UC San Diego

Cell-based therapies hold tremendous potential to treat cancer, inflammatory conditions, and degenerative diseases. However, despite a few notable successes, cell-based therapies face key clinical barriers, including cell source heterogeneity, inefficient directed migration (homing) to target tissues, cell dosing or controllability, inability to affect tissue microenvironments, and patient safety. Bioengineering may potentially enhance cell capabilities, but raises safety concerns regarding the control of genetically engineered cells in vivo. Therefore, bioengineering methods

to improve cell-based therapy efficacy must prioritize predictable behavior and safety.

To address these issues, the Klemke Laboratory developed a platform technology that couples cell biongineering with cell enucleation (mass removal of nuclei) to enhance cellular function and controllability. By adapting methods developed in the 1970s, mesenchymal stromal cells (MSCs) were enucleated by ultracentrifugation in discontinuous Ficoll gradients containing cytochalasin B. Combining cell bioengineering with enucleation both decreases the risk of engraftment and oncogenesis, and provides opportunities for customized pre- and/or post-enucleation bioengineering. We named these bioengineered, enucleated cells “CargocytesTM”.

Cargocytes were characterized by absent nuclei, viability for 2-3 days, functional protein translational machinery, retention of receptor expression, and directional migration, and then further investigated via bioengineering with therapeutically-relevant properties. For in vitro studies, Cargocytes were bioengineered and then examined for secretion of functional cytokines and expression of functional chemokine receptors. Next, Cargocytes in vivo abilities were examined in two independent mouse models. First, bioengineered Cargocytes injected intratumorally in a mouse breast cancer model produced functional antitumor cytokine that slowed tumor progression and increased animal survival. Second, bioengineered Cargocytes injected intravenously in a mouse ear inflammation model both homed to the site of inflammation and produced functional anti-inflammatory cytokine that reduced inflammation. This suggests that bioengineered, enucleated cells are more efficacious than non-engineered cells, and lacking nuclei uniquely contributes to safety and controllability in vivo.

To my knowledge, therapeutic bioengineering of enucleated cells has not been previously described, and Cargocytes represent a unique entity in the cell-based therapy field as either a stand-alone or adjuvant therapy. By retaining biologically important cell-like functions yet with advantages like small size and defined lifespan, Cargocytes potentially resolve key barriers currently limiting cell-based therapies.