Lipid a radiosensitizes hypoxic EMT-6 tumor cells: role of the NF-kappaB signaling pathway.
OBJECTIVE
Lipid A has shown promising immunostimulatory effects in both experimental tumor models and advanced stage cancer patients. This study examines whether lipid A may directly modulate the radioresponse of tumor cells by activating inducible nitric oxide synthase (iNOS) or cyclooxygenase-2 (COX-2) through nuclear factor-kappaB (NF-kappaB) signaling.


METHODS
Hypoxic EMT-6 tumor cells were exposed to lipid A and analyzed for the level of COX-2 and iNOS by Western blotting and enzymatic assays. The hypoxic radioresponse of EMT-6 cells was estimated by clonogenic survival. The activation of NF-kappaB was examined by immunostaining of its p65 subunit and by luciferase reporter gene assay.


RESULTS
Lipid A dose-dependently increased the expression and activity of iNOS with a maximal effect at plasma achievable concentrations of 3-30 micro g/mL. The COX-2 mediated production of prostaglandin E2 was constitutively high and further upregulated by lipid A. The radiosensitivity of hypoxic EMT-6 cells was increased up to 2.5 times and counteracted by the iNOS inhibitor aminoguanidine but not by the COX-2 inhibitor NS-398. The mechanism of radiosensitization was linked to NF-kappaB signaling, because its inhibition by phenylarsine oxide impaired both iNOS activation and radioresponse.


CONCLUSIONS
Lipid A is an efficient hypoxic cell radiosensitizer at plasma relevant concentrations, which provides a rationale to combine lipid A with radiotherapy in further studies.
de Ridder
Mark
M
Academic Hospital Free University Brussels (A.Z.-V.U.B.), Oncology Center, Cancer Research Unit, Brussels, Belgium. Mark.De.Ridder@vub.ac.be
Verovski
Valeri N
VN
Van den Berge
Dirk L
DL
Sermeus
Alexandra B L
AB
Monsaert
Christinne
C
Wauters
Nathalie
N
Storme
Guy A
GA
D015687:Cell Hypoxia / Q000187:drug effects; D045744:Cell Line, Tumor / Q000187:drug effects / Q000528:radiation effects; D051546:Cyclooxygenase 2; D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006146:Guanidines; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism*; D008050:Lipid A / Q000494:pharmacology*; D008565:Membrane Proteins; D016328:NF-kappa B / Q000502:physiology*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D052247:Nitric Oxide Synthase Type II; D009578:Nitrobenzenes / Q000494:pharmacology; D011451:Prostaglandin-Endoperoxide Synthases / Q000378:metabolism*; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology*; D015398:Signal Transduction / Q000502:physiology*; D013449:Sulfonamides / Q000494:pharmacology
D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors; D004791:Enzyme Inhibitors; D006146:Guanidines; D007527:Isoenzymes; D008050:Lipid A; D008565:Membrane Proteins; D016328:NF-kappa B; D009578:Nitrobenzenes; D011838:Radiation-Sensitizing Agents; D013449:Sulfonamides; C080955:N-(2-cyclohexyloxy-4-nitrophenyl)methanesulfonamide; C496319:NOS2 protein, human; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases; C004479:pimagedine