The 2022 International Society for Stem Cell Research (ISSCR) Annual Meeting: Celebrating 20 Years of Achievements

The 20-Year Celebration of International Society for Stem Cell Research

With an overload of excitement and full expectations, the 2022 Annual Meeting of the International Society for Stem Cell Research (ISSCR) took place last June in San Francisco. A special meeting for the following diverse reasons: the first in-person reunion since the outbreak of the COVID-19 pandemic, the launch of its first hybrid format with live virtual attendance across the globe, and the celebration of the society's 20th anniversary since its foundation back in 2002. With around 4000 members from more than 75 countries, ISSCR has played a substantial international role in numerous components of stem cell research, from ethics and regulation matters to ensuring truthful evidence and research integrity in the field.

In these 20 years, the society has witnessed the fast and exhilarating advancement of technologies and has been at the forefront of innovation. All thought, ISSCR has been true to its essence of keeping the focus on translational aspects of stem cell research with sustained support from and to the community, at the interface of academia and industry. And so has been reflected on the vast program put out for this year's edition of the Annual Meeting: seven plenary sessions, multiple track sessions following five main themes (New Technologies; Tissues, Stem Cells, and Regeneration; Cellular Identity; Clinical Applications; and Modeling Development and Disease), and around 900 posters exhibited in the Poster and Exhibit Hall or the Poster Theater through the virtual experience.

The 2022 edition brought together an inspiring combination of speakers from different professional backgrounds and presentations that covered a broad range of topics. Despite the diversity, certain keywords were frequently mentioned across the whole event such as organoids, neuronal reprogramming, disease modeling, and stem cells in the immune system (Box 1).

ISSCR President Melissa Little (reNew, Denmark/Murdoch Children's Research Institute, Australia) opened the meeting by highlighting the many achievements of the stem cell community and presented the awards for Outstanding Young Investigator “Dr. Susan Lim Foundation” to Jennifer Phillips-Clemins (University of Pennsylvania), and the ISSCR Public Service Award to Sean Morrison (UT Southwestern). Following, the Presidential Plenary initiated the presentations featuring speakers selected by the ISSCR President, which cover the future generation of researchers across the breadth of stem cell science as well as known, established leaders in the field.

The first speaker was none other than the founder and first president of the ISSCR, Dr. Leonard Zon (Boston Children's Hospital), who discussed hematopoietic stem cells (HSCs) in the zebrafish model. In the first part of his talk, he focused on the experiments done by his group in the last years studying clonal hematopoiesis. This disorder is very frequent during aging due to the accumulation of mutations, and it affects a large proportion of the elder population, leading to myelodysplastic syndromes and acute myeloid leukemia. For studying this type of cell dominance in their native niche, they developed Tissue Editing With Inducible Stem Cell Tagging Via Recombination (Avagyan et al, 2021). With this tool, it is possible to analyze the competition between normal or mutant stem cells in the same animal without the use of transplantation.

Single-cell transcriptional analysis revealed high expression of proinflammatory genes in mutant mature myeloid cells, converting the bone marrow into a highly inflammatory environment. In addition, they showed enhanced expression of anti-inflammatory genes in progenitor cells of the mutant clone. Overall, this autoregulatory loop between mature and progenitors appears to be a selective pressure needed for clonal expansion. Later, on Dr. Zon's talk, he discussed the interactions between macrophages and HSCs (Wattrus et al, 2022). Macrophages either help expand or remove hematopoietic stem progenitor cells (HSPCs) based on their stress levels during embryogenesis, determining how many HSCs will be present in the adult. This modulation is exercised by very specific interactions between the two cell types and regulates hematopoietic clonality.

The following presentation was given by José Polo (University of Adelaide, Australia): he presented his efforts in understanding human reprogramming using iBlastoids, a reprogrammed 3D model of a blastocyst generated in the laboratory (Liu et al, 2021). He explained the way up to achieving these structures, starting from the combination of single-cell transcriptomics with the reprogramming of fibroblasts to a primed or naive pluripotent state. By reconstructing the entire trajectory of induced pluripotent stem cell (iPSC) reprogramming, his group evidenced independent linear reprogramming trajectories into the human pluripotent states and found that epiblast, hypoblast, and trophectoderm signatures are upregulated during the process.

The following question was how all these cell types interact and communicate in a 3D model. They found that reprogrammed cells can associate and self-organize into spheroid structures, so-called iBlastoids, resembling a blastocyst, with a defined cavity and an inner cell mass, and single-cell transcriptomics confirmed all cell signatures. Importantly, the integration of these data with results coming from blastocysts and other blastoid models showed a high correlation and proper clustering. Finally, he described how iBlastoids can properly model a blastocyst since stem cell lines can be derived, they can model the peri-implantation process in vitro, and trophectoderm can progress into a differentiated trophoblast. These advancements in the field highlight how powerful this tool can be for developmental studies and disease modeling, among other applications.

Continuing the topic of 3D modeling during embryogenesis, Aryeh Warmflash (Rice University) talked about gastrulation, a slightly later stage of development after blastocyst, when the fundamental body plan is shaped. Once gastrulation is initiated, cell fate decisions are mainly controlled by the signaling cascade of three morphogens: BMP, WNT, and NODAL. However, there is still a lack of understanding of how these signals are organized in space and time and how the pluripotent cells of the epiblast execute cell fate decisions. Importantly, the systems for studying these questions in humans are limited. His group has engineered micropatterning systems with human pluripotent stem cells (hPSCs) that recapitulate aspects of gastrulation by controlling the geometry of culture conditions in which cells differentiate into the extraembryonic, pluripotent, and primitive streak fates.

Warmflash and colleagues have shown that BMP signaling is sustained only at the colony edge, that this, signaling triggers an expanding front of WNT, and that this is then followed by a wave of NODAL signaling, leading to the induction of mesodermal differentiation (Chhabra et al, 2019; Heemskerk et al, 2019). With these results, they have shown there are three dynamical events rather than a gradient controlling differentiation, with the focus set on the triggering timings (Liu et al, 2022). Undoubtedly, this system can help study cell interactions that were not modeled in standard culture systems.

As presented in previous talks, it has become clear that the tridimensional organization and disposition of cells are crucial for answering complex questions in the field of stem cells and development. This same mood was continued with the talk from Prisca Liberali (Friedrich Miescher Institute, Switzerland). Her group is interested in understanding how individual cells can create patterned multicellular structures, where each cell plays a role in a space-time coordinated manner. To address this question, she focused on the formation of the intestine by following the self-organization and crypt morphogenesis in intestinal organoids. Starting from adult stem cells, she explained the pipeline for obtaining the organoids that are later imaged and analyzed, compiling imaging for hundreds and hundreds of organoids to infer developmental trajectories.

This process reveals the loss of expression of LGR5 and the switch to a YAP-positive state during organoid development. Also, it appears to recapitulate intestinal regeneration, where YAP heterogeneity is implicated in symmetry breaking (Serra et al, 2019). However, these intestinal organoids show their correct shape during formation since morphogenesis is being ruled by intrinsic forces. The combination of tissue mechanics following a model of spontaneous curvature with mechano-osmotic forces coordinated by the villus gives as a result the correct formation of the crypt (Yang et al, 2021).

The closing speaker for this fantastic session was Christine Mummery (Leiden University Medical Center, Netherlands), a pioneer in PSC differentiation to heart tissue. She emphasized her view of using stem cell models to improve personalized medicine. However, there are roadblocks to achieving this, one being the immature state of cells derived from hPSCs using most differentiation protocols. In the case of cardiac differentiation, cardiomyocytes lack the 3D environment and other cardiac-specific cell types for their proper maturation. To tackle this challenge, Mummery and colleagues combined cardiac fibroblasts, cardiomyocytes, and endothelial cells—all derived from hPSCs—with small molecules to enhance maturity.

Once they obtained these cells, they built cardiac microtissues that display structural, electrical, mechanical, and metabolic maturation (Campostrini et al, 2021). These trilineage models present high utility for disease modeling and drug toxicity, with remarkable advantages like being low tech, scalable, and inexpensive. Dr. Mummery ended her talk and closed the Presidential Plenary putting at the center of attention the fundamental role of cellular context in the development of advanced models based on hPSCs.

Pushing the Boundaries in Stem Cell Therapy and Regeneration

This session focused on research to reprogram and engineer tissue for the purpose of tissue repair and cancer treatment. It started with the winner of the ISSCR Dr. Susan Lim Award for Outstanding Young Investigator, Jennifer Phillips-Cremins (University of Pennsylvania). As a pioneer in developing molecular and computational technologies to dissect how genomes fold at ultra-fine-scale resolution, Dr. Phillips-Cremins applied them to iPSC-derived neurons/organoids and discovered chromatin mechanisms governing synaptic plasticity in the developing and diseased human brain. These include loops/subTADs that are induced by neural circuit simulation and control gene expression during neural differentiation. In the genetic disorder Fragile X syndrome (FXS), where a CGG-repeat tract located at the 5′ end of the FMR1 gene on the X chromosome is expanded, the chromosome repeats attract DNA methylases and form heterochromatin domains, disrupting loops/subTADs in FXS patients and affecting gene expression (Sun et al, 2018; Zhou et al, 2021).

Deepta Bhattacharya (University of Arizona College of Medicine) followed with a talk about engineering pluripotent stem cells to evade and promote immunity. In our bodies, there are cells capable of providing permanent humoral immunity, and these long-lived plasma cells are differentiated from naive B cells. His earlier work showed that 1 μL of immune serum, which contains antibodies corresponding to roughly three long-lived plasma cells, can protect mice against lethal West Nile virus infections Purtha et al, 2011). However, challenges exist to some highly mutated viruses such as HIV, where almost all infections lead to poorly neutralizing antibodies. A small subset of patients who can control HIV well are found to have extremely potent and broadly neutralizing antibodies, although these antibodies take years to develop. How would one develop a prophylaxis to prevent such viral infections?

Engineering plasma cells from ESCs or iPSCs introduced with rearranged antibody genes may be a possible solution. Dr. Bhattacharya then introduced ways to make minimally immunogenic iPSCs, for example, depleting/mutating HLA (β2m, TAP1, CIITA, and CD74) and immune evasion factors (such as MICA/MICB) using CRISPR technologies that allow human cells persist in wild-type immunocompetent mice, and how to turn B cells into long-lived plasma cells by IFNy cytokine-induced glucose uptake.

This was followed by another relevant presentation from Yvonne Chen (UCLA), who talked about engineering next-generation CAR-T cells for cancer immunotherapy. First-generation T cell therapy, mainly CD19 CAR-T cells, faces the problem of antigen escape of tumors, which significantly limits the durability of response and causes cancer relapse in 50%–60% of patients. Yvonne Chen's group developed a bispecific CAR design with both CD19 and CD20 CAR antigens, which allows for an increase in survival (Larson et al, 2022; Zah et al, 2020). Chen's group will be exploring methods to resolve remaining challenges such as graft-versus-host disease, transplant rejection, and immunogenicity arising from repeated CAR-T cell dosing.

Next, Malin Parmar (Lund University, Sweden) talked about cell replacement therapy in Parkinson's disease and focused on using scRNA-seq to analyze graft composition. Similar to fetal ventral mesencephalic (VM) transplant, hESC-derived grafts contain authentic and functional dopaminergic (DA) neurons, non-DA neurons, and glia, and some other cell types. scRNA-seq reveals a high similarity between fetal VM and stem cell-derived grafts and suggests relevant safety information about using stem cell-derived grafts (Fiorenzano et al, 2021). Dr. Parmar also commented on combining a barcoding approach to trace cell origin and map lineage diversification in the graft.

To conclude, Deepak Srivastava (USCF) delivered a talk on using cellular reprogramming to develop regenerative medicine and understand disease mechanisms of congenital heart disease. In mice, cardiac fibroblasts can be transdifferentiated into induced cardiomyocytes and show in vivo functions by combining the three factors Tbx5, Gata4, and Mef2c. In the human model, MEF2c/TBX5 and Myocardin reprogram primary human cardiac fibroblasts into cardiomyocytes (Fu et al, 2013). Dr. Srivastava also highlighted the use of iPSC models of calcific aortic valve disease to allow drug screening and study of disease-associated mechanisms (Gonzalez-Teran et al, 2022).

Programming and Reprogramming

This plenary session was devoted to genetic and epigenetic aspects of programming into somatic and germ cells and reprogramming into pluripotent cells. It opened with an update on using stem cells to make islets. The generation of human β-cells as cell therapy for diabetes has been an area of intense research interest around the world for years. Douglas Melton (Harvard University and Vertex Pharmaceuticals) reported their research progress on making functional β-cells and islets and manipulating genes to induce tolerance to stem cell-derived islets. He emphasized that stem cell-derived islets are functional as they can respond to multiple glucose challenges by secreting insulin, and they can rescue diabetes in animal models.

With impressive progress in stem cell therapy having been made for diabetes, including the well-established directed differentiation protocol into β-cells from stem cells, Douglas presented the work that aims to gain complete mastery over cell composition and move beyond the need for pharmacological immune suppression. Thanks to the scRNA-seq technique, the directed differentiation protocol is proven to be able to develop multiple endocrine cell types. Once the cells complete the differentiation protocol, they are extremely stable as they can be cultured over 5 weeks, maintaining their differentiated and functional states without fate changes. To control the composition of these differentiated functional cells, Melton's group performed CRISPR pooled genetic perturbation screens and identified ∼200 perturbations with statistically significant effects.

Many of these hits turned out to be transcription factors (TF), and PAX4 is one example that its mutation leads to more α-cells with fewer other cell types. Dr. Melton also mentioned the interesting target FBX, an F-box protein functioning in the ubiquitin-proteasome pathway, which can regulate endocrine fate specification as its mutation results in more β-cells. Currently, the best perturbation has not been identified yet, but Dr. Melton stated he was convinced this is the right way to move forward to end up with a monoclonal line that can be eventually used in the clinic. He then briefly mentioned their published work on modeling type 1 diabetes autoimmunity in vitro using hPSCs (Leite et al, 2020) and ended his presentation with the prospect of stem cell-derived islet transplantation for diabetes treatment.

Fiona Watt (King's College London/EMBO, UK) described their recent progress on the role of Gata6 in regulating differentiation of the sebaceous duct lineage. Previously, they used a Gata6 reporter mouse and observed that Gata6 was not only expressed in the junctional zone but also in the sebaceous gland duct that acts as a conduit for the release of sebum onto the skin surface. A vast majority of Gata6+ cells are suprabasal cells and expressed terminal differentiation markers. However, a small population of Gata6+ cells expressing low Gata6 level co-expresses the stem cell marker Lrig1. Lrig1+ stem cells maintain the sebaceous gland and can contribute to additional lineages during wound repair.

They further showed that Gata6+ lineage cells have the same capacity as Lrig1+ lineage cells for long-term epidermal maintenance (Donati et al, 2017). Then to find out how this process works, they carried out scRNA-seq of Gata6+ and Gata6- lineage cells of the postinjury wound cells and the control cells. In unwounded skin, Gata6+ and Gata6- lineage cells mapped primarily to the upper hair follicle, interfollicular epidermis basal cells, and outer hair follicle bulge. In wound epidermis, the Gata6+ and Gata6- lineage cells have a common transcriptional identity. Gata6+ lineage cells in unwounded and wounded epidermis cluster separately, and postinjury, Gata6+ lineage cells dedifferentiate to Lrig1+ stem cells. Such dedifferentiation occurs through reversal of the normal differentiation process rather than occurring through a new pathway.

Following the analysis, they focused on c-Myc, one of the TF whose expression varies with cell states. Depletion of c-Myc showed that the dedifferentiation capacity of Gata6+ lineage cells is dependent on this TF. They further showed that dedifferentiation can also be induced with retinoic acid (RAs) or stretching of unwounded skin, and actin-cytoskeleton remodeling is a key feature of dedifferentiation.

Impressive progress in the pluripotency transition has been made for mouse and human PSCs in the last decade. Austin Smith (University of Exeter, UK) described their recent progress in the pluripotency transition of hPSCs. They previously developed a protocol to establish and propagate naive hPSCs in a medium containing the MEK/ERK inhibitor PD03, tankyrase inhibitor XAV939, aPDCi/z inhibitor Gö6938, and human LIF, constituting PXGL (Dattani et al, 2022). In human, while ERK inhibition can repress the differentiation of naive hPSCs to formative PSCs or hypoblast state, ERK inhibition can also induce trophectoderm. Self-renewal requires a blockade of the trophectoderm lineage. To solve such conflicting effects of ERK inhibition, they showed tankyrase inhibitor XAV939 was able to block the induction of trophectoderm. They then dissected the mechanism of this effect. Tankyrase inhibitor blocks the canonical Wnt/β-catenin signaling.

However, β-catenin null cells remain dependent on XAV as they still readily differentiate to trophectoderm without XAV. XAV can impede YAP/TAZ activation by stabilizing Angiomotin family proteins. Austin then presented that in human naive PSCs, XAV stabilizes Angiomotin family members AMOT and AMOTL2 and reduces nuclear YAP/TAZ. Overexpression of AMOTL2 also blocks trophectoderm induction. They also validated this effect by knocking out AMOTL2 and AMOT, which enables trophectoderm induction. To complete this pathway, they then found that YAP1 knockout cells fail to make trophectoderm and can self-renew efficiently without XAV. These findings show the distinct requirement for tankyrase inhibition in human naive PSCs and highlight the pivotal role of YAP/TAZ in human pluripotency transition.

This year, the ISSCR Anne McLaren Lecture was delivered by Ruth Lehmann (Whitehead Institute/MIT). Her laboratory studies germ cells and is interested in how these cells become specialized and how the cytoplasmic information and cellular organelles are passed from the egg to the next generation. This time, she discussed two interesting guiding mechanisms for germ cells. The first one is about the integration of directed migration cues and self-organized motility that initiate germ cell migration. She first showed that Drosophila melanogaster primordial germ cells (PGCs) migrate with a constant spherical shape and prominent rear polarity. The PGCs utilize autonomous retrograde cortical actin flow and during migration, RhoA activation dominates the front and back polarity of the cells.

RhoGEF2, a microtubule plus-end tracking RhoA-specific RhoGEF, which is enriched at the rear of PGCs, regulates actin flow and polarity with a phosphoregulated feedback loop in these cells. RhoGEF2 depletion slows cortical flow with reduced migration speed, while its activation enhances flow and polarity with guidance impairment, indicating cortical flows must be carefully tuned to stay within a tolerable speed range for accurate migration. Tre1 GPCR controls RhoGEF2 localization during guided migration. RhoGEF2 PDZ and PH domains, instead of the canonical G_α12/13 signaling domains, are required for polarity and migration through a positive feedback loop that augments its basal activity. This feedback loop is regulated by AMPK-dependent phosphorylation, which releases RhoGEF2 from microtubules and anchoring (Lin et al, 2021).

The second mechanism is about isoprenoids, which are conserved between mouse and fly, guiding germ cells to their target. Isoprenoids, such as RAs in vertebrates and juvenile hormones (JHs) in insects, influence the germline lifecycle from meiosis to gametogenesis. Lehmann's group investigated the functions of JHs in early Drosophila germ cell development. With an in vivo JH reporter, they found that JH signaling is first detected in mid-embryogenesis and is enriched near PGCs that are migrating to the developing somatic gonadal precursors. JH can attract germ cells and are sufficient for PGC migration. They further demonstrated that instead of canonical transcription-dependent signaling, an alternative GPCR and Ca++ signaling is required for JH-mediated germ cell migration. Finally, she presented that the vertebrate isoprenoid, RA, like JH, is sufficient for PGC migration in vitro, supporting a broadly conserved role of isoprenoids in reproduction (Barton et al, 2021).

Defining Stem Cells Across Space and Time

Cellular plasticity enables cells to change both through time and space, as the dynamical individual entities they are, and as part of complex structures. For example, so is the case with regeneration, a fundamental process across living organisms. The line-up speakers for the following session approached how fate decisions are taken, guided by extrinsic and intrinsic stimuli that modulate the molecular regulation of all types of cells, both stem and adult cells.

The opening speaker for the fourth plenary session was Tina Mukherjee (Institute for Stem Cell Science and Regenerative Medicine/inStem, India). During her talk, she gave some interesting insights into the noncanonical interplay between the olfactory system and the modulation of the immune system in a model with a refined sense of smell as D. melanogaster. As she detailed, animals sense odors from the environment and release γ-aminobutyric acid (GABA) from the neurons in response to these stimuli. From its metabolism, GABA is broken down to succinate, which increases the levels of Sima/HIF-α and hence triggers the production of lamellocytes, the “giant cells” of the immune system, from hematopoietic progenitor cells.

Next, her group questioned whether fruit flies can sense predators' odors from the environment. For this, they exposed larvae to wasp odor and verified, once as adults, that these preconditioned animals had elevated levels of GABA and were able to produce faster and more competent lamellocytes when exposed again to the same odor. These results prove that the sense of smell can control immune priming (Madhwal et al, 2020).

The following speaker was Sasha Mendjan (IMBA/Vienna BioCenter, Austria). As highlighted before, Dr. Mendjan started his talk by mentioning how self-organization and development are crucial for modeling diseases. His group focuses on organ-specific cell types, patterning, and morphogenesis in cardiac development. Using cardioids developed from hPSCs, they have modeled cardiac mesoderm morphogenesis, recapitulating early stages of ventricle development presenting a hollow chamber and a high proportion of cardiomyocytes (Hofbauer et al, 2021).

As his research moves forward, he stated the importance of studying heart defects and how these cardioids can help bypass the difficulties of having an appropriate 3D model. For this, and after rounds of optimization, his group was set up to develop a platform of cardioids that represent all parts of the human embryonic heart. Even more, the combination of different types of cardioids resulted in the co-development of multichambered cardioids with atria and left and right ventricles (Schmidt et al, 2022). Excitingly, this system shows tremendous advantages for modeling heart diseases in the near future.

Changing the angle of the presentations, Tatjana Sauka-Spengler (University of Oxford, UK) talked about her efforts in identifying regulatory elements by using genome-wide regulatory profiling and reverse engineering global gene regulatory circuits (GRNs) underlying developmental processes. Particularly, she described the use of these systems-level approaches to assemble vagal neural crest (VNC) GRNs (Betancur et al, 2010). The neural crest is a multipotent migratory cell population with a broad range of derivatives that are interesting for therapeutic matters. Specifically, the VNC cells give rise to the enteric nervous system, which controls gut motility and function.

From the results she presented, the transcriptional analysis of VNC subpopulations revealed three distinct signatures corresponding to neural, neuronal, and mesenchymal cells (Ling and Sauka-Spengler, 2019). Epigenetic analysis also indicated different regulatory landscapes, which analyzed through time-revealed unique cis-regulatory programs for each VNC subpopulation (Williams et al, 2019). With this information, it is possible to focus on a precise factor and validate its role in the GRC by studying its interactions and perturbation effects. Eventually, Dr. Sauka-Spengler underlines that these programs can be extrapolated and combined with machine learning for modeling disease phenotypes.

Alejandro Sánchez Alvarado (Stowers Institute) began his presentation by talking about the sources of regenerative capacity in animals. One of the interests of his research group is the transcriptional plasticity not only of stem cells but also of non-stem cells, and the connections between them. Centered in the planarian model, he first summarized how neoblasts with regenerative capacity, which are positive for tetraspanin-1, are actual adult pluripotent cells that act upon regeneration and can present differential gene expression depending on the biological context (Zeng et al, 2018). Going into deeper detail, Sánchez Alvarado and colleagues developed a strategy to characterize transactional plasticity in response to injury defined as transient regeneration-activated cell states (Benham-Pyle et al, 2021).

As they believe, describing the mechanisms that modulate these states is crucial to understanding regeneration across animals. Their results showed cell types and molecules important for regeneration and belonging to all three germ layers, which displayed rare and transient cellular states. Furthermore, they wondered where these cells reside and discovered, by using spatial transcriptomics, that there is a wound-induced association of stem cells with matrix remodeling cells (Benham-Pyle et al, 2022). The presentation ended with an invitation from Dr. Sánchez Alvarado to reflect on the concept of “terminal differentiation” and whether we should shift this idea instead to “stable differentiation,” undoubtedly, one of the many take-home messages from the ISSCR 2022 meeting.

Disentangling Single-Cell Contribution to Organogenesis and Pathology

The session devoted to incorporating recent technological developments for large-scale scRNA-seq and organ engineering to understand development and disease in organs was led off by Muzlifah Haniffa (Wellcome Sanger Institute, UK) with a talk on decoding the developing human immune system. Haniffa explained how her group has profiled multiple peripheral, lymphoid, and primary hematopoietic organs using scRNA-seq combined with antigen receptor sequencing and spatial transcriptomics to understand the dynamics of how the human immune system develops across organs and time.

She then described how this multiomics dataset helped reveal key findings, including some related to changes in HSCs across gestation periods and organs (Jardine et al, 2021), as well as mechanisms by which monocytes exit from the bone marrow and enter the tissue and the role of macrophages in tissue morphogenesis and lymphopoiesis (Suo et al, 2022). She concluded with examples of how they used these datasets of the development of the human immune system to map chromosomal alterations, which allowed them to further our understanding of diseases such as Down syndrome (Jardine et al, 2021) and infant and childhood leukemia (Khabirova et al, 2022).

This was followed by a presentation by April Pyle (UCLA) who discussed her findings focused on understanding how to generate iPSC-derived muscle cells to generate stem cell therapies for muscle wasting diseases. She explained how her group has used a Drop-seq RNA-seq platform to generate a dataset of human limb muscle tissue overlayed with human PSC muscle culture to get a clearer view of what the different cells expressed in embryonic, fetal, and adult myogenesis, as well as when generated in a dish, are.

This revealed that there are five different stages of human myogenesis and that iPSC-derived PAX7+ cells are aligned with stage 3, which occurs during developmental weeks 7–12 (Xi et al, 2020). Building on this, engraftment assays of iPSC-derived skeletal muscle progenitor cells into Pax7 KO mice enabled the generation of regenerating myofibers with human Pax7+ cell association. As such, they developed a very advantageous system to model stages of regeneration happening early after injury, an important step to understanding underlying mechanisms and to develop iPSC-based therapies for muscle wasting diseases.

This was followed by a talk by Xiaoqun Wang (Institute of Biophysics Chinese Academy of Science, China), who presented work on the generation of a cell atlas of the human prefrontal cortex using scRNA-seq to identify developmental trajectory of diverse cells of the prefrontal cortex. They used this dataset to decode the diversity and molecular characteristics of neural progenitors and cell lineages in this region (Zhong et al, 2018). This led to the identification of a primate-specific gene, TMEM14b, as a marker for outer radial glia and a driver of neural progenitor proliferation (Liu et al, 2017).

Remarkably, the expression of this gene in mice induces cortical gyrification, suggesting a contribution to brain evolution. They then developed a cortical brain organoid containing a vascular system that promotes the neurogenesis and maturation of cortical development in vitro, and that reconstructs the vascular system in the mouse cortex and promotes cell survival in the graft (Shi et al, 2020).

To end the session, Janet Rossant, the chair of the selection committee of the ISSCR achievement award, introduced the 2022 recipient, Lorenz Studer (Memorial Sloan Kettering Cancer Center), for his body of work on the development of robust protocols to generate neural progenitor cells, including midbrain dopaminergic progenitors, which he has now brought to an early-stage clinical trial for cell replacement therapy in Parkinson's Disease. Studer detailed two new studies on species-specific timing differences in neuronal maturation using synchronized cortical neuron cultures.

Following a chemical screen for drivers of neuronal maturation, his group identified the GENtoniK cocktail that removes the epigenetic barrier, while triggering neuronal activity (Hergenreder et al, 2022). They further identified chromatin-related factors as barriers slowing down neuronal maturation that, when removed with chemicals, accelerate neuronal maturation in vitro (Ciceri et al, 2022). These findings not only could be extremely useful in in vitro settings to manipulate the timing of human neuronal maturation but also put forward the strong involvement of epigenetics as a driver of species-specific timing differences in neural development.

Technologies That Model and Direct Emergent Cell Behavior in Stem Cell Biology and Regeneration Behaviors

This session focused on new approaches to model, direct, and characterize stem cells and regeneration behaviors. With the integration of classical developmental methods, such technological advances benefit stem cell research. Although single-cell sequencing methods have been widely used today, these tools focus predominantly on exploring the genomes, epigenomes, and transcriptomes of single cells. However, measuring translation at a single-cell level remains a big challenge. Alexander van Oudenaarden (Hubrecht Institute-KNAW and University Medical Center, Netherlands) described their recently published single-cell Ribo-seq method (scRibo-seq) (VanInsberghe et al, 2021), which enables the investigation of translation in individual cells, using micrococcal nuclease footprinting followed by size selection to allow ribosomal profiling.

They first validated their method by showing that ribosome-protected fragments (RPF) are enriched in coding sequences and display a three-nucleotide periodicity. The limitation of a certain amino acid further validates scRibo-seq by showing translational pausing is induced at a subset of the codons encoding the limited amino acid. Interestingly, pausing is only detected in a subpopulation of sequenced cells correlating to their cell cycle states. The classical Ribo-seq has the in-parallel RNA-seq to distinguish the changes happening on RNAs and RPFs. However, the current scRibo-seq cannot distinguish such changes. To solve this problem, they proposed a method to integrate scRNA-seq and scRibo-seq from the same sample and then separate the cells based on positions in the cell cycle, which groups the cells into partially overlapping neighborhoods.

Even though the scRNA-seq and scRibo-seq are not from the same cell, such a combination enables to determine the RNA changes and translation efficiency changes from the neighborhood cells. With this strategy, they identified thousands of genes in the human cell cycle that seem to be translationally controlled. And genes that are translationally repressed in mitosis have structured 5′ untranslated regions (UTRs) with a higher proportion of reads in their 5′ UTRs. They further identified that these 5′ UTRs are enriched with CGC repeats, which might point to RNA G-quadruplexes that could repress translation.

The ubiquitous large datasets containing high-throughput, high-dimensional data of cells pose new challenges in terms of noise, missing data, measurement artifacts, and the “curse of dimensionality.” Smita Krishnaswamy (Yale University) described their data geometric and topological approaches to understanding the shape and structure of developmental data. Their goals are to denoise data by distilling the structure and infer and represent the dynamics of multiple samples and scales (Amodio et al, 2019). They used human embryonic stem cell development as a model. She showed how to obtain useful representations of data, how to combine diffusion geometry with topology, and how to learn dynamics from static snapshot data. In summary, she showed a framework on how to analyze large datasets to uncover insights into stem cell development and regeneration biology (Moon et al, 2019).

Gordana Vunjak-Novakovic (Columbia University) discussed a multiorgan chip platform with human tissue niches linked by vascular flow. Human engineered tissues from pluripotent stem cells can be used to recapitulate organ-level functions, model human pathophysiology, and test drugs. To guarantee physiological relevance, the engineered tissue needs to physiologically communicate and maintain phenotypes for a long time. Dr. Vunjak-Novakovic reported the development and applicability of a tissue chip system in which human tissues, including heart, liver, bone, and skin tissues, can be maintained in their own optimized niche and linked to other tissues for a long time by vascular flow containing circulating cells.

Dr. Vunjak-Novakovic's group designed this multiorgan tissue chip that can provide each tissue with its specialized niche with connecting tissues through vascular flow and selectively permeable endothelial barriers, which can separate the vascular and tissue compartments (Ronaldson-Bouchard et al, 2022). To further explore the potential of this technology, Dr. Vunjak-Novakovic also presented their recent work to use such platforms to recapitulate tissue injury, regeneration, and disease. She gave some examples, including the study of human bone marrow with hematopoiesis, modeling ischemia-reperfusion injury, modeling breast cancer metastasis, and radiation studies.

Satoshi Toda (Kanazawa University, Japan) introduced their research about programming multicellular pattern formation with synthetic cell-cell signaling. To study how cell-cell communications organize complex tissue structures particularly focused on morphogens, Dr. Toda's group developed synthetic morphogen modeling systems. They used GFP as an example of synthetic morphogen, which was expressed from GFP-secreting cells and formed a gradient-like morphogen signaling, that could be sensed by synthetic receptors. Such systems generated patterns reminiscent of those observed in vivo (Toda et al, 2020). Dr. Toda then talked about their recent ongoing work to test precise patterning using a 3D synthetic morphogen system.

They first found that in the 3D system, only interpretation of signal proteins into gene expression was not sufficient for precise gradient, as gradient formation was disturbed with random cell movement. They then expressed E-cadherin in receiver cells and the induced cell adhesion could block cell movement, leading to the formation of a more precise gradient. Such coupling of morphogen signal and cell adhesion could form tissue domains. Their next challenge is to design synthetic circuits for tissue engineering and cell therapy for tissue regeneration. At last, Dr. Toda briefly mentioned their ongoing work of developing an in vitro model of cell therapy for tissue regeneration using intestinal organoids.

The winner of the 2022 ISSCR Momentum Award, Joanna Wysocka (Stanford University), talked about her research journey in stem cell biology. She first briefly mentioned a few studies to highlight the privileged roles of enhancers in mediating developmental gene expression programs, which were published earlier by her group. Then she explained how they used cranial neural crest cells (CNCCs) as a paradigm to study how variation in enhancer sequence or function gives rise to the human phenotypic variation. CNCCs are the major cell type of origin for the developing human face and the protocol to derive human CNCCs from hESCs in vitro was previously established.

To better study cellular anthropology in this question, they also used ape cellular models to study human evolution (Prescott et al, 2015). Genome-wide association studies (GWAS) identified 203 genomic regions associated with different aspects of face shape and the facial shape GWAS variants map to the noncoding genome and are enriched at CNCC enhancers. The analysis of molecular pathways that affect both disease-associated and normal-range facial variation highlighted sensitivity to TF dosage. Therefore, they focused on SOX9, a TF associated with craniofacial variation and disease, and precisely modulated SOX9 dosage in CNCCs with dTAGv-1, to study TF dosage effects.

They found that most SOX9-regulated enhancers and genes respond nonlinearly to dosage. Most SOX9-regulated enhancers are buffered against small to moderate changes in SOX9 dosage and a subset of SOX9-regulated enhancers shows sensitivity to small dose perturbations. They further identified specific cis-regulatory features in dosage-sensitive enhancers and showed that enhancer dosage curves could predict the shape of gene responses. At last, she reported that dosage-sensitive genes are associated with specific cellular and morphological phenotypes, and highlighted the significance of such dosage effects and associated regulatory controls for studies of mechanisms underlying human phenotypic variation in health and disease (Naqvi et al, 2022).

Cell and Gene Therapy in the Clinic

Stem cells have the potential to differentiate into a variety of cells for cell therapy. Meanwhile, viral vectors for gene modifications have provided opportunities to engineer stem cells to custom for the need. Cell and gene therapy has entered clinical trials to test in human patients and this session focused on these exciting advances.

Luigi Naldini (San Raffaele University, Italy) presented a breakthrough in HSC gene therapy. While traditional lentiviral vector allows highly efficient gene replacement and is compatible with long-term HSCs, it suffers from residual insertional genotoxicity. The newly emerged nuclease-based editing technology and base editors have advantages such as in situ gene correction and DNA break-free single/few bases edited, but are still at early clinical stage and their off-target activity needs to be closely evaluated. Therefore, current clinical strategies focus on ex vivo genetic engineering and infusing HSCs back into patients, which require chemotherapy to deplete the original HSPC cells in the patient. Dr. Naldini's group exploited efficient mobilization regimens to exchange engineered HSPC within the original niches so that chemotherapy can be avoided (Omer-Javed et al, 2022).

Transient overexpression of engraftment enhancers (such as CXCR4), coupled with gene editing and the ex vivo culture, provides engineered hematopoietic stem/progenitor cells with migration and homing advantage compared to original mobilized cells, overall conferring a further competitive advantage. This paves the way for broader and safer use of HSPC gene therapy in the clinic.

Masayo Takahashi (Vision Care, Inc., Japan) provided the latest results from the application of iPS-derived cells, like retinal pigment epithelium (RPE), to treat wet age-related macular degeneration. Cell therapy requires collaborations with hospitals and surgeons to ensure successful treatment, and so has shown to be the case for the autologous iPSC-RPE transplantation conducted in 2014 (Mandai et al, 2017). The transplanted cells generated stable RPE sheets in the patient, which have lasted over 7 years to date, and the patient's vision was stabilized after the surgery.

Vision Care, Inc., has also conducted a second clinical research using HLA-matched allogeneic iPSC-RPE transplantation (five cases total) to show the possibility of allogeneic transplantation without systemic immune suppression for elder patients (Sugita et al, 2020). And in the third clinical research, they improved from iPSC-RPE suspension to RPE strip and performed 50 cases since 2020 to evaluate the efficacy of iPSC-RPE treatment (Nishida et al, 2021). The potential issues for the clinical trial are the difficulty to conduct phase 3 clinical trials, the mismatch between costs and benefits currently being an expensive treatment, and incentives for doctors and hospitals as cell therapy causes hospital deficits. The future directions include using retinal organoids that consist of multiple cell types and layers for transplantation (Kuwahara et al, 2019), and using robots to control the variability during RPE differentiation and optimization for the best possible condition.

Bob Valamehr (Fate Therapeutics) presented their platform to make universal natural killer (NK) cell and T cell products from iPSCs for cancer treatment. Impressively, their triple-edited iPSC-derived NK cells (hnCD16+, CAR19+, and IL15-RF) can overcome tumor heterogeneity and antigen escape and gives durable CAR-mediated cytotoxicity in mice (Cichocki et al, 2021).

Nearly two decades ago, Harvard Professor Doug Melton had a vision to make insulin-producing beta cells from stem cells to cure type I diabetes patients with islet cell transplantation, so he decided to switch his laboratory's focus to this direction. This led to years of collective work from academia and industry to advance science (Pagliuca et al, 2014).

For the last talk in this session, Gary Meininger (Vertex Pharmaceuticals) presented the encouraging clinical update on using stem cell-derived, fully differentiated islet cells for type I diabetes, known as drug candidate VX-880. Both patients who received VX-880 infusion had glucose-responsive insulin production and improved glycemic control, as well as showed clinically significant improvements in HbA1c. Unprecedented, the company has reported through a press release that one of the patients does not require insulin use anymore, which confirms achieving insulin independence. These data establish proof of concept and support further development of islet cell replacement therapy.

These advances in clinical practice, which are derived from stem cell research, are the biggest rewards to scientists who work wholeheartedly for decades to address the unmet needs of patients. The field is optimistic that more exciting news will be coming from pharmaceutical companies developing stem cell-related therapies.

Future Perspectives

The field of stem cell research has evolved tremendously since the first ISSCR meeting in 2002 (Fig. 1). The past 20 years have seen a multitude of “firsts” in this field, all converging toward the main goals of further understanding stem cell and differentiation mechanisms for regenerative therapies and to use patient-derived PSCs to better understand disease mechanisms. While the end of the 20th century and early ‘00s were focused on identifying and understanding stem cells, we have witnessed during the past 10 years the fruit of this early research with many HSC and CAR-T cell treatments approved for clinical use and multiple stem-cell based clinical trials currently ongoing to treat, for instance, Parkinson's Disease, macular degeneration, cancer, or diabetes. As such, the next 5 years have great potential for bringing its load of newly approved stem cell therapies to the clinic.

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FIG. 1.

Timeline of milestone achievements in stem cell research before and since the establishment of ISSCR. ISSCR, International Society for Stem Cell Research.

So, what next? The ISSCR community is already working on the next generation of stem cell-based therapies using state-of-the-art technologies such as multiomics, barcoding, and machine learning, to strengthen the link with immunology and to provide better stem cell products for safe and efficient therapies. In addition, as iPSC cultures, organoids, and other 3D systems are getting increasingly high quality, sophisticated, and high-throughput, patient-specific research is booming and the use of stem cells for the development of precision medicine is a major current focus that will see fruition in the next few years. As ISSCR continues its global collaboration to unite stem cell researchers, to set guidelines, and to pursue its advocacy and policy efforts, the next 20 years are looking quite exciting.