Cytarabine (Ara-C) and Daunorubicin (Dnr) forms the backbone of acute myeloid leukemia (AML) therapy. Drug resistance and toxic side effects pose a major threat to treatment success and hence ...alternate less toxic therapies are warranted. NF-E2 related factor-2 (Nrf2), a master regulator of antioxidant response is implicated in chemoresistance in solid tumors. However, little is known about the role of Nrf2 in AML chemoresistance and the effect of pharmacological inhibitor brusatol in modulating this resistance. Primary AML samples with high ex-vivo IC50 to Ara-C, ATO, Dnr had significantly high NRF2 RNA expression. Gene-specific knockdown of NRF2 improved sensitivity to these drugs in resistant AML cell lines by decreasing the expression of downstream antioxidant targets of Nrf2 by compromising the cell's ability to scavenge the ROS. Treatment with brusatol, a pharmacological inhibitor of Nrf2, improved sensitivity to Ara-C, ATO, and Dnr and reduced colony formation capacity. AML cell lines stably overexpressing NRF2 showed increased resistance to ATO, Dnr and Ara-C and increased expression of downstream targets. This study demonstrates that Nrf2 could be an ideal druggable target in AML, more so to the drugs that function through ROS, suggesting the possibility of using Nrf2 inhibitors in combination with chemotherapeutic agents to modulate drug resistance in AML.
Despite the success of Tyrosine kinase inhibitors (TKIs) in treating chronic myeloid leukemia (CML), leukemic stem cells (LSCs) persist, contributing to relapse and resistance. CML Mesenchymal ...Stromal Cells (MSCs) help in LSC maintenance and protection from TKIs. However, the limited passage and self-differentiation abilities of primary CML MSCs hinder extensive research. To overcome this, we generated and characterized an immortalised CML patient-derived MSC (iCML MSC) line and assessed its role in LSC maintenance. We also compared the immunophenotype and differentiation potential between primary CML MSCs at diagnosis, post-treatment, and with normal bone marrow MSCs. Notably, CML MSCs exhibited enhanced chondrogenic differentiation potential compared to normal MSCs. The iCML MSC line retained the trilineage differentiation potential and was genetically stable, enabling long-term investigations. Functional studies demonstrated that iCML MSCs protected CML CD34+ cells from imatinib-induced apoptosis, recapitulating the bone marrow microenvironment-mediated resistance observed in patients. iCML MSC-conditioned media enabled CML CD34+ and AML blast cells to proliferate rapidly, with no impact on healthy donor CD34+ cells. Gene expression profiling revealed dysregulated genes associated with calcium metabolism in CML CD34+ cells cocultured with iCML MSCs, providing insights into potential therapeutic targets. Further, cytokine profiling revealed that the primary CML MSC lines abundantly secreted 25 cytokines involved in immune regulation, supporting the hypothesis that CML MSCs create an immune modulatory microenvironment that promotes growth and protects against TKIs. Our study establishes the utility of iCML MSCs as a valuable model to investigate leukemic-stromal interactions and study candidate genes involved in mediating TKI resistance in CML LSCs.
De novo acute myeloid leukemia (AML) patients with FMS-like tyrosine kinase 3 internal tandem duplications (FLT3-ITD) have worse treatment outcomes. Arsenic trioxide (ATO) used in the treatment of ...acute promyelocytic leukemia (APL) has been reported to be effective in degrading the FLT3 protein in AML cell lines and sensitizing non-APL AML patient samples in-vitro. We have previously reported that primary cells from
FLT3-ITD
mutated AML patients were sensitive to ATO in-vitro compared to other non-M3 AML and molecular/pharmacological inhibition of NF-E2 related factor 2 (
NRF2
), a master regulator of antioxidant response improved the chemosensitivity to ATO and daunorubicin even in non
FLT3-ITD
mutated cell lines and primary samples. We examined the effects of molecular/pharmacological suppression of
NRF2
on acquired ATO resistance in the
FLT3-ITD
mutant AML cell line (MV4-11-ATO-R). ATO-R cells showed increased NRF2 expression, nuclear localization, and upregulation of bonafide NRF2 targets. Molecular inhibition of
NRF2
in this resistant cell line improved ATO sensitivity in vitro. Digoxin treatment lowered p-AKT expression, abrogating nuclear NRF2 localization and sensitizing cells to ATO. However, digoxin and ATO did not sensitize non-ITD AML cell line THP1 with high NRF2 expression. Digoxin decreased leukemic burden and prolonged survival in MV4-11 ATO-R xenograft mice. We establish that altering NRF2 expression may reverse acquired ATO resistance in FLT3-ITD AML.
The ligand-activated transcription factors, nuclear hormone receptors (NHRs), remain unexplored in hematological malignancies except for retinoic acid receptor alpha (
).
Here we profiled the ...expression of various NHRs and their coregulators in Chronic myeloid leukemia (CML) cell lines and identified a significant differential expression pattern between inherently imatinib mesylate (IM)-sensitive and resistant cell lines.
Retinoid-X-receptor alpha (
) was downregulated in CML cell lines inherently resistant to IM and in primary CML CD34
cells. Pre-treatment with clinically relevant RXRA ligands improved sensitivity to IM
in both CML cell lines and primary CML cells. This combination effectively reduced the viability and colony-forming capacity of CML CD34
cells
this combination reduced leukemic burden and prolonged survival. Overexpression (OE) of
inhibited proliferation and improved sensitivity to IM
.
OE cells showed reduced engraftment of cells in the bone marrow, improved sensitivity to IM, and prolonged survival. Both
and ligand treatment markedly reduced BCR::ABL1 downstream kinase activation, activating apoptotic cascades and improving sensitivity to IM. Importantly, RXRA OE also led to the disruption of the oxidative capacity of these cells.
Combining IM with clinically available RXRA ligands could form an alternative treatment strategy in CML patients with suboptimal response to IM.
Chemotherapy resistance leading to disease relapse is a significant barrier in treating acute myeloid leukemia (AML). Metabolic adaptations have been shown to contribute to therapy resistance. ...However, little is known about whether specific therapies cause specific metabolic changes. We established cytarabine-resistant (AraC-R) and Arsenic trioxide-resistant (ATO-R) AML cell lines, displaying distinct cell surface expression and cytogenetic abnormalities. Transcriptomic analysis revealed a significant difference in the expression profiles of ATO-R and AraC-R cells. Geneset enrichment analysis showed AraC-R cells rely on OXPHOS, while ATO-R cells on glycolysis. ATO-R cells were also enriched for stemness gene signatures, whereas AraC-R cells were not. The mito stress and glycolytic stress tests confirmed these findings. The distinct metabolic adaptation of AraC-R cells increased sensitivity to the OXPHOS inhibitor venetoclax. Cytarabine resistance was circumvented in AraC-R cells by combining Ven and AraC. In vivo, ATO-R cells showed increased repopulating potential, leading to aggressive leukemia compared to the parental and AraC-R. Overall, our study shows that different therapies can cause different metabolic changes and that these metabolic dependencies can be used to target chemotherapy-resistant AML.
•AML cell line MV4-11 exposed to ATO/Cytarabine acquire distinct resistant phenotypes and metabolic dependencies.•Cytarabine-resistant cells (AraC-R) rely on mitochondrial metabolism and are sensitive to OXPHOS inhibitor Venetoclax.•ATO-resistant cells (ATO-R) display increased glycolytic capacity and acquisition of leukemic stem cell phenotype.•Glycolytic inhibitor 2DG improves ATO sensitivity in ATO-R cells.
Achieving early molecular response (EMR) has been shown to be associated with better event free survival in patients with chronic phase chronic myeloid leukemia (CP-CML) on Imatinib therapy. We ...prospectively evaluated the factors influencing the 2-year failure free survival (FFS) and EMR to imatinib therapy in these patients including day29 plasma Imatinib levels, genetic variants and the gene expression of target genes in imatinib transport and biotransformation. Patients with low and intermediate Sokal score had better 2-year FFS compared to those with high Sokal Score (p = 0.02). Patients carrying ABCB1-C1236T variants had high day29 plasma imatinib levels (P = 0.005), increased EMR at 3 months (P = 0.044) and a better 2 year FFS (P = 0.003) when compared to those with wild type genotype. This translates to patients with lower ABCB1 mRNA expression having a significantly higher intracellular imatinib levels (P = 0.029). Higher day29 plasma imatinib levels was found to be strongly associated with patients achieving EMR at 3 months (P = 0.022), MMR at 12 months (P = 0.041) which essentially resulted in better 2-year FFS (p = 0.05). Also, patients who achieved EMR at 3 months, 6 months and MMR at 12 months had better FFS when compared to those who did not. This study suggests the incorporation of these variables in to the imatinib dosing algorithm as predictive biomarkers of response to Imatinib therapy.
Early complications post hematopoietic stem cell transplantation (HSCT) such as sinusoidal obstruction syndrome (SOS) and graft versus host disease (GVHD) can be life threatening. Although several ...biomarkers have been identified to correlate with these complications and their response to treatment, these are yet to be used in clinical practice. Here, we evaluated circulating endothelial cells (CECs) (n = 26) and plasma biomarkers (ST2, REG3α, VCAM1, ICAM1, TIM3) (N = 210) at early time points, to determine their association with early complications post‐HSCT. Elevated CEC counts at the end of conditioning was associated with GVHD, indicating endothelial damage during HSCT. Plasma levels of REG3α, VCAM1, ICAM1, and TIM3 on day 14 (D14) and D14 ICAM1 and D28 ST2 were significantly higher in patients with SOS and aGVHD, respectively. Upon sub‐group analysis, D28 ST2, D14/D28 REG3α, and D14 ICAM1 levels were significantly higher in patients with gastrointestinal GVHD, while D28 ST2 was higher in those with skin/liver GVHD. High ST2 levels on D28 was significantly associated with non‐relapse mortality (NRM) and overall survival. Our results suggest that elevated ST2 levels on D28 could predict the likelihood of developing aGVHD and could influence NRM and OS.
PDF
