American Association for Cancer Research
bcd-23-0106_supplementary_figures_1-19_suppsf1.pdf (4.29 MB)

Supplementary Figures 1-19 from Acute Lymphoblastic Leukemia with Myeloid Mutations Is a High-Risk Disease Associated with Clonal Hematopoiesis

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posted on 2024-05-01, 07:21 authored by Caner Saygin, Pu Zhang, Jacob Stauber, Ibrahim Aldoss, Adam S. Sperling, Lachelle D. Weeks, Marlise R. Luskin, Todd C. Knepper, Pankhuri Wanjari, Peng Wang, Angela M. Lager, Carrie Fitzpatrick, Jeremy P. Segal, Mehdi Gharghabi, Sandeep Gurbuxani, Girish Venkataraman, Jason X. Cheng, Bart J. Eisfelder, Oliver Bohorquez, Anand A. Patel, Sheethal Umesh Nagalakshmi, Savita Jayaram, Olatoyosi M. Odenike, Richard A. Larson, Lucy A. Godley, Daniel A. Arber, Christopher J. Gibson, Nikhil C. Munshi, Guido Marcucci, Benjamin L. Ebert, John M. Greally, Ulrich Steidl, Rosa Lapalombella, Bijal D. Shah, Wendy Stock

Supplemental Figure 1. WHO/ICC subtypes for the B-ALL cohort.

Supplemental Figure 2. Lymphoid clonal hematopoiesis (CH) mutations are less common in adults with ALL.

Supplemental Figure 3. Blast percentage in the pre-treatment sample and detected variant allelic frequencies (VAF) of mutations in ALL patients with myeloid mutations.

Supplemental Figure 4. Germline testing in the adult ALL cohort.

Supplemental Figure 5. Associations of gene mutations and WHO/ICC disease subtypes in B-ALL.

Supplemental Figure 6. Overall survival in adults with B-ALL.

Supplemental Figure 7. Kaplan-Meier overall survival curves for Ph-negative B-ALL patients stratified based on the presence of TP53 and non-TP53 MyM (A), and treatment with different chemotherapy protocols (B, C, D). LH, low hypodiploidy.

Supplemental Figure 8. T-lineage ALL with myeloid mutations (MyM).

Supplemental Figure 9. Expression levels of lineage-specific surface markers.

Supplemental Figure 10. Single cell DNA and protein sequencing to study the clonal architecture of Tlineage ALL with MyM.

Supplemental Figure 11. A, Flow plots showing ETP-lymphoblast, myeloid, and mature lymphocytic compartments of ETP-ALL1, which were sorted for DNA extraction and sequencing. B, Distribution of variant allelic frequencies (VAFs) for myeloid (IDH2, DNMT3A) and NRAS mutations across subpopulations of cells in ETP-ALL1.

Supplemental Figure 12. Clonal dynamics of ALL with MyM.

Supplemental Figure 13. Two CH mutations (TP53 and DNMT3A) were detectable seven years before the diagnosis of therapy-related ALL22.

Supplemental Figure 14. Clonal evolution of B-ALL2.

Supplemental Figure 15. Lymphoblasts from B-ALL with MyM are characterized by their resistance to cytotoxic chemotherapy.

Supplemental Figure 16. Curves showing viability of primary human B-ALL samples with TP53 mutation (n= 14), MyM (n= 16) and no MyM/TP53 (n= 24), treated with vincristine, doxorubicin, and blinatumomab at escalating doses.

Supplemental Figure 17. Flow cytometry plots showing ALL22 (TP53-mutated B-ALL) sample treated with escalating doses of blinatumomab.

Supplemental Figure 18. A, CR with MRD negativity rates in Ph-negative B-ALL patients treated with pediatric regimens, stratified based on age, gender, and treatment site. B, CR with MRD negativity rates in Ph-negative B-ALL patients treated with hyper-CVAD, stratified based on age, gender, and treatment site. C, CR with MRD negativity rates in Ph-negative B-ALL patients treated with inotuzumab.

Supplemental Figure 19. A, MHC class I and II antigen expression in blasts from B-ALL with MyM vs B-ALL without MyM/TP53. B, Comparison of genes implicated in blinatumomab resistance in B-ALL with vs without MyM/TP53.


Leukemia and Lymphoma Society (LLS)

Elsa U. Pardee Foundation (EUPF)

Prevent Cancer Foundation (PCF)

Cancer Research Foundation (CRF)

National Institutes of Health (NIH)



Myeloid neoplasms arise from preexisting clonal hematopoiesis (CH); however, the role of CH in the pathogenesis of acute lymphoblastic leukemia (ALL) is unknown. We found that 18% of adult ALL cases harbored TP53, and 16% had myeloid CH-associated gene mutations. ALL with myeloid mutations (MyM) had distinct genetic and clinical characteristics, associated with inferior survival. By using single-cell proteogenomic analysis, we demonstrated that myeloid mutations were present years before the diagnosis of ALL, and a subset of these clones expanded over time to manifest as dominant clones in ALL. Single-cell RNA sequencing revealed upregulation of genes associated with cell survival and resistance to apoptosis in B-ALL with MyM, which responds better to newer immunotherapeutic approaches. These findings define ALL with MyM as a high-risk disease that can arise from antecedent CH and offer new mechanistic insights to develop better therapeutic and preventative strategies. CH is a precursor lesion for lymphoblastic leukemogenesis. ALL with MyM has distinct genetic and clinical characteristics, associated with adverse survival outcomes after chemotherapy. CH can precede ALL years before diagnosis, and ALL with MyM is enriched with activated T cells that respond to immunotherapies such as blinatumomab.See related commentary by Iacobucci, p. 142.

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