Implant for Lymph Node Formation/Regeneration

Abstract

The present invention relates to the field of implants for the formation/regeneration of lymph nodes. In particular, the present invention relates to an implant comprising a biodegradable scaffold and lymph node fragments immobilized therein and/or thereon, to a method of manufacturing such an implant and to uses of such an implant.

Claims

1.-15. (canceled)

16. A method of manufacturing an implant for the formation and/or regeneration of a lymph node in the body of a patient in need thereof, said method comprising the steps of providing a biodegradable scaffold, wherein said biodegradable scaffold is a hollow three-dimensional object having a lumen, and immobilizing from 2 up to 10 lymph node fragments inside and/or on the inner surface of said hollow three-dimensional object, wherein said individual lymph node fragments are arranged close enough to each other to facilitate coalescence of the individual lymph node fragments into one complete functional lymph node; and wherein the size and shape of the lymph node to be formed or regenerated is controlled by the shape and size of the lumen of said biodegradable scaffold.

17. The method according to claim 16, wherein said immobilizing lymph node fragments inside and/or on the inner surface of said hollow three-dimensional object comprises immobilizing from 3 up to 6 lymph node fragments.

18. The method according to 16, wherein said immobilizing lymph node fragments in and/or on the inner surface of said hollow three-dimensional object comprises immobilizing said lymph node fragments inside and/or on the inner surface of said hollow three-dimensional object with fibrin or hyaluronic acid.

19. The method of claim 16, wherein said immobilizing lymph node fragments in and/or on the inner surface of said hollow three-dimensional object comprises embedding said lymph node fragments in a proteinaceous matrix.

20. The method of claim 16, wherein said immobilizing lymph node fragments in and/or on the inner surface of said hollow three-dimensional object comprises immobilizing said lymph node fragments in and/or on the inner surface of said hollow three-dimensional object by wedging said lymph node fragments into cavities present within said hollow three-dimensional object.

21. The method according to claim 16, wherein said lymph node fragments are slices of a lymph node with a thickness of not more than 2 mm, or said lymph node fragments are pieces with a diameter of not more than 3 mm.

22. The method according to claim 21, wherein said lymph node fragments are slices of a lymph node with a thickness of not more than 1 mm, or said lymph node fragments are pieces with a diameter of not more than 2 mm.

23. The method according to claim 16, wherein said biodegradable scaffold is made from polycaprolactone, polyglycolide, polylactide, poly(1,3-trimethylene carbonate) or a copolymer of polycaprolactone and either poly-trimethylene carbonate or polylactide or a copolymer of polycaprolactone, polylactide and polyglycolide, wherein, preferably said biodegradable scaffold is made from polycaprolactone.

24. The method according to claim 16, wherein said biodegradable scaffold comprises holes and/or pores which reach through the three-dimensional structure of said biodegradable scaffold, wherein said holes and/or pores have a diameter in the range of from 20 μm to 2 mm.

25. The method according to claim 16, wherein said biodegradable scaffold has a tubular shape.

26. The method according to claim 16, wherein said biodegradable scaffold has the shape of a tube with a longitudinal slit on one side.

27. The method according to claim 16, wherein said method comprises only steps carried out in vitro.

28. The method according to claim 16, wherein said patient is a human suffering from lymphedema.

29. The method according to claim 16, wherein said lymph node fragments are fragments of a lymph node obtained from said patient.

30. The method according to claim 16, wherein said lymph node fragments are fragments of a lymph node obtained from a body part which does not suffer from lymphedema.

31. The method according to claim 16, wherein said lymph node fragments are fragments of an axillary or inguinal lymph node.

32. An implant for the formation and/or regeneration of a lymph node in the body of a patient in need thereof, said implant comprising a biodegradable scaffold, wherein said biodegradable scaffold is a hollow three-dimensional object, and wherein from 2 up to 10 lymph node fragments are immobilized inside and/or on the inner surface of said hollow three-dimensional object.

33. The implant according to claim 32, wherein said implant comprises from 3 up to 6 lymph node fragments immobilized inside and/or on the inner surface of said hollow three-dimensional object.

34. The implant according to claim 32, wherein said lymph node fragments are immobilized in and/or on the inner surface of said hollow three-dimensional object with fibrin or with hyaluronic acid.

35. The implant according to claim 32, wherein said lymph node fragments are immobilized in and/or on the inner surface of said hollow three-dimensional object by being embedded in a proteinaceous matrix.

36. The implant according to claim 32, wherein said lymph node fragments are immobilized in and/or on the inner surface of said hollow three-dimensional object by having been wedged into cavities present within said hollow three-dimensional object.

37. The implant according claim 32, wherein said lymph node fragments are slices of a lymph node with a thickness of not more than 2 mm, or said lymph node fragments are pieces with a diameter of not more than 3 mm.

38. The implant according claim 36, wherein said lymph node fragments are slices of a lymph node with a thickness of not more than 1 mm, or said lymph node fragments are pieces with a diameter of not more than 2 mm.

39. The implant according to claim 32, wherein said biodegradable scaffold is made from polycaprolactone, polyglycolide, polylactide, poly(1,3-trimethylene carbonate) or a copolymer of polycaprolactone and either poly-trimethylene carbonate or polylactide or a copolymer of polycaprolactone, polylactide and polyglycolide, wherein, preferably said biodegradable scaffold is made from polycaprolactone.

40. The implant according to claim 32, wherein said biodegradable scaffold comprises holes and/or pores which reach through the three-dimensional structure of said biodegradable scaffold, wherein said holes and/or pores have a diameter in the range of from 20 μm to 2 mm.

41. The implant according to claim 32, wherein said biodegradable scaffold has a tubular shape.

42. The implant according to claim 32, wherein said biodegradable scaffold has the shape of a tube with a longitudinal slit on one side.

Description

BRIEF DESCRIPTION OF THE FIGURES

[0088] In the following, reference is made to the figures, wherein:

[0089] FIG. 1 shows an exemplary embodiment of the present invention wherein the biodegradable scaffold has the shape of a sheet or flat cuboid. [0090] (A) Example of a biodegradable scaffold in the shape of a sheet or flat cuboid. [0091] (B) Exemplary embodiment of an implant according to the present invention comprising a biodegradable scaffold having the shape of a sheet or flat cuboid. [0092] (C) Example of such an implant placed in proximity to a blood vessel.

[0093] FIG. 2 shows an exemplary embodiment of the present invention wherein the biodegradable scaffold has a tubular shape. [0094] (A) Example of a biodegradable scaffold with a tubular shape. [0095] (B) Exemplary embodiment of an implant according to the present invention comprising a biodegradable scaffold having a tubular shape, wherein the lymph node fragments are immobilized within the lumen of said tubular shape. [0096] (C) Example of such an implant placed in proximity to a blood vessel.

[0097] FIG. 3 shows an exemplary embodiment of the present invention wherein the biodegradable scaffold has the shape of a tube with a longitudinal slit on one side and wherein the lymph node fragments are immobilized at the inner face of the wall of said tube with a longitudinal slit on one side. [0098] (A) Example of a biodegradable scaffold in the shape of a tube with a longitudinal slit on one side. [0099] (B) Exemplary embodiment of an implant according to the present invention comprising a biodegradable scaffold in the shape of a tube with a longitudinal slit on one side, wherein the lymph node fragments are immobilized at the inner face of the wall of said tube with a longitudinal slit on one side. [0100] (C) Example of such an implant placed around a blood vessel.

[0101] FIG. 4 shows an exemplary embodiment of the present invention wherein the biodegradable scaffold has the shape of a tube with a longitudinal slit on one side and wherein the lymph node fragments are immobilized at the outer face of the wall of said tube with a longitudinal slit on one side. [0102] (A) Example of a biodegradable scaffold in the shape of a tube with a longitudinal slit on one side. [0103] (B) Exemplary embodiment of an implant according to the present invention comprising a biodegradable scaffold in the shape of a tube with a longitudinal slit on one side, wherein the lymph node fragments are immobilized at the outer face of the wall of said tube with a longitudinal slit on one side. [0104] (C) Example of such an implant placed around a blood vessel.

[0105] FIG. 5 shows a regenerated lymph node (bean-shaped structure held with forceps) during explantation carried out 8 weeks after implantation of a biodegradable scaffold with immobilized human lymph node fragments into the inguinal body region of a nude mouse.

[0106] FIG. 6 is a photographic image obtained upon surgically opening the inguinal body region of a nude mouse where a polycaprolacton scaffold with immobilized human lymph node fragments had been implanted 8 weeks before. A full-grown lymph node of considerable size with very good vascularization and connection to the lymphatic system has developed, whereas the biodegradable scaffold has been degraded.

[0107] FIG. 7 shows a histological section of the inguinal body region of an immunodefficient mouse in which a biodegradable scaffold with immobilized human lymph node fragments had been implanted 16 weeks before. The formation of lymph vessels and lymphatic regeneration is observed.

[0108] FIG. 8 shows a histological section of the inguinal body region of an immunodefficient mouse in which a biodegradable scaffold without any immobilized human lymph node fragments had been implanted 16 weeks before. Neither the formation of lymph vessels nor any significant signal of lymphatic regeneration is observed.

[0109] All methods mentioned in the figure descriptions above were carried out as described in detail in the Examples.

DETAILED DESCRIPTION OF EXEMPLARY EMBODIMENTS

[0110] For the purpose of promoting an understanding of the principles of the invention, reference will now be made to the preferred embodiments and specific language will be used to describe the same. It will nevertheless be understood that no limitation of the scope of the invention is intended thereby, such alterations and further modifications in the device and methods and such further applications of the principles of the invention as illustrated therein being contemplated as would normally occur now or in the future to one skilled in the art to which the invention relates.

[0111] Moreover, it is to be understood that features and advantages described with regard to one aspect of the invention may also be implied by other aspects of the invention.

[0112] FIG. 1 shows an exemplary embodiment of an implant according to the present invention and its possible placement relative to a blood vessel. In this embodiment, the biodegradable scaffold 2 of the implant 1 has the shape of a sheet or flat cuboid (FIG. 1 A). Lymph node fragments 3 are immobilized on the surface of the biodegradable scaffold 2, for example by gluing them with fibrin to the biodegradable scaffold, resulting in an implant 1 according to the invention (FIG. 1 B). The implant 1 according to the invention can be placed in the proximity of a blood vessel 4 (FIG. 1 C; blood flow shown by arrows). Due to the proximity to the blood vessel 4, the lymph node fragments 3 are supplied efficiently with oxygen and nutrients. Thus, within a few weeks to months, the individual lymph node fragments 3 grow together, develop blood vessels connecting them to the blood circulation of the body and lymph vessels linking them to the lymph system, and reorganize to form/regenerate a fully functional lymph node that is capable of removing interstitial fluid from the tissue surrounding it.

[0113] FIG. 2 shows another exemplary embodiment of an implant according to the present invention and its possible placement relative to a blood vessel. In this embodiment, the biodegradable scaffold 2 of the implant 1 has a tubular shape (FIG. 2 A). Lymph node fragments 3 are immobilized within the lumen of the tubularly shaped biodegradable scaffold 2, for example by embedding the lymph node fragments 3 in fibrin within the lumen of the biodegradable scaffold 2, resulting in an implant 1 according to the invention (FIG. 2 B). The implant 1 according to the invention can be placed in the proximity of a blood vessel 4 (FIG. 2 C). Due to the proximity to the blood vessel 4, the lymph node fragments 3 are supplied efficiently with oxygen and nutrients, allowing for efficient formation/regeneration of a fully functional lymph node. The size and three-dimensional shape of the lymph node to be formed/regenerated can be influenced by the shape and size of the lumen of said biodegradable scaffold 2 having a tubular shape.

[0114] FIG. 3 shows another exemplary embodiment of an implant according to the present invention and its possible placement relative to a blood vessel. In this embodiment, the biodegradable scaffold 2 of the implant 1 has the shape of a tube with a longitudinal slit on one side (FIG. 3 A). Lymph node fragments 3 are immobilized at the inner face of the wall of the tube with a longitudinal slit on one side formed by the biodegradable scaffold 2, for example by gluing the lymph node fragments 3 with fibrin to the biodegradable scaffold 2, resulting in an implant 1 according to the invention (FIG. 3 B). The implant 1 according to the invention can be placed around a blood vessel 4 (FIG. 3 C). This ensures that the lymph node fragments 3 are stably held in place in very close proximity to the nourishing blood vessel 4, while the shape and size of the lymph node to be formed/regenerated is controlled by the shape and size of the lumen of the biodegradable scaffold 2.

[0115] The embodiment depicted in FIG. 4 differs from that of FIG. 3 by the position of the immobilized lymph node fragments 3: While in the embodiment of FIG. 3 the lymph node fragments 3 are immobilized at the inner face of the wall of the tube with a longitudinal slit on one side, in the embodiment of FIG. 4 they are immobilized at the outer face. This ensures that the lymph node fragments 3 are stably held in place in proximity to the nourishing blood vessel 4, while leaving the lymph node to be formed/regenerated freedom with respect to the three-dimensional shape that it will adopt.

[0116] Not shown in FIGS. 1-4 is the structure of the biodegradable scaffold (holes, pores and/or cavities existing in the scaffold, e.g. between the bars and struts from which the biodegradable scaffold is formed). Typically, the biodegradable scaffold may not have a smooth surface. Moreover, while the lymph node fragments are indicated as spheres in FIGS. 1-4, it is to be understood that the lymph node fragments may have various other shapes or size and typically will be slices or rectangular blocks prepared by cutting a lymph node with a surgical blade.

[0117] As the skilled person will appreciate, various other shapes of the biodegradable scaffold, number, shapes and sizes of the immobilized lymph node fragments, ways to arrange or immobilize the lymph node fragments on the biodegradable scaffold, and spatial arrangements of the implant with respect to the blood vessel than those shown in the exemplary embodiments depicted in FIGS. 1-4 lie within the scope of the present invention, as well.

EXAMPLES

[0118] In the following, reference is made to the examples, which are given to illustrate, not to limit the present invention.

Example 1

[0119] Human lymph nodes were obtained by surgical removal and were mechanically broken up with a scalpel into lymph node fragments. The lymph node fragments were transferred into the lumen of a biodegradable polycaprolacton scaffold in the shape of a hollow cylinder (lymph node fragments obtained from one lymph node per scaffold) and fixed with fibrin glue. A growth of the lymphogenic cells in the 3D matrix structure takes place. DNA analysis reveals that the cells increasingly proliferate and spread throughout the scaffold. Subsequently, the biodegradable scaffold with the immobilized lymph node fragments was transplanted into one of the inguinal regions of immunodeficient nude mice, in close proximity to a blood vessel. A biodegradable scaffold with fibrin glue but no lymph node fragments was implanted into the other inguinal region of the immunodeficient mice so as to provide a control sample. The experiment was carried out with 7 mice.

[0120] After 8 weeks, the site of transplantation was re-opened by surgery and in vivo imaging was carried out by injecting a contrast agent into the respective body region and examining the lymph nodes/lymph vessels in the living mice under anesthesia. It was observed that full-grown lymph nodes of considerable size had formed which were well vascularized and had developed new lymph vessels that connected them to the lymphatic system (FIGS. 5 and 6).

[0121] After 16 weeks potential lymph vessels and high endothelial venules are also observed in the surrounding fatty tissues (FIG. 7). This provides evidence that the environment drains liquid into the regenerated lymph nodes and hence suggests functional activity thereof. Furthermore, the regenerated lymph nodes display similar structures to those characteristic of healthy lymph nodes, in particular lymph follicles, medulla and capsule.

[0122] None of the reported features is significantly observed in the control sample group, that is, in the implants of the scaffold with fibrin glue but without any lymph node fragments (FIG. 8). This suggests that said features can be traced back to the implantation of the lymph node fragments.

[0123] Thus the lymph node fragments are observed to regenerate in mice and to display similar structures to those characteristic of healthy lymph nodes. They furthermore appear to retake their normal working function.

REFERENCES

[0124] Becker C, Assouad J, Riquet M, Hidden G. Postmastectomy lymphedema: long-term results following microsurgical lymph node transplantation. Ann Surg. (2006), 243(3):313-315. [0125] Pabst R, Rothkötter H J. Regeneration of autotransplanted lymph node fragments. Cell Tissue Res. (1988), 251(3):597-601. [0126] Sommer T, Buettner M, Bruns F, Breves G, Hadamitzky C, Pabst R. Improved regeneration of autologous transplanted lymph node fragments by VEGF-C treatment. Anat Rec (2012), 295(5):786-791.