Abstract
Introduction
Off-target effects of induction therapy to treat Acute Lymphoblastic Leukemia (ALL) can lead to hepatotoxicity, cardiotoxicity, and neuropathy, particularly in patients with previous co-morbidities. Incorporation of pegylated L-asparaginase (L-ASP) into pediatric and adult ALL induction regimens has increased remission rate but results in severe hepatotoxicity. Levocarnitine (L-carnitine), a small amino-acid derivative, has been reported in small case series in children and adults with ALL to reduce asparaginase-induced hepatotoxicity. L-carnitine functions as a transport molecule and facilitates long-chain free fatty acid beta-oxidation; exogenous supplementation with L-carnitine may therefore protect hepatocytes through facilitation of fatty acid transport and maintenance of antioxidant balance in mitochondria. However, the effects of L-carnitine on ALL cell survival and possible contributions of L-carnitine to chemotherapy resistance in ALL are not known. Given the ability of L-carnitine to enhance energy production, we sought to investigate whether L-carnitine could compromise the efficacy of common induction chemotherapy agents against ALL cells.
Methods
Human ALL cells, BV173 and RS4;11 were treated with L-carnitine and either daunorubicin (DNR) or vincristine (VCR) for 72 hours (n=4-6). RS4;11 cells were additionally treated with L-ASP (n=4), and current testing to assess dexamethasone in RS4;11 is in progress. Experiments were performed in 96-well culture plates with a seeding density of 90,000-120,000 cells/well (~5 x105 cells/mL). Growth media (RPMI 1640, 1% sodium pyruvate, 1% glutamine, 10% fetal bovine serum) was supplemented to physiologic (50 μM) and supraphysiologic (100-200 μM) levels of L-carnitine (normal reference range: 25-70 μM). Chemotherapy doses were chosen to represent ~EC90 concentrations (BV173: 2 nM VCR and 25 nM DNR; RS4;11: 2 nM VCR, 20 nM DNR, and 0.1 IU/mL L-ASP). After 72 hours, cell viability was measured by trypan blue exclusion and confirmed with spectrophotometric analysis with the Alamar Blue cell viability assay. Data were analyzed using one-way analysis of variance (ANOVA) with significance defined as p<0.05.
Results
Addition of L-carnitine did not significantly affect cytotoxicity of VCR or DNR in BV173 (n=6) or cytotoxicity of VCR, DNR, or L-ASP in RS4;11 (n=4) as measured by trypan blue exclusion at 72 hours (Figure 1A). L-carnitine also did not affect cell viability assessed using Alamar Blue colorimetric assay, presented as the difference in reduction of resazurin (resorufin) compared to untreated RS4;11 or BV173 cells (Figure 1B).
Conclusion
Our data show L-carnitine has no significant effect on the cytotoxicity of VCR, L-ASP, or DNR in two human ALL cell lines. For providers incorporating L-carnitine into induction therapy to reduce asparaginase-induced hepatotoxicity, this data supports its use without compromising the efficacy of induction therapy. Further investigation is necessary to determine whether L-carnitine alters efficacy of glucocorticoid drugs, particularly dexamethasone, and other chemotherapies used in the treatment of ALL and across other ALL subtypes.
No relevant conflicts of interest to declare.
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