EVENTS | VIEW CALENDAR
DNA: A winding string of achievements
Alicia Keys croons into a studio microphone, headphones firmly in place. Elsewhere, a rapper beats about the unfairness of the world in a smackdown with another rap artist. Two hardhats stand at a construction site, reviewing plans for the building that is growing around them, while a scene that smacks of the movie Tron touts the next-generation wonders of a new cell phone.
What do all of these events have in common? DNA.
DNA headphones. The rapper DNA. The DNA3 condominium complex rising in Toronto. The Droid DNA cellphone from HTC.
Walking down the main drag of Alexandria, Va., one is met with the usual signs. A drugstore. A Starbucks. An antiques shop. A Starbucks. A carpeting center. A Starbucks. And the letters D-N-A. Salon DNA, which not only has the name, but also offers a logo design reminiscent of a sequencing gel.
The new world
Crick, Franklin, Watson and Wilkins could not have imagined any of this when they were generating and analyzing the crystallography data that led to the elucidation of the structure of DNA, work that put a face on what was already known as the core of life: A simple, elegant biochemical staircase that scientists and now laypeople have climbed for six decades to see worlds well beyond the vistas imagined by the intrepid, and often conflicting, quartet.
In his 1962 speech at the Nobel Banquet on behalf of himself, Crick and Wilkins (Franklin passed away in 1958 and was therefore ineligible to be recognized by the Nobel committee), James Watson openly recognized the significance of their work in the broadest of terms.
"At that time, we knew that a new world had been opened, and that an old world which seemed rather mystical was gone," he said.
It is important to remember that the quartet didn't discover DNA, as has commonly been misreported by the media. To be completely frank, much of what was technologically accomplished in the decades since could have been achieved in the absence of a known structure—you didn't need to know what insulin looked like to understand how it works.
The structure elucidation clearly established a visual frame of reference, however, that facilitated an understanding of the genetic landscape and perhaps more importantly, a conceptual framework upon which generations of future scientists could dream.
And dream they did, as shown by the timeline "60 years of DNA milestones," (see table at the bottom of this screen) which highlights just a few of the seminal achievements that followed the structure determination and formed the foundation of the biotechnological era in which we now live and work.
Sequence of events
From its humblest beginnings as a multistep process performed by hand, through its first awkward steps of automation, to its dramatic transition into a high-throughput science, DNA sequencing has become the cornerstone of the genetic era, as all other technological advancements are moot without the understanding of what those sequences do and for what they code.
In a recent interview for the 10th anniversary of the completion of the Human Genome Project, Eric D. Green, director of the National Human Genome Research Institute, gave his perspectives on a decade of technological advances.
"What's happened in the arena of DNA sequencing technology development in the last 10 years has been truly spectacular," he opines. "Go back 10 years. We had just generated that first sequence of the human genome, and the active sequencing part took about six to eight years, consumed about $1 billion—that was about the cost for organizing of the sequencing and actually doing the sequencing.
"Fast-forward 10 years, after these spectacular new technologies have been developed, and we're well under $10,000," he continues. "In fact, the current estimates for getting the sequence of a human genome are something on the order of $3,000 to $5,000 and down to $1,000, I think, within a year or two. And remarkably, today, you could do it in a couple of days, and probably by the end of this calendar year, I am being told, within a day."
For Green, though, the real success of these efforts will come with an improved understanding of the genomic basis for human disease. Until recently, he suggests, this understanding has been limited to rare diseases that center around simple genetic paradigms, rather than more complex diseases with multiple genetic components.
Before the Human Genome Project, he says, we knew the genetic basis of about 60 genes involved in rare diseases. By the end of the project, that number exploded to about 2,200. In the 10 years since then, that number has more than doubled up to almost 5,000.
Not wishing to underplay the rare disease efforts, however, he adds, "what's going on with rare genetic diseases has been truly remarkable."
As Green's comments allude, early efforts to improve sequencing technologies focused on increasing throughput to maximize the amount of samples that could be processed in a single run, as the focus of initiatives like the Human Genome Project was to simply catalogue the broadest spray of genomic sequences.
Now that this has been done—or is at least well on its way—the needs within the industry and in medicine have become more refined and more focused on individuals rather than on populations. To that end, companies such as Illumina have adjusted their next-generation sequencing (NGS) technologies to suit not just genomics centers, but also hospital clinics and smaller field laboratories.
"From a clinical perspective, there is great potential for NGS in the management and treatment of human health," said Richard Tothill and colleagues at Melbourne's Peter MacCallum Cancer Centre, in a 2011 review examining the clinical applications of NGS systems from Illumina, Roche and Life Technologies.
"It is easy to imagine that soon every patient will have both their constitutional and cancer genomes sequenced, the latter perhaps multiple times in order to monitor disease progression, thus enabling an accurate molecular subtyping of disease and the rational use of molecularly guided therapies," the authors added.
As suggested in previous articles in DDNEWS, however, and echoed here by Tothill, the new technologies and data streams will require a re-education of clinician who are currently ill-prepared to act upon the NGS results.
"Protocols for dealing with NGS data that guide what and how particular information will be reported and conveyed to the clinician will need to be established," Tothill says.
While applauding the improvements made in speed, throughput and cost, Barrett Bready, CEO of positional sequencing company Nabsys, suggests still more is required to see NGS reach maximal utility.
"While these advances have been impressive and important, many applications of sequence data—in medicine, as well as in basic biological research and agriculture—require similar levels of improvement in data accuracy, information content, reduced data and computational burden and simplified workflow," he said while he prepared for presentations at the Annual Advances in Genome Biology and Technology meeting held in Florida last year.
Many human diseases are the result of large-scale genomic insertions, deletions or duplications, according to John Thompson, Nabsys' director of assay development, information that can be critical to diagnosing and treating patients. Many such variants, however, can be difficult to detect using standard or next-generation sequencing methodologies.
The Nabsys platform uses nanoscale detectors and specific hybridization probes to generate not only sequence information, but also provide a positional reference for the sequence within the genome. Thus, over scales of hundreds of kilobases to megabases, the sequences can be examined within the context of other DNA segments, allowing for an accelerated assembly of de-novo sequences.
Detection and diagnosis
Aside from efforts to sequence entire genomes at increasingly shrinking costs, there has also been a strong effort in the idea of sequencing genomes at increasingly shrinking scales—perhaps even down to the genome of a single cell.
In the April edition of Genome Research, scientists at the J. Craig Venter Institute published their efforts to recover and sequence the genome from a single cell of Porphyomonas gingivalis, a periodontal pathogen they isolated from a biofilm in a hospital sink. Without culturing and within a biofilm population that included 25 different types of bacteria, the researchers were able to sequence and assemble the genome of one literally microbe. Comparing that sequence to cultured strains, the researchers noted 524 unique genes in the biofilm exemplar, some of which may be involved in virulence.
"A vast majority of bacteria in the environment, as well as those associated with the human microbiome, have eluded standard culturing approaches, and therefore, their physiology and gene content are unknown," the authors write. "This leaves a large gap in our knowledge of the potential roles for these organisms in the environment, and also in human health and disease."
As the recent spate of highly publicized hospital-acquired infections indicates, biofilm research is becoming increasingly important as clinicians and scientists attempt to expand their understanding of how these microbes change in becoming part of a biofilm. This knowledge will hopefully lead to insights on how best to fight both biofilm formation on surfaces such as catheters, sinks and medical instruments, and kill the organisms once part of a biofilm.
"Capturing genomes from environmental samples using single-cell approaches could support studies on the prevalence and genotype of pathogens from environmental sources and may ultimately help reveal their possible modes of transmission between the host and environment," the authors conclude.
Sequencing isn't the only DNA technology that is moving clinical research forward. With increasing pressure to provide companion diagnostics with new therapies, several other molecular workhorses continue to ply their trade, including fluorescent in-situ hybridization (FISH) and PCR.
In January, Epizyme announced its collaboration with Roche to develop a PCR-based companion diagnostic to support its efforts with Eisai to progress its EZH2 target for lymphoma. The goal is to identify patients who carry a mutant form of the enzyme involved in cancer proliferation and then treat those patients with their selective inhibitor.
In announcing the effort, Epizyme President and CEO Robert Gould said, "Epizyme is committed to the creation and commercialization of personalized therapeutics and companion diagnostics for patients with genetically defined cancers."
Following through on that pitch, Epizyme in April announced a partnership agreement with Abbott to develop a companion diagnostic for its mixed lineage leukemia candidate EPZ-5676, an inhibitor targeting the DOT1L histone methyltransferase. Under the agreement, Abbott will develop FISH assays to identify patient samples that include oncogenic mutations of DOT1L and identify eligible patients for the inhibitor.
Meanwhile, Dako in March announced it received U.S. Food and Drug Administration (FDA) approval for its HER2 IQFISH pharmDx platform as a companion diagnostic for Genentech's HER-2 positive metastatic breast cancer treatment Kadcyla, an antibody-drug conjugate derivative of Herceptin.
In discussing personalized medicine with DDNEWS last year, Henrik Winther, Dako's vice president of corporate business development, said, "in my personal opinion, I could easily foresee that in seven to eight years' time, you will see no oncology drug being prescribed without having a companion diagnostic attached to it. Looking at the flow of diagnostics tests performed in a pathology lab today, I could also foresee a significant change in favor of companion diagnostics. More and more patient cases are being referred to prognostic and predictive assays simply because you want to be able to provide better treatment and prognosis to the patients."
Spell me a solution
Of course, from a human health perspective, the holy grail of the genomic revolution remains the ability to go into the human body and correct disease-causing errors at their roots: gene therapy.
After some modest successes and high-profile failures in the 1990s—the most famous of the latter being the death of Jesse Gelsinger in 1999—gene therapy research efforts continued, but largely took a back seat to other therapy development efforts. As our understanding of therapy vectors has improved over the intervening years, however, gene therapy is looking at something of a renaissance.
Last November, uniQure's Glybera became the first gene therapy product to be approved by the European Commission. Designed as a treatment for lipoprotein lipase deficiency, Glybera uses an adenoviral vector to introduce a variant of the human lipoprotein lipase gene into patients, facilitating the metabolism of fat-carrying particles in the bloodstream that might otherwise obstruct small blood vessels and can cause acute pancreatitis.
"This therapy will have a dramatic impact on the lives of these patients," said Glybera researcher John Kastelein of the University of Amsterdam. "Currently, their only recourse is to severely restrict the amount of fat they consume. By helping to normalize the metabolism of fat, Glybera prevents inflammation of the pancreas, thereby averting the associated pain and suffering, and if administered early enough, the associated co-morbidities [early-onset diabetes and cardiovascular complications]."
Although the initial push for gene therapy was largely limited to orphan conditions that offered few other treatment options, it is also starting to make clinical progress in the treatment of more widespread conditions.
In March, Japan's AnGes MG announced it received FDA approval on its Special Protocol Assessment (SPA) for its global Phase III study of Collategene, a gene therapy product developed in collaboration with Vical. The agreement hopefully paves the way for success of the trial in critical limb ischemia and thereby opens the door for future regulatory approval.
A month later, researchers at Celladon and Imperial College London announced the initiation of the CUPID2 trial of Celladon's Mydicar gene therapy, an AVV-mediated delivery of the gene for SERCA2A directly into the heart to reverse heart failure and improve heart function.
According to Alexander Lyon, consulting cardiologist from Royal Brompton Hospital and an Imperial College lecturer, "Heart failure affects more than three-quarters of a million people across the U.K. Once heart failure starts, it progresses into a vicious cycle where the pumping becomes weaker and weaker as each heart cell simply cannot respond to the increased demand. Our goal is to fight back against heart failure by targeting and reversing some of the critical molecular changes arising in the heart when it fails."
A legacy of vision
The ripples of the DNA revolution continue to be felt down the biological stream with an 'omics for every biomolecule available, providing ready fodder for publications such as DDNEWS.
Outside of the lab and outside of the clinic, however, the democratization of DNA continues as wider swathes of society embrace its potential, both real and metaphoric.
"We must continue to work in the humane spirit in which we were fortunate to grow up," Watson concluded his Nobel speech. "If so, we shall help insure that our science continues and that our civilization will prevail."
This was clearly an understatement as their science has not only prevailed, it has flourished and evolved in ways the original quartet could never have realized.