Hector Caballero-González | 16 March 2026
- Title: Microfluidics-based single-cell characterization of innovative cellular immunotherapies
- Supervisors: Dr. Laura Rocchi and co-supervisor: Prof. Pier Luigi Zinzani
- Institution: University of Bologna (PhD carried out at Cellply)
- Asbtract: Chimeric Antigen Receptor (CAR)-T cells have fundamentally transformed the landscape of cancer treatment, demonstrating remarkable clinical efficacy in hematological malignancies. Despite achieving unprecedented clinical success, major challenges hamper their broader clinical implementation, including complex mechanisms of action, characterization methodologies, and manufacturing scalability.
CAR-T cell products exhibit substantial phenotypic and functional heterogeneity, which current approaches struggle to capture at the single-cell level. As this heterogeneity critically impacts therapeutic efficacy, there is a pressing need for novel technologies capable of accurately profiling these complex functional dynamics over time. To tackle this challenge, this thesis leverages a novel microfluidic analytical platform combining live-cell fluorescence imaging with an AI-driven analytical pipeline for the simultaneous profiling of thousands of miniaturized tumor-immune co-cultures by integrating cytotoxicity, phenotypic, and kinetic readouts at single-cell resolution.
To mitigate the risk of on-target off-tumor toxicity, adapter CAR-T platforms such as the UniCAR-T system have emerged as a powerful alternative for enhanced controllability and targeting flexibility. However, their single-cell dynamics and serial killing capabilities remain unexplored. This work investigates the single-cell functional landscape of UniCAR-T cells targeting CD33+ acute myeloid leukemia, uncovering distinct functional subsets coexisting within the same UniCAR-T cell population, including a highly potent subset of serial killers. These findings underscore a degree of single-cell functional heterogeneity not detectable by conventional bulk assays. In addition, the present work advances the development of a novel microfluidics-based cytokine release assay with the potential to integrate multifunctional readouts on cytotoxicity, phenotype, and cytokine secretion on individual immune cells.
Overall, the present study offers a novel data-driven framework for the single-cell profiling of immune cell therapies while providing an in-depth characterization of adapter UniCAR-T cells, with the ultimate goal of supporting the design and development of next-generation immune cell therapies.
Simona Mattiussi | 13 March 2026
- Title: Microfluidics-based single-cell characterization of innovative cellular immunotherapies
- Supervisors: Dr. Laura Rocchi and co-supervisor: Prof. Pier Luigi Zinzani
- Institution: University of Bologna (PhD carried out at Cellply)
- Abstract: Abstract: Cancer remains a major global health challenge, with rising incidence and mortality rates worldwide. Despite advances in oncology, early detection, accurate treatment monitoring, and effective therapy for solid tumors remain limited. Theranostics, combining molecular imaging with targeted radionuclide therapy, has emerged as a precision medicine approach to improve patient selection and reduce systemic toxicity. Tumor heterogeneity and the supportive tumor microenvironment, particularly cancer-associated fibroblasts, contribute significantly to therapeutic resistance. Fibroblast activation protein (FAP), highly expressed in these stromal cells and minimally in normal tissues, represents a promising universal target. While FAP inhibitors have shown strong performance in PET imaging, their therapeutic translation is limited by suboptimal tumor retention and pharmacokinetics. Meanwhile, immunotherapies such as CAR T-cell therapy face challenges in solid tumors, prompting the development of controllable platforms like Universal CAR T cells and combination strategies with radionuclide therapy.
This Ph.D. thesis focuses on developing and optimizing novel small-molecule FAP-targeting compounds for theranostic applications. The work improves tumor uptake and retention, explores pretargeting strategies integrating nuclear and near-infrared imaging, and develops a FAP-based module for combined radionuclide delivery and Universal CAR T-cell therapy. Overall, the project advances the field of FAP-targeted theranostics.