Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Truncated guinea pig l-asparaginase variants and methods of use

A technology of asparaginase and variants, applied in chemical instruments and methods, biochemical equipment and methods, medical preparations containing active ingredients, etc., can solve the problem of antibody inactivation and clearance, restricted use, reduced effectiveness, etc. question

Pending Publication Date: 2020-04-21
THE BOARD OF TRUSTEES OF THE UNIV OF ILLINOIS +1
View PDF28 Cites 2 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In fact, the toxic side effects of L-asparaginase treatment severely limit the use of this anticancer drug
[0004] Another disadvantage of using bacterial enzymes as therapeutic agents is their immunogenicity, which can lead to an immediate threat to the patient due to hypersensitivity reactions up to anaphylactic shock
In addition, antibodies produced can inactivate and clear the enzyme drug, reducing or even eliminating its effectiveness

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Truncated guinea pig l-asparaginase variants and methods of use
  • Truncated guinea pig l-asparaginase variants and methods of use
  • Truncated guinea pig l-asparaginase variants and methods of use

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] Example 1: Production of active C-terminally truncated GpA

[0057] Mammalian L-asparaginase such as human L-asparaginase (hASNase1; UNIPROT accession number Q86U10; SEQ ID NO: 2) and guinea pig L-asparaginase (gpASNase1 or GpA; UNIPROT accession number HOWOT5; SEQ ID NO : 1) Contains two domains: an N-terminal domain of ~360 residues in which the L-asparaginase activity resides, and a C-terminal domain of ~200 residues whose function is unknown. For comparison, clinically relevant bacterial L-asparaginases from Escherichia coli and Erwinia chrysanthemi are approximately 350 amino acid residues in length and do not contain this C-terminal domain.

[0058] One of the challenges with injectable biotherapeutics comes from the short half-life, which leads to poor bioavailability. Clearance is mainly caused by proteolysis, renal filtration or neutralization by the immune system. Larger foreign sequence molecules have a greater chance of being detected by the immune system ...

Embodiment 2

[0064] Example 2: Humanized GpA variants produced by directed evolution

[0065]overview. Directed evolution, or the process of mimicking natural evolutionary processes in the laboratory, is widely used to improve enzyme performance. See eg Dalby (2011) Curr. :91-100; Wang & Zhao (2012) Bioresour.Technol.115:117-125. Thanks to advances in library generation, screening techniques, and most importantly a better understanding of the mechanisms of natural protein evolution, large increases in evolved catalytic activity (relative to the starting point, and based on absolute k cat / K m value) (Fasan et al., (2008) J. Mol. Biol. 383:1069-80; Bar-Even et al., (2011) Biochemistry (Mosc.) 50:4402-10).

[0066] To overcome immunogenicity, a human-like L-asparaginase with kinetic properties close to that of type II E. coli L-asparaginase was generated by directed evolution approach. High sequence identity to hASNase1 but low K of gpASNase1 was identified using DNA family shuffling m...

Embodiment 3

[0105] Example 3: Humanized GpA variants produced by a structured-based approach

[0106] As an alternative to DNA shuffling and domain swapping, a structure-based approach was taken to humanize and reduce the immunogenicity of the GpA enzyme. For this approach, a truncated GpA369 variant (SEQ ID NO: 5) was modified. The crystal structure of GpA was examined and predictions were made as to which residues at the surface of the enzyme could be mutated to the corresponding amino acids in hASNase1 and whose presence would not be detrimental to the activity or stability of the enzyme. The candidate surface residues were classified into three groups based on their likelihood of having a possible effect on the activity or stability of GpA (group 1, no effect; group 2, likely to have no effect; and group 3, with possible impact) (Table 6).

[0107] Table 6

[0108]

[0109]

[0110] 1 Reference sequence GpA369 (SEQ ID NO: 5).

[0111] 2 Corresponding residues in reference ...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

Variant guinea pig L-asparaginases which are truncated and humanized are described as are fusion proteins containing the L-asparaginase and use of the L- asparaginases in the treatment of cancers suchas acute lymphoblastic leukemia and acute myeloid leukemia.

Description

technical field [0001] This application claims the benefit of priority to U.S. Provisional Application No. 62 / 544,396, filed August 11, 2017, and U.S. Provisional Application No. 62 / 544,411, filed August 11, 2017, the contents of which are adopted in their entirety Incorporated herein by reference. [0002] This invention was made with Government support under Grant No. EB013685 awarded by the National Institutes of Health and Grant No. BX001919 awarded by the Department of Veteran Affairs. The US Government has certain rights in this invention. Background technique [0003] Certain cancers such as acute lymphoblastic leukemia (ALL) are dependent on the clearance of Asn from the blood, a factor most commonly attributed to the lack / low expression of asparagine synthetase in such cancers. Therefore, L-asparaginase has been identified as a key component in the treatment of these cancers. All commercially available L-asparaginases have dual activity. The main activity, L-asp...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Applications(China)
IPC IPC(8): A61K38/17C12N9/82
CPCC12Y305/01001C12N9/82C07K2319/00A61K38/00A61P35/02
Inventor 阿尔农·拉维海恩-安哈·源阿曼达·沙尔克
Owner THE BOARD OF TRUSTEES OF THE UNIV OF ILLINOIS
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com