Abstract
Therapeutic treatment options for hemoglobinopathies as well as many other monogenic diseases, are typically non-curative and have significant limitations. Autologous gene edited Hematopoietic Stem Cell (HSC) transplant has demonstrated potentially curative effects in recent clinical trials but is limited by low engraftment efficiencies and significant toxicities and malignancies associated with the required myeloablative preconditioning agents. Our overall objective is to establish a safer and more effective autologous transplant method using mRNA enhanced HSCs and a modified conditioning regimen that is non-genotoxic. To achieve this goal, we use the Cell Squeeze® technology to engineer HSCs ex vivo with mRNA to transiently outcompete the endogenous diseased HSCs.
To create HSCs with enhanced engraftment, we delivered mRNA constructs encoding homing receptors such as CXCR4 or survival factors such as BCL2. Furthermore, to provide a competitive advantage against endogenous HSCs, we tested mRNA sequences with selected mutations to create cells resistant to conditioning agents such as Plerixafor, a CXCR4 antagonist, and Venetoclax, a BCL2 inhibitor. By using mRNA-based engineering, after a few days the modified receptors and factors were no longer present, and the gene-edited HSC could be maintained naturally in the now occupied niche.
Our data further demonstrated that the Cell Squeeze® technology is gentle on HSCs compared to alternative engineering approaches. HSCs engineered using Cell Squeeze® technology were functional immediately after engineering as shown by changes in gene expression, colony forming assays, and an 8-week NBSGW mouse engraftment study. Additionally, we showed high levels of gene editing (>75%) and mRNA expression (>85%) in squeezed CD34+ cells.
The Cell Squeeze® technology is scalable, has been implemented in multiple clinical programs, and reliably engineers billions of cells per minute. The generation of HSCs with enhanced engraftment in a clinically translatable manner has the potential to dramatically improve the treatment paradigm of numerous diseases, such as hemoglobinopathies, that can be treated with HSC therapies.
Disclosures
Gilbert:SQZ Biotechnologies: Current Employment, Current holder of stock options in a privately-held company.
Author notes
Asterisk with author names denotes non-ASH members.
