Application of a genome-based predictive CHO model for increased mAb production and Glycosylation control

Monoclonal antibody therapeutics continue to grow in both number and market share with recent forecasts of global sales reaching ~$125MM by 2020. Most mAb products currently on the market are produced using cultured mammalian cells, typically Chinese Hamster Ovary (CHO) cells, which provide the necessary post-translational modifications to make the antibody efficacious. Many post-translational modifications such as the oligosaccharide profile are considered critical quality attributes (CQAs) that must be tightly controlled throughout the manufacturing process to ensure product safety and effectiveness. Therefore, the ability to predict how cell culture media components, including potential contaminants like trace metals, will affect product formation and glycosylation is important from both a process development and process control viewpoint. A detailed genome-based, predictive CHO model from the Insilico Cells™ library was adapted by the reconstruction software Insilico Discovery™ for a representative fed-batch process through a collaborative effort leveraging the computational and experimental expertise of two companies. The final, compartmentalized network model contained 1900 reactions (including transport reactions), 1300 compounds and contains stoichiometric descriptions of anabolic pathways for amino acids, lipids and carbohydrate species. The genome-scale model was constrained using several assumptions on the cell physiology and then used to compute time-resolved flux distributions by the software module Insilico Inspector™. The Insilico Designer™ module was then used to subsequently reduce the large model to a computationally manageable reduced model able to describe all flux distributions using 5 flux modes, of which 4 combined several metabolic functions and one is independently responsible for product synthesis. Using Insilico Designer™, the kinetic parameters of the reduced model were estimated by fitting the model-predicted metabolite concentrations to the experimentally determined values. The calibrated model was able to properly describe the time-dependent trajectories of biomass, product and most metabolites. Simulations using the reduced model were run and a media composition predicted to improve mAb production was identified and experimentally verified. Furthermore, experiments probing the effects of trace metals on product glycosylation were used to extend the model’s glycosylation predictability. The ability to identify both metabolic signatures, as well as media components, that correlate to specific glycan profiles will allow for fine-tuning of desired CQAs and enable more robust control strategies in upstream processes

Multiscale modeling of monoclonal antibody (mAb) production and glycosylation in a CHO cell culture process

The production of recombinant therapeutic monoclonal antibodies (mAbs) using cultured mammalian cells accounts for approximately $80 billion in global sales annually. These antibodies are often produced using Chinese hamster ovary (CHO) cell lines that execute the necessary post-translational modifications (e.g., glycosylation) for the drug to be therapeutically efficacious. Glycosylation is an intracellular, enzymatic process by which glycans (i.e., sugar molecules) are attached to a specific location on the antibody. The structure of the glycans attached to the mAb affects the therapeutic function of the molecule, making glycan distribution a critical quality attribute. Consequently, the ability to predict how variations in process parameters and/or media components affect both product formation and glycosylation is important from both a process development and process control viewpoint. A multiscale, mathematical model describing CHO cell growth and antibody production was developed in MATLAB to provide a quantitative understanding of how to manipulate a cell culture process to improve antibody titer and control glycosylation effectively. At the macro (bioreactor) scale, the model uses Monod growth kinetics to describe cell growth, nutrient/metabolite concentrations, and mAb production; at the micro scale, the glycosylation process in the Golgi apparatus is modeled using a glycosylation reaction network governed by Michaelis-Menten enzyme kinetics. Although both macro and micro scale processes are dynamic, disparate time scales makes it possible to solve the (fast) glycosylation model as a static function of the (slowly changing) macro scale state variables. In this multidisciplinary study, we will present a design of experiments approach for (1) identifying significant factors affecting glycosylation—including concentrations of asparagine, glutamine, and copper in the media, and (2) using these factors as macro scale “inputs” to the micro scale model. Model predictions are validated against an independent data set from a representative industrial mammalian cell culture process. Ultimately, the models we discuss will be valuable for biopharmaceutical process development and model-based control system design

Transdisciplinary learning: Transformative collaborations between students, industry, academia and communities.

Background and objectives of the case An analogy: Imagine you are invited to a dinner party, but instead of a stuffy sit-down affair, your host asks you to bring your favourite ingredient, and together you prepare a delicious feast of unique and distinct flavours. UTS’s transdisciplinary initiatives are changing the shape of higher education and forging innovative partnerships by bringing together diverse professional fields. With a focus on practice-based and problem-focused learning, UTS educational programs combine the strengths of multiple disciplines, industries, public sector organisations, and the community to turn real-world problems into rewarding opportunities for education and also “learning for a lifetime”. In place of the limitations of artificial disciplinary boundaries, transdisciplinary learning practices create synergistic and innovative approaches to grappling with complex applied challenges. Students, researchers, practitioners, community members and other stakeholders combine their knowledge, tools, techniques, methods, theories, concepts, as well as cultural and personal perspectives. By understanding problems holistically, the solutions that emerge are bold, innovative, and creative, as well as mutually beneficial. We view this as the future of education: good to work with, and good to think with — problem solving for (and with) industry and society. The Faculty of Transdisciplinary Innovation is re-imagining how education, research, and professional practice can work together to navigate today’s complex problems, and create commercially attractive and socially responsible futures. We also practice what we preach: for example, staff professional development to enact these models in our own teaching; educational programs to provide experiential learning around problem solving within a rapidly-changing environment involving students from across different disciplines and cultural backgrounds; as well as policy development and research on today’s pressing “wicked problems” with industry and government. Primary objectives of this next practice concept of transdisciplinary learning, include: - To promote a shift in industry-university engagement from producing “knowledge for society” to co-generating “knowledge with society”; - To build a resilient ecosystem for co-learning; - To create and sustain future-oriented degree programs with collaboration between industry, government, and community at the centre, geared to prepare our graduates for the complex challenges of a networked world; - To create an agile and responsive industry-university lab environment for generating and testing new experimental models; - To enable industry – by collaborating with our students and academics – to see their problems from a fresh perspective, often through different and revealing lenses, and to notice opportunities and spot challenges that may have otherwise been overlooked; - To prepare students to lead innovation in a rapidly-changing and challenging world; and - To graduate students who are ‘complexity-fluent’, systems thinkers, creative problem-posers and -solvers, and imaginative, ethical citizens

On-Orbit Results From the NASA Time-Resolved Observations of Precipitation Structure and Storm Intensity With a Constellation of Smallsats (TROPICS) Mission

The NASA TROPICS Earth Venture (EVI-3) CubeSat constellation mission will provide nearly all-weather observations of 3-D temperature and humidity, as well as cloud ice and precipitation horizontal structure, at high temporal resolution to conduct high-value science investigations of tropical cyclones. TROPICS will provide rapid-refresh microwave measurements (median refresh rate better than 60 minutes for the baseline mission) over the tropics that can be used to observe the thermodynamics of the troposphere and precipitation structure for storm systems at the mesoscale and synoptic scale over the entire storm lifecycle. The TROPICS constellation mission comprises four 3UCubeSats (5.4 kg each) in two low-Earth orbital planes. Each CubeSat contains a Blue Canyon Technologies bus and a high-performance radiometer payload to provide temperature profiles using seven channels near the 118.75 GHz oxygen absorption line, water vapor profiles using three channels near the 183 GHz water vapor absorption line, imagery in a single channel near 90 GHz for precipitation measurements (when combined with higher resolution water vapor channels), and a single channel at 205 GHz that is more sensitive to precipitation-sized ice particles. TROPICS spatial resolution and measurement sensitivity is comparable with current state-of-the-art observing platforms. Two dedicated launches (two spacecraft per launch) for the TROPICS constellation mission on Rocket Lab Electron vehicles occurred in 2023 (May 8 and May 26) to place the spacecraft in 32.75-degree inclined orbits at 550 km altitude. Data will be downlinked to the ground via the KSAT-Lite ground network. NASA\u27s Earth System Science Pathfinder (ESSP) Program Office approved the separate TROPICS Pathfinder mission, which launched on June 30, 2021, in advance of the TROPICS constellation mission as a technology demonstration and risk reduction effort. The TROPICS Pathfinder mission has provided an opportunity to checkout and optimize all mission elements prior to the primary constellation mission and is still operating nominally

The NASA Time-Resolved Observations of Precipitation Structure and Storm Intensity with a Constellation of Smallsats (TROPICS) Mission: Results from the Pathfinder Demonstration and Look Ahead to the Constellation Mission

The NASA Time-Resolved Observations of Precipitation structure and storm Intensity with a Constellation of Smallsats (TROPICS) mission will provide nearly all-weather observations of 3-D temperature and humidity, as well as cloud ice and precipitation horizontal structure, at high temporal resolution to conduct high-value science investigations of tropical cyclones. TROPICS will provide rapid-refresh microwave measurements (median refresh rate of approximately 50 minutes for the baseline mission) over the tropics that can be used to observe the thermodynamics of the troposphere and precipitation structure for storm systems at the mesoscale and synoptic scale over the entire storm lifecycle. The TROPICS constellation mission comprises six CubeSats in three low-Earth orbital planes. Each CubeSat will host a high-performance radiometer to provide temperature profiles using seven channels near the 118.75 GHz oxygen absorption line, water vapor profiles using three channels near the 183 GHz water vapor absorption line, imagery in a single channel near 90 GHz for precipitation measurements (when combined with higher resolution water vapor channels), and a single channel at 205 GHz that is more sensitive to precipitation-sized ice particles. TROPICS spatial resolution and measurement sensitivity is comparable with current state-of-the-art observing platforms. Launches for the TROPICS constellation mission are planned in 2022. NASA’s Earth System Science Pathfinder (ESSP) Program Office approved the separate TROPICS Pathfinder mission, which launched into a sun-synchronous orbit (2:00pm LTDN, 530 km) on June 30, 2021, in advance of the TROPICS constellation mission as a technology demonstration and risk reduction effort. The TROPICS Pathfinder mission has provided an opportunity to checkout and optimize all mission elements prior to the primary constellation mission. In this paper, we describe the instrument checkout and calibration/validation plans and progress for the TROPICS Pathfinder mission and discuss first light mission results. All spacecraft and radiometer systems are fully operational as of Launch + 11 months

'The best thing for the baby': mothers' concepts and experiences related to promoting their infants' health and development

Mothers and pregnant women in contemporary western societies are at the centre of a web of expert and lay discourses concerning the ways they should promote and protect the health and development of their foetuses and infants. This article reports the findings from an Australian study involving interviews with 60 mothers. The findings explore in detail four topics discussed in the interviews related to pregnancy and caring for young infants: disciplining the pregnant body; promoting infants’ health; immunisation; and promoting infants’ development. It is concluded that the mothers were highly aware of their responsibilities in protecting their foetuses and infants from harm and promoting their health and development. They conceptualised the infant body as highly vulnerable and requiring protection from contamination. They therefore generally supported the idea of vaccination as a way of protecting their babies’ immature immune systems, but were also often ambivalent about it. The mothers were aware of the judgemental attitudes of others, including other mothers, towards their caring efforts and attempted to conform to the ideal of the ‘good mother’. The emotional dimensions of caring for infants and protecting their health are discussed in relation to the voluntary participation of mothers in conforming to societal expectations

Gravitational Waves From Known Pulsars: Results From The Initial Detector Era

We present the results of searches for gravitational waves from a large selection of pulsars using data from the most recent science runs (S6, VSR2 and VSR4) of the initial generation of interferometric gravitational wave detectors LIGO (Laser Interferometric Gravitational-wave Observatory) and Virgo. We do not see evidence for gravitational wave emission from any of the targeted sources but produce upper limits on the emission amplitude. We highlight the results from seven young pulsars with large spin-down luminosities. We reach within a factor of five of the canonical spin-down limit for all seven of these, whilst for the Crab and Vela pulsars we further surpass their spin-down limits. We present new or updated limits for 172 other pulsars (including both young and millisecond pulsars). Now that the detectors are undergoing major upgrades, and, for completeness, we bring together all of the most up-to-date results from all pulsars searched for during the operations of the first-generation LIGO, Virgo and GEO600 detectors. This gives a total of 195 pulsars including the most recent results described in this paper.United States National Science FoundationScience and Technology Facilities Council of the United KingdomMax-Planck-SocietyState of Niedersachsen/GermanyAustralian Research CouncilInternational Science Linkages program of the Commonwealth of AustraliaCouncil of Scientific and Industrial Research of IndiaIstituto Nazionale di Fisica Nucleare of ItalySpanish Ministerio de Economia y CompetitividadConselleria d'Economia Hisenda i Innovacio of the Govern de les Illes BalearsNetherlands Organisation for Scientific ResearchPolish Ministry of Science and Higher EducationFOCUS Programme of Foundation for Polish ScienceRoyal SocietyScottish Funding CouncilScottish Universities Physics AllianceNational Aeronautics and Space AdministrationOTKA of HungaryLyon Institute of Origins (LIO)National Research Foundation of KoreaIndustry CanadaProvince of Ontario through the Ministry of Economic Development and InnovationNational Science and Engineering Research Council CanadaCarnegie TrustLeverhulme TrustDavid and Lucile Packard FoundationResearch CorporationAlfred P. Sloan FoundationAstronom

Multi-stage genome-wide association study identifies new susceptibility locus for testicular germ cell tumour on chromosome 3q25

Recent genome-wide association studies (GWAS) and subsequent meta-analyses have identified over 25 SNPs at 18 loci, together accounting for >15% of the genetic susceptibility to testicular germ cell tumour (TGCT). To identify further common SNPs associated with TGCT, here we report a three-stage experiment, involving 4098 cases and 18 972 controls. Stage 1 comprised previously published GWAS analysis of 307 291 SNPs in 986 cases and 4946 controls. In Stage 2, we used previously published customised Illumina iSelect genotyping array (iCOGs) data across 694 SNPs in 1064 cases and 10 082 controls. Here, we report new genotyping of eight SNPs showing some evidence of association in combined analysis of Stage 1 and Stage 2 in an additional 2048 cases of TGCT and 3944 controls (Stage 3). Through fixed-effects meta-analysis across three stages, we identified a novel locus at 3q25.31 (rs1510272) demonstrating association with TGCT [per-allele odds ratio (OR) = 1.16, 95% confidence interval (CI) = 1.06-1.27; P = 1.2 × 10-9]