Effects of cytarabine on activation of human T cells - cytarabine has concentration-dependent effects that are modulated both by valproic acid and all-trans retinoic acid.
BACKGROUND
Cytarabine is used in the treatment of acute myeloid leukemia (AML). Low-dose cytarabine can be combined with valproic acid and all-trans retinoic acid (ATRA) as AML-stabilizing treatment. We have investigated the possible risk of immunotoxicity by this combination. We examined the effects of cytarabine combined with valproic acid and ATRA on in vitro activated human T cells, and we tested cytarabine at concentrations reached during in vivo treatment with high doses, conventional doses and low doses.


METHODS
T cells derived from blood donors were activated in vitro in cell culture medium alone or supplemented with ATRA (1 μM), valproic acid (500 or 1000 μM) or cytarabine (0.01-44 μM). Cell characteristics were assessed by flow cytometry. Supernatants were analyzed for cytokines by ELISA or Luminex. Effects on primary human AML cell viability and proliferation of low-dose cytarabine (0.01-0.5 μM) were also assessed. Statistical tests include ANOVA and Cluster analyses.


RESULTS
Only cytarabine 44 μM had both antiproliferative and proapoptotic effects. Additionally, this concentration increased the CD4:CD8 T cell ratio, prolonged the expression of the CD69 activation marker, inhibited CD95L and heat shock protein (HSP) 90 release, and decreased the release of several cytokines. In contrast, the lowest concentrations (0.35 and 0.01 μM) did not have or showed minor antiproliferative or cytotoxic effects, did not alter activation marker expression (CD38, CD69) or the release of CD95L and HSP90, but inhibited the release of certain T cell cytokines. Even when these lower cytarabine concentrations were combined with ATRA and/or valproic acid there was still no or minor effects on T cell viability. However, these combinations had strong antiproliferative effects, the expression of both CD38 and CD69 was altered and there was a stronger inhibition of the release of FasL, HSP90 as well as several cytokines. Cytarabine (0.01-0.05 μM) showed a dose-dependent antiproliferative effect on AML cells, and in contrast to the T cells this effect reached statistical significance even at 0.01 μM.


CONCLUSIONS
Even low levels of cytarabine, and especially when combined with ATRA and valproic acid, can decrease T cell viability, alter activation-induced membrane-molecule expression and decrease the cytokine release.
Institute of Clinical Science, University of Bergen, Bergen, Norway. elisabeth.ersver@hib.no.|Institute of Biomedical Laboratory Sciences, Bergen University College, Nygårdsgaten 112, P.O. Box 7030, N-5020, Bergen, Norway. elisabeth.ersver@hib.no.
Elisabeth
E
Ersvaer
Institute of Clinical Science, University of Bergen, Bergen, Norway. Annette.Brenner@uib.no.
Annette K
AK
Brenner
Institute of Clinical Science, University of Bergen, Bergen, Norway. kristin.vetas@gmail.com.
Kristin
K
Vetås
Institute of Clinical Science, University of Bergen, Bergen, Norway. Hakon.Reikvam@uib.no.
Håkon
H
Reikvam
Institute of Clinical Science, University of Bergen, Bergen, Norway. oystein.bruserud@helse-bergen.no.|Department of Medicine, Haukeland University Hospital, Bergen, Norway. oystein.bruserud@helse-bergen.no.
Øystein
Ø
Bruserud
D051997:ADP-ribosyl Cyclase 1 / Q000096:biosynthesis / Q000187:drug effects; D015703:Antigens, CD / Q000096:biosynthesis / Q000187:drug effects; D000945:Antigens, Differentiation, T-Lymphocyte / Q000096:biosynthesis / Q000187:drug effects; D015704:CD4 Antigens / Q000276:immunology; D016827:CD8 Antigens / Q000276:immunology; D002462:Cell Membrane / Q000187:drug effects / Q000276:immunology; D049109:Cell Proliferation; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003561:Cytarabine / Q000008:administration & dosage / Q000494:pharmacology*; D016207:Cytokines / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D053222:Fas Ligand Protein / Q000378:metabolism; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D037181:Lectins, C-Type / Q000096:biosynthesis / Q000187:drug effects; D008213:Lymphocyte Activation / Q000187:drug effects*; D013601:T-Lymphocytes / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000378:metabolism; D014212:Tretinoin / Q000008:administration & dosage / Q000494:pharmacology*; D014635:Valproic Acid / Q000494:pharmacology*; D000099192:CD69 Antigens
D051997:ADP-ribosyl Cyclase 1; D015703:Antigens, CD; D000945:Antigens, Differentiation, T-Lymphocyte; D015704:CD4 Antigens; D016827:CD8 Antigens; D003561:Cytarabine; D016207:Cytokines; D053222:Fas Ligand Protein; D018840:HSP70 Heat-Shock Proteins; D018841:HSP90 Heat-Shock Proteins; D037181:Lectins, C-Type; D053221:TNF-Related Apoptosis-Inducing Ligand; D014212:Tretinoin; D014635:Valproic Acid; D000099192:CD69 Antigens; C505661:FASLG protein, human