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been utilized in murine and iPS cell to change specific
nucleotides in a gene sequence without changing their
subsequent differentiation properties. 91,92 As more expe-
rience is gained with gene targeting technology, the pro-
tocols will become easier to perform and require even
less cellular manipulations that could induce unantici-
pated negative consequences.
As a platform for treating OI patients, the remaining
challenge for iPS cells is the development of preimplan-
tation cell culture protocols that direct the cells to a level
of differentiation for optimal in vivo bone formation and
a method for delivery of the cells to the site that will pop-
ulate the patient's bone. Neither of these two challenges
has been adequately answered and until substantial
progress is made the potential for a cell/gene therapy for
OI will not be realized.
cells over the OI cells, the ratio of normal to OI will be
more than the chimerism of non-bony tissue, and the
advantage will indicate the degree of engraftment that
will have to be achieved to positively impact the quality
of bone matrix in the OI subject.
Second, various transplantation strategies for direct
engraftment of bone need to be investigated to achieve a
high level and uniform distribution of normal osteoblasts
within the trabecular and endocortical bone. Initially,
BMSC-derived progenitors should be employed because
of their proven ability to engraft as osteoblasts 85 but even-
tually murine iPS cells should be tested in a similar man-
ner. Unless a technical breakthrough develops that allows
either cell source to traverse the lung and home directly
to bone, it is likely that direct introduction of these cells
into individual bones will have to be the route of admin-
istration. In either case, success needs to be judged by
the degree in improved structural properties of the trans-
planted bone and the degree of normal to OI chimerism
that is achieved in the osteoblast and osteocyte popula-
tion. Other stresses on the transplanted bone could be an
induced fracture designed to assess the ability of the trans-
planted cells to respond to an additional demand on their
progenitor potential. Since fractures require an intact peri-
osteum capable of generating the callus of a fracture, 94,95 a
marrow transplant might not be expected to improve the
initial response to a fracture because it would not alter the
cellular composition of the periosteum.
A third use of murine models to investigate a cell
therapy is in the developing fetus or very early neo-
nate where autograft tolerance can be achieved.
Comprehensive testing using the same criteria as dis-
cussed above for adult transplantation needs to be
employed to demonstrate a correlation between osteo-
blast engraftment and bone strength. Other experiments
using ubiquitously expressed GFP reporters need to be
performed to assess the natural history of the infused
cells that engraft and persist in non-osseous tissues and
whether they develop into cells that would change the
function of the engrafted tissues. If this transplantation
approach can be shown to lead to long-term engraftment
in which the infused cells are the major participants in all
aspects of bone growth, modeling, remodeling and frac-
ture repair, then a strong argument could be made that
we should direct our clinical therapies to in utero diagno-
sis and intervention as the best available option of care.
However, until there is solid preclinical data to demon-
strate this is possible, it is unlikely that many physicians
would elect for this type of intervention.
PRECLINICAL TEST PLATFORMS FOR
CELL THERAPY OF OI
Given the potential of a gene correction strategy for
OI, the need for a safe and effective cell delivery pro-
cedure assumes a high level of priority. This is not the
time to have a clinical disaster from a premature use of
iPS cells set back the steady progress that is being made
with this cell source. The following is a roadmap that this
author believes could lead to the development of a safe
and effective preclinical platform that would be the basis
of human trial for a cell-based therapy for OI.
Develop and Understand a Cell-Based Therapy
in Murine Models of OI
Although there is agreement that the mouse does not
fully model human physiology in many aspects of skel-
etal biology, it has been very effective in understanding
the molecular basis of bone and cartilage biology as well
as transplantation biology. Also, it is the only preclinical
model currently available that is tolerant of long-term
human cell engraftment. Thus the mouse is likely to play
a prominent role as an experimental platform for devel-
oping cell-based treatment of OI.
First, it will be necessary to establish the degree of
mosaicism necessary to suppress the impact of osteo-
blasts carrying an OI mutation. A spectrum of mouse
chimera composed of normal and OI morula cells, each
source also embedded with a bone-specific GFP reporter,
needs to be produced and characterized for the structural
properties of bone. 93 Analysis of the resulting adult ani-
mals will indicate the degree of somatic chimerism that
was achieved during the embryo reconstitution. That
result will be compared to the chimerism of the osteo-
blasts that populate the trabecular, cortical and membra-
nous bone. If there is a selective advantage of the normal
Use of Murine Models to Develop Protocols for
Engraftment by Human Osteoblasts
Although in vitro differentiation is frequently
used to demonstrate the multipotential capability of
 
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