Ming Hao Zheng, PhD, DM, FRC Path Centre For Orthopaedic Research, The University of Western Australia; Paul Anderson MD Orthocell Pty Ltd

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1 Autologous Tenocyte Implantation for Refractory Tendonopathy Ming Hao Zheng, PhD, DM, FRC Path Centre For Orthopaedic Research, The University of Western Australia; Paul Anderson MD Orthocell Pty Ltd

2 Tendon rupture the most common sport injury Nirschl 1978 tendonosis/tendinopathy Non Inflammatory Condition Degenerative Process Chronic repetitive tendon loads 230,000 Achilles tendon injuries in USA, p.a, 60% increase last 10 years; AngioFibroblastic Hyperplasia David Beckham A 2 in 100,000 individual Achilles tendon injury rate increased to a 12 in 100,000 individual injury rate in less than 10 years.

3 Normal tendon Normal tendon

4 Tendon Pathology A B C D Tear human rotator cuff samples. A: Disruption of fiber structure and arrangement B: Adipose tissue deposit C: Vascular hyperplasia D: Rounding of nuclei

5 Cell Depletion leads to reduced collagen production, further ECM breakdown, tendon tear, impaired healing response

6 Can Autologous growth factors stimulate the proliferation of tenocyte in tendonopathy? Can Autologous Cell Therapy Improve Tendon Healing and Remodelling in Tendonopathy?

7

8

9

10 Blood do not improve pain and/or Function; PRP has a limited evidence on improvement.

11 JAMA 2010 Jan Stratified, block-randomized, double-blind, placebo-controlled trial; Blinded injection under ultrasound-5ml saline or PRP; Six month and 1 year follow up showed no effect on Visa-A score and ultrasounds.

12 Can Autologous growth factors stimulate the proliferation of tenocyte in tendonopathy? NO Can Autologous Cell Therapy Improve Tendon Healing and Remodelling in Tendonopathy?

13 Sources of cells for tendon regeneration Cell source Tenocytes Epitenocytes Dermal fibroblasts Undifferentiated cells Embryonic stem cells MSC from unbilical cord MSC from blood MSC from fat MSC from bone marrow Advantages Closest to host cell Progenitor cells of tendon, acceptable morbidity of donor site; Non aged-related Easy to recover; acceptable donor site morbidity True pluripotentiality Pluripotentiality Limited pluripotentiality, eazy Limited pluripotentiality, extensive study, reasonable pluripotentiality Disadvantages Age-related reduction; donor site morbidity; Required maturation; Non-homologous use; different lineage differentiation; Teratoma; Low cell No, lack of evidenced Very low cell No, unstable; Low cell No, unstable; higher cell No, reversed differentiation

14 Stem Cells June 2009 Stepwise Differentiation of Human Embryonic Stem Cells Promotes Tendon Regeneration by Secreting Fetal Tendon Matrix and Differentiation Factors Xiao Chen 1, Xing-Hui Song 1, Zi Yin 1, Xiao-Hui Zou 2, Lin-Lin Wang 1, Hu Hu 3, Tong Cao 4, Minghao Zheng 5, Hong Wei Ouyang 1 6 * Human stem cells make bone in tendon after long term implantation

15 Sources of cells for tendon regeneration Cell source Tenocytes Epitenocytes Dermal fibroblasts Undifferentiated cells Embryonic stem cells MSC from unbilical cord MSC from blood MSC from fat MSC from bone marrow Advantages Closest to host cell Progenitor cells of tendon, acceptable morbidity of donor site; Non aged-related Easy to recover; acceptable donor site morbidity True pluripotentiality Pluripotentiality Limited pluripotentiality, eazy Limited pluripotentiality, extensive study, reasonable pluripotentiality Disadvantages Age-related reduction; donor site morbidity; Required maturation; Non-homologous use; different lineage differentiation; Teratoma; Low cell No, lack of evidenced Very low cell No, unstable; Low cell No, unstable; higher cell No, reversed differentiation

16 Cell Characterisation Tenocyte Phenotype Maintenance The tenocyte phenotype is maintained when cultured in vitro NOTE: Increased intensity of in vitro bands due to elevated cell numbers Chen et al (Tiss Eng 2008)

17 Tendon progenitor cells are different from bone marrow MSC and skin fibroblasts. CD44 CD90 Stro-1 CD146 CD18 CD117 CD34 CD45 CD106

18 Figure 1. human tenocyte pellet cultured Chondrogenic medium Figure 2. human tenocyte cultured in osteogenic medium, stained by Alzarin Red

19 BT A U Scaffold only 8w MATT BT Scaffold+cell 8w

20 Autologous tenocyte therapy for Achilles Tendinopathy

21

22

23 Feridex IV cell tracking within the tendon

24 Improvement of Tensile strength By ATI A P<0.05 P<0.05 B

25 Autologous Tenocyte Implantation: the Perth clinical trial

26 Treatment criteria 1. Age: 30~75 Years old and diagnosed with tennis elbows using the following criteria: a. Pain over the outside of the elbow b. Pain when lifting objects c. Pain radiating down the forearm d. Positive of Mills test e. ultrasound confirmed tendinosis /MRI 2. History of tennis elbow longer than 6 months 3. Previously have received conservative treatment and not respond to the treatment 4. Has not received steroid injection within the last 3 months 5. Previously must not have surgical intervention for tennis elbow 6. Without any bone-muscular systematic disease like rheumatic disease, systemic lupus erythematous (SLE), ankylosing spondylosis. 7. Without any history of mental and psychological illness which might prevent patient from giving valid consent or prevent them completing the rial. 8. Patient with pregnancy will not be included. 9. Professional sportsperson will not be included.

27 Clinic Patients Recruitment x 30 Pre-treatment Evaluation Epitendineum Tendon Tissue Harvest Subjective evaluation: Quick DASH UEFS ASES Objective evaluation: MRI Imaging Donor Site Adverse Event Evaluation Orthocell Laboratory Tenocytes Culture Autologous Tenocytes Injection Safety & Tolerability Evaluation 4h,1d,7d, 4,12,24week & 12 months Clinic Received Site Adverse Event Evaluation Early Phase Efficacy Evaluation 4,12,24 week & 12 months Subjective evaluation: Quick DASH UEFS ASES Objective evaluation: MRI Imaging

28 Adverse Events Death or Permanent Disability Systematic Anaphylactic Respond Tendon Rupture Recipient Site Infection Stoping rule Temporary Stop Recruiting Procedures Reviewing Process (By Clinicians and Independent Reviewers) Problem resolved Restart the Trial Problem NOT Resolved Permanent Stop of the Trial

29 Autologous Tenocyte Therapy in Lateral Epicondylitis

30 Autologous Tenocyte Therapy in Lateral Epicondylitis 25 patients completed 1 yr Safety Tolerability No complications of tendon Biopsy No infection/ excessive fibroblastic reaction at injection site

31 Score Quick DASH DASH Initial 4 week 12 week 26 week 52 week Time

32 Score Upper Extremity Functional Scale UEFS Initial 4 week 12 week 26 week 52 week Time

33 Score Mean Pain Score when at worst Max Pain Initial 4 week 12 week 26 week 52 week Time

34 Mean % Increase Grip Strength Time 120 Mean Percentage Increase in Grip Strength from Initial Assessment Grip Strength % % % Percentage Increase Initial 4 week 12 week 26 week 52 week GS_IA-1 month GS_IA-3 month Score GS_IA-6 month Assessment

35 Pain / 10 Average Pain Score following Biopsy Pain Level hr 1day 1wk 4wk 12wk 26wk Assessment

36 Spearman s Correlation DASH UEFS Max Pain Grip Strength DASH (p=0.000) (p=0.000) (p=0.000) UEFS (p=0.000) (p=0.000) (p=0.002) Max Pain (p=0.000) (p=0.000) (p=0.000) Grip Strength (p=0.000) (p=0.002) (p=0.000) 1

37 Autologous Tenocyte Therapy in Lateral Epicondylitis Pre Injection 6m Post Injection

38 Autologous Tenocyte Therapy in Lateral Epicondylitis Pre Injection 6m Post Injection

39 Indication Expansion

40 Conclusion Tendon degeneration is due to depletion of cell pool of functional Tenocytes. Pre-clinical studies showed that Autologous Tenocytes are valuable for tendon repair. First trial of ATI is completed. Safety and level of efficacy are evidenced. Level I randomised double blinded Study for Achilles Tendonopathy

41 Acknowledgment Scientists: Drs Jim Chen, Ben Wu, Craig Willer, Vincent Qin, Jiake Xu, Tim Auckland Surgeons: Allan Wang; Greg Janes, Lars Lidgren, Fiona Wood, David Wood, Ray Crowe Radiologist: Bill Briedhal Mr Paul Anderson, MD of Orthocell.

42 Thank you

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