Support Having Nanostructured Titanium Dioxide Film And Uses Thereof

• US 20080219959A1
• Filed: 01/18/2008
• Published: 09/11/2008
• Est. Priority Date: 07/21/2005
• Status: Abandoned Application

• Current Assignee: Tethis S.p.A.
• Original Assignee: Tethis S.p.A.
• Inventors: Milani, Paolo, Barborini, Emanuele, Bongiorno, Gero Antonio, Piseri, Paolo, Carbone, Roberta, Pelicci, Pier Giuseppe
• Application Number:
  US12/016,716
• Publication Number:
  US 20080219959A1
• Time in Patent Office: Days
• US Class Current: 424/93.21
• CPC Class Codes: A61P 31/12 Antivirals
• CPC Class Codes: A61P 31/18 for HIV
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: B01J 2219/00317 Microwell devices, i.e. having large numbers of wells
• CPC Class Codes: B01J 2219/00527 Sheets
• CPC Class Codes: B01J 2219/00612 the surface being inorganic
• CPC Class Codes: B01J 2219/00621 by physical means, e.g. trenches, raised areas
• CPC Class Codes: B01J 2219/00644 the porous medium being present in discrete locations, e.g. gel pads
• CPC Class Codes: B01J 2219/00659 Two-dimensional arrays
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: B01J 2219/00743 Cells
• CPC Class Codes: C12M 25/14 Scaffolds; Matrices in general C12N5/0068
• CPC Class Codes: G01N 33/551 the carrier being inorganic
• CPC Class Codes: G01N 33/56966 Animal cells
• CPC Class Codes: G01N 33/56983 Viruses

Abstract

The present invention relates to supports for bioassays and the use thereof in cell culturing and in cell-based methods and assays. More precisely, the invention provides solid materials coated with films of nanostructured titanium dioxide suitable for the immobilisation of viruses and for cell-adhesion. The nanostructured TiO<SUB>2</SUB> film-coated support of the invention is particularly useful for the preparation of microarrays for genetic and phenotypic analysis.

Claims

1-10. -10. (canceled)

11. A method for cell infection with viruses in vitro comprising the steps of:

(a) providing a solid support of a biocompatible substrate material being at least partially coated with a nanostructured TiO₂ film; and

(b) culturing cells on the nanostructured TiO₂ film-coated support in the presence of an infecting virus.

12. The method of claim 11 comprising the further steps of:

(c) contacting the nanostructured TiO₂ film-coated support with viruses prior to culturing cells in the support;

(d) contacting the virus adhered on the surface of the nanostructured TiO₂ film-coated support with a cell preparation; and

(e) culturing the cells for a time sufficient for the infection to occur.

15. The method of claim 11 wherein the viruses are retroviruses, adenoviruses, adeno-associated viruses (AAV) and any other viruses that can be utilized as vectors for genetic manipulation of cells.

16. The method of claim 11 wherein the viruses are genetically modified.

13. A method for cell infection with viruses in vitro comprising the steps of:

(a) providing a solid support of a biocompatible substrate material being at least partially coated with a nanostructured TiO₂ film;

(b) immobilising streptavidin, avidin or neutravidin on the nanostructured TiO₂ film coating;

(c) contacting the nanostructured TiO₂ film-coated support with a biotinylated virus so as to form a complex of biotinylated viruses with the immobilised streptavidin, avidin or neutravidin;

(d) contacting the complex with a cell preparation; and

(e) culturing the cells for a time sufficient for the infection to occur.

14. A method for cell infection with viruses in vitro comprising the steps of:

(a) providing a solid support of a biocompatible substrate material being at least partially coated with a nanostructured TiO₂ film;

(b) adding a cell preparation to the nanostructured TiO₂ film-coated support;

(c) culturing the cells for an appropriate period of time;

(d) adding a viral supernatant;

(e) culturing the cells for a time sufficient for the infection to occur.

17. (canceled)

18. Implantable particles or device of a nanostructured TiO₂ film-coated biocompatible material loaded with viruses.

19. A method for gene therapy ex vivo comprising the steps of:

(a) recovering cells to be genetically modified from a patient;

(b) establishing a primary cell culture from the recovered cells;

(c) infecting the cells by providing a solid support of a biocompatible substrate material being at least partially coated with a nanostructured TiO₂ film; and

culturing cells on the nanostructured TiO₂ film-coated support with a virus that carries genetic information;

(d) and re-administering the infected cells to the patient.

21. The method of claim 19 wherein the viruses are retroviruses, adenoviruses, adeno-associated viruses (AAV) and any other viruses that can be utilized as vectors for genetic manipulation of cells.

22. The method according to claim 19 wherein the viruses are genetically modified.

20. A method for gene therapy in vivo comprising the steps of:

(a) providing particles or a device of a nanostructured TiO₂ film-coated biocompatible material;

(b) loading the particles or device with viruses; and

(c) implanting the virus-loaded particles or device into a tissue of a patient.

23. A method for cell replacement therapy comprising the steps of:

(a) providing particles or a device of a nanostructured TiO₂ film-coated biocompatible material;

(b) loading the particles or device with cells to be replaced in a patient; and

(c) implanting the cell-loaded particles or device into a tissue of a patient.

24. The method of claim 23 wherein the cells are genetically modified.

25. A solid support fabricated from a biocompatible substrate material which is at least partially coated with a nanostructured TiO₂ film having viruses and/or cells immobilised on the surface thereof.

26. The solid support of claim 25 wherein the film of nanostructured TiO₂ consists of TiO₂ nanoparticles with a diameter below 20 nm embedded in an amorphous TiO₂ matrix with a density of below 75% of bulk TiO₂ density.

27. The solid support of claim 25 which comprises a slide, dish, flask, plate, coverslip, fiber, foam, particle, membrane, porous scaffold, mesh or implant.

28. The solid support of claim 25 wherein the biocompatible substrate material is glass, plastic, ceramic, metal or a biodegradable or undegradable biopolymeric material.

29. The solid support of claim 25 wherein the viruses are retroviruses, adeno-viruses, adeno-associated viruses (AAV) and any other viruses that can be utilized as vectors for genetic manipulation of cells.

31. The solid support according to claim 25 wherein biotinylated viruses are immobilised on the nanostructured TiO₂ film carrying streptavidin avidin or neutravidin.

32. The solid support according to claim 25 wherein the nanostructured TiO₂ film is in the form of a micro- or nano-pattern.

33. The solid support according to claim 32 wherein viruses carrying different genetic inserts are spotted on the surface of the nanostructured TiO₂ film.

34. A method for the production of a solid support, comprising the steps of:

fabricating the solid support from a biocompatible substrate material;

depositing a nanostructured TiO₂ film at least a portion of the substrate material by nanoparticle deposition from a gas-phase; and

contacting the surface of the nanostructured TiO₂ film with viruses and/or cells.

30. The solid support of claim 34 wherein the viruses are genetically modified.

35. The method of claim 34 wherein the nanoparticle deposition from the gas-phase is carried out by means of supersonic cluster beam deposition (SCBD) using a pulsed microplasma cluster source.

36. A method of virus-mediated gene delivery to cells, comprising the steps of:

(a) providing a solid support of a biocompatible substrate material being at least partially coated with a nanostructured TiO₂ film;

(b) contacting the nanostructured TiO₂ film-coated support with gene delivering viruses;

(c) contacting the virus adhered on the surface of the nanostructured TiO₂ film-coated support with a cell preparation; and

(d) culturing the cells for a time sufficient for gene delivery to occur.