Abstract
BackgroundAcquired glucocorticoid (GC) resistance remains the main obstacle in acute lymphoblastic leukemia (ALL) therapy. The aim of the present study was to establish a novel GC-resistant B-ALL cell line and investigate its biological characteristics.MethodsA cell culture technique was used to establish the GC-resistant cell line from the parental cell, NALM-6. Molecular and cellular biological techniques including flow cytometry, MTT assay, western blotting, DNA fingerprinting analysis and whole transcriptome sequencing (WTS) were used to characterize the GC-resistant cell lines. Nude mice were used for xenograft studies.ResultsThe GC-resistant cell line, NALM-6/HDR, was established by culturing NALM-6 cells under hypoxia for 5 weeks with a single dexamethasone (Dex) treatment. We subcloned the NALM-6/HDR cell lines, and got 6 monoclone Dex-resistant cell lines, NALM-6/HDR-C1, C3, C4, C5, C6 and C9 with resistance index (RI) ranging from 20,000–50,000. NALM-6/HDR and its monoclone cell line, NALM-6/HDR-C5, exhibited moderate (RI 5–15) to high resistance (RI > 20) to Ara-c; low or no cross-resistance to L-Asp, VCR, DNR, and MTX (RI < 5). STR analysis confirmed that NALM-6/HDR and NALM-6/H were all derived from NALM-6. All these cells derived from NALM-6 showed similar morphology, growth curves, immunophenotype, chromosomal karyotype and tumorigenicity. WTS analysis revealed that the main metabolic differences between NALM-6 or NALM-6/H (GC-sensitive) and NALM-6/HDR (GC-resistant) were lipid and carbohydrates metabolism. Western blotting analysis showed that NALM-6/HDR cells had a low expression of GR and p-GR. Moreover, AMPK, mTORC1, glycolysis and de novo fatty acid synthesis (FAS) pathway were inhibited in NALM-6/HDR when compared with NALM-6.ConclusionsNALM-6/HDR cell line may represent a subtype of B-ALL cells in patients who acquired GC and Ara-c resistance during the treatment. These patients may get little benefit from the available therapy target of AMPK, mTORC1, glycolysis and FAS pathway.
Highlights
Acquired glucocorticoid (GC) resistance remains the main obstacle in acute lymphoblastic leukemia (ALL) therapy
NALM-6/HDR cell line may represent a subtype of B-ALL cells in patients who acquired GC and Ara-c resistance during the treatment
Growing NALM-6 cells were cultured under hypoxia condition for 5–6 weeks with or without dexamethasone (Dex) treatment
Summary
Acquired glucocorticoid (GC) resistance remains the main obstacle in acute lymphoblastic leukemia (ALL) therapy. Acquired drug resistance to chemotherapy, especially resistance to glucocorticoids (GCs), remains the main obstacle in ALL therapy [2, 3]. There is a controversy regarding the traditional drug-exposure method for that the resistant phenotype obtained after a long time (6 ~ 16 months, or more) drug exposure and can only remain for several weeks or months after discontinuation of drug exposure, whether or not they can reflect the clinical setting [4]. We constructed a novel and convenient method to establish GC resistant ALL cell lines from GC sensitive ALL cells by mimicking the hypoxic bone marrow (BM) microenvironment [6]. Compared to the traditional methods, the novel method has a high success rate, a single drug exposure, a short duration (5 ~ 6 weeks), and a long maintenance of resistant phenotype without drug re-induction [6]
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