GENETIC CORRECTION OF HEMATOPOIETIC STEM CELLS IN SITU

SPACE
PROVIDED.
Project summary: Knowledge of the molecular basis of blood and immune disorders along with the
availabiltiy of technologies for the genetic manipulation and phenotypic correction of hematopoietic stem
cells provides novel therapeutic opportunities. For example, near universal bone marrow failure in patients
with Fanconi Anemia (FA) results in considerable morbidity and mortality. This coupled with the limited
availability of suitable HLA-matched donors for curative allogeneic stem cell transplantation provides strong
rationale to target autologous hematopoietic stem cells for stable genetic correction. Conventional
approaches to stem cell modification most often utilize in vitro culture of target cell and viral or non-viral
vector systems. This approach is not ideal for HSC from FA patients where a pro-apoptotic tendency and
repopulation deficiency conspire to limit therapeutic chimerism from genetically corrected cells. Alternatively,
dissemination and biosafety concerns present substantial impediments to intravenous injection of vector. I
propose to develop a novel strategy for the systemic in situ delivery of HIV-derived retrovirus particles, to
reconcile barriers to vector delivery and stem cell survival with the therapeutic potential of retroviral HSC
modification. The overall hypothesis underlying my proposal is that the prolonged persistence of vector
particles in hematopoietic cells combined with the inherent homing mechanisms of these cells can provide
for targeted cellular delivery of vector particles to the bone marrow microenvironment. This innovative
strategy is supported by the following observations. 1) Recent studies from our laboratory have revealed the
unexpected uptake and delayed release of HIV vector particles from hematopoietic target cells and their
retained ability for secondary transduction. 2) Hematopoietic cells possess cell surface molecules that direct
their distribution after intravenous injection to the bone marrow. 3) Related approaches in cancer gene
therapy provide precedent for the cellular delivery of particles to xenogenic tumor grafts in mice. The aim of
my project is to elucidate the mechanistic basis for particle delivery, to provide proof of principle that in situ
correction can restore HSC alkylator resistance in Fancc-/- mice, and to limit vector dissemination to non-
targeted tissues.
Relevance: The proposed research is a pre-clinical study that is of direct relevance to curative therapeutic
approaches to single gene disorders affecting hematopoietic stem cells.