A hexameric phosphorothioate oligonucleotide telomerase inhibitor arrests growth of Burkitt's lymphoma cells in vitro and in vivo.

A phosphorothioate oligonucleotide (PS-ODN) with sequence identical to the repeat sequence of the mammalian telomere, 5'-d(TTAGGG)-3', was incubated with a Burkitt's lymphoma-derived (OMA-BL1) cell line. This hexanucleotide inhibits telomerase activity in cell lysates, lengthens cell doubling time, and induces apoptosis. Concatenated repeats (12-, 18-, and 24-mers) of the 5'-d(TTAGGG)-3' motif induce similar cellular responses. Scrambled sequences do not efficiently inhibit telomerase activity or significantly alter cell growth and viability. The in vivo efficacy of this PS-ODN was evaluated in a xenograft human-nude mouse model. Once tumors were established these animals were administered the telomere mimic, 5'-d(TTAGGG)-3', a control scrambled sequence 5'-d(TGTGAG)-3', or saline for 14 days via a subcutaneous osmotic pumps in a blinded study monitoring tumor size with dose and time. A significant decrease in tumor size was observed in animals given 50 micrograms/mouse/day 5'-d(TTAGGG)-3', but not following 5'-d(TGTGAG)-3', relative to the saline-treated animals. The antitumor activity of the 6-mer telomere mimic demonstrated a dose dependency including a reduction in metastatic nodules in the spleen. No activity was observed with the scrambled controls. In addition to antitumor activity we observed an increase in the mouse hematopoietic progenitor cell populations, BFU-e and CFU-GM. These results demonstrated the effects of a short hexameric oligonucleotide telomere mimic in vitro and in vivo and the potential utility of short oligonucleotides as telomerase inhibitors.

[1]  G. Zon,et al.  In vivo treatment of human leukemia in a scid mouse model with c-myb antisense oligodeoxynucleotides. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[2]  J. McDougall,et al.  Telomerase activation by the E6 gene product of human papillomavirus type 16 , 1996, Nature.

[3]  J. Shay,et al.  Detection of telomerase activity in human cells and tumors by a telomeric repeat amplification protocol (TRAP) , 1995 .

[4]  D. Corey,et al.  Inhibition of human telomerase activity by peptide nucleic acids , 1996, Nature Biotechnology.

[5]  G. Morin.  The human telomere terminal transferase enzyme is a ribonucleoprotein that synthesizes TTAGGG repeats , 1989, Cell.

[6]  J. Shay,et al.  Detection of telomerase activity in malignant and nonmalignant skin conditions. , 1996, The Journal of investigative dermatology.

[7]  A Shimizu,et al.  Isolation of cDNA encoding a binding protein specific to 5'-phosphorylated single-stranded DNA with G-rich sequences. , 1993, Nucleic acids research.

[8]  E. Estey,et al.  Telomerase activity in human acute myelogenous leukemia: inhibition of telomerase activity by differentiation-inducing agents. , 1996, Clinical cancer research : an official journal of the American Association for Cancer Research.

[9]  E. Blackburn,et al.  Telomeres, telomerase and cancer. , 1996, Scientific American.

[10]  S. S. Smith,et al.  Hypermethylation of telomere-like foldbacks at codon 12 of the human c-Ha-ras gene and the trinucleotide repeat of the FMR-1 gene of fragile X. , 1994, Journal of molecular biology.

[11]  T. Tomonaga,et al.  Heterogeneous Nuclear Ribonucleoprotein K Is a DNA-binding Transactivator (*) , 1995, The Journal of Biological Chemistry.

[12]  P. Iversen,et al.  Oligonucleotides complementary to c-myb messenger RNA inhibit growth and induce apoptosis in human Burkitt lymphoma cells. , 1996, International journal of oncology.

[13]  F. Eckstein.  Phosphorothioate analogs of nucleotides , 1979 .

[14]  D. Weisenburger,et al.  Characterization of a newly established human burkitt's lymphoma cell line, OMA‐BL‐1 , 1991, International Journal of Cancer.

[15]  W. Telford,et al.  Evaluation of glucocorticoid‐induced DNA fragmentation in mouse thymocytes by flow cytometry , 1991, Cell proliferation.

[16]  J. Szostak,et al.  A mutant with a defect in telomere elongation leads to senescence in yeast , 1989, Cell.

[17]  E. Blackburn,et al.  Runaway telomere elongation caused by telomerase RNA gene mutations , 1995, Nature.

[18]  David M. Prescott,et al.  Inhibition of telomerase by G-quartet DMA structures , 1991, Nature.

[19]  F. Eckstein.  Investigation of enzyme mechanisms with nucleoside phosphorothioates. , 1975, Angewandte Chemie.

[20]  D. Ecker,et al.  Potent and specific inhibition of HIV envelope-mediated cell fusion and virus binding by G quartet-forming oligonucleotide (ISIS 5320). , 1994, AIDS research and human retroviruses.

[21]  J. Shay.  Aging and cancer: are telomeres and telomerase the connection? , 1995, Molecular medicine today.

[22]  H. W. Sharma,et al.  Telomerase as a potential molecular target to study G-quartet phosphorothioates. , 1996, Antisense & nucleic acid drug development.

[23]  D. Ecker,et al.  Combinatorially selected guanosine-quartet structure is a potent inhibitor of human immunodeficiency virus envelope-mediated cell fusion. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[24]  D. Hanahan,et al.  Differential regulation of telomerase activity and telomerase RNA during multi-stage tumorigenesis , 1996, Nature Genetics.

[25]  Michael Simons,et al.  Antisense c-myb oligonucleotides inhibit intimal arterial smooth muscle cell accumulation in vivo , 1992, Nature.

[26]  H. Blum,et al.  In vivo inhibition of duck hepatitis B virus replication and gene expression by phosphorothioate modified antisense oligodeoxynucleotides. , 1993, The EMBO journal.