Cytocentric Visionary: Budd Tucker, University of Iowa
With iPSC and CRISPR, Leave Nobody Behind, Leave Nobody Blind
Dr. Budd Tucker is the Stephen A. Wynn Associate Professor of Regenerative Ophthalmology at the Stephen A. Wynn Institute for Vision Research, University of Iowa, where his lab is working on developing cGMP-compliant production of patient-specific retinal cell regenerative medicine using Xvivo Systems from BioSpherix. The transcript was edited for length.
We see you as a Cytocentric Visionary because you’re providing the best possible environment for cell and tissue production. What made you want to target vision related diseases?
I’ve been in vision research since 2006 when I was a post-doc in the laboratory of Michael Young at Harvard Medical School. My PhD was in neuroscience and I became interested in vision mainly because my wife has an eye disease. So being a neuroscientist, I figured why not work in the retina?
The retina is the ideal site from a therapeutic perspective. Unlike the central nervous system, there’s no myelin sheath in the retina. The photoreceptor cells, which die in the majority of these diseases, connect with an interneuron that is just microns away. It’s a very amenable site for transplantation and cell replacement.
Over the last ten years, I’ve developed a program for photoreceptor cell replacement. It’s patient-based. Everything we do is a translational project for autologous cell replacement or using patient-specific stem cells to evaluate things like pathophysiology, gene- or drug-based therapeutics.
Do you think that in order to be patient-centric, you first have to be cytocentric?
Yes, absolutely. That’s the entire basis for our program. I came to the University of Iowa in 2010, I was recruited by Ed Stone, who’s the world’s leading molecular ophthalmologist. Since then, our team has collected fibroblasts and keratinocytes cell lines from over 1800 patients. That’s 3600 cell lines from individuals with inherited eye disease. So we are as cytocentric as it gets.
Everything in our program is focused on the use of cell lines from patients. Many of our publications are disease modeling in nature, i.e. focused on evaluating pathophysiology of newly identified disease causing genes/mutations. This is where the CRISPR based approach is so powerful. We can now make isogenic controls from individuals with these rare diseases regardless of ethnic background. Unlike age matched controls, isogenic controls vary at the targeted genetic loci only. This allows us to ask for instance if newly identified genetic mutations are truly disease causing or related to ethnic background, i.e. a non-disease causing polymorphism.
When you generate all those primary cell lines are you protecting those cells from room air while those are being isolated?
For our autologous cell work, all cell lines are isolated in the Xvivo System in our non-profit cGMP facility. We bring the skin biopsies in, isolate dermal fibroblasts from each patient, and freeze them down. Then when we want to go on to do the next study we pull them out, expand them, and create iPS cells from them. We’ve generated about 450 iPS cell lines via this process to date.
That’s an astounding number.
It is quite high, but the thing that people kind of take for granted is the value of that primary cell line. This is a human patient who has given us a skin biopsy and we have over 1800! It’s absolutely astounding that we’ve been able to do it. It really speaks for the power of the program that Ed Stone has created here at the University of Iowa.
And it speaks for your ability to communicate effectively to patients, which is not a small thing.
No. One of my most proud papers is certainly not one of the most high impact papers. I have a paper on RPE65-associated LCA. It is a novel intronic mutation in a young child from Haiti. We had no idea if this was actually a disease causing variant or not.
We took skin from the little girl, made retinal pigmented epithelial cells, and showed that this mutation altered normal splicing and as such was in fact predicted to be disease causing and not some rare non-disease causing variant in the Haitian population. Armed with this new knowledge she is now eligible for one of the ongoing RPE65 gene augmentation trials.
So now your group is using CRISPR to actually repair some of the genetic defects that cause vision loss?
We use CRISPR-based genome editing every day. It was another one of these revolutionary tools that, like the iPS cell, completely changed our strategy. It gives us the ability to selectively target and correct a patient’s disease causing mutation, which was the last step required in our entire autologous cell replacement program.
Gene augmentation is the second arm to our program, the one that we’re developing for patients with early stage disease. If you have lots of photoreceptor cells left, the goal is not to replace them, the goal is to prevent them from dying, and of course in rare cases like RPE65 associated LCA, restore function.
Cell therapy, unlike gene therapy, is targeted for late stage disease. So with a patient with an inherited Mendelian disease, something like retinitis pigmentosa, you would first, after generation of the iPS cells, want to correct the patients disease causing gene. We now do that in the Xvivo System. We are actually in the process of developing a robotic system with a group from the Cleveland Clinic, called the Cell-X to selectively pick iPS cells that have been CRISPR corrected. All of that sits inside of the Xvivo unit.
To protect the whole process from variable room air?
The beauty with the Xvivo System is that this is all done in isolation. So you can maintain your cells in this controlled environment. For gene therapy we use HEK293 cells to produce our virus and it’s very consistent.
So I absolutely see the Xvivo units being critical for our process from both the gene augmentation and cell therapy sides. Especially for rare diseases, in which a small dedicated cGMP facility can be used for small batch production.
This helps with cost of production as well?
The rarity of these disorders really necessitates the need for a non-profit approach. Our goal is to make these things as inexpensive as possible, and provide them at cost so that they can be made available to patients in need. That’s the mission behind our program. The goal is to provide the treatment to as many patients as we can. If you walk through any of our labs we all have these pictures of families that are patients of ours here in the clinic at the University of Iowa. Our slogan is “Leave no one behind.”
It doesn’t matter how rare the disease is, it’s still a disease that somebody has. We have a nonprofit, philanthropically funded, leave no one behind strategy and that’s what makes it so unique.
Is that where your legacy is going to lie, in these tremendous individual patient impacts?
Well I hope so. Without Ed Stone and the other physicians seeing patients, my program wouldn’t exist. So I could not do what I do without those people, and to a certain degree it would be very difficult for any of those people to do what I do. So I think that you just touched on something that’s again one of my proudest moments.
It is absolutely true that I’ve been in the right place at the right time a lot of times. The only thing I’ll take credit for was seeing greatness in leaders like Ed Stone and being able to hook my wagon up to theirs. Being able to partner with some of these real leaders to build this program has been absolutely essential.
But now to your question; that is absolutely what I see my legacy being. I see it being using patient-specific/patient-centric approaches and cell-centric approaches to help individuals. This is why I did it. This is the entire reason why I switched from doing work in the spine as a graduate student to the retina as a post-doc. I was approached by Ed Stone in 2010 and he came to me at ARVO, our annual meeting, and asked me if I knew anyone that would be interested in moving to Iowa and I said, “Yeah, I’m definitely movable.”
The next day we met and not a word of a lie, he took one of these big yellow legal pads, put it in front of me, and said, “Write down everything you need to cure blindness and we’ll get it for you.” Sixteen weeks later I was living in Iowa.
So this is my program. This is what I’m all about and it’s not some big altruistic thing because I get a huge amount out of this. I mean the resources I have are incredible! It really has been driven by the Iowa program.
And in the last three years, you’ve published 45 papers.
We’ve been pretty productive.
We look forward to literally seeing what you do next!
See Part 2: Modularity is Key to Cell Therapy Process Flexibility here.
If you would like to be featured in our Cytocentric Visionary Series, contact us. We would love to hear about your work.
- 1. Tucker, B., et al., Using patient specific iPSCs to interrogate the pathogenicity of a novel RPE65 cryptic splice site mutation and confirm eligibility for enrollment into a clinical gene augmentation trial. Translational research: the journal of laboratory and clinical medicine, 2015. 166(6): p. 740.
- 2. Burnight, E.R., et al., Using CRISPR-Cas9 to Generate Gene-Corrected Autologous iPSCs for the Treatment of Inherited Retinal Degeneration. Mol Ther, 2017. 25(9): p. 1999-2013.
About the Author
Alicia D Henn, PhD, MBA
Alicia Henn has been the Chief Scientific Officer of BioSpherix, Ltd for two years. Previously, she was a researcher at the Center for Biodefense Immune Modeling in Rochester, NY. Alicia obtained her PhD in molecular pharmacology and cancer therapeutics from Roswell Park Cancer Institute in Buffalo, NY and her MBA from the Simon School at University of Rochester in Rochester, NY.