Determinants of sensitivity of human T-cell leukemia CCRF-CEM cells to immucillin-H.
Immucillin-H (BCX-1777, forodesine) is a transition state analogue and potent inhibitor of PNP that shows promise as a specific agent against activated human T-cells and T-cell leukemias. The immunosuppressive or antileukemic effects of Immucillin-H (ImmH) in cultured cells require co-administration with deoxyguanosine (dGuo) to attain therapeutic levels of intracellular dGTP. In this study we investigated the requirements for sensitivity and resistance to ImmH and dGuo. (3)H-ImmH transport assays demonstrated that the equilibrative nucleoside transporters (ENT1 and ENT2) facilitated the uptake of ImmH in human leukemia CCRF-CEM cells whereas (3)H-dGuo uptake was primarily dependent upon concentrative nucleoside transporters (CNTs). Analysis of lysates from ImmH-resistant CCRF-CEM-AraC-8D cells demonstrated undetectable deoxycytidine kinase (dCK) activity, suggesting that dCK and not deoxyguanosine kinase (dGK) was the rate-limiting enzyme for phosphorylation of dGuo in these cells. Examination of ImmH cytotoxicity in a hypoxanthine-guanine phosphoribosyltransferase (HGPRT)-deficient cell line CCRF-CEM-AraC-8C, demonstrated enhanced sensitivity to low concentrations of ImmH and dGuo. RT-PCR and sequencing of HGPRT from the HGPRT-deficient CCRF-CEM-AraC-8C cells identified an Exon 8 deletion mutation in this enzyme. Thus these studies show that specific nucleoside transporters are required for ImmH cytotoxicity and predict that ImmH may be more cytotoxic to 6-thioguanine (6-TG) or 6-thiopurine-resistant leukemia cells caused by HGPRT deficiency.
Huang
Min
M
Department of Medicine, Stanford University, Stanford, CA 94305, USA.
Wang
Yanhong
Y
Gu
Jingjin
J
Yang
Jing
J
Noel
Karen
K
Mitchell
Beverly S
BS
Schramm
Vern L
VL
Graves
Lee M
LM
D000970:Antineoplastic Agents / Q000627:therapeutic use*; D017209:Apoptosis / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D003842:Deoxycytidine Kinase / Q000378:metabolism; D003849:Deoxyguanosine / Q000378:metabolism; D004354:Drug Screening Assays, Antitumor; D006801:Humans; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics / Q000378:metabolism; D015458:Leukemia, T-Cell / Q000188:drug therapy*; D008954:Models, Biological; D011684:Purine Nucleosides / Q000627:therapeutic use*; D011683:Purine-Nucleoside Phosphorylase / Q000627:therapeutic use; D011744:Pyrimidinones / Q000627:therapeutic use*
D000970:Antineoplastic Agents; D011684:Purine Nucleosides; D011744:Pyrimidinones; C113101:forodesine; D011683:Purine-Nucleoside Phosphorylase; D007041:Hypoxanthine Phosphoribosyltransferase; D003842:Deoxycytidine Kinase; D003849:Deoxyguanosine