Susan E. Lana, DVM, MS, DACVIM (Oncology)
With the advancement of molecular biology techniques has come a greater need for tumor biorepositories to provide appropriate material for continued discovery. Many current biorepositories in both human and veterinary medicine are a byproduct resource of project or investigator driven studies.1 Samples collected via this mechanism are often specific to a certain disease type and may become available for other investigation only after the primary project is complete.2 They are in many cases linked to a very well annotated data set that makes them an invaluable resource for future studies that may be completely unrelated to the initial project they were collected for. It also raises special considerations about using "left over" tissues for other purposes.2 Another type of tissue bank involves systematic collection of all tissues available from a population, that are then stored for future use. This situation is less common and often lacks an up front scientific question and the financial support that comes from specific projects or grants. As technology advances, using smaller samples for greater discoveries is becoming a reality and the importance of this resource is being recognized at many levels.
Systematic tissue collection and archiving however is not a casual undertaking. Just having access to tissue or a patient population of interest does not constitute a viable program. Other required pieces include a team of personnel trained and dedicated to the process, equipment and supplies, space for storage, computer and data base support, and of course financial resources for all of the above. Each of these components is necessary to make use of the valuable resource that walks through the door each day in the clinic. The information presented below is provided as points to consider for individuals thinking about developing a biorepository.
Client / Patient Interface
During systematic tissue collection every patient who undergoes a biopsy or surgical procedure may be a potential sample donor. It is important to remember however that not all clinical situations are appropriate for tissue collection. Very small tumors or circumstances when diagnostic information would be sacrificed (like tumor margins) are not candidates for collection. Patients with poor performance status may not be good candidates for sample collection. The patients' best interests, care, and well being are always a priority over tissue collection. In our institution, communication occurs early in the process between the surgeon/clinician and the dedicated archiving technician about the possibility of sample collection for each individual case. If any question exists, the patient is not considered a donor.
Client consent is another important part of a biorepository program. Even though the parts that are being banked are often left over pieces, informed consent is still required, although the specifics of this process and regulatory requirements may vary amongst veterinary institutions. In our institution all new client admissions to the oncology service are asked to review and sign the tissue archiving consent form. In many circumstances this is not needed as their pet does not undergo surgery/biopsy or the patient is determined later to not be a good candidate for sampling. Approval of the program should also be obtained from the institutional review board or animal care and use committee if required by the institution. Again, regulations will vary from location to location and will likely differ amongst institutions.
Owner/patient confidentiality is also an important consideration when participating in biorepository efforts. Samples may be anonymized or coded. Anonymized samples generally are those which have no link to donor information and it can not be obtained by the end user of the tissue. Coded samples are indirectly identifiable in that the identity of the donor is not known to the researcher, but can be determined by the provider of the tissues.1 In an ideal setting, the concept of an "honest broker" exists to protect the donor and researcher.3 In this situation through the use of appropriately coded entries and restricted database access, select individuals act as a barrier between confidential patient information obtained during clinical management (name, address, phone numbers, etc) and research information obtained after the samples are used. The ability to cross reference sample identification with sensitive owner or patient information is not removed, just blocked. When new or additional clinical correlate information such as patient outcome becomes available, it can be added at a later date.
Sample Acquisition and Processing
Determining what samples to collect and how to process them is an important step in establishing a tumor bank. For project driven banks these criteria may be predetermined by the study. For systematic collection, methodology for sample processing that has the broadest appeal and is practical usually is the approach taken. Different processing conditions might be better for different intended uses of the samples collected. For example, tissues collected for RNA use are time and degradation sensitive and should be processed quickly. Other samples such as blood may be affected by the anticoagulant that is used.4,5 Standard operating procedures (SOPs) should be written, followed, and revised at regular intervals or as the needs of the research community change. Careful documentation about the sample itself and any deviations from the SOP should be included in the database. In addition, information about sample processing time and the amount of sample present (weight or volume) in each vial or unit is useful to include in the database. This information can help end users select the most appropriate sample set for their research purpose.
Large amounts of data are generated as a biorepository grows. If possible, anticipating future needs such as the desire to interface with other repositories or providing a user web interface may save time, effort, and allows for greater flexibility as a program expands and the needs of the end user change. In addition to sample specific information, storage and tracking information is also needed. Monitoring when samples leave the bank, if they return and in what condition, and who handles those samples is important.
Linking to clinical information is an important feature that makes the samples in the bank even more valuable. Common clinical data elements should be determined so that the most useful information will be obtained. This may include patient demographics and history; tumor stage, grade, or other characteristics important to that disease type; and clinical outcome data.3 Publications providing detailed information and examples of tumor banking experiences exist in the human field and can be used as a starting point when considering how to best proceed from a data management point of view.3,6,7
Evaluating what has been collected is also an important part of the tissue archiving process. From a histopathologic standpoint, not all sections or pieces of a large tumor are likely to be exactly the same because tumors are in general heterogeneous. Reviewing the H&E section to determine things such as percent tumor present in the sample, percent necrosis, percent normal tissue within the specimen and confirming the diagnosis are important. Several large human biorepository groups report the use of a 65% tumor content cut off as an acceptable standard for sample quality.8,9 If a sample is below this cut off it is not released for external use. Sandusky reported that of 1550 samples tested 75% passed histologic quality assurance.8 Tumor types most likely to fail QC evaluation were those taken from very small tumors such as breast and prostate samples. Because many samples collected by biorepository are used for molecular analysis, assuring RNA and DNA quality is also important. In most cases spot checking to ensure the collection procedures result in adequate nucleic acid quality is done, representing different tissue types and storage times. Several techniques have been reported and compared.9 In many cases this type of QC is also done when requested by the end user although an additional cost is charged for these services.
Another point to consider when participating in a biorepository is who has access to the tissues and how is this use determined. In many cases this will depend on the intent and scope of the program. Some resources are for internal use only but this may change over time as the bank grows and is populated by more samples. If the bank is intended for outside use generally an application process with committee review is used to assure that the intent of the tissues' use is reasonable (ie appropriate research), and that the user has the funds and expertise to make use of the samples provided. Materials transfer agreements are recommended for any tissue transfer to protect the interests of both the provider and researcher.
Cost of establishing and then maintaining a tumor bank is also a point to be considered. By far the most expensive part of the process is dedicated personnel to provide sample acquisition and data management support. This is also the piece of the program that ensures its success. Project specific support is often available for this type of archiving effort but those resources are often finite and go away when the project is completed. A minimal charge to recoup operational or processing costs is often requested of the biorepository user. The fee structure may varying depending on the end recipient (for profit or non for profit) and the type of sample requested. In our institution the cost of processing a sample through the wax block stage is approximately $25-30 dollars. This includes supplies and pathology charges and does not include personnel.
Animal Cancer Center Experience
The tissue archiving program at the ACC was formalized in September 2003 and has been actively collecting patient samples since. It began as a project driven tissue repository several years earlier and was continued when that project ended. Client consent is obtained from every new patient seen on the oncology service and IACUC approval has been obtained for sample collection and is renewed yearly. For each patient that undergoes a surgical procedure or biopsy, the following samples are obtained when available: serum, plasma, peripheral blood mononuclear cells (PBMCs), urine, tumor, and normal stromal samples and appropriately processed (frozen, OCT, RNALater®). A tissue block and H&E slide is also maintained for use by the tissue archive lab independent of the general pathology service. In order to accurately catalogue all samples, a database has been developed that is specific to the archiving effort. Patient information is input, samples are coded and tracked for use. Currently, over 800 patient sample sets exist in the archive. In addition to general tumor archiving, trial or investigator specific processing is also done through the tissue archiving core. A full time research associate is dedicated to the tissue archiving core and activities include database entry, patient identification, interaction with surgery technical staff, and tissue and fluid processing. A half-time administrative professional also devotes the majority of her time to the current tumor archiving effort. Duties include database construction and management, sample storage, retrieval, and shipping, as well as responding to inquires about tissue availability from investigators outside of and within the ACC.
Storage for tissues and other specimens is provided by the Biophile® freezer system. This is a fully automated, self contained, -80 degree robotic storage system. Individual bar-coded racks containing sample vials enter the system through an automated airlock that prevents frost build up and negates the need to open the door of the unit. Racks are stored and retrieved according to bar code within 20 seconds of request. Security measures and pass codes restrict sample access. Additional storage is provided at -140 degrees for certain samples.
1. Oosterhuis JW, et al. Nature Reviews Cancer 2002;3:73.
2. Goodman GE, et al. Cancer Epidemiol Biomarkers Prev 2006;15(4):599.
3. Patel AA, et al. BMC Cancer 2006;6:120.
4. Vaught JB. Cancer Epidemiol Biomarkers Prev 2006;15(9):1582.
5. Holland NT, et al. Toxicology and Applied Pharmacology 2005;206:261.
6. Ayers LW, et al. Infectious Agents and Cancer 2007;2:7.
7. Qualman SJ, et al. British Journal of Cancer 2004;90:1115.
8. Sandusky GE, et al. Cell Tissue Banking 2007;8:287.
9. Jewell SD, et al. Am J Clin Pathol 2002;118:733.