Perivascular mesenchymal precursor cells

• US 10,421,946 B2
• Filed: 06/23/2015
• Issued: 09/24/2019
• Est. Priority Date: 03/28/2003
• Status: Active Grant

• Current Assignee: Mesoblast, Inc. (Mesoblast Ltd.)
• Original Assignee: Mesoblast, Inc. (Mesoblast Ltd.)
• Inventors: Gronthos, Stan, Zannettino, Andrew, Shi, Songtao
• Primary Examiner(s): Belyavskyi, Michail A
• Application Number:
  US14/746,960
• Publication Number:
  US 20150368614A1
• Time in Patent Office: 1,554 Days
• Field of Search: None
• CPC Class Codes: A61K 2300/00 Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00
• CPC Class Codes: A61K 35/28 Bone marrow; Haematopoietic stem cells; Mesenchymal stem cells of any origin, e.g. adipose-derived stem cells
• CPC Class Codes: A61K 35/44 Vessels; Vascular smooth muscle cells; Endothelial cells; Endothelial progenitor cells
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61K 9/0014 Skin, i.e. galenical aspects of topical compositions non-active ingredients are additionally classified in A61K47/00; A61K9/0009, A61K9/0021, A61K9/7015, A61K9/7023 take precedence; cosmetic preparations A61K8/00, A61Q; preparations for wound dressings or bandages A61L26/00
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: A61P 9/04 Inotropic agents, i.e. stimulants of cardiac contraction; Drugs for heart failure
• CPC Class Codes: A61P 9/10 for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
• CPC Class Codes: C12N 2500/42 Organic phosphate, e.g. beta glycerophosphate
• CPC Class Codes: C12N 2501/11 Epidermal growth factor [EGF]
• CPC Class Codes: C12N 2501/135 Platelet-derived growth factor [PDGF]
• CPC Class Codes: C12N 2501/165 Vascular endothelial growth factor [VEGF]
• CPC Class Codes: C12N 2501/235 Leukemia inhibitory factor [LIF]
• CPC Class Codes: C12N 2501/39 Steroid hormones
• CPC Class Codes: C12N 2506/1392 from mesenchymal stem cells from other natural sources
• CPC Class Codes: C12N 5/0663 Bone marrow mesenchymal stem cells (BM-MSC)
• CPC Class Codes: C12N 5/0667 Adipose-derived stem cells [ADSC]; Adipose stromal stem cells
• CPC Class Codes: C12N 5/0668 Mesenchymal stem cells from other natural sources
• CPC Class Codes: C12N 5/0691 Vascular smooth muscle cells; 3D culture thereof, e.g. models of blood vessels

Abstract

Mesenchymal precursors cells have been isolated from perivascular niches from a range of tissues utilizing a perivascular marker. A new mesenchymal precursor cell phenotype is described characterized by the presence of the perivascular marker 3G5, and preferably also alpha smooth muscle actin together with early developmental markers such as MUC 18, VCAM-1 and STRO-1^bri. The perivascular mesenchymal precursor cell is multipotential and is shown to form, vascular tissue, as well as bone marrow dentin and pulp. A method of enriching using cell sorting based on these markers is also described.

Claims

1. A therapeutic composition comprising a suspension of culture expanded mammalian mesenchymal precursor cells (MPCs), wherein the culture expanded cells are derived from adipose tissue cells that are multipotent and at least 1% of the culture expanded cells are positive for the surface marker 3G5 and one or both of the surface markers MUC18/CD146 and STRO-1.

2. The therapeutic composition of claim 1, wherein the MPCs co-express the marker alpha-smooth muscle actin.

3. The therapeutic composition of claim 1, wherein the MPCs co-express the marker STRO-1^bri.

4. The therapeutic composition of claim 1, wherein the MPCs co-express a marker selected from, but not limited to, the group comprising THY-1, VCAM-1, ICAM-1, PECAM-1, CD49a/CD49b/CD29, CD49c/CD29, CD49d/CD29, CD29, CD61, integrin beta 5, 6-19, thrombomodulin, CD10, CD13, SCF, STRO-1^bri, PDGF-R, EGF-R, IGF1-R, NGF-R, FGF-R, Leptin-R(Stro-2).

5. The therapeutic composition of claim 1, wherein the MPCs co-express the markers STRO-1^bri, MUC18/CD146, and alpha-smooth muscle actin.

6. The therapeutic composition of claim 1, wherein the MPCs are negative for the hematopoietic markers CD45, CD34, and glycophorin A.

7. The therapeutic composition of claim 1, wherein the MPCs are capable of forming a clonogenic colony and differentiating to three or more mesenchymal tissue types, isolated from a tissue of the group comprising, but not limited to, adipose tissue, teeth, dental pulp, skin, liver, kidney, heart, retina, brain, hair follicles, intestine, lung, spleen, lymph node, thymus, pancreas, bone, ligament, bone marrow, tendon, and skeletal muscle, and which are positive for the surface marker STRO-1^bri.

8. The therapeutic composition of claim 7 wherein the MPCs co-express the marker MUC-18/CD146 or alpha-smooth muscle actin.

9. The therapeutic composition of claim 7 wherein the MPCs co-express a marker selected from, but not limited to, the group comprising THY-1, VCAM-1, ICAM-1, PECAM-1, CD49a/CD49b/CD29, CD49c/CD29, CD49d/CD29, CD29, CD61, integrin beta 5, 6-19, thrombomodulin, CD10, CD13, SCF, STRO-1^bri, PDGF-R, EGF-R, IGF1-R, NGF-R, FGF-R, Leptin-R (STRO-2).

11. The therapeutic composition of claim 1, wherein the mammal is a human.

12. The therapeutic composition of claim 1 wherein the MPCs have the capacity to be induced to differentiate to form cells comprising one or more of at least osteoblast, odontoblast, dentin-producing, chondrocyte, tendon, ligament, cartilage, adipocyte, fibroblast, marrow stroma, osteoclast- and hematopoietic-supportive stroma, cardiac muscle, smooth muscle, skeletal muscle, pericyte, vascular, epithelial, glial, neuronal, astrocyte or oligodendrocyte cell type.

13. The therapeutic composition of claim 1, wherein the composition is enriched for MPCs capable of forming a clonogenic colony and differentiating to three or more mesenchymal tissue types.

14. in the therapeutic composition of claim 13, wherein the MPCs are additionally positive for the marker STRO-1^bri.

15. in the therapeutic composition of claim 13, wherein the MPCs co-express MUC18/CD146, alpha-smooth muscle actin, STRO-1^bri, and 3G5.

16. The therapeutic composition of claim 13, wherein the MPCs are negative for the hematopoietic markers CD34, CD45, and glycophorin-A.

17. The therapeutic composition of claim 13, comprising at least 0.01% MPCs capable of forming a clonogenic colony and differentiating to three or more mesenchymal tissue types.

18. The therapeutic composition of claim 13, comprising at least 0.1% MPCs capable of forming a clonogenic colony and differentiating to three or more mesenchymal tissue types.

19. The therapeutic composition of claim 13, comprising at least 1% MPCs capable of forming a clonogenic colony and differentiating to three or more mesenchymal tissue types.

20. The therapeutic composition of claim 13, comprising at least 0.01% STRO-1^bri MPCs.

21. The therapeutic composition of claim 13, comprising at least 0.1% STRO-1^bri MPCs.

22. The therapeutic composition of claim 13, comprising at least 1% STRO-1^bri MPCs.

23. The therapeutic composition of claim 13, wherein the MPCs have the capacity to be induced to differentiate to form cells comprising one or more of at least osteoblast, odontoblast, dentin-producing, chondrocyte, tendon, ligament, cartilage, adipocyte, fibroblast, marrow stroma, osteoclast- and hematopoietic-supportive stroma, cardiac muscle, smooth muscle, skeletal muscle, pericyte, vascular, epithelial, glial, neuronal, astrocyte, or oligodendrocyte cell type.

24. The therapeutic composition of claim 13, which comprises at least 0.1% cells which express at high levels one or more of the markers STRO-1, 3G5, or MUC18/CD146.

25. The therapeutic composition of claim 13, which comprises at least 1% cells which express at high levels one or more of the markers STRO-1^bri, 3G5, or MUC18/CD146.

26. The therapeutic composition of claim 13, which comprises at least 2% MPCs which express at high levels one or more of the markers STRO-1^bri , 3G5, or MUC18/CD146.

27. The therapeutic composition of claim 13, which comprises at least 5% cells which express at high levels one or more of the markers STRO-1^bri, 3G5, or MUC18/CD146.

28. The therapeutic composition of claim 13, which comprises at least 10% cells which express at high levels one or more of the markers STRO-1^bri, 3G5, or MUC18/CD146.

29. The therapeutic composition of claim 1, wherein at least 5% of the culture expanded cells are positive for the surface marker 3G5.

30. The therapeutic composition of claim 29, wherein the culture expanded cells that are positive for the surface marker 3G5 are also positive for the surface marker STRO-1.

10. The therapeutic composition of claim wherein the MPCs are negative for the hematopoietic markers CD45, CD34, and glycophorin A.