REVIEW ARTICLE The Role of AMPK/mTOR Modulators in the … · 2018. 6. 19. · tivity of protein kinase B/Akt were observed, and siRNA-mediated down-modulation of Akt in HL-60 cells

Send Orders for Reprints to [email protected]

Current Medicinal Chemistry, 2018, 25, 1-20 1

REVIEW ARTICLE

0929-8673/18 $58.00+.00 © 2018 Bentham Science Publishers

The Role of AMPK/mTOR Modulators in the Therapy of Acute Mye-loid Leukemia

Dora Visnjic*, Vilma Dembitz and Hrvoje Lalic

Department of Physiology and Croatian Institute for Brain Research, School of Medicine, University of Zagreb, Salata 12, 10 000 Zagreb, Croatia

A R T I C L E H I S T O R Y

Received: July 14, 2017 Revised: January 01, 2018 Accepted: January 11, 2018 DOI: 10.2174/0929867325666180117105522

Abstract: Differentiation therapy of acute promyelocytic leukemia with all-trans retinoic acid represents the most successful pharmacological therapy of acute myeloid leukemia (AML). Numerous studies demonstrate that drugs that inhibit mechanistic target of ra-pamycin (mTOR) and activate AMP-kinase (AMPK) have beneficial effects in promoting differentiation and blocking proliferation of AML. Most of these drugs are already in use for other purposes; rapalogs as immunosuppressants, biguanides as oral antidiabetics, and 5-amino-4-imidazolecarboxamide ribonucleoside (AICAr, acadesine) as an exercise mi-metic. Although most of these pharmacological modulators have been widely used for decades, their mechanism of action is only partially understood. In this review, we sum-marize the role of AMPK and mTOR in hematological malignancies and discuss the pos-sible role of pharmacological modulators in proliferation and differentiation of leukemia cells.

Keywords: AML, AMPK, mTOR, rapamycin, metformin, AICAR, differentiation.

1. INTRODUCTION

Acute myeloid leukemia (AML) is a heterogenous group of malignant disorders that are characterized by uncontrolled proliferation of blasts that are blocked at an early stage of differentiation. The accumulation of immature myeloblasts in the bone marrow and periph-eral blood occurs at the expense of normal terminally differentiated blood cells resulting in progressive ane-mia, thrombocytopenia and neutropenia. AML is the most common type of acute leukemia in adults with the incidence of 3-5 cases per 100 000, a slight male pre-dominance and a median age at diagnosis of approxi-mately 70 years. The WHO classification that is cur-rently in use, defines 7 main subtypes of AML accord-ing to genetic abnormalities, relationship to previous conditions and therapies, and cytochemical and mor-phological characteristics. The older classification, which is still in use, is the French-American-British

*Address correspondence to this author at the Department of Physi-ology and Croatian Institute for Brain Research, School of Medi-cine, University of Zagreb, Salata 12, 10 000 Zagreb, Croatia; Tel: 385-1-4596-831; Fax: 385-1-4590-207; E-mail: [email protected]

Classification (FAB). FAB classification is based pre-dominantly on cytomorphological features of leukemia cells, and the 8 AML subtypes (M0-M7) defined in such manner have been integrated into the WHO classi-fication.

The standard therapy for acute myeloid leukemia (AML) has not changed significantly for the past four decades and consists of 7-10 days of cytarabine com-bined with 3 days of an anthracycline as a remission induction therapy. In patients <60 years of age, this therapy results in complete remission rates of 60-90%. Several courses of high dose cytarabine or allogeneic hematopoietic stem cell transplantation may be used as a consolidation therapy. In elderly patients for whom intensive therapy is not appropriate, treatment remains unsatisfactory and includes low-dose cytarabine or de-methylation therapy with supportive care [reviewed in 1, 2].

Several new agents including monoclonal antibod-ies, drugs targeting signaling pathways, and epigenetic regulators are in early clinical trials for AML. How-ever, the most successful pharmacological therapy for

Dora Visnjic

Replace

Dora Visnjic

Replace

c

2 Current Medicinal Chemistry, 2018, Vol. 25, No. 00 Visnjic et al.

AML is still retinoic acid-based differentiation therapy of acute promyelocytic leukemia (APL), a particular subtype of AML characterized by t(15;17) transloca-tion. Once fatal disease with a high incidence of early hemorrhagic death, APL has now been successfully treated with all-trans retinoic acid (ATRA) and chemo-therapy inducing complete remission rates of 90% and cure rates of approximately 80% [3]. The clinical out-come of the disease has been further improved by the introduction of arsenic trioxide (ATO) into the treat-ment of refractory or relapsed APL. Moreover, the lat-est results of the clinical studies instituting ATO plus ATRA as a first-line treatment suggest the possibility of leukemia treatment without any DNA-damaging chemotherapy [4].

The biochemical mechanisms responsible for differ-entiation of leukemia cells are not entirely understood. The high sensitivity of APL to ATRA is ascribed to the presence of the fusion protein promyelocytic leukemia (PML)/retinoic acid receptor α (RARα), which is en-coded by a specific chromosomal translocation involv-ing PML gene on chromosome 15 and RARA gene on chromosome 17. In cells containing t(15;17) transloca-tion, the fusion protein PML/RARα acts as a co-repressor recruiting multiple repressive epigenetic modifiers to down-regulate the expression of target genes and to induce a differentiation block. It has been generally assumed that the mechanism of action of pharmacological doses of ATRA includes binding to RARα, release of co-repressors, recruitment of tran-scriptional activators and relief of the differentiation block [5]. However, it should be noted that differentia-tive properties of ATRA were first described in HL-60 cell line [6] established from peripheral blood of a pa-tient suffering from acute myeloblastic leukemia, or the AML-M2 which actually carries no t(15;17) transloca-tion. Therefore, paradoxically, the cells on which dif-ferentiative properties of ATRA were first described would not fulfill the current clinical criteria for ATRA-based treatment. Obviously, the mechanism of ATRA-mediated differentiation of AML could not be solely ascribed to the effects on the PML/RARα protein [7].

Phosphatidylinositol 3-kinase (PI3K)/Akt/ mammal-ian target of rapamycin (mTOR) had been generally considered one of the principal pathways responsible for transmitting proliferative and anti-apoptotic signals in AML [8]. Our previous study on ATRA-treated HL-60 and NB4 cells, the latter containing the typical t(15;17), suggested that PI3K/Akt pathway had some role in differentiative responses of leukemia cells as increases in the level of both nuclear phosphatidyli-

nositol (3,4,5) trisphosphate (PIP3) and phosphatidyli-nositol (3) phosphate, as well as an increase in the ac-tivity of protein kinase B/Akt were observed, and siRNA-mediated down-modulation of Akt in HL-60 cells reduced the expression of differentiation markers in ATRA-treated cells [9]. In both the cell lines, phar-macological inhibitors of PI3K and Akt inhibited pro-liferation, but negatively affected differentiative capac-ity of the cells [10]. In contrast, use of rapamycin, which inhibits mTOR, a more distal component of the pathway, potentiated differentiation of AML cells along granulocytic pathway [10-12]. Physiologically, mTOR is negatively regulated by AMP-activated kinase (AMPK), an evolutionary conserved serine-threonine kinase that is activated whenever the energy level in the cell is low and the ratio of AMP to ATP is high. Once activated, AMPK stimulates ATP-gene- rating pathways (glycolysis and fatty acid oxidation), and inhibits ATP-consuming pathways (gluconeogene-sis and synthesis of fatty acids and cholesterol) through both direct phosphorylation of enzymes and alteration in gene expression [13]. Our recent study demonstrated that 5-amino-4-imidazolecarboxamide ribonucleoside (AICAr, acadesine), an AMPK-activator, enhances ATRA-mediated differentiation in HL-60 and NB4 cells. In monocytic U937 cells, a non-APL AML cell line, AICAr induces the expression of cell surface markers associated with mature monocytes and macro-phages but the mechanisms responsible for AICAr-mediated differentiative effects are still unknown [14].

Recent studies suggested that drugs that target me-tabolism may have some role in the treatment of cancer [reviewed in 15]. Many of these drugs have been stud-ied as modulators of AMPK pathway. In AML cells, several studies demonstrated involvement of PI3K/ Akt/mTOR and AMPK pathways in cell cycle progres-sion, proliferation and survival based on cytotoxicity assays. In this review, we will focus more on the possi-ble role that both AMPK activators and mTOR inhibi-tors may have in the differentiation of leukemia cells. The identification of their mechanisms of action may provide some theoretical basis for further improvement of differentiation therapy of AML.

2. MTOR AS A POTENTIAL THERAPEUTIC TARGET

2.1. Regulation and Functional Role of mTOR

The mechanistic target of rapamycin (mTOR, for-merly named mammalian target of rapamycin) is a ser-ine/threonine protein kinase that was identified as a target for rapamycin, a drug discovered more than 50

AMPK/mTOR in Acute Myeloid Leukemia Current Medicinal Chemistry, 2018, Vol. 25, No. 00 3

Fig. (1).

years ago on the Chilean Easter Island (Polynesian name Rapa Nui) [16]. Although it was originally dis-covered as an antifungal agent, it was soon attributed with immunosuppressive and antitumor activity, but the exact mechanism of action was unknown until the 1990s when genetic screens for rapamycin resistance identified two genes, TOR1 and TOR2 in Saccharomy-ces cerevisiae [17].

mTOR is a catalytic subunit of two functionally dis-tinct protein complexes, mTORC1 (mTOR complex 1) and mTORC2 (mTOR complex 2), which are mammal-ian analogues of yeast complexes TORC1 and TORC2. As shown in Fig. (1), rapamycin-sensitive mTORC1 contains protein Raptor and phosphorylates p70 S6 Kinase 1 (S6K1) and eIF4E Binding Protein (4EBP-1), which are involved in regulation of protein translation [18]. On the other hand, mTORC2 is considered to be rapamycin insensitive, at least upon acute exposure. Although mTORC2 phosphorylates several PKC iso-forms important for cytoskeletal remodeling and cell migration, the most important target is Akt, which is activated downstream of receptor tyrosine kinases.

mTORC1 is at the intersection of many signaling pathways, but one of the best described pathways is phosphatidylinositol 3-kinase class I/Akt/mTOR

(PI3KC1/ Akt/mTOR) downstream of insulin receptor. As shown in Fig. (1), activation of PI3K by growth fac-tors leads to generation of PIP3 which recruits inactive PKB/Akt to the plasma membrane. Once activated, Akt phosphorylates tuberous sclerosis complex 2 (TSC2) and activate mTORC1 through Ras homologue enriched in brain (Rheb). In addition, Akt directly activates mTORC1 through phosphorylation of proline-rich AKT substrate 40 (PRAS40). Activated mTOR allows for normal cell growth by stimulating protein, lipid and nu-cleotide synthesis in case of nutrient abundance and growth factor stimulation. mTOR is a master regulator of translational pathways controlling biosynthesis of proteins crucial for cell survival, like Bcl-2, Mcl-1 and Survivin. The functional role of mTOR in growth, sur-vival, aging, metabolism and cancer have recently been reviewed in depth by several authors [18-21].

The PI3KC1/Akt/mTOR signaling pathway is con-stitutively activated in many sporadic human cancers, but also plays a role in familiar cancer syndromes. In most cases, the activation is due to the amplifica-tion/mutation of catalytic or regulatory subunits of PI3K, receptor tyrosine kinases (RTK), Akt, TSC1/2, mTOR, or deletion/inactivation of tumor suppressors, including phosphatase and tensin homolog (PTEN) and

Dora Visnjic

Note

Figure title and legend are missing


liver kinase B1 (LKB1). Loss-of-function mutation of TSC1/2 is associated with multiorgan hamartomas and multiple benign tumors in tuberous sclerosis and lym-phangioleiomyomatosis; the loss of PTEN, the main negative regulator of PI3K/Akt that removes the 3’-phosphate from PIP3, is mutated in 70% of patients with Cowden syndrome; and LKB1 is mutated in Peutz-Jeghers syndrome characterized by intestinal polips [22]. Using publicly available tumor genome sequencing data Grabiner et al. [23] generated a cata-logue of 33 MTOR mutations in various cancer types that confer pathway hyperactivation and these mutated cells were highly sensitive to rapamycin.

The PI3KC1/Akt/mTOR signaling pathway has been found to be constitutively activated in 50-80% of human primary AML cells [24-28], mostly due to acti-vating mutations in tyrosine kinases upstream of PI3K/Akt axis. Although several studies have demon-strated beneficial effects of drugs targeting either PI3K and/or Akt in preclinical models of AML, results of clinical trials are modest or disappointing [reviewed in 29-31]. Two inhibitors selectively targeting Akt, MK-2206 and UCN-01, had promising effects in preclinical studies, but failed to exert their action on AML patients in clinical trials [32,33]. A recent phase I trial of bu-parlisib, an oral PI3K inhibitor, demonstrated accept-able tolerability and preliminary activity in a subset of patients with advanced leukemia [34].

The activity of mTOR is high in majority of AML samples, even in the absence of an elevated PI3K and Akt signaling. One of the possible Akt-independent mechanisms of mTOR-activation involves an upstream activation of the oncogenic Lyn kinase that has been shown to be augmented in AML cells [35]. Amplified levels of Rheb and Raptor, or down-regulated TSC2 may lead to increased activity of mTORC1. Other pos-sible mechanisms of conbstitutive mTOR activation involve the activity of ERK, WNT or HIF-1 [29-31].

The role of mTOR in promoting leukemia have been documented in several murine models. In a model of leukemogenesis evoked by PTEN deficiency, dele-tion of Raptor, an activator of mTORC1, significantly prolonged survival of mice [36]. In a mouse model of MLL-AF9-driven AML, conditional deletion of Raptor significantly suppressed leukemia progression, pre-vented leukemia initiation and prolonged animal sur-vival [37], and the similar results were obtained when S6K1, downstream target of mTORC1 [38], or Rheb1, an upstream activator of mTORC1 [39], were deleted. Taken together, these results suggested that mTORC1 promoted leukemogenesis and that pharmacological

inhibition of mTORC1 might be a promising strategy in the treatment of leukemia.

2.2. Rapamycin and Analogs

Rapamycin or sirolimus (Rapamune) is approved as an immunosuppressant drug for the prophylaxis of or-gan rejection in patients receiving renal transplants and for treating lymphangioleiomyomatosis. Besides oral formulation, it is also used for coating of drug-eluting coronary stents to prevent recurrence of stenosis. Limi-tations such as poor solubility and pharmacokinetics resulted in the development of its analogs, temsi-rolimus and everolimus that are approved by U.S. Food and Drug Administration for the treatment of advanced renal cell carcinoma. Everolimus has been also ap-proved for several other tumors and for the treatment of advanced-stage, hormone receptor-positive, HER2-negative breast cancer in combination with aromatase inhibitor [40].

Rapamycin binds FK506-binding protein (FKBP12) and their complex acts as an inhibitor of mTORC1, resulting in inhibition of protein translation. Immuno-suppressive properties of rapamycin were first attrib-uted to its antiproliferative activity and promotion of cell cycle, but experiments on T cells showed modest antiproliferative effects of rapamycin and pointed to the role of rapamycin in differentiation and anergy of T cells, as well as a complex set of roles in other immune cell populations. Recently, rapamycin was found to extend animal lifespan and reduce aging-dependent phenotypes, including neurodegenerative disorders. Still, rapamycin has been proven in human subjects to have serious adverse effects like nephrotoxicity, im-munosuppression, insulin resistance, hyperlipidemia and hypercholesterolemia [reviewed in 20, 41-43].

Early after the discovery of immunosuppressive properties, rapamycin was proven to exert antineoplas-tic activity. In the first reports, rapamycin was shown to reduce tumor growth in several transplantable tumor models [44] and to inhibit proliferation of hepatoma cell line through decreased activity of p70 S6K [45]. In hematologic malignancies, the use of rapamycin was first considered for prevention of transplantation-related lymphoma, due to its known effects on lympho-cyte proliferation [46]. Furthermore, antileukemic ac-tivity of rapamycin has been demonstrated against B-acute lymphoblastic leukemia (ALL) [47], childhood ALL [48], and BCR-ABL positive chronic myeloid leukemia (CML) in combination with imatinib [49]. Although preclinical studies have shown cytostatic ef-fects of mTOR inhibitors on primary AML cells [28,

Dora Visnjic

Highlight

Dora Visnjic

Strikeout

Dora Visnjic

Highlight

cell cycle arrest


50], results of the clinical trials using rapamycin de-rivatives as a single agent have shown very limited re-sponse in AML patients treated with sirolimus [28], deforolimus [51] or everolimus [52]. Even with the addition of cytotoxic chemotherapy, sirolimus did not enhance antileukemic effects of chemotherapy [53]. Temsirolimus [54] and everolimus [55] demonstrated acceptable toxicity, but modest clinical effects. Table 1. summarizes results of completed clinical trials using mTORC1 inhibitors for AML and gives a listing of trials currently underway with mTORC1 and PI3K/mTOR inhibitors.

There are several recent reviews regarding the use of mTOR inhibitors in various hematological malig-nancies, including AML, which provide possible ex-planations for limited potential of rapalogs in treating cancer [29,30,56]. Besides the lack of inhibition of all mTORC1 substrates or induction of autophagy, an im-portant role is ascribed to the lack of negative feedback exerted by mTORC1 on the proximal PI3K/Akt signal-ing pathway. As an increase in PI3K/Akt activity has been demonstrated in several rapalogs-treated cells, dual inhibitors of PI3K/mTOR (e.g. PI-103, BEZ235) and/or dual mTORC1/2 inhibitors (OSI-027, PP242) have been used to increase antiproliferative effects. However, although blocking two components of PI3K/Akt/mTOR is more effective in inhibiting prolif-eration, it may compromise differentiation that was also shown to depend on the increase in the activity of PI3K/Akt. In ATRA-treated cells, an increase in the level of PI3K [57], or phosphorylated Akt [9] has been detected in nuclei, and specific down-regulation of Akt [9] or p85a-subunit of PI3K [57] reduced CD11b ex-pression in ATRA-stimulated leukemia cells. In murine AML cells, constitutive activation of Akt promotes myeloid differentiation and these effects were mediated by FOXO [58].

Rapamycin was shown to induce differentiation of variety of cells [59-61]. In leukemia cell lines, an early study reported that rapamycin alone can induce differ-entiation, but these data were obtained at doses that were up to two orders of magnitude greater than the range of therapeutically achievable doses [62]. Several studies later on confirmed that rapamycin alone at doses of 20-50 nM had no effects on the expression of differentiation markers in vitro [10-12,63,64]. How-ever, rapamycin enhanced differentiation of AML cell lines and these effects were not common to all differen-tiation inducers. Rapamycin enhanced the expression of CD11b in response to ATRA [10-12] and vitamin D3 [65], and had no effects in cells treated with ATO [66]

or differentiated in the presence of PMA [10] or DMSO [12]. In typical APL cells, the enhancing effects of rapamycin on cell differentiation has been ascribed to rapamycin-mediated induction of autophagy, which degrades the fusion PML/RARα protein [67], but these effects cannot be responsible for enhancing effects in non-APL cell lines. In non-APL AML cells, cellular mechanisms leading to differentiation induction are poorly understood but seem to depend on the increase in the activity of proximal PI3K/Akt pathway, which is observed in response to ligands of the nuclear receptor family, like ATRA [9,57] and vitamin D3 [68, 69]. Therefore, although combined inhibition of PI3K and mTOR exerted synergistic antiproliferative effects, our previous study showed that combined inhibition dimin-ished differentiative properties of rapamycin in AML cells [10].

3. AMPK AS A POTENTIAL THERAPEUTIC TARGET

3.1. Regulation and Functional Role of AMPK Sig-naling

The kinase activity responsible for phosphorylation and inactivation of 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) and acetyl-CoA carboxylase (ACC) was named “AMP-activated kinase” to denote the new serine/threonine kinase, which is activated by an in-crease in 5’-AMP [70]. AMPK is a heterotrimer com-posed of a catalytic subunit α and two regulatory β and γ subunits; the α-subunit contains a conserved threonine residue (Thr 172) within the kinase domain, phosphorylation of which usually serves as a marker for its activation, and the γ-regulatory subunit provides binding sites for adenine nucleotides. During metabolic stress, AMP and ADP displace ATP from γ-regulatory subunit and induce conformational change that allows phosphorylation of Thr 172 by upstream kinases. Prin-cipal upstream kinases are: liver kinase B1 (LKB1), which is mutated in Peutz-Jeghers syndrome, cal-cium/calmodulin dependent kinase kinase (CAMKKb) and TGFβ-activated kinase (TAK1). Once activated, AMPK regulates metabolic pathways and affects the activity of various proteins involved in aging, cell cycle regulation, cell growth and apoptosis. Among many different targets, AMPK inhibits the activity of mTOR, which provides a link between a lack of ATP and a de-crease in cell growth and protein synthesis. For more in-depth discussion regarding the structure, isoforms, regulation and the functional role of AMPK in metabo-lism and disease, excellent reviews have been recently provided [71-76].


Table 1. Clinical trials of mTORC1, dual mTORC1/2 and PI3K/mTOR inhibitors in AML.

Drug Target Drugs in Combination Clinical Trial Status Results

Deforolimus mTORC1 -

NCT00086125 A Phase II Study of AP23573, an mTOR Inhibitor, in Patients With Relapsed or Refractory Hematologic Malignancies

Completed, Data pub-

lished

Total: 23 patients (AML) Deforolimus was well tolerated.

None of the 22 evaluable patients with AML had CR or PR. a)

Sirolimus mTORC1

MEC (mi-toxantrone,

etoposide, and cytarabine)

NCT00780104 A Prospective Single Institution Pilot

Study Evaluating the Pharmacokinetics of Sirolimus in Combination With MEC (Mitoxantrone + Etoposide + Cytarabine)

in Patients With High Risk Leukemias


lished

Total: 29 patients Sirolimus and MEC is an active

and feasible regimen, the synergy between MEC and sirolimus was

not confirmed.b)

Temsirolimus mTORC1 clofarabine

NCT00775593 An Open Label Phase II Trial of Clo-farabine and Temsirolimus in Older

Patients With Relapsed or Refractory Acute Myeloid Leukemia (AML)


lished

Total: 53 patients The combination of lower-dose

clofarabine and temsirolimus has activity and acceptable toxicity in older adults with AML refractory to or relapsing after initial inten-

sive chemotherapy. CR + CRi=21%, MS all patients = 4.0 mo, MS responders = 9.1 mo. c)

Everolimus (RAD001) mTORC1

daunorubicin, cytarabine

NCT01074086 Multicentric Study of GOELAMS Phase I Evaluation of RAD001 in Association With Aracytine and Daunorubicine in

AML Treatment in Patients Less Than 65 Years in Relapse More Than One Year

After First Complete Remission


lished

Total: 28 patients RAD001 at d1 and d7 of an in-duction chemotherapy regimen

for AML has acceptable toxicity and may improve treatment. CR=

19/28 d)

Sirolimus mTORC1 decitabine

NCT00861874 A Phase I Study of Decitabine in Combi-

nation With Escalating Doses of Ra-pamycin in Patients With Relapsed or Refractory Acute Myeloid Leukemia


lished

Total: 12 patents The combination of decitabine and rapamycin can be safely administered to patients with relapsed/refractory AML. e)

Everolimus (RAD001) mTORC1 azacitidine

ACTRN12610001031055 A Phase Ib/II Clinical Evaluation of the

Safety of Combining the mTOR inhibitor Everolimus with 5-Azacitidine in Acute

Myeloid Leukaemia (AML).


lished

Total: 40 patients Everolimus in combination with

azacitidine is tolerable, with promising clinical activity in advanced AML. OS= 8.5 mo,

ORR= 22.5% f)

Sirolimus mTORC1

MEC (mi-toxantrone,

etoposide, and cytarabine)

NCT00634244 A Phase II Randomized Trial of Car-boplatin and Topotecan; Flavopiridol,

Mitoxantrone and Cytosine Arabinoside; and Sirolimus, Mitoxantrone, Etoposide and Cytosine Arabinoside for the Treat-ment of Adults With Primary Refractory or Initial Relapse of Acute Myelogenous

Leukemia (AML)

Completed, Results un-published

Total: 20 patients CR+CRi= 15% g)

Temsirolimus mTORC1 Standard chemotherapy

NCT01611116 A Double-blind, Placebo-controlled,

Randomized, Multicenter Phase II Trial to Assess the Efficacy of Temsirolimus Added to Standard Primary Therapy in Elderly Patients With Newly Diagnosed

AML

Completed, No results

posted -

(Table 1) contd….


Drug Target Drugs in Combination Clinical Trial Status Results

Sirolimus mTORC1 decitabine, ribavirine

NCT02109744 A Phase I/II Study of Decitabine in

Combination With Sequential Rapamy-cin or Ribavirin in High Risk AML Pa-

tients

Active -


MEC (mi-toxantrone,

etoposide, and cytarabine), idarubicin

NCT01154439 A Phase I Study Investigating the Com-

bination of RAD001 With Standard Induction and Consolidation Therapy in Older Patients With Newly Diagnosed

Acute Myeloid Leukemia (AML)

Active -

Sirolimus mTORC1 azacitidine

NCT01869114 A Phase II Study of Azacitidine and

Sirolimus for the Treatment of High Risk Myelodysplastic Syndrome or Acute

Myeloid Leukemia Refractory to or Not Eligible for Intensive Chemotherapy

Active -

Sirolimus mTORC1 cytarabine, idarubicin

NCT01822015 A Pilot, Pharmacodynamic Correlate

Trial of Sirolimus in Combination With Chemotherapy (Idarubicin, Cytarabine) for the Treatment of Newly Diagnosed

Acute Myelogenous Leukemia

Active -


PKC412 (FLT3 inhibi-

tor)

NCT00819546 A Phase I Trial of Escalating Dose of

RAD001 in Combination With PKC412 in Patients With Relapsed, Refractory or

Poor Prognosis AML or MDS

Active -

BEZ235 PI3K/mTOR -

NCT01756118 A Phase I, Dose-finding Study of the

Oral, Dual Phosphatidylinositol 3(PI3)-Kinase / Mammilian Target of Rapamy-cin (mTOR) Inhibitor BEZ235 in Adult Patients With Relapsed or Refractory

Acute Leukemia

Active -

PF-05212384 (PKI-587) PI3K/mTOR -

NCT02438761 Phase II Evaluating the Efficacy of the Dual Inhibition of Phosphoinositide 3

Kinase (PI3K)/Akt /Mammalian Target Of Rapamycine (mTOR) Signaling

Pathway by PF-05212384 (PKI-587) for Patients With Myeloid Neoplasm Secon-

dary to Chemo-radiotherapy (t-AML/MDS) or de Novo Relapsed or

Refractory AML

Active -

CR – complete remission, CRi - complete remission with incomplete hematologic recovery, PR – partial remission, MS – median survival, OS – overall sur-vival, ORR – overall response rate a) Rizzieri DA, et al. Clin Cancer Res. 2008;14:2756-62. b) Perl AE, et al. Clin Cancer Res. 2009;15:6732-9. c) Amadori S, et al. Br J Haematol. 2012;156:205-12. d) Park S, et al. Leukemia. 2013;27:1479-86. e) Liesveld JL, et al. Leuk Res. 2013;37:1622-7. f) Tan P, et al.Oncotarget. 2016;8:52269-52280. g) https://clinicaltrials.gov/ct2/show/NCT00634244

Most of the functions of AMPK have been de-scribed on the basis of treatment of cell lines and organ cultures with pharmacological activators of AMPK,

such as antidiabetic biguanides, metformin and phen-formin, or 5-amino-4-imidazolecarboxamide ribonu-cleoside (AICAr). Although numerous drugs and xeno-


biotics, including barbiturates, 2-deoxyglucose and resveratrol, have been used as AMPK agonists, based on their ability to phosphorylate AMPK on Thr 172, their mechanisms of action are mostly indirect and only partially elucidated [77]. In addition, an increasing number of studies suggest that the majority of the ef-fects of most commonly used AMPK agonists, met-formin and AICAr, on cell cycle, metabolism and vi-ability are actually AMPK-independent [78, 79].

More reliable way to assess the functional role of AMPK in vivo could be provided by mice knockouts. Data provided by transgenic mice models demonstrated that both AMPK [80] and LKB1 [81] are important for glucose uptake into muscle cells, but have lesser role in physiological regulation of glucose metabolism in car-diac muscle cells [82-83]. However, AMPK allows for the increase in glucose uptake and lactate production in post-ishemic cardiomyocyte [84], while LKB1 deletion does not change the rate of glucose and palmitate oxi-dation after reperfusion of the ishemic heart [83]. Al-though liver-specific LKB1 knockouts show an in-crease in blood glucose level [85], tissue-specific AMPK knockouts are normoglycemic [86]. The differ-ence in phenotype of AMPK and LKB1 knockouts could be explained by the presence of different AMPK isoforms in different tissues and/or the fact that LKB1 has at least 12 different downstream targets, not includ-ing AMPK [as reviewed in 87].

The possible role of AMPK in cancer was suggested as soon as LKB1, a known tumor suppressor, was iden-tified as an upstream kinase [88-90]. In addition, AMPK activates another tumor suppressor complex, TSC2 [91], and thus inhibits both cell growth, through mTOR inhibition, and cell proliferation, through phos-phorylation and activation of p53, which is often mu-tated or its activity disabled in many tumors [92]. The more direct evidence for the role of AMPK as a tumor suppressor was provided by a study on transgenic mice overexpressing c-Myc in B cells in which genetic abla-tion of the α1 catalytic subunit of AMPK accelerates Myc-induced lymphomagenesis [93]. In addition, both direct and indirect AMPK-activators, A769662 and biguanides, delay the onset of tumorigenesis in mice that are prone to the development of tumors due to het-erozygous loss of PTEN combined with reduced ex-pression of LKB1 [94]. However, although phenformin protected against initial tumor development [94], once tumor has been already established, phenformin was more efficient in apoptosis induction in genetically en-gineered mouse model of lung cancer baring LKB1 mutations [95]. In a murine model of AML, AMPK

was shown to confer metabolic stress resistance to leu-kemia initiating cells and to promote leukemogenesis [96]. Furthermore, in a mouse model of T cell acute lymphoblastic leukemia (T-ALL), AMPK deficiency led to leukemia cell death and increased animal sur-vival [97]. Importantly, although these results may suggest that AMPK activation may be useful only for cancer prevention and not for cancer treatment, phen-formin treatment reduced the number and percentage of T-ALL cells in the treated mice and resulted in a sig-nificant increase in overall animal survival further con-firming the hypothesis that majority of antineoplastic effects of AMPK agonists may be AMPK-independent and related to profound metabolic changes.

In AML patients, the functional LKB1/AMPK axis was first demonstrated in primary samples pharmaco-logically treated with metformin in which AMPK acti-vation fully inhibited mTORC1 and reduced oncogenic protein synthesis [98]. Tumor suppressor role of AMPK was further confirmed in a study using targeted knockdown screen of AML cell lines [99]. Of note, mutations of AMPK [100], or its predominant upstream regulator LKB1 [101], are very rare in AML patients and our understanding of the role of AMPK in AML is mostly based on studies using pharmacological modu-lators. In a recent study using GSK621, a more specific AMPK agonist, cytotoxic effect of AMPK activation was not dependent on mTORC1 inhibition but indeed required mTORC1 activation that was unique to AML cells and involved the eIF2α/ATF4 signaling pathway [102]. The activity of mTOR in AML cells may depend on several other factors, including microenvironment or hypoxia as HIF-1 controls myeloid hematopoietic cell function by maintaining mTOR phosphorylation through AMPK [103]. Irrespective of mTOR involve-ment, AMPK has many other downstream targets im-portant in metabolism such as fatty acid metabolism that has been recently described to be activated in acute monocytic leukemia cells co-cultured with bone mar-row adipocytes [104].

The listing of clinical trials currently underway us-ing AMPK modulators for the treatment of hema-tologic malignancies is presented in Table 2.

3.2. Metformin

Metfomin or 1,1-dimethylbiguanide is the first-line medication in the treatment of type 2 diabetes mellitus (T2DM). By chemical structure, biguanides are deriva-tives of guanidines, which gained popularity in 1950s due to their good oral bioavailability and the ability to reduce blood glucose levels without inducing hypoglycemia.


Table 2. AMPK modulators in clinical trials for hematologic malignancies.

Drug Disease Clinical Trial Results

metformin Acute lymphoblastic leukemia

NCT03118128 Effect of the Addition of Metformin Hydrochloride on the

Prognosis of Patients With B-cell Precursor (Ph+ Negative) Acute Lymphoblastic Leukemia With High Expression of

ABCB1 Gene

Completed, no results posted

metformin Relapsed chronic lymphocytic leukemia

NCT01750567 A Phase II Pilot Study of Metformin Therapy in Patients With Relapsed Chronic Lymphocytic Leukemia and Untreated CLL

Patients With Genomic Deletion 11q

Active study

metformin Childhood acute lymphoblastic leukemia

NCT01324180 A Phase I Window, Dose Escalating and Safety Trial of Met-formin in Combination With Induction Chemotherapy in Re-

lapsed Refractory Acute Lymphoblastic Leukemia: Metformin With Induction Chemotherapy of Vincristine, Dexamethasone,

Doxorubicin, and PEG-asparaginase (VPLD)


metformin Recurrent/refractory plasma

cell myeloma and chronic lym-phocytic leukemia

NCT02948283 A Pilot Feasibility Study of Metformin/Ritonavir Combination Treatment in Patients With Relapsed/Refractory Multiple Mye-

loma or Chronic Lymphocytic Leukemia

Active study

metformin Acute myeloid leukemia NCT01849276

A Phase I Study of Metformin and Cytarabine for the Treat-ment of Relapsed/Refractory Acute Myeloid Leukemia

Active study

metformin Lymphoma NCT00659568

A Phase I Study of Temsirolimus in Combination With Met-formin in Advanced Solid Tumours


metformin Diffuse large B cell lymphoma

NCT03200015 Effect of Metformin in Combination With R-CHOP for the First Line Treatment of Patients With Diffuse Large B-cell

Lymphoma

Active study

metformin Lymphoma NCT02145559

A Pharmacodynamic Study of Sirolimus and Metformin in Patients With Advanced Solid Tumors

Active study

AICAR Chronic lymphocytic leukemia

NCT00559624 A Phase I/II Open Label Dose Escalation Study to Investigate

the Safety and Tolerability of Acadesine in Patients With B-cell Chronic Lymphocytic Leukemia

Completed, MTD (single dose)= 210 mg/kg; accept-able safety profile a)

Sodium salicylate

Leukemia, Myelodysplastic Syndromes, Myelodysplas-tic/Myeloproliferative Neo-

plasms

NCT00004245 A Phase I Study of Salicylate for Adult Patients With Advanced

Myelodysplastic Disorders or Acute Myelogenous Leukemia

Completed, acceptable safety

profile b)

MTD – maximum tolerated dose a) Van Den Neste E, et al. Cancer Chemother Pharmacol. 2013; 71: 581–591. b) Klimek VM, et al. Leukemia Res 2012;36:570-574. However, the more potent biguanides phenformin and buformin have not been in clinical use since 1970s because of their lower safety profile mostly due to the increased risk of lactic acidosis. Metformin demonstrated less toxicity and therefore has become the most commonly prescribed oral antidiabetic drug worldwide [105, 106].

The anti-diabetic properties of metformin are com-monly explained by the reduction of hepatic glu-

coneogenesis, the increase in cellular glucose uptake and increased insulin sensitivity [reviewed in 107,108]. These effects of metformin were first associated with the activation of AMPK [109], but later studies in transgenic mice demonstrated that both metformin-mediated decrease of gluconeogenesis [86], and in-crease in glucose uptake were mediated by an AMPK-independent mechanism [110].


In 2005, Evans et al. reported the results of the case-control study showing that the use of metformin re-duced the risk of cancer in a large cohort of patients with T2DM [111]. The finding was further confirmed by several studies and meta-analyses [reviewed in 112], even though the beneficial effects of metformin on cancer incidence appear to be much smaller than first reported, especially in randomized controlled trials and after adjustment for time-related biases. First reports of prospective randomized control trials with a clinical endpoint in which metformin was applied for an on-cological indication demonstrated that the addition of metformin to a standard systemic therapy did not im-prove outcome in patients with advanced pancreatic cancer [113,114]. At the moment, there are more than 120 ongoing clinical trials, 8 out of them for hema-tologic malignancies, which aim at repurposing met-formin for anti-cancer use [115].

The anticancer effects of metformin were first asso-ciated with AMPK since the activation of AMPK, as measured by phosphorylation of Thr 172, have been observed in many models in vitro [116] and in vivo [94]. Metformin was first proposed to activate AMPK indirectly, through inhibition of complex I of mito-chondrial respiratory chain and subsequent decrease in energy production and increase in AMP/ATP ratio [117]. However, further studies showed that metformin can activate AMPK in two different cell lines without any detectable change in cellular ADP-to-ATP ratio [118], and that metformin-mediated activation may employ a more complex mechanism involving reactive nitrogen species, c-Src, phosphatidylinositol 3-kinase [119] and/or protein kinase C-ζ-mediated phosphoryla-tion of LKB1 [120]. To make the story even more complicated, numerous studies in various cancer cell lines [78, 121,122], and mouse models of tumorigene-sis [95] demonstrated that antitumoror effects of bigua-nides are actually AMPK-independent.

Irrespective of AMPK involvement in metformin action, the fact remains that metformin exerts inhibi-tory effects on cancer cells growth in vitro, but does not improve outcome in patients with pancreatic cancer when added as an adjuvant to standard chemotherapy in randomized clinical trials [113, 114]. The simplest explanation for the difference may be that the doses of metformin used in vitro were approximately 1000 times higher than the peak plasma concentrations after oral intake. However, the concentration in plasma was the same as the one achieved in T2DM patients in which epidemiological studies reported beneficial ef-fects of metformin in cancer prevention initially raising

the possibility that the effectiveness of metformin in diabetic patients is maybe due to indirect effects on plasma levels of insulin and insulin-like growth factor 1, which are all known to act in a pro-oncogenic man-ner [123]. This mechanism is supported by a recent meta-analysis that showed a decrease in cancer inci-dence in T2DM patients who were taking metformin or thiazolidinediones but an increase in patients who were taking insulin or insulin secretagogues [124]. However, according to data obtained with high concentrations of metformin in vitro, there might be a rationale for the use of phenformin as more potent biguanide in the treatment of cancer since in these settings potential beneficial effects of the drug on cancer might outweigh its toxic side effects [108].

In the context of hematologic malignancies, met-formin was reported to activate AMPK and inhibit growth of AML cell lines and primary AML cells while sparing normal hematopoiesis ex vivo [98], in-hibit BCR-ABL-expressing cell lines and primary CML cells [125], interfere with the growth and survival of murine PTEN-deficient T cell lymphomas and hu-man T-ALL/T-LL cancer cells [126], and enhance the anti-myeloma effect of bortezomib [127], but the func-tional role of AMPK in metformin effects has not been tested. In lymphoma cell lines, AMPK siRNA experi-ments proved that metformin-mediated inhibition is AMPK-dependent [128], shRNA-mediated down-regulation in T-ALL cell line prevented metformin-induced apoptosis [129] and knockdown of AMPKα1 desensitizes metformin-mediated enhancements of vin-cristine-induced apoptosis in leukemia cells [130]. However, further studies in AML cell lines again showed that metformin mediated effects on apoptosis and proliferation are preserved even in cells with siRNA-downregulated AMPK suggesting AMPK-independent effects [131].

In a recent retrospective study aimed to evaluate whether metformin related cancer benefits reported in solid tumors are also present in AML patients, baseline metformin use provided no significant benefit in AML overall ad disease free survival. Lack of metformin benefit in AML could be ascribed to advanced age but also to the practice of metformin substitution with insu-lin which further confirmed the hypothesis that at least part of the beneficial effects of metformin in cancer prevention is indirect and due to a decrease in plasma insulin level [132].

Apart from having pro-apoptotic effects, metformin has been reported to enhance differentiation in APL cell line NB4 through activation of ERK signaling

Dora Visnjic

Strikeout

Dora Visnjic

Strikeout

Dora Visnjic

Highlight

Dora Visnjic

Highlight

Dora Visnjic

Insert Text

and


pathway [133]. In this study, enhancing effects of met-formin were found to be restricted to APL cells, and only pro-apoptotic effects of metformin were observed in several non-M3 AML cell lines. The effects of met-formin may depend on the basal activity of mTOR, as a downstream target of AMPK [14, 78], or the activity of ERK, which is known to regulate AMPK [134] and increase during differentiation [14, 135]. In monocytic U937 cells with a high basal level of mTOR, no in-crease in the level of differentiation was observed in response to metformin [14], and inhibitory effects of metformin were observed in PMA-mediated differen-tiation of acute monocytic leukemia cell line THP-1 [136]. There is increasing evidence that differential effects of metformin on the differentiation of hema-tologic and immunologic cells with its subsequent im-munomodulatory effects could be another mechanism of the antineoplastic activity of metformin [137,138]. In mouse models of normal hematopoiesis, the addition of metformin to the culture of Lin- Sca-1+ c-Kit+ bone marrow cells decreases differentiation and helps main-tain highly repopulating hematopoietic stem cells in culture [139]. In a preclinical murine model of Fanconi anemia, metformin increased the size of the hema-topoietic stem cell compartment, enhanced quiescence in hematopoietic stem and progenitor cells and delayed the tumor formation [140].

3.3. AICAR

The abbreviation AICAR has been widely used for 5-amino-4-imidazolecarboxamide (AICA) ribonucleo-side or acadesine, although it should be reserved for AICA-ribonucleotide or ribotide, which is the phos-phorylated form of AICA-ribonucleoside or riboside (Fig. 2). Therefore, AICA-riboside should be properly abbreviated as AICAr to denote an exogenous sub-stance which, after entering cells, becomes phosphory-lated by adenosine kinase into AICA-ribotide or AI-CAR [141]. From yeast to man, AICAR (also termed ZMP) is a normal cellular metabolic intermediate in de novo purine synthesis [142]. In humans, AICAR can be found to accumulate in various purine synthesis disor-ders, including Lesch-Nyhan disease [143].

As a cell-permeable nucleoside that shares some structural similarities with adenosine, AICAr was first developed to block adenosine reuptake in the ischemic heart [144]. In 1995, AICA riboside was reported to activate AMPK kinase [145, 146] and since then has been used in numerous studies related to metabolism, insulin signaling pathways and diabetes. Another surge of interest in AICAr was raised when it was shown to act as an “exercise in a pill” by increasing endurance in

sedentary mice [147] and to prevent heat-induced sud-den death in mice carrying mutations of type I ryano-dine receptors [148]. Finally, the most recent interest in another possible use of AICAr was initiated by data demonstrating anti-tumor effects of metformin and various AMPK-agonists in cancer [111, 149].

AICAr inhibits fatty acid synthesis [145], increases fatty acid oxidation [150] and induces hypoglycemic effects in vivo [151-153], which are probably due to an increase of glucose uptake into muscle cells through GLUT4 translocation [154] and an AICA-riboside me-diated inhibition of gluconeogenesis in the liver [151]. Some of these AICAR-mediated metabolic effects are AMPK-independent [155,156], but may be of rele-vance for the potential treatment of type 2 diabetes since the inhibition of fat synthesis and an increase in lipolysis may reduce fat storage and decrease periph-eral resistance to insulin action [74]. However, poor oral bioavailability of AICAr renders it quite unsuitable for the treatment of metabolic disorders like diabetes [157].

On the other hand, clinical trials involving more than 4000 patients with coronary artery bypass graft (CABG) surgery proved that AICAr (or acadesine) was safe and well-tolerated when used as an intravenous agent for the prevention of ischemia-reperfusion injury associated with CABG [158-160]. In the treatment of cardiovascular disorders, AICAr is classified as an adenosine regulating agents since the proposed mecha-nism of AICAr-effects includes the inhibition of adenosine deaminase and an increase in adenosine con-centrations during ischemic conditions [161]. In the list of prohibited substances of The World Anti-Doping Agency (WADA) from year 2011 AICAR was classi-fied as an AMPK agonist, but then moved to the class of “Hormone and metabolic modulators” in the list 2012 [162]. Endurance athletes could obviously benefit from the use of the substance, but it is difficult to esti-mate how widespread is the usage of AICAr as a dop-ing agent since the detection of the abuse of AICAr, as an endogenous substance, in sports is a complex prob-lem [163].

The possible clinical efficacy of AICAr as an anti-cancer agent has been recently tested in hematological malignancies. In year 2003, AICAr was first reported to induce apoptosis of B cell chronic lymphocytic leu-kemia (CLL) cells in vitro at doses that are well toler-ated when achieved in plasma after intravenous injec-tion [164]. Ten years later, results of the first clinical study testing the effects of acadesine in CLL demon-strated that AICAr had an acceptable safety profile and

Dora Visnjic

Strikeout


Fig. (2).

antileukemic activity in patients with poor prognosis [165]. In other hematological malignancies, cytotoxic effects of AICAr in vitro have been demonstrated on B cells from mantle cells lymphoma and splenic marginal zone lymphoma [166], childhood ALL cells [167] and CML [125].

The cellular mechanisms of beneficial effects of AICAr are still partially understood. As previously mentioned, the principal mechanism of action was long assumed to be the activation of AMPK, but more and more studies show that both AICAr and metformin dis-play AMPK-independent effects on cell proliferation, metabolism and differentiation (14, 78, 79, 168]. AI-CAr-mediated apoptosis in CLL cells is not mimicked by phenformin or A-769662, a direct AMPK agonist, and AICAr also potently induce apoptosis in B lym-phocytes from mice lacking α subunit of AMPK [169]. In CML cells, AMPK knockdown by shRNA failed to prevent the effect of acadesine, and acadesine exerted a potent anti-leukemic effect through protein kinase C-dependent autophagyic cell death [168]. In mantle cell lymphoma, acadesine effects may be potentiated in combination with the anti-CD20 monoclonal antibody rituximab [170] or Bcl-2 inhibitors [171].

The possible role of AICAr in differentiation ther-apy has been less investigated, although differentiative properties of AICAr has been described in several cell systems [172, 173], including AICAR-mediated differ-entiation of mouse embryonic stem cells along erythroid lineage [174]. Our recent study demonstrated

that AICAr induced apoptosis, reduced proliferation and enhanced ATRA-mediated differentiation of HL-60 and NB4 cells. In monocytic U937 cells, a non-APL AML cell line, AICAR alone induced the expression of cell surface markers associated with mature monocytes and macrophages. Although we detected time and dose-dependent increase in the level of Thr 172-phosphorylated AMPK, a significant decrease in AMPK expression that was achieved by using com-mercially available siRNA sequences in U937 cells had no significant effects on the AICAr-mediated effects on the number of viable cells or the expression of differen-tiation markers [14]. Of note, no differentiative effects of metformin were observed although both agents ex-erted similar growth-arresting properties in all cell lines and reduced the phosphorylation of p70 S6K as a downstream target to the similar level.

To conclude, the results obtained with AICAr in various preclinical in vitro models, as well as the re-sults of the first clinical studies testing AICAr for CML, suggest that the strategy using AICAr may im-prove therapy of hematological malignancies, including differentiation therapy for AML.

CONCLUSION

Numerous studies support the hypothesis that the inhibition of mTOR and activation of AMPK may have beneficial effects in promoting differentiation and blocking proliferation of acute myeloid leukemia cells. An advantage of possible therapeutic approach based

Dora Visnjic

Strikeout

Dora Visnjic

Note

Figure title and legend are missing.


on the modulation of AMPK/mTOR signaling pathway relies on the fact that majority of drugs that modulate the pathway are already in use for unrelated purposes; rapalogs have been used as immunosuppressive drugs for decades, biguanides are the most widely prescribed oral antidiabetics, and AICAR is used as an exercise mimetic. However, results of clinical trials using rapa-logs in leukemia have led to disappointing results, and no significant improvement was observed when dual mTOR inhibitors or inhibitors that block both PI3K and mTOR component of the pathway were used. Many authors still consider mTOR a promising target in AML mostly due to the fact that the activity of mTOR is high even in those AML patients with no genetic ab-errations of PI3K/Akt. Several combinatorial ap-proaches have been proposed in order to overcome re-sistance mechanisms, including agents targeting RAS/RAF/MEK/ERK axis, histone deacetylase path-ways, the Bcl-2 apoptotic pathways and many others [29, 30]. Recent data showing cytotoxicity of a specific AMPK activator in AML indicated a potential for AMPK-activating agents in the treatment of mTORC1-overactivated leukemia [102].

In this review, we wanted to stress that some com-ponents of this signaling pathways may also have a role in differentiation of leukemia cells. Although ap-proaches aimed to block upstream and/or parallel sig-naling pathways may help to increase cytotoxicity, blocking PI3K/Akt [9, 10] or ERK pathway [14, 135] certainly decreases differentiation of the cells. The concept of differentiation therapy first emerged from the success of ATRA therapy in APL, but is recently revisited again as there are more and more novel strategies and approaches aimed to increase the ability of ATRA to induce myeloid differentiation and apopto-sis in non-APL AML [175] or to transform other dif-ferentiating drugs into more efficient therapies [176]. Rapamycin-mediated inhibition of mTOR enhances differentiation of leukemia cells in response to various agents, including ATRA [10, 11, 177]. In addition, ac-tivation of AMPK with AICAR increases the expres-sion of differentiation markers in parallel with a de-crease in mTOR activity [14]. Therefore, modulation of AMPK and mTOR might have some role in new strategies to induce differentiation of leukemia cells.

Downstream targets of AMPK/mTOR in differen-tiation response are only partially elucidated. Auto-phagy is one of the mechanisms that was reported to be important for rapamycin-mediated enhancement of dasatinib-induced differentiation of leukamia cell lines [177] and ATRA-mediated differentiation of APL cells

[67]. Our recent study confirmed that autophagy in-creased in parallel with differentiation in response to many agents, including AICAR, but differentiation was not abolished by down-regulation of key components of classical or canonical autophagy pathway [178]. An-other possibility is that both rapamycin and AICAR, as pharmacological modulators of AMPK/mTOR, have profound effects on cell metabolism, and there are more and more data pointing to the role of metabolic changes during both proliferation and differentiation of leukemia cells. A recent study that examined metabolic changes in leukemia cell lines (including HL-60 and U937) in response to metformin revealed that AMPK-independent proapoptotic effects of metformin depend on the ability to produce Pasteur effect, i.e. switch to glucose consumption, glycolysis and lactate production [131]. It is possible that the induction of apoptosis in response to metformin might be directly due to its ac-tion on mitochondria because several agents that inhibit mitochondrial electron transport have been recently shown to act synergistically with Bcl-2 inhibitor and inductor of intrinsic apoptosis pathway in AML and ALL cell lines and primary cells [179]. The role of mi-tochondrial metabolism and glycolysis in leukemia cell differentiation in response to AICAr and other differen-tiation agents remains to be determined [180], but sev-eral studies pointed to the role of fatty acid oxidation [181], mitochondrial translation and oxidative phos-phorylation [182], glycolysis [183] or glutaminolysis [184] in AML proliferation and survival. Efforts to identify new therapeutic targets have recently pointed to the possible roles of either dihydroorotate dehydro-genase [185] or one-carbon folate pathway, specifically methylenetetrahydrofolate dehydrogenase-cyclohydro- lase 2 (MTHFD2) [186] in differentiation blockade of AML cells. The most recent clinically successful proof-of-concept that drugs targeting an “oncometabolite” may be useful in differentiation therapy of leukemia was provided by studies testing the effects of ena-sidenib in AML patients having mutations of isocitrate dehydrogenase 1 and 2 (IDH1 and IDH2) [187, 188].

To conclude, both inhibition of mTOR by rapamy-cin and activation of AMPK by AICAR have beneficial effects on leukemia cell differentiation. Better under-standing of their mechanism of action could help to identify new therapeutic targets to overcome myeloid differentiation blockade. As differentiation therapy of APL with ATRA still represents the most successful pharmacological therapy of AML, an important task would be to identify similar differentiation therapy strategies for the remaining 90% of AML patients.


CONSENT FOR PUBLICATION

Not applicable.

CONFLICT OF INTEREST

The authors declare no conflict of interest, financial or otherwise.

ACKNOWLEDGEMENTS

This work has been supported by Croatian Science Foundation under the project IP-2016-06-4581 (to D.V.) and co-financed by the European Union through the European Regional Development Fund, Operational Programme Competitiveness and Cohesion, grant agreeemnet No. KK.011.1.01.0007, CoRE-Neuro.

REFERENCES [1] Khwaja, A.; Bjorkholm, M.; Gale, R.E.; Levine, R.L.; Jor-

dan, C.T.; Ehninger, G.; Bloomfield, C.D.; Estey, E.; Burnett, A.; Cornelissen, J.J.; Scheinberg, D.A.; Bouscary, D.; Linch, D.C. Acute myeloid leukemia. Nat. Rev. Dis. Primers., 2016, 2, 16010.

[2] De Kouchkovsky, I.; Abdul-Hay, M. Acute myeloid leuke-mia: a comprehensive review and 2016 update. Blood Can-cer J., 2016, 6, e441.

[3] Coombs, C.C.; Tavakkoli, M.; Tallman, M.S. Acute pro-myelocytic leukemia: where did we start, where are we now, and the future. Blood Cancer J., 2015, 5, e304.

[4] Lo-Coco, F.; Avvisati, G.; Vignetti, M.; et al. for Gruppo Italiano Malattie Ematologiche dell'Adulto; the German–Austrian Acute Myeloid Leukemia Study Group; and Study Alliance Leukemia. Retinoic Acid and Arsenic Trioxide for Acute Promyelocytic Leukemia. N. Engl. J. Med., 2013, 369, 111-121.

[5] van Gils, N.; Verhagen, H.J.M.P.; Smit, L. Reprogramming acute myeloid leukemia into sensitivity for retinoic-acid-driven differentiation. Exp. Hematol., 2017, doi: 10.1016/j.exphem.2017.04.007.

[6] Breitman, T.R.; Selonick, S.E.; Collins, S.J. Induction of differentiation of the human promyelocytic leukemia cell line (HL-60) by retinoic acid. Proc. Natl. Acad. Sci. U S A., 1980, 77, 2936-2940.

[7] Petrie, K.; Zelent, A.; Waxman, S.; Differentiation therapy of acute myeloid leukemia: past, present and future. Curr. Opin. Hematol., 2009, 16, 84-91.

[8] Martelli, A.M.; Evangelisti, C.; Follo, M.Y.; Ramazzotti, G.; Fini, M.; Giardino, R.; Manzoli, L.; McCubrey, J.A.; Cocco, L. Targeting the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin signaling net-work in cancer stem cells. Curr. Med. Chem., 2011, 18, 2715-2726.

[9] Matkovic, K.; Brugnoli, F.; Bertagnolo, V.; Banfic, H.; Visnjic, D. The role of the nuclear Akt activation and Akt inhibitors in all-trans-retinoic acid-differentiated HL-60 cells. Leukemia, 2006, 20, 941-951.

[10] Mise, J.; Dembitz, V.; Banfic, H.; Visnjic, D. Combined inhibition of PI3K and mTOR exerts synergistic antiprolif-erative effect; but diminishes differentiative properties of rapamycin in acute myeloid leukemia cells. Pathol. Oncol. Res., 2011, 17, 645-656.

[11] Nishioka, C.; Ikezoe, T.; Yang, J.; Gery, S.; Koeffler, H.P.; Yokoyama, A. Inhibition of mammalian target of rapamy-cin signaling potentiates the effects of all-trans retinoic acid

to induce growth arrest and differentiation of human acute myelogenous leukemia cells. Int. J. Cancer, 2009, 125, 1710-1720.

[12] Lalic, H.; Lukinovic-Skudar, V.; Banfic, H.; Visnjic, D. Rapamycin enhances dimethyl sulfoxide-mediated growth arrest in human myelogenous leukemia cells. Leuk. Lym-phoma 2012, 53, 2253-2261.

[13] Hardie, D.G. AMP-activated protein kinase: an energy sen-sor that regulates all aspects of cell function. Genes Dev., 2011, 25, 1895-1908.

[14] Lalic, H.; Dembitz, V.; Lukinovic-Skudar, V.; Banfic, H.; Visnjic, D. 5-Aminoimidazole-4-carboxamide ribonucleo-side induces differentiation of acute myeloid leukemia cells. Leuk. Lymphoma, 2014, 55, 2375-2383.

[15] Hauge, M.; Bruserud, Ø.; Hatfield, K.J. Targeting of cell metabolism in human acute myeloid leukemia--more than targeting of isocitrate dehydrogenase mutations and PI3K/AKT/mTOR signaling? Eur. J. Haematol., 2016, 96, 211-221.

[16] Seghal, S.N. Sirolimus: Its discovery; biological properties; and mechanism of action. Transplantation Proc., 2003, 35, 7S-14S.

[17] Heitman, J.; Movva, N.R.; Hall, M.N. Targets for cell cycle arrest by the immunosuppressant rapamycin in yeast. Sci-ence, 1991, 253, 905-909.

[18] Saxton, R.A., Sabatini, D.M. mTOR signaling in growth; metabolism; and disease. Cell, 2017, 168, 960-976.

[19] Yu, J.S.L.; Cui, W. Proliferation, survival and metabolism: the role of PI3K/AKT/mTOR signaling in pluripotency and cell fate determination. Development, 2016, 143, 3050-3060.

[20] Kennedy, B.K.; Lamming, D.W. The mechanistic target of rapamycin: the grand conducTOR of metabolism and aging. Cell Metabolism, 2016, 23, 990-1003.

[21] Laplante, M.; Sabatini, D.M. mTOR signaling in growth control and disease. Cell, 2012, 149, 274-292.

[22] Chiarini, F.; Evangelisti, C.; McCubrey, J.A.; Martelli, A.M. Current treatment strategies for inhibiting mTOR in cancer. Trends Pharmacol. Sciences, 2015, 36, 124-135.

[23] Grabiner, B.C.; Nardi, V.; Birsoy, K.; Possemato, R.; Shen, K; Sinha, S.; Jordan, A.; Beck, A.H.; Sabatini, D.M. A di-verse array of cancer-associated MTOR mutations are hy-peractivating and can predict rapamycin sensitivity. Cancer Discov., 2014, 4, 554-563.

[24] Xu; Q.; Simpson; S.E.; Scialla; T.J.; Bagg; A.; Carroll; M. Survival of acute myeloid leukemia cells requires PI3 kinase activation. Blood, 2003, 102, 972-980.

[25] Min, Y.H.; Eom, J.I.; Cheong, J.W.; Maeng, H.O.; Kim, J.Y.; Jeung, H.K.; Lee, S.T.; Lee, M.H.; Hahn, J.S.; Ko, Y.W. Constitutive phosphorylation of Akt/PKB protein in acute myeloid leukemia: its significance as a prognostic variable. Leukemia, 2003, 17, 995-997.

[26] Kubota, Y.; Ohnishi, H.; Kitanaka, A.; Ishida, T.; Tanaka, T. Constitutive activation of PI3K is involved in the spon-taneous proliferation of primary acute myeloid leukemia cells: direct evidence of PI3K activation. Leukemia, 2004, 18, 1438-1440.

[27] Grandage, V.L.; Gale, R.E.; Linch, D.C.; Khwaja A. PI3-kinase/Akt is constitutively active in primary acute myeloid leukaemia cells and regulates survival and chemoresistance via NF-kappaB, Mapkinase and p53 pathways. Leukemia, 2005, 19, 586-594

[28] Récher, C.; Beyne-Rauzy, O.; Demur, C.; Chicanne, G.; Dos Santos, C.; Mas, V.M.; Benzaquen, D.; Laurent, G.; Huguet, F.; Payrastre, B. Antileukemic activity of rapamy-cin in acute myeloid leukemia. Blood, 2005, 105, 2527-2534.

[29] Dinner, S.; Platanias, L.C. Targeting mTOR pathway in leukemia. J. Cell. Biochem., 2016, 117, 1745-1752.

Dora Visnjic

Strikeout

Dora Visnjic

Note

agreement No. KK.01.1.01.0007,

Dora Visnjic

Strikeout


[30] Tabe, Y.; Tafuri, A.; Sekihara, K.; Yang, H.; Konopleva, M. Inhibition of mTOR kinase as a therapeutic target for acute myeloid leukemia. Expert Opin. Ther. Targets, 2017, 21, 705-714.

[31] Herschbein, L.; Liesveld, J.L. Dueling for dual inhibition: Means to enhance effectiveness of PI3K/Akt/mTOR inhibi-tors in AML. Blood Rev., 2017, doi:10.1016/j.blre.2017.11.006

[32] Konopleva, M.Y.; Walter, R.B.; Faderl, S.H.; Jabbour, E.J.; Zeng, Z.; Borthakur, G.; Huang, X.; Kadia, T.M.; Ruvolo, P.P.; Feliu, J.B.; Lu, H.; Debose, L.; Burger, J.A.; Andreeff, M.; Liu, W.; Baggerly, K.A.; Kornblau, S.M.; Doyle, L.A.; Estey, E.H.; Kantarjian, H.M. Preclinical and early clinical evaluation of the oral Akt inhibitor, MK-2206, for the treatment of acute myelogenous leukemia. Clin. Cancer Res., 2014, 20, 2226-2235.

[33] Goyo, I.; Perl, A.; Luger, S.; Baer, M.R.; Norsworthy, K.J.; Bauer, K.S.; Tidwell, M.; Fleckinger, S.; Carroll, M.; Sausville, E.A.Phase I study of UCN-01 and perifosine in patients with relapsed and refractory acute leukemias and high-risk myelodysplastic syndrome. Invest. New Drugs, 2013, 31, 1217-1227.

[34] Ragon, B.T.; Kantarjian, H.; Jabbour, E.; Ravandi, F.; Cor-tes, J.; Borthakur, G.; DeBose, L.; Zeng, Z.; Schneider, H.; Pemmaraju, N.; Garcia-Manero, G.; Kornblau, S.; Wierda, W.; Burger, J.; DiNardo C.D.; Andreeff, M.; Konopleva, M.; Daver, N. Buparlisib, a PI3K inhibitor, demonstrates acceptable tolerability and preliminary activity in a phase I trial of patients with advanced leukemias. Am. J. Hematol., 2017, 92, 7-11.

[35] Dos Santos, C.; Demur, C.; Bardet, V.; Prade-Houdellier, N.; Payrastre, B; Recher, C. A critical role for Lyn in acute myeloid leukemia. Blood, 2008, 111, 2269-2279.

[36] Kalaitzidis, D.; Sykes, S.M.; Wang, Z.; Punt, N.; Tang, Y.; Ragu, C.; Sinha, A.U.; Lane, S.W.; Souza, A.L.; Clish, C.B.; Anastasiou, D.; Gilliland, D.G.; Scadden, D.T.; Guer-tin, D.A.; Armstrong, S.A. mTOR complex 1 plays critical roles in hematopoiesis and Pten-loss-evoked leukemogene-sis. Cell Stem Cell, 2012, 11, 429-439.

[37] Hoshii, T.; Tadokoro, Y.; Naka, K.; Ooshio, T.; Muraguchi, T.; Sugiyama, N.; Soga, T.; Araki, K.; Yamamura, K.; Hirao, A. mTORC1 is essential for leukemia propagation but not stem cell self-renewal. J. Clin. Invest., 2012, 122, 2114-2129.

[38] Ghosh, J.; Kobayashi, M.; Ramdas, B.; Chatterjee, A.; Ma, P.; Mali, R.S.; Carlesso, N.; Liu, Y.; Plas, D.R.; Chan, R.J.; Kapur, R. S6K1 regulates hematopoietic stem cell self-renewal and leukemia maintenance. J. Clin. Invest., 2016, 126, 2621-2625.

[39] Gao, Y.; Gao, J.; Li, M.; Zheng, Y.; Wang, Y.; Zhang, H.; Wang, W.; Chu, Y.; Wang, X.; Xu, M.; Cheng, T.; Ju, Z.; Yuan, W. Rheb1 promotes tumor progression through mTORC1 in MLL-AF9-initiated murine acute myeloid leu-kemia. J. Hematol. Oncol., 2016, 9, 36.

[40] U.S. Food and Drug Administration. www.fda.gov (Ac-cessed June 28-30, 2017).

[41] Li, J.; Kim, S.G.; Blenis, J. Rapamycin: one drug; many effects. Cell Metab., 2014, 19, 373-379.

[42] Pollizzi, K.N.; Powell, J.D. Regulation of T cells by mTOR: the known knowns and the known unknowns. Trends in Immunol., 2015, 36, 13-20.

[43] Fantus, D.; Thomson, A.W. Evolving prospectives of mTOR complexes in immunity and transplantation. Am. J. Transplant., 2015, 15, 891-902.

[44] Eng, C.P.; Sehgal, S.N.; Vézina, C. Activity of rapamycin (AY-22;989) against transplanted tumors. J. Antibiot. (To-kyo), 1984, 37, 1231-1237.

[45] Price, D.J.; Grove, J.R.; Calvo, V.; Avruch, J.; Bierer, B.E. Rapamycin-induced inhibition of the 70-kilodalton S6 pro-tein kinase. Science, 1992, 257, 973-976.

[46] Muthukkumar S; Ramesh TM; Bondada S. Rapamycin; a potent immunosuppressive drug; causes programmed cell death in B lymphoma cells. Transplantation, 1995, 60, 264-270.

[47] Brown, V.I.; Fang, J.; Alcorn, K.; Barr, R.; Kim, J.M.; Wasserman, R.; Grupp, S.A. Rapamycin is active against B-precursor leukemia in vitro and in vivo, an effect that is modulated by IL-7-mediated signaling. Proc. Natl. Acad. Sci. U S A, 2003, 100, 15113-15118.

[48] Avellino, R.; Romano, S.; Parasole, R.; Bisogni, R.; Lam-berti, A.; Poggi, V.; Venuta, S.; Romano, M.F. Rapamycin stimulates apoptosis of childhood acute lymphoblastic leu-kemia cells. Blood, 2005, 106, 1400-1406.

[49] Mohi, M.G.; Boulton, C.; Gu, T.L.; Sternberg, D.W.; Neuberg, D.; Griffin, J.D.; Gilliland, D.G.; Neel, B.G. Combination of rapamycin and protein tyrosine kinase (PTK) inhibitors for the treatment of leukemias caused by oncogenic PTKs. Proc. Natl. Acad. Sci. U S A, 2004, 101, 3130-3135.

[50] Xu, Q.; Thomson, J.E.; Carroll, M. mTOR regulates cell survival after etoposide treatment in primary AML cells. Blood, 2005, 106, 4261-4268.

[51] Rizzieri, D.A.; Feldman, E.; Dipersio, J.F.; Gabrail, N.; Stock, W.; Strair, R.; Rivera, V.M.; Albitar, M.; Bedrosian, C.L.; Giles, F.J. A phase 2 clinical trial of deforolimus (AP23573; MK-8669); a novel mammalian target of ra-pamycin inhibitor, in patients with relapsed or refractory hematologic malignancies. Clin. Cancer Res., 2008, 14, 2756-2762.

[52] Yee, K.W.; Zeng, Z.; Konopleva, M.; Verstovsek, S.; Ra-vandi, F.; Ferrajoli, A.; Thomas, D.; Wierda, W.; Apostoli-dou, E.; Albitar, M.; O'Brien, S.; Andreeff, M.; Giles, F.J. Phase I/II study of the mammalian target of rapamycin in-hibitor everolimus (RAD001) in patients with relapsed or refractory hematologic malignancies. Clin. Cancer Res., 2006, 12, 5165-5173.

[53] Perl, A.E.; Kasner, M.T.; Tsai, D.E.; Vogl, D.T.; Loren, A.W.; Schuster, S.J.; Porter, D.L.; Stadtmauer, E.A.; Gold-stein, S.C.; Frey, N.V.; Nasta, S.D.; Hexner, E.O.; Dierov, J.K.; Swider, C.R.; Bagg, A.; Gewirtz, A.M.; Carroll, M.; Luger, S.M. A phase I study of the mammalian target of ra-pamycin inhibitor sirolimus and MEC chemotherapy in re-lapsed and refractory acute myelogenous leukemia. Clin. Cancer Res., 2009, 15, 6732-6739.

[54] Amadori, S.; Stasi, R.; Martelli, A.M.; Venditti, A.; Meloni, G.; Pane, F.; Martinelli, G.; Lunghi, M.; Pagano, L.; Cil-loni, D.; Rossetti, E.; Di Raimondo, F.; Fozza, C.; Annino, L.; Chiarini, F.; Ricci, F.; Ammatuna, E.; La Sala, E.; Fazi, P.; Vignetti, M. Temsirolimus; an mTOR inhibitor; in com-bination with lower-dose clofarabine as salvage therapy for older patients with acute myeloid leukaemia: results of a phase II GIMEMA study (AML-1107). Br. J. Haematol., 2012, 156, 205-212.

[55] Park, S.; Chapuis, N.; Saint Marcoux, F.; Recher, C.; Pre-bet, T.; Chevallier, P.; Cahn, J.Y.; Leguay, T.; Bories, P.; Witz, F.; Lamy, T.; Mayeux, P.; Lacombe, C.; Demur, C.; Tamburini, J.; Merlat, A.; Delepine, R.; Vey, N.; Dreyfus, F.; Béné, M.C.; Ifrah, N.; Bouscary, D.; GOELAMS (Groupe Ouest Est d’Etude des Leucémies aiguës et Autres Maladies du Sang). A phase Ib GOELAMS study of the mTOR inhibitor RAD001 in association with chemotherapy for AML patients in first relapse. Leukemia, 2013, 27, 1479-1486.

[56] Calimeri, T.; Ferreri, A.J.M. m-TOR inhibitors and their potential role in haematological malignancies. Br. J. Hae-matol., 2017, 177, 684-702.


[57] Bertagnolo, V.; Neri, L.M.; Marchisio, M.; Mischiati, C.; Capitani, S. Phosphoinositide 3-kinase activity is essential for all-trans-retinoic acid-induced granulocytic differentia-tion of HL-60 cells. Cancer Res., 1999, 59, 542-546.

[58] Sykes, S.M.; Lane, S.W.; Bullinger, L.; Kalaitzidis, D.; Yusuf, R.; Saez, B.; Ferraro, F.; Mercier, F.; Singh, H.; Brumme, K.M.; Acharya, S.S.; Scholl, C.; Tothova, Z.; At-tar, E.C.; Fröhling, S.; DePinho, R.A.; Armstrong, S.A.; Gilliland, D.G.; Scadden, D.T. AKT/FOXO signaling en-forces reversible differentiation blockade in myeloid leu-kemias. Cell, 2011, 146, 697-708.

[59] Jayaraman, T.; Marks, A.R. Rapamycin-FKBP12 blocks proliferation; induces differentiation; and inhibits cdc2 kinase activity in a myogenic cell line. J. Biol. Chem., 1993, 268, 25385-25388.

[60] Buscà, R.; Bertolotto, C.; Ortonne, J.P.; Ballotti, R. Inhibi-tion of the phosphatidylinositol 3-kinase/p70(S6)-kinase pathway induces B16 melanoma cell differentiation. J. Biol. Chem., 1996, 271, 31824-31830.

[61] Ogawa, T.; Tokuda, M.; Tomizawa, K.; Matsui, H.; Itano, T.; Konishi, R.; Nagahata, S.; Hatase, O. Osteoblastic dif-ferentiation is enhanced by rapamycin in rat osteoblast-like osteosarcoma (ROS 17/2.8) cells. Biochem. Biophys. Res. Commun., 1998, 249, 226-230.

[62] Yamamoto-Yamaguchi, Y.; Okabe-Kado, J.; Kasukabe, T.; Honma, Y. Induction of differentiation of human myeloid leukemia cells by immunosuppressant macrolides (rapamy-cin and FK506) and calcium/calmodulin-dependent kinase inhibitors. Exp. Hematol., 2001, 29, 582-588.

[63] Nishioka, C.; Ikezoe, T.; Yang, J.; Koeffler, H.P.; Yoko-yama, A. Blockade of mTOR signaling potentiates the abil-ity of histone deacetylase inhibitor to induce growth arrest and differentiation of acute myelogenous leukemia cells. Leukemia, 2008, 22, 2159-2168.

[64] Gadhoum, S.Z.; Madhoun, N.Y.; Abuelela, A.F.; Merzaban, J.S. Anti-CD44 antibodies inhibit both mTORC1 and mTORC2: a new rationale supporting CD44-induced AML differentiation therapy. Leukemia, 2016, 30, 2397-2401.

[65] Yang, J.; Ikezoe, T.; Nishioka, C.; Ni, L.; Koeffler, H.P.; Yokoyama, A. Inhibition of mTORC1 by RAD001 (ever-olimus) potentiates the effects of 1;25-dihydroxyvitamin D(3) to induce growth arrest and differentiation of AML cells in vitro and in vivo. Exp. Hematol., 2010, 38, 666-676.

[66] Dembitz, V.; Lalic, H.; Ostojic, A.; Vrhovac, R.; Banfic, H.; Visnjic, D. The mechanism of synergistic effects of ar-senic trioxide and rapamycin in acute myeloid leukemia cell lines lacking typical t(15;17) translocation. Int. J. Hematol., 2015, 102, 12-24.

[67] Isakson, P.; Bjørås, M.; Bøe, S.O.; Simonsen, A. Auto-phagy contributes to therapy-induced degradation of the PML/RARA oncoprotein. Blood, 2010, 116, 2324-2331.

[68] Neri, L.M.; Marchisio, M.; Colamussi, M.L.; Bertagnolo, V. Monocytic differentiation of HL-60 cells is characterized by the nuclear translocation of phosphatidylinositol 3-kinase and of definite phosphatidylinositol-specific phos-pholipase C isoforms. Biochem. Biophys. Res. Commun., 1999, 259, 314-320.

[69] Zhang, Y.; Zhang, J.; Studzinski, G.P. AKT pathway is activated by 1; 25-dihydroxyvitamin D3 and participates in its anti-apoptotic effect and cell cycle control in differentiat-ing HL60 cells. Cell Cycle, 2006, 5, 447-451.

[70] Carling, D.; Clarke, P.R.; Zammit, V.A.; Hardie, D.G. Puri-fication and characterization of the AMP-activated protein kinase. Copurification of acetyl-CoA carboxylase kinase and 3-hydroxy-3-methylglutaryl-CoA reductase kinase ac-tivities. Eur. J. Biochem., 1989, 186, 129-136.

[71] Carling, D. AMPK signalling in health and disease. Curr. Opin. Cell Biol., 2017, 45, 31-37.

[72] Garcia, D.; Shaw, R.J. AMPK: Mechanisms of Cellular Energy Sensing and Restoration of Metabolic Balance. Mol. Cell., 2017, 66, 789-800.

[73] Dasgupta, B.; Chhipa, R.R. Evolving Lessons on the Com-plex Role of AMPK in Normal Physiology and Cancer. Trends Pharmacol. Sci., 2016, 37, 192-206.

[74] Hardie, D.G. AMPK: a target for drugs and natural products with effects on both diabetes and cancer. Diabetes, 2013, 62, 2164-2172.

[75] Hardie, D.G. AMPK--sensing energy while talking to other signaling pathways. Cell Metab., 2014, 20, 939-952.

[76] Hardie, D.G. Molecular Pathways: Is AMPK a Friend or a Foe in Cancer? Clin. Cancer Res., 2015, 21, 3836-3840.

[77] Hawley, S.A.; Ross, F.A.; Chevtzoff, C.; Green, K.A.; Evans, A.; Fogarty, S.; Towler, M.C.; Brown, L.J.; Ogun-bayo, O.A.; Evans, A.M.; Hardie, D.G. Use of cells ex-pressing gamma subunit variants to identify diverse mecha-nisms of AMPK activation. Cell Metab., 2010, 11, 554-565.

[78] Liu, X.; Chhipa, R.R.; Pooya, S.; Wortman, M.; Yachyshin, S.; Chow, L.M.; Kumar, A.; Zhou, X.; Sun, Y.; Quinn, B.; McPherson, C.; Warnick, R.E.; Kendler, A.; Giri, S.; Poels, J.; Norga, K.; Viollet, B.; Grabowski, G.A.; Dasgupta, B. Discrete mechanisms of mTOR and cell cycle regulation by AMPK agonists independent of AMPK. Proc. Natl. Acad. Sci. U S A, 2014, 111, E435-E444.

[79] Vincent, E.E.; Coelho, P.P.; Blagih, J.; Griss, T.; Viollet, B.; Jones, R.G. Differential effects of AMPK agonists on cell growth and metabolism. Oncogene, 2015, 34, 3627-3639.

[80] O'Neill, H.M.; Maarbjerg, S.J.; Crane, J.D.; Jeppesen, J.; Jørgensen, S.B.; Schertzer, J.D.; Shyroka, O.; Kiens, B.; van Denderen, B.J.; Tarnopolsky, M.A.; Kemp, B.E.; Rich-ter, E.A.; Steinberg, G.R. AMP-activated protein kinase (AMPK) �1�2 muscle null mice reveal an essential role for AMPK in maintaining mitochondrial content and glu-cose uptake during exercise. Proc. Natl. Acad. Sci. U S A, 2011, 108, 16092-16097.

[81] Sakamoto, K.; McCarthy, A.; Smith, D.; Green, K.A.; Har-die, D.G.; Ashworth, A.; Alessi, D.R. Deficiency of LKB1 in skeletal muscle prevents AMPK activation and glucose uptake during contraction. EMBO. J., 2005, 24, 1810-1820.

[82] Sung, M.M.; Zordoky, B.N.; Bujak, A.L.; Lally, J.S.; Fung, D.; Young, M.E.; Horman, S.; Miller, E.J.; Light, P.E.; Kemp, B.E.; Steinberg, G.R.; Dyck, J.R. AMPK deficiency in cardiac muscle results in dilated cardiomyopathy in the absence of changes in energy metabolism. Cardiovasc. Res., 2015, 107, 235-245.

[83] Jessen, N.; Koh, H.J.; Folmes, C.D.; Wagg, C.; Fujii, N.; Løfgren, B.; Wolf, C.M.; Berul, C.I.; Hirshman, M.F.; Lo-paschuk, G.D.; Goodyear, L.J. Ablation of LKB1 in the heart leads to energy deprivation and impaired cardiac func-tion. Biochim. Biophys. Acta, 2010, 1802, 593-600.

[84] Russell, R.R.; Li, J.; Coven, D.L.; Pypaert, M.; Zechner, C.; Palmeri, M.; Giordano, F.J.; Mu, J.; Birnbaum, M.J.; Young, L.H. AMP-activated protein kinase mediates ischemic glucose uptake and prevents postischemic cardiac dysfunction; apoptosis; and injury. J. Clin. Invest., 2004, 114, 495-503.

[85] Shaw, R.J.; Lamia, K.A.; Vasquez, D.; Koo, S.H.; Barde-esy, N.; Depinho, R.A.; Montminy, M.; Cantley, L.C. The kinase LKB1 mediates glucose homeostasis in liver and therapeutic effects of metformin. Science, 2005, 310, 1642-1646.

[86] Foretz, M.; Hébrard, S.; Leclerc, J.; Zarrinpashneh, E.; Soty, M.; Mithieux, G.; Sakamoto, K.; Andreelli, F.; Viollet, B. Metformin inhibits hepatic gluconeogenesis in mice independently of the LKB1/AMPK pathway via a de-crease in hepatic energy state. J. Clin. Invest., 2010, 120, 2355-2369.


[87] Momcilovic, M.; Shackelford, D.B. Targeting LKB1 in cancer - exposing and exploiting vulnerabilities. Br. J. Can-cer., 2015, 113, 574-584.

[88] Hawley, S.A.; Boudeau, J.; Reid, J.L.; Mustard, K.J.; Udd, L.; Mäkelä, T.P.; Alessi, D.R.; Hardie, D.G. Complexes be-tween the LKB1 tumor suppressor; STRAD alpha/beta and MO25 alpha/beta are upstream kinases in the AMP-activated protein kinase cascade. J. Biol., 2003, 2, 28.

[89] Woods, A.; Johnstone, S.R.; Dickerson, K.; Leiper, F.C.; Fryer, L.G.; Neumann, D.; Schlattner, U.; Wallimann, T.; Carlson, M.; Carling, D. LKB1 is the upstream kinase in the AMP-activated protein kinase cascade. Curr. Biol., 2003, 13, 2004-2008.

[90] Shaw, R.J.; Kosmatka, M.; Bardeesy, N.; Hurley, R.L.; Witters, L.A.; DePinho, R.A.; Cantley, L.C. The tumor suppressor LKB1 kinase directly activates AMP-activated kinase and regulates apoptosis in response to energy stress. Proc. Natl. Acad. Sci. U S A, 2004, 101, 3329-3335.

[91] Inoki, K.; Zhu, T.; Guan, K.L. TSC2 mediates cellular en-ergy response to control cell growth and survival. Cell, 2003, 115, 577-590.

[92] Jones, R.G.; Plas, D.R.; Kubek, S.; Buzzai, M.; Mu, J.; Xu, Y.; Birnbaum, M.J.; Thompson, C.B. AMP-activated pro-tein kinase induces a p53-dependent metabolic checkpoint. Mol. Cell., 2005, 18, 283-293.

[93] Faubert, B.; Boily, G.; Izreig, S.; Griss, T.; Samborska, B.; Dong, Z.; Dupuy, F.; Chambers, C.; Fuerth, B.J.; Viollet, B.; Mamer, O.A.; Avizonis, D.; DeBerardinis, R.J.; Siegel, P.M.; Jones, R.G. AMPK is a negative regulator of the Warburg effect and suppresses tumor growth in vivo. Cell Metab., 2013, 17, 113-124.

[94] Huang, X.; Wullschleger, S.; Shpiro, N.; McGuire, V.A.; Sakamoto, K.; Woods, Y.L.; McBurnie, W.; Fleming, S.; Alessi, D.R. Important role of the LKB1-AMPK pathway in suppressing tumorigenesis in PTEN-deficient mice. Bio-chem. J., 2008, 412, 211-221.

[95] Shackelford, D.B.; Abt, E.; Gerken, L.; Vasquez, D.S.; Seki, A.; Leblanc, M.; Wei, L.; Fishbein, M.C.; Czernin, J.; Mischel, P.S.; Shaw, R.J. LKB1 inactivation dictates thera-peutic response of non-small cell lung cancer to the metabo-lism drug phenformin. Cancer Cell., 2013, 23, 143-158.

[96] Saito, Y.; Chapple, R.H.; Lin, A.; Kitano, A.; Nakada, D. AMPK Protects Leukemia-Initiating Cells in Myeloid Leu-kemias from Metabolic Stress in the Bone Marrow. Cell Stem Cell, 2015, 17, 585-596.

[97] Kishton, R.J.; Barnes, C.E.; Nichols, A.G.; Cohen, S.; Ger-riets, V.A.; Siska, P.J.; Macintyre, A.N.; Goraksha-Hicks, P.; de Cubas, A.A.; Liu, T.; Warmoes, M.O.; Abel, E.D.; Yeoh, A.E.; Gershon, T.R.; Rathmell, W.K.; Richards, K.L.; Locasale, J.W.; Rathmell, J.C. AMPK Is Essential to Balance Glycolysis and Mitochondrial Metabolism to Con-trol T-ALL Cell Stress and Survival. Cell Metab., 2016, 23, 649-662.

[98] Green, A.S.; Chapuis, N.; Maciel, T.T.; Willems, L.; Lam-bert, M.; Arnoult, C.; Boyer, O.; Bardet, V.; Park, S.; Foretz, M.; Viollet, B.; Ifrah, N.; Dreyfus, F.; Hermine, O.; Moura, I.C.; Lacombe, C.; Mayeux, P.; Bouscary, D.; Tam-burini, J. The LKB1/AMPK signaling pathway has tumor suppressor activity in acute myeloid leukemia through the repression of mTOR-dependent oncogenic mRNA transla-tion. Blood, 2010, 116, 4262-4273.

[99] Jude, J.G.; Spencer, G.J.; Huang, X.; Somerville T.D.D.; Jones, D.R.; Divecha, N.; Somerville, T.C.P. A targeted knockdown screen of genes coding for phosphoinositide modulators identifies PIP4K2A as required for acute mye-loid leukemia cell proliferation and survival. Oncogene, 2015; 34, 1253-1262.

[100] Visconte, V.; Przychodzen, B.; Han, Y.; Nawrocki, S.T.; Thota, S.; Kelly, K.R.; Patel, B.J.; Hirsch, C.; Advani, A.S.;

Carraway, H.E.; Sekeres, M.A.; Maciejewski, J.P.; Carew, J.S. Complete mutational spectrum of the autophagy inter-actome: a novel class of tumor suppressor genes in myeloid neoplasms. Leukemia, 2017, 31, 505-510.

[101] Monteverde, T.; Muthalagu, N.; Port, J.; Murphy, D.J. Evi-dence of cancer-promoting roles for AMPK and related kinases. FEBS J., 2015, 282, 4658-4671.

[102] Suyobert, P.; Poulain, L.; Paubelle, E.; Zylbersztejn, F.; Grenier, A.; Lambert, M.; Townsend, E.C.; Brusq, J.; Nicodeme, E.; Decrooqc, J.; Nepstad, I.; Green, A.S.; Mondesir, J.; Hospital, M.; Jacque, N; Christodoulou, A.; Desouza, T.A.; Hermine, O.; Foretz, M.; Viollet, B.; La-combe, C.; Mayeux, P.; Weinstock, D.M.; Moura, I.C.; Bouscary, D.; Tamburini, J. Co-activation of AMPK and mTORC1 induces cytotoxicity in acute myeloid leukemia. Cell Reports, 2015, 11, 1446-1457.

[103] Yasinska, I.M.; Gibbs, B.F.; Lall, G.S.; Sumbayev, V.V. The HIF-1 transcription complex is essential for transla-tional control of myeloid hematopoietic cell function by maintaining mTOR phosphorylation. Cell. Mol. Life Sci., 2014, 71, 699-710.

[104] Tabe, Y.; Yamamoto, S.; Saitoh, K.; Sekihara, K.; Monma, N.; Ikeo, K.; Mogushi, K.; Shikami, M.; Ruvolo, V.; Ishi-zava, J.; Hail, N.; Kazuno, S.; Igarashi, M.; Matsushita, H.; Yamanaka, Y.; Arai, H.; Nagaoka, I.; Miida, T.; Hayashi-zaki, Y.; Konopleva, M.; Andreeff, M. Bone marrow adipo-cytes facilitate fatty acid oxidation activating AMPK and a transcriptional network supporting survival of acute mono-cytic leukemia cells. Cancer Res., 2017, 77, 1453-1464.

[105] Pryor, R.; Cabreiro, F. Repurposing metformin: an old drug with new tricks in its binding pockets. Biochem. J., 2015, 471, 307-322.

[106] Christodoulou, M.I.; Scorilas, A. Metformin and Anti-Cancer Therapeutics: Hopes for a More Enhanced Arma-mentarium Against Human Neoplasias? Curr. Med. Chem., 2017, 24, 14-56.

[107] Chae, Y.K.; Arya, A.; Malecek, M.K.; Shin, D.S.; Carneiro, B.; Chandra, S.; Kaplan, J.; Kalyan, A.; Altman, J.K.; Pla-tanias, L.; Giles, F. Repurposing metformin for cancer treatment: current clinical studies. Oncotarget, 2016, 7, 40767-40780.

[108] Rosilio, C.; Ben-Sahra, I.; Bost, F.; Peyron, J.F. Metformin: a metabolic disruptor and anti-diabetic drug to target human leukemia. Cancer Lett., 2014, 346, 188-196.

[109] Zhou, G.; Myers, R.; Li, Y.; Chen, Y.; Shen, X.; Fenyk-Melody, J.; Wu, M.; Ventre, J.; Doebber, T.; Fujii, N.; Musi, N.; Hirshman, M.F.; Goodyear, L.J.; Moller, D.E. Role of AMP-activated protein kinase in mechanism of metformin action. J. Clin. Invest., 2001, 108, 1167-1174.

[110] Turban, S.; Stretton, C.; Drouin, O.; Green, C.J.; Watson, M.L.; Gray, A.; Ross, F.; Lantier, L.; Viollet, B.; Hardie, D.G.; Marette, A.; Hundal, H.S. Defining the contribution of AMP-activated protein kinase (AMPK) and protein kinase C (PKC) in regulation of glucose uptake by met-formin in skeletal muscle cells. J. Biol. Chem., 2012, 287, 20088-20099.

[111] Evans, J.M.; Donnelly, L.A.; Emslie-Smith, A.M.; Alessi, D.R.; Morris, A.D. Metformin and reduced risk of cancer in diabetic patients. BMJ., 2005, 330, 1304-1305.

[112] Lee, D.K.; Szabo, E. Repurposing Drugs for Cancer Pre-vention. Curr. Top. Med. Chem, 2016, 16, 2169-2178.

[113] Kordes, S.; Pollak, M.N.; Zwinderman, A.H.; Mathôt, R.A.; Weterman, M.J.; Beeker, A.; Punt, C.J.; Richel, D.J.; Wil-mink, J.W. Metformin in patients with advanced pancreatic cancer: a double-blind, randomized, placebo-controlled phase 2 trial. Lancet Oncol., 2015, 16, 839-847.

[114] Reni, M.; Dugnani, E.; Cereda, S.; Belli, C.; Balzano, G.; Nicoletti, R.; Liberati, D.; Pasquale, V.; Scavini, M.; Mag-giora, P.; Sordi, V.; Lampasona, V.; Ceraulo, D.; Di Ter-

Dora Visnjic

Highlight


lizzi, G.; Doglioni, C.; Falconi, M.; Piemonti, L. (Ir)relevance of Metformin Treatment in Patients with Me-tastatic Pancreatic Cancer: An Open-Label, Randomized Phase II Trial. Clin. Cancer Res., 2016, 22, 1076-1085.

[115] ClinicalTrials.gov, a service of the U.S. National Institutes of Health. clinicaltrials.gov (Accessed July 2017).

[116] Zakikhani, M.; Dowling, R.; Fantus, I.G.; Sonenberg, N.; Pollak, M. Metformin is an AMP kinase-dependent growth inhibitor for breast cancer cells. Cancer Res., 2006, 66, 10269-10273.

[117] El-Mir, M.Y.; Nogueira, V.; Fontaine, E.; Avéret, N.; Rig-oulet, M.; Leverve, X. Dimethylbiguanide inhibits cell res-piration via an indirect effect targeted on the respiratory chain complex I. J. Biol. Chem., 2000, 275, 223-228.

[118] Hawley, S.A.; Gadalla, A.E.; Olsen, G.S.; Hardie, D.G. The antidiabetic drug metformin activates the AMP-activated protein kinase cascade via an adenine nucleotide-independent mechanism. Diabetes, 2002, 51, 2420-2425.

[119] Zou, M.H.; Hou, X.Y.; Shi, C.M.; Kirkpatick, S.; Liu, F.; Goldman, M.H.; Cohen, R.A. Activation of 5'-AMP-activated kinase is mediated through c-Src and phosphoi-nositide 3-kinase activity during hypoxia-reoxygenation of bovine aortic endothelial cells. Role of peroxynitrite. J. Biol. Chem., 2003, 278, 34003-34010.

[120] Xie, Z.; Dong, Y.; Scholz, R.; Neumann, D.; Zou, M.H. Phosphorylation of LKB1 at serine 428 by protein kinase C-zeta is required for metformin-enhanced activation of the AMP-activated protein kinase in endothelial cells. Circula-tion, 2008, 117, 952-962.

[121] Ben Sahra, I.; Regazzetti, C.; Robert, G.; Laurent, K.; Le Marchand-Brustel, Y.; Auberger, P.; Tanti, J.F.; Giorgetti-Peraldi, S.; Bost, F. Metformin; independent of AMPK; in-duces mTOR inhibition and cell-cycle arrest through REDD1. Cancer Res., 2011, 71, 4366-4372.

[122] Yi, Y.; Chen, D.; Ao, J.; Sun, S.; Wu, M.; Li, X.; Bergholz, J.; Zhang, Y.; Xiao, Z.X. Metformin Promotes AMP-activated Protein Kinase-independent Suppression of ΔNp63α Protein Expression and Inhibits Cancer Cell Vi-ability. J. Biol. Chem., 2017, 292, 5253-5261.

[123] Foretz, M.; Guigas, B.; Bertrand, L.; Pollak, M.; Viollet, B. Metformin: from mechanisms of action to therapies. Cell Metab., 2014, 20, 953-966.

[124] Wu, L.; Zhu, J.; Prokop, L.J.; Murad, M.H. Pharmacologic Therapy of Diabetes and Overall Cancer Risk and Mortal-ity: A Meta-Analysis of 265 Studies. Sci Rep., 2015, 5, 10147.

[125] Vakana, E.; Altman, J.K.; Glaser, H.; Donato, N.J.; Pla-tanias, L.C. Antileukemic effects of AMPK activators on BCR-ABL-expressing cells. Blood, 2011, 118, 6399-6402.

[126] Rosilio, C.; Lounnas, N.; Nebout, M.; Imbert, V.; Hagen-beek, T.; Spits, H.; Asnafi, V.; Pontier-Bres, R.; Reverso, J.; Michiels, J.F.; Sahra, I.B.; Bost, F.; Peyron, J.F. The meta-bolic perturbators metformin; phenformin and AICAR in-terfere with the growth and survival of murine PTEN-deficient T cell lymphomas and human T-ALL/T-LL cancer cells. Cancer Lett., 2013, 336, 114-126.

[127] Jagannathan, S.; Abdel-Malek, M.A.; Malek, E.; Vad, N.; Latif, T.; Anderson, K.C.; Driscoll, J.J. Pharmacologic screens reveal metformin that suppresses GRP78-dependent autophagy to enhance the anti-myeloma effect of borte-zomib. Leukemia, 2015, 29, 2184-2191.

[128] Shi, W.Y.; Xiao, D.; Wang, L.; Dong, L.H.; Yan, Z.X.; Shen, Z.X.; Chen, S.J.; Chen, Y.; Zhao, W.L. Therapeutic metformin/AMPK activation blocked lymphoma cell growth via inhibition of mTOR pathway and induction of autophagy. Cell Death Dis., 2012, 3, e275.

[129] Leclerc, G.M.; Leclerc, G.J.; Kuznetsov, J.N.; DeSalvo, J.; Barredo, J.C. Metformin induces apoptosis through AMPK-

dependent inhibition of UPR signaling in ALL lym-phoblasts. PLoS One, 2013, 8, e74420.

[130] Yi, Y.; Gao, L.; Wu, M.; Ao, J.; Zhang, C.; Wang, X.; Lin, M.; Bergholz, J.; Zhang, Y.; Xiao, Z.J. Metformin sensi-tizes leukemia cells to vincristine via activation of AMP-activated protein kinase. J. Cancer, 2017, 8, 2636-2642.

[131] Scotland, S.; Saland, E.; Skuli, N.; de Toni, F.; Boutzen, H.; Micklow, E.; Sénégas, I.; Peyraud, R.; Peyriga, L.; Théodoro, F.; Dumon, E.; Martineau, Y.; Danet-Desnoyers, G.; Bono, F.; Rocher, C.; Levade, T.; Manenti, S.; Junot, C.; Portais, J.C.; Alet, N.; Récher, C.; Selak, M.A.; Carroll, M.; Sarry, J.E. Mitochondrial energetic and AKT status mediate metabolic effects and apoptosis of metformin in human leukemic cells. Leukemia, 2013, 27, 2129-2138.

[132] Ceacareanu, A.C.; Nimako, G.K.; Wintrob, Z.A.P. Missing the benefit of metformin in acute myeloid leukemia: A problem of contrast? J Res. Pharm. Pract., 2017, 6, 145-150.

[133] Huai, L.; Wang, C.; Zhang, C.; Li, Q.; Chen, Y.; Jia, Y.; Li, Y.; Xing, H.; Tian, Z.; Rao, Q.; Wang, M.; Wang, J. Met-formin induces differentiation in acute promyelocytic leu-kemia by activating the MEK/ERK signaling pathway. Bio-chem. Biophys. Res. Commun., 2012, 422, 398-404.

[134] Kawashima, I.; Mitsumori, T.; Nozaki, Y.; Yamamoto, T.; Shobu-Sueki, Y.; Nakajima, K.; Kirito, K. Negative regula-tion of the LKB1/AMPK pathway by ERK in human acute myeloid leukemia cells. Exp. Hematol., 2015, 43, 524-533.

[135] Miranda, M.B.; McGuire, T.F.; Johnson, D.E. Importance of MEK-1/-2 signaling in monocytic and granulocytic dif-ferentiation of myeloid cell lines. Leukemia, 2002, 16, 683-692.

[136] Vasamsetti, S.B.; Karnewar, S.; Kanugula, A.K.; Thatipalli, A.R.; Kumar, J.M.; Kotamraju, S. Metformin inhibits monocyte-to-macrophage differentiation via AMPK-mediated inhibition of STAT3 activation: potential role in atherosclerosis. Diabetes, 2015, 64, 2028-2041.

[137] Eikawa, S.; Nishida, M.; Mizukami, S.; Yamazaki, C.; Na-kayama, E.; Udono, H. Immune-mediated antitumor effect by type 2 diabetes drug, metformin. Proc. Natl. Acad. Sci. U S A, 2015, 112, 1809-1814.

[138] Limagne, E.; Thibaudin, M.; Euvrard, R.; Berger, H.; Cha-lons, P.; Végan, F.; Humblin, E.; Boidot, R.; Rébé, C.; De-rangère, V.; Ladoire, S.; Apetoh, L.; Delmas, D.; Ghiring-helli, F. Sirtuin-1 Activation Controls Tumor Growth by Impeding Th17 Differentiation via STAT3 Deacetylation. Cell Rep., 2017, 19, 746-759.

[139] Liu, X.; Zheng, H.; Yu, W.M.; Cooper, T.M.; Bunting, K.D.; Qu, C.K. Maintenance of mouse hematopoietic stem cells ex vivo by reprogramming cellular metabolism. Blood, 2015, 125, 1562-1565.

[140] Zhang, Q.S.; Tang, W.; Deater, M.; Phan, N.; Marcogliese, A.N.; Li, H.; Al-Dhalimy, M.; Major, A.; Olson, S.; Mon-nat, R.J.; Grompe, M. Metformin improves defective hema-topoiesis and delays tumor formation in Fanconi anemia mice. Blood, 2016, 128, 2774-2784.

[141] Van Den Neste, E.; Van den Berghe, G.; Bontemps, F. AICA-riboside (acadesine), an activator of AMP-activated protein kinase with potential for application in hematologic malignancies. Expert Opin. Investig. Drugs, 2010, 19, 571-578.

[142] Pinson, B.; Vaur, S.; Sagot, I.; Coulpier, F.; Lemoine, S.; Daignan-Fornier, B. Metabolic intermediates selectively stimulate transcription factor interaction and modulate phosphate and purine pathways. Genes Dev., 2009, 23, 1399-1407.

[143] Daignan-Fornier, B.; Pinson, B. 5-Aminoimidazole-4-carboxamide-1-beta-D-ribofuranosyl 5'-monophosphate (AICAR), a highly conserved purine intermediate with mul-tiple effects. Metabolites, 2012, 2, 292-302

Dora Visnjic

Highlight

Dora Visnjic

Highlight

Dora Visnjic

Highlight

be so kind to change font -(italic for name of the journal, BOLD for year, italic for volume)


[144] Mangano D.T. Effects of acadesine on myocardial infarc-tion; stroke; and death following surgery. A meta-analysis of the 5 international randomized trials. The Multicenter Study of Perioperative Ischemia (McSPI) Research Group. JAMA., 1997, 277, 325-332.

[145] Henin, N.; Vincent, M.F.; Gruber, H.E.; Van den Berghe, G. Inhibition of fatty acid and cholesterol synthesis by stimulation of AMP-activated protein kinase. FASEB J., 1995, 9, 541-546.

[146] Corton, J.M.; Gillespie, J.G.; Hawley, S.A.; Hardie, D.G. 5-aminoimidazole-4-carboxamide ribonucleoside. A specific method for activating AMP-activated protein kinase in in-tact cells? Eur. J. Biochem., 1995, 229, 558-565.

[147] Narkar, V.A.; Downes, M.; Yu, R.T.; Embler, E.; Wang, Y.X.; Banayo, E.; Mihaylova, M.M.; Nelson, M.C.; Zou, Y.; Juguilon, H.; Kang, H.; Shaw, R.J.; Evans, R.M. AMPK and PPARdelta agonists are exercise mimetics. Cell, 2008, 134, 405-415.

[148] Lanner, J.T.; Georgiou, D.K.; Dagnino-Acosta, A.; Ain-binder, A.; Cheng, Q.; Joshi, A.D.; Chen, Z.; Yarotskyy, V.; Oakes, J.M.; Lee, C.S.; Monroe, T.O.; Santillan, A.; Dong, K.; Goodyear, L.; Ismailov, I.I.; Rodney, G.G.; Dirksen, R.T.; Hamilton, S.L. AICAR prevents heat-induced sudden death in RyR1 mutant mice independent of AMPK activa-tion. Nat. Med., 2012, 18, 244-251.

[149] Bost, F.; Decoux-Poullot, A.G.; Tanti, J.F.; Clavel, S. En-ergy disruptors: rising stars in anticancer therapy? Onco-genesis, 2016, 5:e188.

[150] Merrill, G.F.; Kurth, E.J.; Hardie, D.G.; Winder, W.W. AICA riboside increases AMP-activated protein kinase; fatty acid oxidation; and glucose uptake in rat muscle. Am. J. Physiol., 1997, 273: E1107-1112.

[151] Vincent, M.F.; Erion, M.D.; Gruber, H.E.; Van den Berghe, G. Hypoglycaemic effect of AICAriboside in mice. Diabe-tologia, 1996, 39, 1148-1155.

[152] Bergeron, R.; Previs, S.F.; Cline, G.W.; Perret, P.; Russell, R.R.; Young, L.H.; Shulman, G.I. Effect of 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside infusion on in vivo glucose and lipid metabolism in lean and obese Zucker rats. Diabetes, 2001, 50, 1076-1082.

[153] Cuthbertson, D.J.; Babraj, J.A.; Mustard, K.J.; Towler, M.C.; Green, K.A.; Wackerhage, H.; Leese, G.P.; Baar, K.; Thomason-Hughes, M.; Sutherland, C.; Hardie, D.G.; Ren-nie, M.J. 5-aminoimidazole-4-carboxamide 1-beta-D-ribofuranoside acutely stimulates skeletal muscle 2-deoxyglucose uptake in healthy men. Diabetes, 2007, 56, 2078-2084.

[154] McGee, S.L.; van Denderen, B.J.; Howlett, K.F.; Mollica, J.; Schertzer, J.D.; Kemp, B.E.; Hargreaves, M. AMP-activated protein kinase regulates GLUT4 transcription by phosphorylating histone deacetylase 5. Diabetes, 2008, 57, 860-867.

[155] Mu, J.; Brozinick, J.T. Jr; Valladares, O.; Bucan, M.; Birn-baum, M.J. A role for AMP-activated protein kinase in con-traction- and hypoxia-regulated glucose transport in skeletal muscle. Mol. Cell, 2001, 7, 1085-1094.

[156] Dzamko, N.; Schertzer, J.D.; Ryall, J.G.; Steel, R; Macaulay, S. L.; Wee, S.; Chen, Z.; Michell, B.J.; Oakhill, J.S.; Watt, M.J.; Jørgensen, S.B.; Lynch, G.S.; Kemp, B.E.; Steinberg, G.R. AMPK-independent pathways regulate skeletal muscle fatty acid oxidation. J. Physiol., 2008, 586, 5819–5831.

[157] Dixon, R.; Gourzis, J.; McDermott, D.; Fujitaki, J.; Dew-land, P.; Gruber, H. AICA-riboside: safety; tolerance; and pharmacokinetics of a novel adenosine-regulating agent. J. Clin. Pharmacol., 1991, 31, 342-347.

[158] Drew, B.G.; Kingwell, B.A. Acadesine; an adenosine-regulating agent with the potential for widespread indica-tions. Expert Opin. Pharmacother., 2008, 9, 2137-2144.

[159] Mangano, D.T.; Miao, Y.; Tudor, I.C.; Dietzel, C.; Investi-gators of the Multicenter Study of Perioperative Ischemia (McSPI) Research Group.; Ischemia Research and Educa-tion Foundation (IREF). Post-reperfusion myocardial in-farction: long-term survival improvement using adenosine regulation with acadesine. J. Am. Coll. Cardiol., 2006, 48, 206-214.

[160] Newman, M.F.; Ferguson, T.B.; White, J.A.; Ambrosio, G.; Koglin, J.; Nussmeier, N.A.; Pearl, R.G.; Pitt, B.; Wechsler, A.S.; Weisel, R.D.; Reece, T.L.; Lira, A.; Harrington, R.A.; RED-CABG Steering Committee and Investigators. Effect of adenosine-regulating agent acadesine on morbidity and mortality associated with coronary artery bypass grafting: the RED-CABG randomized controlled trial. JAMA, 2012, 308, 157-164.

[161] Szentmiklósi, A.J.; Cseppento, A.; Harmati, G.; Nánási, P.P. Novel trends in the treatment of cardiovascular disor-ders: site- and event- selective adenosinergic drugs. Curr. Med. Chem., 2011 18, 1164-1187.

[162] Fischetto, G.; Bermon, S. From gene engineering to gene modulation and manipulation: can we prevent or detect gene doping in sports? Sports Med., 2013, 43, 965-977.

[163] Pokrywka, A.; Cholbinski, P.; Kaliszewski, P.; Kowalczyk, K.; Konczak, D.; Zembron-Lacny, A. Metabolic modulators of the exercise response: doping control analysis of an ago-nist of the peroxisome proliferator-activated receptor δ (GW501516) and 5-aminoimidazole-4-carboxamide ribo-nucleotide (AICAR). J. Physiol. Pharmacol., 2014, 65, 469-476.

[164] Campàs, C.; Lopez, J.M.; Santidrián, A.F.; Barragán M.; Bellosillo, B.; Colomer, D.; Gil, J. Acadesine activates AMPK and induces apoptosis in B-cell chronic lymphocytic leukemia cells but not in T lymphocytes. Blood, 2003, 101, 3674-3680.

[165] Van Den Neste, E.; Cazin, B.; Janssens, A.; Gonzalez-Barca, E.; Terol, M.J.; Levy, V.; Perez de Oteyza, J.; Zachee, P.; Saunders, A.; de Frias, M; Campàs, C. Acadesine for patients with relapsed/refractory chronic lymphocytic leukemia (CLL): a multicenter phase I/II study. Cancer Chemother. Pharmacol., 2013, 71, 581-591.

[166] Campàs, C.; Santidrián, A.F.; Domingo, A.; Gil, J. Acadesine induces apoptosis in B cells from mantle cell lymphoma and splenic marginal zone lymphoma. Leukemia, 2005, 19, 292-294.

[167] Sengupta, T.K.; Leclerc, G.M.; Hsieh-Kinser, T.T.; Leclerc, G.J.; Singh, I.; Barredo, J.C. Cytotoxic effect of 5-aminoimidazole-4-carboxamide-1-beta-4-ribofuranoside (AICAR) on childhood acute lymphoblastic leukemia (ALL) cells: implication for targeted therapy. Mol. Cancer., 2007, 6, 46.

[168] Robert, G.; Ben Sahra, I.; Puissant, A.; Colosetti, P.; Bel-hacene, N.; Gounon, P.; Hofman, P.; Bost, F.; Cassuto, J.P.; Auberger, P. Acadesine kills chronic myelogenous leuke-mia (CML) cells through PKC-dependent induction of autophagic cell death. PLoS One., 2009, 4:e7889.

[169] Santidrián, A.F.; González-Gironès, D.M.; Iglesias-Serret, D.; Coll-Mulet, L.; Cosialls, A.M.; de Frias, M.; Campàs, C.; González-Barca, E.; Alonso, E.; Labi, V.; Viollet, B.; Benito, A.; Pons, G.; Villunger, A.; Gil, J. AICAR induces apoptosis independently of AMPK and p53 through up-regulation of the BH3-only proteins BIM and NOXA in chronic lymphocytic leukemia cells. Blood, 2010, 116, 3023-3032.

[170] Montraveta, A.; Xargay-Torrent, S.; López-Guerra, M.; Rosich, L.; Pérez-Galán, P.; Salaverria, I.; Beà, S.; Kalko, S.G.; de Frias, M.; Campàs, C.; Roué, G.; Colomer, D. Synergistic anti-tumor activity of acadesine (AICAR) in combination with the anti-CD20 monoclonal antibody ri-


tuximab in in vivo and in vitro models of mantle cell lym-phoma. Oncotarget, 2014, 5, 726-739.

[171] Montraveta, A.; Xargay-Torrent, S.; Rosich, L.; López-Guerra, M.; Roldán, J.; Rodríguez, V.; Lee-Vergés, E.; de Frías, M.; Campàs, C.; Campo, E.; Roué, G.; Colomer, D. Bcl-2 high mantle cell lymphoma cells are sensitized to acadesine with ABT-199. Oncotarget, 2015, 6, 21159-21172.

[172] Zang, Y.; Yu, L.F.; Pang, T.; Fang, L.P.; Feng, X.; Wen, T.Q.; Nan F.J.; Feng, L.Y.; Li, J. AICAR induces astroglial differentiation of neural stem cells via activating the JAK/STAT3 pathway independently of AMP-activated pro-tein kinase. J. Biol. Chem., 2008, 283, 6201-6208.

[173] Sun, X.; Yang, Q.; Rogers, C.J.; Du, M.; Zhu, M.J. AMPK improves gut epithelial differentiation and barrier function via regulating Cdx2 expression. Cell Death Differ. 2017, 24, 819-831.

[174] Chae, H.D.; Lee, M.R.; Broxmeyer, H.E. 5-Aminoimidazole-4-carboxyamide ribonucleoside induces G(1)/S arrest and Nanog downregulation via p53 and en-hances erythroid differentiation. Stem Cells, 2012, 30, 140-149.

[175] Johnson, D.E.; Redner, R.L. An ATRActive future for dif-ferentiation therapy of AML. Blood Rev., 2015, 29, 263-268.

[176] De The, H. Differentiation therapy revisited. Nat. Rev. Can-cer., 2017, doi:10.1038/nrc.2017.103.

[177] Xie, N.; Zhong, I.; Liu, I.; Fang, Y.; Qi, X.; Cao, J.; Yang, B.; He, X.; Ying, M. Autophagy contributes to dasatinib-induced myeloid differentiation of human acute myeloid leukemia cells. Biochem.Pharmacol., 2014, 89, 74-85.

[178] Dembitz, V.; Lalic, H.; Visnjic, D. 5-aminoimidazole-4-carboxamide ribonucleoside-induced autophagy flux during differentiation of monocytic leukemia cells. Cell Death Discov., 2017, 3, 17066.

[179] Velez, J.; Pan, R.; Lee, J.T.; Enciso, L.; Suarez, M.; Duque, J.E.; Jaramillo, D.; Lopez, C.; Morales, L.; Bornmann, W.; Konopleva, M.; Krystal, G.; Andreeff, M.; Samudio, I. Biguanides sensitize leukemia cells to ABT-737-induced apoptosis by inhibiting mitochondrial electron transport. Oncotarget, 2016, 7, 51435-51449.

[180] Visnjic, D.; Lalic, H.; Dembitz, V.; Banfic, H. Metabolism and differentiation. Period Biol., 2014, 116, 37-43.

[181] Samudio, I.; Harmancey, R.; Fiegl, M.; Kantarjian, H.; Ko-nopleva, M.; Korchin, B.; Kaluarachchi, K.; Bornmann, W.; Duvvuri, S.; Taegtmeyer, H.; Andreeff, M. Pharmacologic inhibition of fatty acid oxidation sensitizes human leukemia cells to apoptosis induction. J. Clin. Invest., 2010, 120, 142-156.

[182] Skrtić, M.; Sriskanthadevan, S.; Jhas, B.; Gebbia, M.; Wang, X.; Wang, Z.; Hurren, R.; Jitkova, Y.; Gronda, M.; Maclean, N.; Lai, C.K.; Eberhard, Y.; Bartoszko, J.; Spag-nuolo, P.; Rutledge, A.C.; Datti, A.; Ketela, T.; Moffat, J.; Robinson, B.H.; Cameron, J.H.; Wrana, J.; Eaves, C.J.;

Minden, M.D.; Wang, J.C.; Dick, J.E.; Humphries, K.; Nis-low, C.; Giaever, G.; Schimmer, A.D. Inhibition of mito-chondrial translation as a therapeutic strategy for human acute myeloid leukemia. Cancer Cell, 2011, 20, 674-688.

[183] Poulain, L.; Sujobert, P.; Zylbersztejn, F.; Barreau, S.; Stu-ani, L.; Lambert, M.; Palama, T.L.; Chesnais, V.; Birsen, R.; Vergez, F.; Farge, T.; Chenevier-Gobeaux, C.; Fraisse, M.; Bouillaud, F.; Debeissat, C.; Herault, O.; Récher, C.; Lacombe, C.; Fontenay, M.; Mayeux, P.; Maciel, T.T.; Por-tais, J.C.; Sarry, J.E.; Tamburini, J.; Bouscary, D.; Chapuis, N. High mTORC1 activity drives glycolysis addiction and sensitivity to G6PD inhibition in acute myeloid leukemia cells. Leukemia, 2017 Mar 28. doi: 10.1038/leu.2017.81.

[184] Jacque, N.; Ronchetti, A.M.; Larrue, C.; Meunier, G.; Birsen, R.; Willems, L.; Saland, E.; Decroocq, J.; Maciel, T.T.; Lambert, M.; Poulain, L.; Hospital, M.A.; Sujobert, P.; Joseph, L.; Chapuis, N.; Lacombe, C.; Moura, I.C.; Demo, S.; Sarry, J.E.; Recher, C.; Mayeux, P.; Tamburini, J.; Bouscary, D. Targeting glutaminolysis has antileukemic activity in acute myeloid leukemia and synergizes with BCL-2 inhibition. Blood, 2015, 126, 1346-1356.

[185] Sykes, D.B.; Kfoury, Y.S.; Mercier, F.E.; Wawer, M.J.; Law, J.M.; Haynes, M.K.; Lewis, T.A.; Schajnovitz, A.; Jain, E.; Lee, D.; Meyer. H.; Pierce. K.A.; Tolliday. N.J.; Waller. A.; Ferrara. S.J.; Eheim, A.L.; Stoeckigt, D.; Maxcy, K.L.; Cobert, J.M.; Bachand, J.; Szekely, B.A.; Mukherjee, S.; Sklar, L.A.; Kotz, J.D.; Clish, C.B.; Sadreyev, R.I.; Clemons, P.A.; Janzer, A.; Schreiber, S.L.; Scadden, D.T. Inhibition of Dihydroorotate Dehydrogenase Overcomes Differentiation Blockade in Acute Myeloid Leukemia. Cell, 2016, 167, 171-186.

[186] Pikman, Y.; Puissant, A.; Alexe, G.; Furman, A.; Chen, L.M.; Frumm, S.M.; Ross, L.; Fenouille, N.; Bassil, C.F.; Lewis, C.A.; Ramos, A.; Gould, J.; Stone, R.M.; DeAngelo, D.J.; Galinsky, I.; Clish, C.B.; Kung, A.L.; Hemann, M.T.; Vander Heiden, M.G.; Banerji, V.; Stegmaier, K. Targeting MTHFD2 in acute myeloid leukemia. J Exp Med, 2016, 213,1285-306.

[187] Amatangelo, M.D.; Quek, L.; Shih, A.; Stein, E.M.; Roshal, M.; David, M.D.; Marteyn, B.; Rahnamay Farnoud, N.; de Botton, S.; Bernard, O.A.; Wu, B.; Yen, K.E.; Tallman, M.S.; Papaemmanuil, E.; Penard-Lacronique, V.; Thakurta, A.; Vyas, P.; Levine, R.L. Enasidenib induces acute mye-loid leukemia cell differentiation to promote clinical re-sponse. Blood, 2017 pii: blood-2017-04-779447.

[188] Stein, E.M.; DiNardo, C.D.; Pollyea, D.A.; Fathi, A.T.; Roboz, G.J.; Altman, J.K.; Stone, R.M.; DeAngelo, D.J.; Levine, R.L.; Flinn, I.W.; Kantarjian, H.M.; Collins, R.; Patel, M.R.; Frankel, A.E.; Stein, A.; Sekeres, M.A.; Swords, R.T.; Medeiros, B.C.; Willekens, C.; Vyas, P.; To-solini, A.; Xu, Q.; Knight, R.D.; Yen, K.E; Agresta, S.; de Botton, S.; Tallman, M.S. Enasidenib in mutant-IDH2 re-lapsed or refractory acute myeloid leukemia. Blood, 2017, pii: blood-2017-04-779405.

DISCLAIMER: The above article has been published in Epub (ahead of print) on the basis of the materials provided by the author. The Editorial Department reserves the right to make minor modifications for further improvement of the manuscript.

PMID: 29345570

Documents

REVIEW ARTICLE The Role of AMPK/mTOR Modulators in the … · 2018. 6. 19. · tivity of protein kinase B/Akt were observed, and siRNA-mediated down-modulation of Akt in HL-60 cells