Engineered tissues for in vitro research uses, arrays thereof, and methods of making the same

a technology of in vitro research and tissue, applied in the field of in vitro research and development cost of a new pharmaceutical compound, can solve the problems of low rate of new therapeutic discovery, unsustainable r&d cost, and long drug discovery process, so as to increase the number and quality of innovative, cost-effective

Inactive Publication Date: 2013-07-25
ORGANOVO
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The research and development cost of a new pharmaceutical compound is approximately $1.8 billion.
Despite advances in technology and understanding of biological systems, drug discovery is still a lengthy, expensive, and inefficient process with low rate of new therapeutic discovery.

Method used

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  • Engineered tissues for in vitro research uses, arrays thereof, and methods of making the same
  • Engineered tissues for in vitro research uses, arrays thereof, and methods of making the same
  • Engineered tissues for in vitro research uses, arrays thereof, and methods of making the same

Examples

Experimental program
Comparison scheme
Effect test

example 1

Cell Culture

[0188]Smooth Muscle Cells

[0189]Primary human aortic smooth muscle cells (HASMC; GIBCO / Invitrogen Corp., Carlsbad, Calif.) were maintained and expanded in low glucose dulbecco's modified eagle medium (DMEM; Invitrogen Corp., Carlsbad, Calif.) supplemented with 10% fetal bovine serum (FBS), 100 U / mL Penicillin, 0.1 mg / mL streptomycin, 0.25 μg / mL of amphotericin B, 0.01M of HEPES (all from Invitrogen Corp., Carlsbad, Calif.), 50 mg / L of proline, 50 mg / L of glycine, 20 mg / L of alanine, 50 mg / L of ascorbic acid, and 3 μg / L of CuSO4 (all from Sigma, St. Louis, Mo.) at 37° C. and 5% CO2. Confluent cultures of HASMC between passage 4 and 8 were used in all studies.

[0190]Endothelial Cells

[0191]Primary human aortic endothelial cells (HAEC; GIBCO / Invitrogen Corp., Carlsbad, Calif.) were maintained and expanded in Medium 199 (Invitrogen Corp., Carlsbad, Calif.) supplemented with 10% FBS, 1 μg / mL of hydrocortisone, 10 ng / mL of human epidermal growth factor, 3 ng / mL of basic fibroblas...

example 2

NovoGel™ Solutions and Mold

[0200]Preparation of 2% and 4% (w / v) NovoGel™ Solution

[0201]1 g or 2 g (for 2% or 4% respectively) of NovoGel™ (Organovo, San Diego, Calif.) was dissolved in 50 mL of Dulbecco's phosphate buffered saline (DPBS; Invitrogen Corp., Carlsbad, Calif.). Briefly, the DPBS and NovoGel™ are heated to 85° C. on a hot plate with constant stirring until the NovoGel™ dissolves completely. NovoGel™ solution is sterilized by steam sterilization at 125° C. for 25 minutes. The NovoGel™ solution remains in liquid phase as long as the temperature is maintained above 65.5° C. Below this temperature a phase transition occurs, the viscosity of the NovoGel™ solution increases and the NovoGel™ forms a solid gel.

[0202]Preparation of NovoGel™ Mold

[0203]An NovoGel™ mold was fabricated for the incubation of cylindrical bio-ink using a Teflon® mold that fit a 10 cm Petri dish. Briefly, the Teflon® mold was pre-sterilized using 70% ethanol solution and subjecting the mold to UV light f...

example 3

Fabrication of HASMC-HAEC Polytypic Cylindrical Bio-Ink

[0204]To prepare polytypic cylindrical bio-ink, HASMC and HAEC were individually collected and then mixed at pre-determined ratios. Briefly, the culture medium was removed from confluent culture flasks and the cells were washed with DPBS (1 ml / 5 cm2 of growth area). Cells were detached from the surface of the culture flasks by incubation in the presence of trypsin (1 ml / 15 cm2 of growth area; Invitrogen Corp., Carlsbad, Calif.) for 10 minutes. HASMC were detached using 0.15% trypsin while HAEC were detached using 0.1% trypsin. Following the incubation appropriate culture medium was added to the flasks (2× volume with respect to trypsin volume). The cell suspension was centrifuged at 200 g for 6 minutes followed by complete removal of supernatant solution. Cell pellets were resuspended in respective culture medium and counted using a hemocytometer. Appropriate volumes of HASMC and HAEC were combined to yield a polytypic cell susp...

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Abstract

Disclosed are living, three-dimensional tissue constructs for in vitro scientific and medical research, arrays thereof, and methods of making said tissues and arrays.

Description

CROSS REFERENCE TO RELATED APPLICATIONS[0001]This application claims the benefit of U.S. Application Ser. No. 61 / 533,757, filed Sep. 12, 2011, U.S. Application Ser. No. 61 / 533,753, filed Sep. 12, 2011, and U.S. Application Ser. No. 61 / 533,761, filed Sep. 12, 2011, all of which are hereby incorporated by reference in their entirety.BACKGROUND OF THE INVENTION[0002]The research and development cost of a new pharmaceutical compound is approximately $1.8 billion. See Paul, et al. (2010). How to improve R&D productivity: the pharmaceutical industry's grand challenge. Nature Reviews Drug Discovery 9(3):203-214. Drug discovery is the process by which drugs are discovered and / or designed. The process of drug discovery generally involves at least the steps of: identification of candidates, synthesis, characterization, screening, and assays for therapeutic efficacy. Despite advances in technology and understanding of biological systems, drug discovery is still a lengthy, expensive, and ineffi...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): G01N33/50
CPCC12N5/0697A61L27/38C12N2502/27G01N33/5082C12N5/0691G01N33/5088A61L27/34A61L27/3891A61L27/50C12N2502/28C12N2513/00
Inventor MURPHY, KEITHKHATIWALA, CHIRAGDORFMAN, SCOTTSHEPHERD, BENJAMINPRESNELL, SHARONROBBINS, JUSTIN
Owner ORGANOVO
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