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D,L-Methadone causes leukemic cell apoptosis via an OPRM1-triggered increase in IP3R-mediated ER Ca²⁺ release and decrease in Ca²⁺ efflux, elevating [Ca²⁺]_i.

ABSTRACT: The search continues for improved therapy for acute lymphoblastic leukemia (aLL), the most common malignancy in children. Recently, D,L-methadone was put forth as sensitizer for aLL chemotherapy. However, the specific target of D,L-methadone in leukemic cells and the mechanism by which it induces leukemic cell apoptosis remain to be defined. Here, we demonstrate that D,L-methadone induces leukemic cell apoptosis through activation of the mu1 subtype of opioid receptors (OPRM1). D,L-Methadone evokes IP3R-mediated ER Ca²⁺ release that is inhibited by OPRM1 loss. In addition, the rate of Ca²⁺ extrusion following D,L-methadone treatment is reduced, but is accelerated by loss of OPRM1. These D,L-methadone effects cause a lethal rise in [Ca²⁺]_i that is again inhibited by OPRM1 loss, which then prevents D,L-methadone-induced apoptosis that is associated with activation of calpain-1, truncation of Bid, cytochrome C release, and proteolysis of caspase-3/12. Chelating intracellular Ca²⁺ with BAPTA-AM reverses D,L-methadone-induced apoptosis, establishing a link between the rise in [Ca²⁺]_i and D,L-methadone-induced apoptosis. Altogether, our findings point to OPRM1 as a specific target of D,L-methadone in leukemic cells, and that OPRM1 activation by D,L-methadone disrupts IP3R-mediated ER Ca²⁺ release and rate of Ca²⁺ efflux, causing a rise in [Ca²⁺]_i that upregulates the calpain-1-Bid-cytochrome C-caspase-3/12 apoptotic pathway.

SUBMITTER: Lee J

PROVIDER: S-EPMC7806773 | biostudies-literature | 2021 Jan

REPOSITORIES: biostudies-literature

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D,L-Methadone causes leukemic cell apoptosis via an OPRM1-triggered increase in IP3R-mediated ER Ca2+ release and decrease in Ca2+ efflux, elevating [Ca2+]i.

Lee JungKwon J Rosales Jesusa L JL Byun Hee-Guk HG Lee Ki-Young KY

Scientific reports 20210113 1

The search continues for improved therapy for acute lymphoblastic leukemia (aLL), the most common malignancy in children. Recently, D,L-methadone was put forth as sensitizer for aLL chemotherapy. However, the specific target of D,L-methadone in leukemic cells and the mechanism by which it induces leukemic cell apoptosis remain to be defined. Here, we demonstrate that D,L-methadone induces leukemic cell apoptosis through activation of the mu1 subtype of opioid receptors (OPRM1). D,L-Methadone evo ...[more]

PMID: 33441856

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Project description:Mitochondrial Ca2+ handling is accomplished by balancing Ca2+ uptake, primarily via the Ru360-sensitive mitochondrial calcium uniporter (MCU), Ca2+ buffering in the matrix and Ca2+ efflux mainly via Ca2+ ion exchangers, such as the Na+/Ca2+ exchanger (NCLX) and the Ca2+/H+ exchanger (CHE). The mechanism of CHE in cardiac mitochondria is not well-understood and its contribution to matrix Ca2+ regulation is thought to be negligible, despite higher expression of the putative CHE protein, LETM1, compared to hepatic mitochondria. In this study, Ca2+ efflux via the CHE was investigated in isolated rat cardiac mitochondria and permeabilized H9c2 cells. Mitochondria were exposed to (a) increasing matrix Ca2+ load via repetitive application of a finite CaCl2 bolus to the external medium and (b) change in the pH gradient across the inner mitochondrial membrane (IMM). Ca2+ efflux at different matrix Ca2+ loads was revealed by inhibiting Ca2+ uptake or reuptake with Ru360 after increasing number of CaCl2 boluses. In Na+-free experimental buffer and with Ca2+ uptake inhibited, the rate of Ca2+ efflux and steady-state free matrix Ca2+ [mCa2+]ss increased as the number of administered CaCl2 boluses increased. ADP and cyclosporine A (CsA), which are known to increase Ca2+ buffering while maintaining a constant [mCa2+]ss, decreased the rate of Ca2+ efflux via the CHE, with a significantly greater decrease in the presence of ADP. ADP also increased Ca2+ buffering rate and decreased [mCa2+]ss. A change in the pH of the external medium to a more acidic value from 7.15 to 6.8∼6.9 caused a twofold increase in the Ca2+ efflux rate, while an alkaline change in pH from 7.15 to 7.4∼7.5 did not change the Ca2+ efflux rate. In addition, CHE activation was associated with membrane depolarization. Targeted transient knockdown of LETM1 in permeabilized H9c2 cells modulated Ca2+ efflux. The results indicate that Ca2+ efflux via the CHE in cardiac mitochondria is modulated by acidic buffer pH and by total matrix Ca2+. A mechanism is proposed whereby activation of CHE is sensitive to changes in both the matrix Ca2+ buffering system and the matrix free Ca2+ concentration.

Dataset Information

D,L-Methadone causes leukemic cell apoptosis via an OPRM1-triggered increase in IP3R-mediated ER Ca²⁺ release and decrease in Ca²⁺ efflux, elevating [Ca²⁺]_i.

Publications

D,L-Methadone causes leukemic cell apoptosis via an OPRM1-triggered increase in IP3R-mediated ER Ca<sup>2+</sup> release and decrease in Ca<sup>2+</sup> efflux, elevating [Ca<sup>2+</sup>]<sub>i</sub>.

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