The number of reserve samples required by current regulations has been a sore subject for both NDA and ANDA sponsors as well as testing facilities as the cost and storage requirements for the required reserve samples of both test and reference product was overly burdensome.  Now, after twenty years of concern expressed by the industry, the FDA’s revised guidance, titled Handling and Retention of BA and BE Testing Samples (here), is being issued in part as a final guidance and in part as a draft.  It adjusts the reserve sample requirements based on years of practice, new methods of analytical testing, and a change in the FDA’s thinking regarding enforcement policy related to sample retention.  As the Agency stated, “[t]his guidance is a revision of the previously issued final guidance from May 2004 and responds to comments received to the ‘Compliance Policy for the Quantity of Bioavailability and Bioequivalence Samples Retained Under 21 CFR 320.38(c)’ guidance issued in August 2020, now withdrawn upon publication of this guidance.” 

The FDA has reevaluated the regulatory requirement to retain five times the quantity of reserve samples for conducting analytical testing and, based on current advances in analytical testing methods and techniques, the FDA now believes that a reduced number of reserve samples for testing is sufficient.  As a result, the FDA has revised the current Compliance Policy Guide by stating, “Accordingly, at this time and based on FDA’s current understanding of the risks involved, FDA generally does not intend to enforce the requirement to retain a sufficient quantity to perform five times all the release tests required in the application or supplemental application, so long as the recommended lower quantities in this guidance are retained.”  The FDA is taking action by issuing this guidance and revised enforcement policy because it cannot just change regulations.  I am certain that, at some point in the future, the FDA will move to revise the regulations to comply with its current and less burdensome approach. 

“The guidance highlights: 

  • How the test article (T) and reference standard (RS) for BA and BE studies should be distributed to the testing sites. 
  • How testing sites should randomly select samples for testing and material to maintain as reserve samples. 
  • How the reserve samples should be retained. 

Additionally, this guidance: 

  • Addresses the requirement at 21 CFR 320.38(c) to retain reserve samples of sufficient quantity to permit FDA to perform five times all the release tests required in an application or supplemental application. 
  • Describes the conditions under which the Agency does not generally intend to take enforcement action against an applicant or CRO for retaining less than the quantity of reserve samples of the test article and reference standard that were used in the BA or BE study as specified in 21 CFR 320.38(c).” 

There is a section in the guidance that clarifies which samples used for in vitro testing require reserve sample retention and provides examples of those that do not require retention.  For instance, samples used for in vitro dissolution testing typically do not require sample retention because they relate to a waiver provision and not establishment of bioequivalence, but in vitro testing for inhaled products as described in product-specific guidances are generally used to establish bioequivalence and, thus, are subject to sample retention. 

The section on Selection Techniques provides several examples of the proper way for a drug sponsor to submit drug samples to a testing facility or CRO to ensure that the facility may randomly select product to be used in bioequivalence testing.  In addition, this section describes common problems that FDA investigators have seen during investigations. 

The guidance recommends that the following number of reserve samples be retained: 

For in vivo studies:  A minimum quantity of 30 single-dose (SD) or 3 multi-dose (MD) units each of the T and RS (and P if applicable) in the original container in total across all testing sites with at least 1 unit in the original container per treatment (or blinded kit, as applicable) retained from each shipment used in the BA or BE study. 

For in vitro studies:  A minimum quantity of 30 SD or 3 MD units in the original container per batch each of the T and RS in total for all in vitro studies conducted at the testing site with at least 1 unit in the original container each of the T and RS retained from each shipment used in the BE studies. 

This compliance policy is applicable to all reserve samples for BA and BE studies held to date, including reserve samples from previously completed BA or BE studies.” 

There are some caveats regarding multiple testing sites and the differences between in vitro and in vivo testing that are further explained relative to stored reserve samples so be careful to read and understand each of the requirements to make sure that you understand what is expected of your firm.  Reserve samples should be stored at appropriate recommended storage conditions and should be retained by the testing site or independent third-party for a period of at least five years “following the date on which the application or supplemental application is approved, or the date of completion of the BA or BE study if such application or supplemental application is not approved.”  The guidance emphasizes that retain samples should never be returned to the sponsor. 

In the guidance, there are further examples and discussion of the requirements as well as a historical discussion of the genesis of the requirement for retain samples.  It is very comprehensive and should provide sponsors with the necessary information to avoid most (if not all) issues that may befall an applicant should the samples not be handled properly by the sponsor or the testing site.