March 2020, ScientificAmerican.com 51molecular properties that determine drug delivery and distribu-
tion into tissues, and the sequence of bases necessary to target a
specific gene. Different sequences of bases make the information
contained in the drugs distinct, but antisense drugs with the
same chemical modifications tend to behave in similar ways in
the body. “That’s what allows us to move quickly once a platform
is established to deliver to a tissue of interest,” says Jonathan
Watts, a nucleic acid chemist at the University of Massachusetts
Medical School. “By shuffling the sequence of bases, we can dial
in a totally different target using the information from a genome-
sequencing experiment of a patient with a rare disease or from
the genome databases. Being able to use that information intui-
tively and rationally is very powerful.”
A LONG-DISTANCE RUN
the idea of using genetic information to make a drug that could
bind to RNA has been around since 1978. But there were a host of
unanswered questions: How do you make an oligonucleotide into
a drug? Why would binding to RNA produce an effect? Nonethe-
less, the idea was intriguing enough to Crooke that in 1989 he left
his position as head of research and development for SmithKline
(now GlaxoSmithKline) to establish a company dedicated to the
development of antisense technology. He was joined there by his
wife, Rosanne, also a pharmacologist, and by colleagues, including
Bennett and Monia. (Originally called Isis, the company eventually
changed its name, for obvious reasons, to Ionis Pharmaceuticals.)
A handful of other companies started up to pursue antisense
around the same time, but one by one they abandoned the hunt.
The leader of one, Michael Riordan of Gilead Sciences, an -
nounced in 1995 that antisense did not work. For a time it did
seem that the problems of toxicity, off-target effects and a lack
of potency might not be overcome.
But Crooke and his colleagues doggedly solved the scientific
problems one at a time. A long, high wall of patents at Ionis’s
headquarters near San Diego attests to their work. First they
had to develop the necessary chemistry. For example, by modify-
ing a key position (2′) in ribose sugar in the RNA and DNA of
ASOs, they were able to enhance the affinity of the ASOs for RNA
receptors, thereby dramatically reducing the necessary dose.
Other chemical modifications improved safety and tolerability.
They also found that the drugs were not taken up into tissue
when delivered directly into cells in culture, but Ionis scientists
made the leap to testing the drugs in animals anyway. Monia,
who ran drug development for Ionis, vividly remembers the
moment when he looked at a chemical test he was using to mea-
sure levels of a specific RNA and saw almost no trace of it—the
drug had entered cells in most tissues, and they had successfully
knocked down the RNA’s expression.
Time spent working on cancer did not prove all that fruitful,
Bennett says. (Promising, more carefully designed experiments
are in the pipeline, however.) What did work were drugs with
specific targets, usually for rare diseases, for which proof of con-
cept is easier to establish. The earliest ASOs were for diseases of
the eye and, later, the liver, where uptake works particularly
well. The drugs were effective, but they were ultimately not com-
mercially viable, because better solutions came along.
The newest oligonucleotide drugs are designed to tackle rare
diseases. One is Exondys 51, which targets Duchenne muscular
dystrophy, a severe, progressive degenerative disease caused by
mutations in the gene that produces the protein dystrophin.
Annemieke Aartsma-Rus of Leiden University Medical Center
in the Netherlands, who is president of the Oligonucleotide
Therapeutics Society, is an expert in Duchenne and helped to
develop the drug. It has been less spectacular than Spinraza,
but on the strength of early results showing increased dystro-
phin levels, the drug received accelerated regulatory approval.
The company marketing it (in which Aartsma-Rus has a stake)
will need to show by 2021 that it makes a meaningful difference
in how a patient functions.
The first RNAi drug, Onpattro, made by Boston-based bio-
tech company Alnylam Pharmaceuticals, was approved in 2018
for treating a hereditary form of nerve damage. An approved
Ionis ASO drug called Tegsedi treats the same thing. The focus
now for all oligonucleotide therapies is delivering more drug
more productively to more parts of the body. “A lot of people
were in wait-and-see mode,” Aartsma-Rus says. “They now see
that if they don’t start, they’ll have missed the boat.”HOPE FOR THE BRAIN
for a Long time antisense companies largely ignored neurologi-
cal targets because oligonucleotides generally do not cross the
blood-brain barrier. But Bennett thought that delivering them
directly to the cerebrospinal fluid via lumbar puncture might
work. He pushed a skeptical Crooke to let him try. “I had a lot of
reservations, but the idea is to say yes,” Crooke says. “ ‘No’ never
made a drug, and ‘no’ never made anybody better.” They started
exploratory studies with a mouse model of Huntington’s, an
obvious candidate for ASOs because it is directly linked to a
specific mutation. People with Huntington’s carry a repeated
sequence of a triplet of base pairs, CAG, that results in toxic lev-
els of huntingtin protein and causes the progressive breakdown
of brain cells. In mice, Bennett and his colleagues found that
they could reduce levels of the mutant protein. “The mice actu-
ally improved,” Bennett says.
Meanwhile Krainer was investigating SMA. Others had dis-
covered that healthy people have two versions of a critical motor
neuron gene, SMN1 and SMN2, but the latter makes very little
functional SMN protein. People with SMA do not have a func-
tional SMN1 gene, and their broken copy of SMN2 cannot do the
job itself. Stretches of DNA include both “exons,” the coding
sequences that are expressed (hence the “ex” in their name), and
“introns,” the noncoding stretches between exons. A process
called RNA splicing joins the exons together and discards the
introns. The SMN2 gene had a variation that rendered it inac-
tive by causing a particular coding chunk, ex on 7, to be ignored.
Krainer and Bennett surmised that an ASO could force that
instruction chunk to be included. By 2008 they had shown that
the ASO they had created worked in mice by fixing the splicing
defect. The clinical trials in humans followed.
“This is what’s called a disease-modifying therapy,” Krainer
says of Spinraza. “It isn’t just dealing with some symptoms. It’s
getting at the root cause of the disease and changing its course.”
Early intervention is critical. A person with symptoms, such as
Emma Larson, has already lost some motor neurons, which
cannot be restored. But the treatment can prevent the remain-
ing neurons from dying off and bring improvements in motor
function. The success in treating infants has led to a push for
newborn screening for SMA, which now occurs in 16 states.© 2020 Scientific American