Today, we feature Dr. Alessandro Penna from
the University of Padova, whose research focuses on advancing adoptive cell
therapies for prostate cancer.
When standard gene-editing protocols failed
to produce a critical cell line needed to validate his studies, Dr. Penna
turned to Cytosurge’s CellEDIT service. Leveraging our proprietary FluidFM
technology, we successfully delivered the complex knockout required to unblock
his research and allow his team to move forward.
Alessandro Penna, Ph.D. Postdoctoral Researcher. Hosted by the Department of Surgery, Oncology and Gastroenterology (DiSCOG) University of Padova, Italy.
Dr. Alessandro Penna is a Postdoctoral
Researcher in Prof. Antonio Rosato’s group at the University of Padova. Holding
a Ph.D. in Experimental and Clinical Oncology, he specializes in cancer
immunotherapy. His current work focuses on engineering CAR-T strategies to
overcome immunosuppressive microenvironments in solid tumors like prostate
cancer.
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Tell us about your research project.
Our research focuses on the development of cellular therapies for solid tumors, with a specific emphasis on prostate cancer. We utilize PC-3 cell lines as one of our primary models.
Our current project investigates the influence of immunotolerant stimuli, specifically the PD-L1/PD-1 axis. We hypothesized that disrupting this axis could positively influence the therapeutic outcome of our cell therapy. To test this, we needed to knock out the endogenous CD274 (PD-L1) gene in PC-3 cells to see if removing this checkpoint ligand would improve the efficacy of our treatment targeting the PSMA antigen.
What specific technical hurdles did you encounter before contacting CellEDIT?
Our primary obstacle was technical: we simply couldn't get the knockout to work in-house. We spent considerable time troubleshooting standard CRISPR/Cas9 protocols using lipofectamine transfection, testing various cell concentrations and conditions without success.
While our lab is well-versed in genetic engineering, specifically using lentiviral vectors for gene delivery, this specific knockout proved elusive. We found ourselves stuck in a cycle of protocol optimization, losing valuable time that should have been spent on the actual cell therapy research.
How did you learn about CellEDIT, and why did you choose our team to handle this challenge?
After several failed attempts, a colleague identified Cytosurge as a company specializing in difficult cell editing. We realized that to proceed with our actual research, the adoptive cell therapy analysis, we needed to stop troubleshooting the editing process and outsource it to experts.
We chose CellEDIT Service because the Cytosurge team was able to propose a robust strategy. Specifically, the double-guide CRISPR approach (targeting Exon 3 and Exon 4) gave us confidence that we would achieve a complete disruption of the protein, which is essential for our validation experiments.
How was the collaboration with the team throughout the project?
I was very happy with the overall experience. The approach was clear from the beginning, and the support from the team, particularly Dr. Gorka Santos, was excellent. There was constant intercommunication regarding the project's status.
We did face some logistical delays regarding the shipment of cells, but I want to clarify that this was due to customs and external administrative factors, not Cytosurge. Throughout that process, the Cytosurge team was proactive, constantly checking in and providing clear updates. It was a relief to have a partner that was responsive and transparent.
How important was the technical outcome for your research?
It was critical. The report confirmed that we received a homozygous knockout with a complete deletion between Exon 3 and Exon 4. This was exactly what we needed, a cell population that is completely devoid of PDL-1. Without this, I couldn't perform my experiments because any residual PD-L1 expression could compromise the data regarding the therapeutic outcome.
We also received a compound heterozygous clone, which, while not my primary target, offers interesting possibilities for future validation experiments.
Would you recommend Cytosurge to other researchers? And would you work with CellEDIT again?
Absolutely. Having this cell line has accelerated our studies significantly. If we had continued trying to do this in-house, we would have lost much more time.
I will certainly share this positive experience with my colleagues in the lab. As we continue our work on cell therapies, I would definitely consider using Cytosurge again for any future complex editing needs that arise.
Dr. Penna’s experience underscores the value of specialized engineering for difficult-to-edit cancer cell lines. We are proud to support the University of Padova in its quest to improve cellular therapies for solid tumors.
Accelerating Adoptive Cell Therapy Development
How CellEDIT Solved a Critical Knockout Challenge for the University of Padova