Low Bioavailability and High Immunogenicity of a New Brand of E. coli l-Asparaginase with Active Host Contaminating Proteins

Abstract

The drug l-asparaginase is a cornerstone in the treatment of acute lymphoblastic leukemia (ALL). The native E. colil-asparaginase used in Brazil until recently has been manufactured by Medac/Kyowa. Then a decision was taken by the Ministry of Health in 2017 to supply the National Health System with a cheaper alternative l-asparaginase manufactured by Beijing SL Pharmaceutical, called Leuginase®. As opposed to Medac, the asparaginase that has been in use in Brazil under the trade name of Aginasa®, it was not possible to find a single entry with the terms Leuginase in the Pubmed repository. The apparent lack of clinical studies and the scarcity of safety information provided to the hospitals by the drug distributor created a debate among Brazilian pediatric oncologists about issues of safety and efficacy that culminated eventually in a court decision to halt the distribution of the new drug all over the country. Boldrini Children's Center, a non-profit pediatric oncohematology hospital, has conducted its own evaluation of Leuginase®. Mass spectrometry analyses found at least 12 different contaminating host-cell proteins (HCP) in Leuginase®. The presence of two HCP (beta-lactamase and malate dehydrogenase) was confirmed by orthogonal methodologies. The relative number of HCP peptides ranged from 19 to 37% of the total peptides identified by mass spectrometry. In vivo studies in mice injected with Leuginase® revealed a 3 times lower plasma bioavailability and the development of higher antibody titres against l-asparaginase in comparison to Aginasa®-injected animals. The decision to buy a new drug based on its price alone is not safe. Developing countries are especially vulnerable to cheaper alternatives that lack solid quality assurance.

A comparison of asparaginase activity in generic formulations of E.coli derived L‐ asparaginase: In‐vitro study and retrospective analysis of asparaginase monitoring in pediatric patients with leukemia

“…It was the only generic available in the hospital pharmacy at the time of this analysis. The sample was prepared using a method previously described by Zenatti et al Briefly, the sample was reconstituted in normal saline from which 30 μg was aliquoted to which 10 μL urea 8 M and 0.4 μL 250mM Dithiothreitol (DTT) was added. For reduction, the sample was incubated at 56°C for 25 min, following which 0.57 μL 500mM iodoacetamide was added and the sample was incubated for 30 min at room temperature in the dark for alkylation.…”

Science and health for all children with cancer

“…However, native asparaginase (derived from the bacterium Escherichia coli) has been only intermittently accessible in many settings (82). In 2017, anticipating drug shortage of native asparaginase, the Ministry of Health in Brazil changed the national supplier to a foreign manufacturer that offered a new lowercost generic product (83). Concerned by the lack of published data on this new product, investigators in Brazil studied the drug's properties, and found that it was less bioactive, and contained contaminating proteins that increased the risk for immune-related side effects compared with the native asparaginase used previously (83,84).…”

“…In 2017, anticipating drug shortage of native asparaginase, the Ministry of Health in Brazil changed the national supplier to a foreign manufacturer that offered a new lowercost generic product (83). Concerned by the lack of published data on this new product, investigators in Brazil studied the drug's properties, and found that it was less bioactive, and contained contaminating proteins that increased the risk for immune-related side effects compared with the native asparaginase used previously (83,84). Although the nationwide distribution of this new product in Brazil was halted, this drug continues to be manufactured and distributed to other countries around world (83).…”

“…Concerned by the lack of published data on this new product, investigators in Brazil studied the drug's properties, and found that it was less bioactive, and contained contaminating proteins that increased the risk for immune-related side effects compared with the native asparaginase used previously (83,84). Although the nationwide distribution of this new product in Brazil was halted, this drug continues to be manufactured and distributed to other countries around world (83). Moving forward, the WHO's 2018 global initiative will hopefully engage additional partners to increase economies of scale and uphold accountability mechanisms for product availability and quality consistency worldwide.…”

Development of L-Asparaginase Biobetters: Current Research Status and Review of the Desirable Quality Profiles

“…The downstream process must achieve the removal of host cell protein (HCP), process and products related impurities, DNA, buffer, antifoam agents, aggregates, fragments, among others (Jozala et al, 2016; Lopes et al, 2017). Regarding injectable biopharmaceuticals, such as ASNase, the manufacturing process must be even more judicious, since the presence of contaminants, may potentially lead to detrimental clinical consequences (Zenatti et al, 2018).…”