Keywords::
Acute myeloid leukemia (AML) is an aggressive hematologic malignancy and is the most common form of acute leukemia in adults Citation[1]. While there have been advances in the treatment of certain hematological malignancies, the prognosis of AML remains grim. For example, patients older than 60 years have a 2-year survival of less than 20% despite aggressive treatment. Thus, further research is warranted into developing novel targeted AML therapies with greater efficacy and less toxicity than standard chemotherapy regimens.
A challenging aspect of cancer therapy is selectively eliminating the cancer stem cell. Much of the evidence for the cancer stem cell hypothesis has come from studies on hematologic malignancies. Dick and colleagues found leukemia stem cells in a small compartment of the peripheral blood in AML patients Citation[2]. These quiescent malignant cells are capable of self-renewal and differentiation. They are often chemoresistant and are a likely cause of disease relapse. Therefore, therapeutic strategies that target the AML stem cell as well as the bulk AML cell population preferentially over normal stem cells could improve the outcome of this disease.
A custom library of the US FDA-approved drugs for agents cytotoxic to TEX and M9-ENL1 cells was recently screened to identify such therapies. These leukemic cell lines have features of leukemia stem cells, including hierarchal differentiation and marrow repopulation Citation[3,4]. This screen identified tigecycline Citation[5], a recently characterized antimicrobial agent of the novel glycylcycline class that is active against a range of Gram-positive and Gram-negative bacteria, particularly drug-resistant pathogens Citation[6]. Subsequent validation studies demonstrated that tigecycline targeted AML cells and AML stem cells preferentially over normal hematopoietic stem cells in vitro and in vivo.
To understand the antileukemic mechanism of action of tigecycline, the authors performed genome-wide drug-induced haploinsufficiency profiling in Saccharomyces cerevisiae and identified mitochondrial translation inhibition as the mechanism of tigecycline cell death in eukaryotic cells. This finding was validated in AML cell lines and primary AML patient samples. Moreover, it was shown that mitochondrial translation inhibition is functionally important for cell death of leukemia progenitors and stem cells using both chemical and genetic approaches. Finally, it was demonstrated that the heightened sensitivity of AML cells to the inhibition of mitochondrial translation was a derivative of increased mitochondrial mass in these cells Citation[5].
In addition to nuclear DNA, eukaryotic cells have circular mitochondrial DNA located within the mitochondria that is 16.6 kB in length and consists only of exons Citation[7]. Mitochondrial DNA encodes two rRNAs, 22 tRNAs and 13 of the 90 proteins that comprise the mitochondrial respiratory chain and oxidative phosphorylation pathway. Of note, the remaining proteins of the respiratory chain are encoded in the nucleus and imported into the mitochondria where they are assembled into the functional complexes of the electron transport chain.
Mitochondrial protein translation occurs on mitochondrial ribosomes. Mitochondrial ribosomes differ from bacterial and eukaryotic cytosolic ribosomes in their biochemical properties and structure Citation[8]. Compared with bacterial ribosomes, mitochondrial ribosomes have approximately half as much rRNA and over twice the amount of protein. Mitochondrial ribosomal proteins are encoded by nuclear genes and translated in the cytosol. Once translated, these proteins are imported into the mitochondria where they join two rRNA molecules to form the functional ribosomes of the mitochondria. Many of these mitochondrial ribosomal proteins have no similar analogs in bacterial or cytosolic ribosomes. Similar numbers of proteins are in both mitochondrial and cytosolic ribosomes. Although mitochondrial ribosomes differ structurally from cytoplasmic and bacterial ribosomes, they function similarly.
Mitochondrial protein synthesis uses its own initiation and elongation factors Citation[9,10]. Therefore, as part of these studies, the impact of genetically inhibiting mitochondrial translation in AML cells was evaluated. Here, the initiation (IF-3) and elongation (EF-Tu) factors of mitochondrial translation in leukemia cells were knocked down. Interestingly, IF-3 knockdown did not inhibit mitochondrial translation, while EF-Tu did so with similar consequences as chemical inhibition by tigecycline. Therefore, future work exploring the translational control of mitochondria in cancer should carefully assess the functional importance of various initiation and elongation factors in the context of disease.
Over the last decade, there have been multiple advances in understanding the role of mitochondria in cancer physiology Citation[11]. Mitochondria are the powerhouse of the cell, enabling energy production, and therefore, are important for the survival of eukaryotic cells Citation[12]. The Warburg hypothesis implies that malignant cells rely on glycolysis and do not require oxidative phosphorylation for survival. Leukemia cells are unique in their mitochondrial characteristics and their reliance on oxidative phosphorylation.
In the context of leukemia metabolism, recent work highlighted the dependence of leukemia cells on fatty acid oxidation for their supply of energy Citation[13]. These results complement the findings that increased oxidation of fatty acids may render leukemia cells more susceptible to disruption of the oxidative phosphorylation pathways via reductions in mitochondrial protein synthesis.
AML cells and AML stem cells have increased mitochondrial mass and oxygen consumption compared with normal hematopoietic cells. Moreover, AML cells with the highest mitochondrial mass are most sensitive to tigecycline treatment in culture. Therefore, the therapeutic potential of mitochondrial translation inhibition may be best suited to a population of AML patients with higher intrinsic mitochondrial biogenesis Citation[5].
To initially test the proof-of-concept of inhibiting mitochondrial translation in AML, tigecycline may be a useful candidate given its known pharmacokinetics and toxicology in humans as an antimicrobial agent. Tigecycline is routinely administered as a 50-mg intravenous dose every 12 h without significant toxicity, but higher doses have also been used safely. For example, intravenous doses of 300 mg are well tolerated except for mild nausea, resulting in a Cmax of 2.82 µg/ml (5 µM) Citation[14], a concentration within the range required for antileukemic effects. Toxicology studies in animals also suggest a potential for antileukemic activity. Rats receiving >30 mg/kg/day for 2 weeks developed reversible anemia, thrombocytopenia and leucopenia with hypocellular bone marrow Citation[15]. However, these higher concentrations of tigecycline are not used in the treatment of infection, potentially explaining why anticancer activity has not been previously reported with the drug.
A Phase I dose escalation trial of intravenous tigecycline in patients with relapsed and refractory AML is currently underway. In the context of this trial, pharmacodynamics studies are conducted to determine the effects of tigecycline on mitochondrial protein synthesis in the patient's AML cells.
Tigecycline has been shown to have synergistic activity with the standard AML agent daunorubicin when combined, both in vitro and in vivo. Therefore, in later clinical trials, tigecycline could be tested in combination with standard induction or reinduction chemotherapy. In addition, future preclinical and clinical investigation could also evaluate mitochondrial protein synthesis inhibition as a therapeutic strategy for other malignancies with increased dependence on oxidative phosphorylation.
Thus, in summary, AML cells and stem cells appear significantly dependent on oxidative phosphorylation for their supply of energy and have unique mitochondrial characteristics. As such, targeting AML tumor metabolism by inhibiting mitochondrial protein synthesis is a plausible therapeutic strategy.
Financial & competing interests disclosure
This work was supported by the Canadian Stem Cell Network, the Canadian Institutes for Health Research, the Leukemia and Lymphoma Society, the NIH (NCI 1R01CA157456), the Terry Fox Foundation, MaRS Innovation, the Ontario Institute of Cancer Research with funding provided by the Ontario Ministry of Research and Innovation, the Princess Margaret Hospital Foundation, and the Ministry of Long Term Health and Planning in the Province of Ontario. M Škrtic´holds a Canada Graduate Scholarship from the Canadian Institutes of Health Research. AD Schimmer is a Leukemia and Lymphoma Society Scholar in Clinical Research. M Škrtic´ and AD Schimmer are named as inventors on patent applications claiming the use of tigecycline for the treatment of hematologic malignancies. The authors are named as inventors on patent applications claiming the use of tigecycline for the treatment of leukemia. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.
No writing assistance was utilized in the production of this manuscript.
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