Chemistry Development for Phase 1 API Manufacture: A Pragmatic Fit-For-Purpose Approach
Introduction
The development of a synthetic route for clinical API manufacturing is a complex process which encompasses many elements, involves interdepartmental cooperation, and must be performed within constantly evolving and expanding regulatory guidance and good practice. It is therefore important in early phase chemistry development to adopt a pragmatic approach where efforts are focussed on key areas. This focussed approach can support aggressive timelines and the early generation of the API to supply pivotal preclinical toxicology studies, solid form and formulation development, as well as related analytical work and reference standard requirements. Sufficient process control and safety data must also be demonstrated to allow cGMP manufacture in support of a phase 1 clinical trial and to satisfy the CMC requirements of an IMPD/IND submission.
Synthetic route screening and optimisation
It is often the case that the proposed synthetic route for scale-up hinges on the success of a key chemical transformation which may be expected to be challenging. Fast-tracking of material through the synthetic route is therefore desirable to allow key stages to be assessed early. To simplify the reaction impurity profiles, analysis and data interrogation of challenging chemistry, it is important to use input materials of suitable purity, with chromatography often being the most time-efficient method to achieve this. To demonstrate proof of concept, a variety of conditions will be examined, drawing from both literature references and in-house knowledge.
By evaluating the key stages of the synthesis first, it is possible to rapidly make decisions regarding the appropriate synthetic route that offers the best chance of success. This avoids unproductive work on pathways that ultimately do not prove suitable for scale-up. In addition, rapidly progressing material to the API stage naturally supports the generation of samples of all synthetic intermediates that are required for initiating analytical method development and defining a solid form reference data set. The key impurity species may also be observed during this scouting run. This not only aids analytical method development but also allows for subsequent reaction optimisation to minimise their formation and/or demonstrate control during workup and purification.
The success of these early reactions, with respect to literature examples, may often depend on subtle changes in solubility, electronics or pKa associated with the specific target molecule. Therefore it is important to screen a wide range of possible reagents and conditions. This work can be greatly accelerated by adopting carefully considered experimental design and the efficient use of parallel synthesis equipment. Of perhaps even greater importance is the iterative process by which experimental data is tabulated, interrogated and discussed within the project team so that important trends and conclusions can be made to inform future experiments. Time is therefore saved by performing only sensible reactions which serve to prove or disprove a working hypothesis. This helps develop an understanding of key process parameters and ultimately gives a more robust process.
Once starting conditions are identified, some reaction optimisation is typically performed to further optimise basic parameters such as stoichiometry, solvent choice, temperature, reaction time and addition regime. Success is usually judged based on increased yield and reaction throughput or, more importantly, a cleaner reaction profile which gives fewer problem impurities.
Especially pertinent with early phase development is knowing when to stop the optimisation process. Often, suitable conditions are identified fairly early on which, with minor changes, can be the basis of a successful process. If a molecule progresses in the clinic to phase 2 or beyond, further optimisation can be scheduled ahead of future campaigns. Throughout this fit-for-purpose approach, it is important to keep in mind the eventual scale and number of batches required when deciding if suitable conditions have been identified. The time saved on additional reaction screening work can be invested into workup and purification development. This area deserves equal consideration as even the best reactions will generate impurities and by-products that require removal, and an efficient work-up and purification will save on processing time and provide a more robust process.
Reaction workup
The choice of reaction solvent is sometimes influenced by ease of workup, especially where liquid-liquid extractions are utilised. While considering workup steps, particularly if they are based on a literature procedure, it is important to ask what each step is achieving and if it is really required for the process. For example, multiple washes can often be omitted if testing shows a single wash of suitable volume is sufficient. Differences in polarity and ionisability should be utilised as a purification opportunity where they exist.
If the reaction product can be encouraged to solidify or crystallise then solubility screening is always worthwhile, including assessment both at ambient and elevated temperatures. Ideally, a recrystallisation will be identified from a suitable solvent or anti-solvent mixture. Combined with a carefully designed workup this can offer sufficient impurity control and allow removal of chromatography from the process. Isolation of a solid is also preferred since this avoids the requirement for concentration to dryness, which is time-consuming on a large scale. Where this is not possible, telescoping a solution into the next step may be appropriate (either directly or after performing a solvent swap).
Where a solid form exists for the API, developing a form-specific final product recrystallisation is often critical to achieving the target molecule’s impurity control and other critical quality attributes (CQAs). A formal polymorphism study and subsequent crystallisation development program should typically be performed, ideally before large-scale production for toxicological and clinical evaluation. An integrated approach between the chemistry and solid state teams allows problem impurities to be identified early, alternative control strategies to be implemented and useful solubility data and observations transferred back to influence late-stage reactions and workups.
Safety
Consideration of process safety throughout the development cycle begins with a theoretical literature review to detect the presence of known or potential hazards (e.g. reagent incompatibilities, highly exothermic processes, presence of potentially explosive species, known carcinogens and highly toxic reagents with low exposure limits). Where processing and material hazards are unavoidable, a well-defined COSHH review form enables controls appropriate to the intended reaction scale to be deployed. Diligent observations made during small-scale scouting reactions will then confirm the presence of any predicted reaction hazards and serve to capture any unforeseen events before they become problematic (e.g. exotherms, and off-gassing).
To ensure all reaction hazards are captured and well understood a stepwise approach to scale-up is employed (an increase in scale of no more than 5-10x is enforced). A representative batch will then be completed in a jacketed vessel whose design closely mimics that of the larger process. By carefully monitoring and logging overall heat flow, the active process can be reviewed to ensure adequate controls are in place for initial scale-up without the requirement for a formal adiabatic assessment of reaction thermodynamics. This may result in process modifications to ensure a complete reaction is achieved with no dangerous accumulation of reagents or potential for uncontrolled thermal events. This approach is valid for the limited scale-up typically performed at Onyx (up to 50L) with further safety testing generally required to support larger batch sizes (for example when moving towards pilot plant).
Where high-energy species are in use which present a higher risk profile (for example azides with an ’azide number’ approaching but not less than 3), further assessment of stability is always warranted. Initially, differential scanning calorimetry (DSC) thermographs can be collected for starting materials, intermediates and reaction mixtures with any exotherms approaching or exceeding 500 J/g giving cause for concern. Where exothermic events are present, appropriate processing and drying temperatures can be specified (limited to at least 100°C below the onset of any DSC exotherm) alongside any alternative and additional testing or control steps that are considered necessary. Our approach to health and safety at work is discussed in more detail as part of our next blog article.
Impurities
Consideration must also be given to the toxicity of the reagents being used, both in terms of operator safety during processing, and also with respect to the safety of the product. An assessment of any potential mutagenic impurities (PMIs) should always be performed as part of development. These can be present as reagents, reaction products or reaction by-products. Where present, suitable control strategies must be developed in line with ICH M7 guidance either based on a purge argument, analytical testing, or often a combined approach. For further details on Onyx Scientific’s approach to PMIs click here. A separate N-nitrosamine risk assessment is always performed as described here.
The use of LCMS and NMR for routine reaction monitoring often allows for structures to be proposed for any impurity species observed with little additional effort. By tabulating these species and tracking their levels throughout synthesis using the final product HPLC method, a better understanding of the API impurity profile can be gained which becomes increasingly invaluable at later stages in the development pipeline. By performing a pre-validation exercise on the chemical purity HPLC method at the time of development, additional confidence can be gained that the method will be stability indicating and should validate without issue for subsequent GMP usage. This reduces the likelihood of future method change requirements and means a library of representative data from all synthetic campaigns is available for direct comparison and troubleshooting.
Documentation
The last stage of the initial development process is producing documentation to clearly define the process. Technical data including procedures, observations, safety data, raw material and intermediate specifications and relevant analytical methods should be compiled into a Process Description. This document can be used to facilitate the transfer of information to other departments and enable the writing of batch records for both nonGMP and GMP use, as well as the drafting of relevant analytical monographs. A development report is also key for summarising all aspects of the development effort and outlining the thinking behind any decisions made. Sections include a listing of observed impurity species and their fate, PMI and N-nitrosamine risk assessments and any areas where future work may be beneficial.
In conclusion
Leveraging our comprehensive expertise and streamlined workflows, we can deliver fit-for-purpose processes within aggressive timelines. This includes the supply of multi-gram scale advance batches to support ongoing project needs such as pre-formulation and pharmacokinetics. Ultimately the goal is to develop processes that are robust, efficient and safe, while also generating all accompanying paperwork clearly outlining the process and transferring all technical knowledge between departments and to our clients.