Polymorph profiling with X-ray powder diffraction protects patents and patients

By Dr Natalia Dadivanyan, Segment Marketing Manager, Malvern Panalytical 

Small molecules can face big challenges 

Despite the emergence of, and excitement around, advanced therapeutic modalities, traditional small molecule pharmaceuticals still occupy a significant and steadily growing share of the market. A recent report indicated the current market size to be ~$166 billion, and a 5% compound annual growth rate is set to take the sector to a worth of $250 billion by 2028, offering an attractive landscape for developers.  

However, developing and bringing new small molecule products to market is fraught with challenges, and the failure rate remains high, driven primarily by later-stage efficacy, safety, and solubility concerns. While a candidate molecule’s chemical structure is a critical determinant of such characteristics, it isn’t the only one. Just as important, especially for oral solid dosage forms, is the molecule’s solid form — in particular, whether it exhibits polymorphism.  

Polymorphism, defined as the ability of a molecule to exist in two or more different crystal structures, has been the subject of increasing interest and scrutiny in the pharmaceutical sector. Different polymorphs of the same active pharmaceutical ingredient (API) can have profound effects on solubility, permeability, and bioavailability, as well as manufacturability. Combined with the fact that even slight changes in manufacturing processes and storage conditions can induce polymorph transitions, it’s clear to see why thorough, comprehensive polymorphic profiling is critical throughout the entire drug development and manufacturing process. Indeed, this is why polymorphic screening is recommended in the International Committee for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use (ICH) Q6A guidelines, developed to help ensure only the highest quality products are released to market. 

Polymorphic perils and possibilities

Companies who don’t comprehensively understand the presence, structure, and stability of polymorphic variants in their formulations risk safety, efficacy, and quality issues in their product. In the worst case, an undetected polymorphic change in a drug product can impact bioavailability such that the product delivers either a sub-therapeutic or toxic dose, the result of both scenarios potentially being the death of the patient. 

A high-profile case of the consequences of unidentified polymorphism is that of Norvir® (now known by its generic name ritonavir), an anti-viral protease inhibitor shown to be effective in treating patients with human immunodeficiency virus (HIV) and hepatitis C. In 1998, two years after Norvir’s release to market, a second polymorph created during manufacturing was discovered. This form was less biosoluble, meaning its efficacy was significantly reduced, creating lasting problems for patients. As a result, the drug was removed from the market until the manufacturing issues were remedied. The financial losses to the company were significant. 

While the need to profile polymorphs to ensure patient safety is clear, companies also need to keep in mind the business risk presented by undetected polymorphism — particularly with respect to intellectual property (IP) protection and infringement. For instance, if an innovator company fails to identify that its new API contains (or later converts into) an already-patented polymorph, product sales can ultimately be halted, and business activity derailed. A prominent example is that concerning the polymorphic forms of Zantac (ranitidine), a histamine-2 blocker used to lower stomach acid production. GSK originally patented two polymorphic forms of ranitidine. In 1984, TorPharm filed a patent for polymorphic form 1 of ranitidine, whose patent had recently expired. However, upon investigation of the drug product by GSK, a small amount — <0.5% — of polymorphic form 2 which was still under patent, was discovered. This tiny proportion of polymorphic form 2 in the formulation was enough to demonstrate patent infringement and, as a result, TorPharm was blocked from releasing their product until 2002 — at significant estimated revenue loss. 

When seen from the opposite side, unanticipated polymorphism can be a powerful opportunity for generics companies to capture market share while also improving the quality of medicines available to patients. This was seen in the case of atorvastatin, a cholesterol-lowering statin released under patent in 2000. Three years later, a generics manufacturer discovered and filed a patent for a more effective form of the drug not covered under the originator’s patent. The ensuing infringement case launched against the generics company ended in failure, and the generics company went on to sell its new product five years ahead of the originator’s patent expiring. Revenue for the generics company reached around $600 million in the first six months of the product entering the market. 

X-ray powder diffraction shows the big polymorphic picture

Many tools are available to support companies in their efforts to comprehensively identify and characterise polymorphs. One of the leading and most reliable is X-ray powder diffraction (XRPD), which works by detecting the X-ray diffraction patterns of analytes to elucidate their crystallographic structure. Because these diffraction patterns vary across (and are unique to) APIs and their different crystalline forms (Figure 1), XRPD offers analysts the ability to effectively identify, characterise, and monitor polymorphs. Having such insights delivers great benefit throughout the entire drug development journey, from discovery through scale-up, manufacturing, and even storage.

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Figure 1: XRPD diffractograms of two different polymorphs of an API: form I (top) and form II (bottom). The two forms have reflections (peaks) at different 2theta positions (the angle between transmitted and reflected beams), indicating different crystal structures. Different crystalline structures mean that these polymorphs will very likely have different properties such as bioavailability and solubility. Phase behaviour under non-ambient conditions will also be different between the polymorphs.

At the earliest stages of drug discovery, the comprehensive polymorphic profiling enabled by XRPD facilitates deeper investigation of alternative solid forms, including those that might have more favourable physicochemical and pharmacological characteristics. Because most new drugs in development are Biopharmaceutics Classification System (BCS) classes 2 and 4 (i.e., are poorly soluble and permeable), discovering and properly characterising all crystalline forms means substances with the best solubility and bioavailability can be identified and progressed, and those that might pose a risk discarded early. Moreover, because different crystalline forms can impact manufacturing-critical characteristics such as flowability and compressibility, having this complete picture of the polymorphic forms of your molecule means you can identify and progress the form best-suited to large-scale manufacturing.  

Through this — screening early for forms that make more efficacious, more manufacturing-friendly products — developers can foresee and mitigate costly issues, and better build a fail-early paradigm into their programs. Ultimately, this makes for more time- and cost-efficient drug development, but also ensures patients receive a better-optimised, safer therapeutic product. 

On top of this, having a full view of the crystalline forms (and amorphous forms, if existing) of a substance makes good business sense; pharmaceutical organisations with such knowledge can ensure all relevant polymorphs are included in their patent applications, thereby minimising the risk of their competitive standing being undermined. 

XRPD plays well with others 

XRPD, on its own, provides a suite of benefits to those looking to de-risk their small molecule development efforts. But XRPD can also be easily used in tandem with other analytical techniques to provide companies with an even broader understanding of their product, and with it, too, a better capacity to optimize drug development.  

For example, XRPD is often combined with thermal analysis techniques, such as differential scanning calorimetry (DSC) or thermogravimetric analysis (TGA). DSC records the transition temperature when a substance changes its state, making it possible for analysts to distinguish between different crystalline and amorphous forms. Through DSC, the thermal stability of a polymorph can be analysed, helping developers choose more stable polymorphs and better define the manufacturing and storage conditions that minimise the risk of polymorph transitions. Similarly, TGA, which records mass as a result of temperature change, can elucidate the thermal stability of a given API form. Again, such information is highly valuable in identifying optimal crystalline forms and setting conditions for manufacturing and storage. 

Classic XRPD protocols are also now increasingly being combined with X-ray scattering techniques to explore and characterise novel solid forms that offer promising solubility advantages — namely nanosuspensions and amorphous solid dispersions. For analysing nanomaterials, small-angle X-ray scattering (SAXS) is often the method of choice. Here, X-rays are beamed at a sample, and the intensity of scattered X-rays is measured as a function of scattering angle, where the angle is inversely proportional to sample structure size. The technique is highly versatile, and delivers a wealth of information about samples, including nanoparticle size distribution, particle shape, and particle structure.  

For analysis of intrinsically disordered materials, applying the pair-distribution function method (PDF) is particularly useful. This technique uses the complete powder XRD pattern to provide structural information about amorphous, poorly crystalline, nano-crystalline, or nano-structured substances. 

With the broader and deeper structural information provided by combining XRPD with other analytical techniques, pharmaceutical companies are better empowered to make well-informed, future-proof choices in their API development.  

A solid choice for solid form analysis

XRPD operates in a landscape of many other analytical techniques, from single crystal X-ray diffraction (SC-XRD) and three-dimensional electron diffraction (3D ED) to Fourier transform infrared spectroscopy (FTIR) and Raman spectroscopy. Pharmaceutical organisations thus have many techniques to choose from. However, XRPD boasts several significant advantages over competing analytical methods when it comes to exploring pharmaceutical solid forms. 

SC-XRD has been used for more than 70 years, and its ability to determine the absolute structure of a crystal (including atomic bond length, position, and angle, and crystal density and disorder) is highly valued. But while the technique is good for generating such absolute reference data, it is not suitable as a bulk characterisation technique. This is because the process of obtaining a suitable crystal for analysis, one that is both properly-sized and of high enough quality, is very time-intensive. In contrast, XRPD is highly suited to the bulk measurement of powder samples; sample preparation is simple, and the technique can speedily generate a diffractogram that can indicate multiple polymorphs as well as amorphous material. Accordingly, XRPD can be seen to offer a superior analytical solution in the polymorph screening context, providing faster and more holistic insight into an API.  

3D ED is another analytical technique that allows structure determination from individually selected crystals. Compared to SC XRD, 3D ED can work with nanometre-sized crystals, saving the scientists a lot of time and effort on growing crystals large enough for an SC XRD experiment. Recent advances have made 3D and microcrystal ED techniques more viable. However, ED expertise and equipment is still broadly inaccessible, and the technique’s cost is prohibitive for most organisations. XRPD, on the other hand, has been used for many years, and so is both widely accessible and at a price point that is much more affordable, for a broader range of pharmaceutical companies. 

FTIR has been a popular choice with pharmaceutical companies looking to better understand polymorphs, too. The technique is fast, simple to use, and highly sensitive. But again, it lags behind XRPD in its performance: more specifically, it is poorly suited to sample analysis at elevated temperatures. In such conditions, infrared light absorption can degrade the sample, leaving analysis of some structures off-limits. 

Raman spectroscopy is a common analytical technique that is increasingly considered as a complimentary tool to XRPD for solid form characterisation. Raman spectroscopy uses scattered light to measure the energy modes of a sample, providing a molecular fingerprint to aid chemical identification. However, while Raman spectroscopy allows analysts to distinguish between different polymorphs of an API, it does not give insight into the crystal structure of these forms, unlike XRPD. 

Product (re)assurance 

The small molecule pharmaceutical market is exceptionally competitive, and carving a profitable space in it is becoming increasingly challenging. Many of the hurdles that continue to plague developers — achieving favourable solubility, permeability, and bioavailability, and ensuring ideal handling and manufacturing characteristics — can be simplified or overcome entirely with thorough identification and characterisation of a candidate molecule’s polymorphs.  

For pharmaceutical companies looking to comprehensively analyse the solid form of their API and drug product, XRPD stands out as the technique of choice. Offering versatility, simplicity, and an ability to be augmented with complementary analytical techniques, XRPD is a robust tool that can bring significant value to drug development. With it, pharmaceutical developers can de-risk development programs, better serve patients with safer, more efficacious medicines, and rest assured that their IP is more adequately protected.

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