Friday, March 24, 2023
3/24/2023
PROJECT SUMMARY/ABSTRACT The major shortcoming of current cancer therapies is the inability to target the leukemic stem cells, despite initial success in eliminating the bulk of leukemic cells. Primary myelofibrosis (PMF) belongs to the group of myeloproliferative neoplasms (MPNs), the fourth most prevalent hematological malignancy. In the United States, ~100,000 people are living with or in remission from MPNs. The most frequent driver mutation is JAK2V617F. JAK2 inhibitor ruxolitinib remarkably alleviates constitutive symptoms, but does not prolong survival or reverse disease, which progresses to myelofibrosis or transforms to acute myeloid leukemia. Mouse models and patient data suggest that this shortcoming is due to its inability to target the JAK2V617F+ stem cell. This application addresses the urgent need to develop treatments that specifically target JAK2V617F+ stem cells to improve the long-term outcome of current therapies. Previous work on the SERCA K01 award revealed de-regulated expression of integrins, adhesion molecules that integrate the extracellular environment with the actin cytoskeleton, in megakaryocytes of JAK2V617F transgenic mice and PMF patients. The central hypothesis of this proposal is that ¿1 integrin is a key mediator of JAK2V617F+ stem cell maintenance in PMF. In our most recent preliminary studies, we found higher activation of ¿1 integrin in stem cells of JAK2V617F transgenic mice compared to wild-type (WT) controls. Treatment with an anti-¿1 integrin inhibitory antibody HM¿1-1 in vivo was able to dramatically reduce the number of JAK2V617F+ stem cells, with negligible effect on WT animals. However, it is critical to understand what makes the JAK2V617F+ stem cells particularly susceptible to ¿1 integrin inhibition, and whether the stem cell reduction persists long-term. AIM 1 of this proposal, of mechanistic significance, will define how the anti-¿1 integrin antibody HM¿1-1 reduces JAK2V617F+ stem cells in vivo, by addressing two hypotheses: 1) that induction of cell cycle leads JAK2V617F+ stem cells to exit quiescence and differentiate, and 2) that HM¿1-1 induces apoptosis and cell death in JAK2V617F+ stem cells. AIM 2, of translational significance, will address the hypothesis that JAK2V617F+ stem cell reduction upon HM¿1-1 treatment persists long-term, and will lead to reduction of all progeny derived from JAK2V617F+ stem cells. Studies will be performed on JAK2V617F transgenic mice, carrying the human mutation, using state-of-art flow cytometric analysis of hematopoietic stem cells. Importantly, despite its essential role in organism development, conditional deletion of ¿1 integrin in hematopoietic cells of adult mice did not affect blood cell production or maintenance of stem cells. In contrast, as our data suggest, JAK2V617F+ stem cells seem to be particularly dependent on integrin signaling and highly sensitive to its inhibition, offering a possible therapeutic opportunity window. This proposal addresses the unmet need of identifying a molecular target specific to JAK2V617F+ stem cells with potential for therapeutic development. The AIMS of the proposal will provide important preliminary data which will serve as a basis for mechanistic studies to be proposed in an ensuing R01 grant application in support of applicant’s independent research career.
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