Megan McNerney, MD, PhD
Assistant Professor, Department of Pathology, University of Chicago
In Vivo Characterization of a Tumor Suppressor Associated with High-Risk Myeloid Neoplasms
In vivo characterization of a tumor suppressor associated with high-risk myeloid neoplasms
CUX1 is a transcription factor gene frequently inactivated in myeloid cancers and certain solid tumors. Myeloid cancers with loss of CUX1 are difficult to treat and carry a poor prognosis.
The proposed research will define the phenotypes and pathways in hematopoietic cells that are altered by CUX1 inactivation and may therefore identify new therapeutic targets that ultimately improve the outcome for patients with these diseases.
2015 Interim Report
Young Investigator Award
SUMMARY: CUX1 is transcription factor gene frequently inactivated in myeloid cancers and certain solid tumors. Myeloid cancers with loss of CUX1 are difficult to treat and carry a poor prognosis. The proposed research will define the phenotypes and pathways in hematopoietic cells that are altered by Cux1 inactivation and will therefore identify new therapeutic targets that ultimately improve the outcome for patients with these diseases.
The Aim of this proposal is to identify the impact of Cux1 haploinsufficiency on hematopoietic lineage growth and how it exerts tumor suppressor activity.
Publications during first year of this award: I have had a productive publication record over the last year. In the British Journal of Haematology, I reported the spectrum of mutations that co-occur with loss of CUX1 in patients with myeloid neoplasms. I found that oncogenic RAS mutations occur in almost half of these patients. RAS provides a new therapeutic target that we can test in the mouse model that I have developed as a direct result of the CRF award. I co-authored a paper with Bob Grossman’s group which describes the analysis of “big data” on a novel computing infrastructure. I co-authored a manuscript in the journal, Oncogene, by utilizing the genomics data I generated on high-risk myeloid neoplasms. I also contributed to a high-profile paper in Nature Communications, where we report that genetic interactions between cancer mutations can predict patient prognosis.
1. McNerney ME, Brown, CD, Peterson, AL, Banerjee, M, Larson, RA, Anastasi, J, Le Beau, MM, and White, KP. (2014) The spectrum of somatic mutations in high-risk acute myeloid leukemia. British Journal of Haematology. 166:550-556.
2. Heath, AP, Greenway, M, Powell, R, Spring, J, Suarez, R, Hanley, D, Bandlamudi, C, McNerney, ME, White, KP, and Grossman, RL (2014) Bionimbus: A cloud for managing, analyzing and sharing large genomics datasets. Journal of the American Medical Informatics Association. 21:969-975.
3. Voce, D, Schmitt, A, Uppal, A, McNerney, ME, Wahlstrom, J, Bernal, G, Nasiri, A, Yu, X, Crawley, C, White, KP, Weichselbaum, R, and Yamini, B. (2014) NF-kB1 is a haploinsufficient, DNA damage-specific tumor suppressor protein. Oncogene. Epub ahead of print.
4. Wang, X*, Fu, AQ*, McNerney, ME, and White, KP. (2014) A quantitative epistasis map of cancer genes reveals an association between genetic interaction and patient survival. Nature Communications. 5:4828. *Authors contributed equally.
Grants received during first year of this award: I am pleased to report I have had an successful year in grant funding. The CRF award enabled me to bridge the period from my post-doctoral studies to independent investigator, and seeded the research that served as preliminary data for several of these awards. In total, I received $1,036,000 (direct costs) in extramural funding to support my laboratory over the next five years. In addition, I am a co-investigator on two collaborative grants I helped write with Dr. Michelle Le Beau, totaling $1,650,000 in direct costs over five years.
1. K08 Mentored Research Scholar Grant. 1K08CA181254-01A1. PI: M. McNerney. My role: PI. Title: “Determining the role of CUX1 in myeloid neoplasia.” Total direct costs: $811,000. Project period: 9/1/14-8/30/19.
2. V Foundation Scholar Award. PI: M. McNerney. My role: PI. Title: “Transcriptional misregulation in high-risk myeloid neoplasms.” Total direct costs: $200,000. Project period: 11/01/2014-10/31/16.
3. Edward P. Evans Foundation. PI M. Le Beau. My role: Co-investigator. Title: “Synergistic Interaction of the Microenvironment with del(5q) MDS Stem Cells.” Total direct costs: $400,000. Project period: 7/1/2014-6/30/2016.
4. NIH/NCI 1 R01 CA190372-01. PI: M. Le Beau My role: Co-investigator. Title: “Molecular Mechanisms of Myeloid Suppressor Genes on Chromosome 5”. Total direct costs: $1,250,000. Project period: 9/1/14-10/31/19.
5. Riviera Country Club United-4 A Cure. PI: M. McNerney. My role: PI. Title: “Determining the pathways to high-risk acute myeloid leukemia.” Total direct costs: $25,000. Project period: 02/22/14-02/21/15.
Research progress: Partly as a result of this grant, I have received a strong institutional commitment to my independent research program. I have been promoted to Assistant Professor, as of July 1, 2014. I have received a competitive start-up package to support my salary, a technician, and a post-doc for the next three years, in addition to equipment and supplies. I have obtained approximately 650 square feet of wet-laboratory space, a tissue culture room, an office, and two cubicles for dry-lab work. The wet-lab space has three bays, with six benches and six desks, with room to expand. Over the summer I fully equipped my laboratory to perform all the necessary techniques necessary for my research, including biochemical, molecular, and tissue culture equipment and reagents. I hired a Senior Research Technologist, who has two masters degrees, thirteen years of academic research experience, and has been integral to setting up the lab. A Research Professional joined this month, who just finished a very successful post-doc in murine hematopoietic stem cell biology, has many publications, and is now going to dedicate her effort to the work supported by this CRF award.
Over the last year, we have made substantial progress in developing the tools and data necessary to complete the Aims in this proposal.
2016 Final Report
Young Investigator Award
CUX1 is transcription factor gene frequently inactivated in myeloid cancers and certain solid tumors. Myeloid cancers with loss of CUX1 are difficult to treat and carry a poor prognosis. The proposed research was to define the phenotypes and pathways in blood cells that are altered by Cux1 inactivation and therefore identify new therapeutic targets that ultimately improve the outcome for patients with these diseases.
The Aim of this proposal was to identify the impact of Cux1 haploinsufficiency on hematopoietic lineage growth and how it exerts tumor suppressor activity.
Summary. I am delighted with the progress that my laboratory has made on this proposal. We developed state of the art genetically modified mice (shCux1) in which we can inactivate Cux1. We have shown that the mice are indeed behaving as expected, in that they are inactivating Cux1 in blood cell tissues. We have found that the mice are showing signs of blood disease, as we hypothesized. These mice will be used in future experiments as a model for high-risk myeloid neoplasms in humans, to identify new therapies to treat patients.
SubAim 1.1 Identify the role for Cux1 in hematopoietic differentiation. As a direct result of funding from this proposal, we have generated two separate lines of transgenic mice expressing
an shRNA targeting Cux1 (shCux1) and obtained a line expressing a control shRNA. We have extensively validated that the shCux1 mice are indeed expressing the transgene in all blood cell lineages and that knockdown of the Cux1 protein is occurring at the mRNA and protein level in hematopoietic lineages. Studying these mice, we have found that Cux1 is essential for normal development of the mice, if we inactivate Cux1 during development. Due to this finding, we have chosen to knockdown Cux1 in adult mice. We have discovered that shCux1 adult mice have enlarged spleens, likely due to blood cell expansion, as we hypothesized. We have expanded the colony sufficiently to determine what cell types are expanding, using flow cytometery and analysis of all blood cell compartments. We have also launched experiments to follow the mice long term to determine when they develop myeloid neoplasia. We are currently crossing the mice to NRAS G12D mice, as my work has shown that RAS mutations and CUX1 deletions co-occur frequently in high-risk myeloid neoplasia. It is anticipated that NRAS x shCux1 mice will develop myeloid neoplasia that accurately models human disease. In future experiments, we will use these mice as preclinical models for identification of novel therapeutics.
SubAim 1.2 Determine the impact of Cux1 haploinsufficiency on myeloid cell proliferation
As indicated in SubAim 1.1, we have discovered that shCux1 adult mice have enlarged spleens, which is often seen in the setting of blood cell proliferation and expansion. To determine if Cux1 haploinsufficient cells have increased self-renewal capacity, we are performing serial replating, colony forming assays in vitro, using bone marrow from shCux1 and control mice. Preliminary data suggests that Cux1 haploinsufficiency does increase the self-renewal capacity of hematopoietic stem cells. Preliminary data confirms this finding in human acute myeloid leukemia cell lines in vitro. As discussed in SubAim 1.3, we have identified a cell cycle gene signature associated with CUX1 expression levels, further supporting the role of Cux1 in proliferation.
SubAim 1.3 Establish critical elements of the Cux1 dosage-dependent transcriptional targets.
I successfully recruited a computational post-doc to help with these analyses and we have uncovered some interesting results. Compared to other transcription factors, CUX1 has unique DNA binding characteristics. More often than not, it does not bind to its cognate DNA binding motif. It is associated with activating chromatin marks and has a strong association with SWI/SNF components. This suggests that CUX1 may recruit SWI/SNF to remodel chromatin to influence gene expression. We confirmed that CUX1 directly interacts with multiple SWI/SNF components by CUX1 co-ip followed by mass-spec. We hope to perform some confirmatory experiments of this model and publish the findings of this work within the year. We identified CUX1 regulated genes in primary human HSCS by shRNA mediated knockdown of CUX1 in CD34+ human cells. This experiment was performed in triplicate with two independent shRNAs targeting CUX1. We identified 405 differentially expressed transcripts (FDR <0.05). 90% of the genes go down after CUX1 knockdown, strongly suggesting that CUX1 is predominantly a transcriptional activator, in contrast to prior studies in different cell types. Several pathways were differentially influenced by CUX1 knockdown. We are particularly interested in following up on the finding that genes that coordinate HSC quiescence are decreased, suggesting that CUX1 may play a role in stem cell quiescence.
Publications during second year of this award: During Year 1 of this proposal I published one first author paper and co-authored three additional manuscripts. During Year 2 of this proposal I co-authored two manuscripts on the subject of myeloid neoplasia. I co-authored one book chapter on the subject of genomic abnormalities in cancer.
1. Zhao, Z, Chen, C, Rillahan, CD, Shen, R, Kitzing, T, McNerney, ME, Diaz-Flores, E, Zuber, J, Shannon, KM, Le Beau, MM, Spector, MS, Kogan, SC, and Lowe, SW. (2015) Cooperative loss of RAS feedback regulation drives myeloid leukemogenesis. Nature Genetics. 47:539-43.
2. Stoddart, A, Qian, Z, Fernald, AA, Bergerson, RJ, Wang, J, Karrison, T, Anastasi, J, Bartom, ET, Sarver, AL, Wolff, L, McNerney, ME, Largaespada, DA, and Le Beau, MM (2015) Retroviral insertional mutagenesis identifies the del(5q) genes, CXXC5, TIFAB and ETF1, as well as the Wnt pathway, as potential targets in del(5q) myeloid neoplasms. Under review.
3. Vanderweele, D, McNerney, ME, and Le Beau, MM. Chapter 8: Chromosomal aberrations in human cancer. Holland-Frei Cancer Medicine, 9th edition. (2015) Edited by Bast, RC, Croce, CM, Hait, W., Hon, WK, Kufe, DW, Pollock, RE, Weicheselbaum, RR, and Holland, JF. John Wiley & Sons, Inc., Hoboken NJ.
Grants received during second year of this award: I am pleased to report that in addition to the $1,036,000 (direct costs) in extramural funding I received during Year 1 of this proposal, in the last year I have obtained additional funding support. In summary, I have secured sufficient funding to support my research program for the next four years.
University of Chicago Cancer Research Foundation Auxiliary Board. PI: M. McNerney. My role: PI. Total direct costs: $150,000. Project period: 1/01/2015-1/01/2018.
Video: Presentation to the CRF
In Vivo Characterization of a Tumor Suppressor Associated with High-Risk Myeloid Neoplasms