In a study published this year, the failing hearts of rhesus macaques and a human patient were reinvigorated with cells derived from them. Heart cells extracted from the patient were engineered and grown into a patch. When grafted into the patient, the patch promoted new muscle growth and improved the heart’s pumping ability.

As we age, heart muscle cells thicken and lose elasticity. Other cardiovascular disorders, neurodegenerative diseases, and conditions like fibrosis are also marked by cellular damage. The damage is often irreversible as, except for the liver, human organs don’t regenerate themselves. Regenerative therapies, like the engineered heart patches, show that it doesn’t have to be the case.

Building on synthetic biology technologies for engineering and delivering cells to specific locations in the body, these therapies repair and regenerate tissues. Now, researchers are looking for ways to improve their effectiveness and scalability.

Deeper insights into engineered human therapies will be discussed in great detail through various sessions and talks at SynBioBeta: The Global Synthetic Biology Conference, taking place May 6-8 at the San Jose Convention Center.  

Reprogramming Cell Identity

Cell therapies that treat cancers and autoimmune diseases have proliferated over the last decade. More recently, companies have been developing cell therapies with regenerative potential. These typically use stem cells that differentiate into healthy cells and replenish damaged cells in diseased tissues. Such therapies are under development for conditions ranging from Alzheimer’s and muscle atrophy to skin disorders.

Shinya Yamanaka’s discovery of induced pluripotent stem cells (iPSCs)—using a combination of transcription factors to reprogram adult, differentiated cells into pluripotent stem cells—was transformative for regenerative therapies. Subsequently, researchers have learned more about how different transcription factors regulate gene expression and, in turn, trigger cellular programs specific to a particular cell type.

However, cell differentiation is complex and little understood. “When cells go through development, they sequentially hit branch points where they sort of roll the dice to decide which pathway to follow,” said Mark Kotter, founder of UK-based synthetic biology company bit.bio. Consequently, protocols can be long and highly variable. Differentiated cells don’t always stay in the desired state or perform the intended function. Moreover, “it's not always clear whether the final cell type reflects the true human tissue,” said Tim Lu, CEO of the South San Francisco-based biotech Senti Bio.

Researchers are figuring out ways to use these programs to coax stem cells to differentiate into and remain in desired states. While every cell has its identity program, marked by particular transcription factor combinations, few of these are known. “At bit.bio, we built an experimental platform that can screen transcription factors with very high throughput at a massive scale,” said Kotter.

Senti Bio, on the other hand, designs logic-gated synthetic gene circuits. “There are certain transcription factors, microRNAs, and epigenetic genes that are important in dictating cell differentiation and cell function,” said Lu. Gene circuits built with these elements can control how stem cells differentiate.

Cell fate can be manipulated with synthetic transcription factors as well, enabling orthogonal gene regulation systems that don’t interfere with the host cell’s machinery. “These can be modulated by the addition of small molecule drugs,” said Lu. By adding one or a few drugs, multiple genes can be turned on or off inside the cell. “And that works both in vitro, for the manufacturing process, and in vivo,” Lu added.

Scientists are getting better at understanding cell identity as a result of programs. “You can combine subprograms to create cell types that have particular functions that don't exist in nature,” said Kotter.

A relatively newer but incredibly bold kind of regenerative therapy involves engineering animal organs for human transplantation. In the last few years, some patients have received gene-edited pig kidneys and hearts (although none of them survived more than a few months). The organs were edited to remove certain pig genes and introduce human genes to make them compatible with human recipients.

While fully grown gene-edited organs are far from becoming mainstream, mini-organs are now ubiquitous in regenerative medicine. Organoids grown from patient cells are gene-edited to correct the disease-causing mutations and produce healthy cells for transplantation. For off-the-shelf therapies, organoids offer a readily available source of cell lines. Moreover, experimenting on organoids provides insights into organ and disease development that can be brought back to developing regenerative therapies.

Improving Regenerative Cell Therapies

The key to improving regenerative cell therapies is a better understanding of the mechanisms of how stem cells repair and regenerate tissues. “Understanding the cells we’re trying to engineer is an important first step to get to where we want to go,” said Nika Shakiba, a stem cell engineer and synthetic biologist at the University of British Columbia. With advances in omics technologies, scientists can capture information at different scales of biology. This, Shakiba added, allows for the reading out of the lineage of a cell and the genes and proteins being expressed.

The value of omics in regenerative medicine illustrates the complementary roles of synthetic and systems biology. “With systems analysis, we are understanding where to use synthetic biology to tune and module cellular behaviors,” said Mo Ebrahimkhani, a synthetic biologist at the University of Pittsburgh. After reverse engineering the rules of how cells do what they do, they can be applied to forward-engineer more effective therapies.

In addition to greater control over cell differentiation, synthetic gene circuits permit programmable control of other aspects of cell behavior. For instance, therapeutic cells can be designed to respond to their environment. Gene circuits also address an important concern with cell therapies: toxicity. Cell therapies must not have unintended effects and, particularly with off-the-shelf therapies, must avoid triggering the immune system. With gene circuits, bioengineers can strike a balance between function and safety, Ebrahimkhani explained, by ensuring the genetic footprint is enough to perform the function of interest but not too much to induce any toxicity.

Companies are experimenting with a range of strategies to make cell therapies safer. These include engineering cells to confer immunocloaking [to evade the immune system], making universal cell lines, and integrating fail-safe switches that eliminate the transplanted cell population [if it grows uncontrollably],” said Shakiba.

Parallely, technologies to tease out cellular cross-talk are getting better. “Understanding how cells talk to each other and the language of tissue programming and self-organization can improve how we engineer tissues and organisms,” said Ebrahimkhani. Engineering multicellular systems have the potential to bring forth the next generation of cell therapies as well as make organoids that are more robust and reliable and capture the complexity of human organs.

Synthetic gene circuits and gene-edited organoids highlight the importance of gene editing technologies in engineering regenerative therapies. But they could also reveal the fundamental principles of regeneration. For instance, bit.bio creates CRISPR-ready cell lines that can be used to edit a particular cell type in cultures comprising multiple cell types and deconvolute complex behaviors in such cultures. 

Scaling Up Cell Therapies

Scaling up is a massive challenge for manufacturing cell therapies, particularly for patient-derived therapies, regardless of whether they are regenerative or not. This is a massive part of why they are prohibitively expensive. Increasing cell therapy yields requires new bioreactor designs and strategies that optimize stem cell differentiation. “Automation and adapting manufacturing into larger and larger formats is going to be an important part of achieving greater scale,” said Lu. Employing genetic engineering to produce more potent growth factors or have the cell make its growth factors could also improve scalability.

The adoption of AI will also facilitate the production of effective cell therapies at scale. It can design gene circuits with greater control over cell behavior and map varying nutrients and environmental conditions in a bioreactor to cell growth and differentiation. Besides, it can speed up quality control.

Stem cell fitness is another active area of investigation that has implications for the yield and effectiveness of regenerative cell therapies. “We have to understand how perturbations, both to the environment and the cell, influence the ability of a community of cells to get to where we're trying to get them,” said Shakiba. This is critical to ensuring that all cells in a batch reprogram into desired cell types and are fit enough to engraft and do their therapeutic function.

Currently, scientists lack the tools to study how different perturbations impact the fitness and function of stem cell-derived therapies. To overcome this barrier, Shakiba noted, “We need to devise simulation platforms that allow us to do model-aided design with cells and stem cells the way we do in other engineering disciplines.”