Briefing Document


Cellular, Tissue and Gene Therapies Advisory Committee


Meeting #43


Draft Guidance for Industry for minimally manipulated, unrelated allogeneic cord blood, and the regulatory approach to minimally manipulated unrelated allogeneic peripheral blood stem/progenitor cells


March 30, 2007



FDA has regulatory authority over two types of hematopoietic stem/progenitor cells (HPC) that are used for replenishing the bone marrow (i.e., hematopoietic reconstitution) in patients exposed to myeloablative chemotherapy or total body irradiation for treatment of a variety of diseases—those sourced from placental/umbilical cord blood (HPC-C) and those obtained by apheresis (HPC-A).  When the HPC are minimally manipulated, and collected from unrelated allogeneic donors, these cells are regulated as human cells, tissue, and cellular and tissue-based products (HCT/P) subject to licensure.  Other unrelated allogeneic HPC, such as those that are more than minimally manipulated, and certain autologous and related allogeneic HPC, such as those that are more than minimally manipulated, are also regulated as HCT/P subject to licensure; however, these products are not under consideration for this meeting.

Transplant physicians, laboratory scientists, and medical technologists have amassed extensive clinical and non-clinical laboratory experience with HPC-C since the first reported transplant in a child with Fanconi anemia in 1988.  By 1993, large public repositories of HPC-C were established, in New York, Milan, and Dusseldorf.  The use of HPC-C for transplantation is increasing, and now exceeds the use of bone marrow for pediatric transplants (IOM report).  Currently it is estimated that there are over 75,000 cryopreserved, HLA-typed unrelated donor HPC-C in US inventories alone, and over 240,000 HPC-C stored in cord banks worldwide (  Estimates from registry data suggest that over 7000 cord blood transplants have been performed worldwide.


Prior to the advent of large scale banking of HPC-C in public inventories, HLA-matched bone marrow and peripheral blood stem/progenitor cells obtained by apheresis of healthy mobilized donors (HPC-A) were the major sources of HPC for transplantation. Each source has relative limitations and advantages.  For example, HPC-A consist of large numbers of mononuclear cells and therefore are associated with rapid neutrophil and platelet engraftment.  Rapid access to a large inventory of immediately available, HLA-matched unrelated allogeneic HPC is one of the most significant advantages of banked cord blood.


The regulatory approach to both HPC-A and HPC-C has been proposed in several workshops and FDA publications (described below).  Although HPC-A have been used for transplantation for a longer period of time and in greater numbers than HPC-C, the availability of extensive HPC-C manufacturing and clinical outcome data in a public repository allowed for development of recently published draft guidance for industry describing an approach to licensure for these products.


In 1997, we proposed a new regulatory framework for human cellular and tissue-based products, including hematopoietic stem/progenitor cells.  The proposed framework provided a tiered approach to the regulation of human cellular and tissue-based products, now referred to as human cells, tissues, and cellular and tissue-based products (HCT/Ps).  We implemented this approach by promulgating three final rules, which comprise 21 CFR Part 1271.

On January 19, 2001, we published the first final rule entitled “Human Cells, Tissues, and Cellular and Tissue-Based Products; Establishment Registration and Listing” (66 Federal Register 5447) (the “registration and listing final rule”).  The registration and listing final rule requires establishments that manufacture HCT/Ps to register and list their products with the agency.

On May 25, 2004, we published the second final rule entitled “Eligibility Determination for Donors of Human Cells, Tissues, and Cellular and Tissue-Based Products” (69 Federal Register 29786) (the “donor eligibility final rule”).  This final rule requires, with certain exceptions, that a donor eligibility determination be made based on the results of HCT/P donor screening and testing for relevant communicable disease agents and diseases.

On November 24, 2004, we published the third final rule entitled “Current Good Tissue Practice for Manufacturers of Human Cells, Tissues, and Cellular and Tissue-Based Products; Inspection and Enforcement” (69 Federal Register 68612).  The Current Good Tissue Practice (CGTP) requirements govern the methods used in, and the facilities and controls used for, the manufacture of HCT/Ps, including but not limited to all steps in recovery (collection), donor screening, donor testing, processing, storage, labeling, packaging, and distribution.

On May 25, 2005, the three final rules became effective. As described in the registration and listing final rule, we are regulating as biological drugs or devices those HCT/Ps that:

                    o            are more than minimally manipulated (processing alters the biological characteristics of the cells);

                    o            are for a use other than homologous use as reflected by the labeling, advertising, or other indications of the manufacturer’s objective intent;

                    o            the manufacture involves the combination of the cell or tissues with another article, to include water, crystalloids, or a sterilizing, preserving, or storage agent provided the addition of water, crystalloids, or the sterilizing, preserving, or storage agent raises new clinical safety concerns with respect to the HCT/P; or

                    o            has a systemic effect or is dependent upon the metabolic activity of living cells for its primary function, and is not for:

                                             §            autologous use;

                                             §            allogeneic use in a first- or second- degree blood relative; or

                                             §            reproductive use.

We consider unrelated allogeneic hematopoietic stem/progenitor cells to have a systemic effect and are therefore regulating them as biological products and drugs under the Public Health Service (PHS) Act and the Federal Food, Drug, and Cosmetic Act (FDCA).

On January 20,1998 (63 Federal Register 2985), we issued a notice in the Federal Register entitled “Request for Proposed Standards for Unrelated Allogeneic Peripheral and Placental/Umbilical Cord Blood Hematopoietic Stem/Progenitor Cell Products; Request for Comments.”  In the notice, we explained that it may be possible to develop product standards and establishment and processing controls of minimally manipulated unrelated allogeneic hematopoietic stem/progenitor cell products intended for hematopoietic reconstitution, based on existing clinical trial data, or data developed shortly thereafter, demonstrating the safety and effectiveness of the cells.  We further explained that if adequate information were submitted to the docket, we intended to issue guidance in accordance with good guidance practice regulations, 21 CFR 10.115, for establishment controls, processing controls, and product standards.  If such standards could be developed, we suggested that, in lieu of reviewing individual applications containing clinical data, we would propose granting licensure for products shown to meet the issued standards.

To provide a scientific basis for the proposed standards, we requested the submission of comments proposing establishment controls, process controls, and product standards designed to ensure the safety and effectiveness of minimally manipulated unrelated allogeneic hematopoietic stem/progenitor cell products derived from peripheral and cord blood for hematopoietic reconstitution.  Submitted comments were to include supporting clinical and nonclinical laboratory data and other relevant information. To allow sufficient time for the development of data and standards for these products, the notice also announced our intention to phase in implementation of an IND and license application requirements for these products.  We provided a period of two years, until January 20, 2000, for interested persons to submit proposed product standards and establishment and processing controls with supporting clinical and nonclinical data.  At the request of industry, we reopened the comment period for 90 days until July 17, 2000 (65 Federal Register 20825, April 18, 2000). The data submitted to this public repository were exclusively related to cord blood manufacture and transplantation. No data were submitted on peripheral blood HPC manufacture and transplantation.

The Biological Response Modifiers Advisory Committee (BRMAC) met on February 27, 2003, to discuss issues related to the use of unrelated allogeneic hematopoietic stem/progenitor cells derived from placental/umbilical cord blood for hematopoietic reconstitution.  At the meeting, FDA provided an analysis of clinical outcome data submitted to the FDA regarding the safety and effectiveness of cord blood for hematopoietic reconstitution.  Guest experts provided the most recent data on clinical studies of unrelated donor umbilical cord blood transplantation in children and adults.

The committee’s deliberations focused on:

·         factors the agency should consider in determining the safety and effectiveness of placental/umbilical cord blood transplantation for hematopoietic reconstitution;

·         the role of CD34+ cell count in selection of cord blood units for transplantation; and

·         other measures of quality that should be considered.


A transcript and summary minutes of the meeting can be obtained from the CBER Advisory Committee Meeting Transcripts web page (

A task group composed of CBER scientists, medical officers, and regulatory staff reviewed and assessed the submitted information, data presented at the BRMAC and other public meetings, and the large body of published literature on this subject.  The task group prepared and published a draft Guidance for Industry entitled “Minimally Manipulated, Unrelated, Allogeneic Placental/Umbilical Cord Blood Intended for Hematopoietic Reconstitution in Patients with Hematological Malignancies” (1/17/07), recommending ways for manufacturers to apply for licensure of these products, a copy of which has been provided to Advisory Committee Members.

The draft guidance is divided into sections, some of which contain the applicable regulatory requirements and others which describe the license application procedure.  The draft guidance provides detailed information that should be included in the chemistry, manufacturing and controls (CMC) section and the establishment description (ED) section of their biologic license applications.  The CMC section provides guidance on HPC-C description and characterization, methods of manufacturing, container closure system, methods validation/verification procedures, and labeling.  The ED section provides guidance on facility layout, utility systems (water, heating/ventilation/air conditioning), facility controls, computer systems, and prevention of cross-contamination.

The information that follows provides an overview of the specific issues in the Draft Guidance to be addressed in the questions for the Committee.

Chemistry, Manufacturing and Controls Recommendations in the Draft Guidance


The Product standards proposed in Table A are based on product data submitted to the docket, review of the scientific literature, and recommendations received from the Biological Response Modifiers Advisory Committee at the 2003 meeting.  We have proposed the use of CD34+cell dose and the total nucleated cell (TNC) content to demonstrate purity and potency of the HPC-C products.  Applicants for a BLA for HPC-C would be expected to achieve similar product characteristics using the recommended or other appropriate tests in order to rely on the clinical data from the docket instead of submitting their own clinical data.



Table A. Required and recommended Tests and Test Results 1

Product Characteristics 2



(Type and Testing)

Results of Product Testing



Infectious diseases- Testing Required

(21 CFR1271.45 through 1271.90)


Maternal peripheral blood collected within 7 days of cord blood collection-Type and Timing Required (21 CFR 1271.80(a) and (b))


All tests negative except non-treponemal test for syphilis when confirmatory test is negative. (Cytomegalovirus (CMV) results are recorded).


    CMV - Report
Sterility - Bacterial & fungal cultures –Testing Required (21 CFR 211.165(b) and 21 CFR 610.12) Cord blood* and HPC-C (pre-cryopreservation) ** Negative
Hemoglobin Cord blood* No homozygous hemoglobinopathy
Purity and Potency Total nucleated cells (TNC) HPC-C (pre-cryopreservation) = 5.0 x 10 8 TNC *** / unit HPC-C
Viable nucleated cells HPC-C (pre-cryopreservation) =85% viable nucleated cells
Viable CD34+ cells (flow cytometry) HPC-C (pre-cryopreservation) = 1.25 x 10 6 viable CD34+ cells**** / unit HPC-C
Identity Human leukocyte antigen (HLA) Typing Cord blood Report
Confirmatory HLA typing Attached segment of HPC-C Confirms initial typing
Blood Group and Rh Type Cord blood Report
1 Testing, Sample (Type and Timing) and Results are recommended unless specifically noted as


Table A.   Required and recommended Tests and Test Results1

1 Testing, Sample (Type and Timing) and Results are recommended unless specifically noted as required Collected cord blood = cord blood product before undergoing volume reduction

2 The PHS Act requires a demonstration that the product is safe, pure, and potent

* Cord blood= cord blood before undergoing volume reduction

** Sample may be obtained before or after addition of the cryoprotectant

***Based on 20 kg recipient @ ≥2.5 x 107 NC/kg & ≥70% post-thaw recovery = 1.7 x 107 nucleated cells/kg

****Based on CD34+ cells ≥0.25% of TNC prior to freezing



Cord blood processing is currently performed using volume reduction methods.  The most commonly cited volume reduction method was developed by Rubinstein et al (PNAS 1995).  This processing involves the addition of a plasma volume expanding medium, hydroxyethylstarch (Hespan or Pentastarch) to the cord blood and centrifuging to cause the red cells to rouleaux and sediment; this facilitates the separation of red blood cells from the white cells containing the progenitor cells.  The leukocyte-rich supernatant is separated into a plasma transfer bag that is further centrifuged. Surplus supernatant plasma is then expressed into a second plasma bag and the sedimented leukocyte fraction is resuspended in supernatant plasma to a volume of 20 ml.  The leukocyte-enriched HPC-C is then mixed with DMSO (in dextran); DMSO protects the cells from damage as a result of the freezing process.  The HPC-C cell suspension is cryopreserved and then stored in liquid nitrogen, until ready to use for transplantation.  The final volume of the each cryopreserved HPC-C product after volume reduction is between 25-40 ml.  The purpose of volume reduction is to allow for the storage of a large number of HPC-C in a finite amount of space, ultimately resulting in an increased number of available HPC-C.  Some establishments use slight modifications of this volume reduction procedure, however, they essentially achieve the same goal.


Prior to the development of the volume reduction methods, cord blood was processed by alternate procedures.  Any change in methodology has the potential to affect the quality and stability of HPC-C.  For HPC-C processed or manufactured using different procedures than those currently recommended in the draft guidance document, establishments would need to provide separate validation summaries and demonstrate comparability of the previously manufactured products to the currently manufactured products, in order for these HPC-C to be included under the license.  We propose the use of product characteristics such as TNC count, viable CD34+ cells content and colony forming units (CFU), for the comparability data to support licensure of these previously manufactured HPC-C in storage.  Establishments may opt to use alternate product attributes from stability or other studies to demonstrate comparability.  Clinical outcome data on these HPC-C or information from the medical literature may also be cited to support product comparability.


Our recommendation to use TNC count, viable CD34+ cells content and colony forming units (CFU) in comparability studies is supported by the scientific and medical literature.  The nucleated cell dose (Gluckman et al, 1997; Rubinstein et al., 1998) and CD34+ cell dose (Wagner et al., 2002) have been shown to correlate to consistent engraftment. Increases in the nucleated cell dose are shown to be associated with a shortened time to engraftment, whereas the CD34+ cell dose has been associated with predicting the speed of recovery. Hence these two attributes have been used as indices for faster engraftment and disease-free survival in cord blood cell transplantation. 


Most establishments generally enumerate the viable CD34+ cells in the product as a surrogate for committed and uncommitted progenitor cells in the context of validating the collection and processing methods. Assessment of colony forming units (CFU) have also been used as a measure of in-vitro function of the progenitor cells, however, this assay has problems with reproducibility.  Because of this problem some establishments discard HPC-C which do not grow colonies.  The assessment of CFU, although not a good quantitative measure, is an in-vitro function assay that may be useful for validating the processing, storage and shipping procedures, and in comparability studies.  As depicted in Figure 1, a correlation between the numbers of viable CD34+ cells and CFU in HPC-C has been reported (Cairo et al 2005).





Text Box: Log All CFU (x 104)

Figure 1 Correlation of CFU and CD34+ cells in HPC-C.  From; “Cord Blood Log CD34+ and CD34+ Subsets (CD38-, CD61+, CD90+) are Significantly  Correlated to Log Total  CFU, CFU-GEMM, CFU-GM and BFU-E: A Report From The COBLT/NHLBI Program”.  Mitchell S. Cairo, Elizabeth Wagner, Geoff Cohen, John Fraser, LeeAnn Jensen, Shelly Carter, Carmella van de Ven, Nancy Kernan, Joanne Kurtzberg.  Blood, Volume 104, issue 11, November 16, 2004, Abstract #406.



Comparability studies will likely include assays performed on cryopreserved samples from previously manufactured and stored HPC-C.  For HPC-C stored in plastic containers, samples may be obtained from an integral segment of contiguous tubing that is heat-sealed, cryopreserved, and stored under the same conditions as the HPC-C unit.  HPC-C samples may also be stored in separate aliquots, such as in a small volume vial, in separate freezers. These separate aliquots may not have been exposed to the same freezing and storage conditions as the HPC-C unit and thus may not be representative of the unit.  Procedures and conditions for freezing and storage may have an effect on the quality of the HPC-C  Some reports have shown a correlation of test results (such as TNC count, viable CD34+ cells content, CFU) between samples from the segment and those taken directly from the HPC-C container (Rodriguez et al. 2004; Goodwin et al. 2003 ).  Other studies have evaluated cryovial samples and determined that it is representative of the hematopoietic content of the HPC-C unit (Solves et. al. 2004).  Limited information is currently available in the literature on the utility of segments and cryovial samples to assess the quality of the HPC-C unit and to determine whether a correlation exists between tests of small product aliquots stored separately from the HPC-C, and samples obtained directly from the HPC-C.


We request that the Committee discuss the scientific aspects of demonstrating comparability between previously manufactured HPC-C and HPC-C manufactured currently.  (See question 1)


Clinical Indication


The Draft Guidance describes a path to licensure for HPC-C intended for hematopoietic reconstitution in patients with hematological malignancies. Regulatory requirements for licensure of biological products include demonstration of potency. Potency has long been interpreted to include efficacy; in other words, the “specific ability or capacity of the product to effect a given result” (21 CFR 600.3(s)). Therefore the draft guidance is limited to this indication based on data submitted to the public docket, which supports safety and efficacy of HPC-C for hematopoietic reconstitution in patients with hematologic malignancies.


HPC-C transplant outcome data reported to the docket by HPC-C establishments summarized the primary outcomes of engraftment and survival in recipients with a multitude of different underlying life-threatening diseases.  For example, a large proportion of the clinical data were provided by a single center that categorized the disease conditions into 3 basic categories: hematological malignancies, genetic diseases, and acquired marrow failure syndromes, with reported outcomes including rate and speed of engraftment, graft versus host disease, and transplantation-related events.


Table 2: Diseases treated by Cord Blood Transplantation

Rubinstein et al., 1998


From the table it is apparent that about 2/3 of HPC-C were used to treat hematological malignancies, a quarter were used to treat genetic diseases and 8-10% were used to treat acquired marrow failure syndromes such as aplastic anemia and MDS.  Overall, about 15% of all the cord blood transplants in this series were performed for congenital and acquired marrow failure syndromes.


The number of specific genetic and marrow failure diseases treated by cord blood transplantation is large; however, information regarding safety and efficacy of cord blood transplantation is available for relatively few specific diseases.  In contrast, there is considerable clinical data supporting the use of cord blood for hematopoietic reconstitution in patients with hematological malignancies.  We request that the Committee discuss whether there are data available to support additional indications. This will be discussed in question 2 (below).




1. Please discuss the types of data that could be submitted to demonstrate comparability between the previously manufactured HPC-C and HPC-C manufactured currently.  The Draft Guidance states these data could include TNC count, viable CD34+ cell content, and colony forming unit content, other product attributes obtained from stability or other studies, data cited from the medical literature, and clinical outcome data.  In the discussion please address the following:

·         relationship of data obtained from viable CD34+ cell content and colony forming unit content

·         alternate test methods

·         the types of samples that are available from the previously manufactured HPC-C and HPC-C manufactured currently, and how they might impact comparability studies


2. Please comment on the clinical indication described in the Draft Guidance; i.e., hematopoietic reconstitution in patients with hematological malignances, and describe any additional data of which you are aware that could potentially support additional indications.

3. Please discuss any additional comments you have about the recommendations provided in the Draft Guidance to assist cord blood manufacturers in preparing information to be submitted in the Biologics License Application (BLA) for their HPC-C.

4. Please discuss what data would be adequate to demonstrate safety and efficacy of HPC-A, and to consider an approach to licensure of HPC-A similar to the one proposed for cord blood.



1.      Rubinstein P, et al. Processing and cryopreservation of placental/umbilical cord blood for unrelated bone marrow reconstitution. Proc Natl Acad Sci U S  A 1995;92:10119-22

2.      Gluckman E, et al. Outcomes of cord-blood transplantation from related and unrelated donors. Eurocord Transplant Group and the European Blood and Marrow Transplantation Group. N Engl J Med, 1997 Aug;337(6):373-81

3.      Wagner JE, et al. Transplantation of unrelated donor umbilical cord blood in 102 patients with malignant and nonmalignant diseases: influence of CD34 cell dose and HLA disparity on treatment-related mortality and survival. Blood. 2002 Sep;100(5):1611-18

4.      Rodriguez R, et al. Predictive utility of the attached segment in the quality control of a cord blood graft. Biol Blood Marrow Transplant. 2005 Apri;11(4):247-51

5.      Goodwin HS, et al. Long term cryostorage of UC blood units: ability of the integral segment to confirm both identity and hematopoietic potential. Cytotherapy. 2003;5(1):80-6

6.      Solves P, et al. Utility of the segment and cryovial samples for quality control and confirmatory HLA typing in umbilical cord blood banking. Clin Lab Haematol. 2004 Dec;26(6):413-8

7.      Cairo MS, et al. Characterization of banked umbilical cord blood hematopoietic progenitor cells and lymphocyte subsets and correlation with ethnicity, birth weight, sex, and type of delivery: a Cord Blood Transplantation (COBLT) Study. Transfusion. 2005 Jun;45(6):856-66

8.      Rubinstein P et al., Outcomes among 562 recipients of placental blood transplants from unrelated donors. N Engl J Med. 1998 November;339(22):1565-1577