Amy Ritter was Scientific Editor, BioPharm International.

Amy Ritter, PhD

The recent announcement that GlaxoSmithKline will pay a record $3 billion in civil and criminal penalties marks the largest settlement ever incurred by a drug company, and underscores a trend in larger and larger penalties imposed by the Justice Department for off-label drug promotions and violations of the False Claims Act. Earlier this year, Abbott paid $1.6 billion to settle allegations related to a single drug, Depakote, and in 2009 Pfizer paid around $2.3 billion, including $1.3 billion in criminal penalties, for off-label promotion of Bextra. Along with the large monetary penalties, each of these companies signed a corporate integrity agreement, to assure that the companies address their behaviors and prevent them from recurring.

		These megasettlements are being levied against actions that occurred as much as a decade ago, and the question remains as to whether the era of block-buster settlements will fade together with the era of block-buster drugs. In a 

 issued in response to the announced settlement, GSK’s CEO, Sir Andrew Witty, says, “Today brings to resolution difficult, long-standing matters for GSK. Whilst these originate in a different era for the company, they cannot and will not be ignored. On behalf of GSK, I want to express our regret and reiterate that we have learnt from the mistakes that were made. “ He continues, “In the US, we have taken action at all levels in the company. We have fundamentally changed our procedures for compliance, marketing and selling. When necessary, we have removed employees who have engaged in misconduct. In the last two years, we have reformed the basis on which we pay our sales representatives and we have enhanced our ability to ‘claw back’ remuneration of our senior management.“

		Sir Witty’s tone is contrite, but time will tell whether GSK and the other members of Big Pharma who have been caught by the government have really put these behaviors behind them.

Articles

The recent announcement that GlaxoSmithKline will pay a record $3 billion in civil and criminal penalties marks the largest settlement ever incurred by a drug company,

A New Era For Corporate Integrity?

The first steps in the production of any protein are transfecting a cell line, achieving expression of functional protein, and selecting a high-expessing clone for production. Charles H. Squires, PhD, vice-president of discovery and external partnerships at Pfenex Inc., spoke with 

 about advances in protein expression technology, and discussed the challenges yet to be overcome. 

Charles H. Squires, PhD, vice-president of discovery and external partnerships at Pfenex. (JEREN (FRANCE)/FUSION/ CASPAR BENSON/GETTY IMAGES)			

 Can you describe how the process of protein expression has improved or changed over the past 10 years or so? What are one or two key technological advances that have made these changes possible? 

New methods to more rapidly and reliably express proteins have proliferated over the past decade and a half. Innovations in this area have included the improvement of expression hosts to introduce more phenotypic capability to produce active, undegraded protein. For instance, hosts have been modified to overexpress folding modulators or mutated to eliminate proteolytic activities. Also, there has been a large increase in the number of these tools available commercially, putting many effective approaches within the reach of researchers with limited resources. The past several years have also seen the emergence of proprietary platform technologies, each of which offers some advantages in expressing difficult proteins or which delivers an advantageous downstream process. Examples include platforms such as a strain of 

that secretes product to the extracellular space (Wacker Chemie AG), a toolbox of regulated promoters in the yeast 

 (VTU Technologies), and a completely unique 

-based expression platform (Pfenex). The Pfenex platform has proven highly effective in expressing a variety of types of proteins and employs a high-throughput parallel processing technology in which a thousand or more unique expression strains are constructed and analyzed in about a month.  

Similar types of improvements have been made in the screening of mammalian expression hosts for the extent of target protein expression, including fluorescence-activated cell sorting for the selection of productive clones. These technologies are particularly effective in the identification of cell lines expressing high levels of engineered monoclonal antibodies.  

In addition to these types of improvements to host and expression strategy, advances in gene design that go beyond the traditional adherence to codon usage tables have been implemented, such as the technology developed by DNA 2.0. Expression data-taught, host-specific algorithms that modulate DNA sequences while keeping protein amino acid sequence constant may add yet another dimension to the list of effective protein production tools. The community of researchers involved in all aspects of protein production is certainly much better off than it was 20 years ago in its ability to provide solutions to protein expression, but it will take much more work understanding all aspects of the nature of protein expression to arrive at universally applicable principles. Those solutions will necessarily involve an ever widening toolbox of approaches that will need to be accessed through more advanced parallel processing tools. The principle difference now is that a multidimensional approach to gene and protein expression is available to the great majority of researchers, whereas 20 years ago the process was one dimensional and digital, giving only a yes or no result.  

 What parts of the protein-expression process are still rate-limiting, and how do you see these challenges being addressed? What improvements do you expect to see in the future? 

We anticipate that given the continued need for protein supply across the spectrum of applications, many of the existing weak links in protein expression will be strengthened. One of the most pressing unmet needs is production of small amounts of protein for research purposes. Often, dozens of proteins ranging from antigens for preclinical testing to related variants of a target protein for some form of differential testing are required. While only a small amount of protein, a few mg, is generally required for activity evaluation or crystallization for these types of studies, currently all of the cloning steps needed to get even that small amount of protein are identical to those required to assemble a strain or cell line for large-scale biopharmaceutical production. This is highly inefficient and a consistent bottleneck in the process. Clearly, there is a need to develop off-the-shelf solutions to fill this gap. New, precise, and more adaptable tools allowing more rapid cloning and characterization will need to be developed. These tools will necessarily include the development and adoption of more effective high-throughput processing methods. Improvements in genetic, analytical, and processing methods practiced in high throughput, parallel modes will enable the researcher to access more tools and reach a wider range of solutions more rapidly.  

Clearly, for these approaches to have an impact, methods to allow the researcher to identify productive clones at the earliest possible stage must be developed. Better reporter systems that may enable screening, even at the colony or single cell level, would be very valuable. Processing to obtain small amounts of protein would then benefit from the development of more efficient purification tags. These may also be envisioned to include reliable expression reporter sequences, sequences that promote proper protein folding, and ideally be efficiently and universally removable. Pfenex is exploring this segment, adapting its expression platform strategy specifically for custom reagents. Development in this area would be of great benefit not only to the core of researchers producing proteins for research purposes but the benefits would be amply felt in the area of target protein expression as well.  

Another route to solve the problem of small-scale supply would be to adopt a cell-free approach such as that employed by Sutro Biopharma. Such an approach is enabled today by the relatively rapid, commercial availability of synthetic, gene-length DNA sequences. Developing an off-the-shelf cell-free system would jump the cloning and cell growth steps in the current processes and potentially deliver advantages of protein quality as well. An 

system would also be able to take advantage of the reporter and purification technologies mentioned above for 

systems. As more and more dimensions to protein production are added, the researchers' ability to deliver will continue to improve. The challenge will continue to evolve, however, as more and more types of hybrid or completely synthetic proteins are developed that may or may not have some evolutionarily defined path to proper folding and thus activity. 

Improvements in expression platforms and enhanced tools for selecting clones are among the advances of the past few decades.

35th Anniversary Special: Advances in Protein Expression

Nanotechnology has been used to develop innovative formulations to improve bioavailability of drugs,  and have found utility as carriers for either highly toxic payloads, such as chemotherapeutics, or very fragile payloads, such as proteins or nucleic acids. Panayiotis P. Constantinides, PhD, founder and principal of Biopharmaceutical and Drug Delivery Consulting, spoke to 

 about the utility of nanotechnology in formulation strategies, both past and future. 

				 Can you identify one or two key technical advances that have enabled the successful development of nanoparticles as drug delivery vehicles?  

 Within the scope of this discussion, both 1–100 nm and submicron particles (100–1000 nm) are considered as nanoparticles for both oral and non-oral drug delivery applications. The successful development of drug delivery vehicles through advances in pharmaceutical nanotechnology is certainly reflected in the marketed nanoparticulate drug products which are in the submicron range (100–1000 nm). These products include Doxil (liposomal doxorubicin, ALZA/1999), Abraxane (albumin-bound paclitaxel, Abraxis/2005) and NanoCrystal based drug products, Rapamune (Sirolimus, Wyeth/2000), Emend (aprepitant, Merck/2003) TriCor (fenofibrate, Abbott/2004), Megace ES (megestrol, Par/2004) and Invega (paliperidone palmitate, Ortho-McNeil-Janssen/2009). 

 Panayiotis P. Constantinides, PhD, founder and principal of Biopharmaceutical and Drug Delivery Consulting, ppconstantinides@bpddc.com.			

Key technical advances which have enabled the successful development and commercialization of the aforementioned nanoparticulate drug products include advances in biology, materials science and particle engineering, processing, and manufacturing. Top-down and bottom-up manufacturing methods at small and large scale have been applied and tailored to the specific compound and its application. During the top-down process, high shearing homogenization or media-milling is used to reduce particle size in the presence of polymeric and surfactant stabilizers to prevent crystallization and particle growth. In the bottom-up process of manufacturing nanoparticles, the nanoparticle is created from its individual components, using precipitation, coacervation, polymerization, and hydrophobic aggregation in an effort to achieve small size by controlling the thermodynamics and kinetics of nucleation, growth, and chemical reactivity. Layer-by-layer self-assemblies using biopolymers and synthetic polymers create versatile nanostructures for drug and gene delivery. Nanoformulation is becoming an integral part of life-cycle management strategies for product line extensions and as enabling drug discovery technology for new molecular entities.   

Top-down and bottom-up methods to manufacture nanoparticles have been applied to commercial products, the former for the marketed parenteral liposomal Doxil and the NanoCrystal drug products, and the latter for the albumin-bound nanoparticle Abraxane. For these drug products, advances in the characterization of nanoparticles and better understanding of how the ADME properties of compounds administered as nanoparticles are linked to particle size and surface chemistry have also contributed to their successful development and commercialization. In the case of the parenteral cancer chemotherapeutic drug products Doxil and Abraxane, key factors that have also contributed to their successful design and development were advances in nanoparticle targeting strategies based on the fundamental understanding of tumor biology principles, such as tumor vasculature and permeability, pH, and surface antigens.   

 What advances to you expect to see in the future that will improve selectivity or efficacy? 

 Nanoparticle selectivity and efficacy is particularly important in parenteral drug delivery and targeting for cancer and other diseases. Linked to nanoparticle selectivity and efficacy is their potential toxicity. Further advances in cell and tissue imaging techniques and other 

characterization methods of nanomaterials and nanoparticles are necessary to better understand and predict toxicity. To this end, further development and clinical application of multifunctional nanoparticles which combine disease diagnosis with therapy (nanotheranostics) could prove to be very useful. These systems consist of a core which can be magnetic for MRI or quantum dot for optical imaging, a lipidic and/or polymeric shell for intracellular targeting and an outer layer incorporating a surfactant or polymeric stabilizer and a targeting moiety (e.g., antibody, aptamer, or ligand). The drug, small molecule or biologic, can be encapsulated in the core and/or the shell of the particle. 

Advances in image-guided drug delivery will offer several advantageous features and enable scientists and clinicians to monitor and quantify drug release, noninvasively assess drug site accumulation, visualize biodistribution in real time, analyze drug distribution at the target site, predict drug response and evaluate drug efficacy and toxicity. Ultimately and perhaps not in the distant future, these features could be incorporated into microchips and personalized medicines.   

Most nanoparticles accumulate in the liver but depending on their size and charge can also accumulate in the kidney and other tissues. All classes of nanomaterials have extensive tissue retention, particularly carbon nanotubes and quantum dots, and this has toxicological implications. The state of nanomaterials once deposited in the tissue is largely unknown. Furthermore, comparison between and across complex nanoparticles is difficult.        

In reference to nanoparticle toxicity, limited number of nanomaterials have been evaluated to date, and mechanisms for nanomaterial toxicity are actively being pursued. Specific properties of nanometerials, particularly their surface characteristics, contribute to their toxicity. There is a need for nanotoxicity guidelines, and no specific regulations are available at the present time. However, assessing nanomaterial risks should be proactively pursued to identify and analyze potential hazards, assess potential exposure scenarios and evaluate toxicity and analyze and communicate potential risks and uncertainties.  

From a formulation and process development perspective, we need to critically assess which lessons learned from the marketed nanoparticulate drug products can be applied to 1-100 nm particles and what is truly new science and knowledge. For example, new processing and characterization methods are needed along with the establishment of meaningful controls, specifications, and standards for 1–100 nm particles. Further advances in manufacturing methods are required in order to scale up and reproducibly produce complex multifunctional nanoparticles with an acceptable shelf-life.   

Drivers for future growth and commercialization of drug delivery nanoparticles include availability of funding, patent landscape, increased understanding of the pathophysiology of disease, and the need for novel drugs and safe therapies.  

A look back at key nanoformulation advances and what lies ahead for nanoparticle-based drug-delivery systems.

35th Anniversary Special: Nanoformulations

In a 5–4 ruling, the US Supreme Court upheld the provision of the Affordable Care Act requiring all adults to purchase health insurance or pay a penalty—the so-called individual mandate. The court rejected the government’s argument that because health care was a market in which everyone will eventually participate, the government could regulate individual coverage under the Commerce Clause. Instead, the court found the mandate to be constitutional when viewed as a tax. The full text of the court’s decision can be found through CNN’s 

. Chief Justice John Roberts, writing for the majority, sums up the decision as follows:

“The individual mandate cannot be upheld as an exercise of Congress’s power under the Commerce Clause. That Clause authorizes Congress to regulate interstate commerce, not to order individuals to engage in it.  In this case, however, it is reasonable to construe what Congress has done as increasing taxes on those who have a certain amount of income, but choose to go without health insurance. Such legislation is within Congress’s power to tax.”

The court found a second contested provision that required states to expand existing coverage for Medicaid or lose federal funding for the program to be unconstitutional. Chief Justice Roberts writes:

“As for the Medicaid expansion, that portion of the Affordable Care Act violates the Constitution by threatening existing Medicaid funding. Congress has no authority to order the States to regulate according to its instructions. Congress may offer the States grants and require the States to comply with accompanying conditions, but the States must have a genuine choice whether to accept the offer.”

The Biologics Price Competition and Innovation Act

 which provides an abbreviated approval pathway for biosimilars, was enacted as a provision of the Affordable Care Act. There was some concern that if the individual mandate had been found to be unconstitutional, the entire piece of legislation would have been struck down, a fear that is now allayed. The Biotechnology Industry Organization says in their 

, “We also will continue our work with US Food and Drug Administration (FDA) to implement the bipartisan-backed biosimilars pathway that was enacted under the law. Specifically, we will advocate for implementation approaches that ensure patient safety, expand patient access and competition, and provide necessary and fair incentives that will help spur continued biomedical breakthroughs.“ 

In a 5-4 ruling, the US Supreme Court upheld the provision of the Affordable Care Act requiring all adults to purchase health insurance or pay a penaltyâ€”the so-called individual mandate.

US Supreme Court Upholds Individual Mandate for Health Insurance

 on June 26, 2012, that it will be closing its R&D site in Nutley, New Jersey, eliminating approximately 1000 positions. The action is being taken to streamline research activities and to reallocate resources to support programs in early clinical development. According to the company press release, the reorganization will enable Roche to keep R&D costs stable despite a strong increase in the number of clinical development projects in the last 18 months. The company plans to complete the transfer of business operations in Nutley by the end of 2013. Roche’s Genentech division, based in South San Francisco, California, will not be affected by the reorganization.

The site closure will be partially offset by the establishment of a Pharmaceuticals Translational Clinical Research Center on the east coast of the US, which is expected to employ approximately 240 people. The center will support US-based clinical trials and early development programs, support and maintain Roche interactions with FDA, and enhance Roche’s collaborations with US-based partners, such as academic institutions and biotech companies. R&D activities at the Nutley site will be transferred to sites in Switzerland and Germany, which are expected to gain 80 positions.

Roche announced that it will be closing its R&D site in Nutley, New Jersey, eliminating approximately 1000 positions.

Roche to Close US R&D Site

 that it will be closing its venerable Nutlley, NJ R&D site is the latest in a string of mergers and reorganizations that have resulted in the loss of thousands of life-sciences jobs from New Jersey, and an exodus of both R&D and manufacturing from a state known as the home of big Pharma. The mergers of Wyeth with Pfizer and Schering Plough with Merck were among the highlights of a decade that has seen a dramatic contraction in the pharmaceutical industry as a whole. According to 

, 17 of the top 20 pharma companies have a presence in New Jersey, and because the presence was so large to begin with, the effects of the contraction on the New Jersey economy have been acutely felt.

		Ironically, New Jersey’s governor Chris Christie was awarded the 2012 Governor of the Year award by BIO, the biotech industry trade group, because of his biotech-friendly tax policies. While BIO, in their 

 calls New Jersey a biotech cluster, the reality is that the 300 or so biotech companies in New Jersey are spread throughout the state (around 8000 square miles), without the dense concentration found in Boston (around 240 companies within 750 square miles, according to 

) or San Diego. And biotechs tend to be small, employing tens or hundreds of people, so cannot begin to replace the thousands of high-paying jobs that were lost from big Pharma. That the BIO Governor of the Year was awarded in a state where the joke is that the cashier at the grocery story might be a PhD chemist is bound to make a few people cringe.

Roche’s recent announcement that it will be closing its venerable Nutlley, NJ R&D site is the latest in a string of mergers and reorganizations that have resulted in the loss of thousands of life-sciences jobs from New Jersey,