scholarly journals Single Cell Transcriptomics Predicts Relapse in Infants with Acute Lymphoblastic Leukemia

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 3951-3951
Author(s):  
Tito Candelli ◽  
Pauline Schneider ◽  
Patricia Garrido Castro ◽  
Luke A Jones ◽  
Rob Pieters ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Single Cell ◽  
Single Cells ◽  
Lymphoblastic Leukemia ◽  
Gene Expression Pattern ◽  
Disease Classification ◽  
Leukemic Cells ◽  
Response To Treatment ◽  
Bone Marrow Biopsies ◽  
Relapse Risk

Care of infants (<1 year of age) diagnosed with MLL (KMT2A)-rearranged acute lymphoblastic leukemia (MLLr-iALL) suffers from two major drawbacks. First, a poor survival rate due to a high rates of early relapse and chemo-resistance. Additionally, the approximately 30-50% of patients that do survive, suffer from life-long, debilitating side-effects of current treatment. While almost all MLLr-iALL patients show an initial promising response to treatment, two-third of the patients relapse, typically within the first year from diagnosis and while still on treatment. Accurate relapse prediction would allow implementation of treatment strategies that take relapse risk into account, with great potential benefit for all patients. Here, we show that Single-cell RNA sequencing (scRNA-seq) can be valuable for risk stratification and that the abundance of chemo-resistant cells within the diagnosis sample might be a powerful indicator of the likelihood of relapse. We have used scRNA-seq to analyze the response to treatment of leukemic cells in bone marrow biopsies of seven MLLr-iALL patients, expressing either the oncogenic MLL-AF4 or MLL-ENL fusion gene, at the time of initial diagnosis. Three of these patients successfully underwent treatment and remained disease-free during 7 years of follow-up, while in the remaining four cases the disease returned within a year from diagnosis. All samples were subjected to scRNA-seq by FACS index sorting with the aim of identifying differences between early relapsers and long-term survivors. Quantification of the proportion of cells classified by single cell transcriptomics, categorized as either chemo-resistant or chemo-sensitive, accurately predicts the occurrence of future relapse in individual patients. Strikingly, the single cell-based classification is even consistent with the order of relapse timing. Additionally, leukemic cells associated with high relapse risk are typified by a small phenotype, which coincides with an apparent quiescent gene expression pattern. This study clearly and, to the best of our knowledge, for the first time shows how disease classification and treatment management can directly benefit from single cell genomics. It demonstrates how classification based on a pivotal functional characteristic of single cells can be performed, despite individual patient variation. Our results shed light on the subpopulation from which leukemic relapse originates, and opens up opportunities for strong, risk-based strategies for future MLLr-iALL treatment regimens. Disclosures Pieters: medac: Consultancy; jazz farmaceuticals: Consultancy.

scholarly journals Single-cell transcriptomics predicts relapse in MLL-rearranged acute lymphoblastic leukemia in infants

2020 ◽  
Author(s):  
Tito Candelli ◽  
Pauline Schneider ◽  
Patricia Garrido Castro ◽  
Luke A. Jones ◽  
Rob Pieters ◽  
...  
Keyword(s):  
Gene Expression ◽  
Acute Lymphoblastic Leukemia ◽  
Risk Stratification ◽  
Single Cell ◽  
Rna Sequencing ◽  
Lymphoblastic Leukemia ◽  
Leukemic Cells ◽  
List Type ◽  
Relapse Risk ◽  
Single Cell Rna Sequencing

AbstractInfants with MLL-rearranged acute lymphoblastic leukemia (ALL) undergo intense therapy to counter a highly aggressive leukemia with survival rates of only 30-40%. The majority of patients initially show therapy response, but in two-thirds of cases the leukemia returns, typically during treatment. Accurate relapse prediction would enable treatment strategies that take relapse risk into account, with potential benefits for all patients. Through analysis of diagnostic bone marrow biopsies, we show that single-cell RNA sequencing can predict future relapse occurrence. By analysing gene modules derived from an independent study of the gene expression response to the key drug prednisone, individual leukemic cells are predicted to be either resistant or sensitive to treatment. Quantification of the proportion of cells classified by single-cell transcriptomics as resistant or sensitive, accurately predicts the occurrence of future relapse in individual patients. Strikingly, the single-cell based classification is even consistent with the order of relapse timing. These results lay the foundation for risk-based treatment of MLL-rearranged infant ALL, through single-cell classification. This work also sheds light on the subpopulation of cells from which leukemic relapse arises. Leukemic cells associated with high relapse risk are characterized by a smaller size and a quiescent gene expression program. These cells have significantly fewer transcripts, thereby also demonstrating why single-cell analyses may outperform bulk mRNA studies for risk stratification. This study indicates that single-cell RNA sequencing will be a valuable tool for risk stratification of MLL-rearranged infant ALL, and shows how clinically relevant information can be derived from single-cell genomics.Key PointsSingle-cell RNA sequencing accurately predicts relapse in MLL-rearranged infant ALLIdentification of cells from which MLL-rearranged infant ALL relapses occur

Dicentric Translocation (9;12) in Acute Lymphoblastic Leukemia: A Chromosomal Abnormality with an Excellent Prognosis?

2017 ◽  
Vol 103 (1_suppl) ◽  
pp. S44-S46
Author(s):  
Laura Illade ◽  
Victoria Fioravantti ◽  
Maitane Andion ◽  
Carmen Hernandez-Marques ◽  
Luis Madero ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
Chromosomal Abnormality ◽  
Pediatric Patients ◽  
Chromosomal Abnormalities ◽  
Lymphoblastic Leukemia ◽  
Prognostic Significance ◽  
International Study ◽  
Leukemic Cells ◽  
Response To Treatment ◽  
Molecular Alteration

Purpose Recurrent chromosomal abnormalities present in the leukemic cells of pediatric patients with acute lymphoblastic leukemia (ALL) often allow us to classify patients according to their prognosis, in order to establish further treatment. The dicentric translocation (9;12) consists of the rearrangement of the short arms of chromosomes 9 and 12 generating a dicentric chromosome (9;12). Patients with this alteration present a very good response to treatment and an excellent prognosis. Methods We present the case of an adolescent with ALL in which the dicentric translocation (9;12) was observed in the karyotype of the blasts at diagnosis. Results and conclusion Given the excellent results of our patient and most of the series published to date, an international study is necessary to determine the true prognostic significance of this molecular alteration.

scholarly journals Prevalence and Growth Characteristics of Malignant Stem Cells in B-Lineage Acute Lymphoblastic Leukemia

Blood ◽  
1997 ◽  
Vol 89 (10) ◽  
pp. 3735-3744 ◽  
Author(s):  
Hikari Nishigaki ◽  
Chikako Ito ◽  
Atsushi Manabe ◽  
Masa-aki Kumagai ◽  
Elaine Coustan-Smith ◽  
...  
Keyword(s):  
Stem Cells ◽  
Acute Lymphoblastic Leukemia ◽  
Cell Growth ◽  
Single Cells ◽  
Lymphoblastic Leukemia ◽  
Leukemic Cell ◽  
Growth Characteristics ◽  
Leukemic Cells ◽  
Paracrine Factors ◽  
B Lineage

We used a stroma-supported culture method to study the prevalence and growth characteristics of malignant stem cells in acute lymphoblastic leukemia (ALL). In 51 of 108 B-lineage ALL samples, bone marrow-derived stroma not only inhibited apoptosis of ALL cells but also supported their proliferation in serum-free medium. When single leukemic cells were placed in the stroma-coated wells of microtiter plates, the percentage of wells with leukemic cell growth after 2 to 5 months of culture ranged from 6% to 20% (median, 15%; 5 experiments). The immunophenotypes and genetic features of cells recovered from these cultures were identical to those noted before culture. All cells maintained their stroma dependency and self-renewal capacity. Leukemic clones derived from single cells contained approximately 103 to 106 cells after 1 month of culture; other clones became detectable only after prolonged culture. Cell growth in stroma-coated wells correlated with the number of initially seeded cells (1 or 10; r = .87). However, the observed percentages of positive wells seeded with 10 cells always exceeded values predicted from results with single-cell–initiated cultures (P < .003 by paired t-test), suggesting stimulation of leukemic cell growth by paracrine factors. In conclusion, the proportion of ALL cells with clonogenic potential may be considerably higher than previously thought.

scholarly journals Overcoming Glucocorticoid Resistance in Acute Lymphoblastic Leukemia: Repurposed Drugs Can Improve the Protocol

2021 ◽  
Vol 11 ◽  
Author(s):  
Miguel Olivas-Aguirre ◽  
Liliana Torres-López ◽  
Igor Pottosin ◽  
Oxana Dobrovinskaya
Keyword(s):  
Cell Death ◽  
Acute Lymphoblastic Leukemia ◽  
Lymphoblastic Leukemia ◽  
Metabolic Reprogramming ◽  
Leukemic Cells ◽  
Response To Treatment ◽  
Remission Induction ◽  
Central Component ◽  
Apoptosis Resistance ◽  
Treatment Protocols

Glucocorticoids (GCs) are a central component of multi-drug treatment protocols against T and B acute lymphoblastic leukemia (ALL), which are used intensively during the remission induction to rapidly eliminate the leukemic blasts. The primary response to GCs predicts the overall response to treatment and clinical outcome. In this review, we have critically analyzed the available data on the effects of GCs on sensitive and resistant leukemic cells, in order to reveal the mechanisms of GC resistance and how these mechanisms may determine a poor outcome in ALL. Apart of the GC resistance, associated with a decreased expression of receptors to GCs, there are several additional mechanisms, triggered by alterations of different signaling pathways, which cause the metabolic reprogramming, with an enhanced level of glycolysis and oxidative phosphorylation, apoptosis resistance, and multidrug resistance. Due to all this, the GC-resistant ALL show a poor sensitivity to conventional chemotherapeutic protocols. We propose pharmacological strategies that can trigger alternative intracellular pathways to revert or overcome GC resistance. Specifically, we focused our search on drugs, which are already approved for treatment of other diseases and demonstrated anti-ALL effects in experimental pre-clinical models. Among them are some “truly” re-purposed drugs, which have different targets in ALL as compared to other diseases: cannabidiol, which targets mitochondria and causes the mitochondrial permeability transition-driven necrosis, tamoxifen, which induces autophagy and cell death, and reverts GC resistance through the mechanisms independent of nuclear estrogen receptors (“off-target effects”), antibiotic tigecycline, which inhibits mitochondrial respiration, causing energy crisis and cell death, and some anthelmintic drugs. Additionally, we have listed compounds that show a classical mechanism of action in ALL but are not used still in treatment protocols: the BH3 mimetic venetoclax, which inhibits the anti-apoptotic protein Bcl-2, the hypomethylating agent 5-azacytidine, which restores the expression of the pro-apoptotic BIM, and compounds targeting the PI3K-Akt-mTOR axis. Accordingly, these drugs may be considered for the inclusion into chemotherapeutic protocols for GC-resistant ALL treatments.

scholarly journals Decoding Clonal Evolution in Relapsed T Cell Acute Lymphoblastic Leukemia at Single Cell Resolution

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 3482-3482
Author(s):  
Jingliao Zhang ◽  
Yongjuan Duan ◽  
Yanxia Chang ◽  
Yue Wang ◽  
Chao Liu ◽  
...  
Keyword(s):  
T Cells ◽  
Acute Lymphoblastic Leukemia ◽  
T Cell ◽  
Single Cell ◽  
Conflicts Of Interest ◽  
Expression Profiles ◽  
Lymphoblastic Leukemia ◽  
Clonal Evolution ◽  
The Other ◽  
Leukemic Cells

Abstract T-lineage acute lymphoblastic leukemia (T-ALL) comprises approximately 10-15% of pediatric ALL cases with distinct feature in biology and largely inferior outcome compared to B-ALL. Growing evidence has reflected pivotal roles of clonal evolution in T-ALL recurrence, but bulk sequencing may not serve as the perfect model to reliably infer clonal heterogeneities and their immunomodulatory milieu during leukemia development. In this study, single-cell sequencing was applied to uncover leukemic clonal relationships with relapse throughout chemotherapy in T-ALL at a more accurate resolution. We performed bulk whole-exome sequencing for sorted CD7 + BMMCs from 5 pairs of diagnosis-relapse (Dx_Rel) samples, revealing a series of well-reported hotspot mutations in T-ALL. Among those, we observed diagnosis-specific variations and relapse-emerged variations, suggesting the putative correlations with chemo-resistance. Transcriptomic sequencing highlighted additional stemness and metabolic abnormalities underlying leukemic re-occurrence. Incorporated Dx_Rel paired ATAC-seq depicted relapse-specific activated chromatin regions, such as ELK1, ELK4, RUNX1. To dissect clonal diversities within and across the 5 Dx_Rel T-ALL pairs, we carried out high-throughput droplet-based 5'-single-cell RNA-seq (scRNA-seq) and paired T cell receptor sequencing (scTCR-seq). By performing unsupervised clustering of scRNA-seq profiles encompassing 10 samples, we identified 23 distinct T-lineage clusters (Cluster 0-22) based on the two-dimensional UMAP visualization. In 2 out of 5 patients (T593 and T788), diffusion map of T-lineage sub-clusters between diagnostic and relapsed samples appeared to be almost identical, while distinct shifts from diagnosis to relapse in the compositions have been observed in the other 3 out of 5 patients (T956, T723 and T856). Besides, it was noteworthy that two T-cell sub-clusters were concluded as "normal" T cells (Cluster 9 and 12) uniformly presented in both diagnostic and relapsed diffusion of T-cell sub-clusters across 5 Dx_Rel, from which TCR repertoires and expression profiles could well discriminate leukemic cells. Next, we sought to further deconvolute the clonal evolution patterns for T-ALL Dx_Rel pairs. We observed that except in T788 lacking of clonal TCRs, dominant diagnostic clones of the other 4 patients diminished (T593) or vanished (T956, T723, T856) at relapse, sparing newly emerged subclones predominantly substituted at relapse. We clearly depicted two distinct patterns of evolutionary trajectories in these 4 Dx_Rel pairs by comprehensively mapping hierarchical TCR clonotypes onto leukemic clonotypes at single cell levels. Specifically, in T956 and T723, we observed significant outgrowth of incidental diagnostic sub-clones at relapse, whereby surrogate TCR repertoires correspondingly enumerated, suggestive of dynamic shifts in dominant clone over continuous chemo-exposure. Whereas in T593 and T856, expanding clones at relapse were showed up with completely different gene signatures from the diagnostic ones, but dominant clones at diagnosis and relapse were surprisingly presented with identical TCR repertoires. This was undoubtedly informative of leukemic "clonal drift" within which hypothetical intrinsic transformation happened to the same subclones over persistent chemotherapy. Besides, we took advantage of our well-discriminated model to fully delineated the involvement of "normal" T subclusters in leukemic latency and chemo-responsiveness. By analyzing TCR repertoires in combined with expression profiles, we noted that "normal" T cells infiltrated by T-ALL were majorly distributed in CD8-effector sub-clusters compared to those from healthy donor, suggesting a robust leukemic stimulation on effector CD8 signaling in T-ALL microenvironment. Collectively, our presented study accurately distinguished leukemic cells from normal T cells in T-ALL at a single-cell resolution. By tracking transcriptomic profiles within and across Dx_Rel T-ALL pairs, we further identified distinct clonal evolutionary patterns, which may determine diversified fates of leukemic clones in response to therapeutic pressures. In the meantime, we provided a comprehensive phenotypic view on "normal" T cells under leukemic prevalence and re-occurrence, extending significant implications for future precise immunotherapies. Disclosures No relevant conflicts of interest to declare.

scholarly journals Prevalence and Growth Characteristics of Malignant Stem Cells in B-Lineage Acute Lymphoblastic Leukemia

Blood ◽  
1997 ◽  
Vol 89 (10) ◽  
pp. 3735-3744 ◽  
Author(s):  
Hikari Nishigaki ◽  
Chikako Ito ◽  
Atsushi Manabe ◽  
Masa-aki Kumagai ◽  
Elaine Coustan-Smith ◽  
...  
Keyword(s):  
Stem Cells ◽  
Acute Lymphoblastic Leukemia ◽  
Cell Growth ◽  
Single Cells ◽  
Lymphoblastic Leukemia ◽  
Leukemic Cell ◽  
Growth Characteristics ◽  
Leukemic Cells ◽  
Paracrine Factors ◽  
B Lineage

Abstract We used a stroma-supported culture method to study the prevalence and growth characteristics of malignant stem cells in acute lymphoblastic leukemia (ALL). In 51 of 108 B-lineage ALL samples, bone marrow-derived stroma not only inhibited apoptosis of ALL cells but also supported their proliferation in serum-free medium. When single leukemic cells were placed in the stroma-coated wells of microtiter plates, the percentage of wells with leukemic cell growth after 2 to 5 months of culture ranged from 6% to 20% (median, 15%; 5 experiments). The immunophenotypes and genetic features of cells recovered from these cultures were identical to those noted before culture. All cells maintained their stroma dependency and self-renewal capacity. Leukemic clones derived from single cells contained approximately 103 to 106 cells after 1 month of culture; other clones became detectable only after prolonged culture. Cell growth in stroma-coated wells correlated with the number of initially seeded cells (1 or 10; r = .87). However, the observed percentages of positive wells seeded with 10 cells always exceeded values predicted from results with single-cell–initiated cultures (P < .003 by paired t-test), suggesting stimulation of leukemic cell growth by paracrine factors. In conclusion, the proportion of ALL cells with clonogenic potential may be considerably higher than previously thought.

scholarly journals Toward Therapeutic Targeting of Bone Marrow Leukemic Niche Protective Signals in B-Cell Acute Lymphoblastic Leukemia

2021 ◽  
Vol 10 ◽  
Author(s):  
Marjorie C. Delahaye ◽  
Kaoutar-Insaf Salem ◽  
Jeoffrey Pelletier ◽  
Michel Aurrand-Lions ◽  
Stéphane J. C. Mancini
Keyword(s):  
Bone Marrow ◽  
Acute Lymphoblastic Leukemia ◽  
B Cell ◽  
Lymphoblastic Leukemia ◽  
Young Patients ◽  
Chemotherapeutic Agents ◽  
Leukemic Cells ◽  
Response To Treatment ◽  
Hematopoietic Stem ◽  
Cell Acute Lymphoblastic Leukemia

B-cell acute lymphoblastic leukemia (B-ALL) represents the malignant counterpart of bone marrow (BM) differentiating B cells and occurs most frequently in children. While new combinations of chemotherapeutic agents have dramatically improved the prognosis for young patients, disease outcome remains poor after relapse or in adult patients. This is likely due to heterogeneity of B-ALL response to treatment which relies not only on intrinsic properties of leukemic cells, but also on extrinsic protective cues transmitted by the tumor cell microenvironment. Alternatively, leukemic cells have the capacity to shape their microenvironment towards their needs. Most knowledge on the role of protective niches has emerged from the identification of mesenchymal and endothelial cells controlling hematopoietic stem cell self-renewal or B cell differentiation. In this review, we discuss the current knowledge about B-ALL protective niches and the development of therapies targeting the crosstalk between leukemic cells and their microenvironment.

scholarly journals Single-cell DNA amplicon sequencing reveals clonal heterogeneity and evolution in T-cell acute lymphoblastic leukemia

Blood ◽  
2020 ◽  
Author(s):  
Llucia Albertí-Servera ◽  
Sofie Demeyer ◽  
Inge Govaerts ◽  
Toon Swings ◽  
Jolien De Bie ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
T Cell ◽  
Single Cell ◽  
Lymphoblastic Leukemia ◽  
Protein Translation ◽  
Amplicon Sequencing ◽  
Leukemic Cells ◽  
Clonal Heterogeneity ◽  
Cell Acute Lymphoblastic Leukemia ◽  
Notch1 Mutations

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive leukemia that is most frequent in children and is characterized by the presence of few chromosomal rearrangements and 10 to 20 somatic mutations in protein-coding regions at diagnosis. The majority of T-ALL cases harbor activating mutations in NOTCH1 together with mutations in genes implicated in kinase signaling, transcriptional regulation or protein translation. To obtain more insight in the level of clonal heterogeneity at diagnosis and during treatment, we used single-cell targeted DNA sequencing with the Tapestri platform. We designed a custom ALL panel and obtained accurate single-nucleotide variant and small insertion-deletion mutation calling for 305 amplicons covering 110 genes in about 4400 cells per sample and time point. A total of 108,188 cells were analyzed for 25 samples of 8 T-ALL patients. We typically observed a major clone at diagnosis (&gt;35% of the cells) accompanied by several minor clones of which some were less than 1% of the total number of cells. Four patients had &gt;2 NOTCH1 mutations some of which present in minor clones, indicating a strong pressure to acquire NOTCH1 mutations in developing T-ALL cells. By analyzing longitudinal samples, we detected the presence and clonal nature of residual leukemic cells as well as clones with a minor presence at diagnosis that evolved to clinically relevant major clones at later disease stages. Therefore, single-cell DNA amplicon sequencing is a sensitive assay to detect clonal architecture and evolution in T-ALL.

scholarly journals Colony formation in vitro by leukemic cells in acute lymphoblastic leukemia (ALL)

Blood ◽  
1978 ◽  
Vol 52 (4) ◽  
pp. 712-718 ◽  
Author(s):  
SD Smith ◽  
EM Uyeki ◽  
JT Lowman
Keyword(s):  
Bone Marrow ◽  
Acute Lymphoblastic Leukemia ◽  
Bone Marrow Cells ◽  
Lymphoblastic Leukemia ◽  
Lymphoid Cells ◽  
Assay System ◽  
Leukemic Cells ◽  
Specific Cell ◽  
Specific Cell Surface

Abstract An assay system in vitro for the growth of malignant lymphoblastic colony-forming cells (CFC) was established. Growth of malignant myeloblastic CFC has been previously reported, but this is the first report of growth of malignant lymphoblastic CFC. Established assay systems in vitro have been very helpful in elucidating the control of growth and differentiation of both normal and malignant bone marrow cells. Lymphoblastic CFC were grown from the bone marrow aspirates of 20 children with acute lymphoblastic leukemia. Growth of these colonies was established on an agar assay system and maintained in the relative hypoxia (7% oxygen) of a Stulberg chamber. The criteria for malignancy of these colonies was based upon cellular cytochemical staining characteristics, the presence of specific cell surface markers, and the ability of these lymphoid cells to grow without the addition of a lymphoid mitogen. With this technique, specific nutritional requirements and drug sensitivities can be established in vitro, and these data may permit tailoring of individual antileukemic therapy.

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