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HMGN1 plays a significant role in CRLF2 driven Down Syndrome leukemia and provides a potential therapeutic target in this high-risk cohort

Abstract

The genetic basis of the predisposition for Down Syndrome (DS) patients to develop cytokine receptor-like factor 2 rearranged (CRLF2r) acute lymphoblastic leukemia (ALL) is currently unknown. Genes located on chromosome 21 and expressed in hematopoietic cells are likely candidates for investigation of CRLF2r DS-ALL pathogenesis. We explored the high-mobility group nucleosome-binding protein 1 (HMGN1), located in the DS critical region, in an inducible CRISPR/Cas9 knockout (KO) xenograft model to assess the effect of HMGN1 loss of function on the leukemic burden. We demonstrated HMGN1 KO-mitigated leukemic phenotypes including hepatosplenomegaly, thrombocytopenia, and anemia, commonly observed in leukemia patients, and significantly increased survival in vivo. HMGN1 overexpression in murine stem cells and Ba/F3 cells in vitro, in combination with P2RY8-CRLF2, resulted in cytokine-independent transformation and upregulation of cell signaling pathways associated with leukemic development. Finally, in vitro screening demonstrated successful targeting of P2RY8-CRLF2 and HMGN1 co-expressing cell lines and patient samples with fedratinib (JAK2 inhibitor), and GSK-J4 (demethylase inhibitor) in combination. Together, these data provide critical insight into the development and persistence of CRLF2r DS-ALL and identify HMGN1 as a potential therapeutic target to improve outcomes and reduce toxicity in this high-risk cohort of young patients.

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Fig. 1: Knockouts of JAK2 and HMGN1 were generated in SET-2 CRLF2 p.F232C and Jurkat cell lines by CRISPR/Cas9.
Fig. 2: HMGN1 knockout reduces CRLF2 p.F232C leukemic burden in vivo.
Fig. 3: Knockout of HMGN1 rescues leukemic phenotype and increases survival outcomes.
Fig. 4: P2RY8-CRLF2 and HMGN1 cooperate for cytokine-independent growth.
Fig. 5: Characterizing the signaling profile when HMGN1 is overexpressed in CRLF2+ Ba/F3 cells.
Fig. 6: Effect of GSK-J4 treatment on cell signaling of Ba/F3 cell lines expressing CRLF2 and HMGN1.
Fig. 7: Effective targeting of Ba/F3 cell lines expressing CRLF2 and HMGN1 with fedratinib and GSK-J4 combination therapy.

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Data and materials availability

Additional data and requests for resources should be directed to the lead contact, Deborah White (deborah.white@sahmri.com). Materials can be obtained via material transfer agreement from authors’ institutions upon reasonable request to corresponding authors.

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Acknowledgements

Flow cytometry analysis and cell sorting were performed at the South Australian Health Medical Research Institute (SAHMRI) in the ACRF Cellular Imaging and Cytometry Core Facility. The Facility is generously supported by the Detmold Hoopman Group, Australian Cancer Research Foundation, and the Australian Government through the Zero Childhood Cancer Program. Animal models were performed in the Bioresources Core Facility at SAHMRI and we would like to acknowledge the technical support provided. The authors acknowledge the facilities and scientific and technical assistance of the National Imaging Facility, a National Collaborative Research Infrastructure Strategy (NCRIS) capability, at the Preclinical Imaging and Research Laboratories/Bioresources, South Australian Health and Medical Research Institute. This study has been performed as partial fulfillment of the requirement for a Ph.D. degree from the University of Adelaide Faculty of Sciences for E.C.P. Funding for this study was provided by NHMRC, Beat Cancer, and L.F.A. E.C.P. was supported by an RTP University of Adelaide scholarship.

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E.C.P., S.L.H., and D.L.W. conceived of and designed the experiments. B.J.M., P.Q.T., and D.L.W. provided all study materials. C.E.D. and S.O. performed and analyzed murine stem cell experiments. E.C.P. collected, assembled, and analyzed the data and wrote the manuscript. D.L.W., S.L.H., P.Q.T., L.N.E., B.J.M., C.E.D., S.O., D.T.Y., and T.P.H. critically appraised the manuscript. All authors gave final approval of the manuscript.

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Correspondence to Deborah L. White.

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Competing interests

D.L.W. receives research support from BMS, and Honoraria from BMS and AMGEN. D.T.Y. receives research support from BMS & Novartis, and Honoraria from BMS, Novartis, Pfizer, and AMGEN. T.P.H. receives research support from BMS & Novartis, and Honoraria from BMS, Novartis, and Fusion Pharma. None of these agencies have had a role in the preparation of this manuscript. All other authors declare no conflicts of interest.

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Experiments with mice were conducted according to the guidelines of the South Australian Health and Medical Research Institute animal ethics committee. Ethical approval was obtained from each institutional Human Research Ethics Committee and parents or legal guardians gave informed, written consent. This study was conducted in accordance with the Declaration of Helsinki.

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Page, E.C., Heatley, S.L., Eadie, L.N. et al. HMGN1 plays a significant role in CRLF2 driven Down Syndrome leukemia and provides a potential therapeutic target in this high-risk cohort. Oncogene 41, 797–808 (2022). https://doi.org/10.1038/s41388-021-02126-4

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