Suberoylanilide hydroxyamic acid modification of chromatin architecture affects DNA break formation and repair.
OBJECTIVE
Chromatin-modifying compounds that inhibit the activity of histone deacetylases have shown potency as radiosensitizers, but the action of these drugs at a molecular level is not clear. Here we investigated the effect of suberoylanilide hydroxyamic acid (SAHA) on DNA breaks and their repair and induction of rearrangements.


METHODS
The effect of SAHA on both clonogenic survival and repair was assessed using cell lines SCC-25, MCF7, and TK6. In order to study unique DNA double-strand breaks, anti-CD95 antibody was employed to introduce a DNA double-strand break at a known location within the 11q23 region. The effects of SAHA on DNA cleavage and rearrangements were analyzed by ligation-mediated PCR and inverse PCR, respectively.


RESULTS
SAHA acts as radiosensitizer at 1 microM, with dose enhancement factors (DEFs) at 10% survival of: SCC-25 - 1.24 +/- 0.05; MCF7 - 1.16 +/- 0.09 and TK6 - 1.17 +/- 0.05, and it reduced the capacity of SCC-25 cells to repair radiation induced lesions. Additionally, SAHA treatment diffused site-specific fragmentation over at least 1 kbp in TK6 cells. Chromosomal rearrangements produced in TK6 cells exposed to SAHA showed a reduction in microhomology at the breakpoint between 11q23 and partner chromosomes.


CONCLUSIONS
SAHA shows efficacy as a radiosensitizer at clinically obtainable levels. In its presence, targeted DNA strand breaks occur over an expanded region, indicating increased chromatin access. The rejoining of such breaks is degraded by SAHA when measured as rearrangements at the molecular level and rejoining that contributes to cell survival.
Singh
Sheetal
S
Department of Radiation Oncology, University of California at Davis, 4501 X St., Sacramento, California 95817, USA.
Le
Hongan
H
Shih
Shyh-Jen
SJ
Ho
Bay
B
Vaughan
Andrew T
AT
D000107:Acetylation / Q000187:drug effects; D053148:Caspase 3 / Q000378:metabolism; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D002843:Chromatin / Q000187:drug effects*; D002880:Chromosomes, Human, Pair 11 / Q000187:drug effects; D053903:DNA Breaks, Double-Stranded / Q000187:drug effects*; D004260:DNA Repair / Q000528:radiation effects*; D004789:Enzyme Activation; D015321:Gene Rearrangement / Q000187:drug effects*; D006657:Histones / Q000378:metabolism; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology*; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology*; D000077337:Vorinostat
D002843:Chromatin; D006657:Histones; D006877:Hydroxamic Acids; D011838:Radiation-Sensitizing Agents; D000077337:Vorinostat; D053148:Caspase 3