EP0330344A2
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EP
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Method of crosslinking collagen using photoactivatable chemical crosslinkers. Uses of both the such crosslinked collagen or amino acid containing polymers .
Assignee: Cedars-Sinai Medical Center
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Filing: 1989-02-09
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Granted: —
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Expires: 2009-02-09
Abstract
Abstract A method for molecularly crosslinking amino acid containing polymers by photoactivating chemical crosslinkers which have been combined with the polymers. Collagen crosslinked by this method can be used as a bioadhesive for sutureless closures of the skin and eye or as a superhydrated material for contact lenses, moist bandage contact lens, lens or corneal implant material, or as a drug delivery mechanism.
Inventors
- 👤 Anthony Bart Nesburn
- 👤 Michael Gorin
- 👤 Marvin Martinez
- 👤 Maria Cristina Kenney
- 👤 Ezra Maguen
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-10-23 14:03
CPC Classifications
G; G03; G03F; G03F7/00; G03F7/004; G03F7/038; A; A61; A61L; A61L24/00; A61L24/04; A61L24/10; A61L24/102; A61L27/00; A61L27/14; A61L27/22; A61L27/24; C; C08; C08H; C08H1/00; C08H1/06; A61F; A61F2310/00; A61F2310/00005; A61F2310/00365; Y; Y10; Y10S; Y10S128/00; Y10S128/08
Claims Analysis
Total Claims: 26
Claim 1
Independent Claim
98 words
A method of crosslinking two or more amino acid containing polymers with photoactivatable heterobifunctional crosslinking agents, said crosslinking agents having a photoactivatable site and a conventional site, which includes: (a) selecting two or more amino acid containing polymers: (b) combining said amino acid containing polymers with said photoactivatable heterobifunctional crosslinking agents such that the conventional site on the crosslinking agent is bound to the polymer, and the other photoactivatable site on the crosslinking agent is unbound, such that upon photoactivation, crosslinks are formed when the photoactivated site on the crosslinking agent binds to another amino acid containing polymer.
Claim 2
Independent Claim
98 words
A method of crosslinking two or more amino acid containing polymers with photoactivatable heterobifunctional crosslinking agents, said crosslinking agents having a photoactivatable site and a conventional site, which includes: (a) selecting two or more amino acid containing polymers: (b) combining said amino acid containing polymers with said photoactivatable heterobifunctional crosslinking agents such that the conventional site on the crosslinking agent is bound to the polymer, and the other photoactivatable site on the crosslinking agent is unbound, such that upon photoactivation, crosslinks are formed when the photoactivated site on the crosslinking agent binds to another amino acid containing polymer.
Claim 3
Dependent Claim
15 words
A method as claimed in Claim 1, wherein the amino acid containing polymer is collagen.
Claim 4
Dependent Claim
21 words
A method as claimed in Claim 2, wherein the collagen is an Atelocollagen (e.g. Vitrogen 100) or Rat Tail Type I.
Claim 5
Independent Claim
26 words
A method as claimed in any one of the preceding claims, wherein the concentration of the amino-acid containing polymer is between 2.5 mg/ml to 10 mg/ml.
Claim 6
Dependent Claim
38 words
A method as claimed in Claim 4, wherein the concentration is achieved by dialyzing the polymer against acetate buffer at pH 5, or by lyophilizing known quantities of the polymer and the resuspending in .012 HCl or CH₃COOH.
Claim 7
Independent Claim
24 words
A method as claimed in any one of the preceding claims, wherein the pH of the amino-acid containing polymers range between 2.0 and 7.4.
Claim 8
Independent Claim
26 words
A method as claimed in any one of the preceding claims, wherein the photoactivatable heterobifunctional crosslinking agent is a diazo compound or an Aryl or Alkyl-Azide.
Claim 9
Dependent Claim
25 words
A method as claimed in Claim 7, wherein the photoactivatable heterobifunctional crosslinker is 4-azidobenzoic acid N-Hydroxysuccinimide Ester (HSAB) or 6-(-4-azido-2-nitrophenyl-amino) hexanoic acid N-hydroxysuccinimide Ester (SANAH).
Claim 10
Independent Claim
20 words
A method as claimed in any one of the preceding claims, wherein the crosslinking agent is dissolved in a solvent.
Claim 11
Dependent Claim
12 words
A method as claimed in Claim 9, where the solvent is DMSO.
Claim 12
Dependent Claim
17 words
A method as claimed in Claim 10, wherein the concentration of DMSO is not less than 50%.
Claim 13
Independent Claim
25 words
A method as claimed in any one of the preceding claims, wherein the concentration of the photoactivatable heterobifunctional crosslinking agent is between 5mM and 25mM.
Claim 14
Independent Claim
42 words
A method as claimed in any one of the preceding claims, wherein the photoactivation is achieved with a wavelength range of 220 nanometers (nm) to 310nm, with an optimum absorbing wavelength approximately 265mm with a photoactivation time not to exceed 20 minutes.
Claim 15
Independent Claim
31 words
A method as claimed in any one of the preceding claims, wherein the prephotoactivated combination of polymer and crosslinking agent is run through a Sephadex G-25 column to remove unbound crosslinker.
Claim 16
Dependent Claim
29 words
A method as claimed in Claim 14, wherein fractions collected from the Sephadex G-25 column are run through a Spectrophotometer 260-320nm to determine and collect the peak collagen-reagent fractions.
Claim 17
Independent Claim
164 words
A method of crosslinking collagen with photoactivatable heterobifunctional crosslinking agents, said crosslinking agents having a photoactivatable site and a conventional site, which comprises: (a) preparing a 9 mg/ml concentration of collagen molecules at pH 7.2. (b) dissolving said photoactivatable heterobifunctional crosslinking agents in DMSO such that the concentration of the crosslinking agent is 10mM. (c) combining said collagen mixture with said photoactivatable heterobifunctional crosslinking agents, such that the conventional site on the crosslinking agent is bound to the collagen molecule, and the other photoactivatable site on the crosslinking agent is unbound, such that upon photoactivation, crosslinks are formed when the photoactivated site on the crosslinking agent binds to another collagen molecule. (d) running the pre-photoactivated combination of collagen and crosslinking agent in Step (c) through a Sephadex G-25 column such that unbound crosslinking agent is removed. (e) collecting fractions from the Sephadex G-25 column after Step (d) and running said fractions through a Spectrophotometer 260-320nm such that peak collagen-crosslinking agent fractions can be collected.
Claim 18
Independent Claim
164 words
A method of crosslinking collagen with photoactivatable heterobifunctional crosslinking agents, said crosslinking agents having a photoactivatable site and a conventional site, which comprises: (a) preparing a 9 mg/ml concentration of collagen molecules at pH 7.2. (b) dissolving said photoactivatable heterobifunctional crosslinking agents in DMSO such that the concentration of the crosslinking agent is 10mM. (c) combining said collagen mixture with said photoactivatable heterobifunctional crosslinking agents, such that the conventional site on the crosslinking agent is bound to the collagen molecule, and the other photoactivatable site on the crosslinking agent is unbound, such that upon photoactivation, crosslinks are formed when the photoactivated site on the crosslinking agent binds to another collagen molecule. (d) running the pre-photoactivated combination of collagen and crosslinking agent in Step (c) through a Sephadex G-25 column such that unbound crosslinking agent is removed. (e) collecting fractions from the Sephadex G-25 column after Step (d) and running said fractions through a Spectrophotometer 260-320nm such that peak collagen-crosslinking agent fractions can be collected.
Claim 19
Dependent Claim
25 words
A molecularly crosslinked amino acid containing polymer comprising two or more amino acid containing polymers crosslinked to one another by a photoactivated heterobifunctional crosslinking agent.
Claim 20
Dependent Claim
25 words
A molecularly crosslinked amino acid containing polymer comprising two or more amino acid containing polymers crosslinked to one another by a photoactivated heterobifunctional crosslinking agent.
Claim 21
Dependent Claim
23 words
A molecularly crosslinked collagen comprising two or more collagen molecules crosslinked to one another by a photoactivated heterobifunctional crosslinking agent dissolved in DMSO.
Claim 22
Dependent Claim
23 words
A molecularly crosslinked collagen comprising two or more collagen molecules crosslinked to one another by a photoactivated heterobifunctional crosslinking agent dissolved in DMSO.
Claim 23
Independent Claim
76 words
A method of preparing crosslinked bioadhesives comprising: (a) selecting two or more amino acid containing polymers; (b) combining said amino acid containing polymers with said photoactivatable heterobifunctional crosslinking agents such that the conventional site on the crosslinking agent is bound to the polymer, and the other photoactivatable site on the crosslinking agent is unbound, such that upon photoactivation, crosslinks are formed when the photoactivated site on the crosslinking agent binds to another amino acid containing polymer.
Claim 24
Independent Claim
76 words
A method of preparing crosslinked bioadhesives comprising: (a) selecting two or more amino acid containing polymers; (b) combining said amino acid containing polymers with said photoactivatable heterobifunctional crosslinking agents such that the conventional site on the crosslinking agent is bound to the polymer, and the other photoactivatable site on the crosslinking agent is unbound, such that upon photoactivation, crosslinks are formed when the photoactivated site on the crosslinking agent binds to another amino acid containing polymer.
Claim 25
Independent Claim
80 words
A method for preparing superhydrated crosslinked amino acid containing polymers comprising: (a) selecting two or more amino acid containing polymers; (b) combining said amino acid containing polymers with said photoactivatable heterobifunctional crosslinking agents such that the conventional site on the crosslinking agent is bound to the polymer, and the other photoactivatable site on the crosslinking agent is unbound, such that upon photoactivation, crosslinks are formed when the photoactivated site on the crosslinking agent binds to another amino acid containing polymer.
Claim 26
Independent Claim
80 words
A method for preparing superhydrated crosslinked amino acid containing polymers comprising: (a) selecting two or more amino acid containing polymers; (b) combining said amino acid containing polymers with said photoactivatable heterobifunctional crosslinking agents such that the conventional site on the crosslinking agent is bound to the polymer, and the other photoactivatable site on the crosslinking agent is unbound, such that upon photoactivation, crosslinks are formed when the photoactivated site on the crosslinking agent binds to another amino acid containing polymer.
References Cited (Backward Citations)
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