ALS Target: Microglia

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deboer1HRGlia, perhaps the most underappreciated of cell types, are finally getting some attention. A new report in Science Translational Medicine from Kevin Eggan’s group at the Harvard Stem Cell Institute validates the role of microglia in amyotrophic lateral sclerosis – aka ALS, motor neuron disease or Lou Gehrig’s disease — and investigates how to manipulate them to extend life, in a mouse model for now.

Often described as inert scaffolding, bystanders, or background cells, neuroglial cells – glia for short– actually do quite a lot. In the central nervous system, the several types -– astrocytes, oligodendrocytes, ependymal cells, and microglia — provide nutrients and growth factors, mop up excess neurotransmitters from synapses, form the intricate architecture that supports axons, provide insulating myelin, and control the communications of neurons.

The relative numbers of neurons and glia have long been a matter of debate, but consensus seems to be emerging of a near 1:1 ratio. And because neurons don’t divide and glia do, there’s a lot of life and death among the so-called bystanders. Not surprisingly, when brain tumors form, they’re typically glial. Neurons must revert to a more plastic developmental state to veer onto a pathway towards cancer.

When I began coauthoring a human anatomy and physiology textbook years ago, one of the first things I did was to redo the glia section, freeing them from their stereotyped supportive roles and giving them proper respect.

All specialized cells derive from pathways that branch from stem cells. Most glia are cousins of sorts to neurons, descending from division of the same neural progenitor cells. Yet some microglia come from hematopoietic stem cells, the “mother” cells of the bone marrow. Brain cells coming from the bone marrow makes it possible to treat some brain diseases with bone marrow transplant, such as adrenoleukodystrophy. lists a few such efforts for ALS.

Microglia mediate signaling of prostanoids, which include the prostaglandins, hormone-like fatty acids that control inflammation, among other activities. In the brain microglia aggregate near damage, but in ALS, overactive versions of the cells might contribute to the pathology. PET scans show supercharged microglia in the brains of ALS patients, and heightened signaling in their spinal fluid.

Dr. Eggan and colleagues called attention to the possible role of microglia in ALS in 2008. “Now 6 years later, after considerable effort and many long-term experiments, we’ve been able to better pinpoint the source of those signals in the nervous system in an animal model, as well as show that the prediction we made using a stem cell model of disease can hold up to closer scrutiny in the context of a whole animal,” Dr. Eggan said in a news conference yesterday. The research trajectory is in a sense circuitous, demonstrating mechanisms in human cells that are then repeated in mice, to collect evidence to catalyze clinical trials of compounds that intervene in the pathological process.


Human motor neuron from iPS cells from ALS patient (Gist Croft + MacKenzie Weygandt)

Human motor neuron from iPS cells from ALS patient (Gist Croft + MacKenzie Weygandt)

Using human embryonic stem (hES) cells or induced pluripotent stem (iPS) cells is particularly important in studying diseases of the nervous system because neurons don’t divide. Otherwise where would new ones come from in a culture dish?

Contrary to the popular but deficient definition of stem cells as “cells that can turn into any cell type,” the defining characteristic is ability to self-renew – make another stem cell. If stem cells magically “turned into any cell type in the body,” there would quickly be nothing left to keep things going.

A stem cell can self-renew and generate a neural progenitor cell that in turn can divide to give rise to neurons and glia. The Eggan team first created iPS cells from an ALS patient  in 2008.

Microglia apparently harm motor neurons via a specific type of overactive prostanoid receptor. First author and grad student Sophie De Boer and her colleagues at HSCI, Massachusetts General Hospital, and Boston Children’s Hospital, conducted a brilliant series of experiments that show that blocking or removing the errant receptors may be one route to extending survival in ALS.


Human embryonic stem cells

Human embryonic stem cells

In the new study, the investigators exposed human motor neurons derived from ES cells and marked with green fluorescent protein (GFP) to sheets of “toxic” glia from mice that have a form of ALS due to mutation in the superoxide dismutase 1 (SOD1) gene. At last count, 7 human genes had been implicated in the 5 to 10% of cases that are inherited; SOD1 was the first and is the best studied.

The SOD1 toxic microglia killed more than half of the neurons, but chemically blocking the receptors enabled the neurons to survive.

In complementary experiments, activating receptors in normal glia turned them toxic – but only to motor neurons, and not other neuron types. Removing the gene encoding the protein that forms the receptor had the same effect as blocking it chemically. And perhaps most important in a translational medicine sense, a short blast of antagonist had a long-lasting effect on the health of the motor neurons. Plus the effect is seen in actual living animals, increasing lifespan in mice.

The most exciting part of the new work is that it zeroes in on a “druggable” target – the DP1 receptor and its associated signalling pathways. The receptor is a G protein, a membrane-spanning molecule that is already the basis of many drugs. Said Dr. Eggan, “At least two major pharmaceutical companies have significant development programs around this receptor for another indication — niacin induced flushing.” Perhaps these candidate drugs can be retasked to inhibit inflammatory effects in ALS, he added, and possibly teamed with a seizure medication the researchers identified earlier this year that fights ALS by a different route.

640px-2006_Pro_Bowl_tackleALS is like Alzheimer and Parkinson diseases in that a shared phenotype might represent any number of different gene-environment interactions. What’s true for an individual with SOD1 ALS may not be so for a person with a different mutant gene.

The ALS patient I posted about in April, Glenn, didn’t have any known mutations, but he did have a few environmental risk factors. He played football for many years, maybe getting clunked in the head one too many times, and may have been exposed to pesticides when his boyhood home (to which he moved back as an adult) bordered fields of crops. Smoking, aspartame, and exposure to formaldehyde and heavy metals are other suspects. Reports that vitamin E protects against ALS haven’t held up.

BMAA, an amino acid implicated in ALS that is not part of dietary protein.

BMAA, an amino acid implicated in ALS that is not part of dietary protein.

Perhaps the most fascinating causal candidate is exposure to cyanobacteria (aka blue-green algae) living in cycad trees on Guam. The cyanobacteria release an amino acid called BMAA into the soil, where it makes its way through the roots to the seeds of the cycads. After biomagnification through food webs, BMAA causes an ALS-like disease when it binds proteins in the brains of some individuals, who may be genetically-predisposed to the motor neuron damage.


Do people survive ALS? The disease may be so heterogeneous that this might be possible. One commentator on my previous ALS post took me to task for not mentioning this idea, which I admit I was not aware of, but I found his tone disturbing. He wrote:

“C’mon, time to wake up and smell the coffee. People have been solving ALS for years, and the only way to really do it is change diet, psychology and lifestyle.”

He linked to a website offering accounts of recoveries, and then commented on Dr. Eggan’s team’s identification of the seizure drug for possible repurposing:

“While maybe that drug will be of some use somehow, don’t let that distract you from the overall reality of the situation. Stem cells? Eh, just learn to heal.”

There's more to finding a treatment for ALS than just "making it so."

There’s more to finding a treatment for ALS than just “making it so.”

I don’t think the commentator intended to upset ALS patients who haven’t managed to heal themselves, and I understand the value of hope. But an anti-science stance certainly isn’t going to help. I’m reminded of Captain Picard on Star Trek: Next Generation saying “make it so,” or the professor in The Music Man teaching kids to play instruments using the “think” system rather than learning music theory.

Positive thoughts aren’t enough.

Using stem cells to recapitulate the precise choreography of a disease, illuminating the various drug targets and their interactions, and then deploying drugs old and new based on those discoveries, may indeed “make it so,” slowing, vanquishing, and ultimately preventing this heartbreaking and complex disease.

I think that use of stem cells to create “diseases-in-a-dish” will turn out to be their most successful application. The very quality of self-renewal that many news reports omit is what scares me. Stem cells used therapeutically that self-renew, perhaps in places unexpected, could form a tumor.

Coincidentally, I just received an e-mail from Maurie Hill, whose participation in a clinical trial to treat Stargardt disease, a form of visual loss, was the subject of the very first DNA Science post, Human Embryonic Stem Cells Finally Reach Clinical Trials: Maurie’s Story, nearly two years ago. Alas, her vision has not improved at all, although I do not know the findings of the frequent tests she undergoes — just that she’s very disappointed.

But use of stem cells wedded to drug discovery? That’s a can’t-miss! Congrats Eggan lab for the continuing fine work.



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Gene Test Predicts Blindness After LASIK

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Granular_corneal_dystrophy_type_I._Numerous_irregular_shaped_discrete_crumb-like_corneal_opacitiesMillions of people have put aside their eyeglasses or contact lenses thanks to a procedure called LASIK. But for carriers of a rare condition called granular corneal dystrophy (GCD), LASIK can damage the cornea, even causing blindness. The Avellino DNA Dual Test for LASIK Safety, from Avellino Laboratory, can identify individuals genetically predisposed to this complication, offering a great example of an “actionable” genetic test.

Ophthalmologists can provide the cheek swab test to patients considering LASIK, and results are available within 2 days. Company founder and CEO Gene Lee pursued development of the test after learning about the genetic connection in early 2008. It’s now standard-of-care in Korea and Japan, and became available in the US in April. Insurance typically doesn’t cover it (yet), but the test is often incorporated into a work-up for LASIK, and out-of-pocket costs about $100.

“We started doing it last year and we’ve found it to give increased peace of mind and confidence in the procedure. If someone has this condition and it is clinically apparent and visible, the test would just confirm the visual findings. But a number of patients have subclinical findings or none, and this genetic test is the only way to identify the condition,” Richard Rothman, MD, an ophthalmologist who practices in Las Vegas told me. And patients who have the mutation can tell their relatives to be tested before choosing LASIK.




GCD results from mutations in the transforming growth factor beta induced (TGFΒI) gene, which encodes the protein keratoepithelin. The affected part of the cornea, the stroma, consists of extracellular matrix (the goo between cells) and stacks of collagen fibrils, with some other proteins such as fibronectin and integrins, and scant keratocytes that produce the keratoepithelin that keeps the cornea clear. Combine carrying a mutation with stress – such as radiation, hypoxia, chemotherapy, peroxide, or perhaps a laser procedure — and keratoepithelin misfolds into a gunky, amyloid-like mess.

The two types of GCD are due to mutations in different parts of the gene. The type 1 or classic presentation results from a mutation in exon 12, giving a “bread crumb” like appearance to the cornea. Type 2 is known as “Avellino” because the first identified cases, in 1988, lived in the town by that name in Italy. This type resembles a “snowflake icicle” due to a mutation in exon 4 (the exons are the parts of genes that encode protein). Other studies identified patients in Japan and Korea, and then pretty much everywhere.

For individuals who have two mutations (homozygotes), proteins deposit in the corneas during infancy, causing blindness by the teens. The condition is much milder, with later onset, in people with one mutation (heterozygotes). But if vision isn’t very impaired, or begins late in life, then visual loss rather than improvement months to years after LASIK comes as quite a shock. People can carry the mutation and not know it. And that’s where the test comes in.




LASIK (laser-assisted in situ keratomileusis) creates a thin flap in the cornea that is hinged back to reveal the middle layer, where an excimer laser is applied to alter the topography to better focus light rays on the retina. Normally, the intervention activates transforming growth factor beta to repair the wound. But in people with one mutation, the surgery makes keratoepithelin misfold and aggregate at the flap. This action accelerates the corneal dystrophy and may cause another complication in which torn collagen fibers bind various proteins, producing a “sands of Sahara“ effect. Ouch.

Avellino DNA Dual TestGCD is rare. Avellino Laboratory has identified 390 people with one or two mutations among 420,000 being worked up for possible LASIK. More than 30 mutations are known, and they also account for the related condition lattice corneal dystrophy. The nomenclature is still traditionally clinical, based on appearance, but subtyping now reflects genetic distinctions. For families aware of the pattern of inheritance because homozygotes are blind, full gene sequencing is available from several labs.

Every morning, I look through digests of news releases about genetic research from a wonderful service from the American Association for the Advancement of Science called EurekAlert. A few weeks ago, the news release about the LASIK genetic test caught my eye because of its simplicity, its utility, and the fact that it looks for mutations in ONE gene. That’s unusual these days.

(Dept. of Energy)

(Dept. of Energy)

More often, the studies in the news release roster, and therefore those that make the news, deal with big data. They track thousands upon thousands of sites in genomes that vary among individuals, boiling down to a dozen or so that can serve as usually weak risk factors (genome-wide association studies or GWAS); follow epigenetics (sites of methylation) and changes in gene expression; or sequence exomes and genomes. All good of course, but usually not immediately of any help to patients. And many news releases mix up the techniques or write so vaguely that it’s difficult to know what exactly the findings are without reading the technical papers.

Around the time of the LASIK news release was another on a GWAS finding a dozen risk sites for Parkinson’s disease, a report in the Journal of Urology tracking methylation profiles of three genes that predict which prostate cancer biopsies are false negatives, and reports on finding common gene variants behind autism and schizophrenia. Inheriting a mix of variants can set the stage for each condition. An additional, triggering mutation or exposure to an environmental factor might then turn risk into reality.

These are all quite different genetic scenarios than a single gene mutation that makes itself known after surgery injures the eye.

I realize that large-scale investigations take massive and talented teamwork, and that the projects are vital to understanding pathology, which fuels new treatments. But I’m starting to feel an information overload that makes me appreciate the single-gene tests that a patient can benefit from right now. Many have been available for years, but we don’t hear about them as often as the megadata. They’re not news.

Blausen_0290_DeepVeinThrombosisPerhaps the best example of a an actionable single-gene test is the one for factor V Leiden, a mutation in a clotting factor gene. Knowing it’s there means you can avoid dangerous clots by taking blood thinners, wearing special support socks, and avoiding long plane rides or crouching for long periods, as reporter David Bloom did in 2003 when reporting from Iraq in a tank with his legs folded underneath him for hours. He died of a pulmonary embolism following deep vein thrombosis.

I hope that single-gene test panels, such as those for cardiovascular disease or Jewish genetic diseases, don’t become completely buried by the avalanche of genomic megadata.

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Vaccine Memories: From Polio to Autism

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Salk_Thank_You“April 15 – Polio Vaccine Perfected!!!!”

So wrote my mother in 1955, on the “Baby’s Health Record” page of my baby book. I unearthed it a few days ago while looking for some old writing clips.

Just a day earlier, I’d just reported in Medscape on a study finding that adding injected Salk polio vaccine to the oral Sabin vaccine series can boost immunity in parts of the world where polio is still endemic, such as Nigeria, Pakistan, and Afghanistan.

The coincidences continued.


Herd immunity protects a population against an infectious disease when enough individuals are vaccinated to block spread of the pathogen.

Herd immunity protects a population against an infectious disease when enough individuals are vaccinated to block spread of the pathogen.

Among my old article clips I soon found “Polio Eradication  Goal Extended,” in The Scientist from 2000, which had been the target year for ending polio. I’d quoted U.S. Secretary of Health and Human Services Donna Shalala defining herd immunity on a global scale: “No nation is truly free from polio unless every nation is free of polio.”

It’s a message that needs repeating, for a number of infectious diseases.

My Scientist article began with a glimpse of the pre-polio-vaccine days:

An iron lung took over for the flaccid chest muscles of people with polio.

An iron lung took over for the flaccid chest muscles of people with polio.

For parents of young children in the early 1950s, summertime brought the terror of a fever that might explode into ‘infantile paralysis,’ a fear propelled by images of Franklin D. Roosevelt’s battle with the disease. Although only 1 percent of infected individuals developed severe symptoms as the virus invaded spinal cord cells, the number of cases grew large enough to inspire a massive effort to develop a vaccine, including the founding of the March of Dimes in 1938 to specifically battle polio.

For those old enough to remember the Beatles, polio vaccines were part of childhood. By 1955, youngsters in many nations received injections of Jonas Salk’s inactivated polio vaccine. By 1962, children were lining up at school, tongues out to receive pink-stained lumps of sugar impregnated with Albert Sabin’s live, attenuated oral polio vaccine.”

Neither Salk nor Sabin sought patent protection for their inventions, which offer the best protection when teamed. The first vaccine, live but weakened, was actually invented in 1948 by virologist Hilary Koprowski, director of the Wistar Institute in Philadelphia for 30 years. He died last year.

Kathleen Sebelius gives oral polio vaccine to a child in India.

Kathleen Sebelius gives oral polio vaccine to a child in India.

Sabin’s oral vaccine, much more acceptable to the kindergarten set than a shot in the arm, sent attenuated virus directly into the gastrointestinal tract. To develop the oral vaccine, passage through several non-human animal hosts weakened the virus so that it activated immunity in people without causing disease.

Stool from kids who ate the sugar cubes released the weakened poliovirus into sewage, spreading protection even to those who hadn’t been vaccinated, like older people. But the virus can mutate, as live viruses do, sometimes into a form that invades nerve cells. Very very rarely, this causes the acute flaccid paralysis of poliomyelitis.

My sister Edith had the good sense to be born after me, and even though that meant she was a mere penciled-in footnote in my baby book, she had some vaccines for which I suffered through the associated illnesses. We straddled that time when the classic illnesses of childhood began to go away.

When I was 3 I had the measles for a full month. During a house call, the pediatrician prepared my parents for the chance that I might need to be hospitalized, and possibly lose my hearing or end up brain damaged. Fortunately I recovered.

A few years later I watched several people hold down a screaming Edith as she got the new measles vaccine, 2 shots back then. But we both had mumps, chickenpox, and German measles, their vaccines not yet perfected. She reports recalling her delight at expecting a shot and getting a sugar cube.

Because smallpox vaccine was initially derived from cowpox lesions, people feared the vaccine would cause tiny bovines to emerge from the skin. This famous cartoon is from 1802.

Because smallpox vaccine was initially derived from cowpox lesions, people feared the vaccine would cause tiny bovines to emerge from the skin. This famous cartoon is from 1802.

According to my baby book, my smallpox vaccine of April 5, 1955, didn’t take, so I had it again on May 6. Now we don’t hear much about smallpox unless it’s discovered in a back room at a government lab.

May 6, 1955 must have been a bad day for me (and my mother), because I had my first whooping cough (pertussis) vaccine then too, and again a month later. Edith had to have pertussis vaccine again just last week because she’s visiting her about–to-be-born first grandchild in California, where the disease has returned due both to natural waning of the immune response and to people who won’t vaccinate their children. Pertussis can kill a baby.

During my first year I also had 4 diphtheria shots and 1 for tetanus, the DPT vaccine, now called Tdap, yet to have been combined into existence. It is the main target of the anti-vaccine movement.

Thanks, mom, for getting me vaccinated.

Thanks, mom, for getting me vaccinated.

I had my polio vaccine shots in May, June, and October of 1956. Being a toddler, I didn’t know that the vaccine had run into trouble right out of the gate. Thirteen days after its debut in April 1955, the product from Cutter Labs in Berkeley, California, was found to harbor live virus, which infected 94 vaccinated kids and 166 of their close contacts. On May 7, the Surgeon General suspended the polio vaccine program, spearheading government vaccine surveillance. Polio vaccinations resumed in the fall of 1955, much to the relief of millions of parents.

My mother, back in 1955, understood the concept of herd immunity, if not by that name. Grateful and relieved parents throughout the U.S. realized that if enough people were vaccinated, active virus would have nowhere to infect, and polio would go away.

An alternate possible explanation for the rise in autism cases: extended use of folic acid in pregnancy.

An alternate possible explanation for the rise in autism cases: extended use of folic acid in pregnancy.

I hated vaccines because they hurt, but even as a small child wondered how they worked. And because some of my earliest memories are of those first vaccines, I signed on as this year’s keynote speaker for the  March of Dimes student convocation when I learned that the first speaker, in 1971, was Jonas Salk.

At the last talk in the series, at the Institute for Basic Research in Staten Island, Mohammed Junaid, PhD, head of the structural neurobiology lab there, talked to the high school students just before I did. His team’s data may explain the increase in incidence of autism: folic acid supplementation extending well beyond day 28 of gestation, when it prevents neural tube defects (NTDs). So far they’ve got an association (timing) and a possible mechanism (epigenetics). The vaccine connection, of course, is the blame for the uptick in autism cases.

Folic acid

Folic acid

In 1992, the CDC recommended that pregnant women get .4 mg of folic acid daily to prevent NTDs, and in 1998, the US government mandated adding folic acid to grains and cereals to help them do so. (The folate form in foods is poorly absorbed.) But getting this much folic acid from even fortified foods isn’t that easy, so in 2003, clinical management guidelines  from the American College of Obstetricians and Gynecologists called for folic acid in pill form for pregnant women.

Through the 1990s, recommendations stressed folic acid during the perinatal period – a month before conception and 2 to 3 months after. But somehow, perhaps following a more-is-better philosophy, the time to take folic acid supplements during pregnancy stretched. Some women even continue to take the leftovers after giving birth, perhaps reassured by studies countering concerns that the vitamin might cause heart problems, twinning, allergies, and mask B12 deficiency.

Advised WebMD: “The CDC recommends that you start taking folic acid every day for at least a month before you become pregnant, and every day while you are pregnant. However, the CDC also recommends that all women of childbearing age take folic acid every day. So you’d be fine to start taking it even earlier.”

But is it “fine” to continue taking a high dose of folic acid beyond the point at which the neural tube closes?

Might exposure during the second and third trimesters, when brain development accelerates, lead to more subtle symptoms than the open neural tube that early exposure prevents? Like behavioral ones? It makes sense, because folic acid begins the pathway that adds methyl groups to certain genes, silencing their expression – a great example of an epigenetic effect.

Autism enters the picture rather circuitously.

A widely-reported 2013 study presented evidence that folic acid taken from 4 weeks before to 8 weeks after giving birth lowers incidence of autism. Less widely-reported was a study from two years earlier, from Dr. Junaid and his colleagues, who used white blood cells and animal models to look at what happens later in gestation, mimicking the situation of women who continue to take higher doses of the vitamin throughout pregnancy.

Methylation silences genes.

Methylation silences genes.

Indeed, the experiments showed that exposure to high doses of folic acid past the first trimester causes overexpression of a whole bunch of genes. The list includes XIST, the site that controls methylation (silencing) of one entire X chromosome. This might explain why boys with autism outnumber girls. Males are more vulnerable to the effects of both mutation and epigenetic changes because they only have one X chromosome, and are in this genetic sense the weaker sex. Also underexpressed in the presence of folic acid is the FMR1 gene, which lies behind fragile X syndrome, a common cause of autism.

I’d scribbled notes at Dr. Junaid’s talk in April and read the group’s papers, waiting to blog about the work until the results he discussed were published. That happened, also coincidentally, on the day that I found my baby book nestled among my tattered writing clips.

Dr. Junaid’s group reported in PLoS One last week that prolonged prenatal exposure to folic acid in mice is associated with anxiety, hyperactivity, and increased vocalization, but with no effects on learning memory, or social behavior. Male mice were more likely to respond to the excess than female mice. And they identified nine key genes with altered expression. Not exactly autism, but distinct brain changes nonetheless.

The team is conservative in their conclusion that there may be a “loss of benefit” to prolonged folic acid supplementation, writing: “Unregulated high FA supplementation during pregnancy and throughout the life course may have lasting effects, with alterations in brain development resulting in changes in behavior.”

The increase in number of autism cases parallels folic acid use during pregnancy. Coincidence?

Clearly much more work needs to be done on the link, if any, between extended folic acid supplementation and autism. But IMHO this is the very best type of scientific investigation.

It isn’t a “cure” or a “breakthrough” or even “proof,” which are to me the “F” words of reporting on science. The link isn’t coming from celebrities who speak as if they’ve anointed themselves with PhDs in immunology. The work of Dr. Junaid and his colleagues presents a compelling idea, providing enough information to seed a new hypothesis: Can overexposure to folic acid in the second and third trimesters of pregnancy increase risk of developing autism?

That’s what science is: framing new questions.

PikiWiki_Israel_10876_Alyn_Day_CampSo stay tuned. We aren’t nearly done “rounding up the suspects” in causing autism, but meanwhile vaccine phobia is opening the door to the diseases I had as a child, and some much worse.

Do we really want to return to those long-ago summers when parents feared their kids would spike fevers and become paralyzed in a matter of days?

(Thanks to Wikimedia for images and to my mother for my baby book — and getting me vaccinated.)

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The Menopause Toolkit and Summer Sci-Fi

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eggsOne of the worst feelings a woman of a certain age can experience is when someone stupidly asks, “When is your baby due?” and points to the bulge that is not in fact a baby bump, but padding that appears once one’s ovaries have run out of eggs.

And so when researchers from Monash University in Australia published this week, in the journal Climacteric, a “practitioner’s toolkit” to help primary care providers recognize and possibly treat symptoms of menopause, I was intrigued, and then thrilled to discover the phrase “central abdominal fat deposition (even in slim women).” Guilt over the intractable status of my middle began to fade.

I wrote about the menopause toolkit for Medscape, quoting one ob/gyn who loved it, another who thought it stated the obvious. News aggregators parroted the press release, a few dropping in a “breakthrough” or two, as if a woman in the throes of menopause doesn’t know it.

Heraldic_hourglass.svgFiona Jane, MD and Susan Davis, MD and their colleagues created the toolkit, which is more a labyrinth of linked flowcharts than something you’d use to change a tire or clean teeth. The investigators tapped their clinical experience, published diagnostic algorithms, various position statements, and the peer-reviewed literature. The information-packed toolkit is also available in stand-alone form. It targets women over 40, and is so detailed that the International Menopause Society has endorsed it for global use.

I learned a few things.

In addition to discovering that my bulge is due to putting up with low estrogen instead of replacing it (I’m a DES daughter of a mom who had breast cancer, so no thanks, plus some estrogen products come from horse pee), I became aware from the toolkit that “formication” means “crawling sensations on skin.” A biologist might not mind that one so much.

I learned that “dyspareunia” is painful intercourse, which I would have known about had I paid more attention to those Osphena ads on TV. (My husband is in love with the willowy Osphena lady).

I also learned from the menopause toolkit that in addition to hormones served up in all sorts of contraptions and dosages, women take antidepressants, clonidine, gabapentin, eye of newt, and undergo hypnosis and cognitive behavior therapy and something called stellate ganglion blockade to temper what the docs call “vasomotor symptoms” or “hot flushes,” but the rest of us call “hot flashes.”

Having read the flowcharts several times for Medscape, I’d like to make a few recommendations for extending the menopause toolkit.

ISHERB_Caveman1. Some entries require more information than fits in the tiny boxes of the flowchart. Specifically, “no interest in sex” as a symptom may simply reflect the fact that a woman’s partner is an australopithecine. Or she’s exhausted. Or she refuses to take hormones to reinstate someone else’s idea of what should be her sex drive. “Poor sleep?” Please.

2. A few entries could use more graphic descriptions. “Dyspareunia” might be more recognizable as “agony when a giant pole is repeatedly shoved up a desiccated orifice.”

Firetora3. “Hot flush” sounds like a function of a bidet. How about “awakening stark naked drenched in a pool of sweat?”

4. The toolkit mentions “overall diminished well-being” and depression, anxiety, and irritability. Might these so-called symptoms instead be reactions to such external stressors as an adult child returning home? Costly medical care? Pressure at work? Being ignored or taken for granted?

5. Despite the above, I suggest adding “sudden cessation of higher brain functions, particularly recall, reasoning and concentration” as a description of the well-known “menopause brain.”

Water_drops_by_Ximeg_24.12.12-046. The toolkit seems to ignore the sudden loss of bladder control that is a hallmark of menopause, unless it is subsumed under the rather vague “localized urogenital symptoms.” The sounds of a dripping faucet, waves crashing ashore, or rain can all precipitate an event, as can unanticipated laughter. (See Poise pads)

7. Add the vivid dreams, such as iridescent eels writhing out of all body openings.

8. I’ll let my husband Larry have the final word: Another indication of menopause is a partner joining the witness protection program.

PiolinmarcianocolorP.S.: MY TAKE ON SUMMER SCI-FI ON TV: The Leftovers and Under the Dome are explorations of religion with science a mere backdrop, but Extant hit it out of the park.

The Leftovers is the HBO adaptation of Tom Perrotta’s novel that explores the aftermath of a Rapture-like phenomenon that suddenly whisks away 2 percent or so of global humanity. Under the Dome is the second season of the TV version of the Stephen King book, and this week’s episode concluded with a dramatic if anti-science “We are all here for a reason.” After watching the two shows back-to-back I can only conclude that the missing 2 percent from one show somehow ended up under the dome of the other.

In the premier of Extant, on CBS, Halle Berry’s astronaut character Molly Woods returns from a 13-month solo mission preggers. In flashbacks aboard the ship she looks just as great in her form-fitting suit as Sigourney Weaver did in Alien before that reptilian thing blasted out of her non-menopausal middle.

Molly comes home to second husband John (Goran Visnjic aka Dr. Luka Kovac from ER, so glad to see him again!) and his invention, their young son Ethan who is a “humanic,” some sort of android/Damien devil child. After just one episode, Extant seems to have recombined Rosemary’s Baby, V, The Omen, Village of the Damned, Alien, the Exorcist, the X-Files, and even Blazing Saddles.

Ico_specieStay tuned. I know I will.

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Wilson Disease – A Genetic Success Story

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800px-Penny_cents_copper_Lincoln_coin_macroA woman I never met, Ingrid, has taught thousands of students about Wilson disease, an inborn error of copper metabolism. But she never knew it, and I wish I could tell her.

Wilson disease is an inborn error of copper metabolism in which the body can’t mop up excess free copper. The element accumulates in the liver, gets into the bloodstream, and enters the brain. Untreated, it’s invariably fatal.

Also known as “hepatolenticular degeneration,” Wilson disease affects 1 in 30,000 (see the National Organization for Rare Disorders) and is autosomal recessive, inherited from carrier parents and affecting both sexes.

The Wilson Disease International Association has a facebook page, but it isn’t as packed and chatty as those for the other genetic diseases I visit often. And Wilson disease has hardly been in the news for astonishing recent progress in the way that cystic fibrosis and  Duchenne muscular dystrophy have. (Recent posts on DNA science have covered these two diseases.

That’s because Wilson disease is treatable. But it wasn’t when Ingrid became ill and then developed a bizarre set of symptoms.

Wilson disease is an inborn error of copper metabolism. It builds up.

Wilson disease is an inborn error of copper metabolism. It builds up.

I read about Ingrid in 1992, in a magazine article. I did some research and included her story in the first edition of my textbook,  Human Genetics: Concepts and Applications, in a boxed reading on how genetic diseases affect metabolism of nutrients, minerals in her case.

At the risk of plagiarizing myself, the section began:

Ingrid is in her early thirties. She lives in a geriatric ward of a state mental hospital, unable to talk or walk. Although her silly-looking grin and drooling make her seem mentally deficient, Ingrid is alert and communicates by pointing to letters on a board. It is hard to believe that in 1980 she was a vivacious, normal high school senior.

Ingrid suffered increasingly odd symptoms: after the initial abdominal pain and headaches, she developed slurred and gravelly low-pitched speech, loss of balance, and altered handwriting. Following a multi-year diagnostic journey, telltale greenish-brownish rings around Ingrid’s irises finally prompted a psychiatrist to recognize Wilson disease. These Kayser-Fleischer rings are thought to be deposits of copper.

Copper deposits ring the irises in people with Wilson disease. (Wikimedia)

Copper deposits ring the irises in people with Wilson disease. (Wikimedia)

It was only then that helpful treatment began. A drug called penicillamine enabled her to excrete the excess copper in urine the color of bright new pennies,” I wrote.

Penicillamine is a chelating agent, a compound that encloses a metal and escorts it out of the body. (“Chelate” is actually Greek for “crab’s claw.”) Ingrid took the first in a series of drugs to treat Wilson disease. Today patients typically take triethylene tetramine and zinc acetate for four to six months, and then lifelong maintenance on the second drug. (A new clinical trial will evaluate Wilson disease drugs to treat a form of melanoma caused by mutation in the gene BRAF. )

I was amazed at the entry for Wilson disease in Online Mendelian Inheritance in Man (OMIM; the bible for geneticists), from 1956:

Almost overnight, Wilson disease became one of the few inherited metabolic disorders for which there was effective therapy.

We don’t often hear about treatable genetic disease, and if we do, the strategy is often not this straightforward.

The midbrain in Wilson disease resembles a giant panda on MRI.

The midbrain in Wilson disease resembles a giant panda on MRI.

120px-Basal_ganglia_1A brain scan at some point might have revealed that Ingrid had the “giant panda sign“ – the appearance of midbrain structures in later stages of Wilson disease. Early-onset osteoporosis, arthritis, heart disease, and kidney problems also arise, and for 5 percent of people with Wilson disease, severe liver damage makes a transplant vital. Psychiatric symptoms include depression, anxiety, mood swings, psychosis, suicidal thoughts. The person may be confused and clumsy.


New editions of my genetics textbook bloomed into life every two years since that first edition from 20 years ago. Stories about real people came and went as I rewrote every other year, but Ingrid’s tale remained.

So imagine my surprise when last week I received this email, from Phyllis Crane.

Hello Dr. Lewis,
My 16-year-old granddaughter is taking a summer class at the University of California, Riverside. The textbook used is your book Human Genetics (2012). I was browsing through the book and on page 21 read the story about Ingrid, who was diagnosed with Wilson disease.

I have been a speech pathologist since 1972 and a clinical psychologist since 1987. In the early 1980′s, I worked at Metropolitan State Hospital in Norwalk, California in the skilled nursing ward. My responsibility was as a Speech Pathologist. Ingrid was one of my patients. When I read about her in your textbook, I was stunned and of course recalled our time together.

When I worked with Ingrid, it was before computers and we developed augmentative communication tools for our patients who were unable to speak. Ingrid’s inability to speak was due to dysarthria weak muscles). Those communication tools consisted of a picture board and an alphabet board. With computers those tools developed into software with the computer producing synthesized speech.

At the same time that I worked with Ingrid, I was referred another individual with Wilson disease with whom I worked on an outpatient basis. He and Ingrid became aware of each other and started to communicate. When my contract with the state hospital came to an end, the ward was closed and the patients moved to another state hospital in Camarillo, California.

I visited Ingrid one time with my other patient, but then lost contact with her. Eventually Camarillo State hospital also closed and is now the site of one of our state universities. I’m thinking that it was at the time of that closure that Ingrid went to live with a family member.” (This is how the entry in my textbook ends, circa 2011.)

I’m assuming that she is a whiz on the computer now,” Phyllis wrote another time, asking that I give her contact info to Ingrid. My answer, that alas I didn’t know what had become of Ingrid, sent Phyllis on a search.

It didn’t take me long to find her obituary. Sad for me, but Ingrid will live on in your book. She would have been thrilled to know she has had an impact on others,” Phyllis wrote.

Ingrid Innecken, born May 21, 1962, had died on August 1, 2010, peacefully.


1. It is important to evaluate all symptoms and think of the zebras, not just the horses. A depressed and anxious teen may not be like all the others. That tendency to lump may be why 16-year-old David Acott, in the UK, needed an emergency liver transplant just this past May. Diagnosis of Wilson disease had taken 2 years, with doctors not seeing beyond his depression and anxiety, until his eyes yellowed and urine darkened and someone finally diagnosed Wilson disease.

2. Exome and genome sequencing dominate headlines, but good old-fashioned clinical diagnosis will always be important. Ingrid’s diagnosis came from a clinician looking closely at her eyes, and seeing something highly unusual, the unmistakable rings of copper. Diagnosis for Wilson disease is testing for free copper (most of the mineral is bound to a protein called ceruloplasmin) in the blood, urine, and liver. A genetic test confirms mutations in the gene encoding copper-transporting ATPase 2 (ATP7B). And low-tech alphabet boards did for Ingrid what computers do today.

3. Presentations of genetic diseases can be extremely tricky because of their variability. More than 500 mutations have been identified for Wilson disease, including the gamut of point mutations, deletions, duplications, and copy number variants. Mixing and matching of mutations in compound heterozygotes – people with two different mutations – makes for an astounding diversity in age of onset (2-70), different responses to drugs, and lifetimes of changing subsets of symptoms.

4. Today I would have hesitated to describe a person with a disease who might be recognized in my textbook without her permission. We live in a peculiar world of sometimes paralyzing anonymity due to HIPAA, yet details down to the odor of cystic fibrosis farts broadcast, with names, on social media.

(Dept. of Energy)

(Dept. of Energy)

But I’m glad I noticed Ingrid’s story and included it all these years. She will be in the eleventh edition of my human genetics textbook, due out this fall. Thank you Ingrid for teaching the world about Wilson disease.

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Getting to the Bottom of Fecal Transplants

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Fecal bacteria sampleImagine that you are a bacterium, comfortably living inside a human bowel movement.

Suddenly, a chemical attack kills most of your neighbors. As other types of microorganisms arrive and begin to take over the vacated niches, they alter the milieu so that you’re washed out in a sudden stream propelled by a blast of gas. How can your few surviving colleagues back in the colon re-establish the peaceful old community?

An infusion of feces from another body can reboot a healthy microbiome in the large intestine (colon), in a biological gentrification of sorts that’s been well studied and much discussed. Now, Vincent B. Young and his team from the University of Michigan and the Essentia Institute of Rural Health in Duluth report in the May/June issue of mBio the biological functions that “fecal microbiota transplantation” (FMT) alters to restore the neighborhood of the colon.


Clostridium difficile (USDA)

Clostridium difficile can take over when antibiotics cleanse the intestines (NHGRI)

FMT delivers other peoples’ excrement to treat recalcitrant infections of Clostridium difficile, a painful and sometimes lethal condition that sweeps in after antibiotics have altered the gut microbiome. In recent years ”C. diff” infection incidence and severity have been on the rise.

Fecal transplants have been done in cattle (via enema) for a century, and on people, in various settings, since the late 1950s. Marie Myung-Ok Lee’s “Why I Donated My Stool,” in the The New York Times a year ago, traces the approach even farther back. She recounts a DIY experience, doctor-guided, that indeed helped her friend with ulcerative colitis. And the New England Journal of Medicine published the straight poop last year demonstrating efficacy.

Feces are a very accessible research material chock full of bacteria. Along the 5 feet of loops of the colon live some 6,800 bacterial species. In one of the first microbiome studies (the subject of one of my very first blog posts and the classic example I use in my textbook), researchers chronicled the establishment of the gut bacterial community by tracking the contents of soiled diapers from 14 healthy babies for the first year, one the child of the chief investigator.

Studies aren't necessary to distinguish between the BMs of breast vs bottle fed infants -- it's obvious.

Studies aren’t necessary to demonstrate microbiome differences between breast vs bottle fed infants — it’s obvious.

David Relman, Patrick Brown and their colleagues at Stanford University, today a powerhouse of microbiome research, found that the babies’ bacteria were quite different at the outset, but by the end of the year, their communities resembled those in the adult digestive tract. And it was published right here at PLOS.

(I ventured briefly into the realm of the microbiome for Medscape, reporting on distinctions between the circumcised and uncircumcised penile ecosystems.)

In the new study, 14 people who’d suffered at least two C. difficile infections received FMT. And it was, as metagenomic studies tend to be, a tremendously data-rich endeavour.

But before I get to the results, let’s address the product and its delivery system. I usually skim, skip, or read last the Methods section of a paper, but in this case I read it first. Just out of curiosity. And it instantly convinced me that my recent decision to switch from the drip coffee method to a French press was wise.

“Donor stool … was collected 6 hours prior to the procedure and then brought to the clinic for preparation of the stool suspension by laboratory staff. The stool was then combined with 90 ml sterile saline and processed in a blender until a smooth consistency is reached. The suspension was then filtered using a coffee filter twice, yielding 40 to 60 ml of stool suspension to be used for transplantation.”

The product, delivered through a nasogastric tube, looks like a melted frozen coffee drink.



The processing destroys the distinctive morphology of feces as depicted so colorfully in the Bristol Stool Chart, a medical tool that I will readily admit I had not heard of. (You can order a coffee mug festooned with the chart.) Ken Heaton, from the University of Bristol, invented it in 1997. Apparently the presentations of human turds hold clues to digestive health.

The researchers identified the bacterial residents in feces from the 14 participants, before and after treatment, from ribosomal RNA sequences, a tried-and-true way to tell eukaryotes (us) from prokaryotes (them). (No fancy genome sequencing required.) Overall, Bacteroidetes become more abundant while Proteobacteria become less so as new feces take up residence.

But the new investigation also imputed what was going on metabolically – presumably so that one day these exact effects can be mimicked by some more palatable approach. “If we can understand the functions that are missing, we can identify supplemental bacteria or chemicals that could be given therapeutically to help restore proper gut function,” Dr. Young said. It reminds me a little of developing infant formula by trying to recreate human milk.

Enterococcus fecalis, a normal resident of the human colon. (NHGRI)

Enterococcus faecalis, a normal resident of the human colon. (USDA)

The analytical tools used offer quite a data dump. Software called “mothur” identifies “operational taxonomic units” (OTUs), which I assume are something akin to species. Then to get at what these microbes are doing rather than simply what they are, the researchers used HUMAnN (HMP Unified Metabolic Analysis Network), which taps into such resources as the KEGG (Kyoto Encyclopedia of Genes and Genomes). Then something called PICRUSt (Phylogenetic Investigation of Communities by Reconstruction of Unobserved States) provides the “metagenomics contribution,” the acronym evoking the image of a “meadow muffin,” one of my favorite scatological synonyms.

Put another way, what, exactly, does the new crap do?

The analysis found 75 “gene modules” of 5 to 20 genes each. And their functions at first conjured up bad memories of graduate school courses in biochemistry. Things that change as new bacteria move in include:

amino acid synthesis and degradation
pace of the citric acid cycle
function of amino acyl tRNA synthetases (key enzymes in protein synthesis)
vitamin and nucleic acid metabolism

Many altered activities were classed as “environmental information processing,” which I deduced from the details referred to a lot of schlepping of amino acids and sugars.

Spermidine contributes some odor to stools. (Wikimedia)

Spermidine contributes some odor to stools. (Wikimedia)

Also altered pre- and post-transplant were levels of spermidine and putrescine,” “foul-smelling organic compounds” initially isolated from rotting meat and semen, respectively. They produce odors reminiscent of rotting flesh, halitosis, and, despite the name, the piscine-like scent of a vaginal bacterial infection.

Some biochemical pathways that didn’t work well in the throes of a bout with C. diff recovered after the treatment. Other pathways revved up after treatment, such as changes in glutamate and gamma amino butyric acid (GABA) metabolism that indicate stressed bacteria.

But remembering biochem isn’t necessary to follow the terrific mBio paper, because a beautifully clear figure lists the pathways on the left, and color-coded sets of three horizontal bars on the right: red for “pre-FMT,” green for “post-FMT,” and blue for the donor material. The green bars inch along from red to blue as the microbial community recovers.

The study confirmed efficacy. Five of the 14 participants still tested positive for C. diff after treatment, but 3 of them were clinically okay, the fourth improved on vancomycin, and the fifth was lost to follow up when the study ended at 6 months. That’s a 12/14 or 86% success rate.

Perhaps one shouldn't mess with a functioning microbiome. (NHGRI)

Perhaps one shouldn’t mess with a functioning microbiome. (NHGRI)

“The bottom line is fecal transplants work, and not by just supplying a missing bug but a missing function being carried out by multiple organisms in the transplanted feces,” Young said. “By restoring this function, C. difficile isn’t allowed to grow unchecked, and the whole ecosystem is able to recover.”

The treatment brings back “colonization resistance,” which is the ability to fend off pathogens that comes with the natural gut microbiome. All of this confirms my long-held hypothesis that bowel-cleansing regimens make little biological sense. Leave nature be.

In May 2013 the Food and Drug Administration announced that it would regulate FMT as an  investigational new drug, but a public hearing led to loosening of that requirement.

Discussion continues about whether human feces for transplant should be regulated as a drug or as a tissue. Meanwhile, stool banks have been established, procedures are being performed in hospitals to treat C. difficile infections, and I’m sure companies are exploring the potential new market. I ventured into a health food supermarket today just to be sure they aren’t jumping the gun, and to my relief, among the gas suppressors and bowel cleansers, I didn’t find anything resembling stool replacement. I suspect the approach may have a bit of a PR problem, a little like comandeering HIV to deliver gene therapy.

Dr. Young and colleagues call for further research to better define the risks of fecal transplants: viral or bacterial infection or inflammatory bowel disease exacerbation in the short-term, and the effects of replacing the gut microbiome with a “non-self” set of microbes in the long term.

I hope we won’t be seeing excrement elixirs as dinnertime infomercials just yet.

(opening photo courtesy of University of Minnesota, via Wikimedia)

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Catching Up With 3 Rare Disease Families

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"If you hear hoofbeats, think horses, not zebras," goes the medical mantra. Rare diseases are unicorns.

If you hear hoofbeats, think horses, not zebras,” goes the medical mantra. The 7,000 rare diseases are unicorns.

Four-year-old Eliza O’Neill’s viral videos, the subject of my last two blog posts, continue to dominate the news media with another appearance on The Today Show June 17. Hopefully, her family’s fight to fund gene therapy for her rare disease, Sanfilippo syndrome type A, will focus more attention on the entire rare disease community – 30 million people in the U.S. alone.

That’s a lot of families.

Four years ago, I spent the summer getting to know the families whose stories became my gene therapy book. Thanks to social media we’ve stayed in touch, and I’ve met many others. All continue to astonish me. Here’s a catch-up with three families featured in past posts.

Laura King Edwards ran the Thunder Road half marathon blindfolded, in honor of her sister Taylor. Beside her is Dr. Steve Gray, PI of gene therapy trials for two brain diseases.

Laura King Edwards ran the Thunder Road half marathon blindfolded, in honor of her sister Taylor. Beside her is Dr. Steve Gray, PI of gene therapy trials for two brain diseases.

Laura King Edwards posted at DNA Science a year ago about her younger sister Taylor, now 15, who was diagnosed with ceroid lipofuscinosis, neuronal type 1 – aka Batten disease – when she was 7. Recalls Laura:

“In the worst hour of our lives, we learned that my bright-eyed, golden-haired, intelligent sister – a second grader who loved to sing and dance and run and play – would go blind, have seizures, and lose the ability to walk, talk, and swallow food. She would deteriorate … confined to a wheelchair. She would have to have a feeding tube. Eventually, she would die – blind, bedridden, and unable to communicate.”

Laura eloquently captures her sister’s life and her family’s efforts to help fund a gene therapy clinical trial at her blog, Write the Happy Ending. A post from last week is particularly heartbreaking. Rather than charting her sister’s decline with brain scans or mobility tests, Laura notes that in the 6 weeks between haircuts, Taylor lost the ability to walk. Last week, she had to be carried up the stairs to the hairdresser. This week, she’s in the hospital.

To better get into her sister’s head, Laura runs races blindfolded.

“I do the runs for a variety of reasons. I’ve always been a runner, and running helped me face Taylor’s illness when she was first diagnosed. After watching her run the first of two 5Ks with her Girls on the Run team despite battling Batten disease (and she was already blind at that point), I started running in her honor. I mainly run for Taylor to raise awareness, but my runs have also raised money for Taylor’s Tale. The Thunder Road half marathon I ran with Dr. Steve Gray in November raised money for the (gene therapy) project at the University of North Carolina.

I’ve run 18 races for Taylor. Thunder Road was the only race I ran blind, but I went on 18 blind training runs to get ready for it.

635205790291677074During my months of training to become a blind runner and far more so in the months following the race, my sister slipped farther down the chasm of Batten disease. It is a deep, dark chasm. There are no footholds for climbing out, and some days, no light reaches her ledge. And yet, each day she teaches me something new about courage; each day, she imparts some great piece of wisdom without having to say anything at all.

My next challenge is to run a race in all 50 states for Taylor to continue spreading awareness of Batten disease and build support for the rare disease community. I’m kicking it off this summer!”


Ten-year-old Hannah Sames also has a very rare inherited disease of the nervous system, giant axonal neuropathy (GAN). DNA Science told her story about a year ago too.

In GAN, intermediate filaments composed of a protein called gigaxonin overgrow and run askew, hampering nerve function. Hannah is very slowly losing mobility, and suffers from kidney stones and visual loss, as the lack of gigaxonin in various body parts makes its presence known in ebbing motor and sensory functions.

Dr. Gray (behind Laura in the photo above) began working on gene therapy for GAN before he took on the Batten disease project, and the GAN trial is set to begin within the next few months at the NIH Clinical Center. The trial is largely possible due to the constant networking, meeting-holding, and fundraising efforts of Hannah’s family – parents Lori and Matt, and sisters Reagan and Madison. Their Hannah’s Hope Fund (HHF) was born in the days following the diagnosis in 2008. The highlight is the annual ball, held in February in snowy Albany, NY, near the Sames (and my) home. From Lori:

Doris Buffett's Sunshine Lady Foundation donated $500,000 in matching funds to Hannah's Hope Fund for GAN.

Doris Buffett’s Sunshine Lady Foundation donated $500,000 in matching funds to Hannah’s Hope Fund for GAN.

“The Hope and Love Ball began 5 years ago when friends, Todd and Beth Silaika and Tim and Lee Wilson, approached us with the idea. The first formal gala in 2010 netted $90,000 and was a Valentine theme, fitting for February. Other themes followed: Monte Carlo, Mardi Gras, Midnight in Paris, and Candyland this year, which netted more than $165,000.

In 2010, HHF was awarded a $500,000 all-or-nothing matching challenge grant from Doris Buffett’s Sunshine Lady Foundation. The deadline to raise the funds was the night of the Ball. Snow kept Ms. Buffett (Warren’s sister) away the evening when more than 450 HHF supporters celebrated the success of the $1.2 million, 6-month “Hope for a Million” fundraising campaign. Ms. Buffett was the highlight of the event the following year.

To date, HHF has raised $6 million in 6 years, grassroots, with the vast majority of funds spent on the GAN gene delivery Investigational New Drug (IND) work. The FDA placed the protocol on “Active” status at the end of May, awaiting IRB approval of the GAN gene delivery system. Then trial recruitment can begin.

Unfortunately, Hannah, the inspiration of HHF, has a homozygous deletion mutation. She isn’t a candidate for the phase 1 trial because only missense mutation patients will initially be included. Hannah is awaiting the results of a non-human primate study aimed at inducing tolerance to an intracellular transgene in the CNS. If tolerance is achieved, it will likely be 10 months to a year before Hannah can receive gene delivery.”

(Hannah doesn’t make gigaxonin at all, and so introducing it into her spinal cord, via healthy genes in viral vectors, could trigger an explosive immune response. The other kids who will be in the trial make abnormal forms of the protein, and so their immune systems are already alerted that gigaxonin is a “self” protein.)


Michael and Mitchell Smedley and their friends brainstormed the Bike the Basin event.

Michael and Mitchell Smedley and their friends brainstormed the Bike the Basin event.

A few months ago at DNA Science, Kristen Smedley told how she and her husband Mike assembled a research team to pursue gene therapy for the CRB1 form of Leber congenital amaurosis, which has robbed their sons Michael and Mitchell of sight.

But the boys are more interested in having fun than recruiting researchers, so they dreamed up the hugely successful Bike the Basin event, a half-mile race at the Northampton Civic Center Basin in Bucks County, PA. Kristen continues.

“Back in summer 2011 when the Curing Retinal Blindness Foundation launched, I asked my kids to come up with a fundraiser that could get their friends involved and start getting the word out about our big mission. I wanted my boys to take the lead because while it’s nice that so many people want to help them due to their blindness, my guys need to be able to show the world that they can help themselves.

We gathered about 15 of their closest friends at my kitchen table and the boys pitched their idea of a bike event fundraiser. The kids brainstormed ideas of how to make it work (with parents taking notes and serving lots of ice cream) and Bike the Basin was born!

Just under three months later, the first event raised $20,000. The first three BTB events raised just over $200,000 combined, and the goal for 2014 (Oct 5th) is $250,000. We’ve raised about $80K so far!”


Hannah and her sisters and parents.

Hannah and her sisters and parents.

The families who raise funds for gene therapy clinical trials begin with their own relatives in mind and perhaps as a way to channel their anxiety and fear into something productive. But their generosity extends much farther.

As rare disease-based communities form and strengthen, certain individuals emerge as catalysts. Laura King Edwards, Lori Sames, and Kristen Smedley are three.

Gene therapy will almost certainly be too late for Taylor, and possibly for Hannah. But the Smedley boys may one day be able to see. And Eliza O’Neill may find her way into a clinical trial before Sanfilippo syndrome darkens her sunny childhood, thanks to the efforts of the media to share her story, and the kindness of so many strangers. But Eliza is one child, representing one unicorn. There are so many more.

Whatever the future holds, the efforts of these brave families will reverberate for years to come, measured in the numbers of lives improved or saved.

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Eliza’s Journey: Part 2

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Gene therapy is, finally, about to take off! The feeling was palpable at the American Society of Gene and Cell Therapy annual meeting a few weeks ago. It’s been nearly a quarter century since the first experiment in humans.

The drug development pipeline is beginning to swell with early-stage trials, while a few candidates are closing in on FDA approval for marketing. And just as the annual meeting got underway, Francis Collins, director of the NIH, announced his approval of the Institute of Medicine’s late-2013 recommendation to eliminate certain redundant regulatory hurdles in the launch of a clinical trial of gene transfer. That’s what the technology is technically termed before showing efficacy. It’s a way to deliver healthy genes to compensate for those affected by a particular disease.

There’s more encouraging news.

Spark Therapeutics is preparing to commerical gene therapies for Leber congenital amaurosis type 2 and hemophilia B. (courtesy Spark Therapeutics).

Spark Therapeutics is preparing to commercialize gene therapies for Leber congenital amaurosis type 2 and hemophilia B. (courtesy Spark Therapeutics).

In May, Spark Therapeutics, affiliated with the Children’s Hospital of Philadelphia Center for Cellular and Molecular Therapeutics, announced $72.8 million in new funding, with efforts initially in RPE65-related blindness and factor IX deficiency (hemophilia B). Both indications are far along the development trajectory.

Searching for “gene therapy” at brings up 3,547 hits, but that includes terminated and completed projects. Gene Therapy Clinical Trials Worldwide, in the
Journal of Gene Medicine, lists the number of ongoing trials as 1,991. And gene therapy came up several times two days ago during testimony to the House Energy and Commerce Committees about promoting “21st century cures.”

And still more …

The problem of gene therapy strategies that vanquished inherited disease but caused leukemia is disappearing, thanks to retooled or replaced viral vectors (a future post) and tremendous cooperation among research groups in several nations. And my latest jobs list from Linked/In included several biotech/pharma companies seeking experts in regulatory affairs for gene therapies.

Perhaps most important in the evolution of gene therapy are the contributions from families of kids with rare genetic diseases, many of whom have shared their stories on this blog.

Last week’s post introduced the O’Neills, whose viral video has raised more than $1 million towards launching a phase 1/2 clinical trial (safety and possibly efficacy) for gene transfer to treat Sanfilippo syndrome type A. Without intervention, in the next year or two, 4-year-old Eliza will begin to descend into the nightmare of irreversible brain damage as an enzyme deficiency upsets the biochemical balance in her cells’ lysosomes, the debris sacs.

The media, in droves, conveyed Eliza’s need in early April. Most reported that research so far is only in mice, perhaps because results from a clinical trial in France weren’t officially published until May, in Human Gene Therapy, a source that might be off the radar of many media folk.

The French company,, founded by Sanfilippo parents in 2009, delivered genes by catheter directly into the brains of four children. Results were in by June 2013, a month before Eliza was diagnosed. The trial that she may participate in at Nationwide Children’s Hospital in Columbus, Ohio, will deliver the needed genes much less invasively, by intravenous infusion. The two trials use different viral vectors.

Michaël Hocquemiller, PhD, who handles scientific and clinical affairs at Lysogene, shared good news and links: to last month’s publication, FDA’s granting of orphan drug designation to SAF-301 a year ago, and recent financing. Their procedure appears to be safe, but the trial couldn’t show much benefit in so few children over so short a time. Still, “neuropsychological evaluation revealed an improvement of behavior: a strong decrease in hyperactivity, strongly improved sleeping patterns and improved focus and socialization skills in several patients,” he said via email.

Safety in a few kids is a giant first step, and it’s why additional trials are imperative to confirm the findings and demonstrate efficacy.

I haven’t yet spoken to the researchers at Nationwide, but a news release tells how families helped to fund and participated in the natural history studies that are critical to setting parameters to assess whether a gene transfer protocol is working. The project may begin a clinical trial by the end of this year, with a boost in funding vector production from the efforts of the O’Neill family and the generosity of so many caring strangers.

The story of how the family’s video came to be is in itself fascinating. So here is Part 2 of Eliza’s Journey, from Glenn O’Neill.

SavingEliza - instagramWe knew we had to jump right in. We started our own non-profit 501c3 Cure Sanfilippo Foundation (Tax ID: 46-4322131) with no paid employees and all net funding to the cause currently going toward supporting this clinical trial.

Through March we had raised $250,000 from traditional fundraising we continue today. During this time, we kept thinking we would catch a break and meet an “angel” donor, or celebrity, or corporation who would bail us out. It hadn’t happened and we’d only received a long list of declines.

Early in February 2014, I realized our fundraising at this pace wouldn’t get us anywhere close to the goal. How could we get more people to hear Eliza’s story? We had exhausted our Facebook, email, and social media friends. We were out of options.

One very late night in February, out of desperation, I did a simple Google search for “how to make a viral video.” I sent a “shot in the dark” email to the writer of the first article that came up: Karen Cheng – Give it 100.

She responded!

dnaKaren reached out to a few of her friends on our behalf, and long story short, in the last week of March 2014, freelance videographer Benjamin Von Wong and 2 other artists stayed at our house for 8 days, all sleeping on couches. They filmed 40 hours of footage, editing along the way, and charged us nothing! They bonded with our kids and my wife and I joked that it reminded us a bit of our college dorm life. We had the first real laughter since the diagnosis.

When they left, they gave us the 3-minute video of our very personal story, which does a better job explaining the time-critical situation Eliza is facing far better than any words written here could.

This video was released on April 2nd and went viral the following week. Since then, the video has raised over $820,000 with over 16,500 donors. We are closing in on the Most Ever Raised on, which is just over $800,000.

The SavingEliza video has had more than 270,000 views and the video featuring Eliza’s brother Beckham is approaching 100,000 views. Fox News, ABC News, NBC News, the Today Show, MSNBC Live, Al Jazeera America, Huffington Post, BBC, and many others have covered Eliza’s story. (These figures have now been dwarfed. A third video documents the making of the one that went viral.)

Our fundraising had just been catapulted to a completely new level, and honestly, we weren’t ready for it. There had been no real business plan in place for this wild idea and it all happened so quickly. We scrambled for help from friends and family with the increased administration that comes with this. But what we did have now was renewed energy, renewed hope! While there is still a long way to go on funding, we truly felt for the first time that perhaps money would not be a limiting factor. Perhaps we could get there.

In the next weeks, our foundation will be funding the vector production for Sanfilippo Type A, to be used in the clinical trial at Nationwide Children’s Hospital. This was made possible by the kindness of complete strangers around the world, $10, $20, $50 at a time. This is a crucial and time-sensitive step as it takes 6 months to produce these gene therapy viral vector doses.

The O'Neills

The O’Neills

We move now to The Final Step, which is funding the actual clinical trial. We look to raise over $1M more by October 2014.

The amazing researchers at Nationwide Children’s Hospital have been working more than 15 years to get to a point where this disease can be treated. So many incredible families have funded much of this research throughout the years, and we owe them our deepest gratitude. We need this final push now so never again does a parent have to hear “Sanfilippo syndrome” followed by the words “no cure and no treatment.” And never again does a child have to suffer the devastating effects of this disease. Just this week, another beautiful little girl, age 10, is in hospice and her family is making arrangements. It has to stop.

We’ve received so much positive feedback, many with inspirational phrases like “keep going,” “never stop,” and “we are all with you.” We’re in the midst of a historic new model where social media is funding research. Where everyday people, like you and me, from around the world are helping with any amount they are able, to stop this deadly childhood disease this year….and to save Eliza!

Looking ahead, thanks to the help of so many caring strangers.

Looking ahead, thanks to the help of so many caring strangers.

I go back to my journal entry from July 17 of last year, and those words still apply, but in a much different way than they did then. My goal is still to keep Eliza “happy and smiling,” but now for her, it can last a full lifetime.

(Thanks to Glenn O’Neill for family photos)

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Eliza’s Journey: Part 1

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Eliza O'Neill has San Filippo syndrome.

Eliza O’Neill has Sanfilippo syndrome.

If ten science writers were asked to write a book about gene therapy, a biotechnology with roots going back to the 1950s, they could tell ten different stories.

Any account of gene therapy would include the first experiment in humans, on a 4-year-old in 1990. The narrative  would tell of the dual tragedies that until recently defined the field: 18-year-old Jesse Gelsinger, whose immune system lethally rejected treatment for a urea cycle disorder in 1999, and then the young boys who, shortly after, developed leukemia following successful treatment of their inherited immune deficiency. The cancer hadn’t happened in the mice used to clear the gene transfer protocol for clinical trials.

After revisiting the tragedies, my book The Forever Fix: Gene Therapy and the Boy Who Saved It introduces adrenoleukodystrophy (ALD) to showcase family activism, giant axonal neuropathy (GAN) to tell what it takes to get a gene therapy clinical trial up and running, and then Canavan disease to see what happens years after gene therapy. Sandwiching those cases is the experience of Corey Haas, who would surely be blind if not for gene therapy in 2008, when he was 8.

But I could have told, instead, other ongoing stories of the evolving gene therapy — hemophilia Bbattling Batten disease, Wiskott-Aldrich syndrome, metachromatic leukodystrophy, cancers or HIV … I cover some of them here at DNA Science, and at Medscape Medical News, as the successes accrue. It’s finally happening.

Corey Haas, who had gene therapy to treat Leber congenital amaurosis type 2, would never be able to fish in the Hudson near his home had he not had gene therapy. (Nancy Haas)

Corey Haas, who had gene therapy to treat Leber congenital amaurosis type 2, would never be able to fish in the Hudson near his home had he not had gene therapy. By now he would have been blind. (Nancy Haas)

I recently realized, though, after learning of 4-year-old Eliza O’Neill’s future with Sanfilippo syndrome, that The Forever Fix only touched on two types of experiences.

The lucky, like Corey, found gene therapy because a physician knew about a clinical trial, and they fit the criteria. The many more unlucky are the parents who receive a deadly diagnosis and then discover that no one is pursuing treatment. It’s up to them.

The families are a key part of many gene therapy advances. For ALD it’s a pioneering trio of sisters (Amber Salzman, Rachel Salzman, and Eve Lapin). And the soon-to-start gene therapy clinical trial for GAN is largely possible thanks to the Herculean efforts of Lori and Matt Sames. (Their beautiful daughter Hannah, now 10, graces the cover of the new edition of my human genetics textbook, not for the neural destruction of GAN, but for her unusually  kinky curls, also part of the phenotype.)

Exquisite little Eliza represents a third group: a gene therapy clinical trial nearly ready to go that runs out of money. Creating, selecting, and scaling up the viral vector that delivers the healing genes is expensive. Still, Eliza’s parents, Cara and Glenn, didn’t have to start from scratch like the Sames’ and the Salzman sisters. They and other Sanfilippo families are desperately trying to fund the making of the medicine that is gene therapy.

Racing against a rapid killer must go beyond golf tournaments and bake sales, road races and dances, and other classic fundraising routes that work when one has more time. The families need to raise about $2.5 million, and soon. Thanks to a video (Saving Eliza) that went viral in April and the media appearances that followed, donations are coming in. But social media are ephemeral, and the amount raised isn’t enough, yet.


(Dept. of Energy)

(Dept. of Energy)

Gene therapy introduces working copies of a gene that is absent or malfunctioning in a disease, usually aboard a virus or encased in fatty bubbles. Technically it’s “gene transfer” until evidence shows that it works.

The gene therapy for Sanfilippo syndrome uses a viral vector that enters the brain, meaning that the treatment is a simple infusion – not the skull-piercing catheters once used to deliver genes to children who have Canavan disease.

Sanfilippo syndrome is a lysosomal storage disease (LSD). The lysosomes are the “suicide sacs” of the cell, each housing 43 types of enzymes that dismantle specific molecules. A mutation that robs a cell of just one of these enzymes sets into motion an LSD.

Some LSDs, like Tay-Sachs disease, have telltale signs even before birth, if one could look, in addition to the detectable enzyme deficiency. The molecule that the enzyme should destroy builds up, as the molecule that would result from its action ebbs away, like a knot in a filled garden hose causing a backup at one end and a dribble at the other. In Sanfilippo syndrome, aka mucopolysaccharidosis (MPS) type IIIA, the missing enzyme is heparin sulfate sulfatase.

Eliza and Beckham (Glenn O'Neill)

Eliza and Beckham (Glenn O’Neill)

Eliza’s 7-year-old brother Beckham describes his sister’s “very bad disease” more clearly than I just did, in this second video: “It clogs up her brain and that makes her not learn very well. She’s hyper.” The condition is autosomal recessive, inherited from two carrier parents. Therefore each child has a one in four chance of sharing Eliza’s fate.

So here, from Glenn O’Neill, is the first part of Eliza’s story.

We found out the terrible news today. For now, I want to focus on her wonderful personality and life every day. One of my goals is to keep her happy and smiling for as long as possible. I love her so much.

This was my journal entry on the evening of July 17th, 2013. I never kept a journal before this. Earlier that day, our 4-year-old daughter Eliza was diagnosed with a rare terminal genetic disease called Sanfilippo syndrome type A. In one terrifying instant, we were told that we would have to watch Eliza fade away before our eyes. My journal entry words reflected the lack of hope a parent first feels when told their child has a disease that has no cure and no treatment.

Eliza and other children with this disease are missing an essential enzyme for normal cellular function. Over time, a toxic material called heparin sulfate builds up in their brain and body, leading to severe disability and death before they even reach their teens. This disease affects both genders, all races, all countries and continents. It is rare, but it is everywhere and the world needs to know.

Right now Eliza is a fun-loving 4-year-old who loves to talk, sing, run and MOST of all, cuddle. She loves to play dress-up and horse around with her rowdy big brother Beckham, who fortunately does not have Sanfilippo. She is, however, beginning to show signs of the disease in her learning and attention. And if nothing changes, it will only get worse. And quickly.

By age 6, most children with her disease have irreversible brain damage and lose the ability to speak. As the disease continues to tear through her brain and body, she will lose the ability to walk and eventually she won’t even be able to feed herself. Seizures and painful movement disorders will take over. Life expectancy is usually early teens but preceded by this severe disability.

These devastating changes are a 100% certainty if she doesn’t get treated, and soon. It is a parent’s worst nightmare, and an unfair sentence for any innocent child.

After diagnosis, we quickly began looking for researchers working on Sanfilippo, here in the US and around the world. There weren’t many. My wife Cara, who is a pediatrician for special needs children (ironically), made some key contacts who pointed us in the right directions.

What we found was amazing.

The O'Neills

The O’Neills

What we found was HOPE, and near term! There is a gene therapy trial being scheduled for late 2014 that is specifically for children with Sanfilippo syndrome types A and B, the two most common forms of the disease.

Researchers at Nationwide Children’s Hospital in Columbus, Ohio, have shown that the gene therapy stopped the disease in animal models. It is a one-time injection, delivering the gene that encodes the enzyme missing in the disease using the vector AAV9, which has the ability to cross the blood-brain barrier and clear the storage.

This treatment could save Eliza. In addition, this delivery method, if successful, has great possibility to be used in other MPS disorders as well as more common neurological diseases.

One of the main things standing between Eliza and her miracle is money. The trial is lacking funding to make the medicine, administer the treatment, and remain on schedule. Every moment counts as Eliza approaches the tipping point when her disease will take an irreversible turn for the worst.

The total amount needed to fund both production of the medicine (gene therapy viral vector) and conducting the clinical trial for Sanfilippo type A is $2.5 million. What would you do if you knew that money was the only thing standing between your child and her chance at a full and happy life? What would any parent do?

We could not just stand by and watch our little girl lose everything she is, suffer unimaginable pain and frustration and ultimately die. What would we tell her big brother in a few years, when the disease has taken over completely? What would we tell ourselves?

Glenn O’Neill

Eliza’s Journey will continue next week with Glenn’s description of their campaign, which brought Eliza’s plight before the world. I’ll follow up with more on the science. In the meantime, please help. I learned from chatter last week at the American Society of Gene and Cell Therapy annual meeting that one reason why Europe leads the way in gene therapy is that funding comes largely from charity. That means US.
Twitter: @SavingEliza #SavingEliza
Checks can be sent to: Cure Sanfilippo Foundation, PO Box 6901, Columbia, SC 29260

Thanks to Glenn O’Neill for family photos.

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A Checklist for Gene Therapy From the UK Cystic Fibrosis Trial

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A clinical trial is a series of hurdles.

A clinical trial is a series of hurdles, but not in parallel.

Washington, D.C.           I’m at the American Society of Gene and Cell Therapy annual meeting, one of my favorite conferences. The very first talk provided a great example of why it is taking gene therapy so long to reach the clinic — a milestone that hasn’t happened yet in the U.S. The first gene therapy experiment was 24 years ago.

For the first talk, Uta Griesenbach, PhD subbed last minute for Eric Alton, MD to present progress in a phase 2b double-blind placebo-controlled clinical trial of gene transfer (that’s what gene therapy is technically called before results indicate it works) to treat cystic fibrosis (CF). It’s “wave 1” of the effort from the UK Cystic Fibrosis Gene Therapy Consortium, in which more than 80 researchers are participating.

The consortium made headlines about two years ago when the UK Medical Research Council and the National Institute for Health Research refilled their coffers with £3.1 million (US$4.9 million). Funding had been dwindling, perhaps because of the physiological hurdles that CF presents, against the backdrop of gene therapy setbacks. These began with the death of 18-year-0ld Jesse Gelsinger in 1999 days after gene therapy, and the leukemia that’s cropped up in clinical trials for two immune deficiencies. Many investigators had given up on gene therapy for CF – but not the tenacious UK group, led by Dr. Alton.

Cystic fibrosis results from an absent or malformed chloride channel.

Cystic fibrosis results from an absent or malformed chloride channel.

Cystic fibrosis, at the risk of evoking clichés, has been a tough nut to crack. The gene and its encoded protein, the cystic fibrosis transmembrane regulator (CFTR), were discovered in 1989, and my post from April 10 details some of the earlier history of this disease.

The multi-system symptoms – lung congestion and susceptibility to infection, pancreatic insufficiency, male infertility – result from malformed, misfolded, or absent chloride channels. More than 25 clinical trials, involving more than 400 patients, have attempted to deliver functional CFTR genes.

Improvements were limited or transient, because of the nature of the illness. Thick sticky mucus gets in the way, and cells lining the respiratory tract divide so often that normal cell turnover may jettison an altered cell in a cough or swallow too soon to see a sustained effect. Plus, correcting the problem in the airways won’t alleviate the pancreatic clogging that leads to the classic symptom of “failure to thrive.”

Most gene therapy trials use retooled viruses as vectors to deliver the payload, but wave 1 delivered CFTR genes in an aerosol of tiny lipid bubbles, liposomes. They echo the lipid bilayer of a cell membrane so coalesce themselves across that barrier, releasing their cargo inside.

At first I was disappointed when Dr. Griesenbach said she wouldn’t be presenting results. But when she explained why – “we haven’t broken the blinding yet” – I suddenly realized that the journey to demonstrating that a new therapy works is just as interesting as arriving at that destination.

A double-blind, placebo controlled trial design may seem cruel, intentionally depriving people of a possible treatment, but it is essential to demonstrating that a new treatment actually works. There are workarounds – the control group can receive an existing treatment. And trials are revamped, access expanded or FDA approval accelerated if a result is obviously compelling and people are suffering. The cancer drug Gleevec is an example of a drug hurtling towards the market. It happens, but very rarely.

I don’t think Dr. Griesenbach intended to focus on the hurdles researchers must leap to even plan testing a gene therapy, but that’s what held my attention. The reasons help to explain why clinical trials can take years. Following are the questions that needed answers and the concerns that emerged during this CF trial, which has been in progress since 2002.

1. How much of a gene’s function must gene therapy restore?
In gene therapy, a small change can go a long way. That’s the case for a gene transfer approach for the clotting disorder haemophilia B, presented at a news conference by Andrew Davidoff, MD, from St. Jude Children’s Research Hospital. Introducing the gene for clotting factor IX that restores the level to less than 8% of normal activity can free a man from needing to take clotting factor to prevent life-threatening bleeds.

For CF, men whose only symptom is infertility have 10% residual function of the chloride channels. “So if we can achieve some increase, we can have a significant impact,” said Dr. Griesenbach. A 6% increase in lung function might be all that’s necessary.

A liposome is like a bubble of cell membrane.

A liposome is like a bubble of cell membrane.

2. How should researchers pick the best vector and its cargo?

Choosing a vector and making it safe is perhaps the toughest challenge in gene therapy. Investigators must design the delivery method before a phase 1 trial gets underway, and stick to it.

The situation isn’t like getting a new laptop when Apple introduces a new and improved model. Researchers can’t change or tweak a virus, alter the recipe for a liposome, or replace the DNA cargo without going back to square 1, phase 1. It’s one reason why the gamma retroviral vectors that caused leukemia and the adenoviruses that evoked a devastating immune response are still in use, although some have been made “self-inactivating.”

The CF trial used a liposome delivery method developed at Genzyme awhile ago. But the researchers modified the DNA within to decrease the stretches of cytosine and guanine (“CpG islands”) that invite inflammation and they added a bit to extend the effect. That meant starting from scratch in the phase 1 trial, even though the liposome recipe had been used before.

CF affects more than the respiratory system.

CF affects more than the respiratory system.

3. Which endpoints are the most meaningful?

The CF team tests cells lining the nose and airways for chloride transport, finding that it can reach about 20 percent of normal following gene transfer. Other assays include a “lung clearance index” from inhaling a harmless dust and scans that use technetium to show clear areas in the lungs.

But these measures meant little to the trial participants. “The patients said, ‘so what? Will it make my lung disease any better?” Dr. Griesenbach said. “Our program is now hinged around addressing that question. How much improvement is necessary to have a clinical effect?” A quality-of-life questionnaire is now part of the protocol.

4. Which types of patients should a clinical trial enroll?

Should the sickest patients try a new treatment because they are the most desperate, or should the healthiest, because they have a better chance of surviving the experiment? Part of the outcry over the death of Gelsinger that effectively halted the field for two years was the fact that he had not been desperately ill.

The symptoms and natural history of CF dictate the optimal age of trial participants. “In CF we face a dilemma. Very young children have less mucus, but it is harder to measure their increase in lung function. In the full-blown disease patients have lots of thick sputum. It is hard to find the right patients. You need a balance,” Dr. Griesenbach said. They decided on 12 as the minimum age, with average age 22.

Patients received 12 monthly doses, bracketed by 4 additional visits, and the last participant finished just two weeks ago. The monthly intervention was nothing compared to the hours of procedures that people with CF go through on a daily basis to expel mucus.

The CFTR gene is on chromosome 7.

The CFTR gene is on chromosome 7.

I don’t know whether the patients in the UK trial are stratified by mutation, but the development of the blockbuster drug Kalydeco illustrates the importance of distinguishing among the 1600 or so variations of the CFTR gene sequence. Kalydeco corrects misfolding, which affects only some patients with specific mutations, but can be teamed with other drugs to help more. And a new contender for a CF drug is targeted at patients with nonsense mutations, who make no CFTR protein at all.

5. Expect the unexpected.

The researchers determined that they needed 120 patients, and they started with 130, just to be safe. Then in January 2012, the FDA approved Kalydeco. Some participants, understandably, dropped out of the liposome gene transfer study to take the new drug.

6. Think ahead.

So far, CFTR delivery via liposomes seems to be safe. Some of the patients who are feeling better baked cakes for the researchers, although efficacy isn’t known yet. But the consortium is running a parallel “wave 2” using lentivirus (disabled HIV), in case the fatty bubbles aren’t efficient enough or the effect too transient. (I’ll cover HIV as a gene therapy vector in a future post.)

Results are in, Dr. Griesenbach concluded, and will be presented at the North American CF Conference in October. So far the team knows that patients experience a very brief period of fever and decrease in lung function, but recover well. Then some of them improve. A third of patients fully responded, another third had some correction of lung function but not to entirely normal levels, and a third didn’t respond.

The unblinding will reveal whether the gene transfer is responsible for the patients who did the best. And if they are indeed the ones who received functional CFTR genes, then the next chapter – a phase 3 trial – will be up to industry.

It’s easy to see why approval of a gene therapy takes so long!

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