Copyright ©The Author(s) 2015.
World J Med Genet. Feb 27, 2015; 5(1): 1-13
Published online Feb 27, 2015. doi: 10.5496/wjmg.v5.i1.1
Table 1 Current cell mediated gene therapy protocols in clinical and R and D stages
Target diseaseStageGene modificationTarget cellRef.
Hemophilia APreclinicalOct4, Sox2, Klf4FibroblastXu et al[40]
ALSPreclinicalOct4, Sox2, Klf4, c-MycFibroblastDimos et al[55]
AlzheimerPreclinicalOct4, Sox2, Klf4, c-MycFibroblastIsrael et al[46]
ALSPreclinicalOct4, Sox2, Klf4FibroblastChen et al[54]
Cardiac failurePreclinicalGata4, Mef2c, Tbx4FibroblastIeda et al[22]
DiabetesPreclinicalOct4, Sox2, Klf4FibroblastZhang et al[24]
SchizophreniaPreclinicalOct4, Sox2, Klf4, c-MycFibroblast with 15q11.2 delYoon et al[12]
Barth syndromePreclinicalOct4, Sox2, Klf4, c-MycFibroblastWang et al[52]
Parkinson’s diseasePreclinicalOct4, Sox2, Klf4, c-MycFibroblastChung et al[50]
Macular degenerationPhaseIOct4, Sox2, Klf4, c-MycFibroblastKamao et al[14]
Degenerative arthritisPhase II/IIITGF-β1ChondrocyteHa et al[15]
X-linked adrenoleukodystrophyIn the clinicABCD1CD34+ Bone marrow cellCartier et al[36]
Wiskott-aldrich syndromeIn the clinicWASPCD34+ Bone marrow cellAiuti et al[35]
Metachromatic leukodystrophyIn the clinicARSACD34+ Bone marrow cellBiffi et al[37]
Table 2 Different types of production methods based on cells needed per batch
Cells/lot (billions)Cell cultureHarvestFillingFreezing
1T175 flask or hyperflask, 10 layer cell factoryBucket centrifugationHand/manual fillBench top control rate freezer
1010 layer cell factory, hyperstack-12, Xpansion systemsBucket centrifugationSemi automated fill machine or hand/manual fillBench top control rate freezer
50Hyperstack or Xpansion, cell cubeTangential flow filtration, continuous centrifugationAutomated fill machineLarge scale control rate freezer
100Factory automation of Xpansion or Hyperstack technologiesTangential flow filtration of continuous centrifugationAutomated fill machineLarge scale control rate freezer
500Bioreactors using microcarriersContinuous centrifugationAutomated fill machineScale out large scale control rate freezer
Table 3 Suggested release tests advised by Food and Drug Administration for cell product
CategoryTestKey measurement
SterilitySterilityMicrobiological testing
MycoplasmaAfter pooling of cultures but before washing
IdentityIdentityCell identity and assess heterogeneity
PurityImpurityResiduals contaminants
EndotoxinContaminants during the process
PotencySpecific to productRelevant function of the cells
Other testingGeneral safetyCellular therapy products are exempt
ViabilityGenerally > 70%
Cell numberMinimum and maximum cell number
Title 21 of the code of federal regulations applied to cell therapy products
The tissue rules: Part 1271
The biologics requirements: Part 600 and 610
The investigational new drug requirements: Part 312
The drug manufacturing requirements: Parts 211 and 212
Table 4 TG-C release test
Release tests and specifications for hChonJ cell
Cell growth and morphologyVisual (test code 30117)Fibroblast-like: spindle shaped/bipolar or multipolar cells
ViabilityTrypan blue dye exclusion> 70%
(test code 30458)
Mycoplasma1993 Points to consider (test code 30055)Negative
RAP testIn vivo and ELISA (test code 30163)Negative
Sterility (direct inoculation method)21 CFR 610.12 (test code 30744)Negative
EndotoxinLAL (test code 37653)< 5 EU/mL
Release tests and specifications for hChonJb#7 Cell
IdentityEpithelial-like: fried egg or polygonal-shaped appearance
Cell growth and morphologyVisual (test code 30117)
Type II collagen assayRT-PCR (test code 30412)Type II collagen present
TGF-β1 presencePCR (test code 30959)TGF-β1 present
Potency (TGF-β1 assay)ELISA (test code 30444)1-50 ng/105 cells/24 h
ViabilityTrypan blue dye exclusion (test code 30458)> 70%
RCRCo-culture of end of production cells (test codes 30628)Negative
Supernatant amplification (test code 30633)Negative
Mycoplasma1993 points to consider (test code 30055)Negative
Rat antibody productionIn vivo and ELISA (test code 30163)Negative
EndotoxinLAL (test code 37653)< 5 EU/mL
Sterility21 CFR 610.12 (test code 30744)Negative