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Combinations of different medicinal products are widely use in the medical practice and are in fact essential for the treatment of several diseases. The existing regulation for approval of medicinal products offers only a remote possibility for the approval of combination therapies. Usually, active agents are evaluated in terms of their individual safety and efficacy. For exceptional cases, guidelines and regulations exist for the approval of specific combinations, such as fixed combinations, which includes two or more active agents within a single pharmaceutical form. The free combination of individual medications or other medical products is however currently not reflected in the legal framework, meaning a gap between treatment reality and regulatory process exists. A possible approach for closing this gap is the introduction of "therapeutic concepts" which is presented in this thesis. Therapeutic concepts describe the marketing authorisation of a treatment regime consisting of a combination of two or more individual medicinal products for a defined condition or a combination of a medicinal product and a companion device that is compulsory for diagnosis or decision-making, that have been developed and studied together. Therapeutic concepts can be regarded as an integrative approach based on the current framework. The approach would be an addition to the present legislation to meet existing needs. Compared to fixed combinations the free combination of a therapeutic concept would offer additional benefits, such as better dosage adjustment based on the individual patient's need. At present, combinations of medicinal products in certain conditions are for example described in medical guidelines. However, medical guidelines differ greatly in quality from each other, having the status of recommendation and cannot be equated with a market authorisation. Defined regulations for a market authorisation of combinations based on evidence obtained from clinical studies provide a greater knowledge and control on combinations in use and an improved legal certainty compared to medical guidelines. Combinations of different medicinal products have been commonly used as treatment systems, often in complex or multifactorial diseases, such as bacterial infection (e.g. tuberculosis), cardiovascular diseases or cancer. Current research focuses now on genetics-associated diseases, which also often require a complex combination of diagnostics and medicine. This field of research is referred to as personalized medicine. The patient's individual disease and metabolic markers are analysed to stratify patients into subgroups which receive a therapy based on their genetic profile that is more likely to be effective compared to an alternative medication. Due to the complexity of the treatment approach and the involvement of both medicinal products and diagnostics, the field of personalized medicine would profit from approved therapeutic concepts and would be a possible area for the application of therapeutic concepts. The link between medicinal products and medical devices, which are actually subject to different regulations, can be strengthened by therapeutic concepts. For a possible implementation of therapeutic concepts as a new regulatory approval pathway, several aspects have to be considered. The design of pivotal studies for the approval process should allow as much evidence on safety and effectiveness as possible without too many control arms in the study as this might be too time and cost-consuming and requires a high number of participants. Labelling, reimbursement and risk management are particularly challenging for therapeutic concepts. Labelling must be designed in such a way that the individual medicinal product can be identified as part of an approved therapeutic concept. Risk management und vigilance plans should be more extensive to reflect to additional risk caused by the combination.

Elevated greenhouse gases (GHGs) induce adverse conditions directly and indirectly, causing decreases in plant productivity. To deal with climate change effects, plants have developed various mechanisms including the fine-tuning of metabolism. Plant respiratory metabolism is highly flexible due to the presence of various alternative pathways. The mitochondrial alternative oxidase (AOX) respiratory pathway is responsive to these changes, and several lines of evidence suggest it plays a role in reducing excesses of reactive oxygen species (ROS) and reactive nitrogen species (RNS) while providing metabolic flexibility under stress. Here we discuss the importance of the AOX pathway in dealing with elevated carbon dioxide (CO2), nitrogen oxides (NOx), ozone (O3), and the main abiotic stresses induced by climate change. ; This work is supported by the DST-DAAD exchange program between KJG and ARF. This work in the KJG lab is partly supported by a Ramalingaswami Fellowship and IYBA from the Department of Biotechnology, Government of India. IFS has received funding from the European Union's Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant agreement no. 753301. ; Peer reviewed

The interaction of the alternative oxidase (AOX) pathway with nutrient metabolism is important for understanding how respiration modulates ATP synthesis and carbon economy in plants under nutrient deficiency. Although AOX activity reduces the energy yield of respiration, this enzymatic activity is upregulated under stress conditions to maintain the functioning of primary metabolism. The in vivo metabolic regulation of AOX activity by phosphorus (P) and nitrogen (N) and during plant symbioses with Arbuscular mycorrhizal fungi (AMF) and Rhizobium bacteria is still not fully understood. We highlight several findings and open questions concerning the in vivo regulation of AOX activity and its impact on plant metabolism during P deficiency and symbiosis with AMF. We also highlight the need for the identification of which metabolic regulatory factors of AOX activity are related to N availability and nitrogen-fixing legume-rhizobia symbiosis in order to improve our understanding of N assimilation and biological nitrogen fixation. ; FONDECYT from National Agency for Research and Development (ANID) 1191118 ; Chilean Scholarship Program/Becas de doctorado nacional/2017 21180329 ; European Union (EU) 753301

AbstractThe deep sea, the largest ocean's compartment, drives planetary-scale biogeochemical cycling. Yet, the functional exploration of its microbial communities lags far behind other environments. Here we analyze 58 metagenomes from tropical and subtropical deep oceans to generate the Malaspina Gene Database. Free-living or particle-attached lifestyles drive functional differences in bathypelagic prokaryotic communities, regardless of their biogeography. Ammonia and CO oxidation pathways are enriched in the free-living microbial communities and dissimilatory nitrate reduction to ammonium and H2 oxidation pathways in the particle-attached, while the Calvin Benson-Bassham cycle is the most prevalent inorganic carbon fixation pathway in both size fractions. Reconstruction of the Malaspina Deep Metagenome-Assembled Genomes reveals unique non-cyanobacterial diazotrophic bacteria and chemolithoautotrophic prokaryotes. The widespread potential to grow both autotrophically and heterotrophically suggests that mixotrophy is an ecologically relevant trait in the deep ocean. These results expand our understanding of the functional microbial structure and metabolic capabilities of the largest Earth aquatic ecosystem. ; We thank the R/V Hespérides captain and crew, the chief scientists in Malaspina expedition legs, and all project participants for their help in making this project possible. This work was funded by the Spanish Ministry of Economy and Competitiveness (MINECO) through the Consolider-Ingenio program (Malaspina 2010 Expedition, ref. CSD2008-00077). The sequencing of 58 bathypelagic metagenomes was done by the U.S. Department of Energy Joint Genome Institute, supported by the Office of Science of the U.S. Department of Energy under Contract No. DE-AC02 05CH11231 to SGA (CSP 612 "Microbial metagenomics and transcriptomics from a global deep-ocean expedition"). Additional funding was provided by the project MAGGY (CTM2017-87736-R) to S.G.A. from the Spanish Ministry of Economy and Competitiveness, Grup de Recerca 2017SGR/1568 from Generalitat de Catalunya, and King Abdullah University of Science and Technology (KAUST) under contract OSR #3362 and by funding of the EMFF Program of the European Union (MERCLUB project, Grant Agreement 863584). The ICM researchers have had the institutional support of the "Severo Ochoa Centre of Excellence" accreditation (CEX2019-000928-S). High-Performance computing analyses were run at the Marine Bioinformatics Service (MARBITS, https://marbits.icm.csic.es) of the Institut de Ciències del Mar (ICM-CSIC), Barcelona, Supercomputing Center (Grant BCV-2013-2-0001) and KAUST's Ibex HPC. We thank Shook Studio for assistance with figure design and implementation and the anonymous reviewers for their comments to improve this manuscript. This paper is dedicated to the memory of Professor Vladimir B. Bajic, 2019.

Sirtuins are NAD+-dependent deacetylases that facilitate cellular stress response. They include SirT6, which protects genome stability and regulates metabolic homeostasis through gene silencing, and whose loss induces an accelerated aging phenotype directly linked to hyperactivation of the NF-κB pathway. Here we show that SirT6 binds to the H3K9me3-specific histone methyltransferase Suv39h1 and induces monoubiquitination of conserved cysteines in the PRE-SET domain of Suv39h1. Following activation of NF-κB signaling Suv39h1 is released from the IκBα locus, subsequently repressing the NF-κB pathway. We propose that SirT6 attenuates the NF-κB pathway through IκBα upregulation via cysteine monoubiquitination and chromatin eviction of Suv39h1. We suggest a mechanism based on SirT6-mediated enhancement of a negative feedback loop that restricts the NF-κB pathway. ; This work was supported by the Fundació La Marató de TV3 (20133810 to A.V.), the Spanish Ministry of Economy and competitiveness-MINECO (SAF2011-25860, SAF2014-55964R to A.V.) confunded by FEDER funds/European Regional Development Fund (ERDF)-a way to Build Europe, and the Catalan government agency AGAUR (2009-SGR-914 to A.V.) The IDIBELL Proteomics Unit belongs to ProteoRed, PRB2-ISCIII, and is supported by grant PT13/0001/0033 and Cellex Foundation. S.B.B. is the recipient of a 13FIS037 fellowship.

AbstractUncertainty in model predictions of exposure response at low exposures is a problem for risk assessment. A particular interest is the internal concentration of an agent in biological systems as a function of external exposure concentrations. Physiologically based pharmacokinetic (PBPK) models permit estimation of internal exposure concentrations in target tissues but most assume that model parameters are either fixed or instantaneously dose‐dependent. Taking into account response times for biological regulatory mechanisms introduces new dynamic behaviors that have implications for low‐dose exposure response in chronic exposure. A simple one‐compartment simulation model is described in which internal concentrations summed over time exhibit significant nonlinearity and nonmonotonicity in relation to external concentrations due to delayed up‐ or downregulation of a metabolic pathway. These behaviors could be the mechanistic basis for homeostasis and for some apparent hormetic effects.

The European Union (EU) has introduced statutory limits for patulin in fruit products. Species definitions need to be unambiguous for this capability, especially to determine 'weak spots' in food, drink and feed production. Fungi were analysed for the isoepoxydon dehydrogenase (IDH) gene of the patulin metabolic pathway to indicate potential patulin production. In several cases inhibition of PCR product formation was indicated. Dilution as a means of overcoming PCR inhibition and to determine optimal concentrations was assessed and the ramifications for nucleic acid analysis in general are highlighted. The inhibitors may be secondary metabolites. However, inhibition was not involved obviously for Penicillium brevicompactum. The gene was detected frequently from Aspergillus section Clavati and Penicillium subgenus Penicillium species. Some strains within certain species were negative for the gene. Detection of the gene product in Byssochlamys is reported. ; UK Food Standards Agency - contract no. ...

Iron is an essential micronutrient that participates as a cofactor in a broad range of metabolic processes including mitochondrial respiration, DNA replication, protein translation and lipid biosynthesis. Adaptation to iron deficiency requires the global reorganization of cellular metabolism directed to optimize iron utilization. The budding yeast Saccharomyces cerevisiae has been widely used to characterize the responses of eukaryotic microorganisms to iron depletion. In this report, we used a genomic approach to investigate the contribution of transcription rates to the modulation of mRNA levels during adaptation of yeast cells to iron starvation. We reveal that a decrease in the activity of all RNA polymerases contributes to the down-regulation of many mRNAs, tRNAs and rRNAs. Opposite to the general expression pattern, many genes including components of the iron deficiency response, the mitochondrial retrograde pathway and the general stress response display a remarkable increase in both transcription rates and mRNA levels upon iron limitation, whereas genes encoding ribosomal proteins or implicated in ribosome biogenesis exhibit a pronounced fall. This expression profile is consistent with an activation of the environmental stress response. The phosphorylation stage of multiple regulatory factors strongly suggests that the conserved nutrient signaling pathway TORC1 is inhibited during the progress of iron deficiency. These results suggest an intricate crosstalk between iron metabolism and the TORC1 pathway that should be considered in many disorders. ; This work was supported by predoctoral contracts from the Spanish Ministry of Science, Innovation and Universities (MICINN) to AMR and LRA; a fellowship from the "Generalitat de Catalunya" (Spain) to SMM; European Union Funds (FEDER) and MICINN grants BIO2017-87828-C2-1-P to SP, BIO2017-87828-C2-2-P to MATR, BFU2016-77728-C3-3-P to JEPO, and BFU2015-71978-REDT to SP and JEPO; and Regional Government of Valencia PROMETEOII 2015/006 grant to JEPO.

Publisher's version (útgefin grein). ; Background: Lactobacillus reuteri is a heterofermentative Lactic Acid Bacterium (LAB) that is commonly used for food fermentations and probiotic purposes. Due to its robust properties, it is also increasingly considered for use as a cell factory. It produces several industrially important compounds such as 1,3-propanediol and reuterin natively, but for cell factory purposes, developing improved strategies for engineering and fermentation optimization is crucial. Genome-scale metabolic models can be highly beneficial in guiding rational metabolic engineering. Reconstructing a reliable and a quantitatively accurate metabolic model requires extensive manual curation and incorporation of experimental data. Results: A genome-scale metabolic model of L. reuteri JCM 1112T was reconstructed and the resulting model, Lreuteri_530, was validated and tested with experimental data. Several knowledge gaps in the metabolism were identified and resolved during this process, including presence/absence of glycolytic genes. Flux distribution between the two glycolytic pathways, the phosphoketolase and Embden-Meyerhof-Parnas pathways, varies considerably between LAB species and strains. As these pathways result in different energy yields, it is important to include strain-specific utilization of these pathways in the model. We determined experimentally that the Embden-Meyerhof-Parnas pathway carried at most 7% of the total glycolytic flux. Predicted growth rates from Lreuteri_530 were in good agreement with experimentally determined values. To further validate the prediction accuracy of Lreuteri_530, the predicted effects of glycerol addition and adhE gene knock-out, which results in impaired ethanol production, were compared to in vivo data. Examination of both growth rates and uptake- and secretion rates of the main metabolites in central metabolism demonstrated that the model was able to accurately predict the experimentally observed effects. Lastly, the potential of L. reuteri as a cell factory was investigated, resulting in a number of general metabolic engineering strategies. Conclusion: We have constructed a manually curated genome-scale metabolic model of L. reuteri JCM 1112T that has been experimentally parameterized and validated and can accurately predict metabolic behavior of this important platform cell factory. ; This study was supported by the Marine Biotechnology ERA-NET Thermo-Factories project grant number 5178–00003B; the Technology Development fund in Iceland grant number 159004-0612; The Novo Nordisk Foundation in Denmark; and the European Union's Horizon 2020 research and innovation programme under grant agreement No 686070 (DD-DeCaF). ; Peer Reviewed

This article belongs to the Special Issue Plant Respiration. ; The interaction of the alternative oxidase (AOX) pathway with nutrient metabolism is important for understanding how respiration modulates ATP synthesis and carbon economy in plants under nutrient deficiency. Although AOX activity reduces the energy yield of respiration, this enzymatic activity is upregulated under stress conditions to maintain the functioning of primary metabolism. The in vivo metabolic regulation of AOX activity by phosphorus (P) and nitrogen (N) and during plant symbioses with Arbuscular mycorrhizal fungi (AMF) and Rhizobium bacteria is still not fully understood. We highlight several findings and open questions concerning the in vivo regulation of AOX activity and its impact on plant metabolism during P deficiency and symbiosis with AMF. We also highlight the need for the identification of which metabolic regulatory factors of AOX activity are related to N availability and nitrogen-fixing legume-rhizobia symbiosis in order to improve our understanding of N assimilation and biological nitrogen fixation. ; This work was supported by FONDECYT No. 1191118 from National Agency for Research and Development (ANID) and the Chilean Scholarship Program/Becas de doctorado nacional/2017–21180329. I.F.-S. has received funding from the European Union's Horizon 2020 research and innovation program under the Marie Skłodowska-Curie grant agreement no. 753301. ; Peer reviewed

Il lavoro di tesi consiste nello sviluppo di un modello in silico per la simulazione dell'azione del farmaco all'interno del pathway metabolico scelto e uno studio di docking molecolare. Il farmaco preso in considerazione è il Viagra, che contiene il principio attivo Sildenafil. La sua funzione è il rilassamento della muscolatura liscia. Il linguaggio di programmazione considerato per la modellazione è Python. Il software per effetuare il docking molecolare è AutoDock Vina. Con le informazioni ottenute è possibile verificare l'utilizzo del Sildenafil in altre applicazioni cliniche, diverse da quelle approvate dagli enti governativi. The thesis work consists in the development of an in silico model for the simulation of the drug action within the chosen metabolic pathway and a molecular docking study. The drug under consideration is Viagra, which contains the active ingredient Sildenafil. Its function is the relaxation of smooth muscles. The programming language considered for modeling is Python. The software for molecular docking is AutoDock Vina. With the information obtained, it is possible to verify the use of the Sildenafil in other clinical applications, other than those approved by government bodies.

15 pages, 7 figures, supplementary information https://doi.org/10.1038/s42003-021-02112-2.-- All data generated or analyzed during this study are included in this published article (and its supplementary information files). All raw sequences are publicly available at both DOE's JGI Integrated Microbial Genomes and Microbiomes (IMG/MER) and the European Nucleotide Archive (ENA). Individual metagenome assemblies, annotation files, and alignment files can be accessed at IMG/MER. All accession numbers are listed in Supplementary Data 1. The co-assembly for the MAG dataset construction can be found through ENA at https://www.ebi.ac.uk/ena with accession number PRJEB40454, the nucleotide sequence for each MAG and their annotation files can be found through BioStudies at https://www.ebi.ac.uk/biostudies with accession S-BSST457 and also in the companion website to this manuscript at https://malaspina-public.gitlab.io/malaspina-deep-ocean-microbiome/.-- All software used in this work is publicly available distributed by their respective developers, and it is described in "Methods", including the versions and options used. Additional custom scripts to assign taxonomy to the M-geneDB genes and to filter and format FRA results are available through BioStudies at https://www.ebi.ac.uk/biostudies with accession S-BSST457 ; The deep sea, the largest ocean's compartment, drives planetary-scale biogeochemical cycling. Yet, the functional exploration of its microbial communities lags far behind other environments. Here we analyze 58 metagenomes from tropical and subtropical deep oceans to generate the Malaspina Gene Database. Free-living or particle-attached lifestyles drive functional differences in bathypelagic prokaryotic communities, regardless of their biogeography. Ammonia and CO oxidation pathways are enriched in the free-living microbial communities and dissimilatory nitrate reduction to ammonium and H2 oxidation pathways in the particle-attached, while the Calvin Benson-Bassham cycle is the most prevalent inorganic carbon fixation pathway in both size fractions. Reconstruction of the Malaspina Deep Metagenome-Assembled Genomes reveals unique non-cyanobacterial diazotrophic bacteria and chemolithoautotrophic prokaryotes. The widespread potential to grow both autotrophically and heterotrophically suggests that mixotrophy is an ecologically relevant trait in the deep ocean. These results expand our understanding of the functional microbial structure and metabolic capabilities of the largest Earth aquatic ecosystem ; This work was funded by the Spanish Ministry of Economy and Competitiveness (MINECO) through the Consolider-Ingenio program (Malaspina 2010 Expedition, ref. CSD2008-00077). The sequencing of 58 bathypelagic metagenomes was done by the U.S. Department of Energy Joint Genome Institute, supported by the Office of Science of the U.S. Department of Energy under Contract No. DE-AC02 05CH11231 to SGA (CSP 612 "Microbial metagenomics and transcriptomics from a global deep-ocean expedition"). Additional funding was provided by the project MAGGY (CTM2017-87736-R) to S.G.A. from the Spanish Ministry of Economy and Competitiveness, Grup de Recerca 2017SGR/1568 from Generalitat de Catalunya, and King Abdullah University of Science and Technology (KAUST) under contract OSR #3362 and by funding of the EMFF Program of the European Union (MERCLUB project, Grant Agreement 863584). The ICM researchers have had the institutional support of the "Severo Ochoa Centre of Excellence" accreditation (CEX2019-000928-S). High-Performance computing analyses were run at the Marine Bioinformatics Service (MARBITS, https://marbits.icm.csic.es) of the Institut de Ciències del Mar (ICM-CSIC), Barcelona, Supercomputing Center (Grant BCV-2013-2-0001) and KAUST's Ibex HPC ; Peer reviewed

Bacterial photobiohybrids and photosynthesis: Optimizing energy harvesting with bacterial-semiconductor hybrids
Photosynthesis serves as the primary mechanism for converting solar energy into chemical energy and plays a pivotal role in regulating atmospheric oxygen levels and carbon dioxide concentrations, influencing global climate patterns as a result. Traditional photosynthetic pigments, such as chlorophyll, can only absorb specific regions of the electromagnetic spectrum, leaving a significant span of wavelengths of light unutilized. The solar energy-conversion efficiency of photosynthesis typically reaches a maximum of ~6%; however, some plants exhibit efficiencies lower than 1%, depending on the type of organism, environmental conditions, and the specific metabolic pathway involved. While photosynthesis has relatively low efficiency compared to some human-made solar energy technologies, it plays a crucial role in sustaining life on Earth by providing the primary source of energy for most ecosystems.

AbstractObesity is associated with chronic inflammation that affects various organs in the body, including the reproductive system, which is a key factor in male infertility. 1-Deoxynojirimycin (1-DNJ) is a natural alkaloid in mulberry leaves, which has anti-inflammatory capabilities, yet, it's effects on obesity-induced inflammation-related male infertility remain unclear. Therefore, this research investigates the underlying mechanism by which 1-DNJ may mitigate fertility impairment in male mice caused by obesity-related inflammation. Male mice with high-fat diet (HFD)-induced obesity were treated with 1-DNJ or metformin for 8 weeks. Metabolic profiles were evaluated by enzyme method. Reproductive capacity was assessed by sperm viability, motility and counts, immunohistochemistry was performed to evaluate the testicular damage caused by obesity and inflammation. The inflammation was assessed by measuring the levels of tumor necrosis factor α (TNFα), interleukin 1β (IL-1β), and interleukin 6 (IL-6). The activation of IκB kinase β (IKKβ) and nuclear factor κB (NF-κB) was examined using western blot and immunohistochemistry. HFD induced obesity in mice with obvious lipid metabolism disorder. The obese male mice had a decreased testosterone level, impaired sperm motility, and increased inflammatory factors. 1-DNJ treatment improved the testosterone level in the obese mice, ameliorated the testicular structure damage and improve sperm viability. In addition, 1-DNJ treatment inhibited IKKβ/NF-kB signaling pathway and reduced inflammation in obese mice. 1-DNJ can improve the fertility of obese men by reducing obesity as well as obesity-induced inflammation. These findings provide new insights for 1-DNJ to alleviate inflammation caused by obesity and provide future possibilities for treating male infertility.

AbstractWinery effluents containing high ethanol concentrations and diverse organic matter are ideal substrates for producing medium-chain carboxylic acids via fermentation and chain elongation. However, the process needs to be better understood. This study presents novel insights into the bioconversion mechanisms of medium-chain carboxylic acids by correlating fermentation and chain elongation kinetic profiles with the study of microbial communities at different pH (5 to 7) conditions and temperatures (30 to 40 °C). It was found that high productivities of MCCA were obtained using a native culture and winery effluents as a natural substrate. Minor pH variations significantly affected the metabolic pathway of the microorganisms for MCCA production. The maximal productivities of hexanoic (715 mg/L/d) and octanoic (350 mg/L/d) acids were found at pH 6 and 35 °C. Results evidence that the presence of Clostridium, Bacteroides, and Negativicutes promotes the high productions of MCCA. The formation of heptanoic acid was favor when Mogibacterium and Burkholderia were present.
Graphical Abstract