Antisense c-myc fragments induce normal differentiation cycles in HL-60 cells.

BACKGROUND

We have investigated the potential for using antisense technology as a means of delivering treatment for acute myeloblastic leukaemia (FAB-M2) by gene therapy.

MATERIALS AND METHODS

A test recombinant adenovirus vector was constructed containing human c-myc antisense fragments to study the effects of altering c-myc overexpression in the human HL-60 cell line. Control vector contained the human LacZ gene. Transfection efficiency in HL-60 cells was determined using control vector in the presence of protamine sulphate and multiplicity of infection of 100. Morphological and mechanistic changes were assessed using immunohistochemical analysis, flow cytometry and reverse transcription-polymerase chain reaction.

RESULTS

Transfection efficiency of control vector was 79.8% and morphological differences were observed after 72 h in culture. The rate of proliferation of HL-60 cells infected with test vector was inhibited by 73% compared with control following 6 days in culture. Normal terminal differentiation leading to apoptosis was only evident in test vector infected cells. Peak apoptosis (34.7%) was detected at day 6 and cell cycle arrest at days 2, 4 and 6. Expression of c-fos protein was significantly increased in test vector treated cells with a noticeable down-regulation of c-myc expression.

CONCLUSIONS

These data suggest that transfection of a human HL-60 cell line with vector containing c-myc antisense fragments could inhibit proliferation, but induce differentiation and apoptosis. Thus, we believe that further study of this construct is warranted as a potential gene therapy reagent for treatment of acute myeloblastic leukaemia.