TRANSDUCTION OF HUMAN INTERLEUKIN-9 RECEPTOR GENE INTO HUMAN CORD-BLOOD ERYTHROID PROGENITORS INCREASES THE NUMBER OF ERYTHROPOIETIN-DEPENDENT ERYTHROID COLONIES

We constructed a retroviral vector, pLhIL-9RSN, containing cDNA encodi ng the human interleukin-9 receptor (IL-9R) along with a neomycin phos photransferase gene (Neo), In order to study the biological effects of the IL-9R, high titer (1-5 x 10(5) CFU/ml) viral supernatant, generat ed from the packaging cell lines, ecotropic GPE86 and amphotropic PA31 7, was used to transduce the IL-9R gene into sorted populations of CD3 4(++)CD33(-) cells from human cord blood which are highly enriched for erythroid progenitor cells (BFU-E), Colony formation by BFU-E transdu ced with the IL-9R gene and grown without selection in G418 and in the presence of erythropoietin (Epo) and interleukin (IL)-9 was significa ntly increased up to three-fold and the size of the erythroid colonies was significantly increased 50-100% compared to colony formation by m ock virus transduced cells, Moreover, colony formation by IL-9R-transd uced cells was more sensitive to stimulation with lower doses of IL-9 and Epo, Individual colonies formed with or without selection in G418 were evaluated, Proviral integration and mRNA expression were respecti vely assessed by polymerase chain reaction (PCR) and reverse transcrip tase (RT) PCR analysis and were apparent in 93% and 84% of the G418-re sistant colonies and 52% and 48% of the colonies grown in the absence of G418, Our study demonstrates that a functional human IL-9R gene can be efficiently transduced into human cord blood hematopoietic progeni tors using retroviral vectors with increased cytokine-dependent erythr oid colony formation.