For a decade, D. Kacy Cullen—in his labs at Penn and the VA Medical Center—has chipped away at risky research. With his fair share of breakthroughs, as well as challenges, the lifesaving impact of his work is finally in sight.
Today, Cullen, an associate professor of neurosurgery in the Perelman School of Medicine, can count among his many accomplishments the successful development of technology to create the first implantable tissue-engineered brain pathways. It’s an effort that, if translated, could essentially reverse the clock on disease progression for patients with neurodegenerative disorders, such as Parkinson’s disease.
“It’s a radically different approach than most regenerative medicine efforts in the brain,” says Cullen, also director of the Center for Neurotrauma, Neurodegeneration, and Restoration at the VA. “We are building the entire 3D architecture of the construct outside of the brain, and then implanting it as a unit.”
Met with mounds of skepticism for such far-out research, Cullen mostly bootstrapped, with initial help from internal Penn grants, his first five years of work. Ever since, he and his team have not only been able to create the pathways, but also take them through proof-of-concept with efficacy studies in small animal models. Most recently, they have showcased the ability to build these tissue-engineered brain pathways using neurons derived from the human stem cell line.
With three patents, at least eight published papers, $3.3 million in funding, and a productive go with the Penn Center for Innovation’s I-Corps program this past fall, Cullen is ready to take this project’s findings to the next level with the creation of a brand new startup company: Innervace.
“It’s really surreal to think that I’ve been working on this project, this approach, for 10 years now,” says Cullen. “It really was doggedness to just keep pushing in the lab, despite the challenges in getting extramural funding, despite the skepticism of peer reviewers. But we’ve shown that we’re able to do it, and that this is a viable technology.”
Path to pathways
Cullen, originally trained as a biomedical engineer, came to Penn as a post-doctoral fellow in 2007, working in the lab of neurosurgery professor Doug Smith, director of the Center for Brain Injury and Repair.
It’s Smith who Cullen credits for inspiring him to pursue this particular research, which Cullen spun out into his own lab at Penn in 2009. Smith is named a co-founder of Innervace with Cullen.
“My post-doctoral training with Doug Smith was instrumental because he understood the importance of pathways in the brain,” says Cullen. “The majority of the area of the human brain, it’s not nerve cells themselves, it’s the long-distance fibers that are connecting discrete processing centers—unique clusters in the brain.”
And when these pathways—which enable signals to be sent from one region of the brain to another—are lost, Cullen adds, “It’s really devastating for patients.”
Long-distance pathways in the brain form when one is an embryo, when target and source are in relatively intimate proximity, Cullen explains. As the brain expands during embryogenesis and then adolescence, these pathways lengthen to the point where it becomes insurmountable to expect regeneration if they are lost due to trauma, stroke, or a neurodegenerative disease.
But, thanks to Smith’s insight, Cullen became determined to flip this “impossibility” on its head. In his lab, Cullen had set out to tackle something never seen before.
Breaking new ground
Despite many years of effort, scientists have lacked the ability to regenerate long-distance pathways in the brain. So, Cullen took a road less traveled, attempting to build the pathways outside of the body—in vitro.
“You have a lot of different tools at your disposal that allow you to grow long-distance axon tracks, so the concept was to build these long tracks outside the body where we have greater control, we can mitigate many of the inhibitory factors, and really create an ideal environment,” says Cullen. “Then we implant these constructs.”
The tissue-engineered pathways would be personalized—a precision medicine-type therapeutic, Cullen explains. By collaborating with stem cell scientists, such as assistant professor of neurosurgery Isaac Chen, Cullen and his team would pair their newfound microtissue engineering techniques with patients’ very own cells.
“We’d create what are called induced pluripotent stem cells, and then push these to be the specific neuronal population that the patient may be missing,” says Cullen. “We’d be able to grow these into the structure, and hence the function of the brain pathways that the patient is missing.”
The patient would then see a neurosurgeon to have the structure precisely implanted, reconstituting the pathway.
An approach never seen before, it would not only replace the brain cells that a given patient is missing, but in one fell swoop put back the connections that region of the brain had to distant regions in the brain.
“It’s basically recreating the anatomy that a patient is missing on a per patient basis,” says Cullen.
Raising the bar
Cullen has pinpointed Parkinson’s disease as Innervace’s first culprit.
A disease that affects about one million people in the United States, and 10 million worldwide, Parkinson’s offers a rather common, early vulnerability: The loss of dopaminergic neurons in the area of the brain known as the substantia nigra.
These specific neurons have long axons that project to a higher part of the brain—the striatum—one of the most important motor control circuits.
“When this particular pathway succumbs to the disease, that’s when patients experience tremors, trouble with posture, and trouble with balance,” says Cullen.
In a miniature, transplantable form, Cullen and his team have tissue-engineered the first nigrostriatal pathway—which connects the substantia nigra to the striatum—and successfully implanted it into a small animal model.
These findings, described by Cullen as a “herculean effort,” and led by fifth-year bioengineering doctoral candidate Laura Struzyna, a researcher in Cullen’s lab, were recently published in the Journal of Tissue Engineering and Regenerative Medicine.
“The thing about neural tissue engineering is that you are working with living cells, and neurons in particular are a pretty finicky cell type—they just need an excuse to die and your experiment is ruined,” says Struzyna, who studied bioengineering at Duke before coming to Penn. “Working with the living neurons has been the trickiest part of it because before you even get to the point where you are testing hypotheses and seeing the functionality of the constructs, you have to get them to survive long term and mature in the way that they are supposed to.”
Enduring the ups and the downs, the team’s hard work was well worth it: As of late, in the small animal model, they’ve seen outgrowth and histological evidence of integration—something Struzyna says she’s “really excited about.”
A translational proof-of-concept, Cullen and his team have also recently been able to build these pathways at a bigger scale and with actual human stem cells. Their next step is to show efficacy of the implantable construct in a large animal model.
“That will be a big benchmark showing we’re ready to start a clinical trial,” says Cullen. “And we can see that on the horizon in the next several years.”
Gaining credibility
The epitome of a fighter, Cullen kept pushing forward even after being told time and time again that his idea would never work.
“It was really, really tough to get funding, just because this approach is so radical,” he says. “I was basically bootstrapping it all with my startup package from Penn until about 2013.”
Finally, his first grant on the topic came in the form of seed funding from the Penn Neuroscience Center. Specifically designed for early-stage, risky work, the $25,000 grant “definitely accelerated things,” Cullen says.
By 2015, he was able to secure $375,000 from the Michael J. Fox Foundation, followed by $2.1 million from the National Institutes of Health.
“It was a long haul,” he says. “The key was to keep de-risking it by doing it. That took forever.”
In 2017, Cullen received another major grant—$900,000—from the U.S. Department of Veterans Affairs.
Reflecting on his experience, Cullen says, the infrastructure that Penn set up “worked perfectly.”
“Things that were really not ready for prime time for an extramural agency, but were very promising or transformational concepts, the mechanisms existed at Penn to really advance the work and give it a shot in the arm,” says Cullen. “This project is definitely a success story because [what Penn provided] led directly to extramural funding.”
Bench work
Throughout the years, Cullen has utilized resources from the University and Health System, as well as the Corporal Michael J. Crescenz VA Medical Center—located just across the street from campus. With labs at both institutions, made up of a team of 24 graduate and undergraduate students, postdocs, technicians, and fellows, he says he’s gotten “the best of both worlds.”
“We’re able to leverage the best minds and collaborators that Penn uniquely has, recruit and train students, and have world-class lab space,” Cullen says, “but also leverage the strengths of the VA and their intramural programs.”
For Struzyna, she says she feels fortunate to not only get to work with Cullen on groundbreaking discoveries, but while doing so, also have access to his top-quality labs with state-of-the-art equipment.
“I think that a lot of graduate students, their entire project is just in-vitro with cells, but based upon the funding Kacy’s been able to acquire and the environment we’re in, we have access to a lot of sophisticated lab or core equipment,” she says, specifically naming a two-photon microscope, a high-speed imaging microscope, and a confocal microscope. “Being a part of this lab has given me that access and capability that others don’t have, and I feel very lucky for that.”
Justin Burrell’s experience working in Cullen’s labs has been just as rewarding—albeit different. Burrell, a first-year bioengineering Ph.D. student, has been somewhat of a face for the commercialization aspect of Innervace.
With a master’s in neuroscience from Tulane under his belt, Burrell joined Cullen’s team four years ago with the hope of learning the ins and outs of bringing a lab-based idea to market. Under Cullen’s direction, last fall Burrell helped lead Innervace through the Penn Center for Innovation’s I-Corps process.
Open for business
I-Corps, an accelerator program operated by the Penn Center for Innovation (PCI) and supported by the National Science Foundation, offers various participants a two-week-long crash course in business skills. Penn teams with ideas for startups learn the best ways to go about market research, strategic partnerships, and garnering seed funding from potential investors.
The Innervace team—formerly known as ReNeuron—was made up of Burrell, as well as PCI’s Neal Lemon and Derrick Leach, and PCI fellow Sudiksha Sridhar, a master’s student in the School of Engineering and Applied Science’s Department of Chemical and Biomolecular Engineering.
“We were the first group to do a hybrid Penn/PCI I-Corps group, and worked directly with members of PCI’s licensing and marketing offices,” says Burrell. “It was really useful for us to interview and meet with customers, to get a sense of what they are actually looking for—ultimately de-risking our technology for the future.”
Through Innervace’s patent and LLC process, Cullen had already been working closely with Lemon, an associate director of PCI’s licensing group at Penn Medicine. Lemon helps startups get their feet off the ground by reviewing intellectual property, putting cases forward for patenting, and finding licensing partners.
With a Ph.D. in neuroscience as well as an MBA degree, when Lemon joined the University a year ago, he took a close interest in Cullen’s work.
“I was actually surprised he accomplished this, and a little skeptical on how feasible it would be,” says Lemon. “But after reviewing the patent applications, his peer-reviewed publications, and the really good success he’s had at obtaining funding, you quickly realize it’s a legitimate process.”
Now, Lemon is working with Cullen to fast-track Innervace’s development and translation by seeking strategic partners.
“If we can get a potential solution for people suffering from Parkinson’s, and reduce the amount of time it takes to get that solution to a point where we’re delivering treatment, I’ll be happy,” Lemon says.
Pressing forward
Given Cullen’s current workflow and pace of funding, although exceptional, Lemon predicts the technology could take another seven years to develop before it actually gets to a point where it can be tested in a human. But with investment—about $15 million—from a strategic partner, such as a drug or medical device company, that timeline could fall to just four or five years.
“We already have a dedicated site at Penn where the CAR T cell therapy work is being done,” says Lemon. “From an investment point of view, I think it’s pretty attractive that a lot of the infrastructure exists here to get Innervace’s proof-of-concept off the ground.”
Cullen’s goal all along has been to have his research benefit patients, and he’s “worked very hard to build a lab that’s meant to be a translational pipeline,” he says.
The caveat—or, perhaps potential—Cullen notes, is that the world is just now facing the beginning of an era of regenerative medicine: It’s unclear how these different therapies should translate, how the products should look, or how long it should all take. For now, the model of moving these R&D discoveries out of the lab is uncertain.
“We don’t quite have this huge bio-manufacturing area that you might find in electronics or automotive or aerospace, other engineering endeavors,” says Cullen. “It doesn’t quite exist yet for cell and tissue manufacturing, but it will.”
“This is going to be the revolution of the 21st century,” Cullen insists.
Serving, hopefully, as a blueprint “for many other therapies to follow,” Cullen says, is his method of crafting a startup that thrives by partnerships with industry. These strategic partners, for instance, would facilitate venture capital to accelerate development and translation, and include a team with product development and regulatory specialists, and advisers for the eventual clinical trials.
“The idea is to really move something like an eight- to 10-year runway down to closer to a three- to four-year runway,” Cullen says. “And really position Penn as a leader for translational regenerative therapies to greatly impact human health and care.”
Nothing becomes something
The past 10 years have been a whirlwind for Cullen and his team. They took something that “reviewers thought was impossible,” Cullen recalls, and hit breakthrough after breakthrough—“making this a real possibility.”
Even so, he knows it’s inevitable he’ll face challenges going forward, with his research as well as the regenerative medicine field in general.
“We’ve shown at least proof-of-concept for this approach, but it would be really dangerous at this point to extrapolate to people,” he says. “We have our eyes open, we know there are a lot of challenges that we can anticipate, there’s probably even more challenges that we’ve yet to anticipate.”
But, he says, “if we have the funds to put the right team around this and the bodies to accelerate the effort, then we have a shot to be, if it’s successful, helping patients that much sooner.”
Photo on homepage: Researchers in Cullen’s lab, including Justin Burrell (left) and Laura Struzyna (right), are finally seeing their work’s lifesaving potential.
