Antibody-mediated Hsp70 protein therapy.
Intracellular Hsp70 provides cytoprotection against a variety of stressful stimuli, and an effective means of increasing intracellular Hsp70 levels could prove beneficial in the prevention and treatment of a variety of human diseases. A novel protein transduction domain consisting of the single chain Fv fragment of an anti-DNA antibody known to penetrate into living cells and tissues, mAb 3E10, has recently been used to deliver functional proteins to cells. The ability of the single chain Fv fragment to deliver Hsp70 into living cells was tested by generating an Fv-Hsp70 fusion protein. Fv-Hsp70 was produced as a secreted protein in both COS-7 cells and the methylotropic yeast strain Pichia pastoris and was shown capable of penetrating into COS-7 cells and primary rat cortical neurons. Pre-treatment with Fv-Hsp70 protected both COS-7 cells and primary rat cortical neurons against subsequent exposure to hydrogen peroxide. These results provide the first evidence that the Fv fragment of mAb 3E10 is capable of delivering proteins to neurons and indicate its potential in the development of Hsp70 protein therapy.
Hansen
James E
JE
Department of Medicine, San Fernando Valley Program, University of California-Los Angeles, Los Angeles, CA 90095, USA. jameshansen2004@hotmail.com
Sohn
William
W
Kim
Charles
C
Chang
Sophia S
SS
Huang
Natalie C
NC
Santos
Donaldson G
DG
Chan
Grace
G
Weisbart
Richard H
RH
Nishimura
Robert N
RN
D000818:Animals; D000831:Animals, Newborn; D015153:Blotting, Western / Q000379:methods; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology; D002522:Chlorocebus aethiops; D004305:Dose-Response Relationship, Drug; D018840:HSP70 Heat-Shock Proteins / Q000096:biosynthesis / Q000494:pharmacology / Q000502:physiology*; D006801:Humans; D006861:Hydrogen Peroxide / Q000633:toxicity; D007135:Immunoglobulin Variable Region / Q000276:immunology / Q000502:physiology*; D009474:Neurons / Q000187:drug effects / Q000378:metabolism*; D016877:Oxidants / Q000633:toxicity; D021381:Protein Transport / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000502:physiology*; D014161:Transduction, Genetic / Q000379:methods; D014162:Transfection / Q000379:methods
D018840:HSP70 Heat-Shock Proteins; D007135:Immunoglobulin Variable Region; D016877:Oxidants; D011993:Recombinant Fusion Proteins; D006861:Hydrogen Peroxide