Diminished asparaginase turnover due to a germ-line mutation in cathepsin B.

Abstract

9550 Background: ASNase is a key component of multi-agent chemotherapy protocols used in the treatment of pediatric as well as adult acute lymphoblastic leukemia (ALL). Patients that are treated according to fixed shedules show a strong inter individual variation in serum ASNase levels. While underexposure may compromise therapeutic benefits, strongly elevated serum levels may lead to serious adverse effects. In at least 10% of the patients the desired ASNase levels are either not reached or exceeded. Therefore our understanding of ASNase dynamics in vivo needs to be improved. Here we describe the identification of a novel mutation in the gene encoding Cathepsin B to diminished ASNase turnover in a pediatric patient with ALL. Methods: Sequencing, biochemical experiments and immunofluorescence. Results: A 3-year-old girl diagnosed with ALL experienced serious toxicity in the form of hyperammonemic encephalopathy during treatment with ASNase derived from Erwinia chrysantemi. Pharmacokinetic data showed a strongly delayed clearance of the ASNase, which with the standard treatment protocol resulted in extremely high serum ASNase levels. Plasmapheresis was performed and dosage frequencies were adjusted to improve the clinical status of the patient. No underlying metabolic disorder could be diagnosed. Based on a previous report describing the role of proteases in degradation of ASNase in vitro, we hypothesized that the cysteine protease Cathepsin B might be implicated in the strongly reduced clearance of ASNase in this patient. Sequencing of the open reading frame of the Cathepsin B gene (CTSB) revealed a single codon deletion in the germline of the patient, affecting a lysine residue in the carboxy terminus of the protein. Immunofluorescence and biochemical experiments show that this single amino acid deletion leads to a protein product that is retained in the endoplasmic reticulum and is inefficiently processed. Conclusions: ASNase degradation assays show that the mutant Cathepsin B has lost its ability to efficiently degrade ASNase which could explain the high levels of ASNase as observed in our patient.Together, these findings suggest that variations in Cathepsin B activity may influence serum ASNase levels in the patients.