title: Comparison of d-g3139 and its enantiomer L-g3139 in melanoma cells demonstrates minimal in vitro but dramatic in vivo chiral dependency.
abstract: G3139 (Genasense), an 18mer phosphorothioate antisense oligonucleotide targeted to the initiation codon region of the Bcl-2 messenger RNA (mRNA), downregulates Bcl-2 protein and mRNA expression in many cell lines. However, both the in vitro and in vivo mechanisms of action of G3139 are still uncertain. The isosequential L-deoxyribose enantiomer L-G3139, which does not downregulate Bcl-2 expression, was synthesized to study the role of the Bcl-2 protein in melanoma cells. Both D-G3139 and L-G3139 bind nonspecifically to basic fibroblast growth factor with approximately the same K(c), and cause highly effective inhibition of net formation in 518A2 melanoma cells on Matrigel. The uptakes of D-G3139 and L-G3139 in melanoma cells were also similar. However, unlike D-G3139, L-G3139 does not produce poly ADP-ribose polymerase-1 and procaspase-3 cleavage at 9.5 h after the initiation of the transfection, but can activate the intrinsic pathway of apoptosis at approximately 48 h. Furthermore, treatment of A375 melanoma human xenografts in severe combined immunodeficiency (SCID) mice demonstrates that tumor growth is not inhibited by L-G3139, whereas D-G3139 significantly inhibits the rate of tumor growth. Furthermore, the immunostimulatory properties of L-G3139 appear to be nil, which differs dramatically from those of D-G3139. In conclusion, profound differences exist between D-G3139 and L-G3139 in vivo despite their similarities in vitro.
authors.affiliation: Department of Biomedical Engineering, Columbia University, New York, New York, USA.
authors.forename: Johnathan C
authors.initials: JC
authors.lastname: Lai
authors.forename: Bob D
authors.initials: BD
authors.lastname: Brown
authors.forename: Anatoliy M
authors.initials: AM
authors.lastname: Voskresenskiy
authors.forename: Stefan
authors.initials: S
authors.lastname: Vonhoff
authors.forename: Sven
authors.initials: S
authors.lastname: Klussman
authors.forename: Wenzhi
authors.initials: W
authors.lastname: Tan
authors.forename: Marco
authors.initials: M
authors.lastname: Colombini
authors.forename: Risini
authors.initials: R
authors.lastname: Weeratna
authors.forename: Paul
authors.initials: P
authors.lastname: Miller
authors.forename: Luba
authors.initials: L
authors.lastname: Benimetskaya
authors.forename: Cy A
authors.initials: CA
authors.lastname: Stein
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D045304:Cytochromes c / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism; D005434:Flow Cytometry; D006801:Humans; D018664:Interleukin-12 / Q000378:metabolism; D019410:Interleukin-16 / Q000378:metabolism; D008546:Melanoma, Experimental / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D051379:Mice; D008807:Mice, Inbred BALB C; D016513:Mice, SCID; D008858:Microscopy, Phase-Contrast; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D016376:Oligonucleotides, Antisense / Q000378:metabolism / Q000493:pharmacokinetics / Q000494:pharmacology*; D011485:Protein Binding; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics*; D013154:Spleen / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D013237:Stereoisomerism; D023041:Xenograft Model Antitumor Assays
chemical_list: D019410:Interleukin-16; D016376:Oligonucleotides, Antisense; D019253:Proto-Oncogene Proteins c-bcl-2; D016222:Fibroblast Growth Factor 2; D018664:Interleukin-12; D045304:Cytochromes c