33. Newly identified small-RNA

Newly identified small-RNA pathway defends genome against the enemy within

‘Jumping genes’ or transposable elements are long stretches of repetitive DNA that can insert themselves throughout the genome. Normally, they are held inactive by a series of marks along the genome. But when these marks are erased, activated transposons can disrupt critical genes, causing dramatic defects in development. Here are two flowers: on the left is a normal plant where transposons are silent; on the right, transposons have been activated, severely mutating the flower. Researchers at CSHL have discovered a new fail-safe mechanism that prevents transposon activation even when the inactivating marks have been erased.

For a plant to create reproductive cells, it must first erase a key code, a series of tags attached to DNA across the genome known as epigenetic marks. These marks distinguish active and inactive genes. But the marks serve another critical role. They keep a host of damaging transposons, or “jumping genes,” inactive. As the cell wipes away the epigenetic code, it activates transposons, placing the newly formed reproductive cell in great danger of sustaining genetic damage. 

Today, researchers at Cold Spring Harbor Laboratory (CSHL) led by Professor and HHMI Investigator Robert Martienssen announce the discovery of a pathway that helps to keep transposons inactive even when the epigenetic code is erased. 

“Jumping genes” were first identified more than 50 years ago at CSHL by Nobel Prize-winning researcher Barbara McClintock. Subsequent study revealed that jumping genes (or transposable elements) are long, repetitive stretches of DNA. They resemble remnants of ancient viruses that have inserted themselves into their host DNA. When active, transposons copy themselves and jump around in the genome. They can insert themselves right in the middle of genes, thus interrupting them. Scientists have found that more than 50% of the human genome is made up of transposons. Remarkably, in plants, up to 90% of the genome is composed of these repetitive sequences. 

When a transposon is activated, it can insert itself within critical genes, disrupting gene function and causing infertility and many diseases. To combat this ever present threat from within, the cell has devised stringent mechanisms to maintain tight control over transposon activity. The primary mechanism is the epigenetic code, a kind of secondary layer of genetic information that determines how our DNA is used. Epigenetic marks decorate human DNA, delineating active and inactive genes. Regions of the genome that are rich in transposons are heavily marked with inactivating signals, which silence transposons. 

The problem for plants, in particular, is that some cells eliminate nearly all epigenetic marks during reproduction. “The loss of these marks puts the cell in tremendous danger, especially at critical times like reproduction,” says CSHL postdoctoral fellow Kate Creasey, Ph.D., lead author on the paper appearing in Nature today. “There must be another mechanism to prevent this kind of widespread genomic disruption.”

The CSHL team discovered a pathway that does precisely this. The pathway they describe acts as a fail-safe to prevent transposon damage when epigenetic silencing is lost. The cell uses small RNAs, known as microRNAs, to accomplish the task. MicroRNAs were already known to regulate gene expression during development. “Now we show that microRNAs actually target transposons when they are activated, for example in the germline (or reproductive cells),” says Martienssen. “This hints that they may have evolved as a transposon defense mechanism.”

Working in collaboration with Professor Blake Meyers at the University of Delaware, Martienssen and his colleagues found that microRNAs silence transposons through a new class of small RNAs known as easiRNAs. Animals have a similar transposon defense mechanism, says Martienssen. “The pathway we have discovered parallels with small RNA systems in animal germlines (called piwi-interacting RNAs, or piRNAs) which also protect against transposons when the genome is being reprogrammed.”

This work was supported as part of a collaboration with DuPont Pioneer as well as grants from the National Institutes of Health, the Howard Hughes Medical Institute and Gordon and Betty Moore Foundation, and the Cold Spring Harbor Laboratory Shared Resources by the Cancer Center Support Grant. Additional support was provided by fellowships from the Belgian American Educational Foundation and University of Delaware. 

“miRNAs trigger widespread epigenetically activated siRNAs from transposons in Arabidopsis” appears online in Nature on March 16, 2014. The authors are: Kate M. Creasey, Jixian Zhai, Filipe Borges, Frederic Van Ex, Michael Regulski, Blake C. Meyers & Robert A. Martienssen. The paper can be obtained online at: http://dx.doi.org/10.1038/nature13069

About Cold Spring Harbor Laboratory 

Founded in 1890, Cold Spring Harbor Laboratory (CSHL) has shaped contemporary biomedical research and education with programs in cancer, neuroscience, plant biology and quantitative biology. CSHL is ranked number one in the world by Thomson Reuters for the impact of its research in molecular biology and genetics. The Laboratory has been home to eight Nobel Prize winners. Today, CSHL's multidisciplinary scientific community is more than 600 researchers and technicians strong and its Meetings & Courses program hosts more than 12,000 scientists from around the world each year to its Long Island campus and its China center. For more information, visit www.cshl.edu.

32. Bacterial reporters

Bacterial reporters that get the scoop

Inspired by Nature, the team engineered E. coli to sense, record and remember an environmental signal in the gut — and also demonstrated that they can survive and function within the complex environment of the mammalian gut. This work lays the foundation for the use of engineered probiotic bacteria that serve as non-destructive living diagnostics. In this schematic engineered probiotic E. coli have colonized the mammalian intestine and "remember" exposure to an environmental signal, which is indicated by the cells turning blue in color.

It's a jungle in there. In the tightly woven ecosystem of the human gut, trillions of bacteria compete with each other on a daily basis while they sense and react to signals from the immune system, ingested food, and other bacteria.

Problems arise when bad gut bugs overtake friendly ones, or when the immune system is thrown off balance, as in Crohn's disease, celiac disease, and colorectal cancer. Doctors have struggled to diagnose these conditions early and accurately. But now a new engineered strain of E. coli bacteria could deliver status updates from this complex landscape to help keep gastrointestinal diseases at bay.

The new strain non-destructively detected and recorded an environmental signal in the mouse gut, and remembered what it "saw." The advance, reported in the Proceedings of the National Academy of Sciences, could lead to a radically new screening tool for human gut health.

The key to turning E. coli into gut reporters was to insert a well-known genetic switch that flips when it senses a specific environmental cue. This switch confers on the cells the ability to "remember" what they sense for up to a week — long enough for scientists to recover fecal samples and test whether the switch has flipped.

"This achievement paves the way toward living monitors programmed using synthetic gene circuits," said Wyss Institute Core Faculty member Pamela Silver, Ph.D., senior author on the study who is also the Elliott T. and Onie H. Adams Professor of Biochemistry and Systems Biology at Harvard Medical School (HMS). Silver's team included James Collins, Ph.D., who is also a Wyss Core Faculty member and professor of bioengineering at Boston University, as well as other collaborators from the Wyss Institute, Harvard Medical School and Boston University. "It could lead to new diagnostics for all sorts of complex environments."

The approach Silver's team took runs counter to the prevailing dogma in synthetic biology, which is to design genetic systems that drive cell behavior from scratch, said Wyss Institute Senior Staff Scientist Jeff Way, Ph.D., a coauthor on the paper. On the other hand, "Nature has a tried-and-true blueprint for memory systems if you know where to look," Way said. "Why not just accept Nature as it is, and develop the system from there?"

The genetic switch the team inserted in E. coli came from lambda phage, a virus that commonly attacks this bacterium.

After invading E. coli, lambda typically lays low, living in a stealth mode called lysogeny in which its DNA simply hangs out in the E. coli's genome. But when the bacterium's DNA is damaged — and only then — the switch flips, instructing the virus to enter a mode called lysis in which it multiplies inside the cell and breaks through its membrane in a kind of microbial explosion.

"This is a very stable system in Nature," said lead author Jonathan Kotula, Ph.D., a Postdoctoral Fellow at HMS who is also affiliated with the Wyss Institute. "We knew the lambda switch would be a great candidate for the memory element, and we simply tweaked it to meet our needs."

The cells with the engineered lambda switch would not become lytic under any conditions. Kotula and the rest of Silver's team used standard molecular genetic tools to rig the switch such that it turned on only in the presence of an inactive form of the antibiotic tetracycline.

In laboratory experiments, the switch turned on within a few hours of exposure to the antibiotic — and stayed in this 'ON' state inside E. coli for a week or more, even as the bacteria grew and divided. In short, the cells "remembered" that they had seen that molecule in the gut.

"It was truly shocking how cleanly the experiments worked," said Jordan Kerns, Ph.D., a Wyss Institute Postdoctoral Fellow.

But to function as a living diagnostic, the engineered E. coli also had to survive their trip through the gut intact, which meant they had to compete effectively against rival gut microbes.

The engineered strain worked fine in laboratory experiments, but gradually disappeared when the team introduced it into the gut of the mouse itself. It turned out that it had been outcompeted by the animal's native gut bacteria. The team did not fret in the face of this result because they knew that the classical strain of E. coli they used had lived only in the laboratory since the 1940s — losing its ability to compete in the real world, particularly in an environment as challenging as the mammalian gut.

They tackled the problem by isolating a native strain of E. coli from the mouse gut, then engineering its genome to incorporate the switch. The switch in the cells flipped within hours, as it had before, and the cells "remembered" for about a week that they had seen the antibiotic in the gut, Kotula said. Moreover, the population stabilized within the gut, holding its own in the presence of other bacteria.

The team envisions a day when a doctor would give a patient a strain of engineered bacteria as a diagnostic, much as they give a probiotic today. The strain would be rigged to monitor the gut for any number of conditions from inflammation to disease markers. At a follow-up visit, the patient would submit a stool sample, and medical technicians would collect E. coli from the sample and analyze it. Only if the switch (or switches) were on would the doctor perform more invasive tests such as an endoscopy or a colonoscopy.

For now the team is focusing on genetically tweaking the memory element of their system so that the cells remember for even longer periods of time, and engineering it so the switch flips when it senses other chemical signatures as well, such as those of cancer or parasites. In the longer term, their engineered bacteria could sense a disease state and work with other engineered genetic circuits that can produce a specific drug on command, thus producing a dynamic therapy.

"Our increasing appreciation of the role of the microbiome in health and disease is transforming the entire medical field. The concept of using the power of synthetic biology to harness microbes that live in our gut to develop living diagnostic and therapeutic devices is a harbinger of things to come, and Pam's work provides the first proof-of-principle that this is a viable and exciting path to pursue," said Wyss Institute Founding Director Don Ingber, Ph.D., M.D.

This work was funded by the Defense Advanced Research Projects Agency (DARPA) and the Wyss Institute.

PRESS CONTACT

Wyss Institute for Biologically Inspired Engineering at Harvard University
Kristen Kusek
kristen.kusek@wyss.harvard.edu
+1 617-432-8266

31. Chicken bones tell true story

Chicken bones tell true story of Pacific migration

Did the Polynesians beat Columbus to South America? Not according to the tale of migration uncovered by analysis of ancient DNA from chicken bones recovered in archaeological digs across the Pacific.

The ancient DNA has been used to study the origins and dispersal of ancestral Polynesian chickens, reconstructing the early migrations of people and the animals they carried with them.

The study, led by the University of Adelaide's Australian Centre for Ancient DNA (ACAD) and published today in Proceedings of the National Academy of Sciences USA, reveals that previous claims of contact between early Polynesians and South America were probably based on contaminated results. Instead, the new study has identified and traced a unique genetic marker of the original Polynesian chickens that is only present in the Pacific and Island Southeast Asia.

The research team of national and international collaborators, including Australian National University, University of Sydney, and Durham and Aberdeen Universities in the UK, used female-inherited mitochondrial DNA extracted from chicken bones excavated in archaeological digs from islands including Hawaii, Rapa Nui (Easter Island) and Niue.

30. half animal, half plant

Sea anemone is genetically half animal, half plant

Sea anemone shows a genomic landscape surprisingly similar to human genome, but also displays regulatory mechanisms similar to plants

The team led by evolutionary and developmental biologist Ulrich Technau at the University of Vienna discovered that sea anemones display a genomic landscape with a complexity of regulatory elements similar to that of fruit flies or other animal model systems. This suggests, that this principle of gene regulation is already 600 million years old and dates back to the common ancestor of human, fly and sea anemone. On the other hand, sea anemones are more similar to plants rather to vertebrates or insects in their regulation of gene expression by short regulatory RNAs called microRNAs. These surprising evolutionary findings are published in two articles in the journal "Genome Research".
Our appearance, the shape we have and how our body works is, in addition to environmental influences, largely the result of the action of our genes. However, genes are rarely single players, they rather act in concert and regulate each other's activity and expression in gene regulatory networks.

Simple organism with complex gene contentIn the last decades the sequencing of the human and many animal genomes showed that anatomically simple organisms such as sea anemones depict a surprisingly complex gene repertoire like higher model organisms. This implies, that the difference in morphological complexity cannot be easily explained by the presence or absence of individual genes. Some researchers hypothesized that not the individual genes code for more complex body plans, but how they are wired and linked between each other. Accordingly, researchers expected that these gene networks are less complex in simple organisms than in human or "higher" animals.

A measurement of the complexity of gene regulation could be the distribution and density of regulatory sequences in the genome. These motifs on the DNA called enhancers and promoters can bind transcription factors specifically and often regulate the expression of target genes in specific spatio-temporal patterns. "Finding these short motifs in the ocean of nucleotides is far from trivial", explains Ulrich Technau, professor at the Department for Molecular Evolution and Development.

While the genes constitute, in a sense, the words in the language of genetics, enhancer and promoters serve as the grammar. These regulatory elements correlate with certain biochemical epigenetic modifications of the histones, proteins intertwined with the DNA, constituting the chromatin. With the aid of a sophisticated molecular approach called chromatin immunoprecipitation, Hertha-Firnberg-fellow Michaela Schwaiger, member of Technau's team, was able to identify promoters and enhancers on a genome-wide level in the sea anemone and compared the data to regulatory landscapes of more complex and higher model organisms.

Gene regulation comparable to higher animal model systems"Since the sea anemone shows a complex landscape of gene regulatory elements similar to the fruit fly or other model animals, we believe that this principle of complex gene regulation was already present  in the common ancestor of human, fly and sea anemone some 600 million years ago" , Michaela Schwaiger states.

MicroRNAs are important for developmental processes in human…Eventually, gene expression leads to the formation of proteins, the functional effectors in our body. In addition to the control of transcription of DNA to RNA, the expression of a gene can also be regulated on the post-transcriptional level after the RNA is already produced. Here, microRNAs play an important role. MicroRNAs are short regulatory RNAs, which can bind to target RNAs and inhibit their translation or lead to dissociation of the target RNA. In the last years, hundreds of microRNAs were identified in many animals and even more than 1000 microRNAs in human. Many of these have an important role in metabolism and are crucial in developmental processes. Mutations in distinct microRNAs are associated with severe diseases such as cancer. Each microRNA can bind many different RNAs in a sequence specific manner. "We assume that 30 – 50 percent of all human genes are regulated by microRNAs", Ulrich Technau illustrates. However, the evolutionary origin of animal microRNAs is still unclear.

…and in plantsMicroRNAs were also discovered in plants, but it has been assumed that they arose independently from animal microRNAs, since they (1) don't show any sequence similarity to them, (2) have a different biogenesis pathway and (3) have a substantially different mode of action: Plant microRNAs bind only one to a handful of targets with high sequence specificity and induce with the aid of Argonaute proteins the specific cleavage of the target RNA. In collaboration with American, French and Norwegian groups, Ulrich Technau and his team managed to isolate 87 microRNAs from the sea anemone.

Yehu Moran, David Fredman and Daniela Praher from the Technau team were able to show that the microRNAs of the sea anemone depict all the hallmarks of plant microRNAs: They have an almost perfect complementarity to their target RNAs, which are subsequently cleaved and not inhibited like in other animals. Moran also discovered a gene in the sea anemone, HYL-1, which is essential for the microRNA biogenesis in plants and was never detected in any other animal model organism before. Moreover, when one compares the sequences of microRNAs, one microRNA with similarity to a plant microRNA as well as one microRNA with similarity to an animal microRNA can be found. Altogether, these findings suggest the first evolutionary link between microRNAs of plants and animals.

In summary, while the sea anemone's genome, gene repertoire and gene regulation on the DNA level is surprisingly similar to vertebrates, its post-transcriptional regulation is plant-like and probably dates back to the common ancestor of animals and plants. This is the first qualitative difference found between Cnidaria and "higher" animals and the findings provide insight on how important levels of gene regulation can evolve independently. 

29. New Way to Interrogate PTPs

A New Way to Interrogate PTPs

A review of literature describing an assay that could help researchers use protein tyrosine phosphatases in drug development.

Protein tyrosine phosphatases (PTPs) control posttranslational tyrosine phosphorylation, and thus play a critical role in the regulation of many signaling molecules. Despite the disease relevance of PTPs (for example, PTP1B is considered a target for treatment of diabetes type 2; DEP1 and PTPγ are cancer targets) and several PTP inhibitors entering clinical trials (Hardy et al., Curr Oncol 2008;15:5–8), this class of enzymes has been known as one of the “undruggable” targets. This is largely due to the promiscuity of the targets and challenges associated with obtaining potent and specific PTP inhibitors. A need for PTP assays, preferentially with the ability to interrogate both substrate specificity and kinetic parameters in real time, is still highly desirable. Traditional phosphatase assays utilize artificial chromogenic or fluorogenic substrates, which may skew data interpretation. The Malachite green assay, on the other hand, quantifies inorganic phosphate production in an end-point mode, limiting itself for kinetic time course application.

In this article highlighted herein, the authors* present a PTP assay that utilizes a physiologically relevant peptidic substrate that contains 3-nitrophosphotyrosine moiety. The assay is based on the observation that 3-nitrophosphotyrosine is widely accepted by different PTPs, and upon enzyme turnover, the resulting 3-nitrotyrosine can be monitored spectrophotometrically at 415 nm.

After assay validation using two known phosphatase inhibitors, vanadate and NSC87877 (see Figure 2, panel C), the authors further demonstrated several distinct features of the assay. PTP activity profiling and substrate specificity could be conveniently achieved using the assay in plate format (see for example Figure 2, panel B). For instance, between two model cancer targets, DEP1 and PTPγ, their substrate specificity was differentiated among eight different peptidic substrates.

Another advantage of the assay lies in its insensitivity to the presence of background phosphate, such as elements from assay buffer or cell lysates. The same assay was subsequently applied to HEK293 cell lysates that had been treated under four different conditions, including wild type, transfected with PTP1B or inactive PTP1B, nontransfected but with 250 μM vanadate. PTP activity was readily discriminated as expected based on their respective initial reaction velocities, with cell lysates transfected with PTP1B showing the highest activity, while non-transfected lysates treated with vanadate showed the lowest

Figure 2. Sample data for the spectrophotometric PTP assay. All data points are averages of three experiments, errors expressed as SE. (A) Absorption spectra at different concentrations of 3-nitrotyrosine showing the maximum at 415 nm. (B) Initial velocities (v_ini) at different substrate concentrations for PTP1B with a 3-nitrophosphotyrosine peptide substrate derived from SIGLEC2. (C) Inhibition of PTPγ by different concentrations of sodium ortho-vanadate using a 3-nitrophosphotyrosine peptide substrate derived from ZAP70. (D) Initial velocities (v_ini) for HEK293 lysates (WT, nontransfected; PTP1B, transfected with PTP1B; C215S, transfected with inactive PTP1B; VO4, nontransfected in the presence of 250 µM vanadate) using a 3-nitrophosphotyrosine peptide substrate derived from INSR.

Overall, the method reported by van Ameijde et al. represents a welcomed addition to the existing toolbox and drug development programs for protein phosphatases. Potential improvements could include further enhancement in assay sensitivity (such as using fluorescence as a readout and reduce enzyme/substrate consumption).

28. Cancer Stem Cells

Cancer Stem Cells: Making Sense of the Data

The CSC research space is proving to be challenging and fraught with controversy

There is much controversy in the CSC field as to their surface phenotype, intercellular pathways, and role in metastatic progression.

The focus of this GEN Market & Tech Analysis report is to frame the challenging space of cancer stem cells (CSCs). Highlights of this report:

The potential relationship between circulating tumor cells (CTCs) and CSCs is unclear at present—this is important since it underlines the mechanism[s] by which components of the primary tumor migrate to distal sites and induce mets, the drivers for morbidity and mortality in late-stage cancer patients.We have performed market surveys of researchers studying CSCs as a means to understand the surface markers deployed (to characterize CSCs) as well as the intracellular signal transduction pathways being studied in the context of CSCs.Also presented in this report are the various associations of markers with distinct cancer classes documented thusfar—it appears that distinct classes of CSCs with different surface phenotypes may be associated with different classes of cancer and would suggest that the CSC pool is distinct in different cancer classes ("one size does not fit all").This space is an area of continuing industry coverage for us, and in future reports we'll document advances in the CSC space as they occur as well as an update on the clinical trials underway in this field.

27. Top 10 Job-Cutting Companies

Top 10 Job-Cutting Companies of 2013

• #10. Endo Health Solutions

  Number of job layoffs disclosed: About 700 jobs

  Details: Company said June 6 it will eliminate 15% of its workforce worldwide. Company has not stated where jobs will be cut, but suggested at selling, general, and administrative (SG&A) and research job reductions by tying the layoffs to a restructuring whose goals include “streamlining general and administrative expenses, optimizing commercial spend and refocusing R&D onto lower-risk projects and higher-return investments in generics.”

  Why the company did it: Part of a restructuring that included seeking more acquisitions, selling its HealthTronics urological business¹ and branded pharmaceutical early-stage discovery platform, and restructuring its R&D effort. Restructuring was unveiled after company failed to stop a generic version of its Opana ER (oxymorphone HCl) moderate-to-severe pain drug from reaching the market.

  Size of total workforce: Workforce shrinking from a total 4,629 employees as of February 20, 2013.²

• #9. Actavis

  Number of job layoffs disclosed: 813 jobs

  Details: 350 jobs at Actavis Specialty Brands, the company’s business that more than 40 brand pharmaceutical products, primarily in the U.S. and Canada; all 310 jobs at Lincolnton, NC, manufacturing plant, which the company said it will shut down by mid-2015; 88 jobs in Rockaway, NJ, where Warner Chilcott was formerly headquartered; 65 jobs at manufacturing plant in Corona, CA.

  Why the company did it: Actavis has cited need to eliminate staffing and operations overlaps following $8.5 billion acquisition of Warner Chilcott, completed October 1. Actavis said it would continue to employ 750 at U.S. Specialty Brands, which had a pre-layoff workforce of 1,100. Lincolnton, NC, production of prescription drugs will be shifted to an Actavis site in Salt Lake City, while over-the-counter drug production will be shifted to contract manufacturers. No explanation for job cuts at Rockaway, NJ, which Actavis retains as “New Jersey Corporate Office”. Job cuts in Corona, CA, part of a plan to reposition the facility into a “Center of Excellence” for manufacturing oral contraceptives, following a 12-year dispute with the FDA over agency allegations of GMP violations stretching back to predecessor Watson Pharmaceuticals.^3,4

  Size of total workforce: Workforce will shrink from a total of approximately 17,700 employees as of December 31, 2012.

• #8. Takeda Pharmaceutical

  Number of job layoffs disclosed: 1,500 jobs

  Details: Takeda executive disclosed plan for workforce reduction of 5% or about 1,500 positions.⁵

  Why the company did it: Planned job cut would be part of company’s “Project Summit” restructuring initiative, which followed the launch of a generic competitor to Takeda’s diabetes drug Actos. Company has not stated where jobs would be cut, but has identified “four areas of initiatives” in cost-cutting: sales and marketing, production and supply, R&D, and general and administrative.⁶ On February 4, company identified among its cost-cutting measures “consolidation or manufacturing and R&D facilities” in Europe.⁷

  Size of total workforce: Workforce will shrink from a total 31,057 employees as of September 30, 2013.

• #7. Boehringer Ingelheim

  Number of job layoffs disclosed: 1,571 jobs

  Details: All 1,100 jobs at Ben Venue sterile injectables plant in Bedford, OH, which will shut down, the company said October 4; all 240 jobs at API manufacturing plant in Petersburg, VA, announced August 15; 143 sales jobs and 35 administrative jobs in Paris and Reims, France, accounting for 20% of the company’s French subsidiary, disclosed by labor leaders September 9; and 53 jobs at Blanquefort, Gironde, France, following a planned shutdown of the plant operated by subsidiary Labso Fine Chemicals.

  Why the company did it: Company cited projected operating losses at Ben Venue plant of more than $700 million over five years, on top of $350 million spent correcting manufacturing problems like those that led to a 2011 shutdown. Petersburg, VA, plant shut down after company concluded it needed to shrink manufacturing capabilities, then failed to find a buyer for the plant; company also cited overall production overcapacity in the pharma industry and competition from foreign-produced materials. Shutdown of the Blanquefort plant—the last operated by the company in France—was part of a reorganization of the company’s chemical plants designed to reduce manufacturing overcapacity.⁸

  Size of total workforce: Workforce shrunk from 46,228 employees at the end of 2012.

• #6. Eli Lilly

  Number of job layoffs disclosed: Up to 1,624 jobs⁹

  Details: An anticipated, roughly 1,245 full-time U.S sales employees eliminated starting June 30, 2013, in U.S. biomedicines division—of which about 560 “will have opportunities to be placed in open roles and, if so placed, will not be terminated”; as well as 379 fixed-duration contract employees, 97 of whom worked under contracts that ended before, but were extended to, June 30.^9,10

  Why the company did it: A response to “changing customer requirements, evolutions in the U.S. health care environment and the upcoming loss in exclusivity” and projected drop in sales as patent protection expires in December on the antidepressant Cymbalta (duloxetine hydrochloride), and in March 2014 on the osteoporosis drug Evista (raloxifene hydrochloride).

  Size of total workforce: Workforce reduced to 37,925 employees as of the end of 2013.

• • #5. Novartis

    Number of job layoffs disclosed: 1,804 jobs

    Details: All 371 jobs at a respiratory R&D site in Horsham, West Sussex, U.K.; a net 325 jobs lost by eliminating 500 R&D positions companywide while adding 175 jobs in Cambridge, MA;¹¹ all 300 jobs over two years at Lincoln, NE, consumer-drugs plant;¹² all 300 jobs at Alcon contact lens plant in Mississauga, Canada; 262 Ciba Vision workers in Des Plains, IL; about 120 jobs at Novartis Institutes for BioMedical Research’s Alcon Labs campus in Fort Worth, TX; 72 mostly field-based marketing and sales support positions for diabetes and COPD products in the U.K.; 54 global legal, IT, HR, finance, procurement, and vaccines research positions in Emeryville, CA.

    Why the company did it: Part of R&D restructuring designed to cut drug development costs; Lincoln plant being repositioned for production of solids and powder, principally for Sentinel, Excedrin, and Theraflu.¹² Company is consolidating Ciba Vision and Alcon Laboratories, and concentrating U.S.-based eye disease research in Cambridge, MA. Fort Worth production will be scaled down to solid and powdered drugs only. Emeryville jobs were eliminated as part of a realignment of the facility away from vaccine R&D and more toward diagnostics.

    Size of total workforce: Workforce reduced from a total 133,000 employees.

  • #4. Valeant Pharmaceuticals

    Number of job layoffs disclosed: 2,700 jobs

    Details: 10% to 15% reduction in workforce, or up to 2,700 jobs, following its acquisition of Bausch + Lomb for $8.7 billion

    Why the company did it: Part of a restructuring designed to cut costs following the Bausch + Lomb acquisition, in part by consolidating locations. Company has not detailed where it will cut jobs, though Valeant chairman and CEO J. Michael Pearson told employees it will eliminate Bausch + Lomb’s global structure and reduce “significantly” its regional structure to reflect Valeant’s decentralized operating philosophy, which entails a corporate staff of fewer than 100 people.¹³

    Size of total workforce: Workforce reduced from the combined 18,000 employees resulting from Bausch + Lomb acquisition.

  • #3. Teva Pharmaceutical Industries

    Number of job layoffs disclosed: About 5,000 jobs

    Details: 10% reduction in workforce planned by 2017; earlier this year, company committed to carrying out the job cuts as announced October 10 by then-CEO Jeremy Levin, D.Phil., who resigned less than a month later after 17 months in office. Company has not detailed where it will cut jobs and how operations would be affected.

    Why the company did it: Part of a global restructuring intended to increase organization effectiveness, improve manufacturing efficiency, reduce excess capacity, and divest the company of non-core assets. The restructuring also addresses anticipated challenges, such as the launch of a generic version of the company’s top-selling drug, the multiple sclerosis treatment Copaxone.

    Size of total workforce: Workforce to shrink from approximately 45,000 employees as of December 31, 2013.

  • #2. AstraZeneca

    Number of job layoffs disclosed: 3,900 jobs

    Details: 2,300 SG&A jobs, announced March 21, 2013, and 1,600 R&D, announced three days earlier.

    Why the company did it: Part of a comeback strategy by CEO Pascal Soriot following years of clinical setbacks involving drug candidates the company hoped would make up for sales revenues to be lost from “patent cliff” expirations through 2014, when the company will lose U.S. patent protection for two of its biggest selling drugs—its proton pump inhibitor Nexium, and its asthma and COPD medicine Symbicort.

    Size of total workforce: Workforce will shrink from a total of about 51,700 employees (Annual Report and Form 20-F Information 2012). The 2013 layoffs brought to 5,050 the number of jobs—roughly 10% of the current workforce—slated for elimination through 2016 as of December 31, 2013.¹⁴

  • #1. Merck & Co.

    Number of job layoffs disclosed: 8,500 jobs

    Details: 8,500 R&D and commercial operations jobs to be eliminated companywide through 2015, announced October 1. Merck has confirmed or disclosed layoffs as they have occurred: 500 jobs in West Point, PA, an unknown number in Puerto Rico, all 570 employees at a manufacturing plant in Swords, Ireland, and all 152 sales representatives based in Upper Gwynedd, PA who had promoted the antipsychotic drug Saphris (asenapine), effective February 3.^15,16

    Why the company did it: Part of a restructuring designed to help company bolster its pipeline and cut costs, while increasing flexibility through a more agile operating model that will target more spending to what it deems highest-potential growth opportunities.

    Size of total workforce: The new layoffs, on top of an earlier-announced reduction of 8,500 jobs, will shrink by some 20% the company’s global workforce of 81,000 employees.

26. Top 10 U.S. Biopharma Clusters

• #10. Chicago

  While the bottom of a top-10 list is usually not much to brag about, just making the list shows how far the Windy City and its suburbs have progressed in biopharma over the past decade. Chicagoland was eighth in patents granted (360), though the number of applied-for patents published was 800 in 2012, double just four years earlier. The region was ninth in VC funding (about $87 million) and 10th in NIH funding ($48.4 million). It doesn’t have the lab market or workforce size (29,230 within the 560-acre Illinois Medical District) of the larger clusters. And it recently lost the 2016 Biotechnology Industry Organization convention to San Francisco. But the region is home to AbbVie and the U.S. headquarters of Japanese pharma giants Takeda Pharmaceutical and Astellas Pharma. And any region that hosts the annual BIO International Convention three times in less than a decade (2006, 2010, 2013) can’t be that bad.

• #9. Los Angeles

  The nation’s second-largest city is at the center of California’s third biopharma region after San Francisco and San Diego. Yet the Los Angeles region has some notable employers, from Amgen in Thousand Oaks, to California Institute of Technology (Caltech) in Pasadena, and UCLA in, where else? Together they employed 23,054 people in “biomedical” positions (California Biomedical Industry Report 2013), giving the region one of the smallest workforces among the top-10. LA placed ninth in NIH funding ($65.4 million) but climbed to the middle of the pack in patents granted (550).

• #8. Raleigh-Durham, NC (includes Research Triangle Park, NC)

  North Carolina’s biopharma mecca is up toward the middle in NIH grants (sixth at $109.3 million), among the top in jobs (58,000), but decidedly down the list in patents (ninth at 312) and VC funding (eighth at about $150 million). A generation after Research Triangle Park drew pharma manufacturing from the Northeast, the region’s industry is riding a wave of contract research organization growth, anchored by the headquarters of the world’s largest CRO, Quintiles. Last month the Research Triangle Foundation paid $17 million to acquire about 100 additional acres, some of which is expected to draw new biopharmas.

• #7. Philadelphia

  The birthplace of Liberty has sought to leverage its heritage as a pharma mecca to build a more diverse concentration of biopharma businesses and institutions (including 25 medical schools) that has grown to 42,000 jobs (according to regional economic development group Select Greater Philadelphia). Anchors include Philadelphia’s University City Science Center, where earlier this month the University of Pennsylvania agreed to expand to 55,900 square feet in a new office-lab building being co-developed by the center and Wexford Science & Technology. While the region’s NIH grant haul is strong enough to place fourth ($123.4 million), the region was seventh in drawing private capital ($164 million) and in patents (368).

• #6. Seattle

  The home of Starbucks wins plenty of NIH bucks—enough to be ranked third ($147.2 million). And while the city of the Super Bowl-champion Seahawks took home even more VC bucks ($238 million), the region only scored sixth on that measure. That reflects the region’s being anchored more to academic and independent research institutions than local companies; combined, all employers account for nearly all the state’s 35,500 life-sci jobs. Seattle also scored sixth in patents (618) and is also middle-of-the-pack in lab space at five million square feet.

• #5. New York

  The Big Apple is big in grant money—it finished second in NIH funds so far in FY 2014 ($175.1 million), thanks to its concentration of top flight research hospitals, which account for most of the city’s 60,000+ bio jobs (New York City Economic Development Corp.). But New York and its suburbs are only now playing catchup in commercializing that research. The region still has a ways to go, since it’s fifth in patents (730), fifth in VC financing ($326 million), and toward the bottom in lab space (1.7 million square feet)—which could change if 1.1 million square feet now in development gets built as planned.

• #4. Maryland / Suburban Washington, DC

  The home region of NIH is also home to the nation’s longtime academic leader in research grant funding, the Johns Hopkins University. Hopkins’ $69.5 million accounted for more than three-quarters (78.5%) of the grant funding awarded by the agency to the region ($88.6 million), which finished eighth on that measure. In Montgomery County, MD, the center of the region’s biopharma activity, NIH is the largest employer and FDA, third largest. Yet many of Maryland’s 71,600 life sciences jobs (BioMaryland) are at biopharmas, the largest being AstraZeneca’s MedImmune subsidiary and GlaxoSmithKline, which snapped up home-grown Human Genome Sciences in 2012. Maryland/Washington also lags in lab space (6 million square feet), but places a solid fourth in both patents (1,200) and VC funding ($353 million).

• #3. San Diego

  The San Diego region placed third in patents (1,335), VC funding ($386 million), and lab space (13.05 million square feet), hence its ranking here as third. But the city that calls itself “America’s Finest” lagged in NIH funding (7th at $98.4 million) despite its dense concentration of research institutes and one of the larger campuses within the University of California system. The region’s industry group Biocom counted 56,605 jobs in San Diego County in 2012, 143 more jobs than Massachusetts—a figure expected to grow about 6% by this year.

• #2. Boston / Cambridge, MA

  The region that’s home to the World Series-winning Red Sox also outslugged the competition in NIH funds, accounting for about 75% of all Massachusetts’ grant money from the agency ($201.4 million), but lost to San Francisco on patents (2,908) and also finished second to the Bay Area in VC funding ($979 million) and lab space (18.687 million square feet). The region accounts for nearly all the state’s 56,462 industry jobs in 2012 (according to MassBio), but that number should start climbing this year, when the first construction projects conclude in a building boom totaling 3.5+ million square feet of lab and office space valued at more than $2 billion.

• #1. San Francisco Bay Area

  The Bay Area narrowly topped Boston/Cambridge in patents (3,492) and loses out to the Red Sox region in NIH grants ($119.8 million). Jobs-wise it depends who you believe: California’s Economic Development Department lists 110,337 Bay Area “life sciences” jobs in the latest tally (Q1 2013), while a recently released report showed only 47,019. Split the difference, and call the region number one. But there’s no debating San Francisco and its suburbs were hands down number one in VC funding as the only region to top a billion dollars last year ($1.447 billion, to be exact), and also a clear number one in lab space, with a total 29.7 million square feet.

25. Top 10 Multiple Sclerosis Drugs

• #10. Extavia (interferon beta-1b)

  Sponsor/Developer: Novartis

  Dosage strength, form, and frequency: Recommended dose is 0.25 mg injected subcutaneously every other day. Generally, start at .0625 mg (0.25 mL) subcutaneously every other day, and increase over a six week period to 0.25 mg (1 mL) every other day

  Mechanism of action: Unknown

  Indication: Relapsing forms of MS to reduce the frequency of clinical exacerbations

  2013 sales: $159 million

• #9. Aubagio (teriflunomide)

  Sponsor/Developer: Genzyme (Sanofi)

  Dosage strength, form, and frequency: 7 mg and 14 mg tablets, once-daily, with or without food

  Mechanism of action: Pyrimidine synthesis inhibitor

  Indication: Relapsing forms of MS

  2013 sales: About $226 million (€166 million)¹

• #8. Ampyra (dalfampridine) / Fampyra

  Sponsor/Developer: Acorda Therapeutics and Biogen Idec

  Dosage form and strength: 10 mg tablets

  Mechanism of action: Potassium channel blocker

  Indication: Improve walking in patients with MS

  2013 sales: $302.301 million²

• #7. Rebif (interferon beta-1a)

  Sponsor/Developer: EMD Serono and Pfizer

  Dosage strength, form, and frequency: 22 mcg and 44 mcg by injection three times per week. Rebif should be administered, if possible, at the same time (preferably late afternoon or evening) on the same three days, at least 48 hours apart each week. Generally, patients should be started at 20% of the prescribed dose three times per week and increased over a four-week period to the targeted dose, either 22 or 44 mcg three times per week

  Mechanism of action: Unknown

  Indication: Relapsing forms of MS, to decrease the frequency of clinical exacerbations and delay the accumulation of physical disability.

  2013 Sales: About $622 million (€460 million) through September 30, 2013^3,4

• #6. Tecfidera (dimethyl fumarate)

  Sponsor/Developer: Biogen Idec

  Dosage strength, form, and frequency: Starting dose: 120 mg twice a day, orally, for seven days, followed by maintenance dose after seven days: 240 mg twice a day, orally. Capsules must be swallowed whole and intact. Tecfidera can be taken with or without food.

  Mechanism of action: Unknown

  Indication: Relapsing forms of MS

  2013 Sales: $876 million
• • #5. Betaseron (Betaferon; Interferon beta-1b)

    Sponsor/Developer: Bayer HealthCare

    Dosage strength, form, and frequency: Recommended dose is 0.25 mg via injection every other day. Generally, patients are advised to start at 0.0625 mg (0.25 mL) via injection every other day, and increase over a six-week period to 0.25 mg (1 mL) every other day

    Mechanism of action: Unknown

    Indication: Relapsing forms of MS, to reduce the frequency of clinical exacerbations

    2013 sales: $1.05 billion (€779 million) as of September 30, 2013⁵

  • #4. Tysabri (natalizumab)

    Sponsor/Developer: Biogen Idec⁶

    Dosage strength, form, and frequency: 300 mg infused intravenously over approximately one hour, every four weeks

    Mechanism of action: Binds to the α4-subunit of α4β1 and α4β7 integrins expressed on the surface of all leukocytes except neutrophils, and inhibits the α4-mediated adhesion of leukocytes to their counter-receptor(s).

    Indication: Adults with relapsing forms of MS

    2013 sales: $1.67 billion⁷

  • #3. Gilenya (fingolimod)

    Sponsor/Developer: Novartis

    Dosage strength, form, and frequency: Recommended dose: 0.5 mg orally once daily, with or without food

    Mechanism of action: Metabolized into Fingolimod-phosphate, a sphingosine 1-phosphate receptor modulator that binds with high affinity to sphingosine 1-phosphate receptors 1, 3, 4, and 5. Fingolimod-phosphate blocks the capacity of lymphocytes to egress from lymph nodes, reducing the number of lymphocytes in peripheral blood. The mechanism of action on MS is unknown but may involve reduction of lymphocyte migration into the central nervous system.

    Indication: Relapsing forms of MS, to reduce the frequency of clinical exacerbations and to delay the accumulation of physical disability

    2013 sales: $1.9 billion

  • #2. Avonex (interferon beta-1a)

    Sponsor/Developer: Biogen Idec

    Dosage strength, form, and frequency: 30 micrograms once a week is recommended. To reduce the incidence and severity of flu-like symptoms that may occur when initiating Avonex therapy at a dose of 30 micrograms, Avonex may be started at a dose of 7.5 micrograms and the dose may be increased by 7.5 micrograms each week for the next three weeks until the recommended dose of 30 micrograms is achieved.

    Mechanism of action: Unknown

    Indication: Relapsing forms of MS, to slow the accumulation of physical disability and decrease the frequency of clinical exacerbations

    2013 Sales: $3 billion⁸

  • #1. Copaxone (glatiramer acetate)

    Sponsor/Developer: Teva

    Dosage strength, form, and frequency: 20 mg/mL daily by injection; 40 mg/mL three times per week by injection (9)

    Mechanism of action: Not fully understood; believed to act by modifying immune processes that are believed to be responsible for the pathogenesis of MS.

    Indication: Relapsing forms of MS

    2013 sales: $4.328 billion