Leroy Stevens: Fairwell to The Unsung Hero of Stem Cell Research

Leroy-Stevens1-230x300Yesterday a Google Alert popped up with a blast from my past, an obituary for Jackson Lab researcher Leroy C. Stevens. It quoted me calling him “The unsung hero of stem cell research” in an article I wrote 15 years ago for The Scientist. Dr. Stevens passed away on March 28.

Leroy Stevens was born in 1920 in Kenmore, New York, earned a B.S. from Cornell University in 1942 and a doctorate in embryology from the University of Rochester in 1952. I’d discovered his contribution while writing an article for The Scientist published on September 27, 1997, “Embryonic Stem Cells Debut to Little Media Attention.” Unfortunately the headline left off the word “human,” so hardly anyone noticed the article; hES cells blasted into public consciousness some 14 months later.

Back in those pre-electronic days, I trudged to the upper floors of the library at Albany Medical College, where I ironically teach online today, and found a dusty volume that held the treasures of Dr. Stevens’ serial images of developing mouse embryos.

IMG_1086The Scientist has graciously agreed to let me post my tribute from 2000. A more detailed version is in my essay book, “Discovery: Windows on the Life Sciences” (Blackwell Science, 2001). Many thanks to Anne Wheeler for permission to use the photos of her father. He spent his later research years developing mouse models to test chemotherapeutic agents. An update from her follows the post.

A Stem Cell Legacy: Leroy Stevens
The Scientist, March 6, 2000

When Science voted stem cell research its 1999 Breakthrough of the Year, the congratulatory article traced the field’s origin to the 1981 successful culture of mouse embryonic stem (ES) cells. But the roots of exploring these multipotential cells go back considerably farther, to a little-mentioned researcher who worked with mice at the Jackson Laboratory in Bar Harbor, Maine.

FROM CIGARETTES TO TERATOMAS
Leroy Stevens arrived at the lab in 1953, a newly-minted developmental biologist in a premolecular era when the tools of the trade were mostly one’s eyes. The young scientist found himself with an initial assignment that he calls “crazy.”

“The founder of the lab had gotten a grant from a tobacco company, and they wanted him to show that it wasn’t tobacco that was the problem–it was the paper in cigarettes!” recalls Don Varnum, a long-time technician in the lab. So Stevens dutifully dissected cigarettes and exposed mice to the components. One day, he noticed a mouse with a huge scrotum. “We killed it, and looked at the testes, and they had strange things inside.”

This teratoma from the chest has fat blobs, a tooth, a hair, and bone bits.

This teratoma from the chest has fat blobs, teeth, a hair, and bone bits.

The growth included a mishmash of tissue, including hair and teeth. It was a teratoma, and other mice of strain 129 had them too. Intrigued, Stevens pursued federal funding to explore teratomas, and the support lasted a professional lifetime. “This stuff was extremely interesting, and it sure beat studying cigarette papers!” he recalls from his home in rural Vermont, where he has lived near family since suffering a stroke 10 years ago.

Stevens bred strain 129 to select for the teratoma tendency. Then he developed a serial transfer technique to be able to continuously study the rare cells in some of the growths that rendered them cancerous. When and if a teratoma ran out of these “embryonal” cells, it would just sit there, sporting its peculiar mix of hair and teeth, cartilage, and tiny tubules. “We just wanted to keep the tumor alive longer so we could study it. But it took a long time to succeed,” says Stevens.

By passaging teratomas through many hosts and removing them from time to time and cataloging the tissues, the researchers witnessed all that these cells could become. And the odd tissue struggling for identity wasn’t as haphazard as it seemed. When Stevens and Varnum transplanted teratoma bits into the peritoneal cavities of mice, curious growths resembling inside-out embryos formed, called embryoid bodies. The tissues from the teratomas, given signals from the ascites fluid of the peritoneal cavity, seemed to be trying to organize.

The researchers continued to describe the tissues of teratomas and embryoid bodies, supplementing their publications with spectacular atlases of photographs that captured the temporal unfolding of this deranged development. Although the anatomy was clearly off, both out of place and out of time, Stevens and Varnum noted that the events followed a sequence of sorts. “Roy looked at thousands and thousands of mice. He noticed that for tissues to develop, several tissues must be in contact, so that the cells know whether to become liver or kidney,” Varnum recalls.

256px-JacksonLabsSignAlthough Stevens’ and Varnum’s work was purely basic research, the events that they so carefully chronicled, and the biochemicals from without and within that fueled the choreography of early development, are what stem cell researchers and tissue engineers are deciphering today.

TRACING TERATOMA ORIGINS LEADS TO ES CELLS

With a steady supply of teratomas and their tissues identified, Stevens moved on to other questions–where, and when, did development take a wrong turn? And so he began looking backward, seeking the earliest stage when cells in the testes looked different, and then going back a few more days to account for changes not obvious to a human observer. This approach took him to the genital ridge in a 12-day prenatal mouse, which houses primordial germ cells, sperm precursors.

800px-Lab_mouse_mg_3140Then in 1970, Stevens made a leap that would profoundly affect stem cell technology a decade later–he noticed that the primordial germ cells that gave rise to teratomas looked a lot like the cells of considerably earlier embryos. So he transplanted cells from various stages of early strain 129 embryos, including inner cell mass cells (a very early embryo, minus the cells that become extra-embryonic membranes), into testes of adult mice. Some of the early embryo cells gave rise to teratomas! These induced growths looked and acted like spontaneous teratomas, yielding embryoid bodies when transplanted into mouse bellies and displaying an impressive range of tissue types.

Stevens called these cells from early strain 129 embryos that could support differentiation “pluripotent embryonic stem cells”–the origin of the term “ES cell.” But because these cells could give rise to cancerous as well as normal cells, they became known as embryonal carcinoma, or EC cells.

The rest, as they say, was history. But quite a lot transpired between Stevens’ identification of the developmental potentials of primordial germ cells and inner cell mass cells, and the unveiling of the role of the human versions of these cells as ES cells.

Human ES cells (NHGRI)

Human ES cells (NHGRI)

First, Beatrice Mintz and Karl Illmensee, from the Institute for Cancer Research in Philadelphia, visited Stevens to learn his techniques and borrow mice; then they demonstrated that ES cells could give rise to organisms, not just teratomas. (Their surprise announcement of this feat at a meeting floored Stevens, a story unto itself.) Then Martin Evans at Cambridge University and Gail Martin at the University of California, San Francisco, and their coworkers showed that inner cell masses from normal mice could support development too.

The field detoured into genetically altering mouse ES cells, which evolved into knockout technology with the harnessing of homologous recombination to target the genetic changes. Then in the late 1980s, Brigid Hogan, a professor of cell biology at the Vanderbilt University School of Medicine in Nashville, with Peter Donovan’s group at the National Cancer Institute, reignited interest in primordial germ cells by devising culture methods. These cells became known as embryonic germ (EG) or  embryonic stem-like cells.

It wasn’t until November 1998, with publication of the two human ES cell papers, that the media and public were finally diverted from Dolly the cloned sheep sufficiently to notice this much more powerful technology. Attention centered on the two cell sources, and still does.

James Thomson’s group at the Wisconsin Regional Primate Research Center used inner cell masses from in vitro fertilization clinic “leftovers,” while John Gearhart’s group at the Johns Hopkins University School of Medicine used primordial germ cells from aborted fetuses. And the work that led to their success began in a mouse lab nearly half a century ago, with a man with gifted hands and alert eyes who personifies Louis Pasteur’s oft-quoted observation that “chance favors only the prepared mind.”

P.S. A few words from Dr. Stevens’ daughter, Anne Wheeler:

Leroy old“My dad had a major stroke a few days before he retired. It took years for him to ‘come back’, but he did…much to everyone’s surprise. In the past 25 years, he must have been ‘near death’ at least 50 times…strokes, heart attack, bleeding ulcers, etc. He was tenacious, strong, had good genes (apparently!!), and I think his joie de vie, positive attitude and sense of humor brought him back each time. He finally died of congestive respiratory failure. He died peacefully in his sleep, luckily.

He lived on his own, cut his own firewood, had a garden until about 15 years ago. Then, he moved to a ‘retirement’ place where he lived on his own in his own cottage, until just 9 months ago, when he moved to a memory loss center. Every winter, we traveled to Florida (“not that interesting”- LCS), then Costa Rica and Belize. Watching monkeys, sloths, and the ocean waves were some of his favorite things in life.”

He loved being with his 7 grandchildren…lots of birthday parties, weddings, celebrations.”

I’m honored to have had the chance to interview and learn from this great man, who founded a field.

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Autism Gene Discovery Recalls Alzheimer’s and BRCA1 Stories

AutismDiscovery of a new gene behind autism cleverly combines genetic techniques new and classic.

Autism has been difficult to characterize genetically. It is probably a common endpoint for many genotypes, and is a multifactorial (“complex”) trait. That is, hundreds of genes contribute risk to different degrees, as do environmental factors. Research reports implicate either dozens of genes in genomewide sweeps, or focus on a few genes that encode proteins that act at synapses, such as the neuroligins and neurexins.

FEMALE-ENRICHED FAMILIES
Taking cues from the fact that males with autism outnumber females four to one, females are more severely affected, and siblings of females with autism are more likely to also have the condition than siblings of affected males, a team led by Tychele Turner and Aravinda Chakravarti of Johns Hopkins University School of Medicine searched for candidate genes among 13 “female-enriched multiplex families” — FEMFs – that have two or more girls or women with severe autism. The study was published online March 25 in Nature.

Presumably, causative mutations in the female members of these families would have more severe effects. So identifying genes that stand out in their exomes (the protein-encoding part of the genome) and that make physiological sense – that is, affect the brain – could reveal general steps in the beginnings of autism in the broader population. The researchers describe their approach as “modest numbers of samples of rare extreme phenotypes, in contrast to large numbers of typical cases.”

Autism_awareness_ribbon-20051114EXPERIMENTAL RESULTS CONVERGE
The FEMFs indeed revealed 18 candidate genes, four of which emerged as the strongest. The researchers further tested the most likely gene, CTNND2, because it had turned up in other studies. CTNND2 encodes a protein called delta-2 catenin. A series of experiments then led to the following findings:

CLUE 1: Most mutations in humans delete all or part of the gene.

CLUE 2: Knocking out the gene in mice and zebrafish disrupts synapses. Therefore the mutation’s effect is a loss of a normal function, rather than a gain of a new function – and it affects neurons.

CLUE 3: The gene is expressed at 20 times higher level in human fetal brain cells than in human adult brain cells. (This is consistent with the fact that the brain changes that set the stage for autism begin prenatally.)

CLUE 4: The Allen Brain Atlas identified genes with which CTNND2 interacts. They include the usual suspects – proteins that act at synapses or in neural extensions, and in the actin cytoskeleton –  but also a new role, chromatin modification. This means that absence of CTNND2 protein would affect many genes, a broad stroke that could paint the many manifestations of autism.

armadillo(Aside: A key part of the CTNND2 protein is the “armadillo domain,” a 40-amino-acid repeat important in how an embryo passes signals from outside to inside the cell.)

ECHOES OF ALZHEIMER’S AND BRCA1
The new autism study brilliantly uses a handful of unusual families to open a door to the inner workings of autism. Even though the news release calls the FEMF strategy a “novel approach” and “unconventional method,” it actually continues the tradition that first drew me to study genetics – severe or unusual cases that provide insights into disease mechanisms that affect many.

Two examples come to mind: Alzheimer’s disease and breast cancer.

Auguste Deter, the first recognized patient with Alzheimer's disease

Auguste Deter, the first recognized patient with Alzheimer’s disease

The first recognized case of Alzheimer’s disease was Auguste Deter, who began displaying bizarre behavior when she was in her late forties, in the late 1890s. She would scream piteously for hours, often in the dead of night, and traipse around cocooned in bedsheets, propelled by wild hallucinations and delusions. Auguste also had profound memory loss, unable even to write a simple sentence because she’d forget what had just been asked of her. Yet she had glimpses of self-awareness, saying now and then, “I have lost myself.”

Auguste’s terrified husband took her to the Institution for the Mentally Ill and for Epileptics in Frankfurt, where she came under the care of Dr. Alois Alzheimer in 1901. She died five years later, at age 56. In November of that year, after examining her brain, Dr. Alzheimer gave his now-famous lecture on her condition, which was published in 1911 as “eine eigenartige Erkrankung der Hirnrinde” (“a peculiar disorder of the cerebral cortex”).

Alois Alzheimer

Alois Alzheimer

Alas, Dr. Alzheimer’s meticulous and vivid description was lost to history, even as increasing lifespan revealed many people with forms of the condition that Auguste Deter had.

In 1996, psychiatrist Konrad Maurer rediscovered Alzheimer’s medical records for Auguste Deter, and published an analysis in The Lancet. A year earlier, a team from the University of Toronto had identified the presenilin 1 gene in some families with early-onset Alzheimer’s disease. Then in 2013, researchers discovered that Auguste Deter had a presenilin 1 mutation. (See Comment below, this paper is incorrect. Thank you reader!)

256px-BRCA1_enEven more so than the case of Auguste Deter, the new study on autism using female-enriched families reminded me of the 1990 paper in Science introducing the breast cancer 1 gene, better known as BRCA1. That study sought families enriched for early-onset breast cancer.

Mary Claire King famously trolled for susceptibility genes among 329 members of 23 extended families, who included 146 cases of early-onset breast cancer. For anyone who remembers LOD scores (“logarithm of the odds”), a statistic that shows linkage of a phenotype with a particular part of a chromosome, BRCA1 had a good one – 5.98 – signaling something amiss on chromosome 17. Since then, thousands of women and some men have had BRCA1 tests.

CONNECTING THE DOTS THROUGH TIME
Alzheimer DiseaseThe newfound mutations in CTNND2 that may cause or contribute to autism are rare, as are mutations in presenilin 1 among people with Alzheimer’s disease and mutations in BRCA1 among people with breast cancer. But identifying these genes and their pathogenic variants, in the very few patients who serve as canaries-in-the-coalmine, can illuminate at the molecular level how these diseases begin and develop. And that’s a direct route to treating, or at least slowing or controlling, them.

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Universal Newborn Genome Sequencing and Generation Alpha

Newborn Screening

Imagine the day that genome sequencing of all newborns begins. Instantly two cohorts of people will form: the expanding youngest, with a tremendous amount of personal information stored on a cloud, and the shrinking rest of us, with little knowledge of our genomes.

A century from now, possibly everyone will have access to her or his genome data. But until then, how can we prepare to handle the avalanche of information about what I’d call, if I were a science fiction writer, “generation Alpha?”

My idea of the Alphas is inspired by the 1992 dystopian novel “The Children of Men,” by P.D. James. In 1994, all human sperm suddenly die, and 1995 becomes Year Omega. After that, populations plummet and age in the face of global infertility, with the last remaining people, the Omegas, struggling towards inevitable extinction.

(NHGRI)

(NHGRI)

What will happen in our world, in our society, as the Alphas age?

INITIAL INTENT: DIAGNOSIS

Mining sequenced genomes today has the very best of intentions: ending the “diagnostic odysseys” that patients, typically children with rare or one-of-a-kind diseases, endure. But just as opening a magazine can reveal much more than the article one is looking for, a genome sequence provides hundreds of thousands of gene variants that might mean something about a person’s health, perhaps things totally unexpected.

For now, to restrict dissemination of information to the meaningful, the American College of Medical Genetics and Genomics lists 56 “actionable” secondary findings, a minimal menu of genetic conditions which doctors can prevent or treat that show up while looking for something that could explain symptoms. The list will grow as more genes and their variants are identified, and these conditions have already outgrown their initial designation as “incidental.” They’re important.

Thousands of newborns have already had their genomes sequenced, as part of a handful of projects at major research centers. The actual deciphering can take under a day – a lot better than the decade required to sequence the first human genomes. But our understanding of how genotype becomes phenotype lags far behind the ability to decipher the sequence. The value of an “annotated” genome compared to “raw sequence” is like comparing the plot of To Kill a Mockingbird to a pile of word-size pieces cut from a copy of the book. When it comes to genomes, meaning and context are everything.

ATCG's Image with Group of PeopleBEYOND THE USUAL SUSPECTS

The era of looking for what we already know, the “round up the usual suspects” approach to gene identification and disease diagnosis, will gradually end as more human genome sequences and their interpretations are stored in clouds. Our algorithms will ultimately identify all possible gene variants and their interactions – and what they mean at the whole-body level, the phenotype.

My concern is not those “usual suspects,” the well-studied mutations that lie behind single-gene disorders: cystic fibrosis, sickle cell disease, Huntington disease. I fear the fuzzier genetic information. Genome-wide association studies, for example, identify suites of gene variants that signal a good chance that an illness will happen, but not with the power of a clinical diagnosis based on symptoms and biomarkers. The media often trumpet such findings with a false sense of certainty.

(Note on terminology: “gene variant” is a broader, more politically correct term without the negative connotation of “mutation,” which classically means “change in a gene” from the most common form [“wild type”] in a particular population.)

What I fear most isn’t the use of genome information in predicting or diagnosing disease, but in identifying the harder-to-follow, multifactorial traits that are molded by genes and the environment and therefore much more difficult to trace or quantify: intelligence, personality, temperament, talents. Each gene contributes a small amount and to a differing degree to characteristics that aren’t as neatly predictable as the single-gene, Mendelian disorders like the hemophilias.

Newborn_baby_in_hospital_by_Bonnie_GruenbergWill the idea of genetic determinism – that we are our genes – strengthen as the stockpile of genomic information swells through the population, beginning with the youngest? Will the practice become the ultimate example of paternalism, because newborns didn’t provide permission? As they age, can they choose not to know? Will that even be imaginable, as today it’s difficult to envision or remember a time without the Internet?

Choosing not to know will be especially difficult if others have access to genome information. And who should those others be?

ACCESS TO GENOME INFORMATION

Annotated genome sequences could guide pediatricians in troubleshooting problems, providing a powerful new tool in preventive medicine. At the first birthday, a microbiome analysis might identify children with tendencies towards certain conditions, or with insufficiently challenged immune systems.

(NHGRI)

(NHGRI)

Beyond infancy, will availability of genome information fuel stratification as DNA data better predict who is most likely to benefit from a scarce medical resource, and only the young have that information? Years from now, will I be denied a treatment unless I have my genome sequenced to show that I’m just as likely to benefit as a 16-year-old whose genome has been in the electronic medical record since birth?

In a few years, will posh preschools scan applicants’ genome information to select pupils? Will teachers use it to create compatible study groups, or to identify a tendency to bully and treat such a child like future criminals were punished in the dystopian future of the Tom Cruise film Minority Report?

Will standardized test scores be compared to DNA data to deduce whether students are working up to their potential? Will employers look for genomic red flags, the way they stalk Facebook now for evidence of stupidity? This blog has already discussed DNA and dating.

PRIVACY BREACHES

I’m not sure where all this is heading, but it is coming. Widespread newborn genome sequencing could happen within a decade, experts tell me.

Francis Collins wrote in the Wall Street Journal July 7, 2014: “Over the course of the next few decades, the availability of cheap, efficient DNA sequencing technology will lead to a medical landscape in which each baby’s genome is sequenced, and that information is used to shape a lifetime of personalized strategies for disease prevention, detection and treatment.

Is Dr. Collins’ view too narrow? Genome information can be used for purposes other than healthcare. After all, genetic genealogy is based on using landmarks in genomes to identify individuals.

 (NHGRI)

(NHGRI)

Some may say genome data will be secure, we can control access, and limit how much an individual can know about her or his DNA. But did the top executives of Sony Pictures Entertainment last fall ever imagine that all of the company’s as well as their personal e-mails would rain down on the media from the great iCloud in the sky, in 8 humungous and mortifying data dumps?

At least it can be argued that Jennifer Lawrence’s naked photos wouldn’t have gone everywhere if she hadn’t  sent them to a supposedly safe cloud in the first place. But what about the 11 million customers of Premera Blue Cross, whose clinical records, bank account information, and social security numbers may have been released in a cyberattack in May 2014, reported in the media just two days ago?

Privacy breaches have already hampered DNA research. In 2013, Yaniv Erlich, from the Whitehead Institute and his astute student Melissa Gymrek demonstrated their ability to identify people who’d anonymously donated their DNA to the 1000 Genomes Project. They cataloged the short tandem repeats on Y chromosomes that are used in genetic genealogy and matched them to surnames and public information found on Google, such as state and year of birth. Cross-referencing to DNA sequences of cells at the Coriell Cell Repositories and more sleuthing led to women DNA donors. It’s in Science 339:321, unfortunately behind a paywall. And I’ve heard at genetics meetings about children identified by crossreferencing databases that name their rare diseases and their hometowns.

Is the cat out of the bag for genomes already sequenced?

Is the cat out of the bag for genomes already sequenced?

As with Jennifer Lawrence’s revealing images, DNA sequences will be out there, along with a lot of other identifying information. What can we do to ensure that a Sony situation, health insurance leak, or clever use of public databases doesn’t reveal DNA information on a large scale? Late last year Google took on inexpensive genome sequence storage, although raw data may initially be of limited value.

Can we adequately de-identify people and protect the very DNA data that will lay the groundwork for precision medicine? Will the Alphas be the guinea pigs for genome-control? Maybe precision medicine should stick to storing only clinically relevant DNA information. For now.

At a conference to be held April 8-10 at Children’s Mercy, Kansas City, several research groups sequencing newborn genomes as part of an NIH-funded program will meet to discuss results so far and how the information will be used and protected. That’s a great start to what will certainly be an intriguing and important conversation.

(A version of this post appeared on March 16 at the Biopolitical Times blog at the Center for Genetics and Society.)

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CRISPR Meets iPS: Technologies Converge to Tackle Sickle Cell Disease

sickleResearchers from Johns Hopkins University have teamed two powerful technologies to correct sickle cell disease. Linzhao Cheng and colleagues have deployed CRISPR/Cas-9 on iPS cells to replace the mutant beta globin gene.

 

ACRONYMS AND ABBREVIATIONS

CRISPR conjures up images of fried chicken, but it stands for clustered regularly interspaced short palindromic repeats – short repeated DNA sequences interspersed with areas called spacers, like stutters. The pattern attracts an enzyme, Cas9, which is like a molecular scissors that cuts wherever short RNA molecules called “guide RNAs” take it. (For a fuller description, see http://phys.org/news/2014-03-reverse-liver-disorder-mice-mutated.html.)

(Ernesto del Aguiila, NHGRI)

(Ernesto del Aguiila, NHGRI)

CRISPR directs a natural bacterial immune defense to target genes of interest in a wide variety of species. This “genome editing” tool extends gene therapy by not only delivering healthy copies of genes, but removing the bad ones too.

“iPS” stands for induced pluripotent stem cells. Because they are derived from somatic cells, such as skin fibroblasts, iPS cells can yield other cell types while not irking embryo rights advocates. In a lab dish, human iPS cells can reveal a disease’s start, which researchers from the Harvard Stem Cell Institute first accomplished, for cells from patients with ALS, in 2008.

I’ll leave it to the other science bloggers to explain the details of the new paper. Instead I’ll go back in time, for sickle cell disease has been at key crossroads in the field of human genetics. First, the basics.

HOW SICKLING HAPPENS

Hemoglobin

Hemoglobin

Hemoglobin is the protein that carries oxygen in the blood. It consists of two alpha chains and two beta chains.

Sickle cell disease results from a change in codon six in the beta globin gene of CTC to CAC, which replaces the amino acid glutamic acid with valine. That alteration, under low-oxygen conditions, makes the hemoglobin molecules glom into ropelike cables that make the red blood cells that contain them sticky and deformable, then bends them into rigid, fragile, sickle shapes.

The sharply curved cells lodge in narrow blood vessels, cutting off local blood supplies. Once a blockage occurs, behind it oxygen levels fall and sickling speeds up and spreads. The result is great pain in the occluded body parts, especially the hands, feet, and intestines. Bones ache, and depletion of normal red blood cells causes the overwhelming fatigue of anemia.

DISCOVERY OF SICKLED HEMOGLOBIN

Sickle cell disease was the first genetic illness understood at the molecular level. In 1904, young medical intern Ernest Irons noted “many pear-shaped and elongated forms” in a blood sample from a dental student in Chicago who had anemia. Irons sketched this first view of sickled cells and showed his supervisor, James Herrick. Alas, Herrick published the work without including Irons and has been credited with the discovery ever since.

In 1949, Linus Pauling discovered that hemoglobin from healthy people and from people with the anemia, when placed in a solution in an electrically charged field, moved to different positions. Hemoglobin molecules from the parents of people with the anemia, who were carriers, moved to both positions.

In 1956, protein chemist Vernon Ingram chopped up the 146-amino-acid-long beta globin subunit and meticulously sequenced the pieces, and found the sickle cell mutation. It was a little like finding a typo in a sentence rather than on a page.

GLOBIN CHAIN SWITCHING

Fetal hemoglobin has two alpha chains and two gamma chains.

Fetal hemoglobin has two alpha chains and two gamma chains.

Embryos, fetuses, and postnatal humans have different hemoglobin molecules. Two of the molecule’s four subunits are replaced during development in sync with changing levels of oxygen in the environment. The genes that encode the various subunits sit beside each other on chromosome 11, turned on and off from embryo to fetus through infancy, like moving fingers on a flute change the note.

The idea that fetal hemoglobin is different dates to two chemistry papers, from 1866 and 1888, that noted that the fetal version doesn’t fall apart in base. Then in 1934, a paper in the Journal of Physiology, “On the occurrence of two kinds of haemoglobin in normal human blood,” described the distinction.

In 1983, researchers identified the methylation patterns that switch hemoglobin types on and off (no, epigenetics is not a new field). And for 40 years, researchers have been co-opting globin chain switching to treat various  “hemoglobinopathies.” The drug hydroxyurea, for example, de-represses fetal hemoglobin genes, reducing the chance of severe sickling.

THE MALARIA CONNECTION

The relationship between sickle cell disease and malaria is the textbook example of balanced polymorphism: protection against an infectious disease by being a carrier for an inherited disease. DNA Science has covered other examples and more recently in relation to the Ebola epidemic.

The blood of a person with full-blown sickle cell disease is too thick to accommodate the malaria parasite, and the red blood cells too bent to house them. A carrier has enough sickled cells to quell the parasite, but not enough to block circulation and cause the pain and anemia of the inherited disease.

Anthony Allison, a British doctor with expertise in biochemistry and genetics, was investigating blood types in East Africa in 1949, following up a colleague’s observation that sickle cell disease seemed unusually prevalent there. Allison discovered that up to 40% of the members of 35 tribes had sickle cell disease, and all lived in areas where malaria was endemic.

The distributions of sickle cell disease and malaria in Africa coincide.

The distributions of sickle cell disease and malaria in Africa coincide.

Looking at the coinciding maps of the two diseases, he deduced that sickle cell carriers had an advantage. Sure enough, their red blood cells had far fewer malaria parasites than the cells of people with normal hemoglobin. Dr. Allison concluded, “persons with the sickle-cell trait have a considerable natural resistance to infection with Plasmodium falciparum,” in Transactions of the Royal Society of Tropical Medicine and Hygiene, in 1954.

THE DISASTROUS CARRIER SCREEN OF THE 1970s

Use of the term “sickle-cell trait” that Allison referenced, which means heterozygotes (carriers), was to have tragic consequences two decades later. I told the story in this Science Progress post from 2008:

“Sickle cell testing between 1970 and 1972 was mandatory in 12 states and targeted African-Americans. Misunderstanding was rampant, perhaps because the term “sickle cell trait” for carriers suggested that their condition was somehow visible. What they carried was stigma. At the time, the disease could not be prevented, tested for before birth, or treated. So why identify carriers? Fortunately the discrimination was short-lived, thanks to passage of the National Sickle Cell Anemia Control Act in 1972. By the 1980s, antibiotic treatment and bone marrow transplant became possible for this painful, but variable, disease.”

(NHGRI)

(NHGRI)

I learned off-the-record from a geneticist who would know that a few women committed suicide when told they had the “trait,” but told little else.  The sickle cell testing tragedy was a lesson that reverberates today as we debate how to handle reporting of heterozygote status among secondary (“incidental”) findings in sequenced genomes.

TREATMENTS

To better study sickle cell disease, although quite a lot was already known, in the 1990s investigators created several strains of transgenic mice that made human hemoglobin. Because early mouse models didn’t exactly recapitulate the human clinical scenario, various attempts to create a transgenic pig with human hemoglobin happened, but I can’t locate much other than this 1996 paper about a pig with hybrid hemoglobin bearing human alpha chains and pig beta chains.

Stem cell transplants can cure sickle cell disease, and antibiotics following newborn screening can prevent infections. Painkillers and transfusions help. For potential treatments Clinicaltrials.gov lists gene transfer, various supplements and repurposed drugs, and “laying-on-of-hands” in Africa.

CRISPR/iPS blood cells. (Ying Wang, Johns Hopkins Medicine)

CRISPR/iPS blood cells. (Ying Wang, Johns Hopkins Medicine)

This week’s paper about CRISPR and iPS cells may not add much to what we already know about sickle cell disease, but it might provide a way to manipulate oxygen level and watch the hemoglobin subunits segue from embryonic to fetal to adult. That would be cool. It’s also a great proof-of-concept for CRISPR/Cas-9 to cure a single-gene disease in a patient’s own iPS cells, creating a supply of matched replacement cells. Conclude the researchers, “Our results represent a significant step toward the clinical applications of genome editing using patient-derived iPSCs to generate disease-free cells for cell and gene therapies.”

Bravo!

(Much of the history in this post comes from various editions of my textbook, Human Genetics: Concepts and Applications.)

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The Man Who Ate 25 Eggs a Day

SoftBoiledEggHokkaidoTurnipsGarlicButter_(8364603294)Each morning at the retirement community, the healthy 88-year-old man received a delivery of 25 soft-boiled eggs, which he would consume during his day. This had been his way for many years. He’d had one experience of chest pain that might have been angina, but aside from that, he had a healthy cardiovascular system. He recognized that his only problem was psychological: “Eating these eggs ruins my life, but I can’t help it.

I think of the Eggman, a brief case report from 1991 in the New England Journal of Medicine, whenever “news” of cholesterol’s unsuitability as a one-size-fits-all biomarker resurfaces, as it does every few years and did again just last month.

CHOLESTEROL IS AN IMPERFECT BIOMARKER
Fred Kern Jr., MD, a gastroenterologist at the University of Colorado School of Medicine, heard about the man and saw an opportunity to study individual differences in how diet affects serum cholesterol level. And so the 88-year-old egg eater joined such famous patients as French-Canadian explorer Alexis St. Martin and Henrietta Lacks.

St. Martin accidentally shot himself in the abdomen in 1822, and when a hole remained after surgery, provided a window through which US Army surgeon William Beaumont could observe and document how the stomach lining changes during digestion. And of course the celebrated cellular descendants of Henrietta Lacks’ cervical cancer live on in labs throughout the world.

Cholesterol -- not the enemy?

Cholesterol — not the enemy?

First, Dr. Kern tested the man’s lipid levels, which were normal: 200 milligrams per deciliter total cholesterol and 142 for LDL. Then he compared the extent to which the man’s body compensated for the cholesterol overload in the 25 daily eggs to that of 11 volunteers who had their cholesterol tested under their usual eating habits and 16 to 18 days after adding five eggs a day. The Eggman was too obsessed to try to go without to help the study. Dr. Kern’s analysis, albeit limited, revealed a lot about how our bodies differ in what we do with lipids.

The 11 volunteers on average absorbed 54.6% of the dietary cholesterol while on their normal diets and 46.4% while on the egg diet. But the 88-year-old absorbed only 18% of his dietary cholesterol. Plus, his liver made less cholesterol than is common and shunted more of it into bile acids (which aid digestion) than did the livers of the volunteers, a metabolic triple whammy that Dr. Kern called “extremely efficient compensatory mechanisms.”

Basically the man’s metabolism adjusted to the overload in a way that kept his blood lipid levels both healthy and constant, the very definition of homeostasis. And he likely isn’t alone.

Genes lie behind our biochemistry, including lipid metabolism. The Eggman from 1991 is a perfect example of how personalized/precision medicine can not only identify individuals at elevated risk for specific diseases, but also those who are genetically protected. Despite recent presidential recognition and the breathless news coverage it spawned, precision medicine is hardly a new idea. It’s genetics. And geneticists have known for decades that in some families, people can have very high serum cholesterol levels yet healthy hearts.

Back in 1991, the case report of the Eggman intrigued me because I, too, ate eggs every day when everyone was pushing cereal and low-fat diets, and my cholesterol levels were just fine. Nor did any of my relatives have any sort of heart disease, not even hypertension. But I still felt guilty eating omelets.

TRIGLYCERIDES ARE THE ENEMY

Triglycerides -- the enemy

Triglycerides — the enemy

I continued to eat eggs, and like so many in the US in the 1990s who tried to follow low-fat diets, believing the anti-cholesterol mantra, I gained weight. We were drowning in high-fructose corn syrup, added to everything. But when I dug back into the biochemistry I’d learned in grad school, it quickly became apparent that cutting fats was not at all the best way to lower serum cholesterol or to lose weight. So I began researching the Atkins diet and its low-carb brethren.

In 2004, I began the South Beach Diet. Six months later, my weight and triglycerides had plunged.

Why carbs are bad for blood vessel linings is a matter of logic as well as biochemistry. But it’s a bit circuitous, which may have contributed to the persistent perception that cholesterol is the enemy. It isn’t as simple as what you eat showing up unchanged in your blood or on your frame.

Yes, cholesterol is a major part of the plaque that clogs arteries. It comes from the outside (diet; exogenous) or is made in the body (the liver; endogenous). Here is a good review. I’ll summarize.

Triglycerides, sometimes called just “fat,” consist of one glycerol group bonded to 3 fatty acid tails. They are dismantled during digestion, solubilized by bile acids in the intestines, absorbed into the bloodstream at the intestinal villi, and then the freed fatty acids are used to power muscles or are stored in adipose tissue.

Dietary cholesterol is ultimately ferried to the liver. Meanwhile, the liver is making its own cholesterol — from digested triglycerides. So a low-fat diet can reduce dietary cholesterol, and a statin drug can reduce the liver’s production of cholesterol by blocking the rate-controlling enzyme HMG-CoA reductase.

Gray1086-liverBut consider these facts and relationships:

• Most of the cholesterol in the circulation comes from endogenous production (the liver), not from eating cheeseburgers.

• The liver makes cholesterol from triglycerides.

• 95% of lipids in the diet are triglycerides.

• Carbs (the low-fiber white kind: rice, pasta, potatoes) increase triglyceride levels.

Shouldn’t we be limiting carbs, and not dietary cholesterol? It’s the triglycerides that matter. So how did the vilification of dietary cholesterol arise?

Economics.

Might the anti-cholesterol push have something to do with the $29 billion global market for statins? (Despite its suffering from “severe generic erosion” as patents expire.) Rampant fear of cholesterol is making statin manufacturers a lot of money.

MY ANTI-STATIN STANCE
In January 2012, having not had a check-up in years, I saw a new primary care provider, a nurse practitioner. She did a thorough history, and when my cholesterol came back a few days later at the high end of normal and my HDL not as high as it could have been, she insisted that I needed a statin, stat. That plus a low-fat diet, she admonished.

Atorvastatin40mgWhy?” I answered. “I have zero family history of cardiovascular disease and have no other risk factors. I exercise an hour a day. Low HDL is no longer considered a biomarker. And you mean a low-carb diet, correct?”

She dismissed family history, not the best thing to say to a geneticist. Her refusal to consider my personal risk factors, or lack of them, and her prescribing a drug with rare although serious adverse effects was not something that appealed to me. So, “against medical advice,” I went about my low-carb, statin-free lifestyle.

Three months later, the NP called to pitch statins again. Had I thought it over and changed my mind?

I referred her to an article in a recent Lancet showing that 14 genetic markers combined into a risk score to indicate high HDL – supposedly a sign of hearth health – did not lower heart attack risk, in many patients. This wasn’t news; the report confirmed results of another from 2010. In fact a study of an HDL-boosting drug was halted in 2007 because it actually increased “cardiovascular events.” Perhaps low HDL wasn’t a valid surrogate for heart disease after all? Perhaps the NP should have been reading the medical literature instead of believing drug sales reps, but I suspect she didn’t have the time.

A plaque-lined dissected aorta.

A plaque-lined dissected aorta.

Statins do lower endogenous cholesterol synthesis, and do save lives. But wouldn’t they work best if prescribed only to people who actually need them?

My father, for example, was prescribed a statin at age 84, he too with no risk factors, and he developed muscle pain. That was in 2004. But I just checked the Lipitor package insert, and statins are indeed prescribed to people like him, with no personal or family history of heart disease and the only other risk factor his age — and maybe living into one’s 80s indicates a healthy heart!

How many people are being over- or inappropriately treated by taking these drugs? How many health care providers actually determine an individual patient’s risks? I know health insurers are not big fans of genetic testing, but how many years of one-size-fits-all   statin therapy would it take to equal the cost of genetic tests to direct prescribing?

(NHGRI)

(NHGRI)

A TARGET FOR PRECISION MEDICINE

Lots of labs offer tests for single-gene variants that are important in heart health, such as apolipoprotein E (apoE) and angiotensin converting enzyme (ACE). Panels of tests are available too. GENESIS Center for Medical Genetics, Laboratory of Molecular Genetics screens for 8 genes and Vantari Genetics tests expression of several genes to guide drug selection (pharmacogenomics). GeneDx tests for dozens of single-gene conditions that affect cardiovascular health.

In the past, ordering genetic tests for extremely rare diseases involving lipid metabolism when evaluating an average junk food junkie for statin use made little economic sense.  Lecithin cholesterol acyltransferase deficiency, for example, affects fewer than one in a million people. It causes very low levels of HDL cholesterol and very high total cholesterol – bad news! — but no associated cardiovascular disease.

(NHGRI)

(NHGRI)

But the gene-by-gene approach is headed towards extinction, now that the price of exome sequencing (the 85% of the genome that includes most disease-causing genes) has reached the $1,000 mark. A full genome sequencing can now be done in under a day.

Genome analysis takes longer than sequencing, of course, but cloud storage of DNA data will change that. And since tests for single genes that affect lipid metabolism and cardiovascular disease risk have been around for years, why not develop a probe for the relevant subset of the genome to screen statin candidates? Again, could the money insurance companies save on screening out people whom statins wouldn’t help underwrite the cost of sequencing?

Not knowing much about health insurance, I googled to see if any would cover genetic testing to stratify patients for statin use. I didn’t find any, but was thrilled when a statement from one major insurer popped up about genetic testing to identify people at high risk for the severe adverse effect of myopathy, which keeps some people from taking the drugs. Those on high statin doses face a 6-fold increased risk and those over 65 an additional 4-fold increased risk. Some insurers do cover the cost, or for detecting broken-down muscle in the bloodstream every few months, which seems a little late to me.

Paragraph 1 introduces the gene that affects myopathy risk, SLCO1B1. Paragraph 2 claims that “use of statins is associated with approximately 30% reduction in cardiovascular events in a wide variety of populations.” But with genetic testing, aren’t we talking about individuals, not populations?

The document ends with two statements in both italics and boldface. Important! One, that this stuff is complicated so a patient should speak to a physician (patients are dumb, doctors are smart, geneticists don’t exist). Statement two, at the very end after the consumer presumably understands that genetic testing can avoid terrifically painful muscle degradation, I must quote, because it shattered my belief that the insurer was actually going to cover the cost of SLCO1B1 testing, because they bothered to explain it:

Genetic testing for statin-induced myopathy is considered not medically necessary.” This insurer “does not provide coverage for not medically necessary services or procedures.

So there you go. With pharmaceutical giants pushing drugs like statins to a supposedly genetically homogeneous population via doctors who might not have the time or expertise to explore the finer points of lipid genetics, and with insurers years behind the state of DNA science, we might need the hype surrounding precision medicine after all.

Albertus_Verhoesen_Chickens_and_park_vaseCaveat: I’m not an MD. None of the above is meant as medical advice. See your doctor after doing your own research to learn your disease and drug reaction risks. But I’m going to continue to eat my eggs every morning, exercise an hour or more each day, eat veggies and avoid white carbs – and not worry about my cholesterol.

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Fighting Canavan: Honoring Rare Disease Week

baby maxDuring Rare Disease Week, I turn over DNA Science to a family battling a rare inherited disease.

I’ve been following Max Randell, who has Canavan disease, in my human genetics textbook since he was a preschooler – he’s now 17, thanks to gene therapy. Although Max is bright and cherished and happy, life has not been easy.

A year ago DNA Science covered how Max’s experience has inspired his younger brother Alex to become a neuroscientist. My gene therapy book tells the story of how a handful of Jewish people escaping Nazi murderers in a Lithuanian ghetto carried the disease to the US, although like any inherited disease it isn’t confined to one group.

Canavan disease is an enzyme deficiency that melts away the myelin that insulates brain neurons. When Max was born, life expectancy was about 8 years old. But today, with excellent speech, occupational, and physical therapy and earlier diagnosis as doctors become more genetics-savvy, most patients with Canavan disease live into their teens or even their twenties.

Here, in honor of Rare Disease Day 2015, is Ilyce Randell, one of the truly amazing parents I’ve met over the past few years who is finding the funding that is necessary to understand, treat, and possibly even stop these diseases — even if they cannot be reversed.

“Trying to explain to someone what it’s like to raise, love, and save a child born with a rare disease is difficult, and painful beyond words. The world of rare diseases can be dark, depressing and lonely. To raise awareness about a specific rare disease is to let the world into your life, into your deepest hopes and dreams, and even your darkest fears.

Rare_Disease_Day_2015_200pxMy beautiful son Max was diagnosed at 4 months of age with an extremely rare disease called Canavan. My first foray into raising awareness about Canavan disease was not my best moment.

I was still new to raising a sick child. My baby was about 6 months old and lying in his stroller. We were in an elevator when two women began telling me how gorgeous he was (imagine cupid – chubby, electric blue eyes with a pile of golden curls). They asked if he was tired. I looked at them and said, “No, he’s not tired, he’s dying.” The elevator doors opened and I walked out.

Looking back, I should have let them feel the joy of seeing such a beautiful baby, but I hurt and I wanted them to know that even though Max looked beautiful and healthy, he was indeed dying. I think I also needed to try out saying it aloud to strangers. That’s when I realized this would be a long and painful process, and one day, if he lived long enough, the stroller would be a wheelchair and he would not look so healthy anymore.

MaxAt the moment I learned of Max’s diagnosis, I decided I would do anything necessary to save his life. After countless interviews, appearances on talk shows, raising millions of dollars for research, experimental gene therapy, experimental medicines, and a million trips to Capitol Hill, Max has already beat the odds. He is now 17-and-a-half years old.

Along the way, I’ve been explaining what Canavan disease is to anyone who would listen. Part of that explanation is always to let people know that Canavan is a very rare disease, not rare as in affects under 200,000 people, but rare as in a couple hundred kids. There is really no money for research besides what we as families are able to raise.

momandboysathm2We have been very successful garnering attention and funding for Canavan disease, but there is still a long way to go. It can be a sad place, but with the sadness of rare disease I have found immeasurable love so pure that the light it shines brightens even the darkest corner of the world. This fight is exhausting, but my baby was born with a rare disease. My job as his mom is to raise awareness about Canavan.

Ilyce Randell, Director
Canavan Research Illinois
www.canavanresearch.org
PO Box 5823
Buffalo Grove, IL 60089
1-800-833-2194
info@canavanresearch.org

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GAN Gene Therapy Trial Gets Green Light

Lori and Hannah Sames (Dr. Wendy Josephs)

Lori and Hannah Sames (Dr. Wendy Josephs)

Last Friday, February 13th, Lori Sames couldn’t stop crying as she stared at her screen in a coffee shop near her home in Rexford, New York. The announcement had just gone up at clinicaltrials.gov: NCT02362438: Intrathecal Administration of scAAV9/JeT-GAN for the Treatment of Giant Axonal Neuropathy.

Finally.

Translation: recruitment for participants could begin for a clinical trial to test whether genes could be safely transferred into the spinal cord to treat, or at least arrest, a rare neurological disease, giant axonal neuropathy, or GAN. Lori’s 10-year-old Hannah has the disease, and if it weren’t for the family’s efforts, the clinical trial set to begin could still be many years away.

Lori and her husband Matt have raised more than $6 million to get the GAN clinical trial going, in a race against time to save their daughter Hannah and the other 70 or so kids and young people in the world with the disease. Early on they had selected gene therapy as the way to go.

HHFHannah’s Hope Fund was born in Lori and Matt’s home, shortly after the adorable kinky-haired little girl was diagnosed on March 24, 2008. She had just turned four. I’ve chronicled their journey at DNA Sciencehere and here, as well as in my book The Forever Fix: Gene Therapy and the Boy Who Saved It. Lori came up with “forever fix” during our first conversation years ago.

GAN is like ALS in a child. But people aren’t lining up to pour ice over each other’s heads to raise funds, for if ALS is a zebra amongst the horses that are common diseases, then GAN is a unicorn. The clinical trial announcement provides a good description:

The gigaxonin gene lets the body make a protein, gigaxonin, that nerves need to work. Giant axonal neuropathy (GAN) causes a shortage of functional gigaxonin. Nerves stop working normally. This causes problems with walking and sometimes with eating, breathing, and many other activities. GAN has no cure. GAN can shorten life. Researchers want to see if a gene transfer treatment may help people with GAN.”

hockeyHannah needs a wheelchair now, but she’s still very much Hannah. She beautifully sang the national anthem at several local hockey games in recent weeks.

In an ironic and cruel twist, Hannah won’t be among the first to receive the gene therapy. She has a double deletion mutation, one copy inherited from each parent, and so her body doesn’t make gigaxonin at all, not even an altered version of it. And so introducing healthy gigaxonin into her body could trigger an overwhelming immune response.

In the rare disease community, a triumph for one is a triumph for all. So this week before Rare Disease Day, I’d like to share a few comments about what Hannah’s Hope has accomplished.

From clinical trial director Steven Gray, PhD, from the University of North Carolina School of Medicine, at the annual Hope and Love ball for Hannah’s Hope, February 7:

None of this work would have occurred without the people in this room, and the entire Hannah’s Hope community… While we have come a long way, we have a long way to go. Hannah, and patients like her, who are unique in that they make no detectable protein, cannot participate in the Phase 1 trial. Studies are underway to try options for preventing her immune system from interfering with gene delivery. Also, as we learn more, we have more populations of cells we know we need to target in GAN patients beyond this initial delivery to the central nervous system.

From Lori Sames, announcing the start of the trial:

We will know in late April which immune tolerance protocol Hannah will receive, and we want you all with us, every step of the way. We believe in the power of positive thinking and ask that you focus your positive thoughts and prayers on those children who will soon be receiving gene delivery and on Hannah when she is injected, hopefully before the end of the summer.

Taylor King.

Taylor King

A web of shared experience, support, and hope closely connects the rare disease community. Laura King Edwards’ sister Taylor, 16, is battling Batten disease. Laura blogs at Write the Happy Ending and is writing a memoir about Taylor’s fight (it’s great, I’m an early reader).  She blogged recently about the link between Hannah and Taylor:

Nearly five years ago, when my sister could still sing and talk and walk and eat ice cream cones on hot summer days, my mother [Sharon King] met Steve Gray, a young investigator from the University of North Carolina Gene Therapy Center, at a conference in Bethesda. Since 2008 he’d been working on GAN, an ultra-rare, fatal childhood disease that causes progressive nerve death.

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.

Months later, we drove to Chapel Hill to have dinner with him near his lab. We weren’t ready to take the leap then, but Mom had believed in Steve since the first time she heard him speak about his effort to save children from a monster that turned them into quadriplegics unable to eat or breathe on their own. When I sat across from Steve at our booth in a Franklin Street restaurant that day, I believed in him, too.

Our only mistake was that we didn’t fund Steve more quickly; it took Taylor’s Tale three years to sign on as co-funders of a project modeled after the GAN work, announced on World Rare Disease Day in February 2013. In doing so we followed in the footsteps of Hannah’s Hope Fund. Like my mother, Lori Sames entered the fight against GAN with no medical background. But Lori traveled the world, speaking to PhDs and MDs and biotech executives and raising millions of dollars in hopes of saving Hannah and kids like her.

Clinical trials happen all the time. But this one, for a little-known yet devastating disease that affects fewer than 100 children in the world, would never have happened without the dogged determination of Hannah’s family, who raised $6.5 million from their kitchen in upstate New York. It wouldn’t have happened without the genius and iron will of Steve Gray, who took an idea and turned it into a treatment in 6.5 years. This is light speed in the world of science, particularly for an ultra-rare disease like GAN.

Lori Sames at the Recombinant DNA Advisory Committee meeting.

Lori Sames at the Recombinant DNA Advisory Committee meeting.

I won’t forget the night Lori Sames flew to Charlotte to speak to the Taylor’s Tale board, or the day I took a long lunch to watch the webcast of my mother speaking on behalf of Hannah’s Hope Fund at the National Institutes of Health. That day, the Recombinant DNA Advisory Committee (RAC) granted approval for the GAN trial preparations to continue – a vital step in ensuring a better future for children with GAN.

I don’t think a rare disease has ever met a tougher match than Lori Sames or my mom. Lori and my mom looked their child’s rare disease in the face and said, “You will NOT defeat me. I will NOT sit back and let you take my child without a fight.” And because of the choices they’ve made, there is a light at the end of their respective tunnels.

We’ve poured everything into finding a treatment for infantile Batten disease since Taylor was diagnosed in 2006. I know Taylor’s Tale is a big reason why we are far closer to an answer today than we were then. But Taylor’s light is fading, and I’ve had to come to terms with the fact that the work we’re doing is for future Taylors.

I wish I could do more than send Taylor a hippo,

I wish I could do more than send Taylor a hippo.

So today, I try to focus on the good moments I’m able to steal with my sister. She can’t talk to me any longer, but sometimes I can make her laugh.

Hannah and Taylor are in the researchers’ minds when they’re in the lab, and I think that’s part of what drives them to be so good at what they do. I’ll never forget the first time I visited Steve’s lab at UNC. Photos of sick children decorate his office door. He told me that seeing the faces of those kids motivates him.

And though I understand now that we’re probably too late to save my sister, I still believe in this: that one day soon, because people like Steve Gray were motivated by the faces of kids on their office doors, kids like Hannah and Taylor won’t die young.

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Would Charles Darwin Have Used Tinder?

640px-Charles_Darwin_by_G._RichmondIn honor of Charles Darwin’s birthday today, and VD (Valentine’s Day) looming, I’m thinking about how we choose partners. A mobile dating app called Tinder, it turns out, seems to echo sexual selection, Darwin’s idea expounded in The Descent of Man.

This week’s timely post arose when my synapses connected an article just published in Nature’s Scientific Reports Journal, “Risk sensitivity as an evolutionary adaptation,” Darwin’s birthday, and the “Gray Matter” column in last Sunday’s New York Times by Eli J. Finkel, on Tinder.

The app is simple. Images of zillions of potential dates/mates appear on your phone screen. Swipe to the left to reject, swipe to the right to consider.

Judging_of_Best_Pig_in_ShowAfter observing a Tinder demo, the experience reminded me very much of scanning fruit flies for the phenotypes I was looking for. Or judging pigs at a state fair. Each of us likes certain physical traits, and avoids others. Why? What is the basis of attraction?

SETTLE OR WAIT? A DIGITAL EVOLUTIONARY SCENARIO
DNA Science last ventured into the dating arena with “DNA and Dating: Buyer Beware” a few months ago. So when a news release last week pitched “Evolutionary researchers have determined that settling for ‘Mr. Okay’ is a better evolutionary strategy than waiting for ‘Mr. Perfect,’ I was intrigued, with VD coming up. I requested the paper. (Alas I can’t reprint my classic  “If 50 Shades of Grey Had Been Written By A Biology Textbook Author” in honor of tomorrow’s big film debut here because it is far too titillating for the sophisticated readers of PLOS Blogs. But the link still works.☺)

“Risk sensitivity as an evolutionary adaptation” sets up a simulation in a population of digital organisms. The researchers are Arend Hintze, Randy Olson, and leader Chris Adami from Michigan State University and Ralph Hertwig, a psychologist at the Max Planck Institute for Human Development in Berlin. Hintze got the idea to place his imaginary creatures in an evolutionary setting from Hertwig, on visits home to Germany.

As usual the math in the paper scared me, so I zeroed in on the genetics, and found the study’s premise a little weak: “strong evidence that risk-taking behavior has significant genetic components.” Suspicious of genetic determinism, I checked out the two citations.

Gasterosteus_aculeatus_tüskés_pikóOne study refers to the genetic contribution to risk-taking behavior in the three-spined stickleback, a model organism. That paper, actually a book chapter, “describes some of the challenges in studying the genetic basis of individual differences in risk-taking behavior, arguing new insights will emerge from studies which take a whole-genome approach and which simultaneously consider both genetic and environmental influences on the behavior.” So the genomic link to risk-taking behavior, in fish or us, hasn’t yet been established.

The second paper is in the Quarterly Journal of Economics, but is indeed a genetic investigation. It’s a classic twin study (of humans, not fish) that found a heritability of 20% for “experimentally elicited preferences for risk and giving.” Heritability is often misinterpreted to mean the genetic contribution to a trait. Instead, it refers to the genetic contribution to the variability of a trait in a group. So 20% isn’t much.

But the hypothesis of a relationship between risk-taking and evolution is worth exploring, because we likely give a little more thought to mate choice, Tinder notwithstanding, than do fish.

To explore under what conditions mate choice is random or favors the evolutionary currency of ability to have fertile offspring, the researchers followed the choices of digital organisms put computationally through thousands of generations. (Here’s how you can adopt your own digital organism.)

They measured various things, such as conditions and group size. And they found that situations that prompt risk-taking are rare, perhaps once-in-a-lifetime, with a possible high payoff.

Each digital organism made one lifetime decision: 1 = “the safest gamble” or “live” and 0 = the opposite, death or failure to leave fertile offspring. If it lived, the digital organism passed its status (survivability) to offspring.

“Such a life or death decision is akin to a rare lifetime event that has a large
impact on an individual’s fitness, such as mating and mate competition,” the researchers write. The simulation included a low mutation rate, like in life, so that genomes could change and offer new fodder for selection.

ISHERB_Caveman The startling results were that the digital beings were less likely to take risks in collections of fewer than 150 or so – which gibes pretty well with the size of fledgling pre-human groups a million or so years ago. Experiments also simulated small groups within larger ones. “We found that it is really the group size, not the total population size, which matters in the evolution of risk aversion,” Hintze said. Dating in the neighborhood.

Adami translates the results into modern human behavior. “An individual might hold out to find the perfect mate, but run the risk of coming up empty and leaving no progeny. Settling early for the sure bet gives you an evolutionary advantage, if living in a small group. Primitive humans were likely forced to bet on whether or not they could find a better mate. They could either choose to mate with the first, potentially inferior, companion and risk inferior offspring, or they could wait for Mr. or Ms. Perfect to come around. If they chose to wait, they risk never mating.”

In contrast, with Tinder, one might hold out for perfection because the canvas of choices is huge and ever-expanding.

TELEOLOGY

Talk of evolution is often obfuscated by teleology, “the explanation of phenomena by the purpose they serve rather than by postulated causes,” says a standard definition. The phrase “evolves to,” for example, is a 4-letter word for us biology textbook authors, for it implies intent. Natural selection just happens, although it can be directional.

The digital organism paper, I think, has an underlying hint of teleology. The title of the first, unpublished version I read last week said “risk aversion,” which seems to have been changed to “risk sensitivity,” so perhaps the authors were aware of the issue of teleology.

Doubts aside, the findings got me thinking. I imagine myself, an australopithecine perhaps, living in a group of 150 or so, and dating. I’d be, say, 14 or 15 years old, approaching my peak fertility. Would I have had the mental capacity to evaluate who among my limited choices would most likely give me healthy, fertile kids? Do we have that capacity today?

Wading_mooseThat’s where sexual selection comes in, courtesy of Charles Darwin – the traits that attract us tend to be in fertile individuals. (See this article this article for relevant parts of the tome).

Sexually-selected traits are surrogates for fertility. It’s not as if a woman can peer through a man’s jeans into the labyrinth of his seminal vesicles to see whether his spermatids are maturing into healthy swimmers or not.

A female moose might seek a mate with big antlers.

A female 3-spined stickleback favors a male with a red throat.

800px-Bocca_ippopotamo_bioparcoAncient hippos living in what is now the UK grew to enormous lengths (64 feet!) because size seemingly mattered to the ladies.

So while the new study is intriguing in that it matches up the group size of 150 in affecting risk-taking choice with what’s known about early humans, I think the conclusion leads towards teleology – that we did what we did, and do what we do, for a purpose, to intentionally influence evolution.

ValentinesdaytreeI suspect Darwin might not have married his first cousin Emma had he been aware of the work of his contemporary Gregor Mendel. Or if he’d had Tinder.

Happy Valentine’s Day!

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Remembering The Pre-Vaccine Era: The Diseases of Childhood

poxMany of us of a certain age have vivid memories of the “diseases of childhood.”

We remember missing weeks of school, sky-high fevers, spots and pox, cheeks so puffed from mumps that eating was impossible, for days. Our mothers, for they did most of the parenting back then, would intentionally expose us to sick kids, so we’d get the scourges over with ASAP. The lucky among us made it through with just a pockmark or two.

I had the injected Salk polio vaccine as a toddler, but by the time my sister crunched her pink sugarcube of oral polio vaccine years later, I understood why vaccines were part of life. Protect many and you protect nearly all, because the infection can’t spread. It’s just common sense. Vaccination especially protects kids with chronic diseases, like cystic fibrosis, who can’t be immunized, as well as babies too young to have been immunized.

I posted my own “vaccine memories” last July, ending with research revealing a much more likely cause of autism than vaccines. For this post today, I asked a few friends about their memories of the diseases of childhood. First is Robert Marion, MD, a pediatrician at the Albert Einstein College of Medicine and chief of the division of genetics at the Children’s Hospital at Montefiore. He wrote one of my favorite books, Genetic Rounds: A Doctor’s Encounters in the Field that Revolutionized Medicine.

640px-Polio_vaccine_posterFROM A PEDIATRICIAN
“Until the last few years, the campaigns to immunize children against measles, mumps, rubella, diphtheria, pertussis, tetanus, polio, hepatitis B, Haemophilus influenzae, and others, have been so successful that most young pediatricians have never seen a single case. But these conditions were the bread and butter of pediatric practice prior to 1960.

The measles-mumps-rubella (MMR) vaccine was developed in the early 1970s, but each of the components was developed in the 1960s. Rubella (German measles) vaccine in particular was developed because of the devastation of congenital rubella.

Throughout the mid 20th century, epidemics of rubella raged every couple of years. Although the disease itself was mild, pregnant women contracting the virus were at risk to have children with deafness, blindness, congenital heart disease, failure to thrive, intellectual disabilities (then called “mental retardation”), and….yes, you guessed it…..autism. How ironic is it that the vaccine developed to prevent the leading cause of autism became the focus of this pseudoscientific crusade to prevent all humans from using vaccines? Pretty amazing, huh?

We see a fair amount of pertussis because it causes its real harm in very young children, before their full course of immunizations is complete. This is a life-threatening illness in infants, who cough so much that they can’t take in air. They’re at risk to develop hypoxic-ischemic encephalopathy, permanent brain damage, as a result, and every year, there are deaths from pertussis in the very young. About 10 years ago, I learned first hand that the pertussis vaccine does not bring life-long immunity. My son, a teenager, developed whooping cough and required intensive care for a few days. Not a lot of fun.

When I was an intern in the winter of 1979-1980, I cared for an adolescent girl who was brought into the hospital by ambulance, comatose. A few hours earlier, she’d complained of a headache, low grade fever, and became lethargic, confused, and finally unresponsive. Just like that. Seemingly out of the clear blue.

When we examined her, we noted healing scabs on her trunk and legs. On questioning, her parents (who spoke little English) let us know that she’d recently gotten over chickenpox. ‘But that was over a couple of weeks ago,’ they told us, implying that that illness couldn’t possibly be related to this illness. Of course, it was.

She had post varicella encephalitis, a rare complication of chickenpox. This poor girl’s CT scan showed cerebral edema with loss of myelin. Her spinal tap showed increased intracranial pressure.

We admitted her to the ICU. We put a bolt in her skull to measure her intracranial pressure. We treated her aggressively to keep the pressure in a tolerable range, so that her brain would continue to be perfused with blood, giving her mannitol (a powerful diuretic) every time her intracranial pressure spiked. We stayed on top of her.

She remained in this state, in the ICU, unresponsive to all but the most painful stimuli, for about five days. She then started to “lighten up,” raising her level of consciousness. Her intracranial pressure normalized and we were able to remove the bolt. One week after admission, we transferred her from the ICU to the medical floor.

Measles

Measles

The girl remained in the hospital for nearly a month, and then she was transferred to a pediatric rehab hospital. Because of the demyelination, she had lost significant muscle strength and wasn’t able to walk, barely able to sit up on her own. According to her parents, she had regained many of her cognitive skills, but when she left, they told her that she still “wasn’t herself.” I never saw her again, so I’m not sure whether she ever regained all of her function. But it’s clear that all of this could have been avoided had there been a vaccine.”

FROM NON-DOCS

Even routine cases of the childhood diseases were, at the very least, disruptive. Even if a kid wasn’t particularly ill, the disease spread.

“I had chickenpox in the third grade. I got hundreds, in my nostrils, in my throat, on my scalp, everywhere inside and outside. I couldn’t blink without it hurting. I wore gloves at night so I wouldn’t scratch my face. I couldn’t swallow, so I couldn’t eat. When I was finally better and put on shoes, they were too big because I’d lost so much weight. It was horrific. I was 8.” Sharon P., lawyer in NYC

“In 1960, when my father, who had a withered leg from polio, and my mother took me and my sister to get our oral polio vaccine, a line snaked around the block. Every kid in the neighborhood was there and nobody, but nobody, was talking about not getting it.

In high school, I caught rubella and wasn’t sick, just covered with red dots. It was the last day of the school year and I wanted to get stuff from my locker. On my way out, I stopped to say goodbye to my favorite English teacher. I told her I had German measles, thinking it was a sort of joke, and she just shrank away from me and whispered, “I’m pregnant!” I rushed out of the room and went home and stayed there. I felt awful and never saw her again.” Jennie Dusheck, science writer

“My sister and I both got measles midsummer. We were confined to our bedroom. Mom had to keep the drapes closed and we had to wear sunglasses to protect our eyes. I remember all the red splotches and being very itchy and some sort of lotion being applied. We stayed in that room for 7 days, with high fevers, not feeling well at all, and sleeping a lot. It’s hard to believe any parent would want to put their child through that!

Mumps_PHIL_130_loresI didn’t get mumps, but my mother did! It was during May of my senior year in high school. I had a huge term paper due and Mom was going to type it. I was also preparing to go to my senior prom, and give a speech at the National Honor Society induction ceremony. The day before that event, Mom started to get swelling in her face and throat, but the day of the induction ceremony she wrapped a warm scarf around her neck and attended. She could have infected lots of folks by being there. She went to the doctor’s after that and got the mumps diagnosis, and then was instructed to stay in the house. And she did type my paper!” Phyllis Kovall, music teacher

I don’t usually resort to name-calling, but in the case of refusing vaccines, I bow to a New Yorker cartoon by e. flake that shows a doctor examining a spotted child, in front of two befuddled parents: “If you connect the measles it spells out ‘My parents are idiots.’”

Indeed they are. It isn’t cool to be anti-science, or anti-medicine. It’s dumb. And deadly.

UPDATES

eman and flagsI was gratified to read in The New York Times, The New Yorker and elsewhere about the self-appointed neighborhood public health groups in Liberia who have slowed and possibly halted  the Ebola epidemic. Emmanuel Gokpolu, my young friend in Liberia, led one of those efforts, chronicled in How Ebola Kills and previous posts.

pink-150x150DNA Science covered use of mitochondrial DNA from a third party to create embryos free of mitochondrial disease nearly a year ago, when researchers debated the controversial technology at an FDA hearing. The news this week isn’t about the science, but about renewed discussion in the UK. I’ll get back to mitochondria soon, about a way to possibly fight mitochondrial disease without tinkering with embryos.

 

 

 
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Surveying the Genomic Landscape of Modern Mammals

A rhesus macaque and a Tasmanian devil.

A rhesus macaque and a Tasmanian devil.

A study published today in Cell  compares regulatory DNA sequences among 20 species of modern mammals, showcasing how mammalian genomes have found new uses for ancient genes.

The evolution of mammals has been ongoing for about 180 million years, with a burst of numbers and diversity about 66 million years ago. Then, an asteroid impact led to mass extinctions, and the small, scurrying, hairy ones found vacated real estate.

(NHGRI)

(NHGRI)

Fossils help in studying past life, but even more useful is to compare DNA sequences among modern species. The more of a gene or genome sequence a pair of species shares, the more recently they shared an ancestor. That is a more likely explanation for similarity at the DNA level, I think, than that two species ended up with nearly identical sequences by chance. And if researchers know the mutation rate for a specific gene, then they can assign approximate time frames to species’ divergence.

DISSECTING REGULATORY SEQUENCES
Diego Villar, from the University of Cambridge Cancer Research UK, Paul Flicek, head of Vertebrate Genomics at the European Molecular Biology Lab EMBL-EBI, and their colleagues identified promoter and enhancer regions in DNA from liver cells from the selected 20 species of placental mammals.

Promoters are short sequences at the starts of genes that control their transcription into RNA, from which protein is translated. Enhancers are short DNA sequences, typically located a bit away from the genes that they control, that bring different genes together as the chromosome unwinds and loops about itself. So both promoters and enhancers control gene expression, one from near, one from afar.

Tree shrew (wikispecies)

Tree shrew (wikispecies)

The list of participating mammals in the study is diverse, but heavy on primates, rodents, and sea dwellers: human, macaque, vervet and marmoset (primates); mouse, rat, naked mole rat and guinea pig (rodents); dolphin and two whales (cetaceans); and rabbit, tree shrew, pig, ferret, dog, cow, cat, opossum, and Tasmanian devil.

The researchers discovered that promoters tend to have evolved recently – which means over the past 40 million years. In contrast, most enhancers are derived from sequences that have been around for more than 100 million years, but have been co-opted to take on new functions. So species and their genomes can retain what’s worked in the past via natural selection, but can also tap into ancient sequences for new uses.

I’d use the word “repurpose” to describe enhancers’ links to past DNA, but “purpose” is a banned word in evolutionary biology, because it implies intent, which implies a creator. The researchers use the slightly less teleological “redeployment of ancestral DNA.” This approach is different from the gene duplication route to microevolutionary change I learned in graduate school. In that model a gene doubles, perhaps by a slip during DNA replication at a short repeated sequence, and then one copy, through mutation, acquires a novel function that persists if it does no harm.

An evolutionary success

An evolutionary success

It’s reassuring that a genome has more than one way to change, because that’s what evolution is: change. It is not directional, not leading towards someone’s concept of perfection, and we are certainly no more highly evolved, whatever that means, than a bacterium or cockroach. They’ve been around a lot longer than we have.

In 1982, Stephen Jay Gould and Elizabeth Vrba coined the term  exaptation to distinguish novel use of old information from natural selection, which instead results in an adaptation (a trait that makes it more likely to leave fertile offspring.) Exaptation is a little like what my daughter Heather says whenever I want to buy new furniture. “Use something already in the house.

My husband Larry demonstrating his exaptation of kitchen tools to capture rodents. He is a retired chemist with nearly 100 patents.

My husband Larry has exapted kitchen tools to capture rodents. He is a retired chemist with nearly 100 patents.

ALTERNATE VIEWS OF EVOLUTION
Some popular explanations of biological diversity do not take advantage of the endless information to be mined from DNA sequences. Consider the appearance of everything living within the span of a week, plunked down on the lovely Earth by a supernatural force.

According to Genesis, on day 5, God said, “Let the water teem with living creatures, and let birds fly above the earth across the vault of the sky.” Day 5 would seemingly cover the dolphin and pair of whales featured in the Cell paper, but the biblical description is classification by habitat, so includes fish and birds too.

bibleLet the land produce living creatures according to their kinds: the livestock, the creatures that move along the ground, and the wild animals, each according to its kind,” sayeth God on day 6. Towards the end of that great day, we humans came along to lord over everyone else, a situation evidenced by the terms “livestock” and “wild,” a somewhat more subjective taxonomy than comparing DNA sequences and mutation rates. Presumably this day would include the other species in the Cell paper not yet specifically mentioned.

Another skewed view of evolution is the common line-up of creatures, with humans coming after chimps as the most “advanced” species. Cartoonists sometimes put stooped-over office workers in the final slot, or someone hunched over a cell phone. These alignments are offshoots of the “Great Chain of Being,” a religious ordering of everything in the universe.

HaeckelBut evolution tends to be branching, not linear. We share an ancestor with chimps, from around 6 to 7 million years ago, but we didn’t morph directly from them. Even when a lineage appears to be linear, likely offshoots died out along the way, perhaps leaving a bit of themselves lurking in modern genomes, like the Neanderthals.

So now I’ve angered the anti-GMO folk in last week’s post, and the intelligent design crowd in this week’s post. Perhaps I should take next week off.

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