G3139 (oblimersen) may inhibit prostate cancer cell growth in a partially bis-CpG-dependent non-antisense manner

G3139 is an 18-mer phosphorothioate oligodeoxyribonucleotide, which is targeted to the initiation codon region of the bcl-2 mRNA. Although treatment of PC3 prostate cancer cells with G3139, which contains two CpG motifs, causes a dramatic decrease in bcl-2 protein expression after 3 days, it did not result in significant cellular apoptosis, as it does in many other cell lines. The absence of apoptosis was demonstrated by the absence of procaspase 3 cleavage products and of Annexin V cell surface expression. In addition, ATP production and the mitochondrial membrane potential Δψ_m were preserved. Despite this, G3139 significantly inhibited the rate of cellular proliferation in complete media and blocked cloning in soft agar. G4232, a variant of G3139 that down-regulates bcl-2 expression to the same extent but has both CpG cytidines C5 methylated, was only minimally antiproliferative. A series of mismatched G3139-related oligomers were synthesized that could also substantially downregulate bcl-2 protein expression, but only if the CpG motifs were preserved, demonstrating the presence of additional non-antisense mechanisms. G3139 caused production of reactive oxygen species in growth-arrested cells and oxidation of nuclear guanosine to 8-hydroxy-2′- deoxyguanosine, as determined by 1F7 monoclonal antibody staining. Bromodeoxyuridine incorporation studies demonstrated that G3139 induced a G ₁-S entry block and an intra-S-phase block in PC3 cells that persisted as long as 3 days. This finding coincides with the observation that expression of several proteins encoded by S-phase genes, including c-myb and poly(ADP-ribose) polymerase, were significantly reduced. These results illustrate the complexity of the mechanism of action of G3139 in PC3 cells.