Mucin immobilized chromatography
AssigneeVertex Pharmaceuticals Incorporated
InventorsIrina Kadiyala, Riccardo Panicucci
Life of this patent
Estimating the term…
Abstract
An in vitro high-throughput screening method that models the absorption of drugs across the epithelial mucosa is provided. The invention discloses an in vitro mucin immobilized chromatography model to estimate drug permeability coefficients. The invention describes compositions for mucin immobilized chromatography media and methods to prepare this media. The invention also discloses a method to estimate in vitro drug permeability coefficients using mucin immobilized chromatography media. The invention also discloses methods to determine absorption processes in the digestive system and discloses methods of use for mucin chromatography media in column, batch, assay, diagnostic, or high-throughput screening analyses.
Claims77
A composition comprising a mucin-type protein non-covalently immobilized to a solid support matrix, wherein said solid support matrix is surface modified with an amphiphilic molecule.
A composition comprising a mucin-type peptide non-covalently immobilized to a solid support matrix, wherein said matrix is surface modified with an amphiphilic molecule.
A method for preparing a composition comprising a mucin-type protein or a mucin-type peptide non-covalently immobilized to a solid support matrix, wherein said matrix is surface modified with an amphiphilic molecule, comprising the step of:
- mixing a solution comprising a mucin-type protein and/or mucin-type peptide with a solid support matrix that is surface modified with an amphiphilic molecule under conditions in which the mucin-type protein and/or mucin-type peptide is immobilized to the matrix.
The method according to 45, wherein the solvent is polar.
The method according to 46, wherein the polar solvent is acetone or DPBS/Brij-35.
A method for separating one or more components of a mixture, comprising the steps of contacting the mixture with a chromatography medium comprising a mucin-type protein, a mucin-type peptide, or a combination of thereof, wherein said mucin-type protein, mucin-type peptide or combination thereof is immobilized to a solid support matrix, wherein said solid support matrix is surface modified with an amphiphilic molecule, under conditions in which one or more components of the mixture binds to the composition;
- and eluting the chromatography medium under conditions in which one or more components are separated from each other.
A method to estimate the permeability coefficient of a drug comprising the steps of:
- a) providing a chromatography medium, wherein said medium comprises a mucin-type protein, mucin-type peptide, or a combination of thereof immobilized to a solid support matrix, wherein solid support matrix is surface modified with an amphiphilic molecule;
- b) contacting a drug with said chromatography medium;
- c) measuring the degree of binding of the drug on the chromatography medium;
- and
- d) estimating the drug permeability coefficient.
A method to estimate the permeability coefficient of a drug comprising the steps of:
- a) providing a chromatography medium, wherein said medium comprises a mucin-type protein, mucin-type peptide, or a combination of thereof immobilized to a solid support matrix, wherein solid support matrix is surface modified with an amphiphilic molecule;
- b) contacting a drug with said chromatography medium;
- c) recording a retention time of said drug;
- d) determining a membrane diffusion coefficient;
- e) determining a membrane equilibrium constant;
- f) determining a membrane thickness;
- and
- g) calculating an effective drug permeability.
A method of comparing the relative permeability of one or more drugs comprising the steps of:
- a) providing a chromatography medium, wherein said medium comprises a mucin-type protein, mucin-type peptide, or a combination of thereof immobilized to a solid support matrix, wherein solid support matrix is surface modified with an amphiphilic molecule;
- b) contacting a drug with said chromatography medium;
- c) measuring the degree of binding of the drug on the chromatography medium;
- and
- d) comparing the degree of binding to that measured for one or more other drugs.
A method of estimating drug permeability in two or more physiological states comprising the steps of:
- a) providing a first chromatography medium, wherein said medium comprises a mucin-type protein, mucin-type peptide, or a combination of thereof immobilized to a solid support matrix, wherein solid support matrix is surface modified with an amphiphilic molecule and wherein said first chromatography medium mimics the mucus layer of a first physiological state;
- b) providing a second chromatography medium, wherein said medium comprises a mucin-type protein, mucin-type peptide, or a combination of thereof immobilized to a solid support matrix, wherein solid support matrix is surface modified with an amphiphilic molecule and wherein said second chromatography medium mimics the mucus layer of a second physiological state;
- c) contacting a drug with said first and said second chromatography media;
- d) measuring the degree of binding of the drug on the first and second chromatography media;
- and
- e) comparing the degree of binding on said first chromatography medium to that measured for said second chromatography medium.
A method to emulate an absorption process in a system comprising more than one epithelial mucosal layer, comprising the steps of:
- a) providing a series of chromatography media, wherein said media comprises a mucin-type protein, mucin-type-peptide, or a combination of thereof immobilized to a solid support matrix, wherein solid support matrix is surface modified with an amphiphilic molecule;
- wherein each of said media comprises a different amount and/or type of a mucin-type protein, mucin-type peptide or combination thereof, and wherein each of said media mimic the different mucus linings of the system;
- b) contacting a drug with each of said chromatography media;
- c) measuring the degree of binding of the drug on each of said chromatography media;
- and
- d) comparing the degree of binding to that measured for one or more other drugs.
A kit comprising mucin immobilized chromatography media and a standard.
Dependencies are inferred from the claim text.