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Yoon S Chochung - One of the best experts on this subject based on the ideXlab platform.
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an antisense Oligodeoxynucleotide that depletes riα subunit of cyclic amp dependent protein kinase induces growth inhibition in human cancer cells
Cancer Research, 1993Co-Authors: Hiroshi Yokozaki, Giampaolo Tortora, Alfredo Budillon, Scott Meissner, Serge Beaucage, Keizaburo Miki, Yoon S ChochungAbstract:Abstract Enhanced expression of the RI α subunit of cyclic AMP-dependent protein kinase type I has been correlated with cancer cell growth. We provide evidence that RI α is a growth-inducing protein that may be essential for neoplastic cell growth. Human colon, breast, and gastric carcinoma and neuroblastoma cell lines exposed to a 21-mer human RI α antisense phosphorothioate Oligodeoxynucleotide (S-Oligodeoxynucleotide) exhibited growth inhibition with no sign of cytotoxicity. Mismatched sequence (random) S-Oligodeoxynucleotides of the same length exhibited no effect. The growth inhibitory effect of RI α antisense oligomer correlated with a decrease in the RI α mRNA and protein levels and with an increase in RII β (the regulatory subunit of protein kinase type II) expression. The growth inhibition was abolished, however, when cells were exposed simultaneously to both RI α and RII β antisense S-Oligodeoxynucleotides. The RII β antisense S-Oligodeoxynucleotide alone, exhibiting suppression of RII β along with enhancement of RI α expression, led to slight stimulation of cell growth. These results demonstrate that two isoforms of cyclic AMP receptor proteins, RI α and RII β , are reciprocally related in the growth control of cancer cells and that the RI α antisense Oligodeoxynucleotide, which efficiently depletes the growth stimulatory RI α , is a powerful biological tool toward suppression of malignancy.
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differentiation of hl 60 leukemia by type i regulatory subunit antisense Oligodeoxynucleotide of camp dependent protein kinase
Proceedings of the National Academy of Sciences of the United States of America, 1991Co-Authors: Giampaolo Tortora, Stefano Pepe, Timothy Clair, Hiroshi Yokozaki, Yoon S ChochungAbstract:A marked decrease in the type I cAMP-dependent protein kinase regulatory subunit (RI alpha) and an increase in the type II protein kinase regulatory subunit (RII beta) correlate with growth inhibition and differentiation induced in a variety of types of human cancer cells, in vitro and in vivo, by site-selective cAMP analogs. To directly determine whether RI alpha is a growth-inducing protein essential for neoplastic cell growth, human HL-60 promyelocytic leukemia cells were exposed to 21-mer RI alpha antisense Oligodeoxynucleotide, and the effects on cell replication and differentiation were examined. The RI alpha antisense oligomer brought about growth inhibition and monocytic differentiation, bypassing the effects of an exogenous cAMP analog. These effects of RI alpha antisense Oligodeoxynucleotide correlated with a decrease in RI alpha receptor and an increase in RII beta receptor level. The growth inhibition and differentiation were abolished, however, when these cells were exposed simultaneously to both RI alpha and RII beta antisense Oligodeoxynucleotides. The RII beta antisense Oligodeoxynucleotide alone has been previously shown to specifically block the differentiation inducible by cAMP analogs. These results provide direct evidence that RI alpha cAMP receptor plays a critical role in neoplastic cell growth and that cAMP receptor isoforms display specific roles in cAMP regulation of cell growth and differentiation.
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an antisense Oligodeoxynucleotide targeted against the type ii beta regulatory subunit mrna of protein kinase inhibits camp induced differentiation in hl 60 leukemia cells without affecting phorbol ester effects
Proceedings of the National Academy of Sciences of the United States of America, 1990Co-Authors: Giampaolo Tortora, Timothy Clair, Yoon S ChochungAbstract:The type II beta regulatory subunit of cAMP-dependent protein kinase (RII beta) has been hypothesized to play an important role in the growth inhibition and differentiation induced by site-selective cAMP analogs in human cancer cells, but direct proof of this function has been lacking. To address this issue, HL-60 human promyelocytic leukemia cells were exposed to RII beta antisense synthetic Oligodeoxynucleotide, and the effects on cAMP-induced growth regulation were examined. Exposure of these cells to RII beta antisense Oligodeoxynucleotide resulted in a decrease in cAMP analog-induced growth inhibition and differentiation without apparent effect on differentiation induced by phorbol esters. This loss in cAMP growth regulatory function correlated with a decrease in basal and induced levels of RII beta protein. Exposure to RII beta sense, RI alpha and RII alpha antisense, or irrelevant Oligodeoxynucleotides had no such effect. These results show that the RII beta regulatory subunit of protein kinase plays a critical role in the cAMP-induced growth regulation of HL-60 leukemia cells.
Jack S. Cohen - One of the best experts on this subject based on the ideXlab platform.
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Oligonucleotides as therapeutic agents.
Pharmacology & Therapeutics, 1991Co-Authors: Jack S. CohenAbstract:Abstract Oligodeoxynucleotides can act as antisense complements to target sequences of mRNAs to selectively regulate gene expression. Chemically modified analogs that are nuclease-resistant enable this antisense strategy to be utilized in practice. Studies with Oligodeoxynucleotide analogs in cell free systems, and their cellular uptake will be described. Certain analogs have been found to regulate viral and cellular gene expression. However, some also inhibit in a non-specific manner, that may be traced to their selective inhibition of viral and cellular polymerases. A chemically modified Oligodeoxynucleotide analog can be regarded as an informational drug.
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Cellular uptake of antisense Oligodeoxynucleotides
Advanced Drug Delivery Reviews, 1991Co-Authors: Jerzy W. Jaroszewski, Jack S. CohenAbstract:Abstract Synthetic, chemically modified antisense Oligodeoxynucleotides inhibit gene expression in a sequence-specific manner, besides having other, sequence-non-specific effects on cells. In this context, cellular uptake characteristics of these compounds is of particular interest. Radioactive or fluorescence labelling is used to investigate kinetics and intracellular distribution. Oligodeoxynucleotides apparantly enter cells by two mechanisms, an active and a passive; there is evidence for the presence of Oligodeoxynucleotide receptors on the surface of cells. Parameters determining cellular uptake are: type of cells and medium from which the uptake takes place, type of Oligodeoxynucleotide analog, chain length, and presence of linked groups. In cells, the Oligodeoxynucleotides are present mainly in the cytoplasm and in/around the nucleus; the exact distribution pattern appears to vary strongly according to the cellular system and Oligodeoxynucleotide analog used. Chemically modified Oligodeoxynucleotide analogs remain intact inside the cells for days. Future efforts to increase efficacy of antisense Oligodeoxynucleotides should be directed towards design of linked groups increasing and targeting cellular uptake.
Sudhir Agrawal - One of the best experts on this subject based on the ideXlab platform.
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Antisense depletion of RIα subunit of protein kinase A induces apoptosis and growth arrest in human breast cancer cells
Breast Cancer Research and Treatment, 1998Co-Authors: Rakesh K. Srivastava, Aparna R. Srivastava, Yun Gyu Park, Sudhir Agrawal, Yoon S. Cho-chungAbstract:In recent years, several laboratories have explored the possibility of using antisense Oligodeoxynucleotides for specific manipulation of gene expression leading to cancer treatment. The enhanced expression of the RIα subunit of cyclic AMP-dependent protein kinase type I (PKA-I) has been correlated with cancer cell growth. In the present study, the effects of an antisense Oligodeoxynucleotide targeted against RIα subunit of PKA-I on growth inhibition and apoptosis in MDA-MB-231 human breast cancer cells were investigated. The growth inhibitory effects of RIα antisense Oligodeoxynucleotide correlated with a decrease in the RIα mRNA and protein levels. The growth inhibition was accompanied by changes in the cell cycle phase distribution, cell morpbology, cleavage of poly (ADP-ribose) polymerase (PARP), and appearance of apoptotic nuclei. By comparison, mismatched control Oligodeoxynucleotide had no effect. On the basis of these results, it can be suggested that the RIα antisense Oligodeoxynucleotide, which efficiently depletes the growth stimulatory RIα and induces apoptosis/differentiation, could be used as a therapeutic agent for breast cancer treatment.
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Mixed-backbone oligonucleotides as second generation antisense oligonucleotides: In vitro and in vivo studies
Proceedings of the National Academy of Sciences of the United States of America, 1997Co-Authors: Sudhir Agrawal, Zhiwei Jiang, Qiuyan Zhao, Denise Shaw, Allysen Roskey, Lakshmi Channavajjala, Carl Saxinger, Ruiwen ZhangAbstract:Antisense oligonucleotides are being evaluated in clinical trials as novel therapeutic agents. To further improve the properties of antisense oligonucleotides, we have designed mixed-backbone oligonucleotides (MBOs) that contain phosphorothioate segments at the 3′ and 5′ ends and have a modified Oligodeoxynucleotide or oligoribonucleotide segment located in the central portion of the oligonucleotide. Some of these MBOs indicate improved properties compared with phosphorothioate Oligodeoxynucleotides with respect to affinity to RNA, RNase H activation, and anti-HIV activity. In addition, more acceptable pharmacological, in vivo degradation and pharmacokinetic profiles were obtained with these MBOs.
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Inhibition of in vitro transcription by Oligodeoxynucleotides.
Antisense research and development, 1994Co-Authors: Jamal Temsamani, Sudhir Agrawal, Valeri Metelev, Asya Levina, Paul C ZamecnikAbstract:Single-stranded short phosphodiester Oligodeoxynucleotides have been used to inhibit in vitro T7 transcription system. These Oligodeoxynucleotides were complementary to either the 3′-5′ or 5′-3′ strand of the transcription initiation site of a plasmid containing the gag region of HIV. Our results show that incubation of this plasmid DNA with the Oligodeoxynucleotide complementary to the template DNA strand (3′-5′, sense oligo) showed efficient inhibition of transcription. Incubation of this plasmid with the Oligodeoxynucleotide complementary to the 5′-3′ strand (antisense oligo) or a random Oligodeoxynucleotide failed to do so. The inhibition of gag transcription was specific since the sense oligo failed to prevent transcription of a plasmid containing U2 RNA sequences. The inhibition of transcription was not limited to T7 RNA polymerase but was also observed with SP6 RNA polymerase.
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Long-term treatment of human immunodeficiency virus-infected cells with antisense oligonucleotide phosphorothioates.
Proceedings of the National Academy of Sciences, 1993Co-Authors: Julianna Lisziewicz, Paul C Zamecnik, Daisy Sun, V. Metelev, Robert C. Gallo, Sudhir AgrawalAbstract:Abstract The antiviral activity of antisense Oligodeoxynucleotide phosphorothioates complementary to the tat gene, the gag mRNA, and the rev mRNA were studied in a long-term infection model. Three antisense oligonucleotides directed to the splice-acceptor site of the tat gene failed to suppress human immunodeficiency virus type 1 replication at 1 microM concentration in long-term culture. In contrast, two Oligodeoxynucleotide phosphorothioates (28-mer) complementary to the gag and the rev mRNAs inhibited viral replication for > 80 days, and the antiviral activity was sequence- and length-dependent. In addition, after pretreatment of cells we could reduce the concentration of the antisense Oligodeoxynucleotides by > 10-fold and still maintain the inhibition of viral replication. These results suggest that chemotherapy for human immunodeficiency virus type 1 infection with antisense Oligodeoxynucleotide phosphorothioates may be achieved by an initial high-dose treatment followed by a lower maintenance dose.
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Specific inhibition of human immunodeficiency virus type 1 replication by antisense oligonucleotides: an in vitro model for treatment
Proceedings of the National Academy of Sciences, 1992Co-Authors: Julianna Lisziewicz, Paul C Zamecnik, Sudhir Agrawal, Daisy Sun, Mary E. Klotman, Robert C. GalloAbstract:We have developed a culture system, simulating in vivo conditions of human immunodeficiency virus type 1 (HIV-1) infection, to evaluate the long-term efficacy of antisense oligonucleotide treatment. Five oligonucleotide phosphorothioates (28-mers), complementary to different regions of HIV-1 RNA, blocked replication of the virus in a sequence-specific manner at 1 microM concentration. Variations in antiviral activity were seen among the different oligonucleotides, revealing an effect of target selection. Mismatched or random oligonucleotide phosphorothioates delayed, but did not completely inhibit, HIV-1 replication. In the case of inhibition by a splice-acceptor-site antisense Oligodeoxynucleotide, a break-through phenomenon occurred after 25 days of treatment, suggesting the development of an "escape mutant." This result did not occur when the inhibitory Oligodeoxynucleotides were complementary to the primary-sequence areas of the rev-responsive element and rev-1 genes. Sequential treatment of HIV-1-infected cells with a combination of different antisense oligonucleotides, each administered once, also prevented the development of escape mutants. Our results suggest that chemotherapy based on specifically targeted antisense-oligonucleotide phosphorothioates may be an effective method for reducing the viral burden in HIV-1-infected individuals at clinically achievable oligonucleotide concentrations.
Giampaolo Tortora - One of the best experts on this subject based on the ideXlab platform.
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an antisense Oligodeoxynucleotide that depletes riα subunit of cyclic amp dependent protein kinase induces growth inhibition in human cancer cells
Cancer Research, 1993Co-Authors: Hiroshi Yokozaki, Giampaolo Tortora, Alfredo Budillon, Scott Meissner, Serge Beaucage, Keizaburo Miki, Yoon S ChochungAbstract:Abstract Enhanced expression of the RI α subunit of cyclic AMP-dependent protein kinase type I has been correlated with cancer cell growth. We provide evidence that RI α is a growth-inducing protein that may be essential for neoplastic cell growth. Human colon, breast, and gastric carcinoma and neuroblastoma cell lines exposed to a 21-mer human RI α antisense phosphorothioate Oligodeoxynucleotide (S-Oligodeoxynucleotide) exhibited growth inhibition with no sign of cytotoxicity. Mismatched sequence (random) S-Oligodeoxynucleotides of the same length exhibited no effect. The growth inhibitory effect of RI α antisense oligomer correlated with a decrease in the RI α mRNA and protein levels and with an increase in RII β (the regulatory subunit of protein kinase type II) expression. The growth inhibition was abolished, however, when cells were exposed simultaneously to both RI α and RII β antisense S-Oligodeoxynucleotides. The RII β antisense S-Oligodeoxynucleotide alone, exhibiting suppression of RII β along with enhancement of RI α expression, led to slight stimulation of cell growth. These results demonstrate that two isoforms of cyclic AMP receptor proteins, RI α and RII β , are reciprocally related in the growth control of cancer cells and that the RI α antisense Oligodeoxynucleotide, which efficiently depletes the growth stimulatory RI α , is a powerful biological tool toward suppression of malignancy.
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differentiation of hl 60 leukemia by type i regulatory subunit antisense Oligodeoxynucleotide of camp dependent protein kinase
Proceedings of the National Academy of Sciences of the United States of America, 1991Co-Authors: Giampaolo Tortora, Stefano Pepe, Timothy Clair, Hiroshi Yokozaki, Yoon S ChochungAbstract:A marked decrease in the type I cAMP-dependent protein kinase regulatory subunit (RI alpha) and an increase in the type II protein kinase regulatory subunit (RII beta) correlate with growth inhibition and differentiation induced in a variety of types of human cancer cells, in vitro and in vivo, by site-selective cAMP analogs. To directly determine whether RI alpha is a growth-inducing protein essential for neoplastic cell growth, human HL-60 promyelocytic leukemia cells were exposed to 21-mer RI alpha antisense Oligodeoxynucleotide, and the effects on cell replication and differentiation were examined. The RI alpha antisense oligomer brought about growth inhibition and monocytic differentiation, bypassing the effects of an exogenous cAMP analog. These effects of RI alpha antisense Oligodeoxynucleotide correlated with a decrease in RI alpha receptor and an increase in RII beta receptor level. The growth inhibition and differentiation were abolished, however, when these cells were exposed simultaneously to both RI alpha and RII beta antisense Oligodeoxynucleotides. The RII beta antisense Oligodeoxynucleotide alone has been previously shown to specifically block the differentiation inducible by cAMP analogs. These results provide direct evidence that RI alpha cAMP receptor plays a critical role in neoplastic cell growth and that cAMP receptor isoforms display specific roles in cAMP regulation of cell growth and differentiation.
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an antisense Oligodeoxynucleotide targeted against the type ii beta regulatory subunit mrna of protein kinase inhibits camp induced differentiation in hl 60 leukemia cells without affecting phorbol ester effects
Proceedings of the National Academy of Sciences of the United States of America, 1990Co-Authors: Giampaolo Tortora, Timothy Clair, Yoon S ChochungAbstract:The type II beta regulatory subunit of cAMP-dependent protein kinase (RII beta) has been hypothesized to play an important role in the growth inhibition and differentiation induced by site-selective cAMP analogs in human cancer cells, but direct proof of this function has been lacking. To address this issue, HL-60 human promyelocytic leukemia cells were exposed to RII beta antisense synthetic Oligodeoxynucleotide, and the effects on cAMP-induced growth regulation were examined. Exposure of these cells to RII beta antisense Oligodeoxynucleotide resulted in a decrease in cAMP analog-induced growth inhibition and differentiation without apparent effect on differentiation induced by phorbol esters. This loss in cAMP growth regulatory function correlated with a decrease in basal and induced levels of RII beta protein. Exposure to RII beta sense, RI alpha and RII alpha antisense, or irrelevant Oligodeoxynucleotides had no such effect. These results show that the RII beta regulatory subunit of protein kinase plays a critical role in the cAMP-induced growth regulation of HL-60 leukemia cells.
R Narayanan - One of the best experts on this subject based on the ideXlab platform.
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Antisense Oligodeoxynucleotides.
Current opinion in oncology, 1994Co-Authors: C A Stein, R NarayananAbstract:Oligodeoxynucleotides have been proposed as both in vitro and in vivo inhibitors of gene expression because of the specificity of Watson-Crick base pair hybridization. Phosphodiester Oligodeoxynucleotides (normal DNA) cannot be used as drugs because they are nuclease sensitive. Significant efforts have been made to study phosphorothioate Oligodeoxynucleotides, which have a sulfur atom substituted for one of the phosphate oxygen atoms at a nonbridging position. These Oligodeoxynucleotides are nuclease resistant, and over the past year they have entered clinical trials. They have also been extensively examined in vitro and have been targeted to the bcr-abl and bcl2 messenger RNAs among others. Methods to maximize the intracellular Oligodeoxynucleotide concentration have also been devised. However, significant problems remain, including the significant nonsequence specificity of phosphodiester Oligodeoxynucleotides as well as questions of Oligodeoxynucleotide uptake into and compartmentalization within cells. An improvement of our understanding of these phenomena is critical to the elaboration of this technology into a clinical therapeutic modality.
