Abstract: The present invention relates to a novel cell based sensor useful for drug discovery that comprises a cell line with professional regulated exocytosis of secretory granules transfected with a protease as a reporter polypeptide stored in the regulated secretory granules of the cell line with professional regulated exocytosis and having either an endogenous or a heterologous molecule as a modulator of regulated secretory granules exocytosis, such said granule stored protease reporter having at least: a high resistance to conditions already present inside the granules such as low pH and proteolysis by other proteases; enzymatic activity after exocytosis; a highly specific cleavage sequence; a very low level of secretion under unstimulated or basal conditions; and a high signal to background activity in a media compatible with cell culture viability and granule exocytosis for a high throughput robust and sensitive detection.
Type:
Grant
Filed:
July 28, 2010
Date of Patent:
May 24, 2016
Assignee:
CANVAX BIOTECH SL
Inventors:
Elier Paz-Rojas, Maria de Gracia Montero-Peñalvo, Veronica Inmaculada Luna-Guerrero, Fe Isabel Garcia Maceira, Jose Andres Morales-Martínez, Tania Garcia Maceira, Ana Belén Aragón-Gómez, Ana Quesada-Molina, Aurora Maria Marquez-Morales
Abstract: The present invention relates to a novel cell based sensor useful for drug discovery that comprises a cell line with professional regulated exocytosis of secretory granules transfected with a protease as a reporter polypeptide stored in the regulated secretory granules of the cell line with professional regulated exocytosis and having either an endogenous or a heterologous molecule as a modulator of regulated secretory granules exocytosis, such said granule stored protease reporter having at least: a high resistance to conditions already present inside the granules such as low pH and proteolysis by other proteases; enzymatic activity after exocytosis; a highly specific cleavage sequence; a very low level of secretion under unstimulated or basal conditions; and a high signal to background activity in a media compatible with cell culture viability and granule exocytosis for a high throughput robust and sensitive detection.
Type:
Application
Filed:
July 28, 2010
Publication date:
August 8, 2013
Applicant:
CANVAX BIOTECH SL
Inventors:
Elier Paz-Rojas, Maria de Gracia Montero-Peñalvo, Veronica Inmaculada Luna-Guerrero, Fe Isabel Garcia Maceira, Jose Andres Morales-Martínez, Tania Garcia Maceira, Ana Belén Aragón-Gómez, Ana Quesada-Molina