Polymer suitable for use in cell culture

Patent metadata

Field	Value
Publication number	US-2016137977-A1
Application number	US-201414905579-A
Country	US
Kind code	A1
Filing date	Jul 16, 2014
Priority date	Jul 18, 2013
Publication date	May 19, 2016
Grant date	—

Abstract

A process for making an oligo(alkylene glycol) functionalized co-polyisocyanopeptide, wherein the process includes the steps of: i) copolymerizing a first comonomer of an oligo(alkylene glycol) functionalized isocyanopeptide grafted with a linking group and a second comonomer of a non-grafted oligo(alkylene glycol) functionalized isocyanopeptide, wherein the molar ratio between the first comonomer and the second comonomer is 1:500 and 1:30 and ii) adding a reactant of a spacer unit and a cell adhesion factor to the copolymer obtained by step i), wherein the spacer unit is represented by general formula A-L-B, wherein the linking group and group A are chosen to react and form a first coupling and the cell adhesion factor and group B are chosen to react and form a second coupling, wherein the first coupling and the second coupling are independently selected from the group consisting of alkyne-azide coupling, dibenzocyclooctyne-azide coupling, oxanorbornmadiene-based-azide couplings, vinylsulphone-thiol coupling, maleimide-thiol coupling, methyl methacrylate-thiol coupling, ether coupling, thioether coupling, biotin-strepavidin coupling, amine-carboxylic acid resulting in amides linkages, alcohol-carboxylic acid coupling resulting in esters linkages and NHS-Ester (N-Hydroxysuccinimide ester)-amine coupling and wherein group L is a linear chain segment having 10-60 bonds between atoms selected from C, N, O and S in the main chain.

First claim

What is claimed is:

1 . A process for making an oligo(alkylene glycol) functionalized co-polyisocyanopeptide, wherein the process comprises the steps of:
i) copolymerizing a first comonomer of an oligo(alkylene glycol) functionalized isocyanopeptide grafted with a linking group and a second comonomer of a non-grafted oligo(alkylene glycol) functionalized isocyanopeptide, wherein a molar ratio between the first comonomer and the second comonomer is 1:500 and 1:30 and ii) adding a reactant of a spacer unit and a cell adhesion factor to the copolymer obtained by step i), wherein the spacer unit is represented by general formula A-L-B, wherein the linking group and group A are chosen to react and form a first coupling and the cell adhesion factor and group B are chosen to react and form a second coupling, wherein the first coupling and the second coupling are independently an alkyne-azide coupling, dibenzocyclooctyne-azide coupling, oxanorbornadiene-based-azide couplings, vinylsulphone-thiol coupling, maleimide-thiol coupling, methyl methacrylate-thiol coupling, ether coupling, thioether coupling, biotin-strepavidin coupling, amine-carboxylic acid resulting in amides linkages, alcohol-carboxylic acid coupling resulting in esters linkages or NHS-Ester (N-Hydroxysuccinimide ester)-amine coupling and wherein group L is a linear chain segment having 10-60 bonds between atoms selected from C, N, O and S in the main chain.

2 . The process according to claim 1 , wherein group L is chosen from:

where p is 1 to 10, preferably 2 to 5,

where q is 1 to 9, preferably 2 to 5,

where r is 1 to 10, preferably 2 to 5.

3 . The process according to claim 1 , wherein the first coupling is the alkyne-azide coupling.

4 . The process according to claim 1 , wherein the second coupling is the NHS-ester (N-Hydroxysuccinimide ester)-amine coupling or the maleimide-amine coupling.

5 . The process according to claim 1 , wherein group A is represented by formula (VII):

wherein:
n is 0 to 8;
R 3 is [(L) p -Q], hydrogen, halogen, C 1 -C 24 alkyl groups, C 6 -C 24 (hetero)aryl groups, C 7 -C 24 alkyl(hetero)aryl groups or C 7 -C 24 (hetero)arylalkyl groups, the alkyl groups optionally being interrupted by one of more hetero-atoms selected from the group consisting of O, N and S, wherein the alkyl groups, (hetero)aryl groups, alkyl(hetero)aryl groups and (hetero)arylalkyl groups are independently optionally substituted with one or more substituents independently selected from C 1 -C 12 alkyl groups, C 2 -C 12 alkenyl groups, C 2 -C 12 alkynyl groups, C 3 -C 12 cycloalkyl groups, C 1 -C 12 alkoxy groups, C 2 -C 12 alkenyloxy groups, C 2 -C 12 alkynyloxy groups, C 3 -C 12 cycloalkyloxy groups, halogens, amino groups, oxo groups and silyl groups, wherein the alkyl groups, alkenyl groups, alkynyl groups, cycloalkyl groups, alkoxy groups, alkenyloxy groups, alkynyloxy groups and cycloalkyloxy groups are optionally substituted, the alkyl groups, the alkoxy groups, the cycloalkyl groups and the cycloalkoxy groups being optionally interrupted by one of more hetero-atoms selected from O, N and S, wherein the silyl groups are represented by the formula (R 4 ) 3 Si—, wherein R 4 is independently selected from C 1 -C 12 alkyl groups, C 2 -C 12 alkenyl groups, C 2 -C 12 alkynyl groups, C 3 -C 12 cycloalkyl groups, C 1 -C 12 alkoxy groups, C 2 -C 12 alkenyloxy groups, C 2 -C 12 alkynyloxy groups and C 3 -C 12 cycloalkyloxy groups, wherein the alkyl groups, alkenyl groups, alkynyl groups, cycloalkyl groups, alkoxy groups, alkenyloxy groups, alkynyloxy groups and cycloalkyloxy groups are optionally substituted, the alkyl groups, the alkoxy groups, the cycloalkyl groups and the cycloalkoxy groups being optionally interrupted by one of more hetero-atoms selected from O, N and S;
R 1 is independently selected from hydrogen, C 1 -C 24 alkyl groups, C 6 -C 24 (hetero)aryl groups, C 7 -C 24 alkyl(hetero)aryl groups and C 7 -C 24 (hetero)arylalkyl groups; and
R 2 is independently selected from halogen, —OR 6 , —NO 2 , —CN, —S(O) 2 R 6 , C 1 -C 12 alkyl groups, C 1 -C 12 aryl groups, C 1 -C 12 alkylaryl groups and C 1 -C 12 arylalkyl groups, wherein R 6 is as defined above, and wherein the alkyl groups, aryl groups, alkylaryl groups and arylalkyl groups are optionally substituted.

6 . The process according to claim 5 , wherein the spacer unit is represented by formula (IX):

wherein R 1 , R 2 , R 3 and n are as defined above and
L is selected from the group represented by formula (X):

wherein p is 2 to 5.

7 . The process according to claim 1 , wherein the spacer unit is represented by formula (XI):

wherein p is 2 to 5.

8 . The process according to claim 1 , wherein the cell adhesion factor is chosen from a sequence of amino acids.

9 . The process according to claim 1 , wherein the average of the number of the alkylene glycol units on the first comonomer and the second comonomer is at least 3 and at most 4.

10 . The oligo(alkylene glycol) functionalized co-polyisocyanopeptide obtainable by the process according to claim 1 .

11 . A cell culture comprising a hydrogel, comprising the co-polyisocyanopeptide of claim 10 at a concentration of 1.2-3.0 mg/mL and wherein the co-polyisocyanopeptide has a gelation temperature of 18 to 40° C.

12 . The cell culture according to claim 11 , wherein the cell culture comprises at least one of endothelial cells and smooth muscle cells.

13 . A process for culturing cells, comprising the steps of:
a) providing the cell culture according to claim 11 ; b) adding cells to the cell culture at a temperature below the gelation temperature of the hydrogel; and c) culturing the cells.

14 . The process according to claim 13 , wherein the cells are endothelial cells and muscle cells.

15 . A method of making a prevascular system, comprising the steps of: obtaining the cell culture according to claim 12 and making a prevascular system therewith.

16 . The process according to claim 2 , wherein the first coupling is alkyne-azide coupling.

17 . The process according to claim 16 , wherein the second coupling is the NHS-ester (N-Hydroxysuccinimide ester)-amine coupling or the maleimide-amine coupling, wherein group A is represented by formula (VII):

wherein:
n is 0 to 8;
R 3 is [(L) p -Q], hydrogen, halogen, C 1 -C 24 alkyl groups, C 6 -C 24 (hetero)aryl groups, C 7 -C 24 alkyl(hetero)aryl groups or C 7 -C 24 (hetero)arylalkyl groups, the alkyl groups optionally being interrupted by one of more hetero-atoms selected from the group consisting of O, N and S, wherein the alkyl groups, (hetero)aryl groups,

Assignees

• Stichting Katholieke Univ

Inventors

• Rowan Alan Edward
• Eksteen Akeroyd Zaskia Hillet
• Wilson Christopher
• Geutjes Petrus Johannes
• Feitz Wouter F J
• Oosterwijk Egbert

Classifications

• C12N2502/28

  Vascular endothelial cells · CPC title

• C08G69/10

  Alpha-amino-carboxylic acids {(polysuccinimides C08G73/1092)} · CPC title

• C08G69/40

  Polyamides containing oxygen in the form of ether groups (C08G69/12, C08G69/32 take precedence) · CPC title

• C12N5/0691

  Vascular smooth muscle cells; 3D culture thereof, e.g. models of blood vessels · CPC title

• C08G65/33324

  acyclic · CPC title