CIRM interviews Lorenz Studer: 2017 recipient of the Ogawa-Yamanaka Stem Cell Prize [Video]

For eight long years, researchers who were trying to develop a stem cell-based therapy for Parkinson’s disease – an incurable movement disorder marked by uncontrollable shaking, body stiffness and difficulty walking – found themselves lost in the proverbial wilderness. In initial studies, rodent stem cells were successfully coaxed to specialize into dopamine-producing nerve cells, the type that are lost in Parkinson’s disease. And further animal studies showed these cells could treat Parkinson’s like symptoms when transplanted into the brain.

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Lorenz Studer, MD
Photo Credit: Sloan Kettering

But when identical recipes were used to make human stem cell-derived dopamine nerve cells the same animal experiments didn’t work. By examining the normal developmental biology of dopamine neurons much more closely, Lorenz Studer cracked the case in 2011. Now seven years later, Dr. Studer, director of the Center for Stem Cell Biology at the Memorial-Sloan Kettering Cancer Center, and his team are on the verge of beginning clinical trials to test their Parkinson’s cell therapy in patients

It’s for these bottleneck-busting contributions to the stem cell field that Dr. Studer was awarded the Gladstone Institutes’ 2017 Ogawa-Yamanaka Stem Cell Prize. Now in its third year, the prize was founded by philanthropists Hiro and Betty Ogawa along with  Shinya Yamanaka, Gladstone researcher and director of the Center for iPS Cell Research and Application at Kyoto University, and is meant to inspire and celebrate discoveries that build upon Yamanaka’s Nobel prize winning discovery of induced pluripotent stem cells (iPSCs).

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(L to R) Shinya Yamanaka, Andrew Ogawa, Deepak Srivastava present Lorenz Studer the 2017 Ogawa-Yamanaka Stem Cell Prize at Gladstone Institutes. Photo Credit: Todd Dubnicoff/CIRM

Studer was honored at the Gladstone in November and presented the Ogawa-Yamanka Stem Cell Prize Lecture. He was kind enough to sit down with me for a brief video interview (watch it below) a few minutes before he took the stage. He touched upon his Parkinson’s disease research as well as newer work related to hirschsprung disease, a dangerous intestinal disorder often diagnosed at birth that is caused by the loss of nerve cells in the gut. Using human embryonic stem cells and iPSCs derived from hirschsprung patients, Studer’s team has worked out the methods for making the gut nerve cells that are lost in the disease. This accomplishment has allowed his lab to better understand the disease and to make solid progress toward a stem cell-based therapy.

His groundbreaking work has also opened up the gates for other Parkinson’s researchers to make important insights in the field. In fact, CIRM is funding several interesting early stage projects aimed at moving therapy development forward:

We posted the 8-minute video with Dr. Studer today on our official YouTube channel, CIRM TV. You can watch the video here:

And for a more detailed description of Studer’s research, watch Gladstone’s webcast recording of his entire lecture:

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Comparing two cellular reprogramming methods from one donor’s cells yields good news for iPSCs

In 2012, a mere six years after his discovery of induced pluripotent stem cells (iPSCs), Shinya Yamanaka was awarded the Nobel Prize in Medicine. Many Nobel winners aren’t recognized until decades after their initial groundbreaking studies. That goes to show you the importance of Yamanaka’s technique, which can reprogram a person’s cells, for example skin or blood, into embryonic stem cell-like iPSCs by just adding a small set of reprogramming factors.

These iPSCs are pluripotent, meaning they can be specialized, or differentiated, into virtually any cell type in the body. With these cells in hand, researchers have a powerful tool to study human disease and to develop treatments using human cells directly from patients. And at the same time, this cell source helps avoid the ethical concerns related to embryonic stem cells.

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Induced pluripotent stem cell (iPSC) colonies.
Image Credit: Joseph Wu

Still, there has been lingering uneasiness about how well iPSCs match up to embryonic stem cells (ESCs), considered the gold-standard of pluripotent stem cells. One source of those concerns is that the iPSC method doesn’t completely reprogram cells and they retain memory of their original cell source, in the form of chemical – also called epigenetic – modifications of the cells’ DNA structure. So, if a researcher were to make, say, heart muscle cells from iPSCs that have an epigenetic memory of its skin cell origins, any resulting conclusions about a given disease study or cell therapy could be less accurate than ESC-related results. But a report published yesterday in PNAS should help relieve these worries.

The CIRM-funded study – a collaboration between the labs of Joseph Wu and Michael Synder at Stanford University and Shoukhrat Mitalipov at Oregon Health & Science University – carried out an exhaustive series of experiments that carefully compared the gene activity and cell functions of iPSC-derived cells with cells derived from embryonic stem cells. The teams sought to compare cells generated from the same person to be sure any differences were not the result of genetics. To make this “apples-to-apples” comparison, they generated embryonic stem cells using another reprogramming technique called somatic cell nuclear transfer (SCNT).

With SCNT, a nucleus from an adult cell is transferred to an egg which has its own nucleus removed. The resulting cell becomes reprogrammed back into an embryo from which embryonic stem cells are generated – the researchers call them NT-ESCs for short. In this study, the skin cell sample used for making the iPSCs and the cell nucleus used for making the NT-ESCs came from the same person. In scientific lingo, the iPSCs and SCNT stem cells are considered isogenic.

Now, it turns out the NT-ESC reprogramming process is more complete and eliminates epigenetic memory of the original cell source. So why even bother with iPSCs if you have NT-ESCs? There are big disadvantages with SCNT: it’s a complex technique – only a limited number of labs pull it off – and it requires donated human eggs which carries ethical issues. So, if a direct comparison iPSCs and SNCT stem cells shows little difference then it would be fair to argue that iPSCs can replace NT-ESCs for deriving patient-specific stem cells.

And that’s exactly what the teams found, as Dr. Wu summarized it to me in an interview:

“Direct comparison between differentiated cells derived from iPSCs and SCNT had never been performed because it had been difficult to generate patient-specific ESCs by the SCNT method. Collaborating with Dr. Shoukhrat Mitalipov at Oregon Health & Science University and Dr. Michael Snyder at Stanford University, we compared patient-specific cardiomocytes (heart muscle cells) and endothelial (blood vessel) cells derived by these two reprogramming methods (SCNT and iPSCs) and found they were relatively equivalent regarding molecular and functional features.”

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Blood vessel cells derived by iPSC (left) and SCNT (right) reprogramming methods.
Image credit: Joseph Wu

Because the heart muscle and blood vessel cells were similar regardless of reprogramming method, it suggests that the epigenetic memory that remained in the iPSCs is less of a worry. Dr. Wu explained to me this way:

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Joseph Wu

“If iPSCs carry substantial epigenetic memory of the cell-of-origin, it is unlikely these iPSCs can differentiate to a functional cardiac cell or blood vessel cell. Only the stem cells free of significant epigenetic memory can differentiate into functional cells.”

 

Hopefully these results hold up over time because it will bode well for the countless iPSC-related disease studies as well as the growing number of iPSC-related projects that are nearing clinical trials.

Stem cell-derived mini-intestines reveal bacteria’s key role in building up a newborn’s gut

The following factoid may induce an identity crisis for some people but it is true that our bodies carry more microbes than human cells. Some studies in 1970’s estimated the ratio at 10:1 though more recent calculations suggest we’re merely half microbe, half human.

Because microbes are much smaller than human cells they make up only about 1 or 2 percent of our total body mass. But that still amounts to trillions of micro-organisms, mostly bacteria, that live on and inside our bodies. The gut is one part of our body that is teeming with bacteria. Though that may sound gross, you’re very life depends on them. For example, these bacteria allow us to digest foods and take up nutrients that we wouldn’t be able to otherwise.

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E. coli bacteria, visible in this enhanced microscope image as tiny green rods, were injected into the center of a germ-free hollow ball of cells called a human intestinal organoid (inset image, top right). Within 48 hours, the cells formed much tighter connections with one another, visible as red in this image. Image courtesy of University of Michigan.

When we’re first born our intestines are germ-free but overtime helpful bacteria gain access to our gut and help it function, protecting it from infection by the continual exposure to harmful bacteria and viruses. New research out of the University of Michigan Medical School reported in eLife now shows that the initial bacterial infiltration is even more important than scientists previously thought. It appears to play a key role in stimulating human gut cells to shore up the intestine in preparation for the full wave of both micro-organisms and pathogens that are present throughout a person’s lifetime. The finding could help researchers discover methods to protect the gut from diseases like necrotizing enterocolitis, a rare but dangerous infection that strikes newborns.

To reach these conclusions, the research team grew human embryonic stem cells into miniature intestines in the lab. These so-called human intestinal organoids, or HIOs, are structures made up of a few thousand cells that form hollow tubes with many of the hallmarks of a bona fide intestine. The HIOs were first kept in a germ-free environment to mimic a newborn’s intestine. Then a form of helpful E. Coli bacteria, the same that’s often found in an infant’s diaper, was injected into the HIO and allowed to colonize the inside of the intestine.

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A single human intestinal organoid, or HIO — a hollow ball of cells grown from human embryonic stem cells and coaxed to become gut-lining cells. Scientists can use it to study basic gut development, and the effect of microbes on the cells, in a way that mimics the guts of newborn babies. Image courtesy of University of Michigan

The team observed several changes in gene activity shortly after the bacteria was introduced. Within a day or two, genes involved in producing proteins that fight off harmful microbes increased as well as genes that encode mucus production, a key part of protecting the cells that face the inside of the intestine. Other key features of a maturing intestine, such as tighter cell-to-cell connections and lowered oxygen levels were also stimulated by the presence of the bacteria. As co-senior author Vincent Young, M.D., Ph.D. explained in a press release, these results put the team in a position to uncover new insights about intestinal biology and disease:

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Vincent Young

“We have developed a system that faithfully reproduces the physiology of the immature human intestine, and will now make it possible to study a range of host-microbe interactions in the intestine to understand their functional role in health and disease.”

 

The particular mix of microbes found in one person versus another can differ a lot. And the impact of these differences on an individual’s health has been a trending topic in the media. Lead author David Hill, Ph.D., a postdoctoral fellow in the lab of Jason Spence, Ph.D., thinks that’s one specific research path that they aim to investigate with their HIO system:

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David Hill

“We hope to examine whether different bacteria produce different types of responses in the gut. This type of work might help to explain why different types of gut bacteria seem to be associated with positive or negative health outcomes.”

 

Streamlining Stem Cell Therapy Development for Impatient Patients

During this third week of the Month of CIRM, we are focusing on CIRM’s Infrastructure programs which are all focused on helping to accelerate stem cell treatments to patients with unmet medical needs.

Time is money. It’s a cliché but still very true, especially in running a business. The longer it takes to get things done, the more costs you’ll most likely face. But in the business of developing new medical therapies, time is also people’s lives.

Currently, it takes about eight years to move a promising stem cell treatment from the lab into clinical trials. For patients with fatal, incurable diseases, that is eight years too long. And even when promising therapies reach clinical trials, only about 1 out of 10 get approved, according to a comprehensive 2014 study in Nature Biotechnology. These sobering stats slow the process of getting treatments to patients with unmet medical needs.

While a lack of therapy effectiveness or safety play into the low success rate, other factors can have a significant impact on the delay or suspension of a trial. An article, “Why Do Clinical Trials Fail?” in Clinical Trials Arena from a couple years back outlined a few. Here’s a snippet from that article:

  • “Poor study design: Selecting the wrong patients, the wrong dosing and the wrong endpoint, as well as bad data and bad site management cause severe problems.”
  • “Complex protocol: Simple is better. A complex protocol, which refers to trying to answer too many questions in one single trial, can produce faulty data and contradictory results.”
  • “Poor management: A project manager who does not have enough experience in costing and conducting clinical trials will lead to weak planning, with no clear and real timelines, and to ultimate failure.”

CIRM recognized that these clinical trial planning and execution setbacks can stem from the fact that, although lab researchers are experts at transforming an idea into a candidate therapy, they may not be masters in navigating the complex regulatory requirements of the Food and Drug Administration (FDA). Many simply don’t have the experience to get those therapies off the ground by themselves.

Lab researchers are experts at transforming an idea into a candidate therapy but most are inexperienced at navigating the complex regulatory requirements of the Food and Drug Administration (FDA).

So, to help make this piece of the therapy development process more efficient and faster, the CIRM governing Board last year approved the launch of the Translating Center and Accelerating Center: two novel infrastructure programs which CIRM grantees can tap into as they carry their promising candidate therapies from lab experiments in cells to preclinical studies in animals to clinical trials in people. Both centers were awarded to QuintilesIMS which collectively dubbed them The Stem Cell Center.

The Stem Cell Center acts as a one-stop-shop, stem cell therapy development support system for current and prospective CIRM grantees, giving them advanced priority for QuintilesIMS services. So how does it work? When a scientist’s initial idea for a cell therapy gains traction and, through a lot of effort in the lab, matures into a bona fide therapy candidate to treat a particular disease, the next big step is to prepare the therapy for testing in people. But that’s easier said than done. To ensure safety, the Food and Drug Administration requires a rigorous set of tests and documentation that make up an Investigational New Drug (IND) application, which must be submitted before any testing in people take can place in the U.S.

That’s where the Translation Center comes into the picture. It carries out the necessary research activities to show, as much as is possible in animals, that the therapy is safe. The Translating Center also helps at this stage with manufacturing the cell therapy product so that it’s of a consistent quality for both the preclinical and future clinical trial studies. If all goes as planned, the grantee will have the necessary pieces to file an IND. At this stage, the Translating Center coordinates with the Accelerating Center which focuses on supporting the many facets of a clinical study including the IND filing, clinical trial design, monitoring of patient safety, and project management.

Because the work of Translating and Accelerating Centers is focused on these regulatory activities day in and day out, they have the know-how to pave a clearer path, with fewer pitfalls, for the grantee to navigate the complex maze we call cell therapy development. It’s not just helpful for the researchers seeking approval from the FDA, but it helps the FDA too. Because cell therapies are still so new, creating a standardized, uniform approach to stem cell-based clinical trial projects will help the FDA streamline their evaluation of the projects.

Ultimately, and most importantly, all of those gears running smoothly in sync will help leave a lasting legacy for California and the world: an acceleration in the development of stem cell treatment for patients with unmet medical needs.

Building California’s stem cell research community, from the ground up

For week three of the Month of CIRM, our topic is infrastructure. What is infrastructure? Read on for a big picture overview and then we’ll fill in the details over the course of the week.

When CIRM was created in 2001, our goal was to grow the stem cell research field in California. But to do that, we first had to build some actual buildings. Since then, our infrastructure programs have taken on many different forms, but all have been focused on a single mission – helping accelerate stem cell research to patients with unmet medical needs.
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In the early 2000’s, stem cell scientists faced a quandary. President George W. Bush had placed limits on how federal funds could be used for embryonic stem cell research. His policy allowed funding of research involving some existing embryonic stem cell lines, but banned research that developed or conducted research on new stem lines.

Many researchers felt the existing lines were not the best quality and could only use them in a limited capacity. But because they were dependent on the government to fund their work, had no alternative but to comply. Scientists who chose to use non-approved lines were unable to use their federally funded labs for stem cell work.

The creation of CIRM changed that. In 2008, CIRM launched its Major Facilities Grant Program. The program had two major goals:

1) To accommodate the growing numbers of stem cell researchers coming in California as a result of CIRM’s grants and funding.

2) To provide new research space that didn’t have to comply with the federal restrictions on stem cell research.

Over the next few years, the program invested $271million to help build 12 new research facilities around California from Sacramento to San Diego. The institutions used CIRM’s funding to leverage and attract an additional $543 million in funds from private donors and institutions to construct and furnish the buildings.

These world-class laboratories gave scientists the research space they needed to work with any kind of stem cell they wanted and develop new potential therapies. It also enabled the institutions to bring together under one roof, all the stem cell researchers, who previously had been scattered across each campus.

One other important benefit was the work these buildings provided for thousands of construction workers at a time of record unemployment in the industry. Here’s a video about the 12 facilities we helped build:

But building physical facilities was just our first foray into developing infrastructure. We were far from finished.

In the early days of stem cell research, many scientists used cells from different sources, created using different methods. This meant it was often hard to compare results from one study to another. So, in 2013 CIRM created an iPSC Repository, a kind of high tech stem cell bank. The repository collected tissue samples from people who have different diseases, turned those samples into high quality stem cell lines – the kind known as induced pluripotent stem cells (iPSC) – and then made those samples available to researchers around the world. This not only gave researchers a powerful resource to use in developing a deeper understanding of different diseases, but because the scientists were all using the same cell lines that meant their findings could be compared to each other.

That same year we also launched a plan to create a new, statewide network of clinics that specialize in using stem cells to treat patients. The goal of the Alpha Stem Cell Clinics Network is to support and accelerate clinical trials for programs funded by the agency, academic researchers or industry. We felt that because stem cell therapies are a completely new way of treating diseases and disorders, we needed a completely new way of delivering treatments in a safe and effective manner.

The network began with three clinics – UC San Diego, UCLA/UC Irvine, and City of Hope – but at our last Board meeting was expanded to five with the addition of UC Davis and UCSF Benioff Children’s Hospital Oakland. This network will help the clinics streamline challenging processes such as enrolling patients, managing regulatory procedures and sharing data and will speed the testing and distribution of experimental stem cell therapies. We will be posting a more detailed blog about how our Alpha Clinics are pushing innovative stem cell treatments tomorrow.

As the field advanced we knew that we had to find a new way to help researchers move their research out of the lab and into clinical trials where they could be tested in people. Many researchers were really good at the science, but had little experience in navigating the complex procedures needed to get the green light from the US Food and Drug Administration (FDA) to test their work in a clinical trial.

So, our Agency created the Translating (TC) and Accelerating Centers (AC). The idea was that the TC would help researchers do all the preclinical testing necessary to apply for permission from the FDA to start a clinical trial. Then the AC would help the researchers set up the trial and actually run it.

In the end, one company, Quintiles IMS, won both awards so we combined the two entities into one, The Stem Cell Center, a kind of one-stop-shopping home to help researchers move the most promising treatments into people.

That’s not the whole story of course – I didn’t even mention the Genomics Initiative – but it’s hard to cram 13 years of history into a short blog. And we’re not done yet. We are always looking for new ways to improve what we do and how we do it. We are a work in progress, and we are determined to make as much progress as possible in the years to come.

Caught our eye: new Americans 4 Cures video, better mini-brains reveal Zika insights and iPSC recipes go head-to-head

How stem cell research gives patients hope (Karen Ring).
You can learn about the latest stem cell research for a given disease in seconds with a quick google search. You’ll find countless publications, news releases and blogs detailing the latest advancements that are bringing scientists and clinicians closer to understanding why diseases happen and how to treat or cure them.

But one thing these forms of communications lack is the personal aspect. A typical science article explains the research behind the study at the beginning and ends with a concluding statement usually saying how the research could one day lead to a treatment for X disease. It’s interesting, but not always the most inspirational way to learn about science when the formula doesn’t change.

However, I’ve started to notice that more and more, institutes and organizations are creating videos that feature the scientists/doctors that are developing these treatments AND the patients that the treatments could one day help. This is an excellent way to communicate with the public! When you watch and listen to a patient talk about their struggles with their disease and how there aren’t effective treatments at the moment, it becomes clear why funding and advancing research is important.

We have a great example of a patient-focused stem cell video to share with you today thanks to our friends at Americans for Cures, a non-profit organization that advocates for stem cell research. They posted a new video this week in honor of Stem Cell Awareness Day featuring patients and patient advocates responding to the question, “What does stem cell research give you hope for?”. Many of these patients and advocates are CIRM Stem Cell Champions that we’ve featured on our website, blog, and YouTube channel.

Americans for Cures is encouraging viewers to take their own stab at answering this important question by sharing a short message (on their website) or recording a video that they will share with the stem cell community. We hope that you are up for the challenge!

Mini-brains help uncover some of Zika’s secrets (Kevin McCormack).
One of the hardest things about trying to understand how a virus like Zika can damage the brain is that it’s hard to see what’s going on inside a living brain. That’s not surprising. It’s not considered polite to do an autopsy of someone’s brain while they are still using it.

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Microscopic image of a mini brain organoid, showing layered neural tissue and different groups of neural stem cells (in blue, red and magenta) giving rise to neurons (green). Image: Novitch laboratory/UCLA

But now researchers at UCLA have come up with a way to mimic human brains, and that is enabling them to better understand how Zika inflicts damage on a developing fetus.

For years researchers have been using stem cells to help create “mini brain organoids”, essentially clusters of some of the cells found in the brain. They were helpful in studying some aspects of brain behavior but limited because they were very small and didn’t reflect the layered complexity of the brain.

In a study, published in the journal Cell Reports, UCLA researchers showed how they developed a new method of creating mini-brain organoids that better reflected a real brain. For example, the organoids had many of the cells found in the human cortex, the part of the brain that controls thought, speech and decision making. They also found that the different cells could communicate with each other, the way they do in a real brain.

They used these organoids to see how the Zika virus attacks the brain, damaging cells during the earliest stages of brain development.

In a news release, Momoko Watanabe, the study’s first author, says these new organoids can open up a whole new way of looking at the brain:

“While our organoids are in no way close to being fully functional human brains, they mimic the human brain structure much more consistently than other models. Other scientists can use our methods to improve brain research because the data will be more accurate and consistent from experiment to experiment and more comparable to the real human brain.”

iPSC recipes go head-to-head: which one is best?
In the ten years since the induced pluripotent stem cell (iPSC) technique was first reported, many different protocols, or recipes, for reprogramming adult cells, like skin, into iPSCs have been developed. These variations bring up the question of which reprogramming recipe is best. This question isn’t the easiest to answer given the many variables that one needs to test. Due to the cost and complexity of the methods, comparisons of iPSCs generated in different labs are often performed. But one analysis found significant lab-to-lab variability which can really muck up the ability to make a fair comparison.

A Stanford University research team, led by Dr. Joseph Wu, sought to eliminate these confounding variables so that any differences found could be attributed specifically to the recipe. So, they tested six different reprogramming methods in the same lab, using cells from the same female donor. And in turn, these cells were compared to a female source of embryonic stem cells, the gold standard of pluripotent stem cells. They reported their findings this week in Nature Biomedical Engineering.

Previous studies had hinted that the reprogramming protocol could affect the ability to fully specialize iPSCs into a particular cell type. But based on their comparisons, the protocol chosen did not have a significant impact on how well iPSCs can be matured. Differences in gene activity are a key way that researchers do side-by-side comparisons of iPSCs and embryonic stem cells. And based on the results in this study, the reprogramming method itself can influence the differences. A gene activity comparison of all the iPSCs with the embryonic stem cells found the polycomb repressive complex – a set of genes that play an important role in embryonic development and are implicated in cancer – had the biggest difference.

In a “Behind the Paper” report to the journal, first author Jared Churko, says that based on these findings, their lab now mostly uses one reprogramming protocol – which uses the Sendai virus to deliver the reprogramming genes to the cells:

“The majority of our hiPSC lines are now generated using Sendai virus. This is due to the ease in generating hiPSCs using this method as well as the little to no chance of transgene integration [a case in which a reprogramming gene inserts into the cells’ DNA which could lead to cancerous growth].”

Still, he adds a caveat that the virus does tend to linger in the cells which suggests that:

“cell source or reprogramming method utilized, each hiPSC line still requires robust characterization prior to them being used for downstream experimentation or clinical use.”

 

Attractive new regenerative medicine tool uses magnets to shape and stimulate stem cells

The ultimate goal of tissue engineers who work in the regenerative medicine field is to replace damaged or diseased organs with new ones built from stem cells. To accomplish the feat, these researchers are developing new tools and techniques to manipulate and specialize stem cells into three dimensional structures. Some popular methods – which we’ve blogged about often – include the use of bioscaffolds as well as 3D bioprinting . This week, a research team at the Laboratoire Matière et Systèmes Complexes in France has developed an attractive (pun intended!) new tool that uses magnetized stem cells to both manipulate and stimulate the cells into 3D shapes.

The magnetic stretcher: this all-in-one system can both form and mechanically stimulate an aggregate of magnetized embryonic stem cells. Image: © Claire Wilhelm / Laboratoire Matière et systèmes complexes (CNRS/Université Paris Diderot).

The study, reported on Monday in Nature Communications, used embryonic stem cells which were incubated with magnetic nanoparticles. The cells readily take up the nanoparticles which allowed the scientists to group the individual cells using magnets. But first the team needed to show that the nanoparticles had no negative effects on the cells. Comparing the iron nanoparticle-laden stem cells to iron-free cells showed no difference in the cells’ survival and their ability to divide.

It was also important to make sure the introduction of nanoparticles had no impact on the stem cell’s pluripotency; that is, its ability to maintain its unspecialized state. A visual check of the cells through a microscope showed that they grew together in rounded clumps, a hallmark of undifferentiated, pluripotent cells. In addition, the key genes that bestow pluripotency onto embryonic stem cells were still active after the addition of the nanoparticles.

The stem cells’ ability to mature into various cell types, like heart muscle or nerve, is key to any successful tissue engineering project. So, the next important assessment of these magnetized cells was to make sure their ability to differentiate, or specialize, was still intact. The typical first step to differentiating embryonic stem cells is to form so-called embryoid bodies (EBs), which are 3D groups of pluripotent stem cells which begin differentiating into the three fundamental tissues types: mesoderm (gives rise to muscle, bone, fat), ectoderm (gives rise to nerve, hair, eyes), endoderm (gives rise to intestines, liver). Using a popular technique, called the hanging drop method, the team showed that the presence of the nanoparticles did not negatively affect embryoid body formation.

In fact, the use of magnets to form embryoid bodies provided several advantages over the hanging drop method. The hanging drop technique requires multiple, time-consuming steps and the resulting embryoid bodies tend to be inconsistent in size and shape. Use of the magnets, on the other hand, instantaneously assembled the stem cells into consistently round aggregates. And by precisely adjusting the magnetic force used, the scientists could also vary the size of the embyroid body, which is an important variable to control since the embryoid size can impact its ability to differentiate.

While the magnet used to form the embryoid bodies was kept stable, the researchers included another magnet which they could move. With this setup, the team was able to stretch and shape the group of cells without the need of scaffolds or the need to physically contact the cells. Several previous studies, using flat, 2-dimensional petri dishes, have shown that the stiffness and flexibility of the dish can stimulate gene activity by affecting cell shape. In this study, the researchers found that when the magnet was moved in a cyclical pattern that imitates the rhythm of a heart beat, the embryoid bodies were, if you can believe it, nudged toward a heart muscle fate. A press release by France’s National Center of Scientific Research (CNRS), which funded the study, explained the big picture implications of this new technique:

“This “all-in-one” approach, which makes it possible to build and manipulate tissue within the same system, could thus prove to be a powerful tool both for biophysical studies and tissue engineering.”

Extra dose of patience needed for spinal cord injury stem cell therapies, rat study suggests

2017 has been an exciting year for Asterias Biotherapeutics’ clinical trial which is testing a stem cell-based therapy for spinal cord injury. We’ve written several stories about patients who have made remarkable recoveries after participating in the trial (here and here).

But that doesn’t mean researchers at other companies or institutes who are also investigating spinal cord injury will be closing up shop. There’s still a long way to go with the Asterias trial and there’s still a lot to be learned about the cellular and molecular mechanisms of spinal cord injury repair, which could lead to alternative options for victims. Continued studies will also provide insights on optimizing the methods and data collection used in future clinical trials.

Human neuronal stem cells extend axons (green) three months after transplantation in rat model of spinal cord injury. Image: UCSD

In fact, this week a team of UC San Diego scientists report in the Journal of Clinical Investigation that, based on brain stem cell transplant studies in a rat model of spinal cord injury, recovery continues long after the cell therapy is injected. These findings suggest that collecting clinical trial data too soon may give researchers the false impression that their therapy is not working as well as they had hoped.

In this study, funded in part by CIRM, the researchers examined brain stem cells – or neural stem cells, in lab lingo – that were derived from human embryonic stem cells. These neural stem cells (NSCs) aren’t fully matured and give rise to nerve cells as well as support cells called glia. Previous studies have shown that when NSCs are transplanted into rodent models of spinal cord injury, the cells mature into nerve cells, make connections with nerves within the animal and can help restore some limb movement.

But the timeline for the maturation of the NSCs after transplantation into the injury site wasn’t clear because most studies only measured recovery for a few weeks or months. To get a clearer picture, the UCSD team analyzed the fate and impact of human NSCs in adult rats with spinal cord injury from 1 month to 1.5 years – the longest time such an experiment has been carried out so far. The results confirmed that the transplanted NSCs did indeed survive through the 18-month time point and led to recovery of movement in the animals’ limbs.

To their surprise, the researchers found that the NSCs continued to mature and some cell types didn’t fully specialize until 6 months or even 12 months after the transplantation. This timeline suggests that although the human cells are placed into the hostile environment of an injury site in an animal model, they still follow a maturation process seen during human development.

The researchers also focused on the fate of the nerve cells’ axons, the long, thin projections that relay nerve signals and make connections with other nerve cells. Just as is seen with normal human development, these axons were very abundant early in the experiment but over several months they went through a pruning process that’s critical for healthy nerve function.

Altogether, these studies provide evidence that waiting for the clinical trial results of stem cell-based spinal cord injury therapies will require an extra dose of patience. Team lead, Dr. Mark Tuszynski, director of the UC San Diego Translational Neuroscience Institute, summed it up this way in a press release:

Mark Tuszynski, UCSD

“The bottom line is that clinical outcome measures for future trials need to be focused on long time points after grafting. Reliance on short time points for primary outcome measures may produce misleadingly negative interpretation of results. We need to take into account the prolonged developmental biology of neural stem cells. Success, it would seem, will take time.”

Researchers, beware: humanized mice not human enough to study stem cell transplants

A researcher’s data is only as good as the experimental techniques used to obtain those results. And a Stanford University study published yesterday in Cell Reports, calls into question the accuracy of a widely used method in mice that helps scientists gauge the human immune system’s response to stem cell-based therapies. The findings, funded in part by CIRM, urge a healthy dose of caution before using promising results from these mouse experiments as a green light to move on to human clinical trials.

Humanized mice aren’t quite human. Illustration: Pascal Gerard

Immune rejection of stem cell-based products is a major obstacle to translating these therapies from cutting-edge research into everyday treatments for the general population for people. If the genetic composition between the transplanted cells and the patient are mismatched, the patient’s immune system will see that cell therapy as foreign and will attack it. Unlike therapies derived from embryonic stem cells or from another person, induced pluripotent stem cells (iPSC) are exciting because scientists can potentially develop stem cell-based therapies from a patient’s own cells which relieves most of the immune rejection fears.

But manufacturing iPSC-derived therapies for each patient can take months, not to mention a lot of money, to complete. Some patients with life-threatening conditions like a heart attack or stroke don’t have the luxury of waiting that long. So even with these therapies, many researchers are working towards developing non-matched cell products which would be available “off-the-shelf. In all of these cases, immune-suppressing drugs would be needed which have their own set of concerns due to dangerous side effects, like serious infection or cancer. So, before testing in humans begins, it’s important to be able to test various immune-suppressing drugs and doses in animals to understand how well a stem cell-based therapy will survive once transplanted.

But how do you test a human immune response to a human cell product in an animal? Believe it or not, researchers – some of whom are authors in this Cell Reports publication – developed “humanized mice” back in the 1980’s. These mice were engineered to lack their own immune system to allow the engraftment of a human immune system. Over the years, advances in this mouse experimental system has gotten it closer and closer to imitating a human immune system response to transplantation of mismatched cell product.

Close but no cigar, it seems.

The team in the current study performed a detailed analysis of the immune response in two different strains of humanized mice. Both groups of animals did not mount a normal, healthy immune response and so they could not completely reject transplants of various human stem cells or stem cell-based products. Now, if you didn’t know about the abnormally weak immune response in these humanized mice, you might conclude that very little immunosuppression would be needed for a given cell therapy to keep a patient’s immune system in check. But conclusively making that interpretation is not possible, according to team lead Dr. Joseph Wu, director of Stanford’s Cardiovascular Institute:

Joseph Wu. Photo: Steve Fisch/Stanford University

“In an ideal situation, these humanized mice would reject foreign stem cells just as a human patient would”, he said in a press release. “We could then test a variety of immunosuppressive drugs to learn which might work best in patients, or to screen for new drugs that could inhibit this rejection. We can’t do that with these animals.”

To uncover what was happening, the team took a step back and, rather than engrafting a human immune system into the mice, they engrafted immune cells from an unrelated mouse strain. Think of it as a mouse-ified mouse, if you will. When mouse iPSCs or human embryonic stem cells were transplanted into these mouse, the engrafted mouse immune system effectively rejected the stem cells. So, compared to these mice, some elements of the immune system in the humanized mouse strains are not quite capturing the necessary complexity to truly reproduce a human immune response.

More work will be needed to understand the underlying mechanisms of this difference. Other experiments in this study suggest that signals that inhibit the immune response may be elevated in the humanized mouse models. Dr. Leonard Shultz, a pioneer in the development of humanized mice at Jackson Laboratory and an author of this study, is optimistic about building a better model:

“The immune system is highly complex and there still remains much we need to learn. Each roadblock we identify will only serve as a landmark as we navigate the future. Already, we’ve seen recent improvements in humanized mouse models that foster enhancement of human immune function.”

Until then, the team urges other scientists to tread carefully when drawing conclusions from the humanized mice in use today.

CIRM weekly stem cell roundup: minibrain model of childhood disease; new immune insights; patient throws out 1st pitch

New human Mini-brain model of devastating childhood disease.
The eradication of Aicardi-Goutieres Syndrome (AGS) can’t come soon enough. This rare but terrible inherited disease causes the immune system to attack the brain. The condition leads to microcephaly (an abnormal small head and brain size), muscle spasms, vision problems and joint stiffness during infancy. Death or a persistent comatose state is common by early childhood. There is no cure.

Though animal models that mimic AGS symptoms are helpful, they don’t reflect the human disease closely enough to provide researchers with a deeper understanding of the mechanisms of the disease. But CIRM-funded research published this week may be a game changer for opening up new therapeutic strategies for the children and their families that are suffering from AGS.

Organoid mini-brains are clusters of cultured cells self-organized into miniature replicas of organs. Image courtesy of Cleber A. Trujillo, UC San Diego.

To get a clearer human picture of the disease, Dr. Alysson Muotri of UC San Diego and his team generated AGS patient-derived induced pluripotent stem cells (iPSCs). These iPSCs were then grown into “mini-brains” in a lab dish. As described in Cell Stem Cell, their examination of the mini-brains revealed an excess of chromosomal DNA in the cells. This abnormal build up causes various toxic effects on the nerve cells in the mini-brains which, according to Muotri, had the hallmarks of AGS in patients:

“These models seemed to mirror the development and progression of AGS in a developing fetus,” said Muotri in a press release. “It was cell death and reduction when neural development should be rising.”

In turns out that the excess DNA wasn’t just a bunch of random sequences but instead most came from so-called LINE1 (L1) retroelements. These repetitive DNA sequences can “jump” in and out of DNA chromosomes and are thought to be remnants of ancient viruses in the human genome. And it turns out the cell death in the mini-brains was caused by the immune system’s anti-viral response to these L1 retroelements. First author Charles Thomas explained why researchers may have missed this in their mouse models:

“We uncovered a novel and fundamental mechanism, where chronic response to L1 elements can negatively impact human neurodevelopment. This mechanism seems human-specific. We don’t see this in the mouse.”

The team went on to test the anti-retroviral effects of HIV drugs on their AGS models. Sure enough, the drugs decreased the amount of L1 DNA and cell growth rebounded in the mini-brains. The beauty of using already approved drugs is that the route to clinical trials is much faster and in fact a European trial is currently underway.

For more details, watch this video interview with Dr. Muotri:

New findings about immune cell development may open door to new cancer treatments
For those of you who suffer with seasonal allergies, you can blame your sniffling and sneezing on an overreaction by mast cells. These white blood cells help jump start the immune system by releasing histamines which makes blood vessels leaky allowing other immune cells to join the battle to fight disease or infection. Certain harmless allergens like pollen are mistaken as dangerous and can also cause histamine release which triggers tearing and sneezing.

Mast cells in lab dish. Image: Wikipedia.

Dysfunction of mast cells are also involved in some blood cancers. And up until now, it was thought a protein called stem cell factor played the key role in the development of blood stem cells into mast cells. But research reported this week by researchers at Karolinska Institute and Uppsala University found cracks in that previous hypothesis. Their findings published in Blood could open the door to new cancer therapies.

The researchers examine the effects of the anticancer drug Glivec – which blocks the function of stem cell factor – on mast cells in patients with a form of leukemia. Although the number of mature mast cells were reduced by the drug, the number of progenitor mast cells were not. The progenitors are akin to teenagers in that they’re at an intermediate stage of development, more specialized than stem cells but not quite mast cells. The team went on to confirm that stem cell factor was not required for the mast cell progenitors to survive, multiply and mature. Instead, their work identified two other growth factors, interleukin 3 and 6, as important for mast cell development.

In a press release, lead author Joakim Dahlin, explained how these new insights could lead to new therapies:

“The study increases our understanding of how mast cells are formed and could be important in the development of new therapies, for example for mastocytosis for which treatment with imatinib/Glivec is not effective. One hypothesis that we will now test is whether interleukin 3 can be a new target in the treatment of mast cell-driven diseases.”

Patient in CIRM-funded trial regains use of arms, hands and fingers will throw 1st pitch in MLB game.
We end this week with some heart-warming news from Asterias Biotherapeutics. You avid Stem Cellar readers will remember our story about Lucas Lindner several weeks back. Lucas was paralyzed from the neck down after a terrible car accident. Shortly after the accident, in June of 2016, he enrolled in Asterias’ CIRM-funded trial testing an embryonic stem cell-based therapy to treat his injury. And this Sunday, August 13th, we’re excited to report that due to regaining the use of his arms, hands and fingers since the treatment, he will throw out the first pitch of a Major League Baseball game in Milwaukee. Congrats to Lucas!

For more about Lucas’ story, watch this video produced by Asterias Biotherapeutics: