Current Concepts in Cartilage Restoration

COA Center For Joint Preservation May 22, 2016 Disclosures

• I am paid consultant on behalf of Vericel and currently a (KOL) Key Opinion Leader on their behalf Overview

• Articular cartilage disease • Traditional treatments ● Debridement ● Bone-marrow stimulation ● Augmented microfracture (Mfx Plus & Mfx Plus Plus) ● Osteochondral/chondral allograft ● Cell-based repair • Next generation cellular therapy • Emerging technologies

| 3 Articular Cartilage Has Little Intrinsic Ability 1 Water to Repair Itself 6 Cartilage Composition 65%-80%

• Inadequate compensation for cell death or damage2 Chondrocytes Collagen ● Mature chondrocytes rarely divide 1%-5% 10%-15% ● No direct cell supply due to avascular nature Proteoglycans 10%-15% • Healing depends on migration of stem cells from subchondral bone marrow or synovial tissue2 • Role of progenitor cells (mostly in superficial and middle zones) in repair is unknown3,4 • Repair cartilage is inferior to normal cartilage5

1. Minas T. J Bone Joint SurgBr. 2012;94:141-146. 2. Minas T, Glowacki J. In: A Primer in Cartilage Repairand Joint Preservationof the Knee. 2011;8- 21. 3. Pretzel D, et al. Arthritis Res Ther. 2011;13:R64. 4. IwamotoM, et al. Birth Defects Res C Embryo Today. 2013;99:192-202. 5. Pylawka TK, et al. In: Sports Medicine. 2006;418-429. 6. Bhosale Am, Richardson JB. Br Med Bull. 2008;87:77-95. | 4 Untreated Chondral Defects May Lead to a Cycle of Cartilage Degradation and Eventual OA

Weight-bearing shifts to defect shoulders Overload leads to progressive Eventual OA defect enlargement

Untreated defect Cartilage Enzymes cause breakdown further products can degradation trigger synovitis Synovitis liberates catabolic enzymes

Minas T. J Bone Joint Surg Br. 2012;94:141-146. OA = osteoarthritis. Hx & Genetics! | 5 Treating Cartilage Injuries

| 6 Many Options Are Available for Treating Focal Cartilage Defects

MICROFRACTURE OSTEOCHONDRAL DEBRIDEMENT AUTOGRAFT AUTOLOGOUS AND LAVAGE CHONDROCYTE IMPLANTATION OSTEOCHONDRAL ALLOGRAFT

Bentley G, et al. Injury. 2013;44(Suppl1):S3-S10. Image of debridement courtesy of Dr. Brian Cole; images of microfracture, osteochondral autograft, and osteochondral allogaft courtesy of Dr. Christian Lattermann; image of autologous chondrocyte implantation courtesy of Dr. Jack Farr. | 7 Treatment Goals Are to Reduce Symptoms, Improve Function, and Prevent Degeneration1

Palliative1,2 Reparative1-4 Restorative3,5 Techniques that Marrow-stimulation Techniques that relieve or prevent techniques that attempt to recreate pain with little repair result in formation hyaline cartilage at of underlying defect of fibrocartilage the defect site

• Lavage and • Microfracture • Autologous debridement • Augmented chondrocyte therapy • Thermal microfracture • Osteochondral chondroplasty • Microdrilling autograft or allograft

Nonoperative management should be the first-line option for most patients1

1. PylawkaTK, et al. In: Sports Medicine. 2006;418-429.2. Tetteh ES, et al. J Orthop Sports PhysTher. 2012;42:243-253. 3. Kane P, et al. Phys Sportsmed. 2013;41:75-86. 4. Bentley G, et al. Injury. 2013;44(Suppl1):S3-S10. 5.Ebert JR, et al. Osteoarthritis Cartilage. 2008;16:1131-1140. | 8 Several Factors Should Be Considered When Developing an Individualized Treatment Plan1,2

• Age and *Weight3 • Expectations3 Patient • Compliance,3 including rehabilitation4 • Functional level3 • Comorbidities3

• *Chronicity of Lesion, size, and site3 • Underlying contributors to injury4 Injury • Malalignment, ligament instability, meniscus deficiency4

* Obesity and previous knee injury has the highest association with lesion progression 1. Tetteh ES, et al. J Orthop Sports PhysTher. 2012;42:243-253. 2.Kane P, et al. Phys Sportsmed. 2013;41:75-86. 3. Marcacci M, et al. Injury. 2013;44:S11-S15. 4. Gomoll AH, et al. J Bone Joint SurgAm. 2010;92:2470-2490. Alentorn-GeliE. AJSM | 9 Treatment Considerations Include Defect Location and Size, Activity Level, and Prior Treatment Primary Secondary Defect Site Defect Size Treatment Treatment

2 MFX Autograft <2–3 cm Autograft ACI/MACI Femoral Condyle MFX Malalignment Autograft ACI/MACI Meniscal Deficiency >2–3 cm2 ACI/MACI Allograft ACL-PCL Allograft Cartilage Non-Operative Physical LOW HIGH Defect Rehabilitation Demand ACI/MACI MFX Patellofemoral <2–3 cm2 Autograft ACI/MACI alignment Allograft

Patellofemoral >2–3 cm2 ACI/MACI ACI/MACI Allograft Allograft

Adapted from Tetteh ES, et al. J Orthop Sports Phys Ther. 2012;42:243-253 and Cole BJ, et al. J Bone Joint Surg Am. 2009;91:1778-1790. ACI = autologous chondrocyte implantation; ACL = anterior cruciate ligament; MACI = matrix-induced autologous chondrocyte implantation; MFX = microfracture; PCL = posterior cruciate ligament. | 10 Arthroscopic Debridement

| 11 Arthroscopic Debridement

One-step procedure during which damaged cartilage and mediators are removed (usually after lavage)1 • Shaving of rough cartilage and/or smoothening of degenerated meniscus2 • May include osteophyte removal, local synovectomy2 3 • Palliative, not reparative Improves Congruence • May provide temporary symptom relief3 • Less invasive initial treatment for symptomatic focal chondral defects4

1. Krych AJ, et al. Sports Med Arthrosc Rev. 2013;21:23-30. 2. Tetteh ES, et al. J Orthop Sports PhysTher. 2012;42:243-253. 3. Kane P, et al. Physician Sportsmed. 2013;41:75-86. 4. Minas T. J Bone Joint SurgBr. 2012;94:141-146. Image courtesy of Brian Cole, MD. | 12 Bone-Marrow Stimulation/BMS

| 13 Microfracture; MFX

● Unstable and calcified cartilage is removed to form a vertical edge around the defect1 ● An awl is used to create holes in the subchondral bone plate1 ● 3-4 mm apart, 2-4 mm deep ● Blood and fat from the marrow fill the lesion and form a stable “superclot”1,2 ● MSCs in the marrow blood differentiate into fibrochondrocytes2,3 ● Cells make fibrocartilage (not hyaline) 1. Steadman JR, et al. ClinOrtho Relat Res. 2001;(391 Suppl):S362-369. 2. Tetteh ES, et al. J Orthop Sports PhysTher. 2012;42:243-253. 3. Richter W. J Intern Med. 2009;266:390-405. MSC = mesenchymalstromal cells. Image courtesy of Dr. Richard Steadman. | 14 Nanofracture and Microdrilling Are Two Newer Methods of MFX

MFX1,2 MD2 NANOFX1 SUBCHONDRAL BONE PLATE

SUBARTICULAR SPONGIOSA

1. Chen H, et al. J Orthop Res. 2009;27:1432-1438. 2. Benthien and Behrens. IntOrthop. 2013;37:2139-2145. MD = microdrilling; MFX = microfracture; NanoFX = nanofracture. Image adapted from Nanofracture Clinical Monograph; http://www.arthrosurface.com/wp- content/uploads/2013/05/NanoFx_ ClinicalMonograph1.pdf. Accessed March 13, 2014. | 15 Micro CT- Standard MFX vs Nano vs k-wire

COPYRIGHT Walsh WR et al, 2015 Nanofracture and Microdrilling Have Not Been Formally Investigated in Humans1-3

MD vs. MFX With Continuous Irrigation in a Rabbit Model1,2 These findings1,2 MD support the rationale • No heat necrosis1 for creation of the • Greater subchondral hematoma1 NanoFX system • A 1-mm needle is MFX advanced through a • 2x the rate of osteocyte necrosis1 cannulated pick to a • Bone compacted around holes1 depth of 9-mm MD 6-mm depth resulted in better outcomes than MD or MFX 2-mm depth2

1. Chen H, et al. J Orthop Res. 2009;27:1432-1438. 2. Chen H, et al. J Orthop Res. 2011;29:1178-1184. 3. Benthien and Behrens. IntOrthop. 2013;37:2139-2145. MD = microdrilling; MFX = microfracture; NanoFX= nanofracture. | 17 Summary: MFX

● One-stage procedure with a low level of morbidity ● Consistent short-term functional improvement ● Outcomes decline over time ● Limitations include ● Unknown number of MSCs recruited ● Unpredictable fill rate ● Violates subchondral bone and changes architecture ● NanoFX and MD are newer techniques ● Both use deeper holes ● Not yet formally studied in clinical trials

MD = microdrilling; MFX = microfracture; MSC = mesenchymal stromal cell; NanoFX = Nanofracture.

| 18 Augmented Microfracture

| 19 Why do we need to augment MFX Procedures?

• MFX is the most frequently used first-line surgical treatment (130-150k procedures per year) • Straight forward procedure- penetration of the SB to elicit bleeding response ; BMS follows the normal wound healing sequence1 • MFX has demonstrated inconsistent filling of the defect2 Key Point with all biologics- Fill rate is one of the predictors for outcome • Fills with a poor quality tissue and produces highly variable outcomes2 There is an instability of the BM derived clot2 • There is shrinkage and detachment of the platelet-driven clot reaction (Maybe as high as 50%)3 4 • Functional decline with time – 24 to 36mths5 • The critical component of the initial blood clot is to have more abundant BMSCs producing a more adherent voluminous clot6

1-Chevrier et al. Osteoarthritis Cartilage 2007 Mar:15(3) 316-327; 2-Mithoefer K, et al AJSM 2009;37:2053-2063 3- Hoemann et al. JBJS Dec: 87(12) 2671-2686; 4-Johnson et al. Arthroscopy 1991:7 (1) 14-23; 5- Kreuz et al. Osteoarthritis and Cartilage (2006) 14, 1119-1125.; 6-Chevrier et al. Osteoarthritis Cartilage 2011 Jan:19(1) 136-144 | 20 Different Types of Augmented MFX Have Been Reported1-5

HA injections are administered intra- articularlyafter MFX1

Chitosan-GP gel scaffold is applied to dry defect after MFX to stabilize clot2

+ BIOCARTILAGE Micronized cartilage matrix is mixed with MFX PRP and administered after MFX3

MFX site covered with a natural collagen type I/III membrane4

MFX site is covered with cartilage allograft mesh primed for MSC activity5

1. Gunes T, et al. Cartilage. 2012;3:20-26. 2. Stanish WD, et al. J Bone Joint Surg Am. 2013;95:1640-1650. 3. Abrams GD, et al. OperTech Sports Med. 2013;21:116- 124. 4. Anders S, et al. Open Orthop J. 2013;7:133-143. 5. Cartiform Viable Cartilage Mesh. http://www.osiris.com/biosurgery_cartiform.php. Accessed March 11, 2014. AMIC = autologous matrix induced chondrogenesis; HA = hyaluronic acid; MFX = microfracture; MSC = mesenchymal stromal cells; PRP = platelet-rich plasma. | 21 Microfracture Plus- HA

● 3 mos: ● gross and histologic eval of repair tissue showed 3 IJ group had a significantly better fill rate and tissue then 5 IJ; yet, not significantly different vs control ● 6 mos: ● the 3 cohorts were not significantly different

*** key finding- two HA groups had significantly less joint degeneration vs the control group *** Microfracture Plus- BST-CarGel

• 80 patients received MFX alone or with BST-CarGel • Outcomes at 1/5 years ● Fill: 92.8/93.8% with BST-CarGel vs. 85.5/87% with MFX (P=0.017) ● Both groups had similar improvement in WOMAC and SF-36 scores at 5yrs (P<0.48/0.33)

Stanish WD, et al. J Bone Joint Surg Am. 2013;95:1640-1650. MFX = microfracture; WOMAC = Western Ontario and McMaster Osteoarthritis Index. Shive MS, et al. Cartilage. 2015;6:62-72. | 23 Microfracture Plus- BioCartilage

Micronized allograft cartilage implanted into a cartilage defect of Baboon

One week Four weeks

Newly formed Cells Newly GAG content Nine weeks Nine weeks

Material presented by Thomas Temple, University of Miami Tissue Bank 90% of baboons had full regeneration at 9 weeks; no PRP or MFX

Malinin T, et al, Induction of regeneration of AC defects by freeze dried particulate cartilage allografts, ICRS, 2009 Meeting; poster presentation. Microfracture Plus Plus- BioCartilage & ACP

• Scaffold for PRP/BMAC Delivery ● Gets mixed in a syringe with PRP ● Paste is injected into dried defect bed after MFX Tourniquet released ● Sealed with fibrin glue • No data from human trials • Equine model positive fill

Abrams GD, et al. Oper Tech Sports Med. 2013;21:116-124. MFX = microfracture; PRP = platelet-rich plasma. Image: BioCartilage Product Brochure, Arthrex, Inc; 2014. | 25 Why mix with PRP/BMAC?

● 5 IJs of ACP post MXF of full-thickness cartilage injuries: macroscopically, histologically and biomechanically better than MFX alone after 3, 6, & 12 mos

Milano et al. Arthroscopy May;28(5) 2012 688-701; Milano et al Osteoarthritis and Cartilage July 18(7) 2010 971-80 Clinical Studies none, merely case series and cohorts

• SOS Knee ● Multi-centered, 10 pts per site ● Pre-op, 3, 6, 12, 18, 24, 36, 48, 60 mos ● Subjective based outcomes • BioCartilage vs MFX (U of Missouri) ● Matched cohorts: trochlear groove vs FCs • SOS Ankle ● After the knee ***Further studies will delineate optimal indications & efficacy*** Microfracture Plus- AMIC; Autologous Matrix-Induced Chondrogenesis ● The theory is that by combining a MFX with a collagen membrane scaffold it will encourage attachment and development of BMSCs, allowing them to form a stable repair tissue. ● Trying to achieve: -Improved fill capacity -Improved integration- Clot retraction -Activate chondrogenic differentiation

The importance of AMIC, is that it shows introducing A collagen membrane helps to secure the fibrin clot

***Currently, there is no data to suggest that AMIC is superior to MFX alone, nor is there any data to indicate higher quality repair tissue.***

| 28 A RCT Showed Similar Pain and Function Scores With AMIC vs. MFX 150 MFX

100

50

Cincinnati Score 0 150 150 BL Year 1 Year 2

100 Sutured AMIC 100 Glued AMIC

50 50

0 0 BL Year 1 Year 2 BL Year 1 Year 2 Cincinnati Score Cincinnati Score Quality of the regenerate surface and defect cover were similar in all groups

Anders S, et al. Open Orthop J. 2013;7:133-143. AMIC = autologous matrix induced chondrogenesis; BL = baseline; MFX = microfracture.

| 29 Microfracture Plus- Cartiform

• CVOCA- Viable hyaline cartilage mesh primed to optimize interactions with MSC ● Available off the shelf; 2-year shelf life ● 10 or 20mm round discs • After MFX ● Cut material to desired size and shape ● Affix with fibrin glue or sutures ● Cartiform alone does not affect subchondral bone ● No published safety or outcomes data are available ● one goat model is the scientific support Cartiform Viable Cartilage Mesh. http://www.osiris.com/biosurgery_cartiform.php. Accessed March 11, 2014. MFX = microfracture; MSC = mesenchymal stromal cells. Image: CartiformWebsite. | 30 Microfracture Plus Plus- Cartiform & BMAC

• MFC & trochlea • Cartiform grafts soaked in BMAC • BMAC then gelled with Thrombin, CaCl • Grafts sutured c, 6-0 vicryl as needed, cut to fit large defect

Courtesy Scott D. Gillogly, MD Clinical Studies none, merely registry

• Registry • 200 cases- avg 2.4cm2 • Frozen at -80C • 70.5-78% viability • Releasing factors which promote migration of SCs • 65% c mfx, 35% s mfx (BMAC) • 39% c sutures ***Further studies will delineate optimal indications & efficacy***

A novel, cryopreserved, viable osteochondral allograft designed to augment marrow stimulation for articular cartilage repair. Geraghty S, et al. J Orthop Surg Res. 2015 May14;10(1):66. doi: 10.1186/s13018-015-0209-5. PMID: 25968127 Summary: Augmented MFX

• Hypothesized to stabilize the clot, reinforce clot formation by increasing adhesiveness and preventing platelet-derived retraction • Various methods of augmenting MFX are available or in development • Various methods of MFX Plus Plus employed • Data on MFX augmentation are limited: other published studies using different techniques- Dhollander 20121 and Siclari 20122

AMIC = autologous matrix induced chondrogenesis; MFX = microfracture. 1- Dhollander 2012 Knee Surg Sports Traum Arthro Sept 20;(9) 1773-80 2-Siclari 2012 CORR Mar;470(3)910-9 | 33 OATS Procedure

| 34 OATS Outcomes

• Multiple studies show good clinical outcomes • Great return to play data • Limited to size of lesion (auto) • <2 cm?

| 35 Osteochondral Allograft

| 36 Plentiful prospective case series reporting “good to excellent” results for the knee

| 37 19 studies / 644 knees / 93% follow-up 40% traumatic / 30% OCD / 30% other Mean follow-up: 5-years

| 38 •Satisfaction Rate: 86% Overall

| 39 •Little or no arthritis at final follow-up- 65%

| 40 • “Predictably favorable outcomes and high satisfaction - rates at intermediate follow- up” unfortunately, the authors failed to mention which type of grafts were used in these studies- the initial fresh grafts or today’s fresh refrigerated grafts???

| 41 • Future studies should include control groups and established outcome instruments

| 42 Shahal et al

• 38 patients- Military personnel underwent osteochondral allograft ● 28.9% (11/38)- returned to full duty ● 28.9% (11/38)- returned to limited duty ● 42% (16/38)- unable to return to pre-injury level of work

| 43 Osteochondral/Chondral Allograft Engineered plugs

In principle similar to an autograft: Defect is filled with a bone/cartilage composite from a prolonged fresh specimen, or engineered allograft

| 44 Acellular treatment options Scaffold guided tissue regeneration

• Hypothesis is that BMSCs and progenitor cells from the subchondral bone marrow blood (after surgical curettage) are able to migrate inside Cell Guidance and colonize the scaffold • Differentiate along either a chondrogenic or osteogenic lineage based upon the particular physical and chemical gradients composition found Peptides & Growth Factors

Rita Kandel, Bioengineering Conference 2013 Cell-free bi-phasic plugs

Trufit MaioRegen Chondrofix Agili-C

Removed from May enter US? May enter US? Market

Preserved OC Allograft

Poly(D,L-lactide-co-glycolide) Porous three-dimensional composite Calcium sulfate three-layer structure with varying levels of hydroxyapatite and collagen Bone phase composed of calcium carbonate Cartilage region- Collagen I 100% in the aragonite crystalline form, Cartilage Transition region- Hydroxy-40%/Collagen-60% phase is a composite of modified aragonite Bone region- Hydroxy-70%/Collagen-30% and hyaluronic acid (HA). MaioRegen

• Off-the-Shelf Implant • Multi layer scaffold. ● The superficial layer consists of deantigenated type 1 equine collagen and resembles the cartilaginous tissue, while the lower layer consists mostly of Mg-HA and simulates the subchondral bone structure. The middle layer, composed of Mg-HA and collagen, reproduces the tide mark. The bone phase of the implant is composed of calcium carbonate in an aragonite crystalline form, and the cartilage phase is composed of modified aragonite and HA. • Conducting human trials now in Europe • US if proceed with FDA trial

47 Chondrofix

• Engineered bone and cartilage allografts1,2 1 Cartilage cells are • Readily available off the shelf killed in sterilization Bone ● 2 year shelf life ● Eliminates issues with donor tissue Methylene Blue availability and safety2 ● Sterile packaging of decellularized cartilage and bone1 ● 4 diameters (7, 9, 11, 15, 20 mm) with a 10-mm length1 • Prospective clinical trial terminated 1. Bugbee W, et al. J Knee Surg. 2012;25:109-116. 2. GomollAH. Oper Tech Sports Med. 2013;21:90-94. Images courtesy of Dr. Andreas Gomoll.

| 48 Chondrofix 2014 update from LifeNet Meeting • No cysts • Faster recovery and RTP with rapid bone to bone integration (5 to 6 months ) vs fresh osteochondral allografts • 3 pts IKDC 45-80 3 years fu • Approved more if arthro proc • Will chondrofix be tx option for early OA as opposed to focal chondral lesions? Smart Scaffolds for Cell Recruitment • Off-the-Shelf Implant for Hyaline Cartilage Regeneration • Agili-C is an osteochondral bi-phasic implant. ● The bone phase of the implant is composed of calcium carbonate in an aragonite crystalline form, and the cartilage phase is composed of modified aragonite and HA. • Implanted in a press-fit, single-stage procedure. • Animal studies demonstrate excellent bone:cartilage healing • Conducting human trials now in Europe; N=126: 9 24mo PRO, 10 24mo MRI • US in 5 years perhaps if proceed with FDA trial

50 Stimulates Cartilage Regeneration • Invasion and ingrowth of chondrocytes from the periphery, (as well as BMSCs) adhesion, proliferation and differentiation into chondrocytes

• Cartilage flow from the implant periphery towards the center is enhanced by cartilage formation from the chondral phase of the implant, in vivo (caprine model)

51 DeNovo NT Particulated Juvenile Cartilage Allograft • Prepared from donors younger than 13 years1,2 • 3D volume-stable construct1,2 • Cartilage manually minced into 1-cm3 cubes1 ● High chondrocyte viability2 MINCED CARTILAGE ● Secured in defect with fibrin glue2 • Advantages include ● Easier to obtain than OC allograft ● Off-the-shelf product- not really1,2 ● Single-stage procedure2,3 1. Farr J, Yao JQ. Cartilage. 2011;2:346-353. 2. Tompkins M, et al. Oper Tech Sports Med. 2013;21:82-89. 3. Tompkins M, et al. Arthroscopy. 2013;29:1661-1670. Image courtesy of Dr. Kevin Bonner. | 52 Clinical Studies

• Retrospective study (n=16; 2 bilateral) with mean follow-up of 28.8 months Outcome Measure Mean Score ± SD IKDC 73.3 ± 17.6 Pain 84.2 ± 14.2 Symptoms and stiffness 85.0 ± 12.3 KOOS Activities of daily living 88.9 ± 12.9 Sports and recreation 62.0 ± 25.1 Quality of life 60.8 ± 28.6 Tegner 5 (median; range 3–9) VAS pain 1.9 ± 1.4 ● 73% (n=11) knees had normal/nearly normal repair tissue; mean defect fill, 89% (3 knees with >90% fill) Tompkins M, et al. Arthroscopy. 2013;29:1661-1670. IKDC = International Knee Documentation Committee; KOOS = Knee Injury and Osteoarthritis Outcome Score; VAS = Visual Analog Scale. | 53 ● 5 yr tech note from Post market study Cell-Based Cartilage Repair

| 55 Rationale of Using Cultured Chondrocytes for Cartilage Repair

Science Clinical

• Adult chondrocytes can • 4 Year FDA designed redifferentiate up to 3 passages prospective study showed 77% • Animal models have shown the successful outcome after failing implanted chondrocytes to contribute to the repair tissue initial surgical tx • Repair tissue can produce a • 10 Yr Multi-ctr Registry Study repair tissue with hyaline tissue characteristics • 15 Year Brigham Publication • Chondrocytes are the sole cell responsible for producing ECM

“MSCs do not make cartilage!” Arnold Caplan 2015 ICRS & TOBI Meetings

| 56 Autologous Chondrocyte Implantation (ACI)

● Requires rigorous testing before release ü Sterility ü Endotoxins ü Viability ü Identity ü Morphology

Biological Treatment option developed in Goteburg Sweden by Prof’s, Lars Peterson, Anders Lindahl and Mats Brittberg: Using cultured Chondrocytes to develop a durable repair tissue to treat large chondral lesions. Tom Minas completed much of the early scientific and clinical work in the US Brittberg et al. Treatment of Deep Cartilage Defects in the Knee with Autologous Chondrocyte Transplantation, NEJM 1994 ACI- Literature Review- ACI vs Microfracture

• Kreuz et al. 2006 ● 85 patients, average age 39.5 yrs, full thickness cartilage lesions ● All patients underwent microfracture procedures ● All patients had improved ICRS and Mod. Cincinnati scores at 18 months • Significant deterioration in ICRS score after 18 months ● MRI of patients with condyle lesions had best fillèbest clinical scores ● Even in small defects patients failed to see improvements as early as 6 month • Saris et al 2008- ● 118 patients, avg age 33.9 yrs, 61 Microfracture, 57 CCI (ACI) ● Both groups showed significant improvement in clinical outcome short- term ● CCI (ACI) showed superior structural regeneration of cartilage at 1 yr • Superior structural outcome may result in improve long-term clinical | 58 outcome Improvements Have Been Made to ACI Since Its Introduction In the US- the FDA Label requires ACI to be used with a periosteal flap. MACI is not FDA approved in the US, but has received EMA approval

Complications related to Improvements include3 periosteal flap include1 • Less invasive procedure • Hypertrophy • Shorter operative time • Detachment • Less postoperative pain • Delamination • Less surgical-site morbidity • Faster recovery

ACI – 1st Generation MACI– 3rd Generation Cells injected beneath Chondrocytes seeded in a a periosteal flap2 type I/III membrane4

1. Marlovits S, et al. Eur J Radiol. 2006;57:24-31. 2. Bentley G, et al. Injury. 2013;44:S1-S10. 3. Brittberg M. Am J Sports Med. 2010;38:1259-1271. 4. Cortese F, et al. Cartilage. 2012;3:156-164. ACI = autologous chondrocyte implantation; MACI = matrix-induced autologous chondrocyte implantation. Images: Carticel and MACI product Websites. | 59 With Third-Generation ACI, Chondrocytes Are Cultured on a Membrane • In matrix assisted autologous chondrocytes (MACI) ● Autologous chondrocytes adhere to a type I/III collagen membrane ● The MACI implant is cut to the size of the defect and secured with fibrin glue • Advantages include ● Easier application

● ULU- uniform chondrocyte loading MACI ● Arthroscopic possibly? Bentley G, et al. Injury. 2013;44:S3-S10. Images: Sanofi Biosurgery.

| 60 SUMMIT Study Compared MACI With MFX

Demonstrate the Superiority of MACI Implant to MFX Treatment (SUMMIT)

MACI (n=72) MFX (n=72)

Average age: 33.8 years Average lesion size: 4.8 cm2

48 months’ followup: KOOS, MRI, histology, safety

Saris DBF, et al. Am J Sports Med. 2014. KOOS = Knee Injury and Osteoarthritis Outcome Score; MACI = matrix assisted autologous chondrocytes ; MFX = microfracture; MRI = magnetic resonance imaging. | 61 SUMMIT: MACI Repair Resulted in Significantly Better KOOS Scores Than MFX

Baseline 2 Years Baseline 2 Years 100 100 83 80 71 80 61 60 60 49 37 36 40 40

20 20 15 13 Mean KOOS Pain Scores

0 Mean KOOS Function Scores 0 MACI MFX MACI MFX

MACI was associated with significantly better improvement from baseline than MFX in KOOS pain and function scores at 2 years (P <0.001)

Saris DBF, et al. Am J Sports Med. 2014. KOOS = Knee Injury and Osteoarthritis Outcome Score; MACI = matrix assisted autologous chondrocytes ; MFX = microfracture; SUMMIT = Superiority of MACI Implant to MFX Treatment. | 62 Current Phase 3 Clinical Trials in the US

• Confirmatory Study of NeoCart in Knee Cartilage Repair Histogenics-ClinicalTrials.gov Identifier: NCT01066702

A Phase 3, prospective randomized trial comparing the efficacy and safety of an autologous chondrocyte tissue implant (NeoCart) to the surgical intervention microfracture in the treatment of cartilage defects in the knee. • NOVOCART®3D for Treatment of Articular Cartilage of the Knee (N3D) Aesculap Biologics, ClinicalTrials.gov Identifier: NCT01957722

A Phase 3, Prospective, Randomized, Partially Blinded Multi-Center Study to Measure the Safety and Efficacy of NOVOCART 3D Compared to Microfracture in The Treatment of Articular Cartilage of the Femoral Condyle Between 4-6cm2

ClinicalTrials.gov

| 63 Histogenics: Neocart Scaffold assisted ACI

• Estimated Enrollment: 185/245

• Projected Completion: 2017 Primary Completion of Data: JUL/2015

• Primary End Points: KOOS & IKDC Responder Analysis at 12 mos

• Indication: symptomatic femoral AC lesion; no size limitations Engineered Cell-Based Cartilage Repair Produced Better Outcomes Than MFX in a 2-Year Study • NeoCart is a tissue-engineered implant containing chondrocytes in a bovine collagen type-I collagen matrix processed in a bioreactor1 • A phase 2 trial followed patients treated with NeoCart (n=21) or MFX (n=9) for 2 & 5 years1, 2 MFX NeoCart • Mean age (40±9yrs) 40 *P<0.05 * * • BMI (28±4) 30 • Injury acuity (3±5yrs) 20 • Lesion size (NeoCart 287±136mm2 v. MFX 252±135mm2) are similar 10 Mean IKDC Score Change 0 3 6 12 24 Months 1. Crawford DC , et al. J Bone Joint Surg Am. 2012;94:979-989. IKDC = International Knee Documentation Committee; KOOS = Knee Injury and Osteoarthritis Outcome Score; MFX = microfracture. 2. Crawford DC et al. ICRS 2015 presentation. 3. clinicaltrials.gov/show/NCT01400607. Accessed on Dec 16 2013 | 65 Neocart in Comparison to MFX after 5 Years

• 11/60 did not complete the 60-month follow-up • Both groups had similar mean IKDC and KOOS pain scores during study1 • NeoCart was associated with significantly greater ● mean IKDC scores at 12 and 24, but not 60 mos1,2 ● KOOS pain scores at 6, 12 and 24, but not 60 mos1,2 • A phase 3 trial is ongoing3

Compared to MFX, change from baseline was significantly higher in the NeoCart arm at 12, 24, 36, and 48 months for KOOS pain and 12 and 24 months for IKDC (p<0.05) (Table 1)

| 66 Aesculap Biologics: Novocart 3D; Scaffold assisted ACI

• Estimated Enrollment: 25/233; 2 implanted NOV/2014

• Projected Completion: AUG/2021 Primary Completion of Data: JUL/2018

• Primary End Points: co-primary endpoint; KOOS Pain & Function at 24 mos

• Indication: symptomatic femoral AC lesion; 2 - 6cm2 Novocart 3D; Autologous chondrocytes are on a biphasic bovine collagen scaffold seeded with a density of 1.0-4.0 × 106 cells/cm2.

• Cells & scaffold are ready in 21 days • May be performed arthroscopically • The scaffold is placed into the defect upside down; cells are facing the bone • Fixation is achieved with sutures and mini-pins if deemed necessary • Advantages include: easier application, autologous • Clinical data: >8k in Europe, case series: Zak 2 yr fu 23/28 pts & Niethammer graft maturation 3 yr fu 13 pts in AJSM 2014 Prospective Longitudinal Evaluation of 182 pts with Novocart 3D; 5 yr follow-up • Age 35.9 + 9.76 yo • Average defect size of 7.5 cm2 + 3.3 • 50.1% localized degenerative, 21% traumatic, 21% OCD • 79.7% MFC • IKDC scores improved from (44.7 + 18) to (27.6 + 14.9 pts) (p < 0.0001) • 83.3% of patients were responders (> 20% increase in IKDC) • Histology demonstrated hyaline-like cartilage in specimens harvested bw 12 and 42 mos post-op

*Clinical experience with a multi-center prospective trial of Novocart demonstrated significant medium to long term improvements in pts’ pain and function. Outcomes remained stable or improved over time, unlike MFX. Koh J. et al., Prospective Longitudinal Evaluation of 182 Patient With Novocart. ICRS Presentation 2015. Abstract submitted to ORS and provided courtesy of Dr. Daniel Grande Bioprintingde novo Cartilage with ECM-based Bioink Kesti M(1), Grande D(2), Zenobi-Wong M(1) Cartilage Engineering + Regeneration, ETH Zurich, Switzerland, Orthopaedic Research Laboratory, The Feinstein Institute for Medical Research, New York, USA

• Bioprinting is an emerging technology for fabricating complex tissue structures and whole organs. The majority of the bioinks used in bioprinting consists of one or two polymers and are therefore insufficient to mimic the complexity of the native extra cellular matrix (ECM) containing a vast array of morphogens, growth factors and matrix molecules. The potential of ECM bioinks have been shown recently [Pati et al, 2014], however these required the support of a thermoplastic scaffold. Our goal was to develop a bioink that could be incorporated with ECM particles from multiple sources to facilitate 3D bioprinting without the need of structural support materials. The self-supporting scaffolds printed using the ECM bioink demonstrated high precision and overhanging structures. This bioink could be used with ECM particles from many different tissue types to produce complex shapes required for multi-tissue organ printing.

• Figure 1: To demonstrate the versatility of the bioink the hollow cubes (9x6.6x9mm) with 3x4.2mm opening through each face were printed. Scale bar 10mm. • Figure 2: Illustration of 3D printed structure before and after elution of the support material. Scale bar 2.5 mm.

| 70 Patient and Defect Characteristics Are Used to Guide Treatment Selection

B Defects (any size) on the trochlea or femoral condyle1 E 2 T Osteochondritisdissecans T Full-thickness defects on trochlea or femoral condyle2 E R No radiological evidence of osteoarthritis2

Patients aged ≥50 years old2 W Early osteoarthritis2 O 2 R Elevated BMI, smoker S Malalignment2 E Prior MFX3 1. Carticel Website. http://www.carticel.com/ healthcare/about/patient-assessment.aspx. Accessed March 31, 2014. 2. Bentley G, et al. Injury. 2013;44:S3-S10. 3.Minas T, et al. Am J Sports Med. 2009;37:902-908. BMI = body mass index; MFX = microfracture | 71 Conclusions

• Cartilage defects are challenging to repair • Multiple tx options; be comfortable with several ● Factors such as defect size, location, depth and the patient’s physical demands should be considered when developing an individualized treatment plan • New tx options, including novel combinations of existing techniques, continue to be explored • Use an osteochondral technique if bone is abnormal; don’t mess with bone which is normal • Start with WHY- not what and how, but WHY!

| 72 The END

Thank You!

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