Abstract
Venetoclax (VEN) inhibits the prosurvival protein BCL2 to induce apoptosis and is a standard therapy for chronic lymphocytic leukemia (CLL), delivering high complete remission rates and prolonged progression-free survival in relapsed CLL but with eventual loss of efficacy. A spectrum of subclonal genetic changes associated with VEN resistance has now been described. To fully understand clinical resistance to VEN, we combined single-cell short- and long-read RNA-sequencing to reveal the previously unappreciated scale of genetic and epigenetic changes underpinning acquired VEN resistance. These appear to be multilayered. One layer comprises changes in the BCL2 family of apoptosis regulators, especially the prosurvival family members. This includes previously described mutations in BCL2 and amplification of the MCL1 gene but is heterogeneous across and within individual patient leukemias. Changes in the proapoptotic genes are notably uncommon, except for single cases with subclonal losses of BAX or NOXA. Much more prominent was universal MCL1 gene upregulation. This was driven by an overlying layer of emergent NF-κB (nuclear factor kappa B) activation, which persisted in circulating cells during VEN therapy. We discovered that MCL1 could be a direct transcriptional target of NF-κB. Both the switch to alternative prosurvival factors and NF-κB activation largely dissipate following VEN discontinuation. Our studies reveal the extent of plasticity of CLL cells in their ability to evade VEN-induced apoptosis. Importantly, these findings pinpoint new approaches to circumvent VEN resistance and provide a specific biological justification for the strategy of VEN discontinuation once a maximal response is achieved rather than maintaining long-term selective pressure with the drug.
Original language | English |
---|---|
Pages (from-to) | 2127-2141 |
Number of pages | 15 |
Journal | Blood |
Volume | 140 |
Issue number | 20 |
DOIs | |
Publication status | Published - 17 Nov 2022 |
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In: Blood, Vol. 140, No. 20, 17.11.2022, p. 2127-2141.
Research output: Contribution to journal › Article › Academic › peer-review
TY - JOUR
T1 - Single-cell multiomics reveal the scale of multilayered adaptations enabling CLL relapse during venetoclax therapy
AU - Thijssen, Rachel
AU - Tian, Luyi
AU - Anderson, Mary Ann
AU - Flensburg, Christoffer
AU - Jarratt, Andrew
AU - Garnham, Alexandra L.
AU - Jabbari, Jafar S.
AU - Peng, Hongke
AU - Lew, Thomas E.
AU - Teh, Charis E.
AU - Gouil, Quentin
AU - Georgiou, Angela
AU - Tan, Tania
AU - Djajawi, Tirta M.
AU - Tam, Constantine S.
AU - Seymour, John F.
AU - Blombery, Piers
AU - Gray, Daniel H.D.
AU - Majewski, Ian J.
AU - Ritchie, Matthew E.
AU - Roberts, Andrew W.
AU - Huang, David C.S.
N1 - Funding Information: The authors thank the patients who enrolled in the venetoclax clinical trials and for assistance from Naomi Sprigg and Kelli Gray with the collection and curation of patient samples. The authors also thank Casey Anttila, Connie Li, and Stephen Wilcox, the WEHI SCORE team and core facilities (flow cytometry, genomics), and Andrew Mitchell at the Materials Characterisation and Fabrication Platform (MCFP) at the University of Melbourne for mass cytometry support; and Andreas Strasser, Christine White, and Mark van Delft for their critical reading of the manuscript. Illustrations were created with BioRender.com. This work was supported by scholarships, fellowships, and grants from the Australian National Health and Medical Research Council (NHMRC: Program Grants 1016647, 1113577 to A.W.R. 1016701, 1113133 to D.C.S.H. Fellowships 1089072 to C.E.T. 1090236 and 1158024 to D.H.D.G. 1079560 to A.W.R. 1043149, 1156024 to D.C.S.H. Investigator Grant 1177718 to M.A.A. 1174902 to A.W.R.), the Leukemia & Lymphoma Society of America (Fellowship 5467-18 to R.T. Specialized Center of Research (SCOR) grant 7015-18 to A.W.R. and D.C.S.H.), The Australian Research Council (Discovery Project 200102903 to M.E.R.), Cure Cancer and Cancer Australia (grant 1186003 to R.T.), Victorian Cancer Agency (MCRF grant 15018 to I.J.M. and A.W.R. MCRF20026 Fellowship to C.E.T.), Cancer Council Victoria (grants-in-aid 1146518 and 1102104 to D.H.D.G. and 1124178 to I.J.M.), the University of Melbourne (MIRS and MIFRS scholarships to L.T. H.P. and T.M.D.), grants from Snowdome Foundation (M.A.A. and P.B.), Wilson Centre for Lymphoma Genomics (P.B.), Victorian Comprehensive Cancer Centre (support for mass cytometry), Perpetual Impact Philanthropy Founding (IPAP2019/1437 to C.E.T.), the Chan Zuckerberg Initiative DAF (an advised fund of Silicon Valley Community Foundation, grant number 2019-002443 to M.E.R.), and the Australian Cancer Research Foundation. This work was made possible through Victorian State Government Operational Infrastructure Support and the Australian Government NHMRC IRIISS. Contribution: R.T. L.T. M.E.R. A.W.R. and D.C.S.H. devised the research strategy; M.E.R. A.W.R. and D.C.S.H. cosupervised the research; R.T. M.A.A. A.J. H.P. C.E.T. Q.G. A.G. T.T. and T.M.D. performed the experiments; L.T. C.F. A.L.G. H.P. and T.E.L. performed data analyses; J.S.J. developed the tools; R.T. L.T. C.S.T. J.F.S. P.B. D.H.D.G. I.J.M. M.E.R. A.W.R. and D.C.S.H. helped to interpret results; R.T. L.T. A.W.R. and D.C.S.H. wrote the manuscript; and all authors reviewed the manuscript before submission. Funding Information: This work was supported by scholarships, fellowships, and grants from the Australian National Health and Medical Research Council (NHMRC: Program Grants 1016647 , 1113577 to A.W.R., 1016701 , 1113133 to D.C.S.H., Fellowships 1089072 to C.E.T., 1090236 and 1158024 to D.H.D.G., 1079560 to A.W.R., 1043149 , 1156024 to D.C.S.H., Investigator Grant 1177718 to M.A.A., 1174902 to A.W.R.), the Leukemia & Lymphoma Society of America (Fellowship 5467-18 to R.T., Specialized Center of Research (SCOR) grant 7015-18 to A.W.R. and D.C.S.H.), The Australian Research Council (Discovery Project 200102903 to M.E.R.), Cure Cancer and Cancer Australia (grant 1186003 to R.T.), Victorian Cancer Agency (MCRF grant 15018 to I.J.M. and A.W.R., MCRF20026 Fellowship to C.E.T.), Cancer Council Victoria (grants-in-aid 1146518 and 1102104 to D.H.D.G. and 1124178 to I.J.M.), the University of Melbourne (MIRS and MIFRS scholarships to L.T., H.P., and T.M.D.), grants from Snowdome Foundation (M.A.A. and P.B.), Wilson Centre for Lymphoma Genomics (P.B.), Victorian Comprehensive Cancer Centre (support for mass cytometry), Perpetual Impact Philanthropy Founding (IPAP2019/1437 to C.E.T.), the Chan Zuckerberg Initiative DAF (an advised fund of Silicon Valley Community Foundation , grant number 2019-002443 to M.E.R.), and the Australian Cancer Research Foundation. This work was made possible through Victorian State Government Operational Infrastructure Support and the Australian Government NHMRC IRIISS. Publisher Copyright: © 2022 The American Society of Hematology
PY - 2022/11/17
Y1 - 2022/11/17
N2 - Venetoclax (VEN) inhibits the prosurvival protein BCL2 to induce apoptosis and is a standard therapy for chronic lymphocytic leukemia (CLL), delivering high complete remission rates and prolonged progression-free survival in relapsed CLL but with eventual loss of efficacy. A spectrum of subclonal genetic changes associated with VEN resistance has now been described. To fully understand clinical resistance to VEN, we combined single-cell short- and long-read RNA-sequencing to reveal the previously unappreciated scale of genetic and epigenetic changes underpinning acquired VEN resistance. These appear to be multilayered. One layer comprises changes in the BCL2 family of apoptosis regulators, especially the prosurvival family members. This includes previously described mutations in BCL2 and amplification of the MCL1 gene but is heterogeneous across and within individual patient leukemias. Changes in the proapoptotic genes are notably uncommon, except for single cases with subclonal losses of BAX or NOXA. Much more prominent was universal MCL1 gene upregulation. This was driven by an overlying layer of emergent NF-κB (nuclear factor kappa B) activation, which persisted in circulating cells during VEN therapy. We discovered that MCL1 could be a direct transcriptional target of NF-κB. Both the switch to alternative prosurvival factors and NF-κB activation largely dissipate following VEN discontinuation. Our studies reveal the extent of plasticity of CLL cells in their ability to evade VEN-induced apoptosis. Importantly, these findings pinpoint new approaches to circumvent VEN resistance and provide a specific biological justification for the strategy of VEN discontinuation once a maximal response is achieved rather than maintaining long-term selective pressure with the drug.
AB - Venetoclax (VEN) inhibits the prosurvival protein BCL2 to induce apoptosis and is a standard therapy for chronic lymphocytic leukemia (CLL), delivering high complete remission rates and prolonged progression-free survival in relapsed CLL but with eventual loss of efficacy. A spectrum of subclonal genetic changes associated with VEN resistance has now been described. To fully understand clinical resistance to VEN, we combined single-cell short- and long-read RNA-sequencing to reveal the previously unappreciated scale of genetic and epigenetic changes underpinning acquired VEN resistance. These appear to be multilayered. One layer comprises changes in the BCL2 family of apoptosis regulators, especially the prosurvival family members. This includes previously described mutations in BCL2 and amplification of the MCL1 gene but is heterogeneous across and within individual patient leukemias. Changes in the proapoptotic genes are notably uncommon, except for single cases with subclonal losses of BAX or NOXA. Much more prominent was universal MCL1 gene upregulation. This was driven by an overlying layer of emergent NF-κB (nuclear factor kappa B) activation, which persisted in circulating cells during VEN therapy. We discovered that MCL1 could be a direct transcriptional target of NF-κB. Both the switch to alternative prosurvival factors and NF-κB activation largely dissipate following VEN discontinuation. Our studies reveal the extent of plasticity of CLL cells in their ability to evade VEN-induced apoptosis. Importantly, these findings pinpoint new approaches to circumvent VEN resistance and provide a specific biological justification for the strategy of VEN discontinuation once a maximal response is achieved rather than maintaining long-term selective pressure with the drug.
UR - http://www.scopus.com/inward/record.url?scp=85139146598&partnerID=8YFLogxK
U2 - https://doi.org/10.1182/blood.2022016040
DO - https://doi.org/10.1182/blood.2022016040
M3 - Article
C2 - 35709339
SN - 0006-4971
VL - 140
SP - 2127
EP - 2141
JO - Blood
JF - Blood
IS - 20
ER -