Career goals: Dr. Agarwal's long term goal is to elucidate novel molecular mechanisms in leukemia pathogenesis so that this knowledge can be applied to the discovery of new therapies resulting in improved clinical outcomes. Dr. Agarwal's career goal is to establish an independent laboratory focused on understanding cancer pathogenesis. For the K99 Pathway to Independence Award, Dr. Agarwal has described a five-year training program that will allow her to acquire the expertise needed to become a successful independent cancer researcher. During this award, she will be able to expand her knowledge in cancer biology, acquire expertise in a variety of novel techniques to be used in her future research, and generate a body of data as a basis for future studies and applications for extramural funding. Knowledge acquired and data produced during the K99/R00 award will be instrumental in achieving her long term goal. Environment: The Oregon Health &Science University Knight Cancer Institute has 165 primary faculty investigators with expertise across a diverse spectrum of fields. Dr. Agarwal's mentor, Dr. Brain Druker, is the Director of the Knight Cancer Institute. Dr. Druker has over 20 years of experience in the field of cancer research and has mentored numerous students and fellows to independent investigator status. He will continue to provide intellectual and leadership training to Dr. Agarwal for the transition to a career as an independent researcher. Dr. Agarwal has gathered an excellent team of collaborators and advisors with the combined expertise to advise her on all aspects of the proposed study as well as on her career. In addition, the Knight Cancer Institute's infrastructure provides research training and enthusiastic support to post-doctoral fellows who are preparing themselves for a successful faculty career. Dr. Agarwal believes that the proposed project is well-suited to launch her independent research career in the field of leukemia pathogenesis. Research: The objective of the proposed study is to determine the mechanism of p27 deregulation in chronic myeloid leukemia (CML) and to establish the role of p27 in CML pathogenesis. CML is a disease of hematopoietic stem cells caused by BCR-ABL, a constitutively active tyrosine kinase that is the result of the 9;22 translocation. Most patients with early stage disease achieve durable responses upon treatment with imatinib, a small-molecule inhibitor of BCR-ABL. However, in the advanced stages of disease, drug resistance and relapse are frequent. Further, recurrence of active disease is common if therapy is stopped. Therefore identification of additional molecular targets suitable for therapeutic intervention may allow us to develop novel treatment strategies designed to overcome disease resistance and eradicate residual disease. Earlier studies showed that BCR-ABL promotes unregulated cell cycle progression and cell proliferation by impairing the function of p27, a tumor suppressor and a regulator of cyclin dependent kinase. However, the detailed mechanism of p27 deregulation remains to be clearly defined. Dr. Agarwal's preliminary data suggests that, in primary CML cells, p27 deregulation involves both downregulation in the nucleus and increased cytoplasmic mislocalization. While nuclear p27 appears to be under the control of BCR-ABL kinase activity, imatinib fails to decrease cytoplasmic p27 levels, suggesting that this process is regulated in a kinase-independent fashion. The low nuclear-to-cytoplasmic p27 ratio in CML progenitors is reminiscent of findings in several types of solid cancers, where such ratios are associated with a poor prognosis. Dr. Agarwal proposes that increased cytoplasmic p27 levels contribute to BCR-ABL- mediated leukemogenesis in CML. In support of this, Dr. Agarwal shows that lack of p27 decreases disease latency in a murine CML model, while experimentally reducing cytoplasmic p27 levels with forced nuclear localization of p27 prolongs survival of leukemic mice. These findings are consistent with a tumor suppressor function of nuclear p27 and a concomitant oncogenic function of cytoplasmic p27. These results also suggest that restoring nuclear p27 expression and reducing cytoplasmic expression may counteract BCR-ABL-induced cellular transformation. In total, Dr. Agarwal's preliminary findings have led to a hypothesis in which BCR-ABL disrupts p27 function by simultaneously inhibiting its nuclear tumor suppressor function and promoting its cytoplasmic oncogenic function. This hypothesis will be tested by three carefully designed specific aims that utilize both in vitro and in vivo approaches: 1) Dr. Agarwal will apply molecular and cellular biology tools to determine the mechanisms by which BCR-ABL upregulates cytoplasmic p27. 2) She will dissect the role of nuclear and cytoplasmic p27 for BCR-ABL-driven leukemogenesis using p27 transgenic murine leukemia models. 3) She will delineate the signaling mechanism(s) by which cytoplasmic p27 mediates cellular transformation of human CML cells by testing the effect of cytoplasmic p27 on invasion and survival pathways. Dr. Agarwal's work will lead to an improved understanding of the role of p27 in CML pathogenesis and provide a platform for developing new approaches to treat CML. Since disruption of physiological p27 function is a common theme in human cancers, findings from this study may have implications beyond CML.
I propose that loss of nuclear p27 tumor suppressor function and a concomitant increase in cytoplasmic p27 oncogenic activity promote leukemogenesis in chronic myeloid leukemia (CML). Therefore, elucidating the mechanism of p27 deregulation and dissecting the differential roles of cytoplasmic and nuclear p27 in CML will lead to an improved understanding of disease pathogenesis and provide a rational strategy to restore p27 function in CML and other cancers.