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Oby: From the campus of Harvard Medical School, this is ThinkResearch, a podcast devoted to the stories behind clinical research. I'm Oby.

Brendan Keegan: And I'm Brendan. And we are your hosts. ThinkResearch is brought to you by Harvard Catalyst, Harvard University's Clinical and Translational Science Center.

Oby: And by NCATS, the National Center for Advancing Translational Sciences.

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Brendan Keegan: In the first episode of our miniseries, "Focusing on Translational Innovator," guest host Hardeep Ranu described the high-risk, high reward-nature of Dr. Elena Aikawa's project to understand the process of calcification in heart valves. In this second installment of the series, Dr. Aikawa and Mark Blaser, a research fellow in her lab, join Hardeep to discuss the importance of the support from Translational Innovator and where Doctor Aikawa's research is today. Doctor Elena Aikawa is the director of the Vascular Biology program at Brigham and Women's Hospital.

Hardeep Ranu: OK. So why did you apply for the award that you did, for the one working with me? You can also talk about the previous one as well in-- the microscopy one. But yeah.

Elena Aikawa: So they actually, interestingly, both awards are somehow related. They were not two separate studies. They were interconnected studies. And I think this is very important because we got this award two years in a row.

And actually, then it was extended for another year. So it really gave us the opportunity to complete the study in a way. So we published it with Mark as a co-first author for the paper in Nanomaterials. And I think it was extremely exciting.

So why did we do that? I don't know how much our listeners know about heart valve disease, probably very little. But heart valve disease is very-- a very big problem and an unmet medical need because there is no treatment for heart valve disease. And there are no good models to study heart valve disease, neither in vitro nor in vivo.

So because of that-- and the only treatment for heart valve disease is surgical or transcatheter valve replacement. So we need to understand that it's open heart surgery or invasive valve replacement. So that's why we needed to create some kind of model to study this disease in order to provide the drug to the community for the treatment because there is, again, no good models available. And no therapy is available.

So we came up with an idea that-- first of all, for the first part of the proposal several years ago, Josh Hutcheson, who is now an independent investigator, and myself investigated microcalcifications. And we came up with the idea how to grow microcalcifications in three-dimensional culture. So that gave us the boost to start by printing of those little tiny valves and imply microcalcifications-- or implicate microcalcifications into these bioprinting structures.

And why it was important? Because that could serve as a first three-dimensional model of valve and eventually for calcific aortic valve disease. And that was very, very novel. Actually, we patented this 3D structures and how we can grow microcalcifications.

And eventually when Mark joined the lab, we decided to go further and not just have little valves with microcalcifications which we could individually treat with different inhibitors or different-- to have osteogenic medium to induce those microcalcification growth, and so on. So when came, he came with the idea that we can extend this by printing to multi-well format. And that would allow us to do high throughput screening. So maybe Mark, you can speak from here, take from here.

Mark Blaser: Absolutely. And so for me, my background is engineering, but also valve pathobiology. And so this is a really interesting opportunity for me to meld those two areas and to be able to put something interesting out in that interdisciplinary space between the two of them.

And so from our perspective, the Catalyst grant gave us access to a bunch of high-tech, cutting-edge imaging technology that we didn't have available on site but that allowed us to do a lot of work in terms of validating the high-throughput approach, validating the pathobiology we develop inside of these small bioprinted structures. It led us to collaborations with people like Robert Langer at MIT to be able to push this bioprinting technology to the next level.

And then, like Elena was saying, it really potentiates using these structures for drug discovery because the reality is that in the drug discovery field, typically you're screening millions of molecules against cells growing in flat 2D sheets. And those flat 2D sheets really don't recapitulate the microenvironment of the aortic valve. The valve is 3D. It's got different stiffnesses of tissue.

The cells inside of it respond to those stiffness in different ways when they undergo disease development. There's fluid flow on either side of the valve. And all these things influence how the valve responds to both disease, but also potentially to drugs. And so the idea was that one of the reasons that valve disease doesn't have drugs is simply the fact that our 2D screening approaches up to now haven't mimicked all of that interesting biomechanical biology.

So what we did then is we used 3D bioprinting to print arrays of heart valves on a chip, or valve organoids, or whatever sort of fancy buzzword you want to use to describe what we were doing here, but printing these highly reproducible arrays of small 3D structures into 96 well plates, for example. And in that format you can use each of those single wells to screen these millions of candidate drug molecules in a setup that is much more reflective of the needed in vivo environment of the heart valve. And so that was the approach that we took. That was one of the reasons that we were so successful as catalysts is we could leverage all the imaging and technology at HCBI and all the collaborators we had available to us to get that across.

And the other thing I want to mention is that from my practical side of things, me being a young fellow starting up in Elena's lab, it also had a lot of practical benefits for me. It was an opportunity to build a track record of grant funding and being able to write successful grants. Even in this relatively small pilot context, it's still really important to build that up.

It opened up access to the NCATS Reviewer Exchange Program for me. And so now I have a small but growing track record of being experienced as an NIH program peer reviewer, so other things from a career development point of view that have been side benefits to participate in this pilot program as well.

Hardeep Ranu: Yeah. Could you-- you mentioned that it's relatively small. There's $50,000. But you've got a lot out of it. Can you just talk about what you used the money for. Because clearly it was, like I said, well spent.

Elena Aikawa: Thank you. Thank you for noticing that actually. It's good. I didn't think about it. But it's actually a good point. Yes, indeed, we were able to patent our first 3D structures. Second, we published this paper in Nanomaterials with Mark as a co-author.

And then actually very, very recently, just because of one of the students who worked on that original project, Jessica Ruiz-- I don't know, you probably even didn't meet her. She was the first, first, first student who worked with George-- we just published PNS paper, where we actually used these small bio-printed structures, introduced microcalcifications in those structures, and treated them with bisphosphonates. And we showed that those microcalcifications could be reversible.

And I think this, in my opinion, is almost like breakthrough-- I'm sorry, breakthrough in the field because we still don't know whether microcalcifications or calcification in vivo, in humans, or in animals is reversible. But we were able to demonstrate it by imaging and by morphology of those microcalcifications. So this is what how many things, in addition to what Mark mentioned, we were able to get from that. And I agree, it's much more than $50k.

But how did we use $50k? We used it mostly for the imaging of Harvard imaging facility. And that was very helpful for us because we could do so many things. We could look at the calcium and phosphate inside those vesicles. We were confirming that they actually indeed calcify in vesicles.

We did this three-dimensional structure of the vesicles. And for the first time, we were actually able to see how microcalcifications form at the level of one single individual vesicle. So they aggregate with each other, they merge, and then they [? nucleate ?] [? hydroxy ?] [inaudible] and they become microcalcifications. So we were able to look at the [? kinetic ?] formation of microcalcifications using the Harvard imaging facility.

And otherwise, there is no modalities, other modalities on the market which allows us to look at those [? kinetic ?] of microcalcification formation and also how they reverse and become not harmful anymore. Mark?

Mark Blaser: Yeah. I'll just tag onto that. So basically that allowed us to access ultra high resolution confocal microscopy that allows us to really look at single vesicles undergoing the calcification process, like Elena was saying. It also allowed us to access some really cutting edge multiphoton modalities that allowed us to look deeper into tissues from our 3D bioprinted structures but also animal model tissues, to allow us to look more deeply into those to look at calcification forming deep inside these tissues, and in such a manner that you wouldn't be able to see from normal confocal microscopes, for instance.

And the third was the cell discoverer and its subsequent updates at HCBI, which is a high-content imaging system that specifically allows you to image frequently, repeatedly, and automatically over long periods of time multiple wells in a dish. So you could load your 96-well plate in there and ask it to image each well of the dish every hour for 20 days. And you can start to gain a rapidly automated, quantitative set of data for your bioprinted array. So that was what we were able to use to generate all this data in terms of how our bioprinted arrays were calcifying over time and how different molecules were able to modulate that classification. So those were the big things at HCBI.

We also used some of the funds to allow us to do some pilot sequencing at some of the Harvard facilities. And so that allowed us to validate some of the omics approaches we applied to validate disease development and disease inhibition in these arrays. And the last thing we did is we got access to some really cutting-edge scanning electron microscopy and transmission electron microscopy from Harvard [? cause ?] as well that allows us to validate some of the extracellular vesicle biology and calcification biology that we see in these tiny structures that are really ultra-high resolution. So those are the specifics that allowed us to really leverage these data sets into a validated and sort of tie a bow on this project and be able to publish it.

Hardeep Ranu: Yeah, that's great. That's really great. You can be honest about this, but what, if any, value did the project management or the meeting every four to six weeks-- in the meetings when we got everybody together, did you think there was a value added in that or not? Hopefully it's yes, but.

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Elena Aikawa: I can start, and Mark can finish because I think he has individual meetings with you, so he can talk about that. But I can talk about PI meeting. It with presentations from PIs and their teams.

So in my opinion, I mean, despite that it's actually time-consuming and you really need to spend half a day for discussions, but it gave us so much opportunity to look at what other labs are doing, and meet with other PIs, and meet with you guys with Catalyst leadership, including you and Gary Gray, who really-- I felt personally so much support from the Catalyst team. So that encouraged us to move forward, despite that was-- again, those meetings were indeed time-consuming.

But learning from others, talking to Catalyst leadership, seeing what other groups are doing, and which level of research they are at, and collaborating with them, we have a few collaborators, that was amazing. We also met with the group of people, patients who had scleroderma at that time. That was probably most, I don't know, heartbreaking event for me. And Mark, you were there, right? We were together.

Mark Blaser: Yes, I was.

Elena Aikawa: Yeah. And those patients really stimulated our thoughts. We met with several people after that to extend our collaboration and think how we can potentially treat scleroderma, even though it's not necessarily the same as the valve disease. But it has the same problem.

Those people have fibrosis and calcification. So potentially mechanisms could be similar. And this is something what I'm still looking forward to investigating in my future studies.

Mark Blaser: Thanks, Elena. And so I would say, in addition to what Elena said about the large update meetings, I totally agree, it was a really nice opportunity to just cross-incubate ideas amongst a group of PIs and Fellows that are really interested in innovation because that's a slightly separate focus in some sense to traditional biomedicine. So just being around people that had a similar focus, a similar idea, a similar set of values in terms of wanting to push the research in that direction, as well as the standard publishing approach, it was energizing, and it was nice to see.

I think Elena and I both developed some collaborators out of that, had great ideas, feedback from people that had backgrounds not at all associated with basic science that we're working on, but had ideas for local resources. Try this lab, try this sequencing facility, try these people with this microscope. They might be able to fix your problem.

Because we've got everything you would ever want in terms of biomedical research here in Boston. But knowing the specific person to go to who can solve your problem really quickly is-- that's like the lost art-- or the tricky bit of information that can save you months when it comes to putting a project together. So that was really beneficial.

Elena Aikawa: But I think Mark is completely right. Especially I think it was very helpful for postdoc levels because they learned from each other. They learned how to present to a different audience, not just their own peers, not just other postdocs. But actually, we had other PIs from different fields who are not doing the similar research.

So you needed to give a presentation in the language so everybody would understand that. So I think that was a very good experience actually for both, for PIs and postdocs as well. Sorry, Mark, I just talked over.

Mark Blaser: No, that sounds like the exact thought that I was finishing, the idea that speaking to industry, speaking to VCs, speaking to those sorts of people requires a different approach. Those people care about different things and different deliverables. And it was nice to at least get that ingrained into our heads early on before we would ever be going to approach those people was a nice little reminder and wake up call.

And then the other thing I'll say, Hardeep, about the project management and the meetings that we had regularly, for me, it was really consistent with the approach that I know the engineering industry takes, the project management, scheduling, deliverables, timelines. And so it was-- I think, A, for anyone in biomedical science who's interested in going into industry in the future, this is the sort of thing that we all have to get used to. And so experiencing it now is just a nice early introduction to that. And B, these are the sorts of things that are well proven to keep people on track and to have productive projects and have deliverables at the end of the time.

And so I think it was great. We don't always get that level of accountability, let's say, or-- that's not quite the right word, but being able to stick on task and stick to a timeline. And so I think from that respect, it was really helpful to have, I forget, was it once every month, once every two months, something like that.

Hardeep Ranu: It was like four to six weeks, probably every five weeks, something like that.

Mark Blaser: Yeah. Even just a check in every month or two and a troubleshooting check-in, are there issues, what can we do to help, who can we put you in contact with to get you over this particular developmental hump that you're dealing with right now? I think it also helped that no issues languished in the background. They were always brought to light, and we could overcome them regularly. And so I think that was a nice little addition.

And it's also nice to have someone external from the lab to talk to about that, just a new voice, a new person to bump those ideas off of. It's really useful. And it's a great opportunity and a nice part of the system that was set up.

Elena Aikawa: Yeah. Thank you for that, Hardeep. And thank you for your guidance so far.

Mark Blaser: Absolutely.

Hardeep Ranu: Yeah. So, well, for me, I enjoy it because I get to-- well, A, I get to visit a different building, different surroundings and see what other people's labs are like. And I also get to hear about your research. And for me, in terms of my lifelong learning, it's always a benefit for me to be, oh, OK, now I've learned something, the surface level of this particular type of research.

And Mark, you were saying something about innovation. And you're an engineer doing biology-- biomedical research. So one thing that I have just read is this book called Range by David Epstein.

And it's about people who are a generalist. So they have a broader background than somebody who's done something for-- the same thing forever. It's sort of the Tiger Woods versus Roger Federer is the most I think relevant analogy.

And so one of the things that I've been thinking about is in the world of the people I've worked with, how much-- it's like being able to look at something and say, OK, this is a problem, but I've seen this elsewhere when I was fishing or something like that, or whatever it is. It could be something in that way, that the broader the background, but the more innovation you can get because you're not looking at something with this tunnel vision, single view.

Mark Blaser: Absolutely. And I hope hiring committees are all built up with people like you in the future for me, Hardeep. So I'll take that. Absolutely. I totally agree.

I think the reality is for-- I'll opine for a bit. But I think science is becoming more and more interdisciplinary. We're tackling bigger and bigger problems.

The low-hanging fruit is more and more frequently gone. And it's only going to get harder moving forward. And these big problems are not simple. And they will require interdisciplinary skills and expertise.

Elena and I are working on other projects right now trying to get other things off the ground. We have, what, Elena, 10, 15 collaborators from across the world, all different backgrounds, from clinical to engineering, all to tackle these big questions that have evaded science for a long time. And I think it's the only way moving forward that people are going to be able to really start to make a dent in these big, intractable problems.

And so for me, I do think of myself as a generalist. You're absolutely right. I have some formal engineering training, some formal biomedical training, and a lot of biomedicine training that I've learned on the go.

And so I sit in this weird area where I know a bit about drug production, from my chemical engineering background. I have this biostatistics knowledge, but I can't call myself a biostatistician. I do omics here. I'm not a mass spectrometrist. I do some bioengineering, but I'm not a bioengineer.

But I have the ability to speak to these different groups in a reasonably educated way. I know enough about what they value and what they don't value. And I can act as a conduit between these different experts to make sure that everyone's heard, everyone's seen, everyone knows how to best contribute to move that project forward. And I think people like that, if I may toot my own horn a little bit, people like that are going to be important to pushing these multidisciplinary teams forward in the future to solve these sorts of problems.

Hardeep Ranu: Yeah. I think well said, Mark. But don't forget that it requires big vision of the PI.

[laughing]

Mark Blaser: Absolutely. Absolutely. That's putting that team together in the first place, right?

Elena Aikawa: I always try to build my team and hire people from different backgrounds. I never tried to have this team from the same background so they will tackle the same problem from the same vision, from the same view. So that's why Mark is a part of the team because of his bioengineering background.

And bioengineering is very, very important for the valve field because valve is not just tissue. It's tissue, which is moving. And tissue is moving in dynamic surroundings. So I think that requires very specific knowledge about many other things, not about how cells are working.

I think innovation also has been a very big part of my team. From the beginning, when I started-- when I received my first grant, I incorporated many modalities, which I previously don't use. I'm a pathologist by training, not even close to what Mark just described.

But I incorporated and learned molecular imaging, and for the first time, identified microcalcifications in the valve and other sclerotic issues, which people even didn't know about, because calcification was just calcification, like a big [? beak ?] of calcium. But actually, no one even thought how calcification would build.

And without innovation, we cannot move forward. Without innovation we cannot find a treatment for our patients. And without innovation, we cannot build very strong teams with different backgrounds. So that's my kind of vision here.

Hardeep Ranu: Yeah. No, it's great because also it's about getting that funding when you're trying to do something different, which is what scientists, I think, frequently come up against. It sort of, well, this is out of your field, so also we're not going to fund it. But I think hopefully that that will start to change, that it's innovation and creativity, you have to allow it to happen.

Elena Aikawa: And exactly. So this is very well said, Hardeep, because I don't know if you're aware of it, but let's say three or five years ago, NIH didn't want to find any high-risk grants and didn't want to find any high-throughput screening or any screening. They call it a "fishing expedition," right? And so no one paid attention to these kind of grants. No one actually even applied for those kinds of grants. But now I think the situation at NIH is changing little by little.

But what was beneficial with you guys, with Catalyst, that you took this high-risk project, very innovative one at every point, and allowed us to not only continue doing that, but also communicate with others within the Catalyst, like yourself, and the other leadership members, and PIs, and postdocs from those groups, which helped us to think forward and think next month, next two months, next half a year, what else we can do, what we can learn from other teams, and what we can incorporate in our project.

So there is no such a thing within NIH. So you get your four-year grant-- before it was five. Now it's just four. You get your four-year grant, and you never meet anyone. You kind of cooked in your own team, and you're producing your own ideas.

Of course, you read, and you're going to be meeting. But it's different. It's not like you meeting with the group of peers who is thinking about similar things, innovative ideas or something like that.

I had something similar only once within the NIH when we applied for RFA. And I think 10 investigators within heart valve disease got RFA. And just because we were so enthusiastic, we were able to meet every year as a team and talk about our discoveries and how to move forward.

But it was all about calcific aortic valve disease. It was about the same-- it was same team, same theme, and same people who I know already for the last two decades. But Catalyst brought it to a little bit different level. And I think that was very innovative.

Hardeep Ranu: Yeah. Yeah.

Elena Aikawa: That's great. It was so nice seeing you, and talking to you, and talk about our research.

Hardeep Ranu: Yeah, you too.

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Brendan Keegan: Thank you for listening. If you've enjoyed this podcast, please rate us on iTunes and help us spread the word about the amazing research taking place across the Harvard community.

Oby: To learn more about the guests on this episode, visit our website, catalyst.harvard.edu/thinkresearch.