Clinical relevance of in vitro chemoresistance in childhood acute myeloid leukemia

Yamada, Shoichi; Hongo, Teruaki; Okada, Shuichi; Watanabe, Chieko; Fujii, Yasuhisa; Ohzeki, Takehiko

doi:10.1038/sj.leu.2402305

Cited by 41 publications

(38 citation statements)

References 25 publications

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“…The results of this assay have previously shown a good correlation with clinical results of leukemia treatment. 15,16,[22][23][24][25] We show that aplidin indeed is cytotoxic for leukemic cells in a dose-dependent fashion and in relatively low concentrations (10 À9 M ¼ nanomolar) as compared to other drugs we have previously tested in the MTT assay. These in vitro concentrations correlate with the drug levels reached in patients treated with pharmacologically appropriate doses of Aplidin.…”

Section: Discussionmentioning

confidence: 99%

“…The assay results correlate with clinical and cell biological features, as well as with clinical outcome after single agent and after combination chemotherapy. [14][15][16][17][18][19][20][21][22][23][24][25][26] To determine the potential of aplidin as a cytotoxic agent in pediatric leukemia, we have tested bone marrow (BM) and peripheral blood (PB) samples of children with different types of leukemia and of children without leukemia in the MTT assay and compared the results with in vitro cytotoxicity of known antileukemic agents.…”

Section: Introductionmentioning

confidence: 99%

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In vitro cytotoxicity of aplidin and crossresistance with other cytotoxic drugs in childhood leukemic and normal bone marrow and blood samples: a rational basis for clinical development

et al. 2003

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To determine the potential of aplidin as a cytotoxic agent in pediatric leukemia, we tested bone marrow (BM) and peripheral blood (PB) samples (n ¼ 72) of children with different types of leukemia and healthy children in the methyl-thiazol-tetrazolium assay. Also, we compared these results with other cytotoxic drugs. Aplidin was cytotoxic in vitro at nanomolar concentrations, in a dose-dependent fashion. L-carnitine, that is applied in clinical studies to prevent myotoxicity caused by aplidin, had no effect on aplidin cytotoxicity in vitro. Aplidin cytotoxicity in vitro was not different when initial and relapsed acute lymphoblastic leukemia (ALL) or initial ALL and initial acute myeloid leukemia were compared. However, normal BM (n ¼ 19) and PB (n ¼ 13) cells were more resistant to aplidin than leukemic cells (median two-to seven-fold, P ¼ 0.001 and median four-to 11-fold, Po0.0001, respectively). In leukemia samples, no significant crossresistance between aplidin and other cytotoxic drugs was found, except for a trend for correlation with 2 0 ,2 0 -difluorodeoxycytidine (q ¼ 0.71, P ¼ 0.02). In normal BM samples, significant crossresistance with the epipodophyllotoxins was found, which is not readily explained by the currently known mechanisms of action of aplidin. In conclusion, we show that aplidin has selective cytotoxicity in vitro towards childhood leukemia cells and generally lacks crossresistance with other known cytotoxic drugs, which warrants clinical studies.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

In vitro cytotoxicity of aplidin and crossresistance with other cytotoxic drugs in childhood leukemic and normal bone marrow and blood samples: a rational basis for clinical development

et al. 2003

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“…2 Several studies have shown that in vitro resistance to this drug is an independent prognostic factor in ALL. [3][4][5][6][7][8] Also a poor early in vivo response to L-asparaginase as a single drug has been linked to an unfavorable outcome in pediatric ALL. 9 The administration of L-asparaginase results in the deamination of asparagine into aspartic acid leading to a rapid and complete depletion of serum asparagine, which ultimately affects the intracellular asparagine levels.…”

Section: Introductionmentioning

confidence: 99%

Pharmacokinetic, pharmacodynamic and intracellular effects of PEG-asparaginase in newly diagnosed childhood acute lymphoblastic leukemia: results from a single agent window study

et al. 2008

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L-asparaginase is an effective drug for treatment of children with acute lymphoblastic leukemia (ALL). The effectiveness is thought to result from depletion of asparagine in serum and cells. We investigated the clinical response in vivo of 1000 IU/m 2 pegylated (PEG)-asparaginase and its pharmacokinetic, pharmacodynamic and intracellular effects in children with newly diagnosed ALL before start of combination chemotherapy. The in vivo window response was significantly related to immunophenotype and genotype: 26/38 common/pre B-ALL cases, especially those with hyperdiploidy and TELAML1 rearrangement, demonstrated a good clinical response compared to 8/17 T-ALL (P ¼ 0.01) and BCRABL-positive ALL (P ¼ 0.04). A poor in vivo clinical window response was related to in vitro resistance to L-asparaginase (P ¼ 0.02) and both were prognostic factors for long-term event-free survival (hazard ratio 6.4, P ¼ 0.004; hazard ratio 3.7, P ¼ 0.01). After administration of one in vivo dose of PEG-asparaginase no changes in apoptotic parameters or in intracellular levels of twenty amino acids in leukemic cells could be measured, in contradiction to the changes found after in vitro exposure. This may be explained by the rapid removal of apoptotic cells from the circulation in vivo. One additional dose of PEG-asparaginase upfront ALL treatment did not lead to other severe toxicities.

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“…Results of MTT assays indicated that AML-M5 is as sensitive to L-asparaginase as B-precursor ALL. 22,25 We intend to apply RQ-PCR to more clinical patients to evaluate AS expression levels as a useful marker on which to base clinical decisions regarding L-asparaginase therapy. …”

Section: Discussionmentioning

confidence: 99%

“…After a 4-day incubation in a 5% CO 2 atmosphere at 37°C, the results were not evaluated if the OD in control wells was below 0.050 to avoid inconsistencies. 22 …”

Section: Mtt Assay Of Cell Lines and Blast Cells From Patientsmentioning

confidence: 99%

Establishment of Real-Time Polymerase Chain Reaction Method for Quantitative Analysis of Asparagine Synthetase Expression

Irino¹,

Kitoh²,

Koami³

et al. 2004

The Journal of Molecular Diagnostics

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We established a real-time quantitative PCR (RQ-PCRAsparagine is not an essential amino acid obtained from outside the body because it is synthesized by using the hydrolysis energy of ATP from aspartic acid and glutamine via asparagine synthetase (AS). Even when the asparagine supply is reduced, normal cells can compensate by synthesizing L-asparagine. However, lymphoblastic cells require external asparagine for growth as they lack sufficient AS activity. 1-3 Thus, L-asparaginase is effective against childhood acute lymphoblastic leukemia (ALL) during the induction of remission or the intensification phases of treatment. 4,5 Asparagine in the blood, cerebrospinal fluid and bone marrow is depleted by L-asparaginase. A reduction of asparagine leads to cell death, since exposure to L-asparaginase in vitro induces the fragmentation of DNA and morphological changes typical of apoptosis in a mouse lymphoma cell line 6 and in NIH3T3 cells. 7 An asparagine deficiency can be evoked by the intracellular depletion of glutamate and glutamine. 7 The apoptosis of leukemia cells induced by L-asparaginase may be associated with cell cycle arrest in the G1 phase because DNA strand breaks can be seen in G1 phase cells as early as 8 hours after L-asparaginase exposure. 8 L-asparaginase activates AS expression and the overexpression of human AS protein can induce the L-asparaginase-resistance phenotype. 9 The expression of AS and sensitivity to L-asparaginase were correlated not only in leukemic, but also in ovarian cancer cells. 10 These observations indicate the importance of monitoring AS activity as a marker for clinical decisions regarding L-asparaginase therapy. The present study established real-time quantitative PCR (RQ-PCR) as a method of measuring AS expression. We discuss the clinical application of this method compared with immunohistochemistry, Western blotting, and enzyme activity. Materials and Methods Cell Lines and Culture ConditionsK562, HL60, U937, and MOLT4 were purchased from the American Type Culture Collection (ATCC, Manassas, VA). All cells were suspension-cultured in RPMI 1640 medium (Invitrogen Corp., Carlsbad, CA) containing 10% fetal bovine serum (FBS; Sigma, St. Louis, MO) and penicillin and streptomycin under a 5% CO 2 atmosphere. MOLT4/R 11 and U937/R 1 were established by sequential

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Clinical relevance of in vitro chemoresistance in childhood acute myeloid leukemia

Cited by 41 publications

References 25 publications

In vitro cytotoxicity of aplidin and crossresistance with other cytotoxic drugs in childhood leukemic and normal bone marrow and blood samples: a rational basis for clinical development

In vitro cytotoxicity of aplidin and crossresistance with other cytotoxic drugs in childhood leukemic and normal bone marrow and blood samples: a rational basis for clinical development

Pharmacokinetic, pharmacodynamic and intracellular effects of PEG-asparaginase in newly diagnosed childhood acute lymphoblastic leukemia: results from a single agent window study

Establishment of Real-Time Polymerase Chain Reaction Method for Quantitative Analysis of Asparagine Synthetase Expression

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