Hematopoietic stem-progenitor cell (HSPC) transplantation is an established therapy for many malignant and non-malignant diseases. HSPCs are harvested clinically from 3 sources: G-CSF mobilized adult peripheral blood (PBSC), bone marrow (BM), and umbilical cord blood (UCB). Because erythrocytes increase both the risk of harmful side effects in transplant patients and the cost of cryopreservation, they must be depleted from the harvested HSPC tissues. The major problem in UCB transplantation is the low total number of HSPCs in the small volume (100-300 ml) of UCB units. This leads to high risk for delayed engraftment or engraftment failure (with attendant high mortality, morbidity and costs), especially in larger children or adult patients. Current techniques, including density gradient centrifugation and differential sedimentation, result in incomplete erythrocyte depletion and may lose 25% leukocytes (on average) during processing. Since success and speed of engraftment depend on the numbers of leukocytes and HSPCs per recipient body weight, it is essential to develop new cell separation methods to ensure high yields of pure, viable leukocytes and HSPCs from harvested UCB. The commercial goal of this STTR project is to improve stem cell banking and transplantation by marketing an efficient and robust processing system that results in superior recoveries of viable leukocytes and HSPCs, focusing on UCB as an immediate objective but involving a microfluidic cell separation technology that can be modified in the future to further purify both HSPCs and other stem cell types. We will take advantage of microfluidic deterministic lateral displacement (DLD), in which the paths cells take through the microfluidic system is based on size and is deterministic, i.e. absolutely determined, not subject to random processes. The key innovation of this STTR project is the use of DLD to deplete erythrocytes from UCB for hematopoietic transplant;this is a new clinical use, making this application commercially novel as well as clinically beneficial. The research strategy leverages a collaboration among micromechanical systems experts at GPB Scientific LLC;hematopoietic cell biology and cell processing experts at University of Maryland (Civin lab);and microfluidics experts at Princeton University (Sturm-Austin lab).
In Specific Aim 1, the team will optimize the efficacy of erythrocyte depletion and leukocyte recovery from UCB, solving cell clumping events which can block microfluidic flow and increasing microfluidic throughput to clinically relevant volumes.
In Specific Aim 2, the system will be modified for aseptic clinical us such that in the subsequent Phase II project, this high- volume microfluidic blood separation system can be used to assess HSPCs in vitro and in vivo to determine if their functionalities are conserved after erythrocyte depletion. The technology will also be extended for use with PBSC and BM harvests. The value proposition is clear: the GPB technology should deliver greater numbers of higher quality transplant grafts (i.e. more grafts with more HSPCs) that will lead to greater transplant success.
There is a critical unmet need for rapid, efficient methods to deplete erythrocytes and recover leukocytes from G-CSF mobilized peripheral blood (PBSC), bone marrow (BM), and especially umbilical cord blood (UCB), prior to cryopreservation. Incomplete erythrocyte removal from transplant grafts increases the risk of harmful side effects in hematopoietic stem cell transplants, while poor recovery of viable leukocytes and CD34+ cells reduces engraftment success and limits the treatable patient population. Development of a novel, highly efficient system to remove erythrocytes and purify leukocytes would raise the quality of UCB and other transplant grafts, thereby significantly improving patient outcomes.
|D'Silva, Joseph; Austin, Robert H; Sturm, James C (2015) Inhibition of clot formation in deterministic lateral displacement arrays for processing large volumes of blood for rare cell capture. Lab Chip 15:2240-7|