Analytical and regulatory CMC strategies for oligonucleotide therapeutics

At TIDES Europe in November 2020, Thomas Rupp, Owner & Principal at Thomas Rupp Consulting UG spoke about what makes the CMC of oligonucleotides so unique, the regulatory background and the origin and the nature of impurities. Here, we look at the highlights of the presentation.

Vitravene oligo drug established a new class of therapeutics in 1993. From a regulatory standpoint they are chemical entities, but they are bigger than typical small molecules. They are large polyanionic compounds that carry features of both typical drugs and biologics. They are produced by chemical syntheses which is a predictable, but impure, process. Like biological entities, oligos are difficult to fully characterize because of their molecular composition and because of their heterogeneity.

Since 1993, the US FDA has regulated oligos as big small molecules and not as biologics. That means, in the US, most new oligo drug applications go to the Center for Drug Evaluation and Research (CDER), and in Germany, the equivalent is the Federal Institute for Drugs and Medical Device (BfArM).

There are exceptions which may be received by the Center for Biologics Evaluation and Research (CBER) in the US or the Paul-Ehrlich-Institut (PEI) in Germany. For example, if oligos are used as an adjuvant in a vaccine or if they have an aptameric structure which is not sequence dependent.

Guidelines on oligonucleotide impurities

There are no international guidelines or specifications for control strategies of impurities. Like synthetic peptides, oligos were excluded from the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) guidelines Q3A & Q3B, which refer to impurities in drug substances (DS) and drug products (DP), and the Q6A specifications for new DS and DP. They are also excluded in the foreword of the ICH guidelines.

As there are no formal guidelines for therapeutic oligos on limits and thresholds of impurities, they are discussed with the respective regulatory authorities on a case by case basis. Such discussions are based upon the safety and class complexity of the molecule and the formulation. Sponsors have the flexibility of adapting guidelines for other entities (synthetic peptides for example, biologics, antibodies) with appropriate justification for the oligo.

In terms of how the usual ICH Q3A requirements on impurities can be extrapolated to oligos, Rupp focused on starting materials, byproducts, and degradation products. Dealing with oligo impurities requires a thorough understanding of the quality of the raw materials, and the chemistry during synthesis and downstream processing. Understanding the stability of the oligo compound during the syntheses, downstream processing and during the storage is important. Degradation pathways should be known.

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Impurity classification

A 2017 white paper by the oligo safety working group (OSWG) suggested to categorize impurities into four different impurity classes and assess safety and potency only for impurities classified as critical and non-natural. According to Rupp, this whitepaper is well recognized by drug sponsors, and agencies when defining CMC control strategies.

The publication contains a useful decision tree for impurity classification. Accordingly, class I impurities are impurities that are also major metabolites. The structure and the sequence are the same as the parent compound. Class II impurities contain only structural elements found in naturally occurring nucleic acids. Class III; impurities that are sequence variants of the parent oligo. That is base deletions or additions and also deaminations. Class IV are impurities that contain structural elements not found in the parent oligo or in naturally occurring nucleic acids. These are base modifications, backbone modifications, depurinated species, and unidentified impurities and this is the only class that requires a safety assessment if the impurities are above the threshold.

An impurity identification threshold of 1% and a qualification threshold of 1.5% is suggested. Typically, industry uses a 0.5% threshold for identification and 1.5% for the qualification.

Analytical methods

Rupp recommends, firstly, to avoid impurity formation, if at all possible, because not all impurities are easily purified out. In a good control strategy, you would use orthogonal analytical methods to support identification and separation of different impurities.

If critical impurities are detected above the threshold level, the performance of supporting in vitro studies to allow proper classification and comparison to the parent compound and to assess the safety in vitro and in vivo are suggested. It is a good idea to trace back identified impurities to the relevant process step and perform selective post-development to avoid the formation of these impurities.

ICH Q3 states that any molecule that is different to the parent molecule (full-length oligo) is categorized as impurity. Oligos are excluded from ICH Q3A (impurities in DS) and IHC 3B (impurities in DP) and there are no explicit guidelines on process-related impurities available. According to the OSWG classification system, class IV impurities require a safety assessment. Analysis by high-resolution LC-MS can allow tracing the origin of each impurity back to the respective process step and allows for successive process optimization.

Further Reading

Capaldi, Daniel, et al. “Impurities in Oligonucleotide Drug Substances and Drug Products.” Nucleic Acid Therapeutics, vol. 27, no. 6, 1 Dec. 2017, pp. 309–322., doi:10.1089/nat.2017.0691.

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