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Production of Therapeutic Enzymes by Lentivirus Transgenesis

  • María Celeste Rodríguez
  • Natalia Ceaglio
  • Sebastián Antuña
  • María Belén Tardivo
  • Marina Etcheverrigaray
  • Claudio PrietoEmail author
Chapter
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 1148)

Abstract

Since ERT for several LSDs treatment has emerged at the beginning of the 1980s with Orphan Drug approval, patients’ expectancy and life quality have been improved. Most LSDs treatment are based on the replaced of mutated or deficient protein with the natural or recombinant protein.

One of the main ERT drawback is the high drug prices. Therefore, different strategies trying to optimize the global ERT biotherapeutic production have been proposed. LVs, a gene delivery tool, can be proposed as an alternative method to generate stable cell lines in manufacturing of recombinant proteins. Since LVs have been used in human gene therapy, clinical trials, safety testing assays and procedures have been developed. Moreover, one of the main advantages of LVs strategy to obtain manufacturing cell line is the short period required as well as the high protein levels achieved.

In this chapter, we will focus on LVs as a recombinant protein production platform and we will present a case study that employs LVs to express in a manufacturing cell line, alpha-Galactosidase A (rhαGAL), which is used as ERT for Fabry disease treatment.

Keywords

ERT LSDs Fabry disease Lentiviral Vectors (LVs) rhαGAL 

Abbreviations

E. coli

Escherichia coli

ER

endothelial reticulum

PEG

polyethylene glycol

FDA

Food and Drug Administration

LSD

lysosomal storage disease

MPS

mucopolysaccaridosis

ERT

enzyme replacement therapy

LVs

lentiviral vectors

CHO

Chinese Hamster Ovary

Neu5Gc

N-glycolylneuraminic acid

BHK-21

baby hamster kidney cells

HEK293

Human Embryonic Kidney 293

EMA

European Medicines Agency

CAP

CEVEC´s Amniocyte Production

ICH

International Conference of Harmonization

TGE

transient gene expression

DHFR

dihydrofolate reductase

GS

glutamine synthetase

MTX

methotrexate

MSX

methionine sulfoximine

HPRT

hypoxanthine phosphoribosyl transferase

RMCE

recombinase-mediated cassette exchange

FRT

flippase recognition target sites

ZFNs

zinc finger nucleases

TALENs

transcription activator-like effector nucleases

NHEJ

non-homologous end-joining

HDR

homology-directed repair

CRISPR

clustered regularly interspaced short palindromic repeats

SpCas9

Streptococcus pyogenes Cas9 endonuclease

gRNA

guide RNA

trcRNA

trans-acting antisense RNA

PAM

protospacer adjacent motif

S/MARs

matrix attachment regions

UCOEs

Ubiquitous Chromatin Opening Elements

PEI

Polyethylenimine

RT

reverse transcriptase

IN

integrase

TG

transgene

VSV

Vesicular Stomatitis Virus

LTR

long terminal repeats

SIN

self-inactivating

RRE

rev response elements

cPPT

Central Polypurine tract

WPRE

post-transcriptional regulatory element of the woodchuck

qPCR

real time polymerase chain reaction

MOI

multiplicity of infection

CAR

chimeric antigen receptor

RCL

replication-competent lentivirus

ELISA

enzyme-linked immunosorbent assay

PERT

product-enhanced reverse transcriptase

LOD

limit of detection

M6P

phosphate-6-O-mannose

GlcNAc

N-acetylglucosamine

M6PR

M6P receptor

C6S

chondroitin-6-sulfate

KS

keratan sulfate

GALNS

N-acetylgalactosamine-6-sulfate sulfatase

LAL

Lysosomal acid lipase

Gb3

globotriaosylceramide

rhαGAL

recombinant human alpha galactosidase A

LP

lentiviral particle

IEX

ionic exchange

HIC

Hydrophobic interaction chromatography

RP-HPLC

reversed phase high performance liquid chromatography

IEF

isoelectrofocusing

HPAEC-PAD

high-pH anion-exchange chromatography with pulsed amperometric detection

WAX

weak anion exchange

Man

Mannose

Gal

Galactose

GlcNAc

N-acetylglucosamine

Fuc

Fucose

4MU-α-Gal

4-Methylumbelliferyl α-D-galactopyranoside

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Copyright information

© Springer Nature Singapore Pte Ltd. 2019

Authors and Affiliations

  • María Celeste Rodríguez
    • 1
  • Natalia Ceaglio
    • 1
  • Sebastián Antuña
    • 2
  • María Belén Tardivo
    • 2
  • Marina Etcheverrigaray
    • 1
  • Claudio Prieto
    • 3
    Email author
  1. 1.Cell Culture LaboratoryUNL, CONICET, FBCBSanta FeArgentina
  2. 2.Zelltek S.A.Santa FeArgentina
  3. 3.Cell Culture LaboratoryUNL, FBCBSanta FeArgentina

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