Technical and regulatory opportunities and challenges for cell and gene therapies in low earth orbit: a status report

Abstract

The unique advantages exhibited by microgravity in enhancing the biological and chemical interactions for cells and tissues have come into greater focus following a quarter century of biological investigation aboard the International Space Station (ISS) in low earth orbit (LEO). One of its primary biomedical research purposes has been to investigate and mitigate the health risks faced by astronauts during prolonged spaceflight. However, as this status report describes, the hundreds of experiments aboard the ISS have also produced a vast quantity of knowledge opening up new possibilities for improving therapeutic modalities for the unmet medical needs of patients on earth. Among its many functions and capabilities, the ISS has been a preeminent biomedical research laboratory for biotechnology and drug development. Public–private partnerships have created the necessary collaborations and supplied the resources to conduct sophisticated biomedical experiments which have led to the improvement in the applications of stem cell biology, gene therapies, tissue engineering, and regenerative medicine. Technological advancements have also resulted in 3D bio printing of soft tissues such as blood vessels and micro physiological systems (Tissue on a Chip) using cells organized in a predetermined architecture. Nanomaterials assembled in microgravity formed with increased homogeneity and bioactivity can function as delivery platforms for cancer therapeutics or may be shaped into extracellular matrix supporting tissue regeneration therapies. Given these exciting innovations, and with the expectation that a robust regulatory framework will emerge, sustainable biomanufacturing in LEO is poised to unlock a transformative economic potential and accelerate the development of advanced next-generation therapeutics.

Low earth orbit (LEO) is a zone which extends from an altitude as low as 100 miles (160 km) to an altitude of 1,200 miles (2,000 km) above the surface of the earth. Within LEO, at an altitude of 250 miles, the 356 ft. International Space Station (ISS) has been in continuous operation since 2000, circling the earth every 93 minutes at 17,500 miles per hour (Figure 1) [1].

FIGURE 1. The ISS has been circling in LEO since 2000.

Photo courtesy of NASA.

Aboard the ISS astronaut-scientists observe 16 sunrises and sunsets every 24 hours. As the ISS nears its 2030 retirement and potential deorbiting, over 4,000 research investigations have been undertaken by numerous scientific organizations from 108 countries [2]. Expeditions carrying experiments and scientists continue to be transported to the ISS sponsored by governmental, academic, medical, and commercial organizations all of whom utilize this celestial laboratory to better understand the effects of microgravity, radiation and a near continuous vacuum on human physiology, biology, material, and the physical sciences [3]. While microgravity may be simulated on earth, long term environmental conditions of microgravity cannot be duplicated. The vast knowledge gained aboard the ISS over the past quarter century have shown that the properties exhibited in microgravity can have a dramatic effect upon spaceflight mediated risks of radiation, isolation, and confinement, distance from earth, gravity effects, and the hostile and closed environments that affect the physical, chemical, and biological systems of astronauts (Box 1) [4–9].

BOX 1. Space-flight mediated health hazards.

Molecular and cellular features

• Oxidative stress

• DNA damage

• Mitochondrial dysregulation

• Epigenetic and gene regulation changes

• Telomere length dynamics

• Passive osteogenic differentiation

• Microbiome shifts

• Hemoglobin degradation

• Shortened cell cycles

Systemic and physiological health risks

• Cardiovascular deconditioning and dysregulation

• CNS impairments

• Increased blood–brain barrier permeability

• Increased cancer risk

• Muscle degeneration

• Osteoarthritis and bone loss

• Cartilage degradation

• Defects in wound and bone fracture healing

• Immune dysfunction

• Increased liver disease and lipid dysregulation

• Circadian rhythm dysregulation

• Space associated neuro-ocular syndrome

• Altered mechanics of blood flow

Spaceflight mediated health hazards collectively affect multiple biological systems spanning space radiation, microgravity, confinement/isolation, a hostile/close environment, and distance from earth. These areas require scientific advancement to enable deep space exploration [5–9].

Mitigation of these risks will lead to plans for long-term residency of humans on the moon and preparations for long duration space travel to the planet Mars [10]. Planning for the maintenance of crew health and provision of medical care when emergency evacuation is not possible is an essential part of human led explorations to the moon and beyond. These goals will require system development and biomedical technological innovations to ensure the health of astronauts in the years ahead [11]. The ISS is entering its third decade of use as the world’s preeminent orbital microgravity innovation laboratory. The efforts of the scientists working in areas of biology, physics, biomedicine, materials, earth, and space science have produced innumerable research discoveries and demonstrations of advanced technology, all of which will continue to return significant benefits to humanity on earth. This status report is intended to provide an overview of the current state of knowledge specific to the cell and gene therapy sector of LEO and to summarize those areas under active investigation in which the microgravity environment has been shown to exert an impact on the function of cells and tissues.

Much of the knowledge gained from research conducted in space over the past quarter century will contribute to improvements for life on earth. As a result, the creation of new and rapidly evolving initiatives is underway whereby the manufacturing of advanced materials and pharmaceutical products can take advantage of the unique environmental properties of LEO [12–16]. In anticipation of these new initiatives and possibilities, National Aeronautics and Space Agency (NASA) and the International Space Station National Laboratory (ISSNL) have partnered with a number of commercial organizations to facilitate the installation of research facilities aboard the ISS in support of business models promoting the creation of manufacturing platforms in LEO. Malshe describes the key drivers for a commercial space infrastructure for the servicing and assembly of orbital manufacturing facilities [17]. These include:

• Resource consumption limits due to population growth

• Human exploration advancements

• Declining launch costs

• Evolving in-space policies

• Geopolitics

• Advanced spacecraft accessibility

• Demand for space technology platforms [17]

In fact, a number of companies are vying for the opportunity to replace the ISS so as to continue research objectives under conditions of microgravity. Among those companies that have evaluated the impact, value creation, and feasibility, and are anxious to design, build, and launch their facilities into LEO are Vast Space, Blue Origin, StarLab Space, Sierra Space, SpaceX, and Axiom Space.

BIOTECHNOLOGY AND DRUG DEVELOPMENT IN SPACE

Presently, over 300 commercial entities have been established focused on the development and support of space manufacturing efforts in areas including advanced materials, biofabrication, and biotechnology [18]. The number of patents using the terminology ‘microgravity’ in the title or abstract rose from 21 in 2000 to 155 in 2020 [19]. Of these, a number are committed to the areas of biotechnology, biological research, and drug development of therapeutic modalities including cell and gene therapy, nanomaterial therapeutics, biologics, and medical devices [15,20,21]. To date, over 900 research articles have been published on biological and biotechnological adaptations to microgravity [22]. Opportunities for expanding business through space manufacturing of biopharmaceuticals including cell and gene therapies have been deemed feasible as revenue projections for making certain products in orbit exceed the operational costs of space manufacturing [23]. For example, McKinsey estimates that in orbit production of pharmaceutical products at maturity could reach an annual revenue at maturity of between 2.8 and 4.2 billion dollars [19]. With respect to biomanufacturing, Sharma determined revenues projected through 2035 and broke down the biomanufacturing market into five subsegments including the manufacturing of cell and tissue therapies (Figure 2) [24]. One company, Varda Space, has recently raised $90 million toward this effort in developing spacecraft that can autonomously manufacture active pharmaceutical ingredients with an ability to deliver these materials back to earth [25]. Deloitte consultants, who have studied the potential for commercialization and industrialization of space predicted that by the year 2035 there will be a vibrant economy situated in LEO resulting in an annual market of 312 billion dollars, an 8-fold increase in today’s economic value of LEO [16].

FIGURE 2. Biomanufacturing in LEO market sub-segmentation revenue projection.

The LEO biomanufacturing market for regenerative medicine is broken down into five primary sub-segments: cell and tissue tools and diagnostics; cell and tissue therapy; bioprinting; cell therapy biomanufacturing; and organoids. Projections are for the next 15 years. Redrawn with permission from Sharma et al. [24].

CHARACTERISTICS OF A MICROGRAVITY ENVIRONMENT

The force of gravity exists in space. In fact, the gravitational field where most human spaceflight occurs is significant. At 250 miles above the earth’s surface the gravitational field is 88.8% of ‘normal gravity’ (1 g) at sea level. Spacecraft that orbit the earth maintain their position in orbit through the force of gravity and the speed with which the craft is traveling. That speed, 17,500 miles per hour, sufficient to maintain a state of continuous free-fall is considered to be a ‘state of orbit’ [26]. The two forces, centripetal force from the circular motion of spacecraft orbiting around the earth at 17,500 miles per hour and the gravitational force pulling the spacecraft toward earth are equal and balanced. While the ISS can be considered in a state of free fall its high horizontal velocity ensures that the station’s flight trajectory will never touch the earth’s surface. The ISS provides a microgravity value around 10⁻⁵ g, periodically increasing to 10⁻⁴ g during a re-boost phase that occurs approximately once per month when the ISS adjusts its orbit, an event that lasts about 30 seconds. Objects inside the ISS appear to be floating or ‘weightless’ due to this state of ‘microgravity’ [27].

This phenomenon has been studied extensively within aircraft that fly parabolic arcs to create brief periods of microgravity or in zero gravity research facilities containing drop towers in which test packages can be dropped into a vacuum to create brief moments of weightlessness [28]. Life appeared on earth 4 billion years ago adapting to the planet’s gravitational pull. However, in an environment of microgravity where there is a lack of an up or down, significant alterations in the biochemistry of living things, including man become evident [15].

A quarter of a century of research within the ISS has provided an extensive record in understanding the effects microgravity has on the myriad of physical, chemical, and biological properties of life. The characteristics of microgravity are summarized in NASA’s Microgravity Science on the ISS: A Primer for new Researchers (Box 2) [26].

BOX 3. Characteristics of microgravity.

• Absence of convection: there is no convection due to differences in relative densities

• Absence of sedimentation: In microgravity substances of different relative densities, such as water and oil, will disperse evenly

• Absence of buoyancy: buoyancy becomes insignificant in the space environment, light and heavy materials can be mixed uniformly

• Absence of hydrostatic pressure: almost no hydrostatic pressure exists in microgravity

• Containerless float: In microgravity liquids can float in the air without a container

• Dominance of diffusive properties: In microgravity diffusion is the dominant process, a gentler mixing that enables more perfect, uniform, and precise structures at the level of individual molecules and groups of atoms

• Dominance of materials surface tension: Microgravity allows surface tension features to dominate for more precise adhesion, contact, and interactions between layers of similar and dissimilar constituents

The ISS as a Biomedical Research Laboratory for Cell and Gene Therapy.

Reproduced from [26].

The biological laboratory facilities installed aboard the ISS provide access to a microgravity environment and are made available to allow for biological experimentation. Among its many scientific functions, the ISS is well equipped as a biological and biomedical research laboratory (Figure 3) [29,30].

FIGURE 3. A 3D floating laboratory in space.

NASA astronaut, Sunita Williams works on StemCellEX-H1, a technology for in-space production of human stem cells that are used as therapies for certain blood diseases and cancers. Photo taken on Oct 2, 2024, courtesy of NASA.

Each space agency including NASA, European Space Agency (ESA), Japan Aerospace Exploration Agency (JAXA), Russian Federal Space Agency (Roscosmos) and the Canadian Space Agency (CSA) has contributed to the construction and operation of the ISS and has included in its instrument designs, laboratory research facilities to conduct biological and biotechnological experiments supporting cell and gene therapy applications. Flexible modular racks and lockers are dedicated to biological research and come with sophisticated instrumentation to support the experiment packages routinely transported to the ISS by institutions from around the world. A partial list of the basic equipment available for biological experimentation of cells and tissues aboard the ISS include:

• Life Sciences Glovebox (LSG; biosafety cabinet)

• Refrigerators/freezers

• Animal housing facilities for small mammals

• Centrifuges

• Incubators

• Growth chambers

• Greenhouses

• Microscope

• Aquariums

• UV spectrophotometers

Specialized equipment developed by commercial partners and NASA’s Ames Research Center and provided to academic research groups have been designed to facilitate biological research in microgravity environments. These devices include the Bioculture System, Space Automated Bioproduct Laboratory (SABL), BioCell, Microscope Platform, PCR analysis equipment (Wetlab-2), and DNA sequencer.

BioCulture System

The BioCulture system was developed by NASA’s Ames Research Center and Tissue Genesis, Inc. to replace the Cell Culture Module that was originally designed for use in the Space Shuttle Program. The flight proven design and lessons learned were incorporated into a design that would be conducive to bioresearch studies in microgravity involving both stem cell and specialized cell lines. The system consists of a docking station, command module, gas supply assembly that houses 10 individually controlled experiment cassettes (Figure 4). Each cassette is a hollow fiber bioreactor and provides structural support, power, data, gas supply, incubator, and refrigerator compartments. The BioCulture system has since been used to support a wide diversity of tissue, cell, and microbiological cultures and experimental methods [6].

FIGURE 4. The BioCulture System and one of the ten hollow fiber bioreactor cassettes designed for cell culture studies within the Bioculture System.

Each cassette provides the structural support, power, data, gas supply, incubator and refrigerator compartments. Photo courtesy of NASA.

Academic and commercial researchers use the Bioculture System to study a wide range of biological processes in microgravity that are relevant to human health. These experiments have delivered a greater understanding about how gravity affects the physiology, biochemistry, genetics and gene expression of living cells, tissues, and microbes for purposes of drug discovery, countermeasure analyses, or to study infectious disease processes [30]. Studies of tissue engineering, regeneration, and wound healing are also possible applications of this system (Figure 5).

FIGURE 5. An astronaut-scientist conducting cell culture studies by manipulating the hollow fiber bioreactor within the Life Sciences Glovebox (LSG).

Photo courtesy of NASA.

Space Automation Bioproduct Laboratory (SABL)

The Space Automation Bioproduct Laboratory designed and built by Bioserve Space Technologies, Inc. is a dual function incubator/freezer that supports space life science experiments on the ISS for conducting cell culture and other biological experiments. The instrument provides advanced incubator technology, and active CO₂ in support of mammalian cell culture. Three units have been installed aboard the ISS.

BioCell

Bioserve Space Technologies, Inc. also designed cell culture hardware to function aboard the ISS in place of cell culture flasks or multiwell culture plates. The BioCell meets the strict NASA guidelines for safety and biological containment. The devices are compatible with the microscopy platform and the plate reader components. The Biocell supports fluid injections, media exchanges, fixation, and culture preservation.

BioServe Microscope Platform (Figure 6) [31]

FIGURE 6. NASA astronaut K Rubins at the microscope platform aboard the ISS.

She is studying the effects of spaceflight on human induced pluripotent stem cell-derived cardiomyocyte structure and function [31]. Photo courtesy of NASA.

A Nikon Eclipse TS100 professional inverted microscope allows for bright field and phase-contrast microscopy and is capable of providing full high-definition Imaging. The microscope is equipped with high quality objectives ranging in power from 2× to 40×. Additional objectives and other add-ons can be flown to the ISS on a per-experiment basis.

Polymerase Chain Reaction (PCR) analysis (Wetlab-2)

Wetlab-2 is a research platform for conducting real-time quantitative gene expression analysis aboard the ISS. This facility enables spaceflight genomic studies involving a wide variety of biospecimen types in the unique microgravity environment of space. WetLab-2 was developed at NASA’s Ames Research Center and enables the traditional use of quantitative PCR, such as measuring gene transcription or rapid detection of gene targets that indicate infectious disease, cell stress, changes in cell cycle, growth and development, and/or genetic abnormality. The Wetlab-2 facility includes a commercial PCR instrument (Cepheid SmartCycler^®) that can perform up to 16 PCR reactions in parallel, a sample transfer tool for retrieving samples from culturing hardware and a set of fluidic modules to enable sample preparation work. Researchers working in a weightless environment can use the full facility to produce quantitative PCR information or extract RNA from their samples for analysis on the ground or by other facilities available on the ISS.

DNA sequencer (MinION)

Sequencing is a technology that addresses several critical spaceflight needs: infectious disease diagnosis, population metagenomics, gene expression changes, and accumulation of genetic mutations. Based on size, power, and ease of use considerations, the MinION^™ DNA sequencer (Oxford Nanopore Technologies, Oxford, UK) was the most spaceflight-ready of commercially available sequencers. This device sequences DNA and RNA by measuring current changes caused by nucleic acid molecules passing through protein nanopores embedded in membranes; the change in current is diagnostic of the sequence of the DNA or RNA occupying the pore at a given time [32].

PUBLIC–PRIVATE PARTNERSHIPS IN FOSTERING OUTER SPACE INNOVATIONS IN BIOTECHNOLOGY AND BIOPRODUCTION

The recognition that space is a catalyst for economic growth and that commercialization will populate LEO and beyond with human activity, a number of public-private research and development partners have been established focused on broadening our biological knowledge of cells and tissues in space environment [24,33]. These partnerships serve to stimulate open-source research improving intellectual activity and productivity through input sharing, labor pooling and cross fertilization of ideas and knowledge in a concept known as ‘agglomeration externality’ [33]. Collaborations in the area of biotechnology and bioproduction have investigated how sustained microgravity influences cellular behavior including pluripotency, multipotency, cell division, cytokine and growth factor secretion, differentiation, cell to cell interactions, tissue development and regeneration, aggregate interactions in the context of the whole organism, and changes to stem cell proliferation rates [34]. These collaborations have produced an enormous amount of information leading to a greater understanding of stem cell properties and cell behavior in microgravity. Major medical centers have participated in public–private partnerships that are focused on space-based programs for stem cell science [24,31,35–40]. Among the institutions participating in these partnerships includes such prestigious medical centers as: Cedars-Sinai Regenerative Medicine Institute, Stanford University Consortium for Regenerative Medicine, the University of California at San Diego, Center for Regenerative Biotherapeutics and Department of Laboratory Medicine and Pathology, Mayo Clinic, Loma Linda University School of Medicine, New York Stem Cell Foundation Research Institute, Department of Molecular Medicine Scripps Research Institute and Emory University School of Medicine and the Children’s Healthcare of Atlanta and Department of Biomedical Engineering, Georgia Institute of Technology. The investigations into human stem cell science in space undertaken by these organizations are relevant to the study and treatment of human disease on earth.

A number of large pharmaceutical manufacturers have also been committed to space-based research programs, many conducting research on board the ISS. Pharmaceutical developers currently investing in space research include the following:

• AstraZeneca: nanoparticle drug delivery systems for therapeutic cancer vaccines

• Bristol Myers Squibb: protein crystallization

• Merck and Co: monoclonal antibodies (pembrolizumab, Keytruda^®) as crystalline suspensions to enhance drug delivery. Protein crystal growth is smaller, much purer and consistent leading to lower viscosity with better injectability

• Gilead: increase COVID-19 therapeutic remdesivir to improve drug efficiency and reduce risk profile

• Amgen: preclinical trial of two osteoporosis drugs, Evenity^® and Prolia^®, on mice in microgravity

• Eli Lilly and Company: dosed mice with a muscle boosting antibody before their trip to the ISS and found that the treatment pre-empted the atrophying effect of microgravity on muscles

• Schering-Plough Research Institute: microgravity experiments on alpha interferon, Intron A^®, produced large quantities of high-quality crystals

To assist these organizations in deploying their experimental packages for execution in the ISS, a number of commercial companies have been focused on developing or modifying the scientific tools in accordance with NASA’s design and safety specifications to ensure compatibility with the systems onboard the ISS. These organizations include Space Tango, Axiom Space, Sierra Space, BioServe Space Technologies, and Redwire Space. Examples of those commercial entities with significant commitments to the development of therapeutic modalities and the biological and pharmaceutical development of manufacturing platforms in LEO include:

• Varda Space Industries: collaborating with pharmaceutical companies to improve their drugs and develop therapies by taking advantage of the unique properties of space and then returning those materials to earth

• Axiom Space: as an example of a commercial entity having sustained presence in space. Exclusive access to a module of the ISS was awarded by NASA. Axiom Space is creating an innovation platform for the in-space production of advanced materials and biomedical products that support the development of a robust commercial economy in LEO and beyond

• BioServe Space Technologies: has been designing and developing space flight certified equipment for over 34 years. Affiliated with the University of Colorado. BioServe has expertise in cultivation of mammalian cell and tissue culture, tissue engineering, organoids, bioreactors, and organ-on-a-chip technologies. Most recently, developing the ‘BioServe In-space Cell Expansion Platform’ or BICEP which is currently under evaluation aboard the ISS

• Space Tango: is focused on the design, certification, and operation of systems across space platforms, automated data collection and space manufacturing. Projects ongoing within the ISS have included layer-by-layer deposition, development of stem cells, tissue chips, organoid manufacturing, and 3D bio-printing platforms. Their activity is focused on agility, automation and reusability and minimal reliance by crew for all space platforms using their standardized CubeLab hardware

• Eascra Biotech: in-space fabrication of DNA-inspired Janus base nanomaterials (JBNs) for RNA therapeutics and cartilage tissue repair achieving the product development of a Technology Readiness Level (TRL) 7 and a Market Readiness Level (MRL) 5. Successful completion of these initial studies will provide the foundation for continued development of JBN technology development that has the potential to provide significant benefit to the industry and patients across a wide variety of therapeutic applications

• MicroQuin: crystallization of transmembrane proteins which regulate a cell’s internal environment and its eventual death. Awarded ISSNL and Boeing funded MassChallenge accelerator program to crystallize transmembrane proteins

• LamdaVision: fabricated artificial retinas intended to restore vision in people who are blind takes advantage of microgravity to deposit atoms-thick protein films on a polymer membrane

• Angiex, Inc.: treatment targeting the blood supply of tumor cells, which kills cancer cells by depriving them of oxygen and nutrients. The company’s Angiex cancer therapy investigation takes advantage of the space station’s microgravity environment to culture endothelial cells, which line the walls of blood vessels, to see whether they might provide a valid model to help develop safer and more cost-effective cancer treatments

• Neuronix, sponsored by the ISSNL, demonstrates the formation of 3D neuron cell cultures in microgravity and tests a neuron-specific gene therapy. Gene therapy shows promise as a potential treatment for people with paralysis and neurological diseases such as Alzheimer’s and Parkinson’s, but the 3D models needed to test these therapies do not form in Earth’s gravity. Creating 3D cell cultures in microgravity could provide a platform for drug discovery and gene therapy testing

Government organizations involved in space based biological research to support the long-term goals of space exploration and commercial development of space include:

• NASA—In Space Production Applications (InSPA) portfolio (advanced materials, tissue engineering, and biomanufacturing):

• Technology Readiness Levels (TRL): a type of measurement system used to assess the maturity level of a particular technology. Each technology project is evaluated against the parameters for each technology level and is then assigned a TRL rating based on the project’s progress. There are nine technology readiness levels. TRL 1 is the lowest and TRL 9 is the highest

• Marketing Readiness Levels (MRL): Market Readiness Level refers to how ready the product or service is to be taken to market as a commercial offering for a group of customers. MRL frameworks tend to include an idea about a perceived need in the market-to-market leader. MRL 1 is the lowest level and MRL 9 is the highest level

• Space Biosciences Division, NASA Ames Research Center, Moffett field, CA Studies of Somatic/Embryonic Stem cells for long duration Space flight

• ISSNL (Center for Advancement of Science in Space (CASIS) (2024 fiscal year budget—$5 million for NASA to pursue cancer related research on the ISS) The Center for the Advancement of Science in Space^™ (CASIS) is the non-profit organization that manages the ISSNL receiving at least 50%of the US research allocation on the ISS to facilitate research that benefits humanity. NASA manages the other 50% and focuses on research for space exploration purposes

One public–private partnership recently established has been the launch of the Astrobiotechnology Hub, a consortium consisting of academic, industry and government participants [41]. Under the aegis of the Sanford Stem Cell Institute of the University of California, San Diego, this group is focused on translating the basic research findings of stem cell biology in space. By taking advantage of the properties presented in LEO, this consortium coordinates clinical trials and develops commercial products through biomanufacturing of novel drugs, biofilms, and stem cell therapies.

BIOMEDICAL APPLICATIONS AND RESEARCH OBJECTIVES UNDER STUDY AT THE ISS

Long-term space missions will expose crew members, their cells, as well as their micro-biomes to prolonged periods of microgravity, ionizing radiation, and environmental stressors for which almost no earth-based organism have evolved to survive [7]. Applications for stem cell research, tissue engineering and regenerative medicine are being advanced within the environment of microgravity [6,42]. The objective is to create new business models that would attract capital investment for a robust commercial use of LEO as proposed by the ISSNL with the following objectives:

• Exploit the benefits of stem cell research in the microgravity environment for therapeutic applications on Earth

• Demonstrate an organoid or multicellular culture system to model human diseases that can be used for testing therapeutics

• Develop or leverage existing systems on the space station for the production of tissues or other biocompatible materials for regenerative medicine

Requests for investigative programs have been issued to focus on stem cell properties, tissue chips, organoids, and 3D bio fabrication [42,43].

Physiological effects of microgravity on stem cell biology

Pluripotency is the ability to transform induced pluripotent stem cells (iPSC) into tissue cells. Earth’s gravity causes challenges of maintaining pluripotency of iPSC during their production including expansion and growth of cell populations [6,39]. On earth, 2D cultures conditions do not entirely recapitulate the native environment of the human body. However, in microgravity it appears that 3D cell growth more closely resembles how cells grow within the human body [6,39,43–45].

Exposure to microgravity also causes significant mechanical unloading of mammalian tissues, resulting in rapid alterations of their physiology, which poses a significant risk for long-duration manned spaceflight [6]. The immediate degenerative effects of spaceflight understood best are those studied during short-term LEO experiments, and include rapid microgravity adaptive bone and muscle loss, loss of cardiovascular capacity, defects in wound and bone fracture healing, and impaired immune function. Over the long-term, exposure to microgravity may cause severe deficits in mammalian stem cell-based tissue regenerative health, including osteogenesis, hematopoiesis, and lymphopoiesis, as well as significant stem cell-based tissue degeneration in amphibian tail and lens regeneration [6]. In 2013, the ISSNL issued a Request for Information for partners interested in conducting stem cell research in a microgravity environment. The goal of the request was to leverage a LEO-based platform to gain insights into the control and optimization of stem cell pluripotency and multipotency, proliferation and expansion, genomic and epigenomic integrity, differentiation, and maturation [6,24].

Studies would be performed to enhance the growth of large amounts of safe and high-quality clinical grade stem cells with minimal cell differentiation and to evaluate the feasibility of successful harvest and transport of the space expanded stem cells back to earth (Figure 7) [6,39,43–45]. Experiments would be conducted and protocols standardized in accordance with the International Society for Stem Cell Research (ISSCR) to encourage the growth of stem cells in space for patient use on earth and to ensure the safety and efficacy of space produced stem cell therapeutic modalities [34]. Goals for this research would be focused on improving astronaut health for long duration space travel but also to gain insights into developmental biology of stem cells and their potential use in disease modeling or drug screening. The research would also be geared to developing approaches for high-throughput biomanufacturing of stem cell therapies, capability of working autonomously and remotely, employing miniaturization, microfluidics, robotics, machine learning, and artificial intelligence [2].

FIGURE 7. Testing the effects of microgravity on stem cell culture and downstream applications.

A better understanding of human disease and improvements in clinical therapies and biomanufacturing are emerging from parallel cell culture studies conducted in microgravity and on earth. Reproduced with permission from Arzt et al. [35].

The types of stem cells that have been studied in space have included mesenchymal stem cells (MSC), hematopoietic stem cells (HSC), cardiomyocytes derived from induced pluripotent stem cells (hiPSC-CM), cardiovascular progenitor cells (CPC), and neural stem cells (NSC) [37]. However, over the past quarter century many more types of stem cells and specialized cells have been the focus of innumerable experiments by academic centers (Box 3) [24,31,34–40,43]. Categories of stem cell research and disease entities conducted in LEO are shown in Box 4.

BOX 3. Studies of stem cells and specialized cells conducted in LEO [24,31,35–40,43].

Stem cell type

• Mesenchymal stem cells (MSC)

• Hematopoietic stem cells (HSCs)

  • Cardiomyocytes derived from induced pluripotent stem cells
  • Cardiac progenitor spheres derived from induced pluripotent stem cells
• Cardiovascular progenitor cells (CPCs)

  • Neural stem cells (NSCs)
  • Embryonic stem cells
  • Pig fetal liver stem cell line (PICM-19)
  • Cancer stem cells (CD133⁺)
  • Oligodendrocyte progenitors

Specialized cells

• Retinal pigmented epithelia (ARPE-19 cells)

• Human aortic smooth muscle cells (HASMCs)

• Human cardiomyocyte line (AC16)

• Endothelial cells (EA.hy926)

• Human dermal microvascular endothelial cells (HMEC-dBL)

• Human microvascular endothelial cells (HMEC-1))

• Peripheral blood mononuclear cells (PBMCs)

• T cells (CD8 and CD4)

• Lymphocytes

• Neutrophils

• Monocytes

• Dendritic cells (DC)

• B cells (CD19 and lymphocyte depleted (LD)

• M1 and M2 Macrophages

• Primary T cells

• Primary macrophages

• Human chondrocytes

• Meniscus fibrochondrocytes

• Thyroid cells (FRTL5)

• Primary human dermal fibroblasts

• Primary skin tissue from C57BL/6J Mus musculus mic

BOX 4. Categories of stem cell research and disease entities conducted in LEO [42].

Mesenchymal stem cells (MSCs)

• MSCs grown in space maintain their morphology, phenotype, and proliferation capabilities

• Enhanced immunosuppressive properties were observed

• Microgravity may inhibit differentiation, preserving the stemness of MSCs, which is beneficial for clinical applications

• Space-grown MSCs could be used to treat central nervous system diseases, such as spinal cord injuries, due to increased neural development markers in MSCs grown in microgravity

Hematopoietic stem cells (HSCs)

• Microgravity affects HSC proliferation and differentiation, with an observed preservation of stemness

• Space-grown HSCs showed suppressed erythropoiesis and increased macrophage differentiation

• Applications include potential therapies for anemia and other blood-related disorders

Cardiomyocytes derived from induced pluripotent stem cells (iPSCs)

• iPSC-derived cardiomyocytes grown in space demonstrated structural and functional integrity

• Microgravity enhanced gene pathways related to mitochondrial function and calcium signaling

• Applications include advanced cardiac repair therapies and models for studying spaceflight-induced cardiac remodeling

3D models of the human brain derived from iPSCs

• Complex human models containing iPSC-derived neurons, astrocytes, oligodendrocytes, and microglia can be used for disease modeling and drug discovery

• Applications focus on regenerative therapies for neurodegenerative diseases like Alzheimer’s and Parkinson’s

Cardiovascular progenitor cells (CPCs)

• CPCs in space displayed increased DNA repair capabilities and enhanced differentiation into cardiac tissues

• Applications focus on cardiac regeneration and repair through enriched and functional cardiomyocytes

Recently, to address the on-earth limitations and challenges of expanding umbilical cord blood derived stem cells, in-space expansion of hematopoietic stem cells is being evaluated for its technical, economic, and commercial capabilities [46]. Specialized bioreactor technology designed and deployed to the ISS is being evaluated for its ability to expand hematopoietic stem cells collected on Earth from umbilical cord blood or adult mobilized peripheral blood hematopoietic stem cell populations, cryopreserved and transported to future commercial space platforms. Once expanded in microgravity stem cell products can be cryopreserved and returned to earth for clinical use. The novel spaceflight culture system designed by BioServe Space Technologies is termed the ‘BioServe In-Space Cell Expansion Platform’ (BICEP). The hope is that hematopoietic stem cells expanded in space can improve upon the quantity, quality, cell type distribution, genetic stability, function, and clinical safety of earth produced stem cell products.

The BICEP technology has the capability of multiple bioprocessing functions including thawing of cryopreserved cells, seeding into specialized cells to initiate cellular expansion with media supply and control fluids. Cells are incubated at 37 °C in an atmosphere of 5% CO₂ after which the expanded cells are harvested after 10 days, cryopreserved and returned to earth [46].

TISSUE ENGINEERING AND REGENERATIVE MEDICINE

Tissue engineering efforts in space has long been a major goal. Before the advent of the ISS, 3D constructs of chondrocytes on scaffolds comprised of polyglycolic acid were grown in bioreactors aboard the Russian Mir space station [47]. The environment in space yielded cartilage constructs composed of viable cartilage cells expressing proteoglycans and type II collagen, markers for hyaline cartilage [47]. The shape, structure, composition, and function of these cartilage constructs produced under conditions of microgravity were consistent with those grown on earth. The utilization of space and the absence of gravity facilitates the rapid maturation and acceleration of cell growth mimicking the aging process allowing investigators to study changes in cells due to aging.

Tissue engineering in microgravity has been given special emphasis [11,48,49]. This unique environment is seen to allow for the production of delicate tissue constructs through bio fabrication. The environment of space can facilitate the maturation and strengthening of growing 3D tissues without collapsing into less useful 2D forms as gravity would produce on earth. These characteristics have been exploited by a number of organizations who wish to improve patient care and to better understand cell behavior in order to advance regenerative medicine [11,49].

Terrestrial-based cell culture techniques face limitations in the complexity and consistency of cell systems that can be developed. Without the shearing and sedimentation forces present on earth, microgravity allows creation of larger, complex, and more delicate tissues, such as blood vessels, which can enable the formation of sophisticated organoid systems [11,48]. While tissue engineering has many potential applications, efforts often face limitations in culturing tissues resembling those in the body, advancing the research into organ growth, or attaining higher accuracy to validate personalized drug testing.

Tissues grown on earth are constrained by gravity which results in flattening and deformation of 3D constructs. However, those grown in an environment of microgravity offer specific advantages. For example, larger tissue constructs are allowed to form without special restriction into 3D structures termed spheroids. Moreover, evidence suggests that tissues grown in space can elicit a response similar to the aging process. This characteristic can accelerate drug development and disease modeling of cellular function [50].

Organoids are self-organizing 3D aggregates of cells differentiated from stem cells and whose spherical shape and cellular structure may resemble full organs [50,51]. These organoids or spheroids in a tissue culture environment can serve as simplified organ systems which can be used for accurate and scalable disease modeling and drug testing investigations (Figure 8) [52]. Organoids can also be used as tissue batches for regenerative medicine applications. The integrated biological function in organoids serves as a powerful model of human disease states, and applications of this kind of these advanced in vitro systems could enable a wide variety of experiments conducted in microgravity. For example, experiments have been conducted with stem cell derived brain and neural organoid models by the National Stem Cell Foundation to better understand mechanisms behind neurodegenerative diseases such as Parkinson’s and primary progressive multiple sclerosis [38]. There may also be applications for use in personalized medicine and potentially address the shortage of organs for transplantation [50].

FIGURE 8. The dynamics of cell formation into spheroids in an in vitro model of tumor metastasis under conditions of microgravity are presented in this schematic.

When gravity is restored, spheroids will reattach to a substrate. Reproduced with permission from Grimm [52].

Culturing stem cells and progenitor cells in microgravity stimulates proliferation as well as preserving ‘stemness’, helping to maintain population numbers in culture [48]. In addition, microgravity forces cells to interact and anchor to each other promoting the development of tridimensional cultures, producing larger and wider cell clusters with higher order structures (Figure 8) [51,52]. Microgravity therefore could be a platform to optimize conditions for large-scale production of organoids and spheroids for research, regenerative medicine applications, and pre-clinical testing of drug candidates [50].

3D bio-printing

Tissues expressing specific architecture such as muscles, vasculature, nerve tissues and heart valves can be created through 3D bio-printing [53]. The 3D Biofabrication Facility (BFF) and Advanced Space Experimental Processor (ADSEP) developed by Redwire Space and installed aboard the ISS worked on creating knee meniscus, muscle, vasculature, and nerve tissues [54,55]. Heart valves can be created as microgravity allows tissue to retain its shape [54,55]. In fact, a bio printer developed in Finland by Brinter AM Technologies is currently under modification by Redwire Space to meet the stringent requirements of compatibility with systems aboard the ISS. In support of future long duration deep space missions these devices would have the capability of producing replacement damaged tissues when access to earth bound medical facilities would be impossible. These devices would also enhance our knowledge of the biological mechanisms of tissue regeneration and aging.

On earth, gravity constrains engineered tissue by deforming and flattening 3D constructs while in microgravity cells are able to form complex 3D structures without the need for structural support. These structures are similar to tissues naturally found in the human body and facilitates the study of accelerated disease modeling, cell behavior, especially the effects of aging, and may have a role in advancing regenerative medicine and testing the effects of new drugs. Without the need for scaffold matrixes a variety of mechanisms can be applied in space to produce soft human tissues, such as blood vessels [54,55]. Larger tissues may be constructed by utilizing biofabrication capabilities that enable the production of 3D structures [54,55]. These technologies can pave the way for the development of therapies for repair or replacement of damaged tissues and organs [53]. Long-term success of biofabrication may enable potential medical breakthroughs, including the creation of patient-specific replacement tissues or patches and could ultimately help reduce the current shortage of donor organs.

More recently, the Wake Forest University School of Medicine’s Institute for Regenerative Medicine (WFIRM) made significant advancements in the innovations for constructing biological models containing vascular tissues. Using 3D bio-printing technology these investigations were able to mimic vascularized liver tissue constructs [56,57]. NASA has selected Wake Forest Institute for Regenerative Medicine (WFIRM), of Winston-Salem, North Carolina, for a Phase 1 program In Space Production Applications (InSPA) award that takes advantage of the microgravity environment of space to develop and validate a platform and strategy for manufacturing vascularized and perfused liver tissue [56]. The proposed work will leverage microgravity for manufacturing clinical scale liver tissue constructs with intrinsic vascular networks that allow perfusion and integration into the recipient’s peripheral circulation for the treatment of liver disease [57].

3D tissue chips

Tissue chips are small devices, similar in size to a USB drive, engineered to grow human cells on an artificial scaffold to model the structure and function of human tissues and organs [9,58]. Also termed ‘microphysiological systems’ (MPS), tissue chips are constructed using human cells organized in a predetermined architecture and are designed to replicate facets of the physical environment cells experience inside the body, providing higher accuracy models that can lead to advancements in predictive medicine and personalized healthcare. By better replicating the complexity of the tissue architecture, tissue chips are also leveraged for therapeutic screening for multiple disease indications [9,59]. In microgravity, tissue chips have the potential to capture accelerated disease conditions on a complex tissue level to advance our understanding of the mechanisms behind disease progression, thereby increasing opportunities for drug development. Tissue chip devices provide a more financially accessible alternative to convention in vivo models with precise control over microenvironmental cues (e.g., shear stress, oxygen gradients, biomolecule delivery). This democratizes access for advanced screening models with even higher specificity and the opportunity for integrated monitoring systems for complex analysis. Tissue chip applications have been utilized by a number of investigational groups collaborating with the ISSNL to advance the study the effects of microgravity upon the human body including blood–brain barrier function mechanisms, accelerated deterioration of muscle, bone loss representative of osteoporosis, decreased cardiopulmonary function, and immune deficiency, all of which have been observed and documented in space [59–62]. Use of tissue chip applications for these studies can accelerate the understanding of ageing while revealing targets that potentially can reverse these processes. Three examples developed and placed on test within the ISS have been ‘heart on a chip’, ‘tumor on a chip’, and ‘cartilage on a chip’.

Heart on a chip

Cedars-Sinai investigations have pioneered new technology to test chemotherapies and other cancer drugs for heart toxicity. Cardiomyocytes and vascular endothelial cells derived from induced pluripotent stem cells can screen for drug-induced alterations in cardiovascular cell function and survival. Specialized 3D chips containing these cell types are enclosed in separate chambers but are connected with channels that allow the introduction of fluids and facilitate the interaction of the cells. These unique test systems allow the formation of mature heart muscle cells and vascular cells which together form a test platform for precise drug toxicity studies [63].

Tumor on a chip

Another NASA funded organization, Encapsulate, Inc. developed an automated ‘tumor on a chip’ which allows cancer cells to be evaluated for their response to chemotherapeutic agents prior to their administration to patients [51]. In this way the most effective chemotherapeutic agent can be selected for a patient’s specific cancer. Again, challenging these systems in microgravity facilitates the study of cancer cells since cells form 3D structures which more closely resemble the growth and behavior of cells within the human body [51]. The biomimetic microenvironment of the tumor within the chip in an automated system can control cell growth, maintenance, and be accurately monitored simulating the environment of the tumor within the human body [51].

Cartilage on a chip

A physiologically relevant joint model was successfully reproduced and tested in a microgravity environment. Viable and reproducible human cartilage, bone, and synovium cultures were generated [58,62,64]. This resulted in a reproducible baseline for one orthopedic condition, post traumatic osteoarthritis. With this ‘cartilage on a chip’ innovation treatment effects of drugs used to treat inflammation, and pain can more accurately be assessed. Cartilage repair strategies can also be assessed in space. This technology can be utilized on earth for treatment of post-traumatic osteoarthritis (PTOA) in athletes, a common medical problem especially in female athletes.

In an effort to extend the longevity of MPSs to a minimum of 6 months NASA is collaborating with the US FDA, National Institutes of Health (NIH), and the Biomedical Advanced Research and Development authority (BARDA). This extended lifespan would allow researchers to investigate the effects of acute and chronic stressors in a spaceflight environment and allow for longer duration studies to better assess:

• Disease models

• Drug development

• Clinical trial designs

• Chemical and environmental exposures and countermeasures

• Physiological changes due to the spaceflight environment [65]

Gene therapy

The ISSNL has sponsored opportunities to utilize the microgravity environment as a platform for the development of gene therapies [66,67]. The first gene therapy and investigational ophthalmic therapy has been pioneered by Oculogenex, Inc. in the testing of a novel gene therapy to prevent and possibly even reverse vision loss from age related macular degeneration (AMD), a leading cause of blindness in older adults [66,68]. This gene therapy technology addresses the root cause of dry macular degeneration by targeting the epigenetic switch which plays a fundamental role in retinal homeostasis on the mitochondrial enhancement of a cellular response to oxidative stress [66,69]. This approach restores the functionality of damaged cells and prevents senescence and death of retinal cells. Partially funded by NASA because astronauts can be afflicted with spaceflight associated neuro-ocular syndrome, forty female mice treated with the gene therapy were sent to the ISS with an equal number treated and remaining on earth. Exposure to microgravity by the gene therapy treated mice will accelerate the oxidative stresses that encourage the onset of AMD.

Axonis Therapeutics is developing a neuroregeneration gene therapy designed to silence the expression of PTEN, an inhibitory protein that suppresses the ability of axons to regrow after injury. The gene therapy was targeted to central nervous system (CNS) neurons only since the gene also plays a role in the suppression of growth in other non-neural cells [70]. Using an AAV as a viral vector and tailoring the vector’s gene promoter only CNS neurons would be targeted. By deleting the expression of PTEN, the CNS neurons are reprogrammed back into a state of growth to allow regrowth of damaged axons. The model was successfully tested in rodent models. Aboard the ISS, experiments were designed to exploit microgravity and create a 3D model of the human brain by co-culturing iPSC derived mature neurons and astrocytes and forming brain organoids [67]. This achievement would result in a CNS model in order to test the gene therapy. Without an artificial matrix substrate and other growth factors creating similar models on earth would be difficult.

When vials of the mature neurons, astrocytes and AAV vectors sent to the ISS were combined along with a fluorescent protein gene for microscopic visualization the astronauts would be able to evaluate the rate of self-assembly of the brain organoids culture. After 72 hours the neurons and astrocytes were seen to self-assemble into 3D organoids. Importantly, the functions of the neuron specific AAV gene therapy vector in suppressing the PTEN protein could also be visualized. The successful demonstration of the gene therapy would now be used to justify the chemistry, manufacturing and control elements needed to plan and execute clinical investigations [66].

NANOMATERIALS

Nanomaterials hold a strong potential for a number of therapeutic applications [71–74]. The FDA has approved a number of nanomedicines to include therapeutics for cancer, skin conditions, and regenerative medicine [75]. Janus Base Nanomaterials (JBNs) are noncentrosymmetric monomers that possess the Watson-Crick hydrogen bond ‘donor-acceptor’ motifs present in DNA. Mimicking DNA base pairs, a family of JBN monomers achieve controlled self-assembly at ambient temperatures to form a collection of 2D nano-rosettes which further assemble into 3D nanotubes. Each JBN monomer is 400 Da and the self-assembly process is achieved by suspending thousands of monomers in aqueous solution without the need for catalysts or cross linkers. The final resulting Janus Base Nanotube (JBNt) maintains supramolecular helicity and stability through inter-rosette hydrogen and π-π bonds. When self-assembled in the presence of biomolecules such as proteins, the resulting structure includes a biocompatible and mechanically robust nano matrix (JBNm) that mimics natural extracellular matrix [76]. Because of the characteristic of self-assembly Janus base nanomaterials are seen as ideal for in-space manufacturing.

As an adjunct to tissue regeneration and cartilage repair the in-space fabrication of JBNms was recently demonstrated [77,78]. When creating JBNm on Earth, gravity-driven sedimentation can limit the assembly process. However, when manufacturing JBNm in microgravity, the lack of these forces allows for increased homogeneity and bioactivity (Figure 9). Scientists at the University of Connecticut’s Nanomedicine Laboratory and Eascra Biotech have embarked upon studies aboard the ISS to evaluate this therapeutic nanomaterial; their goal is to overcome the decay of cartilage caused by the effects of microgravity and lack of mechanical loading that affect astronauts living and working in space especially on long duration missions [76–78].

FIGURE 9. Transmission electron microscope images of JBNm strands manufactured in space and on earth.

The width of the JBNm bundles manufactured in space were significantly larger than those manufactured on Earth. In space JBNs demonstrated improved homogeneity and scaffold assembly, increasing cell bioactivity indicating low toxicity with high biocompatibility. The mission supported methodology for manufacturing nanomaterials in space and successfully demonstrated the promise of utilizing microgravity for improved JBN assembly and bioactivity [77].

Cartilage damage, whether through trauma or arthritis continues to be an unmet medical need on earth as well as in space [79–81]. Articular cartilage defects are seen in 60–66% of knees undergoing arthroscopy and osteoarthritis is a chronic and debilitating joint disease affecting over 600 million individuals worldwide over the age of 40 [82]. Natural hyaline cartilage, when damaged, has a limited ability for self-repair due to the absence of pluripotent cells, a sparse distribution of chondrocytes, no lymphatic drainage or nerve distribution and the lack of vasculature [79–81].

JBNms manufactured in space which utilize microgravity to minimize the effects of sedimentation were produced with improved homogeneity, low toxicity, and high cell biocompatibility [78]. This process can provide a better understanding of the disease mechanisms that promote cartilage degeneration in space and demonstrated a novel approach for improving tissue engineering of cartilage repair on earth.

FDA REGULATION OF IN SPACE BIOMANUFACTURING FACILITIES

In 1984, writing in the Food, Drug, Cosmetic Law Journal, Robert Altman reiterated that, biological products would be the “most promising drugs for production in space” [83]. To exploit the benefits of a microgravity environment and facilitate the separation of impurities in the drug production process, a goal of the space program was to place in LEO a fully functioning automated pharmaceutical manufacturing laboratory. To achieve this goal, Altman discusses the problems confronting industry and the FDA and its role in supporting space technology. At that time, in order to attract investment from the private sector legislation had been introduced into the US Congress to relax existing FDA regulations. However, these legislative actions were unsuccessful. Only two companies, McDonnell Douglas Astronautics Co and Johnson and Johnson expressed interest in commercializing LEO for drug production in space [83].

FDA had not previously considered its role in space technology and the impact new technology would have in its regulatory scheme. FDA had to consider if its regulatory mandate in protecting the public health would discourage private investment in space programs or if the challenges emanating from the research, development, manufacture, and processing of pharmaceutical products in outer space would impede the entrance of private industry into the space market.

Thirty-four years later those concerns continue to confront industry in its plans to commercialize outer space. In 2018, the FDA and NASA issued a Memorandum of Understanding (MOU 225-8-027) that establishes formal communication between the two agencies to “discuss providing technical expertise for planning, performance, or review in areas of mutual interest”. Under this MOU, the two agencies “seek opportunities to participate in collaborative efforts, in furtherance of their respective objectives and as permitted under appropriate statutory authority and applicable law, as resources permit to:

• Look for potential collaborative studies on the utilization of already funded FDA projects that would enhance NASAs medical risk reduction exploration research

• Encourage space related health research through the exchange of expertise, scientific and technical information, date, and publications

• Discuss providing technical expertise for planning performance, or review in areas of mutual interest, subject to program priorities and availability of fund and personnel

• Facilitate and enhance research and development activities by either agency, including distributing information on research opportunities such as NASA Research announcements

• Coordinate publicity of mutually reinforcing activities, publications, and research results

• Include representatives from FDA and NASA in workshops, including NASA’s Human research Program Investigator’s Workshop, working groups, seminars, and other related activities” [84]

In 2021, this collaboration intensified when NASA solicited science investigations from multiple government agencies, including FDA, to extend the longevity of 3D tissue chips and microphysiological systems for modeling acute and chronic stressors in astronauts during long duration spaceflight. FDA’s chief scientist, Rear Admiral Denise Hinton commented that:“FDA remains deeply engaged in identifying and fostering strategies that can bring alternative testing methods such as microphysiological systems to FDA for integration into the review process, collaboration with our partners in the public and private sectors has been critical to advancing our efforts in this area, particularly with respect to medical countermeasures.” [61]. In the view of FDA these technical innovations could also advance the way drugs can be investigated and reviewed by regulators. Captain Tracy MacGill, Director of Medical Countermeasures (MCM) Regulatory Sciences noted that, “We expect that extending the lifespan of the microphysiological systems will provide more relevant and predictive models, for example, this will enable us to look at the effects of drugs or other FDA regulated products over a longer duration in both normal cells and those with acute and chronic diseases, the research had the potential to provide a wider window into safety and efficacy of a variety of medical products.” [61]. The purpose of this NASA–FDA collaboration is to study a wide variety of biological changes including neurotoxic stressors, radiation exposure, and acute and chronic exposures to drugs that could result in unanticipated discoveries to improve the operational capabilities and medical status of astronauts as benefiting patients back on earth.

The regulatory implications of bio manufacturing in LEO are challenging, with fundamental regulatory and legal questions that will require answers and policy decisions in the not-too-distant future. In a March 13, 2024 article entitled, ‘Are FDA astronauts coming soon? Implication of the revolution in space based drug manufacturing’ the author poses the following questions:

• Will FDA need to recruit astronauts to inspect space based manufacturing facilities?

• What are the intellectual property opportunities and risk for space made drugs?

• Would a generic or biosimilar version of a space made drug also be required to be manufactured in space [85]?

The advantages and scientific rationale of space-based manufacturing have been identified through years of investigations by major pharma organizations conducted aboard the ISS over the last quarter century. For example, the quality and consistency of drug substances comprised of protein crystals have been shown to be of higher quality and consistency when formed in microgravity [86]. Removal of impurities is facilitated by manufacturing that can overcome the earth-bound effects of convection and sedimentation. Affordable access to space and the need for new and improved existing drugs have justified the current investment climate.

REMOTE REGULATORY ASSESSMENT (RRA) OF BIOMANUFACTURING FACILITIES IN LOW EARTH ORBIT

As a result of the COVID epidemic regulatory agencies have adopted criteria and procedures for remote regulatory assessments [87]. In recent publications FDA, through the Office of Study Integrity and Surveillance (OSIS), have developed a variety of surveillance tools and new oversight approaches formalized criteria for such assessments, although not considered on site GMP compliance inspections, but would rather support FDA’s review of marketing applications. The Remote Regulatory Assessment criteria are comparable to the format of an inspection and could be adapted for facilities operating in LEO [88]. These adaptations could include visits to on earth manufacturing facilities where inspectors could become familiar themselves with the on orbit equipment and operations. FDA inspectors could then request specific documentation and review the following:

• Records of specific lots or batches and product specific information, such as product quality reports

• Summaries of batches manufactured in LEO and their disposition

• Visualize electronic systems with Read-only access to electronic databases

• Standard operating procedures and records on quality systems

• Interview relevant staff

These reviews could take place at the company’s on earth location or remotely facilitated by interactive technological advancements on autonomous manufacturing, telemetry, internet connectivity, video conferencing platforms, screen sharing, remote livestreaming, or pre-recorded video of on orbit operations. Additional discussion of FDA personnel with pharmaceutical developers can improve upon design of the criteria by which FDA could gain access to on orbit manufacturing operations in order to assess regulatory compliance. Following such an assessment the FDA would not issue an FDA 483 (Report of Observations), but RRA observations would be shared in writing and discussed at a close-out meeting.

Despite the location of manufacture for drug substances, FDA will expect and rely upon its core principles of CMC (Chemistry, Manufacturing and Controls) [89]. Information will be reviewed at the Pre IND/IND stage to assess identity, strength, safety, quality and purity of the drug substance and drug product [89]. Sponsors would need to supply the preliminary critical quality attributes for the drug substance, along with the primary structure, control of starting materials, preliminary manufacturing process and controls for DS/DP, physical characterization data, assay/impurities test methods such as sterility testing and endotoxin methodologies, preliminary DP formulation, etc. A change to the drug development process that now includes a manufacturing facility site change to a low orbit location would require a comprehensive risk assessment with data that demonstrates comparability and process optimization resulting from an environment of microgravity [90].

Appropriate clinical trial designs may be modified if they occur in deep space locations including personalized clinical trials used to assess the safety and efficacy of ultra-rare diseases or Phase 1 trials which entail micro-dosing of a medication over a short time period. The appropriate clinical design that would be executed in space would have to be discussed at a pre-IND meeting [91].

TRANSLATION INSIGHT: THE ECONOMIC POTENTIAL OF BIOMANUFACTURING IN LEO

Economic potential forecasting of biomanufacturing in LEO in terms of revenue and growth have been assessed by a number of expert analysts [12,15,16,19,92–96]. One analyst predicts that by the year 2040 over $1 trillion of the global economy will move into space [20]. CNBC reported that in 2023 $12.5 billion was raised with 39 merger and acquisition deals made across the sector [97].

The cost to place a satellite into LEO aboard a Delta E rocket in the 1960s was $168,000/kg [98]. Today the cost to launch a satellite into LEO from a SpaceX Falcon Heavy rocket is approximately $1500/kg, about 30X less than the launch cost of a NASA Space Shuttle [99]. It is now estimated that the cost to launch SpaceX’s next vehicle, the Starship Rocket, can place a satellite into orbit for a cost per kilogram of $100 [94]. Space has now become economically feasible and a source of value and a return on investment for a number of business sectors including pharmaceutical development and biomanufacturing [11]. Harvard Business Reviews reported that in 2019, 95% of the estimated $366 billion in revenue earned in the space sector was for the ‘Space for earth’ economy—goods or services produced in space for use on earth [93]. Venture capital is flowing into the commercialization of space, $15 billion in 2021 according to the space consultancy BryceTech [95]. The retirement and deorbiting of the ISS will be replaced by an ever complex technical infrastructure being assembled in orbit by a number of commercial entities including fully autonomous orbiting drug manufacturing vehicles. In 2016, Axiom Space, Inc., was awarded a contract for exclusive access to a module of the ISS. This has allowed Axiom to build its own module for commercial activity on the ISS with plans to have it independently operated when the ISS is retired in 2030 [100].

Deloitte analysts predict that the commercial potential of LEO will require the following incentives to:

• Deliver lower cost, higher cadence human-related access to space

• Significantly increase down-mass (mass of materials returned from space) capacity and industrialization of on-orbit manufacturing operations

• Establish multiple on-orbit destinations for human-rated depot-centric, and other mission specific activities

• Better align the resources and complementary technical capabilities of public and private sector players

• Enable access to LEO and execute missions and activities at the speed of business [101]

For a quarter century, cutting edge research and development has been conducted in space aboard the ISS since its initial launch and construction in the early 2000s. Companies like Bristol Myers Squibb and Merck have used the environmental qualities of microgravity to optimize drug efficacy by improving crystal formulation [102]. While much of the medical advances being made in space have an initial focus on lunar colony residency or the 2–3 year commitment for a round trip journey to Mars, the innovations and discoveries will also have a direct effect on the improvement of healthcare on earth (Figure 10) [10,11].

FIGURE 10. A timeline for the biomedical and medical infrastructure of a life support system over the next decades of space exploration is proposed in this schematic.

Over the next decade, dominated by the construction of planetary habitats on the moon and, later on, Mars, it very likely that innovations in regenerative biomaterials, patches for skin damage repair, injectable systems and acellular therapies would eventually lead to the manufacture of tissue parts and functional organ replacements. Telemedical, robotic, and remote medical assistance capability will be developed and perfected over the next decades. Reproduced with permission from Lordachescu [11].

Numerous public-private partnerships have completed proof of principle studies for the 3D bio printing of tissues and organs, and manufacture of ‘organs on a chip’, stem cell cultivation, and creation of metabolically functional and vascularized heart tissue among others [9,34]. Knowledge of the blood brain barrier, immunoscenescence, pulmonary infection, cardiac dysfunction, post-traumatic osteoarthritis, proteinuria, kidney stones, and inflammation of the intestine and a host of other pathological conditions that might affect astronauts on long duration space flights are also important health problems for large segments of the human population.

Tissue substitutes, nanomaterials for tissue scaffolds and efficient drug delivery, hemostatic agents or biomaterials such as dental fillers will all be available for astronaut crews to manage tissue damage and medical emergencies in deep space. These innovations will also find uses on earth for military medical professionals, or emergency medical first responders in the nation’s emergency rooms [11].

New therapeutic applications and modalities for stem cell derived products that might include scaffolds and matrices, cell–cell and cell–matrix interactions, stem cell and tissue engineering and reprogramming, cellular immunotherapies, organoid development, cellular biomanufacturing, or system integration between biological components will require protocols and procedures to ensure safety and efficacy [39]. Regulatory review and approvals will have to be adapted to include the therapeutic translation using space as a manufacturing platform. Remote regulatory assessments to ensure appropriate chemistry, manufacturing, and quality control elements in accordance with regulatory compliance requirements must be present and verifiable, especially for their suitability in clinical trials as gene or cell therapies [34]. These new protocols and procedures will also be germane to the production and regulation of artificial tissues and organs in space [11].

Development of patient specific gene therapy will benefit from the knowledge that many cell types including induced pluripotent stem cells grow faster in space and that microgravity fosters natural 3D stem cell growth, mimicking the human body’s environment more effectively than earth-based 2D cultures. Space-based research also enhances our ability to understand differentiation, proliferation, and tissue regeneration. Biomedical applications will range from drug discovery to regenerative medicine, disease modeling, and biomanufacturing in space for clinical use on earth [2,3,103,104].

Continued research and validation will be needed to fully understand the complex effects of microgravity on cellular function [103]. The future decades of biomedical commercialization of space will provide the opportunity for the synergies of cell and gene therapy, tissue engineering and microgravity to offer the types of innovations that will transform treatment strategies for human healthcare and medicine on earth [103,104].