Cellular resistance to a nitric oxide releasing glutathione S-transferase P-activated prodrug, PABA/NO.
PABA/NO is a diazeniumdiolate selectively activated by glutathione S-transferase P (GSTP) to release nitric oxide (NO) and is a potent inducer of protein S-glutathionylation, a redox-sensitive post-translational modification of cysteine residues. Using a procedure that incrementally increased exposure of cells to PABA/NO, an acquired drug resistant human promyelocytic leukemia HL60 cell line (HL60(PABA)) that exhibited 1.9-fold resistance to the drug (IC(50) 15 μM vs ~8 μM for wild-type) was created. HL60(PABA) cells had a decreased growth rate attributable to altered cellular differentiation, as measured by increased expression of CD11b; decreased expression of CD14; decreased nuclear to cytoplasmic ratios and a condensation of nuclear chromatin. This was accompanied by alterations in both plasma and mitochondrial membrane potentials. Both GSTP expression and nitric oxide release were reduced two-fold, while increased expression levels of genes involved in the unfolded protein response (UPR) were evident in HL60(PABA) cells. Wild type cells treated with PABA/NO had increased levels of protein S-glutathionylation and JNK activation, while JNK was constitutively active in HL60(PABA) cells and these cells had reduced levels of S-glutathionylation. By removing PABA/NO from the growth medium, HL60(PABA) cells reverted to sensitivity within 21 days suggesting that resistance was not genetically stable. Mechanistically, PABA/NO resistance is mediated through reduced levels of GSTP resulting in reduced NO release and its subsequent alterations in cellular response to nitrosative stress.
Hutchens
Steven
S
Department of Pharmaceutical and Biomedical Sciences, Medical University of South Carolina, Charleston, SC, USA.
Manevich
Yefim
Y
He
Lin
L
Tew
Kenneth D
KD
Townsend
Danyelle M
DM
D010129:4-Aminobenzoic Acid / Q000737:chemistry / Q000494:pharmacology; D001391:Azo Compounds / Q000737:chemistry / Q000494:pharmacology*; D016923:Cell Death / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D048430:Cell Shape / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D019008:Drug Resistance, Neoplasm / Q000187:drug effects*; D005978:Glutathione / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism*; D018922:HL-60 Cells; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D009569:Nitric Oxide / Q000378:metabolism*; D011355:Prodrugs / Q000737:chemistry / Q000494:pharmacology*; D056811:Unfolded Protein Response / Q000187:drug effects / Q000235:genetics; D062366:para-Aminobenzoates
D001391:Azo Compounds; C485370:O(2)-(2,4-dinitro-5-(N-methyl-N-4-carboxyphenylamino)phenyl 1-N,N-dimethylamino)diazen-1-ium-1,2-diolate; D011355:Prodrugs; D062366:para-Aminobenzoates; D009569:Nitric Oxide; D005982:Glutathione Transferase; D048031:JNK Mitogen-Activated Protein Kinases; D005978:Glutathione; D010129:4-Aminobenzoic Acid