Matlab and Python software to consider a time-dependent input signal (e.g. a kinase nuclear translocation) and several model hypotheses, along with simulated single-cell data. We illustrate different approaches (e.g. deterministic and stochastic) to identify the mechanisms and parameters of the same model from the same simulated data. For each approach, we explore how uncertainty in parameter space varies with respect to the chosen analysis approach or specific experiment design. We conclude with a discussion of how our simulated results relate to the integration of experimental and computational investigations to explore signal-activated gene expression models in yeast (Neuert et al 2013 Science 339 584-7) and human cells (Senecal et al 2014 Cell Rep. 8 75-83)5.

Impact of formaldehyde, acetaldehyde, and N-(3-(Dimethylamino)propyl)methacrylamide on the efficacy of the human derived coagulation factor IX.

Polymer-borne leachables such as formaldehyde, acetaldehyde, and N-3-(Dimethylamino)propyl)methacrylamide (DMAPMA) may interact with therapeutic proteins. In this study, the leachables were spiked into human derived coagulation factor IX (FIX) at concentrations of 1, 10, 50, 100, and 500 µg/mL, corresponding to a leachable - FIX ratio of 0.5, 5, 25, 50 and 250 %, respectively. The spiked samples were visually inspected, and pH was measured. No visual effects were observed, and pH was within the drug product's specified range. Recovery experiments were performed and no loss of leachables was identified. Protein structure analysis revealed that formaldehyde reacted with lysine contained in two different positions of FIX, in a concentration-dependent manner starting at 10 µg/mL (5 %). The clotting activity of FIX was measured. The activity of the samples spiked with 500 µg/mL (250 %) of formaldehyde dropped by more than half. The activity of the samples spiked with acetaldehyde began to drop at 50 µg/mL (25 %) and continued to decline in concentration-dependent manner. DMAPMA did not impair the activity of FIX. The findings conclude that depending on the concentration, some leachables may react with or modify therapeutic proteins, potentially causing an undesired pharmacological effect however, this is specific to each protein.

Investigation of SAMD1 ablation in mice.

SAM domain-containing protein 1 (SAMD1) has been implicated in atherosclerosis, as well as in chromatin and transcriptional regulation, suggesting a versatile and complex biological function. However, its role at an organismal level is currently unknown. Here, we generated SAMD1-/- and SAMD1+/- mice to explore the role of SAMD1 during mouse embryogenesis. Homozygous loss of SAMD1 was embryonic lethal, with no living animals seen after embryonic day 18.5. At embryonic day 14.5, organs were degrading and/or incompletely developed, and no functional blood vessels were observed, suggesting failed blood vessel maturation. Sparse red blood cells were scattered and pooled, primarily near the embryo surface. Some embryos had malformed heads and brains at embryonic day 15.5. In vitro, SAMD1 absence impaired neuronal differentiation processes. Heterozygous SAMD1 knockout mice underwent normal embryogenesis and were born alive. Postnatal genotyping showed a reduced ability of these mice to thrive, possibly due to altered steroidogenesis. In summary, the characterization of SAMD1 knockout mice suggests a critical role of SAMD1 during developmental processes in multiple organs and tissues.

Dose Recommendations for Common Drugs in Patients with Liver Cirrhosis: A Systematic Literature Review.

BACKGROUND AND OBJECTIVES: The presence of liver cirrhosis affects the selection and dosing of drugs metabolised by the liver as doses have to be adjusted to the remaining liver function. This is a major challenge in clinical practice as specific guidelines are lacking. The aim of this study was to identify drugs for which recommendations on selection and dose adjustments for patients with cirrhosis exist by assessing the literature according to certain quality standards, paying particular attention to the suitability of these recommendations for clinical practice. METHODS: A systematic literature review included peer-reviewed publications that were published by October 2020 in PubMed in the English language and aimed to generate recommendations on dose adjustment in patients with liver cirrhosis. Subsequently, the identified publications were checked for reporting quality against the relevant reporting guidelines and the Oxford Centre for Evidence-Based Medicine Levels of Evidence. Finally, all specific dose recommendations were extracted, compared with the specifications of the Summaries of Product Characteristics and mapped according to the Anatomical Therapeutic Chemical/Defined Daily Dose Index. RESULTS: Eighteen publications covering a total of 1145 dose recommendations for 481 active substances were identified. There were 706 recommendations for 316 substances sufficiently specific for application in clinical practice. For 22 active substances, the specific recommendations were consistent across multiple publications, of which only six were also consistent with the respective Summaries of Product Characteristics. CONCLUSIONS: As the majority of dose recommendations were not sufficiently specific or even contradictory, there is an urgent need for the definition of standard parameters for a uniform assessment of drugs in liver cirrhosis. In addition, dose recommendations should be aligned by suitable methods.

Understanding WT1 Alterations and Expression Profiles in Hematological Malignancies.

WT1 is a true chameleon, both acting as an oncogene and tumor suppressor. As its exact role in leukemogenesis is still ambiguous, research with model systems representing natural conditions surrounding the genetic alterations in WT1 is necessary. In a cohort of 59 leukemia/lymphoma cell lines, we showed aberrant expression for WT1 mRNA, which does not always translate into protein levels. We also analyzed the expression pattern of the four major WT1 protein isoforms in the cell lines and primary AML blasts with/without WT1 mutations and demonstrated that the presence of mutations does not influence these patterns. By introduction of key intronic and exonic sequences of WT1 into a lentiviral expression vector, we developed a unique tool that can stably overexpress the four WT1 isoforms at their naturally occurring tissue-dependent ratio. To develop better cellular model systems for WT1, we sequenced large parts of its gene locus and also other important myeloid risk factor genes and revealed previously unknown alterations. Functionally, inhibition of the nonsense-mediated mRNA decay machinery revealed that under natural conditions, the mutated WT1 alleles go through a robust degradation. These results offer new insights and model systems regarding the characteristics of WT1 in leukemia and lymphoma.

Peptide-mediated inhibition of the transcriptional regulator Elongin BC induces apoptosis in cancer cells.

Inhibition of protein-protein interactions (PPIs) via designed peptides is an effective strategy to perturb their biological functions. The Elongin BC heterodimer (ELOB/C) binds to a BC-box motif and is essential for cancer cell growth. Here, we report a peptide that mimics the high-affinity BC-box of the PRC2-associated protein EPOP. This peptide tightly binds to the ELOB/C dimer (kD = 0.46 ± 0.02 nM) and blocks the association of ELOB/C with its interaction partners, both in vitro and in the cellular environment. Cancer cells treated with our peptide inhibitor showed decreased cell viability, increased apoptosis, and perturbed gene expression. Therefore, our work proposes that blocking the BC-box-binding pocket of ELOB/C is a feasible strategy to impair its function and inhibit cancer cell growth. Our peptide inhibitor promises novel mechanistic insights into the biological function of the ELOB/C dimer and offers a starting point for therapeutics linked to ELOB/C dysfunction.

Earth beyond six of nine planetary boundaries.

This planetary boundaries framework update finds that six of the nine boundaries are transgressed, suggesting that Earth is now well outside of the safe operating space for humanity. Ocean acidification is close to being breached, while aerosol loading regionally exceeds the boundary. Stratospheric ozone levels have slightly recovered. The transgression level has increased for all boundaries earlier identified as overstepped. As primary production drives Earth system biosphere functions, human appropriation of net primary production is proposed as a control variable for functional biosphere integrity. This boundary is also transgressed. Earth system modeling of different levels of the transgression of the climate and land system change boundaries illustrates that these anthropogenic impacts on Earth system must be considered in a systemic context.