N(1)-guanyl-1,7,-diamineoheptane, an inhibitor of deoxyhypusine synthase, suppresses differentiation and induces apoptosis via mitochondrial and AMPK pathways in immortalized and malignant human oral keratinocytes.
BACKGROUND
Although N(1)-guanyl-1,7,-diamineoheptane (GC7), an inhibitor of deoxyhypusine synthase, has been shown to inhibit cell growth, the mechanism of its action is not completely understood. In this study, we investigated the mechanisms of the effects of GC7 on cell growth, differentiation and apoptosis in relation to adenosine monophosphate-activated protein kinase (AMPK) activation, as AMPK is known to be a possible target for cancer treatment.


METHODS
The effects of GC7 on the growth of immortalized human oral keratinocytes (IHOK) and primary oral cancer cells (HN4), was investigated using MTT assay, Western blotting, cell cycle analysis, DNA fragmentation and expression of apoptotic pathway proteins.


RESULTS
N(1)-guanyl-1,7,-diamineoheptane inhibited cell proliferation in a time- and dose-dependent manner in IHOK and HN4 cells. GC7 treatment decreased the expression of differentiation markers, such as involucrin, CK13 and CK19. The major mechanism of growth inhibition by GC7 treatment was induction of apoptosis, which is supported by sub-G(1) phase arrest, annexin V-FITC staining and DNA fragmentation analysis. GC7 treatment increased the cytosolic level of cytochrome c and resulted in caspase-3 activation. GC7 treatment also resulted in a strong activation of AMPK. Furthermore, specific AMPK activator blocked the GC7-induced growth inhibition effect, as well as apoptosis.


CONCLUSIONS
These results demonstrate that GC7 blocks immortalized and malignant keratinocyte cell proliferation and differentiation by inducing apoptosis through the mitochondrial and AMPK pathways. On the basis of these observations, we propose that a strategy combining GC7 and AMPK inhibition could be developed into a novel chemotherapeutic modality in oral cancer.
Lee
S-K
SK
Department of Oral & Maxillofacial Pathology, College of Dentistry, Wonkwang University, Iksan, South Korea.
Lee
J
J
Lee
S-I
SI
Bae
W-J
WJ
Lee
Y-M
YM
Park
J-S
JS
Lee
S-K
SK
Park
S-J
SJ
Min
S-K
SK
Kim
E-C
EC
D055372:AMP-Activated Protein Kinases / Q000187:drug effects*; D017304:Annexin A5 / Q000187:drug effects; D017209:Apoptosis / Q000187:drug effects*; D002277:Carcinoma / Q000473:pathology*; D053148:Caspase 3 / Q000187:drug effects; D018797:Cell Cycle Proteins / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D045304:Cytochromes c / Q000187:drug effects; D053938:DNA Fragmentation; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000008:administration & dosage / Q000494:pharmacology*; D016193:G1 Phase / Q000187:drug effects; D006147:Guanine / Q000008:administration & dosage / Q000031:analogs & derivatives* / Q000494:pharmacology; D006801:Humans; D053535:Keratin-13 / Q000037:antagonists & inhibitors; D053539:Keratin-19 / Q000037:antagonists & inhibitors; D015603:Keratinocytes / Q000187:drug effects*; D008928:Mitochondria / Q000187:drug effects*; D009061:Mouth Mucosa / Q000187:drug effects* / Q000473:pathology; D009062:Mouth Neoplasms / Q000473:pathology*; D000587:Oxidoreductases Acting on CH-NH Group Donors / Q000037:antagonists & inhibitors*; D011498:Protein Precursors / Q000037:antagonists & inhibitors
D017304:Annexin A5; D018797:Cell Cycle Proteins; D004791:Enzyme Inhibitors; D053535:Keratin-13; D053539:Keratin-19; C100667:N(1)-guanyl-1,7-diaminoheptane; D011498:Protein Precursors; D006147:Guanine; C032253:involucrin; D045304:Cytochromes c; D000587:Oxidoreductases Acting on CH-NH Group Donors; C063507:deoxyhypusine synthase; D055372:AMP-Activated Protein Kinases; D053148:Caspase 3