Soluble Bone Marrow Stroma Factors Improve the Efficiency of Retroviral Transfer of the Human Multidrug Resistance 1 Gene to Human Mobilized Peripheral Blood Progenitor Cells

Abstract

Hematopoietic stem cells (HSCs) are a potential target for the retrovirus-mediated transfer of chemotherapeutic drug resistance genes. For integration of the proviral DNA in the HSC genome cell division is required. In the bone marrow (BM) hematopoiesis occurs in the vicinity of stroma cells. Soluble stroma components were shown to play a permissive role for the proliferation of lineage-committed and primitive hematopoietic progenitors in conjunction with cytokines. We investigated the effect of stroma-conditioned medium (SCM) of the FBMD1 cell line on the gene transfer rate of the human multidrug resistance 1 (MDR1) gene contained in the retroviral SF-MDR vector into human mobilized peripheral blood progenitor cells (PBPCs) from tumor patients (n = 14) during transwell transduction in the presence of the recombinant fibronectin fragment CH-296. Addition of SCM during transduction increased the gene transfer efficiency into myeloid lineage-committed colony-forming cells by an average of 1.5-fold (p = 0.02) as detected by an SF-MDR provirus-specific polymerase chain reaction (PCR). These data were paralleled by significantly (p = 0.04 to p = 0.007) higher proportions of MDR1-expressing myelo-monocytic progeny after transduction in SCM plus interleukin 3 (IL-3), IL-3/Flt3 ligand (FL), IL-3/IL-6/FL, or IL-3/IL-6/stem cell factor (SCF) when compared with transductions without SCM as measured by rhodamine-123 exclusion. A similar trend was observed for SCM employed in combination with IL-3/IL-6/SC F/FL or FL/thrombopoietin (TPO)/SCF during transduction. The latter combination plus SCM yielded the highest proportion, 19.16 +/- 3.10% Rh-123dull cells. The beneficial effect of SCM on transduction efficiency was confirmed in additional four patients' samples, using a serum-free viral supernatant transduction protocol. As soluble BM stroma factors are able to increase the efficiency of retrovirus-mediated gene transfer into committed progenitor cells, beyond that achieved with fibronectin fragment CH-296, their effect on gene transfer into primitive repopulating hematopoietic cells may also prove beneficial.