Big Gene Patent (Busting) News???

Well now, isn't this an interesting development.  As covered by many news outlets (NYT, Wired, Genomeweb), US District Court Judge Robert Sweet has invalidated several US patents, sometimes referred to as the "BRCA1/2 patents", held by the University of Utah and Myriad Genetics.  From Judge Sweet's decision: "Products of nature do not constitute patentable subject matter absent achange that results in the creation of a fundamentally new product."  Judge Sweet's decision is here (PDF) via Genomics Law Report.  Here is the ACLU's take.

Here is a brief summary of what follows: The ruling is remarkable.  Various commentators and reporters remark upon it.  They get confused.  I try to clarify.  Then we get to a truly revolutionary part of the decision: it's about science!  And a little bit about law.  Finally: so what if a few patents are invalidated? 

Didn't See That Coming.  But I Can't Complain.

Last month, I noted that I was skeptical that the ACLU and other plaintiffs would be so successful in one go.  So I am surprised, but I am certainly not disappointed.  But I am not surprised, while being somewhat disappointed, that the coverage of the decision is so confused and confusing.  This confusion arises, I suspect, because the wording of Judge Sweet's decision is not entirely straightforward in places, and this has led to analyses that are insufficiently careful.  More on these points below.

DISCLAIMER: Please recall in what follows that I am but a humble physicist by training (oh yes, yes, we're all very humble), not a lawyer.  But I have written some stuff about patents on genes, and at least a few people (some of whom are IP law lawyers) think my analysis doesn't suck a lot.

First, over at Genomics Law Report (GLR), John Conley and Dan Vorhaus have a great analysis with a nice title: "Pigs Fly: Federal Court Invalidates Myriad's Patent Claims".  I won't bother to repeat their discussion.  If you are interested in this issue, please read that post as well as Dan Vorhaus' initial post analyzing the decision.  In particular, the reader might want to attend closely Vorhaus and Conley's observations about the potential for appeals, the likelihood of success in that endeavor, and the applicability of the ruling in other jurisdictions.

The short summary of what's transpired so far in the case is that Judge Sweet has invalidated a small number of claims, in a summary judgement ruling that so far applies only in the Southern District of New York.  Assertions that this is the end of the world for companies that hold gene patents are rather overblown.

There's Too Much Confusion, But Here is Some Relief

But now onto some of the confusing bits alluded to above.  The confusion starts, surprisingly, at GLR.  Here are Conely and Vorhaus:  "In the broader policy debate surrounding gene and biotechnology patents, however, this decision is the latest, unmistakable shot across the bow of gene patent holders, particularly those such as Myriad Genetics that have developed businesses around patent-protected genetic tests supported by exclusive rights in underlying gene patents."  Hummm...  Maybe not so much, actually.  Let me get straight to the point: there is a rather substantial difference between a "gene patent" that claims naturally occurring sequences and one that claims sequences that are not natural. 

Here is one way to think about the issues under discussion: in my one hand, I have a piece of isolated DNA that is identical in sequence to one in your body.  It is the same genetic sequence, so it carries the same information.  Indeed, for it to be useful in a test tube for the purposes of diagnosis, it must have both the same information content and the same function as the sequence in your body.  In fact, it only works as a diagnostic tool because it is the same as what is in your body.  As I noted in my earlier post, this is sort of the opposite of invention, and I have never understood why natural genes can be patented.  (Note: Judge Sweet hits this point quite squarely, but not until p.124 of his ruling.)  In my other hand, I have a piece of isolated DNA that is solely the result of human manipulation -- "human ingenuity" -- consisting of a sequence that does not exist in nature.  Both pieces of DNA are isolated, but they derive from very different sources, and are derived by very different means. Unfortunately, everybody discussing the present decision, including Judge Sweet in the early pages of his decision, seems to be a tad careless about the distinction, which leads many people down a rabbit hole.  (There is an extended discussion of the definition of "isolated DNA" and of the BRCA1/2 genes on p.90-92.)

Here is where it starts: Judge Sweet sets up his decision in the first couple of pages focusing specifically on the BRCA1/2 genes, and slightly more generally on isolated human genes: "Are isolated human genes and the comparison of their sequences patentable?" (p.2)  He continues: "Two complicated areas of science and law are involved: molecular biology and patent law.  The task is to seek the governing principles in each and to determine the essential elements of the claimed biological compositions and processes and their relationships to the laws of nature."

This sounds great.  Judge Sweet is clearly referring specifically to certain human gene sequences named in the patents in question.  Alas, on the next page he switches his language to address the specific assertions of the plaintiffs that ""isolated DNA" containing human BRCA1/2 sequences" are not patentable.  The basic contention here is that because the isolated DNA as described in the patents does the same thing inside the body as outside the body -- it is an information storage medium -- there is no difference between the two forms of DNA and therefore the isolated DNA in question cannot be patented.  Judge Sweet concludes (p.4):

DNA represents the physical embodiment of biological information, distinct in its essential characteristics from any other chemical found in nature. It is concluded that DNA's existence in an 'isolated' form alters neither this fundamental quality as it exists in the body not the information it encodes.  Therefore, the patents at issue directed to "isolated DNA" containing sequences found in nature are unsustainable as a matter of law and are deemed unpatentable subject matter.

The judge thereby switches within a couple of paragraphs very seamlessly from language referring only to human genes to language referring seemingly to all "isolated DNA".  It takes another 100 pages to get to a true clarification, and I'll bet very few people have read that far, or followed all the byways and cross-references (p.100): "...The issue presented by the instant motions with respect to the composition claims is whether or not claims directed to isolated DNA containing naturally-occurring human sequences [emph added] fall within the products nature exception.  ...It is concluded that the composition claims-in-suit are excepted."

In other words, Judge Sweet very specifically ruled that the claims on isolated DNA containing naturally occurring sequences are not valid.  Even more specifically, the ruling only applies to the motion in question by the plaintiffs, namely to invalidate the patents on BRCA1/2 held by Myriad et al.  Judge Sweet pointedly cites Diamond vs. Chakrabarty (p.109) -- a case that affirmed the patentability of "genetically engineered" organisms -- in limiting his ruling to the patentability of naturally occurring genes.  The ruling has no applicability outside that subject matter, and therefore has little applicability to, for example, much of anything that might come out of synthetic biology (unless you are talking about a synthetic DNA version of a naturally occurring gene).  Nor, for that matter, does the ruling have any say about any bit of DNA altered to be different from a natural sequence.  Which means that the ruling has very little to do with most patents on DNA, and therefore has very little to do with most of the industry surrounding those patents -- more on this below.

(Side note, as I read through the decision: Myriad's lawyers didn't do themselves any favors by making generally unpersuasive assertions aimed as broadside attacks against the plaintiffs' arguments.  As noted in my previous post on this case "Whither Genome Patents?", the defendants' assertions that patents serve as necessary incentives for scientific research are complete bunk.  Defense attorney Brian Poissant previously argued that "women would not even know they had BRCA gene if it weren't discovered" under a system that incentivizes patents.  I say again, as calmly as I can, bull pucky.  For example, see the publicly funded Human Genome Project.  See also the fact that BRCA2 was sequenced first in academic labs rather than by Myriad, who somehow managed to patent it anyway.  See also the many  BRCA1/2 assays independently developed in academia, the use of which Myriad repeatedly quashed through cease-and-desist letters, as recounted in detail in the decision.  But here is Judge Sweet himself (p.76): "According to Myriad, its policy and practice has been and still is to allow scientists to conduct research studies on BRCA 1 and BRCA 2 freely, the result of which has been the publication of [over 8600 papers] representing the work of over 18,000 scientists."  (It wasn't clear to me whether Myriad's legal team itself provided these numbers -- but if they did: bad legal tactics, fellas.)  In other words, 18,000 scientists have managed to produce a substantial body of work without any promise whatsoever of remuneration based on a patent for BRCA1/2.  Unless, of course, you count keeping your job through the promise of not being sued by Myriad.)  

It's Science!  And Science Always Wins -- Eventually, But May be Delayed By Appeals.

There is another very interesting angle to Judge Sweet's decision.  Andrew Pollack, writing in the New York Times, suggests that the most revolutionary part of the decision is where Judge Sweet recognizes that DNA carries information.  Pollack quotes Rebecca Eisenberg, a law professor at the University of Michigan: "There isn't a whole lot of doctrinal support" for considering DNA as information rather than as a chemical.  That, for me, is a truly eye opening perspective.  Not because I didn't know about it before -- unfortunately, that view is all too prevalent among IP lawyers -- but rather because it is being defended and suggested as a possible grounds for appeal.  True, it may be precedent, but that does not mean it is good precedent.

Here's the thing: There may not be much "doctrinal support" for considering DNA as information, but there is a rather overwhelming amount of scientific and technical support for considering DNA as information rather than as a chemical, say starting with the vast majority of molecular biology and biochemistry papers published in Science, Nature, Cell, PNAS, and any other relevant journal you can think of.  For all of the last six decades, no less.  Oh, and then all those silly textbooks.  The genetics and molecular biology ones, obviously; not the law textbooks.

Judge Sweet, in my humble opinion, already smacked this one out of the park on p.4: "The facts relating to molecular biology are fundamental to the patents at issue and to the conclusions reached.  Consequently, in the findings which follow, the discussion of molecular biology precedes the facts concerning the development, application, and description of the patents."  (Whoa there!  Science and reason trump the law of man!  Or science and reason trump the law of lawyers?  Damn, now that is a novel legal theory.  And a welcome one.  Don't tell Sen. James Inhofe.) 

Unfortunately, Pollack misses this angle, and promulgates further the confusion that Judge Sweet's ruling spells doom for the biotech industry: "Some biotechnology investors and executives say that lack of patent protection for DNA could diminish investment in the field and remove incentives for companies to develop tests."  Never mind that, as described above, Judge Sweet's ruling applies only to patents on naturally occurring genes, which should ameliorate the concerns of most of the "some biotechnology investors and executives".  It is nonetheless true that diagnostics companies that rely on patents claiming naturally occurring sequences may have to reevaluate their business plans.  (For instance, they may want to be especially careful in issuing cease-and-desist letters, lest the ACLU and company get busy again.)  And it may be true that this small fraction of biotech businesses may have difficulty raising capital -- but time will tell.  If it turns out that development of new diagnostic assays lags as a result of more patents on human genes being invalidated, then we will have something real to talk about.  We might consider developing public policy around alternate incentives.  Until there is a demonstrated concern, however, it isn't clear to me that we should be so concerned about the fate of private investors who gambled on patents whose validity has long been questioned.

What Is The Real Impact Going To Be? 

To reiterate the numbers from my earlier post: of the roughtly 2% of US GDP that is derived from biotech, at a rough guess I would put only 1% of the total (so .01% of US GDP) in the molecular diagnostics category that depends explicitly on excluding other uses of patented human genes.  A few billion dollars a year, in other words, might be at risk.  But somebody is going to do the tests, and Judge Sweet's decision lists a variety of tests that cost about 1/3 of Myriad's; that is, before Myriad shut them down with cease-and-desist letters.  If you eliminate those patents, we might have to come up with some other way to incentivize the development and testing of assays.  Prizes come to mind as a fine thing to try.  They work.  Academics and garagistas will be happy to compete for those prizes, I am sure.

But the rest of the biotech industry shouldn't be concerned about this ruling, frankly.  They might even celebrate the fact that they now have access, potentially, to a whole bunch more genes that are naturally occurring.  Not just in humans, mind you, but any organism.  This opens up a rather substantial toolbox for anybody interested in using biological technologies derived from viruses, bacteria, plants, etc.  If it holds up over the long run, Judge Sweet's decision should accelerate innovation.  That is definitely a good thing.

DIY Cleanroom

Gizmodo and Make are both pointing to Bill Morris's DIY Cleanroom.  Compare it to the hoods shown in my post on Garage Biology in Silicon Valley a couple of days ago.

Morris reports that his goal is a Class 10,000 hood, a specification that is slightly more involved than I had remembered.

In any event, Morris' hood would be of great use to those doing cell culture at home.  I suspect you are going to want a better filter, for nabbing smaller contaminants, maybe higher airflow, and perhaps some way to hack up a laminar-flow set-up.

Cool.

Whither Gene Patents?

Wired and GenomeWeb (subscription only) have a bit of reporting on arguments in a case that will probably substantially affect patents on genes.  The case is Association of Molecular Pathology , et al. v. US Patent and Trademark Office, otherwise known as "the BRCA1 case", which seeks to overturn a patent held by Myriad Genetics on a genetic sequence correlated with breast cancer.

Here is a brief summary of what follows: I have never understood how naturally occurring genes can be patentable, but at present patents are the only way to stake out a property right on genes that are hacked or, dare I say it, "engineered".  So until IP law is changed to allow some other form of protection on genes, patents are it.

The ACLU is requesting a summary judgment that the patent in question be overturned without a trial.  Success in that endeavor would have immediate and enormous effect on the biotech industry as a whole, and I doubt the ACLU is going to get that in one go.  (Here is the relevant recent ACLU press release.)

However, the lawsuit explicitly addresses the broader question of whether any patents should have been granted in the first place on human genes.  This gets at the important question of whether isolating and purifying a bit of natural DNA counts as an invention.  Myriad is arguing that moving DNA out of the human genome and into a plasmid vector counts as sufficient innovation.  This has been at the core of arguments supporting patents on naturally occurring genes for decades, and it has never made sense to me for several reasons.  First, changing the context of a naturally occurring substance does not constitute an invention -- purifying oxygen and putting it in a bottle would never be patentable.  US case law is very clear on this matter.  Second, moving the gene to a new context in a plasmid or putting into a cell line for expression and culturing doesn't change its function.  In fact, the whole point of the exercise would be to maintain the function of the gene for study, which is sort of the opposite of invention.  Nonetheless, Myriad wants to maintain its monopoly.  But their arguments just aren't that strong.

GenomeWeb reports that defense attorney Brian Poissant, argued that "'women would not even know they had BRCA gene if it weren't discovered'under a system that incentivizes patents."  This is, frankly, and with all due respect, a manifestly stupid argument.  Mr. Poissant is suggesting that all of science and technology would stop without the incentive of patents.  Given that most research doesn't result in a patent, and given that most patent application are rejected, Mr. Poissant's argument is on its face inconsistent with reality.  He might have tried to argue more narrowly that developing a working diagnostic assays requires a guarantee on investment through the possession of the monopoly granted by a patent.  But he didn't do that.  To be sure, the assertion that the particular gene under debate in this case would have gone undiscovered without patents is an untestable hypothesis.  But does Mr. Poissant really want the judge to believe that scientists around the world would have let investigation into that gene and disease lie fallow without the possibility of a patent?  As I suggested above, it just isn't a strong argument.  But we can grind it further into the dust.

Mr. Poissant also argued "that if a ruling were as broadly applied here as the ACLU would like then it could 'undermine the entire biotechnology sector.'"  This is, at best, an aggressive over generalization.  As I have described several times over the past couple of years (here and here, for starters), even drugs are only a small part of the revenues from genetically modified systems.  Without digging into the undoubtedly messy details, a quick troll of Google suggests that molecular diagnostics as a whole generate only $3-4 billion a year, and at a guess DNA tests are probably a good deal less than half of this.  But more importantly, of the nearly ~2% of US GDP (~$220-250 billion) presently derived from biological technologies, the vast majority are from drugs, plants, or bacteria that have been hacked with genes that themselves are hacked.  That is, both the genes and the host organisms have been altered in a way that is demonstrably dependent on human ingenuity.  What all this means is that only a relatively small fraction of "the entire biotechnology sector" is related to naturally occurring genes in the first place.   

I perused some of the court filings (via the Wired article), and the defense needs to up its game.  Perhaps they think the weight of precedent is on their side.  I would not be as confident as they are. 

But neither is the plaintiff putting its best foot forward.  Even though I like the analysis made comparing DNA patents to attempts to patent fresh fruit, it is unclear to me that the ACLU is being sufficiently careful with both its logic and its verbiage.  In the press release, ACLU attorey Chris Hansen is quoted as saying "Allowing patents on genetic material imposes real and severe limits on scientific research, learning and the free flow of information."  GenomeWeb further quotes the ACLU's Hansen as saying "Patenting human genes is like patenting e=mc2, blood, or air."

As described above, I agree that patenting naturally occurring genes doesn't make a lot of sense.  But we need some sort of property right as an incentive for innovators.  Why should I invest in developing a new biological technology, relying on DNA sequences that have never occurred in nature, if anybody can make off with the sequence (and revenues)?  As it happens, I am not a big fan of patents -- they cost too damn much.  At present, the patent we are pursuing at Biodesic is costing about ten times as much as the capital cost of developing the actual product.  Fees paid to lawyers account for 90% of that.  If it were realistically possible to engage the patent office without a lawyer, then the filing fees would be about the same as the capital cost of development, which seems much more reasonable to me.

I go into these issues at length in the book.  Unfortunately, without Congressional action, there doesn't seem to be much hope for improvement.  And, of course, the direction of any Congressional action will be dominated by large corporations and lawyers.  So much for the little guy.

Video from The Economist's World in 2010 Festival

The Economist has posted video from the World in 2010 Festival, held in Washington DC in early December.  The Innovation panel is below, with me (Biodesic), Dean Kamen (DEKA Research), Dwayne Spradlin (Innocentive), and Kai Huang (Guitar Hero), moderated by Mathew Bishop (The Economist).  (Here is a link to video selections from the rest of the event.)  I was chatting with a reporter a few days ago who observed that everyone else on the panel is quite wealthy -- hopefully that bodes well for me in 2010.  But maybe I am destined always to be the odd man out.  C-Span is re-running the video periodically on cable if you want to watch it on a bigger screen, but I can't seem to find an actual schedule.  (Here is their web version: Innovation in 2010.)

I have a couple of general thoughts about the event, colored by another meeting full of economists, bankers, and traders that I attended in the last week of December.  I met a number of fantastically accomplished and interesting people in just a few hours, many of whom I hope will remain lifelong friends. 

First, I have to extend my thanks to The Economist -- they have been very good to me over the last 10 years, beginning in 2000 by co-sponsoring (with Shell) the inaugural World in 2050 writing competition.  (Here is my essay from the competition (PDF).  It seems to be holding up pretty well, these 10 years later, save the part about building a heart.  But at least I wasn't the only one who got that wrong.)

Here is a paraphrased conversation over drinks between myself and Daniel Franklin, the Executive Editor of the newspaper.

Me:  I wanted to thank you for including me.  The Economist has been very kind to me over the past decade.
Franklin: Well, keep doing interesting things.
Me:  Umm, right.  (And then to myself: Shit, I have a lot of work to do.)

On to the World in 2010 Festival.  The professional economists and journalists present all seem to agree that we have seen the worst of the downturn, that the stimulus package clipped the bottom off of whatever we were falling into, and that employment gains going forward could be a long time in coming.  Unsurprisingly, the Democratic politicians and operatives who turned up crowed about the effects of the stimulus, while the Republicans who spoke poo-pooed any potential bright spots in, well, just about everything.

At the other meeting I attended, last week in Charleston, SC, one panel of 10 people, composed Federal reserve and private bankers, traders, and journalists couldn't agree on anything.  The recovery would be V shaped.  No, no, W shaped.  No, no, no, reverse square root shaped (which was the consensus at The World in 2010 Festival).  No, no, no, no, L shaped.  But even those who agreed on the shape did not agree on anything else, such as the availability of credit, employment, etc.

Basically, as far as I can tell, nobody has the slightest idea what the future of the US economy looks like.  And I certainly don't have anything to add to that.  Except, of course, that the future is biology.

Here is John Oliver's opening monologue from the Festival.  He was absolutely hilarious.  Unfortunately you can't hear the audience cracking up continuously.  I nearly pissed myself.  Several times.  (Maybe the cocktails earlier in the evening contributed to both reactions.)

Back to Innovation in 2010.  Dean Kamen had this nice bit in response to a question about whether the imperative to invent and innovate has increased in recent years (see 36:20 in the C-Span video): "7 billion people can't be recipients, they have to be part of the solution.  And that is going to require advanced technologies to be properly developed and properly deployed more rapidly than ever before."

To this I can only add that we are now seeing more power to innovate put into the hands of individuals than has ever occurred in the history of humanity.  Let's hope we don't screw up.

Can't believe it's finally here.

At long last, the printed copies of the book showed up yesterday.  Should be on shelves pretty soon.

A Spot of Tea at The Economist

While in London last month I sat down for tea and an interview with Geoff Carr at The Economist (permalink).  We covered a lot of ground, though as befitting a brief chat over tea we didn't get too deep in to the weeds.  Alas, they cut out the bit where the good Dr. Carr tried to put milk in my peppermint tea.  A topic for discussion in itself.

NYT on Systems Biology, Eric Schadt, and Sage Bionetworks

The Times is running a nice profile piece on Eric Schadt and his work at Rosetta and now Sage Bionetworks.

Biodesic evaluated systems biology investments for a large organization about 18 months ago, and Schadt's approach makes more sense to me -- by far -- than anything else we looked at.  I sat in on the pitch that Schadt and Stephen Friend made to that sameorganization, and it was crystal clear to me that Sage -- now residing at the Hutch here in Seattle -- should be on the receiving end of piles of money.  The stacks of Nature Group publications Schadt is accumulating suggest he is on to something, and it appears that his methods can be used to make predictions about the behaviors of complex networks.  Time and experimentation will tell, of course.  The open source aspect is a huge bonus.

Schadt's move to Pacific Biosciences is interesting because during his talk he suggested that genome sequencing provides enough information about variation to fuel his statistical methods for predicting interactions not just between genes but between tissues -- he is working at the level of describing the behavior of networks of networks.  It seems he will now have access to plenty of data.

Data and References for Longest Published sDNA

Various hard drive crashes have several times wiped out my records for the longest published synthetic DNA (sDNA).  I find that I once again need the list of references to finish off the edits for the book.  I will post them in the open here so that I, and everyone else, will always have access to them.

Year	Length	Refs
1979	207	Khorana (1979)
1990	2100	Mandecki (1990)
1995	2700	Stemmer (1995)
2002	7500	Cello (2002)
2004.4	14600	Tian (2004)
2004.7	32000	Kodumal (2004)
2008	583000	Gibson (2008)

1979
Total synthesis of a gene
HG Khorana
Science 16 February 1979:
Vol. 203. no. 4381, pp. 614 - 625
http://www.sciencemag.org/cgi/content/abstract/203/4381/614

1990
A totally synthetic plasmid for general cloning, gene expression and mutagenesis in Escherichia coli
Wlodek Mandecki, Mark A. Hayden, Mary Ann Shallcross and Elizabeth Stotland
Gene Volume 94, Issue 1, 28 September 1990, Pages 103-107
http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6T39-47GH99S-1J&_user=10&_rdoc=1&_fmt=&_orig=search&_sort=d&_docanchor=&view=c&_acct=C000050221&_version=1&_urlVersion=0&_userid=10&md5=84ca7779ff1489d5e18082b9ecb80683

1995
Single-step assembly of a gene and entire plasmid from large numbers of oligodeoxyribonucleotides
Willem P. C. Stemmer, Andreas Crameria, Kim D. Hab, Thomas M. Brennanb and Herbert L. Heynekerb
Gene Volume 164, Issue 1, 16 October 1995, Pages 49-53
http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6T39-3Y6HK7G-66&_user=10&_rdoc=1&_fmt=&_orig=search&_sort=d&_docanchor=&view=c&_acct=C000050221&_version=1&_urlVersion=0&_userid=10&md5=83620e335899881aac712a720396b8f2

2002
Chemical Synthesis of Poliovirus cDNA: Generation of Infectious Virus in the Absence of Natural Template
Jeronimo Cello, Aniko V. Paul, Eckard Wimmer
Science 9 August 2002: Vol. 297. no. 5583, pp. 1016 - 1018
http://www.sciencemag.org/cgi/content/abstract/1072266

2004
Accurate multiplex gene synthesis from programmable DNA microchips
Jingdong Tian, Hui Gong, Nijing Sheng, Xiaochuan Zhou, Erdogan Gulari, Xiaolian Gao & George Church
Nature 432, 1050-1054 (23 December 2004)
http://www.nature.com/nature/journal/v432/n7020/full/nature03151.html

Total synthesis of long DNA sequences: Synthesis of a contiguous 32-kb polyketide synthase gene cluster
Sarah J. Kodumal, Kedar G. Patel, Ralph Reid, Hugo G. Menzella, Mark Welch, and Daniel V. Santi
PNAS November 2, 2004 vol. 101 no. 44 15573-15578
http://www.pnas.org/content/101/44/15573.abstract

2008
Complete Chemical Synthesis, Assembly, and Cloning of a Mycoplasma genitalium Genome
Daniel G. Gibson, Gwynedd A. Benders, Cynthia Andrews-Pfannkoch, Evgeniya A. Denisova, Holly Baden-Tillson, Jayshree Zaveri, Timothy B. Stockwell, Anushka Brownley, David W. Thomas, Mikkel A. Algire, Chuck Merryman, Lei Young, Vladimir N. Noskov, John I. Glass, J. Craig Venter, Clyde A. Hutchison, III, Hamilton O. Smith
Science 29 February 2008: Vol. 319. no. 5867, pp. 1215 - 1220
http://www.sciencemag.org/cgi/content/abstract/1151721

Another Step Toward DIYStemCells

(18 June 2009: Lightly edited for clarity.)

The June 5 issue of Cell Stem Cells has a brief report describing the use of four proteins to reprogram human fibroblasts into induced pluripotent stem cells (iPSCs).  I think this is a pretty important paper, as it dispenses with any sort of genetic manipulation of the target cells or any use of plasmids to insert new "control circuitry", or any chemical manipulation whatsoever.

As expected, it is getting easier to produce iPSCs, and the authors of the paper ("Generation of Human Induced Pluripotent Stem Cells by Direct Delivery of Reprogramming Proteins") note that their work demonstrates the elimination of "the potential risks associated with the use of viruses, DNA transfection, and potentially harmful chemicals and in the future could potentially provide a safe source of patient-specific cells for regenerative medicine".

Kim et al used four recombinant human proteins to turn human newborn fibroblast cells (purchased from ATCC -- see the Supplemental Data) into iPSCs, where each of the proteins was fused to a nine amino acid long "cell-penetrating peptide" (CPP) that facilitated the importation of the proteins across the cell membrane.  The procedure was not particularly efficient, but after multiple treatments the authors produced cells that could differentiate into many different kinds of human tissues.

Here are a couple of thoughts about the paper.  Note that in what follows I have only had a few sips of my first cup of coffee today, and my brain is still quite fuzzy, but I think I am mostly coherent.  You can be the judge.

First, the authors did not use mature cells from adults, so don't expect this paper to lead to replacement organs and tissues tomorrow.  The use of cells from newborns makes a great deal of sense for a first go at getting protein-based reprogramming to work, as those cells have already been demonstrated to be relatively easy to reprogram.  The published procedure required many weeks of effort to produce iPSCs, and authors note that they have quite a ways to go before they can produce stem cells at the same efficiency as other techniques.

Nonetheless, it works.

Second, the paper describes PCR-based cloning of human genes to add the CPP sequences, along with a fair amount of bench manipulation to generate cells that made each of the four reprogramming proteins.  All the sequences for those proteins are online, as are the sequences for the CPPs, so generating the corresponding genes by synthesis rather than cloning would now cost less than $10K, with delivery in 2-4 weeks.  In another year, it will probably cost no more than $5K.  (How long will it be before these proteins show up in the Registry of Standard Biology Parts?)

Third, the authors did not use purified reprogramming proteins to generate iPSCs, but rather used whole cell extracts from cells that produced those proteins.  Thus the concentrations of the reprogramming proteins were limited to whatever was in the cell extract.  This might critically affect the efficiency of the reprogramming.  Presumably, the authors are already working on generating cultured cell lines to produced the reprogramming proteins in larger quantities.  But if you wanted to do it yourself, it looks like you might "simply" have to order the appropriate sequences from Blue Heron already cloned into the human expression plasmid pCDNA3.1/myc-His A, which is available from Invitrogen.  This would add a couple of hundred dollars to the cost because Blue Heron would have to play around with a proprietary plasmid instead of the public domain plasmids they usually use to ship genes.  You would then follow the recipe from the Supplementary Data to transform a protein production cell line to make those proteins.  Or perhaps you have a favorite recipe of your own.  Here is something I don't get -- it looks like that particular expression plasmid adds a His tag to the end of the gene, so I don't understand why Kim et al didn't try a purification step, but maybe that is underway.

Fourth, if you wanted to do this at home, you could.  You should expect to fail many times.  And then you should expect to fail some more.  And then, assuming your human cell culture technique is up to snuff, you should expect to eventually succeed.  You might want to wait until the inevitable paper showing how to do this with adult differentiated skin cells is published.

And then what?

You will have an autologous stem cell line that you can use to produce tissues that are, immunologically speaking, identical to those in your body.  What should you do with them?  I would suggest you show them off at cocktail parties, brag about them on Facebook, and then destroy them with bleach and an autoclave.  In lieu of an autoclave a microwave would probably do just fine.

But I expect that at least some of you will try to follow a recipe to generate some sort of human tissue, or even to simply inject those cells in your own bodies, which will result in all kinds of crazy teratomas and other tumors.  To quote Harold Ramus, "that would be bad".  So don't do that.  Just because DIYStemCells are cool doesn't mean you should actually use them yourself.  But I know some of you will anyway.  That is the future of biological technologies, for better or worse.

Stem_Cells@Home or DIYStemCells?

I'm in Cambridge, UK, and mostly on local time.  Mostly.  Spring is very pleasant here.

Here are a couple of interesting things that I've come across recently.

The FDA is considering regulating autologous stem cells as prescription drugs.  These cells are removed from a patient, multiplied in culture, and then reintroduced at a site of injury.  The culture step, reportedly, gets the FDA all in a lather with the desire for control.  According to the author of a story at h+ magazine, this could drastically slow down adoption and use, and potentially relegate the the technology to large corporate interests.  The story, and an accompanying interview with a physician, argues that self-regulation of stem cell treatments as a medical practice (which the FDA is not chartered to regulate) is a far better choice.

If the FDA does go the route of asserting (or, rather, attempting to assert) its might, it suggests to me that once again the powers that be are not sufficiently in tune with the progress of technology.  To wit: here is Attila Chordash's homebrew procedure from MAKE for isolating placental stem cells (I met Attila a few years ago at SciFoo and have participated with him in some IFTF activities -- smart fellow).  News this past year has been full of various ways to produce induced pluripotent stem (iPS) cells, ranging from retroviral reprogramming, to drug-controlled lentiviruses, to plasmid-mediated reprogramming. Skin cells were turned into iPSs early in 2008 (here is an earlier summary at Nature Reports Stem Cells).  Last November, a paper in PNAS showed a single synthetic prophage containing 4 genes was sufficient to turn a mouse fibroblast into an iPS cell, and showed that the method could be used to generate human iPS cells from human keratinocytes.  Each of these steps is said to demonstrate an increase the controllability of the reprogramming, increase the uniformity of the resulting population of cells, and decrease the difficulty.

This is not to say that any step in the reprogramming is simple.  From personal experience I can testify that culturing even "stable" human cell lines can be challenging at times.  But, by definition, as published methods to reprogram cells are repeated and refined this will demonstrate a progression from iPS cell production as an art into a technology.  The plasmid-mediated programming, in particular, strikes me as a promising route to a widespread technology because it does not depend upon, or result in, integration of the plasmid into the host chromosome.  Moreover, it will be trivial to synthesize new genes for use in the plasmid as better recipes come along.  So how long before these cells will be used in therapies?

A recent review in Science by Gurdon and Melton identifies some interesting challenges:

The future value of reprogrammed cells is of two kinds. One is to create long-lasting cell lines from patients with genetic diseases, in order to test potentially useful drugs or other treatments. The other is to provide replacement cells for patients. To be therapeutically beneficial, replacement cells will probably need (i) to be provided in sufficient numbers; (ii) to carry out their function, even though they are not normally integrated into host tissues; and (iii) to be able to produce the correct amount of their product.

A human adult has about 10¹⁵ cells, and the liver contains about 10¹⁴ cells. To create this number of cells starting from a 10^-4 success rate of deriving iPS cells from skin would require an enormous number of cell divisions in culture, although the prolonged culture of ES-like cells provides a valuable amplification step. However, many parts of the human body need a far smaller number of cells to improve function. An example is the human eye retina, in which only 10⁵ cells could be of therapeutic benefit.

Will introduced cells be useful even if not "properly" integrated into the host? Most organs consist of a complex arrangement of several different cell types. The pancreas, for example, contains exocrine (acinar) cells, ductal cells, and at least four kinds of hormone-secreting cells in the endocrine islet. Replacement endocrine cells can provide useful therapeutic benefit even if not incorporated into the normal complex pancreas cell configuration. In some cases, introduced cells can have functionally beneficial effects, even if indirectly. It is not yet clear whether introduced cells will be correctly regulated to produce the desired amount of product.

There is obviously a great deal of science to do before iPS cells are used on a regular basis to produce therapies. Nonetheless, therapy is already beginning around the world.  Medical tourism to China for stem cell treatments is increasingly common, even for children.

Clearly, the technology is so promising that families are willing to go to considerable sacrifice to obtain treatment.  Which brings us back to the FDA and regulation.  I have to wonder what the Feds are thinking.  I would certainly agree with anyone who suggests that stem cells are a powerful technology, and that treatments should be safe.  But any regulatory or policy step that reduces access and slows progress in the US is simply going to send people overseas for treatment.  Then, as the technology becomes ever simpler to learn and use, a back-room market will open up in the States.  
 
So, I wonder, as the technology matures, how long before we get DIYStemCells, Stem_Cells@Home, or HomebrewStemCells?  As methods are published to harvest candidate cells and turn them into autologous iPS cells, how long will it be before athletes looking for an edge, the curious, and the truly ill, all start trying this for themselves?  I am by no means arguing that this is a good idea, and I strongly suspect that the better course is to ensure that people have access to the technology through physicians who know what they are doing.  But without that access, a black market, with all of the shadows and horrors envisioned by William Gibson and others, is inevitable.

Wouldn't it be simpler, and vastly safer, to make sure that everyone has access to skills and materials?  This seems like another arena in which pushing for an Open Biology makes a great deal more sense than the alternative.