Cellular Therapies for Cellular Therapies for Repair and Regeneration of Repair and Regeneration of Joint Surfaces—Product Joint Surfaces—Product Characterization and Characterization and Testing Testing Malcolm Moos Jr., M.D., Ph.D. Malcolm Moos Jr., M.D., Ph.D. Medical Officer Medical Officer CBER/OCTGT/DCGT CBER/OCTGT/DCGT [email protected][email protected]
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Malcolm Moos Jr., M.D., Ph.D. Medical Officer CBER/OCTGT/DCGT [email protected]
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Cellular Therapies for Repair Cellular Therapies for Repair and Regeneration of Joint and Regeneration of Joint
Surfaces—Product Surfaces—Product Characterization and Characterization and
““Measure twice, cut once”:Measure twice, cut once”: Up-front investment in each area Up-front investment in each area
may prevent costly waste of time may prevent costly waste of time and resources on sub-optimal and resources on sub-optimal process or variable/ineffective process or variable/ineffective productproduct
Better characterization makes Better characterization makes many things easiermany things easier
In vitroIn vitro culture is not required culture is not required
HACDC in HACDC in regenerating muscleregenerating muscle
Product TestingProduct Testing
Is guided by detailed Is guided by detailed understanding of the product understanding of the product and the manufacturing and the manufacturing processprocess
Should ensure consistency of Should ensure consistency of process and product process and product
Should predict Should predict in vivoin vivo activityactivity
Product TestingProduct Testing
——An example of An example of new alternativesnew alternatives
Terminal markers may not predict biological functionTerminal markers may not predict biological functionCharacterization of functional chondroprogenitors Characterization of functional chondroprogenitors Qualification by meaningful biological responsesQualification by meaningful biological responses
0
20
40
60
80
100
0 20 40 60 80 100
Islet cell transplantation in Islet cell transplantation in diabetic micediabetic mice
Live/dead Staining (%)Live/dead Staining (%)
0 20 40 80 10060
OC
R/D
NA
(%
)O
CR
/DN
A (
%)
20
40
80
100
60
0
Theoreticalcorrelation
OOxygen xygen CConsumption onsumption RRate and dye ate and dye exclusion are exclusion are uncorrelateduncorrelated
(K. Papas, University of Minnesota Department of Surgery, in preparation)
QResponse to Rat Islet Transplants in Diabetic Balb/C Mice (Anti-CD4)
2 / 0 / 0 1 / 0 / 0
2 / 0 / 0
2 / 0 / 0
6/ 0 / 0
4/ 0 / 0
3 / 0 / 0
3 / 0 / 0
1 / 0 / 10 / 1 / 2
1 / 0 / 1
2 / 0 / 0
1 / 0 / 0
0 / 1 / 0
1 / 0 / 0
3 / 0 / 0
2 / 0 / 0
3 / 0 / 0
all curemixedall fail
3 / 0 /0
2 / 0 / 0
1 / 0 / 2
1 / 0 / 0
0
4
5
6
0 50 100 150 200 250 300 350 400 450
100800
700
600
500
400
300
200
100
0
0 50402010 6030 8070 90
0 / 1 / 1
0 / 0 / 4
0 / 0 / 6
0 / 0 / 3
0 / 0 / 2
1
2
3
0 / 0 / 30 / 0 / 2
Cure rates and days to cure in mice Cure rates and days to cure in mice receiving 1,000 human IE according to receiving 1,000 human IE according to
ATP/DNAATP/DNA
00
2020
4040
6060
8080
100100
00 1010 2020 3030 4040
Days after transplantDays after transplant
Cu
re r
ate
(%)
Cu
re r
ate
(%)
ATP < 110ATP < 110 pmol/µg DNApmol/µg DNA
ATP ATP >> 110 110 pmol/µg DNApmol/µg DNA
P=0.008P=0.008
(B. Hering, University (B. Hering, University of Minnesota of Minnesota Department of Department of Surgery, with Surgery, with permission) permission)
The nude mouse modelThe nude mouse model
Dell’Accio et al, A&R, 2001Dell’Accio et al, A&R, 2001
The ShmooThe Shmoo
——Al Capp, ca. 1948Al Capp, ca. 1948
Instructions go both Instructions go both waysways
DonorDonor RecipientRecipient
Donor-Host Donor-Host Interactions?Interactions?
Cell productCell product
Articular surfaceArticular surface
Subchondral boneSubchondral boneRemnants of calcified cartilageRemnants of calcified cartilage
CoverCover
Synovial Joint FluidSynovial Joint Fluid
Competence Competence FactorsFactors
gene expression
Biological Response
P
β-actin
ALK-1
BMP-2
Col2a1
FGFR-3
Population doublings FI 2 4 10 12
Safranin O
Molecular profile of the stable chondrocyte
His
tolo
gy s
core
His
tolo
gy s
core
Gene scoreGene score
-5-5 -3-3 -1-1 11 33 55
00
11
22
33
Relation of Gene score to Relation of Gene score to Stable Cartilage FormationStable Cartilage Formation
Stable cartilageStable cartilage
Space—the final Space—the final frontier?frontier?
Molecular Molecular Boundaries in Tissue Boundaries in Tissue
FormationFormation
400x Thomas and Moos, 2005 (submitted)
Thought QuestionsThought Questions
Acceptance, selection, or instruction of Acceptance, selection, or instruction of starting materialstarting material
Consider competence factors sufficient Consider competence factors sufficient to define chondroprogenitors (rather to define chondroprogenitors (rather than characteristics of terminal than characteristics of terminal differentiation)differentiation)
Distribution of characteristics in 3D may Distribution of characteristics in 3D may be criticalbe critical
Qualification of potential tests against Qualification of potential tests against appropriate biological responses appropriate biological responses in in vivovivo
•Criteria for obtaining starting tissueCriteria for obtaining starting tissue
•Characteristics of functional chondroprogenitor cellsCharacteristics of functional chondroprogenitor cells
•Analytical methods to determine these characteristicsAnalytical methods to determine these characteristics
•Qualification of these tests in preclinical modelsQualification of these tests in preclinical models
•Potency assaysPotency assays
•Special issues for cells contained in natural or artificial matricesSpecial issues for cells contained in natural or artificial matrices