DNA: A winding string of achievements

Examining 60 years of scientific progress since the discovery of DNA

Randall C Willis
Alicia Keys croons into a studio microphone, headphonesfirmly in place. Elsewhere, a rapper beats about the unfairness of the world ina smackdown with another rap artist. Two hardhats stand at a construction site,reviewing plans for the building that is growing around them, while a scenethat smacks of the movie Tron toutsthe 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 complexrising in Toronto. The Droid DNA cellphone from HTC.
Walking down the main drag of Alexandria, Va., one is metwith 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 notonly has the name, but also offers a logo design reminiscent of a sequencinggel.
The new world
Crick, Franklin, Watson and Wilkins could not have imaginedany of this when they were generating and analyzing the crystallography datathat led to the elucidation of the structure of DNA, work that put a face onwhat was already known as the core of life: A simple, elegant biochemicalstaircase that scientists and now laypeople have climbed for six decades to seeworlds well beyond the vistas imagined by the intrepid, and often conflicting,quartet.
In his 1962 speech at the Nobel Banquet on behalf ofhimself, Crick and Wilkins (Franklin passed away in 1958 and was thereforeineligible to be recognized by the Nobel committee), James Watson openlyrecognized the significance of their work in the broadest of terms.
"At that time, we knew that a new world had been opened, andthat an old world which seemed rather mystical was gone," he said.
It is important to remember that the quartet didn't discoverDNA, as has commonly been misreported by the media. To be completely frank,much of what was technologically accomplished in the decades since could havebeen achieved in the absence of a known structure—you didn't need to know whatinsulin looked like to understand how it works.
The structure elucidation clearly established a visual frameof reference, however, that facilitated an understanding of the geneticlandscape and perhaps more importantly, a conceptual framework upon whichgenerations of future scientists could dream.
And dream they did, as shown by the timeline "60 years ofDNA milestones," (see table at the bottom of this screen) which highlights just a few of the seminalachievements that followed the structure determination and formed thefoundation of the biotechnological era in which we now live and work.
Sequence of events 
From its humblest beginnings as a multistep processperformed by hand, through its first awkward steps of automation, to itsdramatic transition into a high-throughput science, DNA sequencing has becomethe cornerstone of the genetic era, as all other technological advancements aremoot without the understanding of what those sequences do and for what theycode.
In a recent interview for the 10th anniversary of thecompletion of the Human Genome Project, Eric D. Green, director of the NationalHuman Genome Research Institute, gave his perspectives on a decade oftechnological advances.
"What's happened in the arena of DNA sequencing technologydevelopment in the last 10 years has been truly spectacular," he opines. "Goback 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 $1billion—that was about the cost for organizing of the sequencing and actuallydoing the sequencing.
"Fast-forward 10 years, after these spectacular newtechnologies 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 aresomething on the order of $3,000 to $5,000 and down to $1,000, I think, withina year or two. And remarkably, today, you could do it in a couple of days, andprobably by the end of this calendar year, I am being told, within a day."
For Green, though, the real success of these efforts willcome with an improved understanding of the genomic basis for human disease.Until recently, he suggests, this understanding has been limited to rarediseases that center around simple genetic paradigms, rather than more complexdiseases with multiple genetic components.
Before the Human Genome Project, he says, we knew thegenetic basis of about 60 genes involved in rare diseases. By the end of theproject, that number exploded to about 2,200. In the 10 years since then, thatnumber 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 trulyremarkable."
As Green's comments allude, early efforts to improvesequencing technologies focused on increasing throughput to maximize the amountof samples that could be processed in a single run, as the focus of initiativeslike the Human Genome Project was to simply catalogue the broadest spray ofgenomic sequences.
Now that this has been done—or is at least well on itsway—the needs within the industry and in medicine have become more refined andmore focused on individuals rather than on populations. To that end, companiessuch as Illumina have adjusted their next-generation sequencing (NGS)technologies to suit not just genomics centers, but also hospital clinics andsmaller field laboratories.
"From a clinical perspective, there is great potential forNGS in the management and treatment of human health," said Richard Tothill andcolleagues at Melbourne's Peter MacCallum Cancer Centre, in a 2011 reviewexamining the clinical applications of NGS systems from Illumina, Roche andLife Technologies.
"It is easy to imagine that soon every patient will haveboth their constitutional and cancer genomes sequenced, the latter perhapsmultiple times in order to monitor disease progression, thus enabling anaccurate molecular subtyping of disease and the rational use of molecularlyguided therapies," the authors added.
As suggested in previous articles in DDNEWS, however, and echoed here by Tothill, the new technologiesand data streams will require a re-education of clinician who are currentlyill-prepared to act upon the NGS results.
"Protocols for dealing with NGS data that guide what and howparticular information will be reported and conveyed to the clinician will needto be established," Tothill says.
While applauding the improvements made in speed, throughputand cost, Barrett Bready, CEO of positional sequencing company Nabsys, suggestsstill 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 biologicalresearch and agriculture—require similar levels of improvement in dataaccuracy, information content, reduced data and computational burden andsimplified workflow," he said while he prepared for presentations at the AnnualAdvances in Genome Biology and Technology meeting held in Florida last year.
Many human diseases are the result of large-scale genomicinsertions, deletions or duplications, according to John Thompson, Nabsys'director of assay development, information that can be critical to diagnosingand treating patients. Many such variants, however, can be difficult to detectusing standard or next-generation sequencing methodologies.
The Nabsys platform uses nanoscale detectors and specifichybridization probes to generate not only sequence information, but alsoprovide a positional reference for the sequence within the genome. Thus, overscales of hundreds of kilobases to megabases, the sequences can be examinedwithin the context of other DNA segments, allowing for an accelerated assemblyof de-novo sequences.
Detection anddiagnosis 
Aside from efforts to sequence entire genomes atincreasingly shrinking costs, there has also been a strong effort in the idea ofsequencing genomes at increasingly shrinking scales—perhaps even down to thegenome of a single cell.
In the April edition of GenomeResearch, scientists at the J. Craig Venter Institute published theirefforts to recover and sequence the genome from a single cell of Porphyomonas gingivalis, a periodontalpathogen they isolated from a biofilm in a hospital sink. Without culturing andwithin a biofilm population that included 25 different types of bacteria, theresearchers were able to sequence and assemble the genome of one literallymicrobe. Comparing that sequence to cultured strains, the researchers noted 524unique genes in the biofilm exemplar, some of which may be involved invirulence. 
"A vast majority of bacteria in the environment, as well asthose associated with the human microbiome, have eluded standard culturingapproaches, and therefore, their physiology and gene content are unknown," theauthors write. "This leaves a large gap in our knowledge of the potential rolesfor these organisms in the environment, and also in human health and disease."
As the recent spate of highly publicized hospital-acquiredinfections indicates, biofilm research is becoming increasingly important asclinicians and scientists attempt to expand their understanding of how thesemicrobes change in becoming part of a biofilm. This knowledge will hopefullylead to insights on how best to fight both biofilm formation on surfaces suchas catheters, sinks and medical instruments, and kill the organisms once partof a biofilm.
"Capturing genomes from environmental samples usingsingle-cell approaches could support studies on the prevalence and genotype ofpathogens from environmental sources and may ultimately help reveal theirpossible modes of transmission between the host and environment," the authorsconclude.
Sequencing isn't the only DNA technology that is movingclinical research forward. With increasing pressure to provide companiondiagnostics with new therapies, several other molecular workhorses continue toply their trade, including fluorescent in-situhybridization (FISH) and PCR.
In January, Epizyme announced its collaboration with Rocheto develop a PCR-based companion diagnostic to support its efforts with Eisaito progress its EZH2 target for lymphoma. The goal is to identify patients whocarry a mutant form of the enzyme involved in cancer proliferation and thentreat those patients with their selective inhibitor.
In announcing the effort, Epizyme President and CEO RobertGould said, "Epizyme is committed to the creation and commercialization ofpersonalized therapeutics and companion diagnostics for patients withgenetically defined cancers."
Following through on that pitch, Epizyme in April announceda partnership agreement with Abbott to develop a companion diagnostic for itsmixed lineage leukemia candidate EPZ-5676, an inhibitor targeting the DOT1Lhistone methyltransferase. Under the agreement, Abbott will develop FISH assaysto identify patient samples that include oncogenic mutations of DOT1L and identifyeligible patients for the inhibitor.
Meanwhile, Dako in March announced it received U.S. Food andDrug Administration (FDA) approval for its HER2 IQFISH pharmDx platform as acompanion diagnostic for Genentech's HER-2 positive metastatic breast cancertreatment Kadcyla, an antibody-drug conjugate derivative of Herceptin.
In discussing personalized medicine with DDNEWS last year, Henrik Winther, Dako'svice president of corporate business development, said, "in my personalopinion, I could easily foresee that in seven to eight years' time, you willsee no oncology drug being prescribed without having a companion diagnosticattached to it. Looking at the flow of diagnostics tests performed in apathology lab today, I could also foresee a significant change in favor ofcompanion diagnostics. More and more patient cases are being referred toprognostic and predictive assays simply because you want to be able to providebetter treatment and prognosis to the patients."
Spell me a solution
Of course, from a human health perspective, the holy grailof the genomic revolution remains the ability to go into the human body andcorrect disease-causing errors at their roots: gene therapy.
After some modest successes and high-profile failures in the1990s—the most famous of the latter being the death of Jesse Gelsinger in1999—gene therapy research efforts continued, but largely took a back seat toother therapy development efforts. As our understanding of therapy vectors hasimproved over the intervening years, however, gene therapy is looking atsomething of a renaissance.
Last November, uniQure's Glybera became the first genetherapy product to be approved by the European Commission. Designed as atreatment for lipoprotein lipase deficiency, Glybera uses an adenoviral vectorto introduce a variant of the human lipoprotein lipase gene into patients,facilitating the metabolism of fat-carrying particles in the bloodstream thatmight otherwise obstruct small blood vessels and can cause acute pancreatitis.
"This therapy will have a dramatic impact on the lives ofthese patients," said Glybera researcher John Kastelein of the University ofAmsterdam. "Currently, their only recourse is to severely restrict the amountof fat they consume. By helping to normalize the metabolism of fat, Glyberaprevents inflammation of the pancreas, thereby averting the associated pain andsuffering, and if administered early enough, the associated co-morbidities[early-onset diabetes and cardiovascular complications]."
Although the initial push for gene therapy was largelylimited to orphan conditions that offered few other treatment options, it isalso starting to make clinical progress in the treatment of more widespreadconditions. 
In March, Japan's AnGes MG announced it received FDAapproval on its Special Protocol Assessment (SPA) for its global Phase IIIstudy of Collategene, a gene therapy product developed in collaboration withVical. The agreement hopefully paves the way for success of the trial incritical limb ischemia and thereby opens the door for future regulatoryapproval.
A month later, researchers at Celladon and Imperial CollegeLondon announced the initiation of the CUPID2 trial of Celladon's Mydicar genetherapy, an AVV-mediated delivery of the gene for SERCA2A directly into theheart to reverse heart failure and improve heart function.
According to Alexander Lyon, consulting cardiologist fromRoyal Brompton Hospital and an Imperial College lecturer, "Heart failureaffects more than three-quarters of a million people across the U.K. Once heartfailure starts, it progresses into a vicious cycle where the pumping becomesweaker and weaker as each heart cell simply cannot respond to the increaseddemand. Our goal is to fight back against heart failure by targeting and reversingsome 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 downthe biological stream with an 'omics for every biomolecule available, providingready fodder for publications such as DDNEWS.
Outside of the lab and outside of the clinic, however, thedemocratization of DNA continues as wider swathes of society embrace itspotential, both real and metaphoric.
"We must continue to work in the humane spirit in which wewere fortunate to grow up," Watson concluded his Nobel speech. "If so, we shallhelp insure that our science continues and that our civilization will prevail."
This was clearly an understatement as their science has notonly prevailed, it has flourished and evolved in ways the original quartetcould never have realized. 
(click here for the rest of this Special Report on the history of DNA and a look toward the next 60 years...)

Randall C Willis

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