Patricia Van Arnum was executive editor of Pharmaceutical Technology.

Patricia Van Arnum

A new study of both computer-created and natural proteins suggests that the number of unique pockets, sites where small-molecule pharmaceutical compounds can bind to proteins, is small, thereby, offering an explanation of the difficulty in mitigating side effects in drug design.  

Studying a set of artificial proteins and comparing them to natural proteins, researchers at the Georgia Institute of Technology (Georgia Tech) concluded that there may be no more than about 500 unique protein pocket configurations that serve as binding sites for small-molecule ligands, according to a May 20, 2013 university press release. "Our study provides a rationalization for why a lot of drugs have significant side effects because that is intrinsic to the process," said Jeffrey Skolnick, a professor in the School of Biology at Georgia Tech and co-author of the recent study (1). "There are only a relatively small number of different ligand-binding pockets. The likelihood of having geometry in an amino-acid composition that will bind the same ligand turns out to be much higher than anyone would have anticipated. This means that the idea that a small molecule could have just one protein target can't be supported." 

Binding pockets on proteins are formed by the underlying secondary structure of the amino acids, which is directed by hydrogen bonding in the chemistry, which allows formation of similar pockets on many different proteins even those that are not directly related to one another. "You could have the same or very similar pockets on the same protein, the same pockets on similar proteins, and the same pockets on completely dissimilar proteins that have no evolutionary relationship," said Skolnick in the release. "In proteins that are related evolutionarily or that have similar structures, you could have very dissimilar pockets. This helps explain why we see unintended effects of drug and opens up a new paradigm for how one has to think about discovering drugs." 

The implications for medicinal chemistry could be significant. To counter the impact of unintended effects, drug developers will need to know more about the available pockets to avoid affecting binding locations that are also located on proteins crucial to life processes. If the unintended binding takes place on less crucial  proteins, the side effects may be less severe. In addition, drug development could also move to a higher level by examining the switches that modulate the activity of proteins beyond binding sites. "The strategy for minimizing side effects and maximizing positive effects may have to operate at a higher level," Skolnick said. "You are never going to be able to design unintended binding effects away. But you can minimize the undesirable effects to some extent." 

In their study, the researchers used computer simulations to produce a series of artificial proteins that were folded, but not optimized for function. Using an algorithm that compared pairs of pockets and assessed the statistical significance of their structural overlap, they analyzed the similarity between the binding pockets in the artificial proteins and the pockets on a series of native protein, according to the release. The artificial pockets all had corresponding pockets on the natural proteins, suggesting that the simple physics of folding has been a major factor in development of the pockets. "This is the first time that it has been shown that side effects of drugs are an inherent, fundamental property of proteins rather than a property that can be controlled for in the design," Skolnick added. "The physics involved is more important than had been generally appreciated." 

, online, DOI: 10.1073/pnas.1300011110, May 20, 2013.  

Articles

A recent study of computer-created and natural proteins suggests the number of sites where small-molecule drugs can bind to proteins is limited, thereby narrowing how to mitigate side effects through drug design.

Small-Molecule Drug Development: Recent Study Suggests Limited Binding Sites

Bringing a drug candidate to successful commercialization is always a challenging task, but it is one that has been made more difficult through the increase of poorly water-soluble drugs. It is estimated that 40% of new drug compounds may be regarded as poorly soluble with that percentage even higher for certain therapeutic classes (1). Improving the oral absorption and bioavailability of poorly soluble drugs is crucial for pharmaceutical companies seeking to bring efficacious drugs to patients in dosing regimens and product forms that are easy to use, affordable, and facilitate patient compliance. 

Image Courtesy of Catalent Pharma Solutions			

To understand the extent of this challenge and the technologies for bioavailability/solubility enhancement, 

 partnered with the Catalent Applied Drug Delivery Institute (see sidebar, "Advancing drug delivery") in the second annual 

Catalent–Pharmaceutical Technology Landscape Drug Delivery Survey

. The study surveyed formulation scientists involved with oral product development in pharmaceutical companies in the United States, Canada, and Europe to understand their key concerns and the technologies used to resolve problems of solubility and bioavailability. The results showed that there are a myriad of technical concerns attendant to solubility/bioavailability and many factors have to be taken into consideration in deciding which approach is optimal for a given API. 

solubility/bioavailability enhancement remains a major issue. Ninety-two percent of the 2013 survey respondents have worked with Biopharmaceutics Classification System (BCS) Class II (low solubility, high permeability) or Class IV (low solubility, low permeability) compounds, and half always or often work with these compounds.  

A poorly soluble compound adds to the complexity of formulation development. "Countless poorly soluble compounds never reach human clinical studies," comments Kurt Nielsen, PhD, chief technology officer and senior vice-president of R&D at Catalent Pharma Solutions. "Better understanding the chemistry of the drug, solubilizers (e.g., polymers, surfactants, and lipids) and the various salt/crystal forms can get more drug candidates through preclinical testing." 

The survey results confirm these observations. The two top problems identified by respondents when working with poorly soluble drugs were optimizing the drug-release profile (71% of respondents cited) and stability (66% cited) (see 

). More than half (58%) cited difficulties in identifying excipients with optimal properties, and 49% identified excipient–API interactions as a concern. As would be expected, permeability and absorption in the gastrointestinal tract were also key issues.  

Figure 1:  Problems encountered when developing oral formulations of poorly soluble APIs. (ALL FIGURES COURTESY OF AUTHORS)			

provides more in-depth insight. Fifty-three percent of respondents said that stability of the API was either "always" or "often" a challenge, and 46% cited inter- and intra-patient variability. Other formulation challenges, such as dose uniformity and the food effect on absorption, also were commonly encountered problems (see 

). "Often times, the bioavailability of a poorly soluble drug increases when given with high-fat meals," explains Catalent's Nielsen. "The changes in bioavailability induced by food can negatively impact assessment of safety and efficacy. In addition, eliminating the food effect simplifies dosing instructions for patients," he adds. 

Figure 2: Frequency of challenges when developing a formulation for a poorly soluble API. 			

Other concerns, such as swallowing/taste masking, were cited as frequent problems by 35% of respondents. "Child- and senior-friendly products rely on taste masking for dosing convenience," says Nielsen. "Poorly soluble drugs often need multiple technologies in the same engineered particle to achieve the required taste profile."  

Given the complexity of formulation challenges when working with poorly soluble drugs, the survey examined the importance of factors in deciding on a solubilization strategy. The physiochemical properties of the API and safety were both major factors (see 

). Interestingly, the depth and availability of expertise, including third-party expertise, were significant factors. Ninety-three percent of respondents said that internal expertise and knowledge were "very important" or "somewhat important," and access to third-party expertise and knowledge were commensurately important (see 

. More than half (53%) said bioavailability and solubility issues were a reason to partner.  

Figure 3: Importance of factors in choosing a solubilization strategy. 			

The survey also asked respondents to evaluate solubilization/bioavailability enhancement strategies (see 

). "Enhancing bioavailability is multifactorial and involves approaches to optimize the API, the formulation, and the processing technology to develop the final product form," explains Nielsen. "These factors are codependent and synergistic in most cases, so a 'best-in-class' toolkit for solubility enhancement includes multiple technologies," he adds. For example, a solubilization approach may first address salt formation and excipient selection. Other technologies, such as particle milling/micronization, nanoparticles, spray-drying, hot-melt extrusion, liquid-filled capsules, and softgels, can be further evaluated for suitability for a given formulation challenge.  

Figure 4: Evaluation of strategies/technologies in addressing poor solubility.  			

Ralph Lipp, PhD, head of Lipp Life Sciences, member of the advisory board of the Catalent Applied Drug Delivery Institute, and formerly vice-president of pharmaceutical sciences R&D at Eli Lilly, offered several examples of commercial drugs to illustrate the diversity of solubility/bioavailability enhancement strategies (3). The HIV treatment Kaletra (ritonavir and lopinavir) uses solid dispersions produced by hot-melt extrusion, and the antifungal drug Sporanox (itraconazole) is produced as a solid dispersion by spray-drying. The antinausea drug Emend (aprepitant) uses particle engineering (i.e., nanocrystals/media milling) to improve bioavailability. And the protease inhibitor Fortovase (saquinavir) is formulated in a self-emulsifying drug-delivery system in a softgel capsule (3).  

These advances in solubility and bioavailability enhancement are an integral part of meeting the needs of patients for efficacious drugs. "Several essential medicines based on advanced oral drug-delivery technologies provide significant value to patients around the globe today already," says Lipp. "In light of the current trend toward low solubility and poor bioavailability drug candidates in the pipelines of innovators, this class of technologies is of increasing relevance. Formulation scientists applying proven and novel oral drug-delivery technologies will significantly contribute to enabling the next generation of important medicines," concludes Lipp. 

 1. A. Fahr and D. Douroumous, "Preface" in 

Drug Delivery Strategies for Poorly Water Soluble Compounds

, A. Fahr and D. Douroumous, Eds. (John Wiley & Sons, Chichester, UK, 2013), p xxiii.  

 3. R. Lipp, "The Innovator Pipeline: Bioavailability Challenges," presented at DCAT Week, Mar. 14, 2013. 

Editorial podcast: strategies for bioavailability enhancement

Kurt Nielsen, PhD, chief technology officer and senior vice-president of R&D at Catalent Pharma Solutions, discusses solubilization strategies and bioavailability enhancement with 

 Executive Editor Patricia Van Arnum. Listen to the podcast at 

A multifaceted approach is needed to resolve the myriad of challenges in developing oral formulations of poorly soluble drugs.

Industry Perspectives: Achieving Solutions for the Challenge of Poorly Water-Soluble Drugs

Chirality plays an important role in pharmaceutical compounds, and methods for detecting and quantifying chirality remains challenging. Researchers at Harvard University and the Center for Free-Electron Laser Science (CFEL) and the Max Planck Institute in Germany recently reported on the use of enantiomer-specific detection of chiral molecules by microwave spectroscopy.  

The researchers pointed out that the spectroscopic methods of circular dichroism and vibrational circular dichroism are commonly used in analyzing chiral molecules. These methods, however, have limitations in electric dipole approximation as the resultant weak effects produce weak signals and  require high sample densities (1). In contrast, nonlinear techniques probing electric-dipole-allowed effects have been used for sensitive chiral analyses of liquid samples. Influenced by these methods, the researchers carried out nonlinear resonant phase-sensitive microwave spectroscopy of gas-phase samples in the presence of an adiabatically switched nonresonant orthogonal electric field. They used this technique to map the enantiomer-dependent sign of an electric dipole Rabi frequency onto the phase of emitted microwave radiation (1). They describe theoretically how this results in a sensitive and species-selective method for determining the chirality of cold gas-phase molecules. They implemented the approach experimentally to distinguish between the

 enantiomers of 1,2-propanediol and their racemic mixture. They reported that this technique produced a large and definitive signature of chirality and has the potential to determine the chirality of multiple species in a mixture. 

"We can soon measure mixtures of different compounds and determine the enantiomer ratios of each," said Melanie Schnell, co-author of the study in a CFEL release. In a next step, the researchers plan to apply the technique in a broadband spectrometer at CFEL that could then measure the enantiomer ratios in mixtures of substances. In the longer run, the method opens the door to develop a technique for separating enantiomers.  

 1. D. Patterson, Melanie Schnell, and John M. Doyle,

Microwave spectroscopy overcomes some of the limitations of circular dichroism and vibrational circular dichroism in analyzing enantiomers.

Microwave Spectroscopy for Enantiomeric Detection of Chiral Molecules

 to Boehringer Ingelheim for cGMP violations for the manufacture of APIs and finished pharmaceuticals at its facility in Ingelheim am Rhein, Germany. The letter, issued on May 6, 2013, was based on an inspection of the facility Nov. 5–12, 2012 and subsequent response by the company based on that inspection in which FDA cited insufficient corrective actions.

	Specifically, FDA said that the company’s quality unit failed to investigate critical deviations in the manufacturing of APIs by not thoroughly investigating the source of foreign particles in an API and by not implementing timely and appropriate corrective and preventive actions (CAPA). FDA said that lots contaminated with foreign particles were manufactured in 2008 and 2009, but that Boehringer Ingelheim did not begin a formal project to implement comprehensive corrections to mitigate the presence of foreign particles until July 2012. FDA also said contaminated lots of the API were used to produce capsules and that the company failed to conduct thorough complaint investigations regarding the presence of foreign particles found in its APIs.

	FDA has requested that the company provide a list of API batches produced after the implementation of the July 2012 CAPA and to indicate whether foreign particles have been observed. FDA also requested that the company inform the agency of appropriate corrective actions taken by its quality unit to prevent foreign-particle contamination in its APIs.

	On the finished product side, FDA said the company failed to reject drug products that did not meet established standards or specifications by not rejecting multiple batches of capsules that were contaminated with foreign particles. The letter said that during 2010 and 2012, several lots containing a given API were contaminated with extrinsic foreign particles that were found during weighing or by visual inspection and were used for finished-drug manufacturing. The size and weight of the particles ranged from approximately 200 microns to approximately 5 mm, from 0.4 micrograms to approximately 9 mg, respectively. The agency said that it disagreed with the company’s decision to use the API because the contaminated batches met the final specifications and noted that use of appropriate manufacturing, controls, quality standards, and systems for investigations of atypical events (e.g., contamination) are all essential parts of a robust quality system.

	Lastly, FDA cited the firm for failing to thoroughly investigate any unexplained discrepancy or failure of a batch or any of its components to meet specifications, whether or not the batch had already been distributed. FDA said that the company failed to determine the cause of out-of-specification (OOS) results for a lot of Spiriva HandiHaler, which failed the uniformity of a delivered-dose test specification for a nine-month stability test and 12-month stability interval. “It was only after the 12-month OOS result that your firm decided to initiate a product recall for this Spiriva lot,” said FDA in its letter. “We are concerned about the management decision to allow adulterated product to remain in the market between the 9- and 12-month stability stations.” FDA has requested that the company inform the agency of its action to prevent recurrence as well as provide a retrospective evaluation of all lots currently in the stability program and assess whether an OOS was obtained at any testing interval.

, Gerhard Koeller, head of corporate division quality at Boehringer Ingelheim, said: “Boehringer Ingelheim is taking this issue very seriously and is fully committed to working with the FDA on a comprehensive action plan that addresses the FDA’s concerns, ensuring that it implements corrective and preventive actions to optimize its quality systems.”

Company receives notice from FDA for not fully investigating foreign particles in APIs and finished products made from its facility in Ingelheim am Rhein, Germany.

FDA Issues Warning Letter to Boehringer Ingelheim for cGMP Violations

Contract Manufacturing Arrangements for Drugs: 

Agreements, which describes the agency’s current thinking on defining, establishing, and documenting the responsibilities of each party (or all parties) involved in contract cGMP manufacturing of drugs. 

In particular, the draft guidance describes how parties involved in the contract manufacturing of drugs can use quality agreements to delineate their responsibilities and ensure drug quality, safety, and efficacy. This guidance applies to the commercial manufacturing of APIs (drug substances or their intermediates), finished drug products, combination products, and biological drug products. For the purposes of this guidance, the term “manufacturing” includes processing, packing, holding, labeling operations, testing, and operations of the quality unit.

For purposes of the draft guidance, in discussing the roles and responsibilities of the parties in contractual relationships, the “owner” is defined as the party that introduces (or causes the introduction of) a drug into interstate commerce, whether such drug is covered by a marketing application/license or not. The “contracted facilities” are the outside entities performing manufacturing operations for the product owner. Some of the manufacturing operations performed by such contracted facilities include: formulation, fill–finish, chemical synthesis, cell culture and fermentation (including biological products), analytical testing and other laboratory services, and packaging and labeling. The draft guidance specifies that the owner is responsible for ensuring that drugs introduced for interstate commerce are neither adulterated nor misbranded as a result of the actions of their selected contracted facilities, and all contracted facilities must ensure compliance with applicable cGMP for all manufacturing, testing or other support operations performed to make a drug(s) for the owner. 

The draft guidance describes how contract manufacturing operations fit within the larger scheme of pharmaceutical quality systems and presents FDA’s current thinking on the roles and responsibilities of entities involved in contract manufacturing arrangements. FDA says the draft guidance incorporates provisions from ICH guidelines, including ICH Q7 

Good Manufacturing Practice Guidance for Active Pharmaceutical Ingredients,

The draft guidance notes that although written quality agreements are not explicitly required under existing cGMP, owners and contracted facilities can draw on quality management principles to carry out the complicated process of contract drug manufacturing by defining, establishing, and documenting the responsibilities of all parties involved in drug manufacturing, testing, or other support operations. Accordingly, FDA recommends that owners and contracted facilities implement written quality agreements as a tool to delineate responsibilities to ensure the quality, safety, and effectiveness of drug products.

Comments and suggestions regarding the draft guidance should be submitted within 60 days of publication in the 

 of the notice announcing the availability of the draft guidance. 

The draft guidance describes how quality agreements can be used to delineate the responsibilities of contract manufacturers involved in the cGMP manufacture of APIs and finished drug products.

FDA Issues Draft Guidance on Contract Manufacturing

 FDA’s quality-by-design (QbD) initiative is a systematic approach to designing and developing pharmaceutical formulations and manufacturing processes to ensure predefined product quality. Understanding the critical quality attributes of the ingredients in a formulation, including excipients, and the critical process parameters in the manufacturing process of the finished drug product is crucial for successfully implementing QbD. 

 on Thursday May 23 from 10:00 AM  to 11:00 AM EST to examine the regulatory and technical considerations in implementing QbD for coatings in modified-release formulations, including strategies to mitigate the risks associated with alcohol-induced dose-dumping and the application of alcohol-resistant coatings.

		The panelists for the webcast include Peter Timmins, PhD, Executive Director, Drug Product Science and Technology, Research And Development, Bristol-Myers Squibb, who will provide insight and case examples of implementing QbD for modified-release formulations. Om Anand, Senior Reviewer and Dissolution Focal Point in the Division of Bioequivalence II, Office of Generic Drugs, Center for Drug Evaluation and Research, US Food and Drug Administration and also the Office of Generic Drugs liaison to the United States Pharmacopeia (USP) and USP Expert Panel member, will offer a regulatory perspective on 

 performance testing of oral modified-release generic products, inclusive of consideration for alcohol-induced dose-dumping. And Rina Chokshi, Project Leader for Controlled Release Technology, FMC Biopolymer, will discuss how to design controlled-release formulations solutions that remain robust in the presence of alcohol and provide further insight about the critical material, formulation and performance attributes of alcohol-resistant coatings.

 which will be live on Thursday May 23 from 10:00 AM EST to 11:00 AM EST and available for on-demand viewing afterward, may be found

FDA’s quality-by-design (QbD) initiative is a systematic approach to designing and developing pharmaceutical formulations and manufacturing processes to ensure predefined product quality.