Biology subjects

Tolison, H.

Publications and source records attributed to Tolison, H..

3 recordsLinked to original sources

bioRxiv2025

siRNA Mediated Genetic Perturbation of Primary Human Leukemia Stem and Progenitor Cells

Acute myeloid leukemia (AML) and myelodysplastic syndromes (MDS) are aggressive hematologic malignancies with poor outcomes. Leukemia stem and progenitor cells (LSPCs) are a subset of cells within the bulk tumor thought to be responsible for initiating disease and causing relapse. LSPCs evade chemotherapy partially due to their quiescent state. Therefore, studying this cell subpopulation is critical to identify new disease targets that can better the outcomes of AML/MDS patients. The use of RNAi and genetic approaches is technically challenging in primary leukemia cells and particularly LSPCs. Overcoming this technical hurdle could greatly expand the breath of pre-clinical and mechanistic examination of LSPCs. In this study, we describe a methodology to efficiently introduce siRNA, resulting in effective gene knock down in LSPCs. After isolation of LSPCs from primary patient samples, we showed that electroporation of these cells does not affect cell viability significantly. Further, using siGLO green transfection indicator, we show that RNA is introduced into the nucleus of these cells. siRNA transfection leads to efficient knockdown of target genes and has subsequent biological relevant activity. We have identified a method to effectively knock down genes in leukemia stem and progenitor cells, opening up new avenues to examine LSPC biology in human specimens.

cancer biology↗

bioRxiv2023

Targeting Acute Myeloid Leukemia Stem Cells Through Perturbation of Mitochondrial Calcium

We previously reported that acute myeloid leukemia stem cells (LSCs) are uniquely reliant on oxidative phosphorylation (OXPHOS) for survival. Moreover, maintenance of OXPHOS is dependent on BCL2, creating a therapeutic opportunity to target LSCs using the BCL2 inhibitor drug venetoclax. While venetoclax-based regimens have indeed shown promising clinical activity, the emergence of drug resistance is prevalent. Thus, in the present study, we investigated how mitochondrial properties may influence mechanisms that dictate venetoclax responsiveness. Our data show that utilization of mitochondrial calcium is fundamentally different between drug responsive and non-responsive LSCs. By comparison, venetoclax-resistant LSCs demonstrate a more active metabolic (i.e., OXPHOS) status with relatively high steady-state levels of calcium. Consequently, we tested genetic and pharmacological approaches to target the mitochondrial calcium uniporter, MCU. We demonstrate that inhibition of calcium uptake sharply reduces OXPHOS and leads to eradication of venetoclax-resistant LSCs. These findings demonstrate a central role for calcium signaling in the biology of LSCs and provide a therapeutic avenue for clinical management of venetoclax resistance. SignificanceWe identify increased utilization of mitochondrial calcium as distinct metabolic requirement of venetoclax-resistant LSCs and demonstrate the potential of targeting mitochondrial calcium uptake as a therapeutic strategy.

cancer biology↗

bioRxiv2022

A novel type of monocytic leukemia stem cell revealed by the clinical use of venetoclax-based therapy

The BCL-2 inhibitor venetoclax has recently emerged as an important component of acute myeloid leukemia (AML) therapy. Notably, use of this agent has revealed a previously unrecognized form of pathogenesis characterized by monocytic disease progression. We demonstrate that this form of disease arises from a fundamentally different type of leukemia stem cell (LSC), which we designate as monocytic LSC (m-LSC), that is developmentally and clinically distinct from the more well-described primitive LSC (p-LSC). The m-LSC is distinguished by a unique immunophenotype (CD34-, CD4+, CD11b-, CD14-, CD36-), unique transcriptional state, reliance on purine/pyrimidine metabolism, and selective sensitivity to cladribine. Critically, in some instances m-LSC and p-LSC subtypes can co-reside in the same AML patient and simultaneously contribute to overall tumor complexity. Thus, our findings demonstrate that LSC heterogeneity has direct clinical significance and highlights the need to distinguish and target m-LSCs as a means to improve clinical outcomes with venetoclax-based regimens. Statement of SignificanceThese studies identify and characterize a new type of human acute myeloid leukemia stem cell (LSC) that is responsible for monocytic disease progression in acute myeloid leukemia (AML) patients treated with venetoclax-based regimens. Our studies describe the phenotype, molecular properties, and drug sensitivities of this unique LSC subclass.

cancer biology↗