Centre for Cancer Research - Research Publications

Permanent URI for this collection

Search Results

Now showing 1 - 7 of 7
  • Item
    No Preview Available
    Single-cell multiomics reveal the scale of multilayered adaptations enabling CLL relapse during venetoclax therapy
    Thijssen, R ; Tian, L ; Anderson, MA ; Flensburg, C ; Jarratt, A ; Garnham, AL ; Jabbari, JS ; Peng, H ; Lew, TE ; Teh, CE ; Gouil, Q ; Georgiou, A ; Tan, T ; Djajawi, TM ; Tam, CS ; Seymour, JF ; Blombery, P ; Gray, DHD ; Majewski, IJ ; Ritchie, ME ; Roberts, AW ; Huang, DCS (AMER SOC HEMATOLOGY, 2022-11-17)
    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.
  • Item
    Thumbnail Image
    Single-cell sequencing demonstrates complex resistance landscape in CLL and MCL treated with BTK and BCL2 inhibitors
    Thompson, ER ; Nguyen, T ; Kankanige, Y ; Markham, JF ; Anderson, MA ; Handunnetti, SM ; Thijssen, R ; Yeh, PS-H ; Tam, CS ; Seymour, JF ; Roberts, AW ; Westerman, DA ; Blombery, P (ELSEVIER, 2022-01-25)
    The genomic landscape of resistance to targeted agents (TAs) used as monotherapy in chronic lymphocytic leukemia (CLL) is complex and often heterogeneous at the patient level. To gain insight into the clonal architecture of acquired genomic resistance to Bruton tyrosine kinase (BTK) inhibitors and B-cell lymphoma 2 (BCL2) inhibitors in CLL, particularly in patients carrying multiple resistance mutations, we performed targeted single-cell DNA sequencing of 8 patients who developed progressive disease (PD) on TAs (either class). In all cases, analysis of single-cell architecture revealed mutual exclusivity between multiple resistance mutations to the same TA class, variable clonal co-occurrence of multiple mutations affecting different TAs in patients exposed to both classes, and a phenomenon of multiple independent emergences of identical nucleotide changes leading to canonical resistance mutations. We also report the first observation of established BCL2 resistance mutations in a patient with mantle cell lymphoma (MCL) following PD on sequential monotherapy, implicating BCL2 as a venetoclax resistance mechanism in MCL. Taken together, these data reveal the significant clonal complexity of CLL and MCL progression on TAs at the nucleotide level and confirm the presence of multiple, clonally independent, mechanisms of TA resistance within each individual disease context.
  • Item
    No Preview Available
    Clonal hematopoiesis, myeloid disorders and BAX-mutated myelopoiesis in patients receiving venetoclax for CLL
    Blombery, P ; Lew, TE ; Dengler, MA ; Thompson, ER ; Lin, VS ; Chen, X ; Nguyen, T ; Panigrahi, A ; Handunnetti, SM ; Carney, DA ; Westerman, DA ; Tam, CS ; Adams, JM ; Wei, AH ; Huang, DCS ; Seymour, JF ; Roberts, AW ; Anderson, MA (AMER SOC HEMATOLOGY, 2022-02-24)
    The BCL2 inhibitor venetoclax has established therapeutic roles in chronic lymphocytic leukemia (CLL) and acute myeloid leukemia (AML). As BCL2 is an important determinant of survival of both myeloid progenitor and B cells, we investigated whether clinical and molecular abnormalities arise in the myeloid compartment during long-term continuous venetoclax treatment of CLL in 89 patients (87 with relapsed/refractory CLL). Over a median follow-up of 75 (range 21-98) months, persistent cytopenias (≥1 of neutropenia, thrombocytopenia, anemia) lasting ≥4 months and unrelated to CLL occurred in 25 patients (28%). Of these patients, 20 (80%) displayed clonal hematopoiesis, including 10 with therapy-related myeloid neoplasms (t-MNs). t-MNs occurred exclusively in patients previously exposed to fludarabine-alkylator combination therapy with a cumulative 5-year incidence of 10.4% after venetoclax initiation, consistent with rates reported for patients exposed to fludarabine-alkylator combination therapy without venetoclax. To determine whether the altered myelopoiesis reflected the acquisition of mutations, we analyzed samples from patients with no or minimal bone marrow CLL burden (n = 41). Mutations in the apoptosis effector BAX were identified in 32% (13/41). In cellular assays, C-terminal BAX mutants abrogated outer mitochondrial membrane localization of BAX and engendered resistance to venetoclax killing. BAX-mutated clonal hematopoiesis occurred independently of prior fludarabine-alkylator combination therapy exposure and was not associated with t-MNs. Single-cell sequencing revealed clonal co-occurrence of mutations in BAX with DNMT3A or ASXL1. We also observed simultaneous BCL2 mutations within CLL cells and BAX mutations in the myeloid compartment of the same patients, indicating lineage-specific adaptation to venetoclax therapy.
  • Item
    No Preview Available
    Outcomes of patients with CLL sequentially resistant to both BCL2 and BTK inhibition
    Lew, TE ; Lin, VS ; Cliff, ER ; Blombery, P ; Thompson, ER ; Handunnetti, SM ; Westerman, DA ; Kuss, BJ ; Tam, CS ; Huang, DCS ; Seymour, JF ; Roberts, AW ; Anderson, MA (ELSEVIER, 2021-10-26)
    Covalent Bruton tyrosine kinase inhibitors (BTKi's) and the B-cell lymphoma 2 (BCL2) inhibitor venetoclax have significantly improved outcomes for patients with chronic lymphocytic leukemia (CLL), especially those with biologically adverse disease. Patients with CLL resistant to their first targeted agent (TA) can be effectively treated with the alternative class. However, relapses are expected with second-line TA therapy, and the clinical challenge of double class-resistant disease is now emerging with increasing frequency. To define the characteristics and outcomes of patients with double class-resistant disease, we retrospectively analyzed 17 patients who developed progressive disease (PD) on both TA classes for CLL (venetoclax, then BTKi, n=12; BTKi, then venetoclax, n = 5). The cohort was heavily pretreated (median lines of prior therapy, 4) and enriched for adverse disease genetics (complex karyotype, 12 of 12 tested [100%]; del(17p)/TP53 mutations, 15 of 17 [88%]). The median time to progression on prior venetoclax was 24 months (range, 6-94 months) and was 25 months (range, 1-55 months) on prior BTKi. Progression on second-line TA was manifest as progressive CLL in 11 patients and as Richter transformation in 6. The median overall survival after progression on second-line TA was 3.6 months (95% confidence interval, 2-11 months). Patients with double class-resistant CLL have a dismal prognosis, representing a group of high unmet need.
  • Item
    No Preview Available
    Immune recovery in patients with mantle cell lymphoma receiving long-term ibrutinib and venetoclax combination therapy
    Davis, JE ; Handunnetti, SM ; Ludford-Menting, M ; Sharpe, C ; Blombery, P ; Anderson, MA ; Roberts, AW ; Seymour, JF ; Tam, CS ; Ritchie, DS ; Koldej, RM (AMER SOC HEMATOLOGY, 2020-10-13)
    Combination venetoclax plus ibrutinib for the treatment of mantle cell lymphoma (MCL) has demonstrated efficacy in the relapsed or refractory setting; however, the long-term impact on patient immunology is unknown. In this study, changes in immune subsets of MCL patients treated with combination venetoclax and ibrutinib were assessed over a 4-year period. Multiparameter flow cytometry of peripheral blood mononuclear cells showed that ≥12 months of treatment resulted in alterations in the proportions of multiple immune subsets, most notably CD4+ and CD8+ effector and central memory T cells and natural killer cells, and normalization of T-cell cytokine production in response to T-cell receptor stimulation. Gene expression analysis identified upregulation of multiple myeloid genes (including S100 and cathepsin family members) and inflammatory pathways over 12 months. Four patients with deep responses stopped study drugs, resulting in restoration of normal immune subsets for all study parameters except myeloid gene/pathway expression, suggesting long-term combination venetoclax and ibrutinib irreversibly affects this population. Our findings demonstrate that long-term combination therapy is associated with immune recovery in MCL, which may allow responses to subsequent immunotherapies and suggests that this targeted therapy results in beneficial impacts on immunological recovery. This trial was registered at www.clinicaltrials.gov as #NCT02471391.
  • Item
    No Preview Available
    Undetectable peripheral blood MRD should be the goal of venetoclax in CLL, but attainment plateaus after 24 months
    Lew, TE ; Anderson, MA ; Lin, VS ; Handunnetti, SM ; Came, NA ; Blombery, P ; Westerman, DA ; Wall, M ; Tam, CS ; Roberts, AW ; Seymour, JF (ELSEVIER, 2020-01-14)
    The highly selective BCL2 inhibitor venetoclax achieves deep responses in patients with relapsed or refractory (R/R) chronic lymphocytic leukemia (CLL), including undetectable minimal residual disease (uMRD). We retrospectively reviewed 62 patients with CLL treated with venetoclax to investigate the performance of peripheral blood (PB) compared with bone marrow (BM) assessment of MRD; the kinetics, clinicopathological associations, and longer-term outcomes of uMRD attainment and recrudescence; and the ability of venetoclax dose escalation to deepen responses. Among 16 patients who achieved PB uMRD and had contemporaneous BM assessments, 13 (81%) had confirmed BM uMRD, and patients with PB uMRD had outcomes at least as favorable as those with BM uMRD for time to progression, overall survival, and MRD recrudescence. Excluding 2 patients lacking earlier assessment, the median time to PB uMRD was 18 (range, 5-26) months, with 90% of instances achieved by 24 months. There was no new PB uMRD attainment after 24 months without treatment intensification. The dominant association with earlier attainment of uMRD was concurrent rituximab (P = .012). Complex karyotype was associated with inferior uMRD attainment after 12 months of therapy (P = .015), and patients attaining uMRD whose disease harbored TP53 abnormalities demonstrated a trend toward earlier recrudescence (P = .089). Of patients who received venetoclax dose escalations, 4 (27%) of 15 achieved improvements in response. For patients with R/R CLL receiving venetoclax, PB uMRD commonly correlates with BM uMRD and is associated with a comparable longer-term prognosis. Concurrent rituximab augments uMRD attainment, but dose escalation and further treatment beyond 24 months infrequently deepen responses.
  • Item
    Thumbnail Image
    Structures of BCL-2 in complex with venetoclax reveal the molecular basis of resistance mutations
    Birkinshaw, RW ; Gong, J-N ; Luo, CS ; Lio, D ; White, CA ; Anderson, MA ; Blombery, P ; Lessene, G ; Majewski, IJ ; Thijssen, R ; Roberts, AW ; Huang, DCS ; Colman, PM ; Czabotar, PE (NATURE PUBLISHING GROUP, 2019-06-03)
    Venetoclax is a first-in-class cancer therapy that interacts with the cellular apoptotic machinery promoting apoptosis. Treatment of patients suffering chronic lymphocytic leukaemia with this BCL-2 antagonist has revealed emergence of a drug-selected BCL-2 mutation (G101V) in some patients failing therapy. To understand the molecular basis of this acquired resistance we describe the crystal structures of venetoclax bound to both BCL-2 and the G101V mutant. The pose of venetoclax in its binding site on BCL-2 reveals small but unexpected differences as compared to published structures of complexes with venetoclax analogues. The G101V mutant complex structure and mutant binding assays reveal that resistance is acquired by a knock-on effect of V101 on an adjacent residue, E152, with venetoclax binding restored by a E152A mutation. This provides a framework for considering analogues of venetoclax that might be effective in combating this mutation.