Probiotics in Clinical Research: Stability Studies Matter

IPA AdminIPA In The Lab, Uncategorized

By Josh Baisley, B.Sc., Associate Director of Clinical Trials, Nutrasource

The importance of stability studies is highlighted in cases where a human intervention trial  demonstrates equivocal results, or worse yet, negative results. When there was no evidence to demonstrate that the probiotic survived for the duration of the study period, it could be plausible that results were due the premature expiration of the study product. However, this may not be the only factor, thus a second trial may or may not prove successful.  If on the other hand, one had evidence to demonstrate the product was stable for the duration of the study and the study still showed equivocal results or negative results, it could not be attributed to the viability of the investigational product.  As with nearly all factors involved with planning clinical research, stability testing provides a level of risk management.

Demonstrating the stability of a probiotic being used in clinical research is necessary to ensure that the strain(s) meet specifications for the duration of the clinical study period. The amount of data required will be dependent on the expected duration of the study, that is the time from the date the product is manufactured to the time the last subject enrolled has their last study visit/last data collected.  This provides the longest time period the product was being investigated during the clinical research study.

Regulatory bodies such as Health Canada (and the FDA if filing an IND (see 21 CFR 312.23 (a)(7)(ii and iv))), require stability data to be submitted to support the clinical study, or where there is insufficient data, the Sponsor must commit to conducting real-time stability (at minimum) to demonstrate stability of the probiotic through the duration of the study. This requirement is a strategy to mitigate risk during the clinical investigation, as it is expected that the Sponsor will take action to replace the study product should it show signs that it prematurely expires prior to the end of the clinical study so as to not jeopardize any outcomes being assessed in the clinical trial.  If it is necessary to replace the study product during the clinical investigation, or use multiple lots of product, the final report must list the subjects and batch number(s) each received.

Monitoring real-time stability during the clinical trial is required by regulatory bodies when there is no existing information on prior lots of the specific formulation (including packaging) to be used in the study; however, there is often no consideration given to activities that occur in advance and during the clinical trial which can negatively impact the products stability. Many probiotic formulations require refrigeration.  Variations in storage conditions could have a negative impact on the viability of microorganisms that require refrigeration. Therefore, consideration must be given to transport of the clinical study product prior to initiating the study (for example, transport to the CRO and transfer from the CRO to the site) as well as how clinical trial subjects handle study supplies.  With respect to the clinical trial subjects, will they follow the storage instructions, what if they were to carry it around during the day at work, what if the bottle of study product was left in the car on a hot summer day, etc.).

To address this, Sponsors should consider sending research sites additional product that will be used to test survivability of the probiotic through shipment to the CRO/site, and during storage at the site. It may also be prudent to analyze some of the products returned by subjects to understand if the probiotics survived (met minimum specification requirements) for the duration the products were in their possession.  This provides evidence that the probiotics were maintained under conditions that ensure their survival for the duration of the study including through shipping, handling and return.

While regulatory bodies expect that stability testing will be conducted under temperatures for which the product is expected/labelled to. To fully understand attributes of the probiotic culture, it may also be prudent to test the product under temperatures/conditions of stress and determine viability. This provides the Sponsor with additional information to determine impact of an variance seen in the handling of clinical study product, either during shipment, at the CRO/site or how the subject has handled the product during the dosing phase.  Stability information should be gathered to understand how the probiotic strain(s) respond to all conditions that it may be exposed to during the clinical investigation (shipping, warehousing, how subjects will handle them, etc.).

Additional ways to mitigate risk related to stability of the investigational probiotic in the context of clinical research includes: shipping products in validated packaging with temperature tails/data loggers, to demonstrate conditions the probiotic was exposed to during transit (to and from the CRO and or site(s)). Data loggers can record temperatures on a pre-set schedule such as every 5 minutes, every minute or in fractions of minutes.  Data loggers provide information on any excursions to the specified storage conditions as well as duration of excursion.  The data can then be compared to existing stability data to determine acceptability of the probiotic prior to use in the clinical study, or after use in the clinical study. When shipping internationally, us a shipper that ensures appropriate storage also when the product is ‘in customs’.

When conducting stability studies, guidelines such as ICH Q1A(R)2 should be followed. It is important that if your product has nuances that prevent standard ICH guidelines from being followed for stability testing, that justification for any modification to the standard protocol is well documented and justified.  Stability testing should be conducted in the packaging material proposed to be used in the clinical investigation and under similar conditions.

As with all best practices there may be nuances that warrant consideration for your specific product, and some practices that may not be applicable due to the specific nature of your probiotic. For example, accelerated stability testing may not be appropriate for some probiotic strains due to the requirement to expose the product to high temperature and humidity, thus accelerated studies may be excluded from the stability test profile when scientifically justified.  In any case, accelerated stability testing is only indicative and does not replace real time stability testing.

In summary, the following best practices warrant consideration, especially for probiotics being used in clinical research that are sensitive to extreme conditions (e.g. heat or cold sensitive):

  • Ship using a validated shipping container and include a digital data logger
  • Ship using a company specializing in cold chain management; in particular with international shipping
  • Include extra bottles of product in the shipment for retains/returns for testing viability (e.g. prior to initiating trial, at end of clinical investigation to ensure quality of product for duration of study);               extends to shipping to CRO as well as site(s)
  • Conduct real-time studies concurrently with the clinical investigation (if new product or lack of information to support duration of study), allowing substitution of a new lot during trial if stability of product starts to drift
  • Collect and test random subject returns, provides confidence that study product was held appropriate by study subjects
  • Depending on the requirements of the probiotic, it may be necessary to provide the CRO and site(s) with coolers for transport if the probiotic is sensitive to heat exposure. This may also be an important logistic depending on location of the study site, or timing (season) the study will be conducted over.