Pharmaceutical Forced Degradation (Stress Testing) Endpoints: A Scientific Rationale and Industry Perspective.

Forced degradation (i.e., stress testing) of small molecule drug substances and products is a critical part of the drug development process, providing insight into the intrinsic stability of a drug that is foundational to the development and validation of stability-indicating analytical methods. There is a lack of clarity in the scientific literature and regulatory guidance as to what constitutes an "appropriate" endpoint to a set of stress experiments. That is, there is no clear agreement regarding how to determine if a sample has been sufficiently stressed. Notably, it is unclear what represents a suitable justification for declaring a drug substance (DS) or drug product (DP) "stable" to a specific forced degradation condition. To address these concerns and to ensure all pharmaceutically-relevant, potential degradation pathways have been suitably evaluated, we introduce a two-endpoint classification designation supported by experimental data. These two endpoints are 1) a % total degradation target outcome (e.g., for "reactive" drugs) or, 2) a specified amount of stress, even in the absence of any degradation (e.g., for "stable" drugs). These recommended endpoints are based on a review of the scientific literature, regulatory guidance, and a forced degradation data set from ten global pharmaceutical companies. The experimental data set, derived from the Campbell et al. (2022) benchmarking study,1 provides justification for the recommendations. Herein we provide a single source reference for small molecule DS and DP forced degradation stress conditions and endpoint best practices to support regulatory submissions (e.g., marketing applications). Application of these forced degradation conditions and endpoints, as part of a well-designed, comprehensive and a sufficiently rigorous study plan that includes both the DS and DP, provides comprehensive coverage of pharmaceutically-relevant degradation and avoids unreasonably extreme stress conditions and drastic endpoint recommendations sometimes found in the literature.

Assessing the Relevance of Solution Phase Stress Testing of Solid Dosage Form Drug Products: A Cross-Industry Benchmarking Study.

Stress testing (also known as forced degradation) of pharmaceutical products has long been recognized as a critical part of the drug development process, providing foundational information related to intrinsic stability characteristics and to the development of stability-indicating analytical methods. A benchmarking study was undertaken by nine pharmaceutical companies and the Brazilian Health Regulatory Agency (Agência Nacional de Vigilância Sanitária, or ANVISA) with a goal of understanding the utility of various stress testing conditions for producing pharmaceutically-relevant chemical degradation of drugs. Special consideration was given to determining whether solution phase stress testing of solid drug products produced degradation products that were both unique when compared to other stress conditions and relevant to the formal drug product stability data. The results from studies of 62 solid dosage form drug products were compiled.  A total of 387 degradation products were reported as being observed in stress testing studies, along with 173 degradation products observed in accelerated and/or long-term stability studies for the 62 drug products.  Among these, 25 of the stress testing degradation products were unique to the solution phase stress testing of the drug products; however, none of these unique degradation products were relevant to the formal stability data. The relevant degradation products were sufficiently accounted for by stress testing studies that included only drug substance stressing (in solution and in the solid state) and drug product stressing (in the solid state). Based on these results, it is the opinion of the authors that for solid dosage form drug products, well-designed stress testing studies need not include solution phase stress testing of the drug product in order to be comprehensive.

Inhibition of N-Nitrosamine Formation in Drug Products: A Model Study.

Nitrosamines, in the absence of toxicological data, are regarded as potential mutagens and need to be controlled at nanogram levels in drug products. Recent high profile product withdrawals have increased regulatory scrutiny of nitrosamine formation assessments for marketed products and for new drug applications. Formation of nitrosamine in drug product is possible when nitrite and vulnerable amines are present. Nitrite is often present as an impurity in excipients at ppm levels, whereas vulnerable amines, if present, stem mainly from the drug substance or its major impurities. In the event a drug product were to contain a major source of vulnerable amines (such as a moiety in the drug substance), it would be desirable to have an inhibitor which could be added to the formulation to minimize nitrosamine formation.  This work demonstrates, for the first time, that the inhibition of nitrosamine formation in oral solid dosage forms is indeed feasible with suitable inhibitors. Five inhibitors investigated (ascorbic acid, sodium ascorbate, α-tocopherol, caffeic acid, and ferulic acid) showed >80% inhibition when spiked at ∼1 wt% level. This work has also shown the potential use of amino acids (glycine, lysine, histidine) as inhibitors of nitrosamine formation in solution.

In-Use Photostability Practice and Regulatory Evaluation for Pharmaceutical Products in an Age of Light-Emitting Diode Light Sources.

This article describes how the increased use of energy-efficient solid-state light sources (e.g., light-emitting diode [LED]-based illumination) in hospitals, pharmacies, and at home can help alleviate concerns of photodegradation for pharmaceuticals. LED light sources, unlike fluorescent ones, do not have spurious spectral contributions <400 nm. Because photostability is primarily evaluated in the International Council of Harmonization Q1B tests with older fluorescent bulb standards (International Organization for Standardization 10977), the amount of photodegradation observed can over-predict what happens in reality, as products are increasingly being stored and used in environments fitted with LED bulbs. Because photodegradation is premised on light absorption by a compound of interest (or a photosensitizer), one can use the overlap between the spectral distribution of a light source and the absorption spectra of a given compound to estimate if photodegradation is a possibility. Based on the absorption spectra of a sample of 150 pharmaceutical compounds in development, only 15% would meet the required overlap to be a candidate to undergo direct photodegradation in the presence of LED lights, against a baseline of 55% of compounds that would, when considering regular fluorescent lights. Biological drug products such as peptides and monoclonal antibodies are also expected to benefit from the use of more efficient solid-state lighting.

Enrichment of Relevant Oxidative Degradation Products in Pharmaceuticals With Targeted Chemoselective Oxidation.

The ability to produce and isolate relatively pure amounts of relevant degradation products is key to several aspects of drug product development: (a) aid in the unambiguous structural identification of such degradation products, fulfilling regulatory requirements to develop safe formulations (International Conference on Harmonization Q3B and M7); (b) pursue as appropriate safety evaluations with such material, such as chronic toxicology or Ames testing; (c) for a specified degradation product in a late-stage regulatory filing, use pure and well-characterized material as the analytical standard. Producing such materials is often a resource- and time-intensive activity, either relying on the isolation of slowly formed degradation products from stressed drug product or by re-purposing the drug substance synthetic route. This problem is exacerbated if the material of interest is an oxidative degradation product, because typical oxidative stressing (H2O2 and radical initiators) tends to produce a myriad of irrelevant species beyond a certain stress threshold, greatly complicating attempts for isolating the relevant degradation product. In this article, we present reagents and methods that may allow the rapid and selective enrichment of active pharmaceutical ingredient with the desired oxidative degradation product, which can then be isolated and used for purposes described above.

Iron(III)-Mediated Oxidative Degradation on the Benzylic Carbon of Drug Molecules in the Absence of Initiating Peroxides.

Metal ions play an important role in oxidative drug degradation. One of the most ubiquitous metal ion impurities in excipients and buffers is Fe(III). In the field of oxidative drug degradation chemistry, the role of Fe(III) has been primarily discussed in terms of its effect in reaction with trace hydroperoxide impurities. However, the role of Fe(III) acting as a direct oxidant of drug molecules, which could operate in the absence of any hydroperoxide impurities, is less common. This work focuses on Fe(III)-induced oxidation of some aromatic drug molecules/drug fragments containing benzylic C-H bonds in the absence of initiating peroxides. Alcohol and ketone degradates are formed at the benzylic carbon atom. The formation of a π-stabilized cation radical is postulated as the key intermediate for the downstream oxidation. Implications are briefly discussed.

Surface-enhanced Raman scattering substrate based on a self-assembled monolayer for use in gene diagnostics.

The development of surface-enhanced Raman scattering (SERS)-active substrates for cancer gene detection is described. The detection method uses Raman active dye-labeled DNA gene probes, self-assembled monolayers, and nanostructured metallic substrates as SERS-active platforms. The mercaptohexane-labeled single-stranded DNA (SH-(CH(2))(6)-ssDNA)/6-mercapto-1-hexanol system formed on a silver surface is characterized by atomic force microscopy. The surface-enhanced Raman gene (SERGen) probes developed in this study can be used to detect DNA targets via hybridization to complementary DNA probes. The probes do not require the use of radioactive labels and have a great potential to provide both sensitivity and selectivity. The effectiveness of this approach and its application in cancer gene diagnostics (BRCA1 breast cancer gene) are investigated.

High-acidity determination in salt-containing acids by optical sensors. The scope of a dual-transducer approach and the Hammett acidity function.

A dual-transducer approach based on sol-gel optical sensors was recently reported to measure acid and salt concentrations, C(acid) and C(salt), in concentrated aqueous LiCl-HCl, CaCl2-HCl, and AlCl3-HCl solutions (C(acid) at 5-6 M; C(salt) < or = 2 M). The scope of this new approach has been studied in salt-containing HCl solutions with C(acid) at 2-9 M, and factors that influence sensor responses and accuracy have been investigated. A linear relationship between (deltaA/deltaC(salt))C(acid) and (dA/dC(acid))C(salt)=0, which is the basis of this dual-transducer approach, was found to lead to an empirical linear relationship between (deltaH0)C(acid) and (deltaC(salt))C(acid) (H0: Hammett acidity function of the indicator encapsulated in the sensor).