Ex Vivo Expansion of SCID Leukemia-Initiating Cells Using NANEX Nanofiber Scaffold

Abstract 4994

The concept of the leukemic stem cell (LSC) has gained wide acceptance since it was first definitively established using a NOD-SCID xenotransplantation model nearly 15 years ago. LSCs, which are functionally defined as SCID leukemia-initiating cells (SL-ICs), are believed to possess biological properties that render them resistant to conventional chemotherapy. Although there is still much debate over how to phenotypically define LSCs, there is general agreement that LSCs are rare in acute myeloid leukemia (AML), which has hindered efforts to develop LSC-targeted therapies. In order to provide researchers and pharmaceutical companies with an ample supply of LSCs for testing, methods are needed to generate large numbers of LSCs from patient samples.

Ex vivo expansion of LSCs in culture is one approach that offers tremendous promise for increasing cell numbers for research and drug development. Conditions that enable efficient expansion of normal hematopoietic stem cells (HSCs) can be used as a starting point for developing an optimal culture system for LSCs. The natural bone marrow microenvironment maintains HSCs in close contact with a complex network of stromal cells and extracellular matrix, likely indicating that cell-cell and cell-matrix interactions play an important role in maintaining their stem cell phenotype. With the goal of mimicking the bone marrow stem cell niche, Arteriocyte, Inc. has developed a 3-D NANEX nanofiber based cell culture substrate. The NANEX substrate is designed to provide topographical and substrate-immobilized biochemical cues that act in synergy with media additives to enhance HSC proliferation.

Here, we present our recent work with the NANEX platform towards achieving a high yield ex vivo expansion of LSCs. Using common LSC markers, including CD34, CD38, CD117, and CD123, we quantify and characterize NANEX-expanded leukemic cells using flow cytometry. We compare NANEX to standard tissue culture polystyrene and demonstrate that NANEXÔ significantly improves LSC expansion and reduces clonogenic phenotype loss during ex vivo culture. Additionally, we show that NANEXÔ-expanded cells engraft in NOD-SCID mice and, through limiting dilution analysis, quantify the increase in SL-ICs as a result of culture on NANEXÔ. Our data indicates that NANEX technology provides a robust ex vivo expansion of SL-ICs and, with further development, offers great potential for use in LSC-targeted drug development.