This document summarizes several case studies using mesenchymal stem cells (MSCs) for regenerative medicine applications: 1. A pre-clinical study using ovine MSCs implanted in a critical-sized bone defect in sheep tibias showed effective bone regeneration compared to autograft or no cell controls. 2. A large animal study treating induced osteonecrosis of the femoral head in sheep with ovine MSCs implanted at the lesion site showed persistence of cells differentiating into bone and effective treatment compared to controls. 3. A Phase I/IIa clinical trial in humans with osteonecrosis of the femoral head treated with implanted human MSCs showed safety and potential efficacy based on imaging follow-up

1. TERÀPIES AVANÇADES
Dr. Joaquim Vives

2. Advanced Therapies
Cells (cell therapy)1
Tissues (tissue engineering)3
Genes (gene therapy)2

3. Advanced Therapies

4. Advanced Therapies
 No immunogenicity
 1 donor/patient, 1 dose
 Expensive
 Time-consuming
 1 donor, several doses
 Reasonable cost
 Off-the-self
 Possible immunogenicity

5. Regenerative Medicine
Sharpe et al. in: Cell Therapy GxP (2015) [Vives&Carmona eds]

6. The cellular basis
SELF-RENEWAL
DIFFERENTIATION
STEM CELL

7. Potentiality

8. Development

9. Potentiality&Development
Pluripotent: ES cells
Totipotent: zygote
Multipotent: HPC
MSC
Oligopotent : lymphoid stem cells
Unipotent : germ cells
Terminally differentiated cells. i.e. Fibroblasts
+POTENTIALITY-
DEVELOPMENT

10. Blastocyst
From Dr. L. Batlle-Morera, CRG

11. 11
EMBRYONIC STEM CELLS

16. Fetus

17. 17
NEURAL STEM CELLS
9

18. Adult organism

19. MESENCHYMAL STROMAL CELLS

20. Qualities of MSC for clinical use
20
Reis et al (2016) Front Immunol 7, 500 Nombela-Arrieta et al (2011)
Nat Rev Mol Cell Biol 12, 126

21. From Dr. L. Rodríguez (BST)
Dominici, M. et al. (2006). The International
Society for Cellular Therapy position statement.
Cytotherapy 8, 315–317.

23. 23
Wagner, W. et al. (2005). Exp. Hematol. 33, 1402–1416

24. Neonatal tissue
UC
AI: UC-MSC
Mechanical
processing

25. Neonatal tissue
Oliver-Vila & Coca et al (2015) Cytotherapy

26. Potentiality

27. Potentiality&Development
Pluripotent: ES cells
Totipotent: zygote
Multipotent: HPC
MSC
Oligopotent : lymphoid stem cells
Unipotent : germ cells
Terminally differentiated cells. i.e. Fibroblasts
+POTENTIALITY-
DEVELOPMENT

28. Waddington’s epigenetic landscape

32. iPSC
32

33. Applications
BASIC RESEARCH
DISEASE MODELLING
DRUG SCREENING

34. Applications

35. Applications
Cherry & Daley (2012) Cell 148, 1110

36. Cell Culture Technology
Sart et al (2014) Cytotechnology 68,709

38. Cell Culture Technology
Robey et al (2015) Bone 70,87

39. Cell Culture Technology

40. Cell Culture Technology

41. Overview of BST’s Advanced Therapies
Auto BM-MSC
Gonarthrosis
Degenerative meniscus
Combined
with bone
scaffolds
ONFH
Spinal Fusion
Pseudoarthrosi
sCriopreserved Multiple
Sclerosis
Spinal cord
injury
GvHD
Cardiomyopathy
Proximal femoral
fractures in the
elderly
Combined with
bone scaffolds
Deficiency of specific
immunity
Allo BM-MSC
(cryopreserved)
Allo WJ-MSC
(cryopreserved)
Virus-specific
activated T cells
Auto
Allo

42. Product development overview
Caminal et al (2014) New
Biotech 31, 492
Codinach et al (2016)
Cytotherapy 18, 1197
Soler et al (2016) Knee
23, 647
Vives et al (2015)
Cytotherapy 17, 1009
Vives et al (2015) BMC Proceedings 9, O9

43. BACKGROUNDOn-going Autologous BM-MSC-based CT at BST
Femoral head osteonecrosis
• Study name: XCEL-MT-10-01
• EudraCT: 2010-023998-18
• Phase: I-II b
• Number of participants: 23/24
• Finished; pending results
Spinal fusion
• Study name: XCEL-MT-10-02
• EudraCT: 2010-023999-12
• Phase: I-II b
• Number of participants: 66
• Finished; pending results
Pseudo arthrosis
• Study name: XCEL-PSART-01
• EudraCT: 2013-005025-23
• Phase: I-II b
• Number of participants: 19/20
• Finished; pending results
MSC
sources
XCEL-MT-OSTEO-
ALPHA

44. TERÀPIES AVANÇADES
Dr. Joaquim Vives

45. Case study
GLP GMP Study description Experimental system
Dose; Route of
administration
X
Proof of concept: Bone regeneration in a model of
critical-sized segmental tibial defect
Ovis aries
(Ripollesa breed, ♀); critical
sized defect in tibia
5.8x106 ±0.1x106
oMSC/cc; implant
Pilot study: Bone regeneration in a model of
femoral head osteonecrosis
Ovis aries
(Ripollesa breed, ♀);
avascular ONFH
5.4x106 ±0.3x106
oMSC/cc; implant
X X
Biodistribution of hMSC in an immunodeficient
mouse model
Mus musculus
(NRG, ♀ and ♂)
4x105 hMSC; tail-vein
injection
X X
Analysis of protooncogen expression levels,
hTERT activity, senescense, G-banding karyotype,
and CGH arrays on clinical grade hMSC
In vitro assays N/A

46. Case study
o Experimental groups :
 Autograft (gold standard)
 Bone construct without
cells (control without cells)
 XCEL-MT-OSTEO-ALPHA
(ovine) (test item)
o Procedures:
 Critical-sized defect (=2.5xØ cm)
in tibia.
 Euthanasia at 3 months post-
treatment.
 Clinical follow-up and
histological/biomechanical
analysis of the tibias.
o Conclusions:
 Large animal model.
 Extreme conditions.
 Safe.
 Effective as an alternative to
autologous bone, avoiding the
associated morbidity to the gold
standard.
Score histology
Gold
standard
(I)
Without
oMSC
(II)
With
oMSC
(III)
P < 0,1
Biomechanical Analysis

47. Case study
o Experimental groups:
 A: Core decompression
(gold standard)
 B: Bone construct without
cells (control without MSC)
 C: XCEL-MT-OSTEO-
ALPHA (ovine) (test item)
o Procedures:
 Induced ONFH 6 weeks pre-
treatment.
 Euthanasia 3 months post-
treatment.
 Clinical follow-up and histological
analysis of the femoral heads.
o Conclusions:
 Large animal model.
 Non-invasive techniques for the
clinical follow-up.
 Safe.
 Effective.

48. Case study
o Experimental groups:
 XCEL-MT-OSTEO-ALPHA
(eGFP labelled-ovine MSC)
o Procedures:
 Induced ONFH 6 weeks pre-
treatment.
 Euthanasia 3 months post-
treatment.
 Clinical follow-up and histological
analysis of the femoral heads.
o Conclusions:
 Cells applied differentiate into
bone and persist.

49. Case study
GMP GCP Study description Experimental system
Dose; Route of
administration
X X
Mesenchymal Stem Cells in Osteonecrosis of the
Femoral Head: a Phase I/IIa Clinical Trial (EUDRA-
CT:2010-023998-18; ClinicalTrials.gov Id: NCT01605383)
Human (♀ and ♂); ARCO grade I
or II ONFH
3-10x105 hMSC/cc of
bone; impant

50. Case study
TEP (F18)
Pre-Op 6 m 12 m
MRI