Junior Research Fellow, Christian Medical College
Approaches to overcome drug resistance in Acute Myeloid Leukaemia
Acute myeloid leukaemia is a heterogenous aggressive blood cancer with relatively poor outcome after induction chemotherapy.This is mainly due to development of drug resistance by the malignant cells and also because of malignant cells which have stem cell(leukaemia stem cells[LSC]) like characteristics. These resistant cells evade chemotherapy and give rise leukaemia again a phenomenon know as relapse. Many factors contribute to the development of drug resistance such as enhanced DNA damage repair, high proliferative capacity,increased expression of anti-apoptotic factors, increased drug effluxing capacity,abberrant metabolic function and increased anti-oxidant capacity.Apart from these factors, the microenvironment the healthy cells that surround these malignant cells act as guardian and prevent the damage caused by chemotherapeutic drugs. Our lab preliminary focuses on cell intrinsic factors that help in clearance of anti-oxidants,drug efflux,differentiation, proliferation of malignant cells and strategies to eliminate leukaemic stem cells. We have reported the role of NRF2 (a master regulator of anti-oxidants)in mediating chemo resistance in leukemic cells also the role of Drug transporter ABCB6 in chemo resistance. Currently we are investigating the role of nuclear receptors a cell intrinsic factors known to be involved in differentiation and proliferation . These nuclear receptors can be activated and deactivated using known clinically available molecules.This makes these receptors attractive targets in the leukaemia setup.
Abstract: It is well established that expression of multi-drug resistance (MDR) proteins (MDR1, BCRP, MDR3, MRP1 and LRP) in leukemic blasts correlates with AML patients clinical response. Assuming that leukemic stem cells (LSC) are resistant to chemotherapy and responsible for relapse it might be clinically relevant to evaluate the expression level of MDR proteins in LSC and relate it to the clinical outcome.Bone marrow samples from 26 patients with de novo AML were labeled with antibodies to distinguish CD34+CD38-CD123+ LSC population and with antibodies against MDR1, BCRP, MDR3, MRP1 or LRP proteins. Multicolor flow cytometry was applied to evaluate the expression of MDR proteins in blasts and LSC.Nine of 26 patients with AML attained CR (30%). High negative correlation was found between MDR1 and LRP expression in blasts and the patients remission. MDR proteins were expressed more frequently in LSC than in leukemic blasts. High negative correlation was also observed between remission achievement and MRP1 expression in LSC.Our data presents for the very first time the high negative correlation between MRP1 protein expression in LSC and AML patients' remission. It does strongly suggest that MRP1 expression in LSC is an adverse prognostic marker in patients with de novo AML. This article is protected by copyright. All rights reserved.
Pub.: 15 Aug '17, Pinned: 05 Oct '17
Abstract: Acute myeloid leukemia (AML) is a heterogeneous malignant disorder derived from the myeloid hematopoietic cells that accounts for ~80% of all adult acute leukemia. Numerous studies have shown that drug resistance not only exists against conventional chemotherapeutic drugs, but also limits the efficacy of new biological agents. Therefore, it is important to identify the mechanisms behind chemoresistance and seek therapeutic strategies to enhance efficacy in AML chemotherapy. MicroRNA (miR)-217 has been recognized as a tumor suppressor that is downregulated in various types of cancer, however the mechanisms behind the expression and function of miR-217 in AML have not yet been recognized. The expression of miR-217 was determined by quantitative polymerase chain reaction (qPCR). Following transfection with miR-217 mimics, an MTT assay, chemosensitivity assay, cell apoptosis assay and western blot analysis were performed in AML cell lines. Functional assays were also performed to explore the effects of endogenous Kirsten rat sarcoma viral oncogene homolog (KRAS) in AML. The results revealed that miR-217 was downregulated in patients with AML. Overexpression of miR-217 inhibited cellular proliferation and enhanced cell chemosensitivity to doxorubicin by the cell apoptosis pathway in AML cells. A dual-luciferase reporter assay demonstrated that KRAS was a direct target gene of miR-217 in vitro. qPCR and western blot analysis revealed that miR-217 negatively regulated KRAS protein expression, but had no impact on KRAS mRNA expression. Knockdown of KRAS expression markedly suppressed AML cellular proliferation, and enhanced cell chemosensitivity to doxorubicin via the cell apoptosis pathway. These findings indicate that miR-217 functions as a tumor suppressor in AML by directly targeting KRAS. Therefore, miR-217-based therapeutic strategies may provide a novel strategy for the enhancement of efficacy in the treatment of AML.
Pub.: 11 Jun '17, Pinned: 05 Oct '17
Abstract: 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.
Pub.: 16 May '17, Pinned: 05 Oct '17
Abstract: Drug resistance and relapse are considered to be the major reasons for treatment failure in acute myeloid leukemia (AML). There is limited data on the role of ABC transporter expression on in vitro sensitivity to cytarabine (Ara-C) and daunorubicin (Dnr) in primary AML cells.RNA expression levels of 12 ABC transporters were analyzed by real-time quantitative PCR in 233 de novo adult acute myeloid leukemia patients. Based on cytarabine or Dnr IC50, the samples were categorized as sensitive, intermediate and resistant. Role of candidate ABC transporter RNA expression on in vitro cytotoxicity, treatment outcome post therapy as well as the influence of various prognostic markers on ABC transporter expression were analyzed.Expression of ABCC3 and ABCB6 were significantly higher in Dnr-resistant samples when compared with Dnr-sensitive samples. Increased ABCC1 expression was associated with poor disease-free survival in this cohort of patients.This comprehensive analysis suggests ABCC1, ABCC3, ABCB6 and ABCA5 as probable targets which can be modulated for improving chemotherapeutic responses.
Pub.: 24 Jan '17, Pinned: 05 Oct '17
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