Research Spotlight: Salvatore M. Caruso, PhD
Salvatore M. Caruso, PhD, the latest PhD graduate of the Tsang lab at Columbia Ophthalmology, has always had a passion for genomic engineering. He first learned about the ongoing gene therapy/gene editing revolution as an undergrad at Johns Hopkins studying Chemical and Biomolecular Engineering, where he joined a research lab focused on enhancing biomanufacturing processes for AAVs (adeno-associated viruses) and lentiviruses—two key viral vectors for gene delivery.
After working as a process engineer at Paragon Bioservices (now Catalent), helping clinical-stage companies develop gene therapies into transformative medicines, he decided to pursue his graduate degree in Biomedical Engineering under the mentorship of Dr. Stephen Tsang. Recently, Dr. Caruso, together with his collaborators inside and outside the Tsang lab, published “Ablating VHL in rod photoreceptors modulates RPE glycolysis and improves preclinical model of retinitis pigmentosa” in The Journal of Clinical Investigation. As co-first author, he shared with us details about his milestone research experience.
Tell us about yourself. How did you decide to pursue a doctorate in biomedical engineering?
After graduating from Johns Hopkins, I joined a manufacturing company in Baltimore called Paragon (now part of Catalent) as a process engineer, where most of our clients were clinical-stage companies looking to develop gene therapies into transformative medicines. I found that role particularly rewarding, especially when our clients would stop by and show us the impact of our work via patient testimonials and trial updates. At the time, Zolgensma, a gene therapy for spinal muscular atrophy, had just been approved, and our Baltimore site had entered into an agreement to help manufacture the new therapy. News reports of a toddler who had received the therapy and gained the ability to walk were starting to circulate, and that had everyone in the industry feeling proud of their work and excited about the potential of gene therapy. While I enjoyed the manufacturing side of things, I became more interested in how gene therapies were designed and the theoretical work that goes into conceptualizing a drug. After speaking with my group’s director and some of the other scientists on the team, the majority of whom had PhDs, it became clear that going back to school was the best route to develop those skills. So, I decided to apply with a focus on PIs known for genomic engineering.
What drew you to Dr. Tsang’s lab?
It was a mixture of personal intention and scientific coincidence. Due to the delivery constraints of gene editing, the costs of manufacturing, and the perhaps controversial immune privilege of the eye, the ophthalmology space has been a major forefront for gene therapy. Dr. Tsang has been a pioneer in gene and cell therapy within ophthalmology, and so I believe a mixture of his curiosity, willingness to explore the potential of gene editing, and its serendipitous success in the eye shaped my decision to join the lab. As for Dr. Tsang as a mentor, even with an ever-growing lab, he still somehow manages to keep an open-door policy and is there whenever you need him. His style of mentorship gives his students creative freedom and the ability to regulate how much guidance they may need, which was especially important to me when I was looking around at different programs. Ultimately, it was a mixture of the people and science that convinced me to join the team.
Tell us about the main findings of your paper "Ablating VHL in rod photoreceptors modulates RPE glycolysis and improves preclinical model of retinitis pigmentosa."
Several groups, including our team, have previously reported on the potential of metabolic reprogramming as a therapeutic avenue for treating retinitis pigmentosa (RP). Due to its high genetic heterogeneity and the rarity of individual variants, developing commercially viable therapies for RP has been historically challenging. However, many groups are starting to consider the problem somewhat differently and, instead of targeting the underlying genetic variant that may only treat <1% of all RP patients, are now targeting conserved mechanisms of degeneration that could be applied to multiple, if not all, RP genetic backgrounds.
In our work, we expand on the potential of targeting cell metabolism and the universal metabolic dysregulation that underpins RP across various genetic backgrounds. Likely due to the imbalances that occur during the initial degeneration of the retina, multiple teams have reported an accumulation of glucose in the RPE and a metabolic starvation of the distal photoreceptors—neuronal cells responsible for our vision. These cells rely on the RPE to shuttle their preferred food source (glucose) to them from the choroid’s blood supply. As a result, photoreceptors have been shown to shift their metabolism away from basal states to counteract the effects of “greedy” RPE cells. This can include photoreceptor cells becoming more ATP-efficient with their limited glucose supply, increasing oxidative phosphorylation and decreasing aerobic glycolysis, which is the more abundant process in healthy photoreceptors.
To counteract this process, our group demonstrated the potential therapeutic benefits of knocking out the VHL gene, a negative regulator of HIFs, which in turn stimulates aerobic glycolysis and dampens oxidative phosphorylation. Using functional and histological endpoints, we observed a delay in retinal degeneration within a mouse model of RP when the VHL gene was specifically knocked out in the photoreceptors. In collaboration with the Hurley lab at the University of Washington, we were able to perform a mixture of in vivo and ex vivo metabolic tracings that showed a positive correlation between ablating VHL, upregulating glycolysis in the photoreceptors, and therapeutic effects. Lastly, we were also able to show metabolic alterations in the RPE cells of treated mice despite no genetic manipulation of these cells, indicating a metabolically sensitive mechanism of communication from photoreceptors to the RPE.
Did this paper tie in with your dissertation work? If so, how? If not, tell us a little more about your dissertation work.
Yes, this paper directly tied into the first half of my thesis. Another aspect of the project we want to explore in the future is seeing if we can suppress glycolysis in the RPE, allowing for more glucose to reach the photoreceptors. To this end, we’ve proposed targeting HIF1A for ablation in the RPE, given the canonical roles of HIFs we observed in our studies.
While less universally applicable, the second half of my thesis explored the use of allele-specific CRISPR and prime editing systems to treat the most common forms of autosomal dominant RP associated with mutations in the rhodopsin gene. This approach looks to target a non-pathogenic SNP that is highly heterozygous so that the therapeutic remains viable regardless of where the mutation exists in the rhodopsin gene. While we’ve generated some great preliminary proof-of-concept data in a humanized mouse model of the disease, this work is still ongoing and will hopefully be published in the near future.
What roles did you and your co-authors each play in writing this paper?
My co-authors were absolutely instrumental in authoring this paper. Xuan Cui, my co-first author, was the real driving force behind much of the early work done here, including the creation of mouse models and exploring the therapeutic effects of knocking out VHL. Other co-authors from the Tsang lab also provided invaluable help in surgical procedures, molecular biology techniques, or critical insight into the interpretation of experimental results and future directions. Several members from the Hurley lab were instrumental in performing GC-MS, a complex but critical technique essential for measuring metabolic tracings and studying the impacts of our intervention on cell metabolism.
What were some of the unique challenges of this project?
Like all science, there were more than a handful of setbacks throughout the project. Developing the experimental mouse models with the exact genetic backgrounds we needed (i.e., the correct gene primed for knockout, exclusively expressing the knockout machinery in the target cells, and retaining the genes essential for disease modeling) was very labor-intensive. On top of that, we were dealing with resource constraints due to COVID and had to scale back our mouse colonies, which meant creating the same mouse lines often more than once. Managing the limited mice became challenging as well, given the breadth of experimental techniques used and the number of samples needed. It quickly made me realize that now more than ever we need to adequately fund science so that we don’t waste precious and valuable time on artificial bottlenecks that may inadvertently increase the long-term cost of research.
When doing the research, were there any results that were particularly surprising or a turning point for the project?
I think what really surprised us was seeing the metabolic alterations in the RPE, despite no genetic perturbation within those cells. There is growing evidence that the various cell types within the retina have distinct metabolic roles, and that these roles are woven together into metabolic interdependencies that give rise to the complex biological functions which allow us to see. It was very exciting to see firsthand that metabolic coupling dynamic at play and to watch how the rescue effects we observed in the photoreceptors were potentially dependent on changes in the RPE.
What are the next steps for you?
Looking back, I had a somewhat non-traditional career path to academia and never quite dreamed about being a professor (although I do often play with the idea of returning one day after accomplishing my personal goals). I loved my time in industry and am looking forward to returning there to leverage my understanding of early-stage drug development that I developed during my PhD. I was fortunate to be exposed to the investing side of the biotech industry during my time at Columbia, and so I would like to make my way back there. Ideally, I would one day be in a role where I can invest in or actively advise on the groundbreaking technologies that are coming out of universities daily and impacting the way healthcare is delivered.