SUBSTRATES CONTAINING POLYPHOSPHAZENE AS MATRICES AND SUBSTRATES CONTAINING POLYPHOSPHAZENE WITH A MICRO-STRUCTURED SURFACE

• US 20090117637A1
• Filed: 09/25/2008
• Published: 05/07/2009
• Est. Priority Date: 01/11/2001
• Status: Active Grant

• Current Assignee: Celonova Biosciences, Inc. (Boston Scientific Corporation)
• Original Assignee: Celonova Biosciences, Inc. (Boston Scientific Corporation)
• Inventors: Grunze, Michael, Fritz, Ulf
• Granted Patent: US 9,080,146 B2
• Time in Patent Office: Days
• US Class Current: 435/180
• CPC Class Codes: A61L 27/34 Macromolecular materials
• CPC Class Codes: A61L 27/3687 characterised by the use of chemical agents in the treatment, e.g. specific enzymes, detergents, capping agents, crosslinkers, anticalcification agents
• CPC Class Codes: A61L 27/3895 using specific culture conditions, e.g. stimulating differentiation of stem cells, pulsatile flow conditions
• CPC Class Codes: C08G 79/025 Polyphosphazenes
• CPC Class Codes: C08L 85/02 containing phosphorus
• CPC Class Codes: C09D 185/02 containing phosphorus
• CPC Class Codes: C12N 2533/30 Synthetic polymers thermoreactive polymers, e.g. PNIPAm, C12N2539/10
• CPC Class Codes: C12N 2535/10 Patterned coating
• CPC Class Codes: C12N 5/0068 General culture methods using substrates for specific animal cell type C12N5/06

Abstract

This disclosure relates to substrates containing at least one polyphosphazene with a forming surface as matrices for producing biological materials that can be implanted in a mammal. The disclosure also relates to a method for producing such substrates and substrates containing polyphosphazene with a micro-structured surface.

Claims

1. A method of regulating cellular adhesion at a substrate, the method comprising:

a) contacting at least a portion of a substrate with at least one adhesion promoter to provide a treated substrate;

b) contacting the treated substrate with a solution comprising a known concentration of at least one polyphosphazene;

c) removing the solvent from the polyphosphazene solution in contact with the treated substrate to provide a polyphosphazene micro-structured substrate; and

d) controlling the degree of cellular adhesion at the polyphosphazene micro-structured substrate by;

i) controlling the polyphosphazene film thickness or surface coverage of the polyphosphazene micro-structured substrate;

ii) controlling the size or aspect ratio of the structural elements of the polyphosphazene micro-structured substrate;

iii) controlling the spatial density of the structural elements of the polyphosphazene micro-structured substrate;

or

iv) any combination thereof.

2. A method of regulating cellular adhesion at a substrate according to claim 1, wherein the polyphosphazene has the formula

3. A method of regulating cellular adhesion at a substrate according to claim 2, wherein at least one of R¹ to R⁶ is an alkoxy group substituted with at least one fluorine atom.

4. A method of regulating cellular adhesion at a substrate according to claim 2, wherein at least one of R¹ to R⁶ is selected from OCH₃, OCF₃, OCH₂CH₃, OCH₂CF₃, OCH₂CH₂CH₃, OCH₂CH₂CF₃, OCH₂CF₂CF₃, OCH(CF₃)₂, OCCH₃(CF₃)₂, OCH₂CF₂CF₂CF₃, OCH₂(CF₂)₃CF₃, OCH₂(CF₂)₄CF₃, OCH₂(CF₂)₅CF₃, OCH₂(CF₂)₆CF₃, OCH₂(CF₂)₇CF₃, OCH₂CF₂CHF₂, OCH₂CF₂CF₂CHF₂, OCH₂(CF₂)₃CHF₂, OCH₂(CF₂)₄CHF₂, OCH₂(CF₂)₅CHF₂, OCH₂(CF₂)₆CHF₂, or OCH₂(CF₂)₇CHF₂.

5. A method of regulating cellular adhesion at a substrate according to claim 2, wherein the polyphosphazene is poly[bis(2,2,2-trifluoroethoxy)]phosphazene.

6. A method of regulating cellular adhesion at a substrate according to claim 2, wherein the polyphosphazene has a molecular weight of at least 70,000 g/mol.

7. A method of regulating cellular adhesion at a substrate according to claim 1, wherein step d) is effected by:

a) increasing the polyphosphazene concentration in the polyphosphazene solution used to contact the treated substrate;

b) repeating steps b) and c) of claim 1 any number of times;

or

c) decreasing the polyphosphazene concentration in the polyphosphazene solution used to contact the treated substrate.

8. A method of regulating cellular adhesion at a substrate according to claim 1, wherein the concentration of the polyphosphazene solution in step b) is up to about 200 mg/mL.

9. A method of regulating cellular adhesion at a substrate according to claim 1, wherein the polyphosphazene micro-structured substrate has a polyphosphazene film thickness of up to about 1000 nm.

10. A method of regulating cellular adhesion at a substrate according to claim 1, wherein the spacing density of the void structure within the polyphosphazene micro-structured substrate is from about 10,000 voids/100 μ

m² to 0 voids/100 μ

m².

11. A method of regulating cellular adhesion at a substrate according to claim 1, wherein the polyphosphazene surface coverage of the polyphosphazene micro-structured substrate is from about 50% to 100%.

12. A method of regulating cellular adhesion at a substrate according to claim 1, wherein the adhesion promoter is selected from:

13. A method of regulating cellular adhesion at a substrate according to claim 1, wherein the adhesion promoter is selected N-methyl-aza-2,2,4-trimethylsilacyclopentane;

2,2-dimethoxy-1,6-diaza-2-silacyclooctane;

(3-trimethoxysilylpropyl)diethylene tri amine;

(3-aminopropyl)trimethoxysilane (APTMS);

N-(3-(trimethoxysilyl)propyl)methanediamine;

N¹,N²-bis(3-(trimethoxysilyl)propyl)ethane-1,2-diamine;

1,3,5-tris(3-(trimethoxysilyl)propyl)-1,3,5-triazinane-2-4-6-trione;

or any combination thereof.

25. A method of regulating cellular adhesion at a substrate according to claim 24, wherein step e) is effected by:

a) increasing the polyphosphazene concentration in the polyphosphazene solution used to contact the treated substrate;

b) repeating steps b) and c) of claim 1 any number of times;

or

c) decreasing the polyphosphazene concentration in the polyphosphazene solution used to contact the treated substrate.

14. A method of regulating cellular adhesion at a substrate, the method comprising:

a) contacting a substrate with a solution comprising a known concentration of at least one polyphosphazene;

b) removing the solvent from the polyphosphazene solution in contact with the substrate to provide a polyphosphazene micro-structured substrate; and

c) controlling the degree of cellular adhesion at the polyphosphazene micro-structured substrate by;

i) controlling the polyphosphazene film thickness or surface coverage of the polyphosphazene micro-structured substrate;

ii) controlling the size or aspect ratio of the structural elements of the polyphosphazene micro-structured substrate;

iii) controlling the spatial density of the structural elements of the polyphosphazene micro-structured substrate;

or

iv) any combination thereof.

15. A method of regulating cellular adhesion at a substrate according to claim 14, wherein the polyphosphazene has the formula

16. A method of regulating cellular adhesion at a substrate according to claim 15, wherein at least one of R¹ to R⁶ is an alkoxy group substituted with at least one fluorine atom.

17. A method of regulating cellular adhesion at a substrate according to claim 15, wherein at least one of R¹ to R⁶ is selected from OCH₃, OCF₃, OCH₂CH₃, OCH₂CF₃, OCH₂CH₂CH₃, OCH₂CH₇CF₃, OCH₂CF₂CF₃, OCH(CF₃)₂, OCCH₃(CF₃)₂, OCH₂CF₂CF₂CF₃, OCH₂(CF₂)₃CF₃, OCH₂(CF₂)₄CF₃, OCH₂(CF₂)₅CF₃, OCH₂(CF₂)₆CF₃, OCH₂(CF₂)₇CF₃, OCCH₂CF₂CHF₂, OCH₂CF₂CF₂CHF₂, OCH₂(CF₂)₃CHF₂, OCH₂(CF₂)₄CHF₂, OCH₂(CF₂)₅CHF₂, OCH₂(CF₂)₆CHF₂, or OCH₂(CF₂)₇CHF₂.

18. A method of regulating cellular adhesion at a substrate according to claim 15, wherein the polyphosphazene has a molecular weight of at least 70,000 g/mol.

19. A method of regulating cellular adhesion at a substrate according to claim 14, wherein step d) is effected by:

a) increasing the polyphosphazene concentration in the polyphosphazene solution used to contact the substrate;

b) repeating steps a) and b) of claim 14 any number of times;

or

c) decreasing the polyphosphazene concentration in the polyphosphazene solution used to contact the substrate.

20. A method of regulating cellular adhesion at a substrate according to claim 14, wherein the concentration of the polyphosphazene solution in step a) is up to about 200 mg/mL.

21. A method of regulating cellular adhesion at a substrate according to claim 14, wherein the polyphosphazene micro-structured substrate has a polyphosphazene film thickness up to about 1000 nm.

22. A method of regulating cellular adhesion at a substrate according to claim 14, wherein the spacing density of the void structure within the polyphosphazene micro-structured substrate is from about 10,000 voids/100 μ

m² to 0 voids/100 μ

m².

23. A method of regulating cellular adhesion at a substrate according to claim 15, wherein the polyphosphazene surface coverage of the polyphosphazene micro-structured substrate is from about 50% to 100%.

24. A method of regulating cellular adhesion at a substrate, the method comprising:

a) contacting at least a portion of a substrate with at least one adhesion promoter to provide a treated substrate;

b) contacting the treated substrate with a solution comprising a known concentration of at least one polyphosphazene;

c) removing the solvent from the polyphosphazene solution in contact with the treated substrate to provide a polyphosphazene micro-structured substrate;

d) observing a degree of cellular adhesion at the polyphosphazene micro-structured substrate; and

either

e) adjusting the degree of cellular adhesion at the polyphosphazene micro-structured substrate relative to that obtained in step c) by;

i) increasing or decreasing the polyphosphazene film thickness or surface coverage of the polyphosphazene micro-structured substrate;

ii) increasing or decreasing the size or aspect ratio of the structural elements of the polyphosphazene micro-structured substrate;

iii) increasing or decreasing the spatial density of the structural elements of the polyphosphazene micro-structured substrate;

or

iv) any combination thereof.

or

f) maintaining the degree of cellular adhesion at the polyphosphazene micro-structured substrate by preserving the polyphosphazene micro-structure on the substrate as provided in step c).

26. A method of regulating cellular adhesion at a substrate according to claim 24, wherein the adhesion promoter is (3-aminopropyl)trimethoxysilane, the polyphosphazene is poly[bis(2,2,2-trifluoroethoxy)]phosphazene, and adjusting the degree of cellular adhesion at the polyphosphazene micro-structured substrate is effected by:

a) increasing the polyphosphazene concentration in the polyphosphazene solution relative to that of step b) to decrease cellular attraction;

or

b) decreasing the polyphosphazene concentration in the polyphosphazene solution relative to that of step b) to increase cellular attraction.

27. A substrate comprising a polyphosphazene micro-structured surface, wherein:

a) the polyphosphazene micro-structured substrate has a polyphosphazene film thickness from 1 nm to 300 nm;

b) the spacing density of the void structure within the polyphosphazene micro-structured substrate is from 2,500 voids per 100 μ

m² to 100 voids/100 μ

m²; and

c) the surface coverage of the polyphosphazene micro-structured substrate is from 50% to 100%.

28. A substrate comprising a polyphosphazene micro-structured surface according to claim 27, wherein:

a) the polyphosphazene micro-structured substrate has a polyphosphazene film thickness from 1 nm to 300 nm;

b) the spacing density of the void structure within the polyphosphazene micro-structured substrate is from 2,200 voids per 100 μ

m² to 150 voids/100 μ

m²; and

c) the surface coverage of the polyphosphazene micro-structured substrate is from 75% to 100%.

29. A method of producing a biological implant, the method comprising contacting a form-building substrate with a biological starting material comprising cells to be cultured, wherein:

a) at least a portion of the substrate comprises a polyphosphazene micro-structured surface; and

b) the polyphosphazene has the formula