Targeting mutations that trigger acute myeloid leukaemia (AML) has emerged as a refined therapeutic approach in recent years. Enasidenib (Idhifa) is the first selective inhibitor of mutated forms of isocitrate dehydrogenase 2 (IDH2) approved against relapsed/refractory AML. In addition to its use as monotherapy, a combination trial of enasidenib with standard intensive induction therapy (daunorubicin + cytarabine) is being evaluated. This study aimed to decipher enasidenib off-target molecular mechanisms involved in anthracycline resistance, such as reduction by carbonyl reducing enzymes (CREs) and drug efflux by ATP-binding cassette (ABC) transporters. We analysed the effect of enasidenib on daunorubicin (Daun) reduction by several recombinant CREs and different human cell lines expressing aldo-keto reductase 1C3 (AKR1C3) exogenously (HCT116) or endogenously (A549 and KG1a). Additionally, A431 cell models overexpressing ABCB1, ABCG2, or ABCC1 were employed to evaluate enasidenib modulation of Daun efflux. Furthermore, the potential synergism of enasidenib over Daun cytotoxicity was quantified amongst all the cell models. Enasidenib selectively inhibited AKR1C3-mediated inactivation of Daun in vitro and in cell lines expressing AKR1C3, as well as its extrusion by ABCB1, ABCG2, and ABCC1 transporters, thus synergizing Daun cytotoxicity to overcome resistance. This work provides in vitro evidence on enasidenib-mediated targeting of the anthracycline resistance actors AKR1C3 and ABC transporters under clinically achievable concentrations. Our findings may encourage its combination with intensive chemotherapy and even suggest that the effectiveness of enasidenib as monotherapy against AML could lie beyond the targeting of mIDH2.

• Keywords
• ABC transporters, AKR1C3, AML, Enasidenib, IDH inhibitor,
• MeSH
• ATP-Binding Cassette Transporters metabolism   MeSH
• Adenosine Triphosphate MeSH
• Leukemia, Myeloid, Acute * drug therapy   genetics   MeSH
• Anthracyclines MeSH
• Cytarabine therapeutic use   MeSH
• Daunorubicin * pharmacology   MeSH
• Isocitrate Dehydrogenase genetics   metabolism   therapeutic use   MeSH
• Humans MeSH
• Antibiotics, Antineoplastic therapeutic use   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• ATP-Binding Cassette Transporters MeSH
• Adenosine Triphosphate MeSH
• Anthracyclines MeSH
• Cytarabine MeSH
• Daunorubicin * MeSH
• enasidenib MeSH Browser
• Isocitrate Dehydrogenase MeSH
• Antibiotics, Antineoplastic MeSH

Midostaurin is an FMS-like tyrosine kinase 3 receptor (FLT3) inhibitor that provides renewed hope for treating acute myeloid leukaemia (AML). The limited efficacy of this compound as a monotherapy contrasts with that of its synergistic combination with standard cytarabine and daunorubicin (Dau), suggesting a therapeutic benefit that is not driven only by FLT3 inhibition. In an AML context, the activity of the enzyme aldo-keto reductase 1C3 (AKR1C3) is a crucial factor in chemotherapy resistance, as it mediates the intracellular transformation of anthracyclines to less active hydroxy metabolites. Here, we report that midostaurin is a potent inhibitor of Dau inactivation mediated by AKR1C3 in both its recombinant form as well as during its overexpression in a transfected cell model. Likewise, in the FLT3- AML cell line KG1a, midostaurin was able to increase the cellular accumulation of Dau and significantly decrease its metabolism by AKR1C3 simultaneously. The combination of those mechanisms increased the nuclear localization of Dau, thus synergizing its cytotoxic effects on KG1a cells. Our results provide new in vitro evidence of how the therapeutic activity of midostaurin could operate beyond targeting the FLT3 receptor.

• Keywords
• AKR1C3, AML therapy, Anthracyclines, Midostaurin, Multidrug resistance,
• MeSH
• Leukemia, Myeloid, Acute drug therapy   enzymology   genetics   pathology   MeSH
• Biotransformation MeSH
• Daunorubicin metabolism   pharmacology   MeSH
• HCT116 Cells MeSH
• Enzyme Inhibitors pharmacology   MeSH
• Colorectal Neoplasms drug therapy   enzymology   genetics   pathology   MeSH
• Humans MeSH
• Aldo-Keto Reductase Family 1 Member C3 antagonists & inhibitors   genetics   metabolism   MeSH
• Antineoplastic Combined Chemotherapy Protocols pharmacology   MeSH
• Staurosporine analogs & derivatives   pharmacology   MeSH
• Drug Synergism MeSH
• fms-Like Tyrosine Kinase 3 antagonists & inhibitors   genetics   metabolism   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• AKR1C3 protein, human MeSH Browser
• Daunorubicin MeSH
• FLT3 protein, human MeSH Browser
• Enzyme Inhibitors MeSH
• midostaurin MeSH Browser
• Aldo-Keto Reductase Family 1 Member C3 MeSH
• Staurosporine MeSH
• fms-Like Tyrosine Kinase 3 MeSH

Over the last few years, aldo-keto reductase family 1 member C3 (AKR1C3) has been associated with the emergence of multidrug resistance (MDR), thereby hindering chemotherapy against cancer. In particular, impaired efficacy of the gold standards of induction therapy in acute myeloid leukaemia (AML) has been correlated with AKR1C3 expression, as this enzyme metabolises several drugs including anthracyclines. Therefore, the development of selective AKR1C3 inhibitors may help to overcome chemoresistance in clinical practice. In this regard, we demonstrated that Bruton's tyrosine kinase (BTK) inhibitors ibrutinib and acalabrutinib efficiently prevented daunorubicin (Dau) inactivation mediated by AKR1C3 in both its recombinant form as well as during its overexpression in cancer cells. This revealed a synergistic effect of BTK inhibitors on Dau cytotoxicity in cancer cells expressing AKR1C3 both exogenously and endogenously, thus reverting anthracycline resistance in vitro. These findings suggest that BTK inhibitors have a novel off-target action, which can be exploited against leukaemia through combination regimens with standard chemotherapeutics like anthracyclines.

• Keywords
• AKR1C3, Bruton’s tyrosine kinase, acalabrutinib, anthracyclines, ibrutinib, multidrug resistance,
• Publication type
• Journal Article MeSH