9-arabinofuranosylguanine and nelarabine

GW506U78 or nelarabine (Glaxo-SmithKline) is a nucleoside analog that is rapidly converted by cells of lymphoid lineage to its corresponding arabinosylguanine nucleotide triphosphate (araGTP). The triphosphate form of araG acts as a substrate for DNA polymerases and araG gets incorporated into the DNA, resulting in inhibition of DNA synthesis and subsequent cytotoxicity. It has been shown that nelarabine has activity as a single agent in patients with T-cell malignancies that have relapsed or are refractory to other therapy. The ongoing research on nelarabine has earned fast-track status from the U.S. Food and Drug Administration (FDA) for treatment of patients with T-cell acute lymphoblastic leukemia and lymphoblastic lymphoma who have not responded to or whose disease has progressed during treatment with at least two standard regimens. It is likely that nelarabine will be a useful drug in the treatment of leukemic diseases in the future and therefore nelarabine is an interesting drug to study further. Here we present an overview of what is known about the mechanism of action of nelarabine and its status in clinical trials.

Topics: Arabinonucleosides; Clinical Trials as Topic; DNA Repair; Dose-Response Relationship, Drug; Drug Resistance; Drug Therapy, Combination; Hematologic Neoplasms; Humans; Leukemia-Lymphoma, Adult T-Cell; Phosphorylation; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Treatment Outcome; Vidarabine

To characterize the pharmacokinetics of nelarabine (506U78), the water-soluble prodrug of 9-beta-D-arabinofuranosyl guanine (ara-G), and ara-G in pediatric and adult patients with refractory hematologic malignancies. Ara-G is phosphorylated within leukemic cells to form ara-G triphosphate (ara-GTP), which acts to terminate DNA chain elongation, resulting in cell death.. The pharmacokinetics of nelarabine and/or ara-G were evaluated in 71 patients (25 pediatric and 46 adult patients) on the first day of therapy. Blood was collected at specified times for the determination of plasma nelarabine and ara-G concentrations.. There were no statistically significant differences in the pharmacokinetics of nelarabine between any of the groups of patients. The harmonic mean half-life (t1/2) of nelarabine in pediatric and adult patients was 14.1 minutes and 16.5 minutes, respectively. The maximum concentrations (C(max)) of ara-G occurred at or near the end of the nelarabine infusion. The C(max) of ara-G ranged from 11.6 micromol/L to 308.7 micromol/L at nelarabine doses of 5 to 75 mg/kg and was linearly related to the nelarabine dose. No statistically significant differences were noted for the pharmacokinetic parameter estimates of ara-G between adult male and female patients. In children versus adults, the dose-normalized C(max), time of the C(max), and the steady-state volume of distribution of ara-G were similar. However, the clearance of ara-G was higher in pediatric patients (0.312 L.h(-1).kg(-1)) as compared with adult patients (0. 213 L.h(-1).kg(-1)) (P <.001). The t1/2 of ara-G was shorter in pediatric patients as compared with adult patients (2.1 hours v 3.0 hours; P <.01).. Nelarabine is an effective prodrug of ara-G, allowing systemic concentrations of ara-G that result in clinical activity.

Topics: Adolescent; Adult; Aged; Antineoplastic Agents; Arabinonucleosides; Child; Child, Preschool; Female; Hematologic Neoplasms; Humans; Male; Middle Aged

In vitro investigations with arabinosylguanine (ara-G) demonstrated potent cytotoxicity to T-lymphoblastoid cell lines. The goals of the present study were to evaluate GW506U78, a prodrug of ara-G, against human hematologic malignancies and to determine its pharmacokinetics in plasma and cells.. During a phase I multicenter trial of GW506U78, 26 patients were treated at M.D. Anderson Cancer Center (MDACC). Daily doses between 20 and 60 mg/kg were administered for 5 days. Parallel plasma and cellular pharmacokinetic studies were conducted.. Complete (n=5) or partial remission (n=5) was achieved in T-cell acute lymphoblastic leukemia (T-ALL), T-lymphoid blast crisis, T-lymphoma, and B-cell chronic lymphocytic leukemia (B-CLL) (n=13). In contrast, patients with B-ALL, B-lymphoma, acute myelogenous leukemia (AMI), or T-CLL did not respond. Peak plasma concentrations of GW506U78 and ara-G were dose-dependent. The elimination of GW506U78 (half-life [t1/2]=17 minutes) was faster than the elimination of ara-G (t1/2=3.7 hours). Median peak concentrations of ara-GTP were 23, 42, 85, and 93 micromol/L at 20, 30, 40, and 60 mg/kg, respectively. T-lymphoblasts accumulated significantly (P=.0008) higher peak arabinsylguanosine triphosphate (ara-GTP) (median, 140 micromol/L; n=7) compared with other diagnoses (median, 50 micromol/L; n=9) and normal mononuclear cells (n=3). The ara-GTP elimination was slow in all diagnoses (median, > 24 hours). Responders accumulated significantly (P=.0005) higher levels of ara-GTP (median, 157 micromol/L) compared with patients who failed to respond (median, 44 micromol/L).. GW506U78 is an effective prodrug and a potent agent for hematologic malignancies with major efficacy in T-cell diseases. The pharmacokinetics of ara-GTP in leukemia cells are strongly correlated with clinical responses to GW506U78.

Topics: Adult; Antineoplastic Agents; Arabinonucleosides; Arabinonucleotides; Child; Child, Preschool; Dose-Response Relationship, Drug; Guanosine Triphosphate; Hematologic Neoplasms; Humans; Leukemia, B-Cell; Leukemia, T-Cell; Multicenter Studies as Topic; Prodrugs; Time Factors; Treatment Outcome

The nucleoside analogue nelarabine, the prodrug of arabinosylguanine (AraG), is effective against T-cell acute lymphoblastic leukaemia (T-ALL) but not against B-cell ALL (B-ALL). The underlying mechanisms have remained elusive. Here, data from pharmacogenomics studies and a panel of ALL cell lines reveal an inverse correlation between nelarabine sensitivity and the expression of SAMHD1, which can hydrolyse and inactivate triphosphorylated nucleoside analogues. Lower SAMHD1 abundance is detected in T-ALL than in B-ALL in cell lines and patient-derived leukaemic blasts. Mechanistically, T-ALL cells display increased SAMHD1 promoter methylation without increased global DNA methylation. SAMHD1 depletion sensitises B-ALL cells to AraG, while ectopic SAMHD1 expression in SAMHD1-null T-ALL cells induces AraG resistance. SAMHD1 has a larger impact on nelarabine/AraG than on cytarabine in ALL cells. Opposite effects are observed in acute myeloid leukaemia cells, indicating entity-specific differences. In conclusion, SAMHD1 promoter methylation and, in turn, SAMHD1 expression levels determine ALL cell response to nelarabine.

Topics: Antineoplastic Agents; Arabinonucleosides; Biomarkers, Tumor; Cell Line, Tumor; DNA Methylation; Drug Resistance, Neoplasm; Gene Expression Regulation, Leukemic; Humans; Precursor B-Cell Lymphoblastic Leukemia-Lymphoma; Precursor T-Cell Lymphoblastic Leukemia-Lymphoma; Promoter Regions, Genetic; SAM Domain and HD Domain-Containing Protein 1

Nine-beta-D-arabinofuranosylguanine (ara-G), an active metabolite of nelarabine, enters leukemic cells through human Equilibrative Nucleoside Transporter 1, and is then phosphorylated to an intracellular active metabolite ara-G triphosphate (ara-GTP) by both cytosolic deoxycytidine kinase and mitochondrial deoxyguanosine kinase. Ara-GTP is subsequently incorporated into DNA, thereby inhibiting DNA synthesis.. In the present study, we developed a novel ara-G-resistant variant (CEM/ara-G) of human T-lymphoblastic leukemia cell line CCRF-CEM, and elucidated its mechanism of ara-G resistance. The cytotoxicity was measured by using the growth inhibition assay and the induction of apoptosis. Intracellular triphosphate concentrations were quantitated by using HPLC. DNA synthesis was evaluated by the incorporation of tritiated thymidine into DNA. Protein expression levels were determined by using Western blotting.. CEM/ara-G cells were 70-fold more ara-G-resistant than were CEM cells. CEM/ara-G cells were also refractory to ara-G-mediated apoptosis. The transcript level of human Equilibrative Nucleoside Transporter 1 was lowered, and the protein levels of deoxycytidine kinase and deoxyguanosine kinase were decreased in CEM/ara-G cells. The subsequent production of intracellular ara-GTP (21.3 pmol/107 cells) was one-fourth that of CEM cells (83.9 pmol/107 cells) after incubation for 6 h with 10 μM ara-G. Upon ara-G treatment, ara-G incorporation into nuclear and mitochondrial DNA resulted in the inhibition of DNA synthesis of both fractions in CEM cells. However, DNA synthesis was not inhibited in CEM/ara-G cells due to reduced ara-G incorporation into DNA. Mitochondrial DNA-depleted CEM cells, which were generated by treating CEM cells with ethidium bromide, were as sensitive to ara-G as CEM cells. Anti-apoptotic Bcl-xL was increased and pro-apoptotic Bax and Bad were reduced in CEM/ara-G cells.. An ara-G-resistant CEM variant was successfully established. The mechanisms of resistance included reduced drug incorporation into nuclear DNA and antiapoptosis.

Topics: Antineoplastic Agents; Apoptosis; Arabinonucleosides; Cell Line, Tumor; DNA; DNA Replication; Drug Resistance, Neoplasm; Humans; Precursor T-Cell Lymphoblastic Leukemia-Lymphoma

Forodesine and nelarabine (the pro-drug of ara-G) are 2 nucleoside analogues with promising anti-leukemic activity. To better understand which pediatric patients might benefit from forodesine or nelarabine (ara-G) therapy, we investigated the in vitro sensitivity to these drugs in 96 diagnostic pediatric leukemia patient samples and the mRNA expression levels of different enzymes involved in nucleoside metabolism. Forodesine and ara-G cytotoxicities were higher in T-cell acute lymphoblastic leukemia (T-ALL) samples than in B-cell precursor (BCP)-ALL and acute myeloid leukemia (AML) samples. Resistance to forodesine did not preclude ara-G sensitivity and vice versa, indicating that both drugs rely on different resistance mechanisms. Differences in sensitivity could be partly explained by significantly higher accumulation of intracellular dGTP in forodesine-sensitive samples compared with resistant samples, and higher mRNA levels of dGK but not dCK. The mRNA levels of the transporters ENT1 and ENT2 were higher in ara-G-sensitive than -resistant samples. We conclude that especially T-ALL, but also BCP-ALL, pediatric patients may benefit from forodesine or nelarabine (ara-G) treatment.

Topics: Antineoplastic Agents; Arabinonucleosides; Cell Line, Tumor; Child; Deoxycytidine Kinase; Deoxyguanine Nucleotides; Drug Resistance, Neoplasm; Equilibrative Nucleoside Transporter 1; Equilibrative-Nucleoside Transporter 2; Gene Expression; Humans; In Vitro Techniques; Leukemia, Myeloid, Acute; Leukemia, Prolymphocytic, B-Cell; Phosphotransferases (Alcohol Group Acceptor); Precursor T-Cell Lymphoblastic Leukemia-Lymphoma; Prodrugs; Purine Nucleosides; Purines; Pyrimidinones; RNA, Messenger; RNA, Neoplasm

The 9-beta-D-arabinofuranosylguanine (ara-G), an active compound of nelarabine, demonstrates potent cytotoxicity specifically on T-cell malignancies. In cells, ara-G is phosphorylated to ara-G triphosphate (ara-GTP), which is subsequently incorporated into DNA, thereby inhibiting DNA synthesis. Because ara-GTP is crucial to ara-G's cytotoxicity, the determination of ara-GTP production in cancer cells is informative for optimizing nelarabine administration. Here, we developed a new, sensitive isocratic-elution HPLC method for quantifying ara-GTP. Samples were eluted isocratically by using phosphate buffer at a constant flow rate. Ara-GTP was clearly separated from other nucleotides by using an anion-exchange column and it was quantitated by its peak area at 254 nm. The standard curve was linear with low variability and a sensitive detection limit (10 pmol). Furthermore, due to ara-G's specificity to T-cells we hypothesized that nelarabine might be effective against adult T-cell leukemia (ATL). The ara-GTP production was compared between T-lymphoblastic leukemia CCRF-CEM and ATL cell lines in vitro. When CEM cells were incubated with ara-G, the ara-GTP production increased in a concentration- and time-dependent manner. In contrast, 5 ATL cell lines accumulated lower ara-GTP in the same condition. While ara-G inhibited the growth of CEM cells with a 50% growth inhibition concentration of 2 microM, the inhibitory-concentration values were >1 mM in 8 of the 12 ATL cell lines. This ineffectiveness appeared to correspond with the low ara-GTP production. The present study is the first to evaluate the potential of ara-G against ATL cells; our results suggest that nelarabine would not be effective against ATL.

Topics: Antineoplastic Agents; Arabinonucleosides; Arabinonucleotides; Biomarkers; Cell Proliferation; Chemical Fractionation; Chromatography, High Pressure Liquid; Guanosine Triphosphate; Humans; Leukemia-Lymphoma, Adult T-Cell; Prodrugs; Reference Standards; Sensitivity and Specificity; Treatment Outcome; Tumor Cells, Cultured

Nelarabine is a water-soluble prodrug of the cytotoxic deoxyguanosine analog ara-G, to which it is rapidly converted in vivo by adenosine deaminase. Nelarabine has shown activity in the treatment of T-cell malignancies, especially T-cell acute lymphoblastic leukemia. Preliminary data suggested that nelarabine might penetrate into the CSF. We therefore studied the CSF penetration of nelarabine and ara-G in a nonhuman primate model that has been highly predictive of anticancer drug distribution in humans.. Nelarabine (35 mg/kg, approximately 700 mg/m2) was administered over 1 h through a surgically implanted central venous catheter to four nonhuman primates. Blood (four animals) and ventricular CSF (three animals) samples were obtained at intervals for 24 h for determination of nelarabine concentrations, which were measured by HPLC-mass spectrometry.. The nelarabine plasma AUC (median+/-s.d.) was 2,820+/-1,140 microM min and the ara-G plasma AUC was 20,000+/-8,100 microM min. The terminal half-life of nelarabine in plasma was 25+/-5.2 min and clearance was 42+/-61 ml/min/kg. The terminal half-life of ara-G in plasma was 182+/-45 min. In CSF the terminal half-life of nelarabine was 77+/-28 min and of ara-G was 232+/-79 min. The AUCcsf:AUCplasma was 29+/-11% for nelarabine and 23+/-12% for ara-G.. The excellent CSF penetration of nelarabine and ara-G supports further study of the contribution of nelarabine to the prevention and treatment of CNS leukemia.

Topics: Animals; Antineoplastic Agents; Arabinonucleosides; Chromatography, High Pressure Liquid; Macaca mulatta; Male; Mass Spectrometry; Prodrugs