<i>In vivo</i> stabilization of a less toxic asparaginase variant leads to a durable antitumor response in acute leukemia

Abstract

Asparagine is a non-essential amino acid since it can either be taken up via the diet or synthesized by asparagine synthetase (ASNS). Acute lymphoblastic leukemia (ALL) cells do not or minimally express ASNS which makes them completely dependent on extracellular asparagine for their growth and survival. This dependency makes ALL cells vulnerable to treatment with L-asparaginase, an enzyme that hydrolyzes asparagine. To date, all clinically approved L-asparaginases have a significant L-glutaminase co-activity, associated with non-immune related toxic side effects observed during therapy. Therefore, reduction of L-glutaminase co-activity with concomitant maintenance of its anti-cancer L-asparaginase effect may effectively improve the tolerability of this unique drug. Previously, we designed a new alternative variant of Erwinia chrysanthemi (ErA; Erwinaze) with decreased L-glutaminase co-activity, while maintaining its L-asparaginase activity, by the introduction of three key mutations around the active site (ErA-TM). However, Erwinaze and our ErA-TM variant have a very short half-life in vivo. Here, we show that the fusion of ErA-TM with an albumin binding domain (ABD)-tag significantly increases its in vivo persistence. In addition, we evaluated the in vivo therapeutic efficacy of ABD-ErA-TM in a B-ALL xenograft model of SUP-B15. Our results show a comparable long lasting durable anti-leukemic effect between the standard-of-care PEG-asparaginase and ABD-ErA-TM L-asparaginase, but with fewer co-glutaminase related acute side effects. Since the toxic side effects of current L-asparaginases often result in treatment discontinuation in ALL patients, this novel ErA-TM variant with ultra-low L-glutaminase co-activity and long in vivo persistence may have great clinical potential.

Substrate Affinity Is Not Crucial for Therapeutic L-Asparaginases: Antileukemic Activity of Novel Bacterial Enzymes

“…The actions of any enzyme should also be tested in vivo to verify its stability, immunogenicity, and antineoplastic properties. Currently, reports from studies in animal models are available only for the commercial enzymes EcAII or ErAII [69][70][71], and there are no data for Nth-hydrolases (class 2) or Rhizobium etli-type (class 3) proteins.…”

Back to the future: the amazing journey of the therapeutic anti-leukemia enzyme asparaginase <i>Erwinia chrysanthemi</i>

“…Despite the approval of the Erwinia c. ASNase as a second-line ASNase since 2011, important difficulties in the manufacturing process of Erwinia c. ASNase have occurred over the last two decades and this has led to global supply shortages of Erwinia c. ASNase. 52 , 53 For this reason, a recombinant Erwinia c. ASNase (namely JZP-458) utilizing a novel Pseudomonas fluorescent technology expression platform has recently been developed; 52 it has the same amino acid sequence as Erwinia c. ASNase and results in no immunologic cross-reactivity to E. coli -derived ASNase products. The recombinant technology-based production allows a stable and enhanced production process thus avoiding the supply problems with the non-recombinant Erwinia c. ASNase.…”

Acute Pancreatitis in Children with Acute Lymphoblastic Leukemia Using L-Asparaginase: A Review of the Literature

“…Normal cells may manufacture L-asparagine for growth using the transaminase enzyme, which converts oxaloacetate into the intermediate aspartate, which then transfers an amino group from glutamate to oxaloacetate, producing ketoglutarate and aspartate. Finally, the enzyme asparagine synthetase transforms aspartate to asparagine in healthy cells [7].…”