Protein engineered extracellular vesicles

A technology of mesenchymal stem cells and cells, applied in the field of extracellular vesicles, can solve the problems of lack of loading and bioactive delivery disclosure

Pending Publication Date: 2020-08-28
EVOX THERAPEUTICS LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In fact, while WO2016 / 044947 superficially describes the modification of exosomes to target lysosomal target cells by using fusion protein constructs containing (i) a lysosomal targeting sequence fused to (ii) an exosomal protein, Enzymosomes, the application conspicuously lacks disclosure related to the loading and bioactive delivery of actual therapeutic mRNA and / or protein-based formulations

Method used

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  • Protein engineered extracellular vesicles
  • Protein engineered extracellular vesicles
  • Protein engineered extracellular vesicles

Examples

Experimental program
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Effect test

example 1

[0073]Example 1. Genetic engineering of amniotic epithelial (AE) cells obtained from postpartum placenta with a polynucleotide construct encoding a polypeptide construct comprising NPC1 protein and EV-enriched protein (using lentiviral transduction) N-terminal portion of inhabitin (PNC1 S) (SEQ ID NO 2). AE-EVs produced from AE cells were harvested, purified, and assessed in vitro for their ability to enhance cholesterol transport in patient-derived fibroblasts. From figure 1 It can be seen that NPC1 AE-EV significantly reduces cholesterol levels in target cells by delivering large amounts of biologically active NPC1 protein to cells. Interestingly, when using only AE-EVs and MSC-EVs (and no other cell sources), it was possible to achieve a dose-dependent reduction in cholesterol accumulation by genetically engineering AE cells to express the NPC1 protein itself, in the absence of Any EV-enriched polypeptides (data not shown). Both EV-producing cells and their corresponding...

example 2

[0074] Example 2. Effect of Wharton's jelly MSC-derived EVs (WJ-MSC-EV, available from Hua obtained from Dun's gum) for genetic engineering. Cystine transporter-containing WJ-MSC-EVs (CYS-EVs) were harvested from hollow-fiber bioreactor cell cultures of WJ-MSCs, purified, and expressed in vitro on patient-derived dermal fibroblasts (PDSF) An assessment was performed to assess the reduction of cystine accumulation in an in vitro model of cystinosis. figure 2 It was shown that a significant reduction in cysteine ​​was seen upon application of WJ MSC EVs comprising a CD63-cystine transporter polypeptide construct. As shown in Example 1, WJ-MSCs and WJ-MSC-EVs were positive for Hsp70 proteins, especially Hsp70-8, and for tetraspanins CD63 and CD81.

example 3

[0075] Example 3. In vivo assessment of AEs positive for CD44, SSEA4, CD133, CD24, and Hsp70-8 and engineered to comprise multiple NPC1-inhabitin polypeptide constructs in a double knockout mouse model of Niemann-Pick disease -EV. The engineered AE-EVs showed significant therapeutic effects on both liver and CNS lesions ( image 3 ). Non-engineered AE-EVs also showed therapeutic efficacy, as inferred from the high content of various heat shock proteins in AE-EVs.

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[0077]

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PUM

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Abstract

The present invention relates to extracellular vesicles (EVs) as a novel therapeutic approach to lysosomal storage disorders (LSD). More specifically, the invention relates to the use of various protein engineering strategies for improving loading of hard-to-load LSD-related proteins and targeting of the resultant EVs to tissues and organs of interest.

Description

technical field [0001] The present invention relates to engineered extracellular vesicles (EVs) as a novel approach to the treatment of lysosomal storage disorders (LSDs). More specifically, the present invention relates to the use of various protein engineering strategies to improve the loading of difficult-to-load LSD-associated proteins and the targeting of the resulting EVs to tissues and organs of interest. Background technique [0002] Lysosomal storage diseases are an important subgroup of inborn errors of metabolism. It is estimated that 1 in every 5,000 live births has LSD, however, due to undiagnosed and misdiagnosed cases, this number is thought to be much higher. All LSDs are caused by the accumulation of specific macromolecular or monomeric compounds within the organelles of the endosome-autophagy-lysosome system. In addition to errors in stromal storage, LSD is often associated with neurological disorders and psychiatric abnormalities, accounting for a large ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K35/50A61K35/28A61K9/50A61P3/00C12N5/073C12N5/0775A61K31/727A61K31/4545A61K31/724A61K31/445A61K31/7016
CPCA61K9/5068A61K35/50C12N5/0605C07K2319/01A61P3/00A61K35/28A61K31/445A61K31/4545A61K31/70A61K31/7016A61K31/724A61K31/727C07K14/47C07K14/79C07K14/70596C07K2319/00A61K2300/00C12N2509/10
Inventor J.黑安P.伦丁A.乔根斯
Owner EVOX THERAPEUTICS LTD
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