All-trans-retinoic acid increases cytosine arabinoside cytotoxicity in HL-60 human leukemia cells in spite of decreased cellular ara-CTP accumulation.
BACKGROUND
Accumulation of the cytosine arabinoside (ara-C) metabolite ara-C-triphosphate (ara-CTP) in leukemic blast cells is considered to be the main determinant of ara-C cytotoxicity in vitro and in vivo. Retinoids such as all-trans-retinoic acid (ATRA) have been shown to increase the sensitivity of acute myelogenous leukemic (AML) blast cells to ara-C. To investigate the mechanism of this sensitisation, the hypothesis was tested that ATRA augments cellular ara-CTP levels in human-derived myelogenous leukemia HL-60 cells.


METHODS
The effect of ATRA and 13-cis-retinoic acid on ara-CTP accumulation and ara-C-induced apoptosis was studied. Ara-CTP levels were measured by high-performance liquid chromatography (HPLC), cytotoxicity by the tetrazolium (MTT) assay, and apoptosis by occurrence of DNA fragmentation (gel electrophoresis), cell shrinkage and DNA loss (flow cytometry).


RESULTS
Pretreatment of HL-60 cells with ATRA (0.01-1 microM) caused a significant decrease in intracellular ara-CTP levels; e.g., incubation for 72 hours with ATRA 1 microM prior to one hour ara-C 10 microM reduced ara-CTP levels to 41% +/- 4% of control. Similar results were obtained after preincubation with 13-cis-retinoic acid. In spite of decreased ara-CTP levels, the cytotoxicity of the combination was supraadditive and ATRA augmented ara-C-induced apoptosis.


CONCLUSIONS
At therapeutically relevant concentrations ATRA increased ara-C cytotoxicity and ara-C induced apoptosis but this augmentation is not the corollary of elevated ara-CTP levels. The feasibility of ara-C treatment optimisation via strategies other than those involving elevation of ara-CTP levels should be investigated further.
Freund
A
A
Department of Pediatric Hematology and Oncology, University of Münster Medical School, Germany.
Rössig
C
C
Lanvers
C
C
Gescher
A
A
Hohenlöchter
B
B
Jürgens
H
H
Boos
J
J
D000704:Analysis of Variance; D000964:Antimetabolites, Antineoplastic / Q000008:administration & dosage; D000971:Antineoplastic Combined Chemotherapy Protocols / Q000032:analysis / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D001085:Arabinofuranosylcytosine Triphosphate / Q000378:metabolism*; D003561:Cytarabine / Q000008:administration & dosage*; D053938:DNA Fragmentation / Q000187:drug effects; D019008:Drug Resistance, Neoplasm; D004587:Electrophoresis, Agar Gel; D005434:Flow Cytometry; D018922:HL-60 Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D014212:Tretinoin / Q000008:administration & dosage* / Q000032:analysis; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism
D000964:Antimetabolites, Antineoplastic; D003561:Cytarabine; D001085:Arabinofuranosylcytosine Triphosphate; D014212:Tretinoin