2 '-C-cyano-2 '-deoxy-1-beta-D-arabino-pentofuranosylcytosine: A novel anticancer nucleoside analog that causes both DNA strand breaks and G(2) arrest

The mechanism of 2'-C-cyano-2'-deoxy-1-beta -D-arabino-pentofuranosylcytosi ne (CNDAC) action was investigated in human lymphoblastoid CEM cells and my eloblastic leukemia ML-1 cells. CNDAC was metabolized to its 5'-triphosphat e and incorporated into DNA, which was associated with inhibition of DNA sy nthesis. After incubation of cells with [H-3]CNDAC, metabolites were detect ed in 3'-->5' phosphodiester linkage and at the 3' terminus of cellular DNA . Specific enzymatic hydrolysis of DNA demonstrated that the parent nucleos ide and its 2'-epimer 2'-C-cyano-2'-deoxy-2-ribo-pentofuranosylcytosine acc ounted for approximately 65% of the total analogs incorporated into DNA and essentially all of the drug in the 3'-->5' phosphodiester linkage. In cont rast, all detectable radioactivity at 3' termini was associated with 2'-C-c yano-2',3'-didehydro-2',3'-dideoxycytidine. This de facto DNA chain-termina ting nucleotide arises from an electronic characteristic and cleavage of th e 3'-phosphodiester bond subsequent to the addition of a nucleotide to the incorporated CNDAC moiety by beta -elimination, a process that generates a single strand break in DNA. Investigation of the biological consequences of these actions indicated that, after incubation with cytostatic concentrati ons of CNDAC, cell cycle progression was delayed during S phase, but that c ells arrested predominantly in the G(2) phase. This differed from the S pha se-arresting actions of ara-C and gemcitabine, other deoxycytidine analogs that inhibit DNA replication but do not cause strand breaks. Thus, once inc orporated into DNA, the CNDAC molecule appears to act by a dual mechanism t hat 1) delays the progress of further DNA replication, but 2) upon addition of a deoxynucleotide results in the conversion of the incorporated analog to a de facto DNA chain terminator at the 3' terminus of a single strand br eak. It is likely that DNA strand breaks trigger cell cycle arrest in G(2).