Outside movie theaters, there are two main aspects to the current story of vaccines. First comes the eternal search for new vaccines that can either protect against heretofore untreatable ailments or improve upon the immunity given by predecessor vaccines. New methodologies for bottom-up vaccine design also figure into this search. And second comes the quest for new and better methods of vaccine production.
One of the defiant diseases which vaccine researchers have focused on is anthrax. The U.S. Department of Health and Human Services’ Biomedical Advanced Research and Development Authority (BARDA) has contracted with Vaxin Inc. of Rockville, MD, to conduct studies and develop manufacturing processes for their new vaccine, AdVAV.8 Based on the work of Vaxin founder De-chu Tang, AdVAV will be administered as a nasal spray. Said Vaxin chief executive officer Bill Enright, “For a small company like Vaxin, having an award of this magnitude is really validating for our technology.”9
Cancers of several types present promising territory for vaccine treatment. Currently, the FDA has approved treatments against the aforementioned HPV: Gardasil, manufactured by Merck; and Cervarix, from GlaxoSmithKline.10 Merck also manufactures Recombivax, which protects against the hepatitis B virus (HBV).11
The efficacy of these approved vaccines has led to nascent assaults against many other types of cancer. Currently, the clinical trials database maintained by the National Cancer Institute is tracking trials of vaccines against bladder cancer, brain tumors, breast cancer, cervical cancer, Hodgkin’s lymphoma, kidney cancer, melanoma, multiple myeloma, leukemia, lung cancer, non-Hodgkin’s lymphoma, pancreatic cancer, prostate cancer, and solid tumors.12
But cancers are not the only target for vaccine researchers. One area crying out for the vaccine approach involves the so-called “emerging and neglected diseases” most often found in developing countries. Seattle BioMed, the largest independent, non-profit organization devoted solely to infectious disease research, has identified the following scourges as candidates for vaccines: African sleeping sickness, amebiasis, Chagas’ disease, leishmaniasis, mycobacterium avium complex, toxoplasmosis, and general fungal infections.13,14 Professor Richard Moxon of the Oxford Vaccine Group concurs, calling for the development of at least 20 new vaccines in the near-future, including perhaps even one against insulin-dependent diabetes.15
But the Big Three among theoretically preventable diseases, according to Seattle BioMed and Moxon, are malaria, HIV/AIDS and tuberculosis.13 Perfecting vaccines against this trio could save hundred of millions of lives over the long run.
Progress is slow yet encouraging in the campaign against HIV/AIDS, despite the disappointing failure of Merck’s experimental HIV vaccine in 2007.16 Two recent projects hold out hope.
David Graham, a molecular biologist at The Johns Hopkins University in Baltimore, MD, has learned that stripping a cholesterol membrane from HIV takes away the virus’ ability to disrupt communication among the body’s disease-fighting cells. “By stealing cholesterol from the envelope of the virus, we can neutralize the subversion,” said Dr. Graham. “We’ve broken the code; we can shut down the type of interference that HIV is having on the immune system.” He believes this insight into the mechanism of HIV subversion is essential in the creation of any vaccine.17
Meanwhile, however, without reference to Graham’s work, early phase trials of a HIV vaccine dubbed MVA-B, conducted by the Spanish Superior Scientific Research Council (CSIC), have resulted in 90% of volunteers developing an immunological response against the virus. Mariano Esteban, head researcher from CSIC’s National Biotech Center, explained that the insertion of four HIV genes into a previously used vaccine (MVA) for smallpox “is like showing a picture of the HIV so that [the human immune system] is able to recognize it if it sees it again in the future.”18
On the somewhat more tractable malaria front, a first step toward global immunization is Seattle BioMed’s genetically attenuated whole parasite (GAP) malaria vaccine, currently undergoing human trials.19 Seattle BioMed isn’t alone in its focus on this mosquito-borne scourge. GlaxoSmithKline, with support from the Bill and Melinda Gates Foundation, continues to field test its RTS, S vaccine in Africa, achieving results that show a high protection rate. GSK chief executive officer Andrew Witty believes that much “will become clearer in the next two years or so.”16,20
A pioneer in the field of systems biology, Dr. Alan Aderem, who in 2012 is slated to become Seattle BioMed’s president, has placed an emphasis on new and more efficient methods of vaccine development, involving “a clear understanding of the precise mixture of immunological responses that must occur if a vaccine is to induce a strong protective reaction.”16 Likewise, the antigens that provoke the immune response must be intelligently winkled out from the start, rather than laboriously derived through sheer trial and error.
This approach is manifest most clearly in a third malaria vaccine — MSP3 — currently being field tested in Burkina Faso and Mali by Dr. Pierre Druilhe from the Pasteur Institute.21 Not only was the vaccine systematically engineered, but novel production methods have been employed. Dr. Druilhe explained, “In my opinion, the rational discovery process for vaccines has yet to be started. What we did was the opposite of what everyone else did. We saw that we could not study the 5,000 items pertaining to Plasmodium [the malaria-causing parasite]. It was hopeless to study them one by one. But human beings exposed to malaria acquire protection after about seven hundred malaria attacks. We found a mechanism for the protection that had not been described before: a cooperation between antibodies and leukocytes. When we found this mechanism we used it to screen out the whole genome of the parasite, to fish out the target antigen that matched the antibodies. That’s how we found MSP3.”
And what about producing it in sufficient quantities for the clinical trials? “We don’t culture our vaccine, we synthesize a protein by chemical means” said Dr. Druilhe. “It’s a totally different platform, fast and very affordable. We can also produce hundreds of millions of doses at 10 cents a dose. And protein synthesis is improving in speed and cost every year. Ten cents could become three times less in five years from now. People think of the synthetic peptides of 20 or 30 years ago, when they were small. That’s completely outdated. Today we are producing synthetic proteins in a single synthesis of some two hundred amino acids. And you can link them together when you need larger. They exhibit a high degree of purity too.”
Dr. Druilhe’s emphasis on new means of production speaks to an industry-wide revolution that is changing the way vaccine manufacturers work.
A vital first step in crafting and marketing new vaccines is being able to reliably assay their immunogenic potency and purity, and to chart their molecular composition and cellular interactions with living organisms. The firm of PerkinElmer is giving researchers and manufacturers the tools they need to accurately guide their development and production.
Martina Bielefeld-Sevigny, vice president and general manager Drug Discovery and Research Reagent Solutions at Perkin Elmer, and Vincent Dupriez, a principal scientist at PE, took the time to inform us of the latest offerings from their company.
PerkinElmer has long provided the pharmaceutical industry with powerful off-the-shelf toolkits — hardware, software, reagents, and training, with customization if necessary — but Ms. Bielefeld-Sevigny pointed to a notable recent shift in the customer base: “A few years ago, with the trend toward more biologics and vaccines, we discovered that our technologies are not only very useful for small molecule discovery but also very well suited for the large-molecule world. Some features of high-throughput screening technologies are perfectly suited for the biopharmaceutical and vaccine world.”
Mr. Dupriez added, “If a customer needs a very specific assay that is not in our catalog, we can develop that.”
Moving into new frontiers, PerkinElmer recently acquired Caliper Life Sciences, Inc.; Caliper’s microfluidic "lab on a chip" technology will further expand the products and technologies PerkinElmer can offer to the vaccines market. But even now, Mr. Dupriez related, PE offers many products particularly suited for vaccine development. The extremely sensitive Alpha assay technology allows for the selection of the most promising molecules for subsequent development, and feeds into 3-D modeling software that allows identification of the best candidates in silico. The Britelite Ultra-High Sensitivity Luminescence Reporter Gene Assay System is being used by the CDC in its quest for an anti-AIDS vaccine.22 Mr. Dupriez also cited other technology offerings.
And behind all this hardware, massive amounts of sophisticated software allow deep, intelligent parsing of the data. The increased focus and commitment to integrated solutions and data analysis is reflected by recent acquisitions including CambridgeSoft, Inc. “Analyzing the data is as crucial as generating the data,” observed Ms. Bielefeld-Sevigny.23
But even taking advantage of the smart tools that a company like PerkinElmer supplies, vaccine production demands immense ingenuity, attention to detail, and continuous refinement of technology.
Traditionally, vaccine production has involved three possible substrates: chicken eggs, human/mammalian cell lines in bioreactors, or bacteria and yeast in fermentors.24 The limitations and failures of the oldest method, chicken eggs, was seen most clearly during the recent H1N1 Swine Flu pandemic. Needing 160 million doses, the U.S. Department of Health and Human Services ended up with fewer than 30 million, due to an unexpectedly low dose-per-egg production volume. Clearly, 21st-century needs can no longer be met exclusively with this 50-year-old technology.25
Dr. Magda Marquet, founder and co-chair of Althea Technologies, believes in the radical virtues of the technology embraced by Althea: e. coli and plasmids in fermentors producing DNA vaccines: proteins expressed by the chosen DNA in its plasmid vessel, with the goal of triggering an immune response in the recipient. This approach is faster, cheaper and results in purer product, she contends.26,27
Dr. Marquet outlined the pioneering methods in operation at Althea: “We are the U.S. leader in this field, producing many lots of DNA vaccine. We have a lot of experience in this area, and our facilities offer services for pre-clinical batches — Phase I, Phase II, and Phase III — and very recently we got FDA approval to manufacture to the commercial level as well. We employ different facilities depending on the scale and compliance level. Our clinical facility can handle up to 100 liter production scale. Our commercial facility can handle up to 1,000 liter production, and its compliance level is a lot more stringent. A third facility handles R&D.”
How does a fermentation run begin? “In some cases, our customers have their plasmids already optimized,” Dr. Marquet said. “In other cases, we do the optimization and strain selection, looking at different strains of e.coli for better yield, better stability of the product and better expression of the product. We also have the capability to use yeast, but for other kinds of biologics, not vaccines.”
One very important step in the process is sterilizing the system between batches. Dr. Marquet explained, “Once the fermentation run is finished, we have to clean and sterilize the equipment. For the cleaning validation we use a very stringent method. We identify [any remaining plasmid traces] by PCR.”
On the output end, Althea puts its products through several tests. “The first test is for identity: is this the right product? Then, purity, to make sure there no contamination from endotoxins or chromosomal DNA or RNA or proteins. Then we also test for potency.”
All of this daily activity takes on an environmental aspect too. “We have a major effort to make our company more and more green. We have a company-wide effort in that arena, rewarding our employees for helping us to improve. We go the extra mile to make sure that our operations are safe for our employees and the environment.”
What lies ahead for vaccine makers? Dr. Marquet is optimistic. “The world of vaccines has changed tremendously,” she remarked. “A couple of decades ago, people did not really want to hear about vaccines. They were not very fashionable. Now they are very important in the pharmaceutical industry, especially for developing countries. There is a renewal in the world of vaccines. Usually, companies that work in vaccine production are well-funded and they are doing quite well.”
Still, even after a vaccine is produced, by whatever method, a final step remains. If the establishment that cultured the vaccine does not do fills, how does it get containerized?
Enter a CMO such as Grand River Aseptic Manufacturing, which president Gregory Gonzales describes as “a relatively young and lean company.” Having opened its doors in January, 2011, Grand River offers expertise in refining vaccine formulations. Able to fill standard and non-standard containers of all types, it also helps ensure vaccine stability with its lyophilization process (important for promoting long shelf life in Third World countries lacking a reliable cold chain for their vaccine stocks). Specializing in small batches for clinical trials, but with larger production capacity, Grand River offers its customers a chance to view fills of their product remotely over the internet, thanks to a closed-circuit monitoring system. All of this takes place in their Class 100 (ISO 5) facility, “more sterile than an operating room.”
The founding of Grand River on the Grand Valley State University campus in Grand Rapids, MI, is a testament to the power of cooperation between government agencies, educational institutions and private enterprise. “Our initial money came from a grant from the state of Michigan. We were a joint venture between the Van Andel Research Insititute and Grand Valley State University.” Extrapolating from Van Andel’s own needs to a wider demand for small-scale API production facilities, and utilizing donated buildings from Steelcase, Inc., the young firm has quickly positioned itself in the market.
A longer-established firm of the same type, but one just as innovative, is JHP Pharmaceuticals. Daniel Leone, JHP’s senior director, Business Development, took the time to elaborate on the company’s role in vaccine production.
“It’s a meaningful and growing portion of the business. Filling these vaccines is a very complex and quality-driven type of activity.” Mr. Leone noted that animal-derived vaccines from egg technology can only be handled by a few specialized firms, according to FDA regulations, thus contributing to the bottlenecks with that old tech. “But there are more forward-looking types of biopharmaceutical product development that we are filling on either a clinical or commercial scale.”
“This year we started up what’s called a clinical filler, a smaller scale filling line. Products in clinical stage have a high API cost, so you have to make sure you get as many vials out of a fill as possible. A smaller line minimizes line loss [irretrievable product left in the network at shutdown]. And using all disposable parts helps in the turnaround time for cleanup.” Capacious cold storage facilities (minus 40° to minus 80°), as well as the ability to maintain the cold chain during a fill, are vital.
Lastly, JHP offers customized formulation abilities on site. It can precisely adjust ratios of excipients and other non-API components before the fill begins. “At JHP we are very able and willing to work with specialized formulation equipment. Our customers are trusting us to use and store their equipment. You have to be very accomplished for them to feel comfortable with that.”
This tradition of close consultation and cooperation among vaccine researchers, vaccine makers, and CMOs can only lead to a new Golden Age of medicines for a range of humanity’s needs.
- http://en.wikipedia.org/wiki/HPV_vaccine#Opposition_in_the_ United_States
Paul Di Filippo is the author of numerous science fiction novels and short fiction collections, and his criticism has appeared in The Washington Post. He can be reached at email@example.com.