L-Asparaginase Purification

Tundisi, Louise Lacalendola; Coelho, D.; Zanchetta, Beatriz; Moriel, Patrícia; Pessoa, Adalberto; Tambourgi, Elias Basile; Mazzola, Priscila Gava

doi:10.1080/15422119.2016.1184167

Cited by 23 publications

(14 citation statements)

References 69 publications

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“…The presence of cytoplasmic and membrane bound proteins possibly indicates a deficiency in the extraction process of the crude periplasmic protein. Additionally the high amount of HCPs could be due to insufficient purification of the final product . Though not examined in our study, high levels of HCPs can lead to drug antibody reactions and cause inactivation of L‐asparaginase along with a higher chance of hypersensitivity reactions .…”

Section: Discussionmentioning

confidence: 88%

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

Sankaran

Sengupta

Purohit

et al. 2020

Brit J Clinical Pharma

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Aims L‐asparaginase is an essential medicine in the treatment of pediatric acute lymphoblastic leukemia (ALL) and the quality of generic formulations is an area of concern. We compared nine generic formulations of L‐asparaginase available in India with the innovator. Methods The quality of formulations was assessed by measuring 72‐hour trough asparaginase activity in children with ALL during induction following administration of 10,000 IU/m2 of L‐asparaginase. In‐vitro analysis of the label claim was assessed by measuring activity of three generic formulations. Liquid chromatography‐mass spectrometry (LC/MS) was used to determine the amount of host contaminant proteins (HCPs) in the formulations. Results Between March 2015 to June 2018, 240 samples from 195 patients were analyzed. The number of samples analyzed ranged from 7–66 per generic brand (median: 18) and seven of the innovator. The proportion of generic formulations that failed to achieve a predefined clinical threshold activity of 50 IU/L ranged from 16.7% (2/12) to 84.9% (28/33) in the highest activity to lowest activity generic respectively. On other hand, all innovator samples had activity greater than 50 IU/L. In‐vitro asparaginase activity in the three generic formulations tested ranged from 71.4–74.6% of the label claim (10,000 IU) compared to 93.5% for the innovator. LC/MS analysis of generic 5 identified 25 HCPs with a relative peptide count of 27.1% of the total peptides. Conclusions Generic formulations had lower asparaginase activity which raises serious clinical concerns regarding their quality. Until stringent regulatory enforcement improves the quality of these generics, dose adaptive strategies coupled with therapeutic drug monitoring need to be considered.

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Section: Discussionmentioning

confidence: 88%

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

Sankaran

Sengupta

Purohit

et al. 2020

Brit J Clinical Pharma

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“…Downstream processes present a large number of potentially critical process parameters that show interactions across unit operations and have to be investigated before scale-up (Meitz et al, 2014). Since 60–80% of the total production costs of biopharmaceuticals is usually associated with downstream processing, it has become crucial to investigate how to replace traditional methods with efficient and cost-effective alternative techniques for recovery and purification of drugs, decreasing the number of downstream unit operations (Buyel and Fischer, 2014; Tundisi et al, 2017). The integration of downstream unit operations is widely used for ASNase purification, through low resolution purification steps, i.e., fractional precipitation, aqueous biphasic systems, centrifugation and membrane-based purification (dialysis) (Zhu et al, 2007; Mahajan et al, 2014; El-Naggar et al, 2016) and high resolution purification steps, i.e., chromatographic processes.…”

Section: L-asparaginase Manufacturing: Production Process and Purificmentioning

confidence: 99%

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

Brumano

Silva

Costa-Silva

et al. 2019

Front. Bioeng. Biotechnol.

144

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L-Asparaginase (ASNase) is a vital component of the first line treatment of acute lymphoblastic leukemia (ALL), an aggressive type of blood cancer expected to afflict over 53,000 people worldwide by 2020. More recently, ASNase has also been shown to have potential for preventing metastasis from solid tumors. The ASNase treatment is, however, characterized by a plethora of potential side effects, ranging from immune reactions to severe toxicity. Consequently, in accordance with Quality-by-Design (QbD) principles, ingenious new products tailored to minimize adverse reactions while increasing patient survival have been devised. In the following pages, the reader is invited for a brief discussion on the most recent developments in this field. Firstly, the review presents an outline of the recent improvements on the manufacturing and formulation processes, which can severely influence important aspects of the product quality profile, such as contamination, aggregation and enzymatic activity. Following, the most recent advances in protein engineering applied to the development of biobetter ASNases (i.e., with reduced glutaminase activity, proteolysis resistant and less immunogenic) using techniques such as site-directed mutagenesis, molecular dynamics, PEGylation, PASylation and bioconjugation are discussed. Afterwards, the attention is shifted toward nanomedicine including technologies such as encapsulation and immobilization, which aim at improving ASNase pharmacokinetics. Besides discussing the results of the most innovative and representative academic research, the review provides an overview of the products already available on the market or in the latest stages of development. With this, the review is intended to provide a solid background for the current product development and underpin the discussions on the target quality profile of future ASNase-based pharmaceuticals.

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“…Various methods reported for purification of microbial l-asparaginase have been listed in Table 3. A combination of downstream unit processes is extensively used for purification of l-asparaginase, which includes nonchromatographic processes or low-resolution purification steps that include centrifugation, fractional precipitation, aqueous biphasic systems, membrane-based purification, and chromatographic processes or high-resolution purification steps that include ion-exchange, gel filtration, and affinity chromatography [35,71,120,121].…”

Section: Downstream Processingmentioning

confidence: 99%

“…potassium phosphate and PEG are mixed at specific concentrations in a solution. PEG-phosphate is the most commonly used ATPS for protein separations[121]. Santos et al used a polymer salt based ATPS separation after ammonium sulfate precipitation for purification of periplasmic l-asparaginase from E. coli.…”

mentioning

confidence: 99%

A comprehensive review on microbial l‐asparaginase: Bioprocessing, characterization, and industrial applications

Chand

Mahajan

Prasad

et al. 2020

Biotech and App Biochem

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l‐Asparaginase (E.C.3.5.1.1.) is a vital enzyme that hydrolyzes l‐asparagine to l‐aspartic acid and ammonia. This property of l‐asparaginase inhibits the protein synthesis in cancer cells, making l‐asparaginase a mainstay of pediatric chemotherapy practices to treat acute lymphoblastic leukemia (ALL) patients. l‐Asparaginase is also recognized as one of the important food processing agent. The removal of asparagine by l‐asparaginase leads to the reduction of acrylamide formation in fried food items. l‐Asparaginase is produced by various organisms including animals, plants, and microorganisms, however, only microorganisms that produce a substantial amount of this enzyme are of commercial significance. The commercial l‐asparaginase for healthcare applications is chiefly derived from Escherichia coli and Erwinia chrysanthemi. A high rate of hypersensitivity and adverse reactions limits the long‐term clinical use of l‐asparaginase. Present review provides thorough information on microbial l‐asparaginase bioprocess optimization including submerged fermentation and solid‐state fermentation for l‐asparaginase production, downstream purification, its characterization, and issues related to the clinical application including toxicity and hypersensitivity. Here, we have highlighted the bioprocess techniques that can produce improved and economically viable yields of l‐asparaginase from promising microbial sources in the current scenario where there is an urgent need for alternate l‐asparaginase with less adverse effects.

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L-Asparaginase Purification

Cited by 23 publications

References 69 publications

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

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

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

A comprehensive review on microbial l‐asparaginase: Bioprocessing, characterization, and industrial applications

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