Nucleotide and oligonucleotide derivatives as enzyme and nucleic acid targeted irreversible inhibitors, biochemical aspects

Abstract

Sequence specific modification of nucleic acids with reactive oligonucleotide derivatives, complementary addressed modification, can provide an efficient approach for specific inactivation of certain cellular nucleic acids. In experiments with ascites tumor Krebs II cells and alkylating oligothymidylate derivatives it was found that alkylating oligonucleotide derivatives enter the living cell and modify complementary sequences in cellular nucleic acids with high efficiency. Complementary addressed modification of poly(A) sequences in cellular RNA with oligothymidylate derivatives was investigated in detail. The results of experiments on alkylation of cellular nucleic acids are consistent with complementary addressed modification of poly(A) sequences in cellular DNA. These results are supported by experiments on modification of chromatin DNA in which it was found that chromatin DNA interacts with oliogothymidylate derivatives more readily than the isolated double stranded DNA. It was found that alkylating oligonucleotide derivatives complementary to a sequence in immunoglobulin mRNA of MOPC 21 cells arrest the cellular immunoglobulin synthesis. Alkylating oligonucleotide derivatives complementary to RNAs of fowl plague virus inhibit virus multiplication in cell culture.

Oligonucleotide N-alkylphosphoramidates: synthesis and binding to polynucleotides

“…In recent years, numerous reports have described the synthesis of oligonucleotides modified to contain adjuvants useful in DNA binding (Letsinger & Schott, 1981;Asseline et al, 1984;Héléne et al, 1985;Thuong et al, 1987;Letsinger et al, 1988), site-selective alkylation (Vlassov et al, 1985;; Zarytova et al, 1986;Iverson & Dervan, 1987), or cleavage (Boutorin et al, 1984;Chu & Orgel, 1985;Dreyer & Dervan, 1985; Le Doan et al, 1986;Boidot-Forget et al, 1986. It seemed likely that the chemistry described here should be applicable for the synthesis of such modified oligonucleotides, and we attempted to prepare a representative dinucleoside phosphoramidate (26).…”

Mechanism of oligonucleotide uptake by cells: involvement of specific receptors?

“…They have been considered as a potential new generation of drugs, capable perhaps of inhibiting various pathogens and of regulating specific gene expression by inhibiting the translation of mRNA molecules in a highly specific manner (1)(2)(3)(4). However, a prevailing view is that cells are not very permeable to oligonucleotides.…”

“…We also investigated the stability of the internalized oligomers. Experiments involving the reactive 4-[N-(2-chloroethyl)-N-methyl]aminobenzyl phosphamide derivative of oligodeoxynucleotides [general formula ClRCH2NHpTTFr..., where R = -CH2CH2N(CH3)C6Hr---(1)]t were also undertaken: reagents of this type have recently been used by us as inhibitors of the influenza and tick-borne encephalitis viruses (4,15). In the present study, these reagents were used to chemically modify the putative cellular receptors binding oligonucleotides.…”

Reactive oligonucleotide derivatives as gene-targeted biologically active compounds and affinity probes

“…In experiments with influenza virus (Vlassov et al, 1986a), alkylating derivatives of oligonucleotides targeted to the universal 5'-terminal sequence of the genomic viral RNAs (RpCCTTGTTTCT, I00/~M) decreased the virus titer in the infected chicken fibroblasts by 4.5 logarithm units as compared to the control. Similar enhancement of antiviral activity due to this derivation was observed in experiments with HIV-1 and a Tick-borne encephalitis virus (Pogodina et al, 1988;Abramova et al, 1990).…”