ING1 and 5-azacytidine act synergistically to block breast cancer cell growth.
BACKGROUND
Inhibitor of Growth (ING) proteins are epigenetic "readers" that recognize trimethylated lysine 4 of histone H3 (H3K4Me3) and target histone acetyl transferase (HAT) and histone deacetylase (HDAC) complexes to chromatin.


RESULTS
Here we asked whether dysregulating two epigenetic pathways with chemical inhibitors showed synergistic effects on breast cancer cell line killing. We also tested whether ING1 could synergize better with chemotherapeutics that target the same epigenetic mechanism such as the HDAC inhibitor LBH589 (Panobinostat) or a different epigenetic mechanism such as 5-azacytidine (5azaC), which inhibits DNA methyl transferases. Simultaneous treatment of breast cancer cell lines with LBH589 and 5azaC did not show significant synergy in killing cells. However, combination treatment of ING1 with either LBH589 or 5azaC did show synergy. The combination of ING1b with 5azaC, which targets two distinct epigenetic mechanisms, was more effective at lower doses and enhanced apoptosis as determined by Annexin V staining and cleavage of caspase 3 and poly-ADP-ribose polymerase (PARP). ING1b plus 5azaC also acted synergistically to increase γH2AX staining indicating significant levels of DNA damage were induced. Adenoviral delivery of ING1b with 5azaC also inhibited cancer cell growth in a murine xenograft model and led to tumor regression when viral concentration was optimized in vivo.


CONCLUSIONS
These data show that targeting distinct epigenetic pathways can be more effective in blocking cancer cell line growth than targeting the same pathway with multiple agents, and that using viral delivery of epigenetic regulators can be more effective in synergizing with a chemical agent than using two chemotherapeutic agents. This study also indicates that the ING1 epigenetic regulator may have additional activities in the cell when expressed at high levels.
Department of Biochemistry and Molecular Biology, University of Calgary, Calgary, Alberta, Canada.
Satbir
S
Thakur
Xiaolan
X
Feng
Zhong
Z
Qiao Shi
Amudha
A
Ganapathy
Manoj
M
Kumar Mishra
Peter
P
Atadja
Don
D
Morris
Karl
K
Riabowol
D000818:Animals; D000964:Antimetabolites, Antineoplastic / Q000627:therapeutic use; D001374:Azacitidine / Q000627:therapeutic use*; D001943:Breast Neoplasms / Q000188:drug therapy* / Q000628:therapy*; D045744:Cell Line, Tumor; D005260:Female; D006801:Humans; D006877:Hydroxamic Acids / Q000627:therapeutic use; D007211:Indoles / Q000627:therapeutic use; D000076023:Inhibitor of Growth Protein 1; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D016513:Mice, SCID; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D000077767:Panobinostat; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism*; D023041:Xenograft Model Antitumor Assays
D000964:Antimetabolites, Antineoplastic; D006877:Hydroxamic Acids; C501503:ING1 protein, human; D007211:Indoles; D000076023:Inhibitor of Growth Protein 1; D047908:Intracellular Signaling Peptides and Proteins; D009687:Nuclear Proteins; D025521:Tumor Suppressor Proteins; D000077767:Panobinostat; D001374:Azacitidine