Progress on Cell-Based Therapies

Spinal cord injury and HIV are the targets of two innovative cell-based therapies, with the planned HIV treatment relying on gene-therapy to produce in vivo RNA interference (RNAi).  In a very short news piece in Nature (subscription required), "Pioneering HIV treatment would use interference and gene therapy", Erika Check writes that;

If the FDA says yes, [John Rossi and his team at  at City of Hope's Beckman Research Institute] will test the therapy on five HIV patients who have a blood cancer called lymphoma. They will treat the patients' lymphoma with aggressive chemotherapy and a bone-marrow transplant — a normal procedure. But before the transplant, they will use gene therapy to add stretches of DNA to stem cells in the bone marrow. It is hoped that molecules encoded by the added genes will trigger the cells' RNAi defences against HIV.

The trial is different from the RNAi trials already under way, because the molecules used in those studies remain in the body for only a short time. The City of Hope researchers will deliver DNA packaged into a gene-therapy vector that could persist in patients for months or even years.

The Recombinant DNA Advisory Committee is having Rossi perform additional safety tests before giving the OK for the trial.  If this works out, it will demonstrate a remarkably powerful way to alter human physiology through the permanent (?) addition of a new RNAi pathway.  The strategy pursued by Rossi, et al., would provide a pool of stem cells that produce lymphocytes immune to HIV.  Since HIV shows some tropism for neural and other tissues, the treatment may not completely rid patients of the virus, but as lymphocytes carrying HIV die out at least the patient would have a source of healthy immune cells.  As this research goes forward, we can expect significant press coverage because the technique will probably find immediate use in treating many other chronic diseases.

The work on repairing spinal cord injury through cord blood cell transplantation has received surprisingly little press.  In an article in Cytotherapy, K-S Kang et al., demonstrate that multipotent stem cells (MSC) derived from umbilical cord blood colonized the site of a spinal cord injury in a 37-year old women who had been a paraplegic for almost 20 years.  The MSCs were amplified in vitro and then surgically transplanted to the site of the injury.

Prior to treatment, the patient showed no somatosensory or motor activity in her lower body, and nerve conduction studies confirmed the extent of the damage.  After transplantation, the patient regained significant sensation within 2 weeks and could maintain an upright posture.  She was able to move her lower legs shortly thereafter.  Nerve conduction studies were used to confirm the extent of recovered electrical activity.  CT and MRI demonstrated regeneration of the spinal cord.

My neurophysiology is more than a tad rusty, which means the import of some specific things reported in the paper isn't immediately clear to me, but the overall results are enough to make anyone take notice; a previously paralyzed patient is now able to at least feel stimulation in her lower limbs, maintain an upright posture unassisted, and has regained some motion in her lower legs.

The specific mechanisms behind the recovery must now be determined, including how the MSCs produce such dramatic improvement.  The authors also note that they cannot rule out the surgery as effecting some recovery.  But the demonstrated increase in electrical activity and motion is extraordinary.  And you have to imagine that the ability to maintain an upright position unassisted for the first time in 20 years is by itself an enormous gift.

This is just one patient, and just one paper, so lots of work is required before anything like this becomes standard treatment.  It is also unclear what the long term effects of the procedure and the new cells will be.  Then there is the little problem that in the U.S. working with stem cells is a tad problematic, regardless of their source.  This study notably, took place in Korea.

Nonetheless, what fantastic news.

Big Day for Bird Flu News

Today brings news that a live, infectious strain of the 1918 flu has been reconstructed in the lab.  The press has responded smartly this time (New York Times, AP via Wired News, CNN, The New Scientist) -- with fairly decent reporting -- no doubt in part because President Bush addressed the situation in a press conference, suggesting that the U.S. military might be involved in managing a pandemic.  There is quite the political hullabaloo in Washington DC, too boot, with the New York Times reporting that in response to closed door briefing last week a Pentagon appropriations bill has been boosted by USD 3.9 Billion solely for dealing with the flu (it's unclear from the story whether that money is intended for use by the Pentagon or by the NIH).  Politicians, and political parties, are evidently trying to outdo one another in being out front on this issue, despite the fact that we are hopelessly behind.  No surprise there.

As far as the biology goes, for those who have been paying attention, or even just reading this blog, there isn't that much new in today's reports.  As related by the Times, papers in Science and Nature basically confirm at least part of the molecular detective story told by Oxford et al., namely that the 1918 flu jumped directly from birds to humans.  Thus, "The Swine Flu" is a misnomer for the disease caused by this particular bug.  There is no further progress on figuring out when and where the bug evolved, as far as I can tell.

The Nature paper, from Jeffrey Taubenberger's group, describes his work in extracting flu sequences from preserved lung tissue and from a corpse frozen in permafrost.  This paper is a bioinformatic comparison (i.e. no experiments) that characterizes the 1918 flu polymerase genes.  The Science article describes infecting mice with a reconstructed virus, which turned out to be considerably more lethal than expected.  The article will be out on 7 October.  I will write more when I've had a chance to carefully ready both papers.

The publication of the viral sequence, with accompanying descriptions of how to reconstruct live virus, obviously raise questions about safety.  Every story above mentions that the investigators and journal editors balanced the benefits and threats, and asked for a review from the National Science Advisory Board for Biosecurity (NSABB), before going to press.  Fortunately, everyone came to a conclusion in favor of publishing.  Press reports, including one in Nature, give voice to critics of publishing the sequence and construction methods.  In particular, there are complaints that the 1918 strain could be reconstituted for use as a weapon or that it could simply escape back into the wild.  I am obviously not the only one not much convinced by these arguments.  While very few people alive today have been exposed to the 1918 strain, related strains are often included in annual flu vaccines.  So humans is no longer immune naive for that set of bugs.  As for the use of 1918 as a weapon, the reverse genetics required to produce a live RNA virus from DNA plasmids are decidedly non trivial at the time being.  No doubt, this process will get simpler, but this isn't something you are going to do in your garage.

 

Tamiflu Resistant H5N1?

CNN is reporting that Tamiflu-resistant strains of H5N1 are appearing in Asia, and that "resistance to anti-flu drugs [is] growing worldwide."  It is a typical CNN story, and therefore leaves one wanting more facts, but at least it's enough to start a Google adventure.

In another story, CNN is repeating the forecast by Dr. David Nabarro of up to 150 million deaths from H5N1.

Taking Issue With Henry Niman

Henry Niman has made quite a lot of bother about the Avian Flu in the last year at his site Recombinomics.  It is still unclear whether he has a better handle on the future of the virus than does the WHO, the CDC, the FAO, and the UN, in part because he has yet to publish anything in a peer reviewed article.  But it is clear some folks are getting touchy about Niman's vocal assertions of doom -- here is a blog entry rebutting some of Niman's claims, and otherwise taking him to task for his behavior.  There's more, for those interested in this sort of thing, at drmartinwilliams.com.

UPDATE (24 October 2005):  Here is a previous post of mine that includes an attempt to figure out whether Niman has interesting or useful contributions to the Avian Flu problem, and a post attempting to sort out the difference, if any, between recombination and reassortment.  Also an early commentary on how little data we have about what evolutionary mechanisms result in pandemic flu strains.

UPDATE (1 November 2005):  Here is a post illustrating the gap between Niman's claims and conclusions based on sequencing data from the World Health Organization.

PowderMed's H5N1 DNA Vaccine

The news service at Nature is reporting ("Bird flu vaccine not up to scratch" -- subscription required) that an egg-based whole virus Avian Flu vaccine, recently announced with fanfare as solving all our problems, is unlikely to be useful.  Fortunately, there is an alternative.  Alas, regulatory issues may prevent PowderMed from distributing it's DNA vaccine for the Avian Flu for some years yet.

The whole virus vaccine was announced with great fanfare just a few months ago.  But as I've written previously (here and here), egg-based vaccine production will never be sufficient for rapid responses to quickly evolving viruses like the flu.  Moreover, to produce a decent immune response the whole virus vaccine must be administered in 2 doses, each 6 times larger than an annual flu shot.  While this is in part because humans have never been exposed to an H5 virus (we are "immune naive"), it also appears that it just isn't a great vaccine.

While it is true that enthusiasm for DNA vaccines has gone through a bit of a boom and bust cycle, early results requiring high dose intramuscular injection are not representative of how the current technology works.  Genes coding for antigens for new viruses are slotted into a plasmid vector that has been proved safe in humans, the plasmids are loaded onto micron-sized gold particles, and the particles are injected into the skin using a high-pressure helium blast.  At Bio-ERA, we've been studying the vaccine and its utility, and it looks like the real deal.  An article in Red Herring quotes the CEO of PowderMed as saying;

What we really believe we’ve got is not just a vaccine; we actually have the ability to produce a capability for a country to cover anything really.  We have designed, with the help of contract manufacturers, a facility that would be able to produce 150 million doses in three months.

The key to the value of PowderMed technology is that the DNA vaccine is delivered directly in the nucleus of dendritic cells in the epidermis.  By getting dendritic cells to express coat proteins from pathogens and then present those proteins in complexes with MHC molecules, the vaccine directly stimulates a cellular immune response; T-cells are thereby primed to recognize and dispose of the virus and infected cells.

Vaccine production in chicken eggs or in cell culture requires at least 6 months to even begin cranking out doses, and requires significant infrastructure to do so.  PowderMed suggests that within three months of sequencing a new pathogen they can have vaccines ready to go.  But my estimates suggest it could be much faster than this.  Included in PowderMed's estimate is the time required to load the vaccine into their proprietary delivery system (a helium powered injector about the size of a flashlight).

My own estimate of the time required to fabricate the plasmids, followed by enzymatic amplification, is more like a week or two.  Injection of the vaccine does require particular technology (a "gene gun") but as it happens those have been used for ~10 years to genetically modify plants and animals.  There are gene guns scattered all across the developed and developing world.  If we had to, if the Avian Flu started to cause real problems in the human population, we could synthesize the vaccine in a widely distributed fashion (anywhere around the globe where people have access to large scale DNA synthesis) and deliver it using gene guns.  True, those instruments were intended for research use only, and were not designed for (or at least not marketed for) use on humans.  But if things start to go south, I'll be first in line.

The four stages of adopting new ideas.

Perhaps the best summary of life in academia that I have ever heard:

There are four stages of adopting new ideas

The first is “It’s impossible”
The second is “Maybe it’s possible, but it’s weak and uninteresting”
The third is, “It’s true and I told you so”
And the fourth is, “I thought of it first”

From H. Koprowski, "Vaccines and sera through plant biotechnology," Vaccine, 23 (2005), 1757-1763.

Synthetic Biology for HIV prevention: "A live microbial microbicide for HIV"

In the latest issue of Proceedings of the National Academy of Sciences, Rao et al demonstrate a fascinating, and probably immensely useful, application of genetic modification.  They altered human commensal strain of E. coli to excrete proteins that prevent HIV from infecting immune cells.

This isn't the first time bacteria have been genetically modified to carry antigens, antibodies, or, as in the present example, peptides that directly interfere with a pathogen's mode of infection.  But it is particularly interesting because the authors chose as a delivery strain a bug that is available as an over the counter probiotic supplement used to treat irritable bowel disease, cholitis, and Crohn's Disease.  The strain, "Nissle 1917", has thus already been demonstrated safe for use in humans, and is distributed in capsules intended for oral ingestion that can be easily manufactured and then stored at room temperature.

It's important to note that while this paper shows the bacterium prevents HIV infection in cell culture, and that the bacterium survives in the intestines of mice while secreting "inhibitory concentrations of the anti-HIV peptide onto mucosal surfaces of the gastrointestinal tract", it does not actually demonstrate prevention of infection in an animal model, let alone humans.  This experiment will no doubt require considerable review by institutional committees, and perhaps by the NIH itself (which is likely, since the work took place at the National Institute of Allergy and Infectious Disease).

Nonetheless, this is a quite sophisticated application of biological technology to address human needs.  The anti-HIV peptide is itself a synthetic product, being a fusion of hemolysin-A and a fragment of gp-41, the later chosen because it binds to the protein complex on HIV that enables it to dock with and then enter human cells.  Hemolysin-A served as the "shipping tag"; it is part of a protein excretion system already present in E. coli.  Thus this work demonstrates the modification of a bacterial strain -- one already known to be well tolerated in humans -- by exploiting an extant protein export system to excrete a synthetic protein cargo that may provide significant protection against HIV.

Like Jay Keasling's work to manufacture anti-malarial drugs in E. coli, Rao et al are on the path to producing organisms that can be easily and inexpensively grown in culture, followed by harvest and packaging of therapeutic compounds or of the bugs themselves.  This production work is similar to commercial efforts in India, China, and other developing countries, and helps pave the way to distributed biological manufacturing(PDF).  There is clearly lots of work left to do before humans are given genetically modified bacteria as preventative microbicides, but the world it is achangin'.

Cheap drug and stem cell trials in China

"Leaders and laggards in the stem cell enterprise," in the 30 June 05 issue of Nature Biotechnology, describes the global distribution of policy and science of stem cell research.  The story notes that;

The wild cards in all this are China and South Korea. Both have extremely talented scientists, but both are (perhaps unfairly) singled out for lax ethical standards and an uninformed public. With fewer shackles on the momentum of ES cell research, the two countries could potentially accelerate products into clinical trials much faster than the rest of the world.

But just because progress might be made in the clinic, it isn't clear that drugs (including cell-based therapies) produced in China will find an easy time in U.S. and European markets.

In "China beckons to clinical trial sponsors", Hepeng Jia writes that;

The clinical trials market is opening up in China...Low cost and ease of access to patients are the main incentives for clinical trial outsourcing...Ying Zhang, marketing director at Beijing-based clinical trial contract research organization (CRO) Excel Medical Technology, estimates that the cost of a clinical trial for a new drug in China is only half of the amount in the United States or Western Europe owing to lower labor and infrastructure costs.

There are, however, a few challenges, including, "poor data standardization, delays in gaining trial authorization from regulators and questionable ethical standards," and a, "poor level of compliance with Western standards."

Despite the harmonization of standards at some clinical trials facilities, it remains to be seen whether the US Food and Drug Administration or European Medicines Agency will accept data obtained from clinical trials carried out in China. Already foreign organizations like Quintiles, CCBR and Canadian company MDS of Toronto, have been seeking endorsement of agencies like the American College of Pathologists to gain recognition for their work there.

Large pharmaceutical corporations are moving into the Chinese market via acquisition.  Alla Katsnelson reports (Nature Biotechnology 23, 765 (2005)) that the Israeli generic manufacturer Teva Pharmaceuticals is buying Tianjin Hualida Biotechnology Pharmaceutical Co, in part because of "regulatory purgatory" in the United States.  "By tackling less strictly regulated markets first, Teva could gain more expertise in manufacturing biogenerics and profit from those markets directly."

Writes Katznelson;

The first problem is that products made in China are arguably unlikely to be approvable in Western countries [by the FDA and EMEA].  'Standards in the US are so much higher that there's just no comparison'.
    Getting non-Western facilities up to production standards is a long and expensive process...  For example, requirements for manufacturing specific such as water purity may not be important clinically, but the FDA is likely to insist on them.

Yet Teva is evidently pursuing a long-term strategy, because while the FDA has for several years been issuing assurances that, "creating a regulatory pathway for biogenerics is a top priority...its reactionary approach has not inspired confidence in the generics industry of late;"

RA used to mean regulatory affairs, but now it means risk aversion," says Alan Liss, senior director of biotech at Duramed Research in Pennsylvania. "While we're talking, China and India will be supplying the rest of the world with products," he adds.

But the story concludes that this same sort of transition occurred in India, that Indian drug development and manufacturing efforts were "eventually folded in the Western regulatory framework," and that "in ten years, companies will use China not only as a local market but as a springboard for global opportunities -- providing talent and infrastructure at a good price."

Gong Yidong, writing in the 29 July issue of Science, notes the many motivations for operating in China;

China's growing appetite for Western drugs--the current $15 billion market is expected to quadruple by 2010, and then double again by 2020--has certainly caught the attention of every drug company. So has its cheap but skilled scientific labor force. Not only do Ph.D.s receive annual salaries of $10,000 or less, but the most expensive aspect of drug development--clinical trials--costs an estimated 30% less in China than in the United States or Europe. And then there is its growing prowess in science. "I'd say that setting up our own research lab there is only a matter of time," Novartis CEO Dan Vasella remarked this spring. "It's not so much a need as it is a hunger to take advantage of the opportunities."

With many companies opening R&D labs in country, combined with the increasing level of education and the desire to take a larger role on the world state, we're likely to see remarkable ferment developing new treatments in China.