Cationic porphyrins: novel delivery vehicles for antisense oligodeoxynucleotides

Abstract

Cationic porphyrins form stable complexes with oligodeoxynucleotides. To evaluate delivery, we used a 20mer phosphorothioate oligomer (Isis 3521) targeted to the 3'-untranslated region of the PKC-alpha mRNA, and complexed it with porphyrin. The expression of PKC-alpha protein and mRNA in T24 bladder carcinoma cells was reduced by approximately 80 +/- 10% at a concentration of oligomer of 3 microM, and 9 microM porphyrin. The expression of PKC-beta1, -delta and -straightepsilon isoforms was unaffected by this treatment, but elimination of PKC-zeta protein and mRNA were observed. However, treatment with the porphyrin complex of Isis 3522, an oligomer which is directed at the 5' coding region of the PKC-alpha mRNA, was equally effective as Isis 3521 with respect to PKC-alpha, but did not affect PKC-zeta protein or mRNA levels. Since Isis 3521 has an 11-base region of complementarity with the PKC-zeta mRNA, wheras Isis 3522 has only a 4-base region, the effect of Isis 3521 on PKC-zeta protein and mRNA expression may be due to irrelevant cleavage. Depending upon the desired application, this new strategy may offer several advantages over other methods of antisense oligodeoxynucleotide delivery including efficiency, stability, solubility, relatively low toxicity and serum compatibility. Porphyrins may thus be a potentially useful delivery vehicle for antisense therapeutics and/or target validation.

(-)-Epigallocatechin‑3‑gallate inhibition of Epstein‑Barr virus spontaneous lytic infection involves downregulation of latent membrane protein 1

“…The EBV LMP1-targeted DNAzyme DZ1 is an LMP1-targeted DNAzyme that binds and cleaves LMP1 RNA in a highly sequence-specific manner and was synthesized by Oligos Etc., Inc. (Wilsonville, OR, USA). Tetra (4-methylpyridyl) porphyrine was purchased from Frontier Scientific, Inc. (Logan, UT, USA) (31). Control oligonucleotides (Takara Biotechnology Co., Ltd., Dalian, China) were designed by inverting the catalytic core sequence, as described previously (31)(32)(33).…”

“…Tetra (4-methylpyridyl) porphyrine was purchased from Frontier Scientific, Inc. (Logan, UT, USA) (31). Control oligonucleotides (Takara Biotechnology Co., Ltd., Dalian, China) were designed by inverting the catalytic core sequence, as described previously (31)(32)(33). Mixtures were incubated for 15 min at room temperature to form transfection complexes.…”

Effect of EBV LMP1 targeted DNAzymes on cell proliferation and apoptosis

“…The DNAzymes/tetra(4-methylpyridyl) porphyrine (TMP) mixtures were made at a charge ratio of 1 with 2 mM DNAzyme oligonucleotides as described. 20 The mixture was then added to the cells and incubated at 371C for 4 hours, followed by the addition of complete medium to the wells and further incubation for 20 hours.…”

“…To facilitate delivery of DNAzyme oligonucleotides into cells, a cationic porphyrin, TMP, was used as a transfection reagent for intracellular delivery. 20 In order to increase DNAzyme stability in cells, two phosphorothioate linkages were incorporated into each of the arms in DNAzymes, which has been shown to be an efficient way to protect DNAzyme oligonucleotides from enzymatic degradation. 22 When the transfected cells were subjected to fluorescent microscopic analysis, efficient delivery was observed using the TMP-complexed phosphorothioate-Dz.…”

A Therapeutic Approach to Nasopharyngeal Carcinomas by DNAzymes Targeting EBV LMP-1 Gene

“…TMP, was used as a transfection reagent for intracellular delivery [29], and TMP was shown to be a simple and efficient transfection reagent due to its high complexing efficiency, low cellular toxicity and preferential nuclear uptake [27,30].…”