Targeting the Mitochondrial Metallochaperone Cox17 Reduces DNA Methylation and Promotes AML Differentiation through a Copper Dependent Mechanism
Notice bibliographique
Résumé
Abstract The majority of mitochondrial proteins are encoded in the nucleus, translated in the cytoplasm and imported into the mitochondria. A subset of cysteine-rich proteins destined for the mitochondrial intermembrane space are oxidized and folded by the Mitochondrial IMS Assembly (MIA) pathway. We found that genes encoding substrates of the MIA pathway are over-expressed in leukemic stem cells compared to bulk AML cells. Therefore, we assessed the effects of inhibiting the MIA pathway in AML by targeting the FAD linked sulfhydryl oxidase ALR, an integral part of the MIA machinery. Knockdown of ALR with shRNA reduced the growth and viability of OCI-AML2, TEX and NB4 leukemia cells. In addition, ALR knockdown reduced engraftment of TEX cells into mouse marrow, demonstrating an effect on the leukemia initiating cells. The small molecule selective ALR inhibitor, MitoBloCK-6 killed AML cells (IC50 of 5-10 mM) and preferentially reduced the clonogenic growth of primary AML cells over normal hematopoietic cells. MitoBloCK-6 treatment (80 mg/kg i.p. 5 of 7 days x 2 weeks) of mice engrafted with primary AML cells strongly reduced the leukemic burden without changing mouse body weight, serum chemistries, or organ histology. In contrast, MitoBloCK-6 did not change engraftment of normal cord blood in similar experiments. As evidenced by secondary transplants, MitoBloCK-6 also targeted leukemic stem cells. As expression levels of ALR substrates are increased in AML stem cells we assessed the effects of ALR inhibition on differentiation in AML. Genetic or chemical inhibition of ALR induced the differentiation of AML cells as evidenced by changes in gene expression, increased differentiation associated CD surface marker expression and increased non-specific esterase. Interrogation of the effects of ALR inhibition on its substrates identified the mitochondrial copper chaperone, Cox17 as the primary downstream target in leukemic cells. Genetic or chemical inhibition of ALR selectively reduced levels of Cox17 protein. Validating the functional importance of these findings, knockdown of Cox17 phenocopied ALR inhibition and reduced AML proliferation and induced AML differentiation. Cox17 is a copper metallochaperone that promotes respiratory chain complex IV assembly by loading copper into the respiratory complex. COX17 knockdown slightly reduced the complex IV enzymatic activity, but did not change basal oxygen consumption or ROS production. However, COX17 knockdown increased intracellular levels of free copper 16-fold as measured by atomic mass spectrometry. Copper is a known inhibitor of adenosylhomocysteinase, a key enzyme involved in the preservation of S-adenosylmethionine (SAM):S-adenosylhomocysteine (SAH) ratio in cells. SAM is a global methyl donor and is critical for DNA methylation. Knockdown of COX17 or ALR inhibition with MitoBloCK-6 decreased levels of SAM and reduced DNA methylation in AML cells. Likewise, the enzymatic activity of adenosylhomocysteinase was reduced in OCI-AML2 cells after MitoBloCK-6 treatment. Importantly, co-treatment with the copper chelator, penicillamine, rescued reductions in SAM, DNA methylation, and cell viability after COX17 knockdown or MitoBloCK-6 treatment. Thus, we have discovered a novel copper-dependent mechanism by which mitochondrial pathways regulate epigenetics and stemness in AML. Moreover, inhibitors of ALR or COX17 may be a novel therapeutic strategy to promote the differentiation of AML cells and stem cells. Disclosures Schimmer: Otsuka Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Medivir AB: Research Funding; Jazz Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees.
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Comment cette classification a été obtenuedéplier
Prédiction machine sur la base complète
Imitation des enseignantsNi prévalence calibrée, ni vérité terrain. Validation humaine à venir. Le volet Gemma est une étiquette directe du modèle pour chaque travail de la base, lue sur la notice réduite au titre. Le volet Codex est un classifieur appris des 10 348 étiquettes directes de Codex et calibré sur les taux pondérés de l'échantillon; les champs sans appui suffisant ne portent aucun appel Codex. Le mode candidate est l'union des deux volets; le consensus est leur intersection. Ces sorties portent le statut machine_predicted_unvalidated et ne sont pas des étiquettes humaines.
Scores du classifieur distillé par catégorie (deux têtes)
| Catégorie | Codex | Gemma |
|---|---|---|
| Métarecherche | 0,000 | 0,000 |
| Méta-épidémiologie (sens strict) | 0,000 | 0,000 |
| Méta-épidémiologie (sens large) | 0,000 | 0,000 |
| Bibliométrie | 0,000 | 0,000 |
| Études des sciences et des technologies | 0,000 | 0,000 |
| Communication savante | 0,000 | 0,000 |
| Science ouverte | 0,000 | 0,000 |
| Intégrité de la recherche | 0,000 | 0,001 |
| Charge utile insuffisante (le modèle a refusé de juger) | 0,002 | 0,000 |
Scores machine (provisoires)
Les deux têtes enseignantes du modèle étudiant, lues sur ce travail. Un score ordonne la base pour la relecture; il n'affirme jamais une catégorie, et le statut de validation accompagne chaque rangée tel quel.
Scores de référence d'un modèle non mature (critères de maturité non atteints, 7 itérations). Un score ordonne; il n'affirme jamais une catégorie.
score_only:v0-immature-baseline · tel quel depuis la passe de notation : score_only signifie que le nombre peut ordonner les travaux, et qu'aucune étiquette de catégorie n'en découleClassification
machine, non validéePrédiction automatique; un appel candidat d’une seule source (Gemma direct ou Codex distillé), pas un consensus.
Le détail, modèle par modèle et score par score, se trouve en fin de page sous « Comment cette classification a été obtenue ».