HIGH-EFFICIENCY RETROVIRAL GENE-TRANSFER INTO MURINE HIGH-PROLIFERATIVE-POTENTIAL CELLS CYCLE-ACTIVATED BY CYTOSINE-ARABINOSIDE

We investigated cytosine arabinoside (Ara-C) as a potential agent for in vivo cycle activation of hematopoietic progenitors for the purpose of retroviral-mediated gene transfer. C(57)Bl mice were treated intrap eritoneally with one of three regimens of Ara-C: a single 1,750 mg/kg dose (regimen 1), a 1,750 mg/kg dose on day 0, and a 1,500 mg/kg dose on day 2 (LD(50)) (regimen 2), or a 1,750 mg/kg dose on day 0 and a 1, 500 mg/kg dose on day 3 (regimen 3). The high-proliferative-potential cells (HPPC)/10(5) cells were 47.0 +/- 7.5 pretreatment. The post-trea tment HPPC cloning efficiencies were 40.6 +/- 3.4, 83.6 +/- 6.1, and 2 0.4 +/- 3.2 HPPC/10(5) cells on days 1, 2, and 4, respectively, with r egimen 1; 60.0 +/- 7.9, 194.0 +/- 9.6, and 103.0 +/- 11.0 HPPC/10(5) c ells 1, 2, and 4 days after the second Ara-C dose, respectively, with regimen 2; and 266 +/- 13.4, 132 +/- 23.9, and 118.0 +/- 5.7/10(5) cel ls 1, 2, and 4 days after the second Ara-C dose, respectively, with re gimen 3. The transduction efficiency of HPPC from untreated animals wi th N2 viral supernatant was 4.9 +/- 5.8%. The post-treatment HPPC tran sduction efficiencies were 22.4 +/- 8.4%, 22.7 +/- 11.7%, and 0.4 +/- 0.6%, 1, 2, and 4 days after treatment with Ara-C, respectively, with regimen 1; 41.9 +/- 21.8%, 54.8 +/- 11.5%, and 0% 1, 2, and 4 days aft er the second Ara-C dose, respectively, with regimen 2; and 22.3 +/- 3 .8%, 33.3 +/- 12.4% and 0% 1, 2, and 4 days after a second Ara-C dose, respectively, with regimen 3. In vitro Ara-C susceptibility assays sh owed substantial cycle activation of HPPC 24-48 hr after the last Ara- C doses of the different regimens. These preliminary in vitro data sug gest that Ara-C administration may be useful for cycle activating earl y hematopoietic progenitors to increase the efficiency of retroviral t ransduction. These results set the stage for in vivo animal studies th at may lead to useful applications in future human gene therapy protoc ols.