Hematopoietic stem cells (HSCs) reside in the bone marrow where they throughout life sustain continuous blood production through a controlled balance of differentiation and self-renewal. Transplantation of HSCs from a healthy person can replace a defective hematopoietic system of a patient thereby curing the patient for life. HSCs have found increasing therapeutic application, e.g. in hematologic malignancies and hematopoietic genetic disorders. This applies not only to the allogeneic transplantation setting but also to the autologous setting where advances in genetic engineering technologies have enabled autologous gene therapies. However, major challenges remain in both settings pertaining to the scarcity of HSCs, as well as the cells being partially refractory to precise gene correction.
In this research proposal, I will address these challenges by leveraging the unique power of repurposed CRISPR/Cas systems for precise transcriptional manipulation of HSCs. In these systems, the normal DNA-cleaving ability of the Cas9 enzyme is disabled (dCas9) while transcriptional activators or inhibitors are fused to dCas9. By targeting the dCas9-effector proteins to transcriptional start site regions by sgRNA programming, gene transcription can be activated (CRISPRa) or inhibited (CRISPRi). Complex transcriptional engineering is readily achieved using multiple sgRNAs and orthogonal CRISPR systems for simultaneous CRISPRa and CRISPRi.
I will apply these technologies to investigate and enhance therapeutically relevant HSC pathways, namely homologous recombination for precise gene editing, self-renewal, and bone marrow homing. These biological phenomena have previously been studied with techniques that do not have the same elegant properties and therapeutic relevance as CRISPRa/i. With this new state-of-the-art method for precisely controlling gene expression, I will study and manipulate genetic pathways to overcome long-standing challenges in HSC therapies.
Fields of science
- HORIZON.1.1 - European Research Council (ERC) Main Programme