Regenerative medicine is an area of intensive investigation, based on the observation that the differentiation and de-differentiation of stem cells along tissue-specific pathways can be induced in the laboratory. One of the most active applications has been in cardiology where a number of trials have been conducted using a variety of marrow cell subpopulations delivered by infusion or intracardiac injection [4
]. Early studies of this type often were neither placebo controlled nor blinded, were conducted at a single site, and did not use standardized cell doses. The CCTRN aims to address these issues by using multiple sites to accrue a larger number of patients to standardized blinded studies. In order to provide the cell products for these trials, two approaches were available. In the first, cells could be processed at a central location and shipped to the clinical sites. However, the requirement to administer fresh cells, and the restricted time window available for the protocols (no longer than 12 hours between marrow harvest and product administration), made this impossible. The alternative was to utilize cell processing facilities associated with the clinical sites, or located nearby, and to develop standardized processing and testing procedures. Once this decision was made, the Sepax device was selected to provide a more uniform automated cell separation procedure, since variability in recipient outcome has been attributed to differences in cell quality [10
] and processing techniques [11
], although this has been disputed [12
]. The Sepax device has been used extensively for cord blood processing [13
] and the density gradient program has been used to prepare cells for cardiac applications, where it was shown to produce separations comparable or superior to those obtained using manual procedures. An extensive study by Atkas et al using Sepax produced separations similar to those from the CCTRN processors [7
] and the product composition. In this earlier study a third wash of the cells in the Sepax was shown to improve platelet depletion and the current Sepax software incorporates this additional wash step.
This comparability to manual techniques was also confirmed by the group at MDACC [13
] who conducted the initial training program for the CCTRN cell processors ( and ). This program demonstrated that comparable results could be obtained by various individuals processing the same starting marrow, both at the initial training, and when repeating the procedures using the same device in their own facilities. In the subsequent clinical applications the Sepax device generally performed well. There were two technical issues, in response to which Biosafe introduced modifications to both the software and the instruction manual and subsequently there have been no further problems.
The CCTRN trial design required that randomization to receive the cell product or placebo was conducted after the product had been prepared; therefore, product information is available for all sixty intended recipients, regardless of whether they ultimately received a placebo. The final products all met release criteria and the use of Gram staining and the Endosafe device for endotoxin testing [8
] reduced the turnaround time for testing to about one hour. FOCUS product preparation was more involved since the product had to be drawn into syringes that were to be handled in a sterile field in the catheterization laboratory.
It was recognized that standardization of additional testing i.e. flow cytometric analysis and colony assays between facilities was more difficult to achieve without extensive validation, and it was decided that these should be handled primarily by the biorepositories in Minnesota (for banking and flow cytometry) and Florida (for colony and functional assays). Basic flow and CFU assays were, however, also performed at the processing centers and these data are included in this report.
The use of centralized testing laboratories required development of careful information dissemination between the processing staff, the trial coordinating center, the CPQC staff and the two biorepositories to alert them to upcoming samples, shipment tracking information, delays etc. This was facilitated through the CPQC who also developed and validated the shipping conditions to address the wide range of temperatures, ranging between Florida in summer and Minnesota in winter. This group also provided centralized quality oversight. Batch records were reviewed prior to randomization of the patient by quality assurance staff at each site and then sent on to CPQC for formal review. They were also responsible for approving any planned deviations (such as filtering the Sepax product if any fibrinous strands or clumps were detected) and for performing on-site audits of the facilities.
This study has demonstrated the feasibility of conducting a multicenter cardiac cell therapy study involving multiple processing sites. As these types of protocols become more widespread this model can be further developed in cases where it is not possible or desirable to cryopreserve the effector cells for shipment. The study has been facilitated by centralized training and quality control, by the use of a more uniform automated cell processing system and by rapid release testing techniques. The CCTRN design has now been extended to include satellite clinical centers up to 128 miles from the main clinical center. This has necessitated validation of transport conditions for shipping the separated cells, but further extends the utility of this system for extending access to cellular therapies.