NIH:Scientists first described the sickle-shaped red blood cells that
give sickle cell disease its name more than a century ago. By the 1950s,
the precise molecular and genetic underpinnings of this painful and
debilitating condition had become clear, making sickle cell the first
“molecular disease” ever characterized. The cause is a single letter
“typo” in the gene encoding oxygen-carrying hemoglobin. Red blood cells
containing the defective hemoglobin become stiff, deformed, and prone to
clumping. Individuals carrying one copy of the sickle mutation have
sickle trait, and are generally fine. Those with two copies have sickle
cell disease and face major medical challenges. Yet, despite all this
progress in scientific understanding, nearly 70 years later, we still
have no safe and reliable means for a cure.
Recent advances in CRISPR/Cas9 gene-editing tools, which the blog has highlighted in the past,
have renewed hope that it might be possible to cure sickle cell disease
by correcting DNA typos in just the right set of cells. Now, in a study
published in Science Translational Medicine, an NIH-funded
research team has taken an encouraging step toward this goal [1]. For
the first time, the scientists showed that it’s possible to correct the
hemoglobin mutation in blood-forming human stem cells, taken directly
from donors, at a frequency that might be sufficient to help patients.
In addition, their gene-edited human stem cells persisted for 16 weeks
when transplanted into mice, suggesting that the treatment might also be
long lasting or possibly even curative.
CRISPR/Cas9 technology uses small RNA
molecules to guide a scissor-like enzyme to a specific DNA sequence.
This DNA editing tool allows researchers to target a gene in a stem cell
and snip the precise spot where an error in the sequence occurs. Once
the double-stranded DNA is cut, the disease-causing typo can be replaced
with the correct sequence, allowing the stem cell to produce healthy
normal cells that can potentially cure the condition.
Lab studies had shown that CRISPR/Cas9 could be used to correct
sickle cell mutations in experimental cell lines. But modifying a
sufficient number of blood-forming stem cells in hopes of treating
patients with their own edited cells had proved a far greater technical
challenge. While some editing was achieved in a small fraction of cells,
those edited cells tended to disappear over time for reasons that
aren’t yet well understood.
In the new study, Jacob Corn at the University of California,
Berkeley, and colleagues sought to find a more reliable and efficient
method. Using human stem cells isolated from whole blood samples, they
started by using an electric shock to open up pores in the cell
membrane. With those pores open, the researchers could introduce the
CRISPR/Cas9 complex into the cells along with bits of guide DNA encoding
the correct hemoglobin sequences.
Corn and his colleagues capitalized on the fact that the cutting
enzyme hangs on to one of the two DNA strands for a time, leaving the
second strand free. Using their knowledge of the sequence contained
within that loose bit of DNA, they designed short and inexpensive DNA
“fillers” to come in and correct the sequence to the normal state with a
high rate of success.
Using this method, the researchers found that they could replace the
incorrect sequence in more than a third of human stem cells. Those
cells, which had started out with two copies of the sickle mutation,
went on to produce red blood cells containing healthy hemoglobin.
However, given the challenges of obtaining blood-forming stem cells from
people with sickle cell disease, they didn’t have enough corrected
human cells to transplant into mice for testing.
To get past this limitation, the researchers turned the protocol
around to generate sickle-trait stem cells. They took stem cells from
healthy patients and edited in the sickle cell mutation. When about a
million of those CRISPR/Cas9-treated cells were transplanted into mice, a
small but significant number—2 to 4 percent—of the human stem cells
took up residence in the animals’ bone marrow and persisted for at least
16 weeks. That’s good news because previous studies have suggested that
fixing the sickle cell gene in just 2 to 5 percent of bone marrow stem
cells could be enough to benefit patients.
Genetic blood diseases are prime candidates for potentially
therapeutic gene-editing approaches because a patient’s cells can be
easily accessed, edited, and then replaced. Sickle cell disease is an
obvious first choice, in part because the condition affects millions of
people around the world—100,000 in the United States alone [2]. But
other even rarer single gene disorders of blood cells, including various
types of anemia and immune deficiencies, are also fair game.
As promising as these results are, much more work is needed in the
laboratory before the new approach can move forward for a possible
clinical trial to treat people with sickle cell disease. In the nearer
term, Corn and colleagues hope their success will inspire researchers in
labs around the world to explore gene editing for other challenging
health conditions.