6794

STK11

LKB1|PJS|hLKB1;serine/threonine kinase 11;STK11;serine/threonine kinase 11

liver kinase B1|polarization-related protein LKB1|renal carcinoma antigen NY-REN-19|serine/threonine-protein kinase 11|serine/threonine-protein kinase LKB1|serine/threonine-protein kinase STK11

19p13.3

protein-coding

Germ-line mutations of LKB1 and PTEN tumor suppressor genes underlie the phenotypically related Peutz-Jeghers syndrome (PJS) and Cowden disease (CD), respectively. To analyze possible developmental roles of PTEN and LKB1, we have studied their mRNA expression during mouse embryonic development (E7-17.5) by in situ hybridization. Ubiquitous expression of both genes during early stages (E7-11) became more restricted in later embryonic development (E15-19) where LKB1 and PTEN showed prominent overlapping expression in e.g. gastrointestinal tract and lung. In contrast, LKB1 was selectively expressed at high levels in testis and PTEN was prominently expressed in skin epithelium and underlying mesenchyme. These results indicate that LKB1 and PTEN display largely overlapping expression patterns during embryonic development. Moreover, a high expression of these genes was observed in the tissues and organs affected in PJS and CD patients and in PTEN+/- mice.

PURPOSE: LKB1 (also called STK11) is a recently identified tumor suppressor gene in which its mutation can lead to Peutz-Jeghers syndrome, characterized by gastrointestinal polyps and cancers of different organ systems. Weak expression of this gene does occur at a certain frequency in sporadic breast cancer. This indicates that LKB1 gene may relate to the tumorigenesis of breast cancer. EXPERIMENTAL DESIGN: To investigate the function of the LKB1 gene in sporadic breast cancer, we reintroduced LKB1 into breast cancer cell lines which lack the LKB1 gene. Also, we examined the LKB1 protein expression in human breast cancer samples. RESULTS: We found that reintroducing LKB1 into breast cancer cell lines suppresses cell growth by G(1) cell cycle block. The LKB1-mediated G(1) cell cycle arrest is caused by up-regulation of the expression of p21(WAF1/CIP1) in breast cancer MDA-MB-435 cells. We also demonstrated that low LKB1 protein expression correlates with higher histological grade (P = 0.013), larger tumor size (P = 0.001), progesterone receptor status (P = 0.048), and presence of lymph node metastasis (P = 0.003). Furthermore, LKB1 low expression was associated with a higher relapse rate (P = 0.002) and a worse OS (P = 0.008). CONCLUSIONS: LKB1 plays a role in tumor suppressor function in human breast cancer. LKB1 expression may be a useful prognostic marker in human breast cancer.

BACKGROUND: The AMP-activated protein kinase (AMPK) cascade is a sensor of cellular energy charge that acts as a metabolic master switch and inhibits cell proliferation. Activation requires phosphorylation of Thr172 of AMPK within the activation loop by upstream kinases (AMPKKs) that have not been identified. Recently, we identified three related protein kinases acting upstream of the yeast homolog of AMPK. Although they do not have obvious mammalian homologs, they are related to LKB1, a tumor suppressor that is mutated in the human Peutz-Jeghers cancer syndrome. We recently showed that LKB1 exists as a complex with two accessory subunits, STRAD alpha/beta and MO25 alpha/beta. RESULTS: We report the following observations. First, two AMPKK activities purified from rat liver contain LKB1, STRAD alpha and MO25 alpha, and can be immunoprecipitated using anti-LKB1 antibodies. Second, both endogenous and recombinant complexes of LKB1, STRAD alpha/beta and MO25 alpha/beta activate AMPK via phosphorylation of Thr172. Third, catalytically active LKB1, STRAD alpha or STRAD beta and MO25 alpha or MO25 beta are required for full activity. Fourth, the AMPK-activating drugs AICA riboside and phenformin do not activate AMPK in HeLa cells (which lack LKB1), but activation can be restored by stably expressing wild-type, but not catalytically inactive, LKB1. Fifth, AICA riboside and phenformin fail to activate AMPK in immortalized fibroblasts from LKB1-knockout mouse embryos. CONCLUSIONS: These results provide the first description of a physiological substrate for the LKB1 tumor suppressor and suggest that it functions as an upstream regulator of AMPK. Our findings indicate that the tumors in Peutz-Jeghers syndrome could result from deficient activation of AMPK as a consequence of LKB1 inactivation.

AMP-activated protein kinase (AMPK) is a highly conserved sensor of cellular energy status found in all eukaryotic cells. AMPK is activated by stimuli that increase the cellular AMP/ATP ratio. Essential to activation of AMPK is its phosphorylation at Thr-172 by an upstream kinase, AMPKK, whose identity in mammalian cells has remained elusive. Here we present biochemical and genetic evidence indicating that the LKB1 serine/threonine kinase, the gene inactivated in the Peutz-Jeghers familial cancer syndrome, is the dominant regulator of AMPK activation in several mammalian cell types. We show that LKB1 directly phosphorylates Thr-172 of AMPKalpha in vitro and activates its kinase activity. LKB1-deficient murine embryonic fibroblasts show nearly complete loss of Thr-172 phosphorylation and downstream AMPK signaling in response to a variety of stimuli that activate AMPK. Reintroduction of WT, but not kinase-dead, LKB1 into these cells restores AMPK activity. Furthermore, we show that LKB1 plays a biologically significant role in this pathway, because LKB1-deficient cells are hypersensitive to apoptosis induced by energy stress. On the basis of these results, we propose a model to explain the apparent paradox that LKB1 is a tumor suppressor, yet cells lacking LKB1 are resistant to cell transformation by conventional oncogenes and are sensitive to killing in response to agents that elevate AMP. The role of LKB1/AMPK in the survival of a subset of genetically defined tumor cells may provide opportunities for cancer therapeutics.

The LKB1 (also called serine/threonine kinase 11) tumor suppressor gene was cloned in 1998 by linkage analysis of Peutz-Jeghers cancer syndrome patients. Mammalian LKB1 has been implicated as a regulator of multiple biological processes and signaling pathways, including the control of cell-cycle arrest, p53-mediated apoptosis, Wnt signaling, transforming growth factor (TGF)-beta signaling, ras-induced cell transformation, and energy metabolism. The Caenorhabditis elegans and Drosophila melanogaster LKB1 homologs, termed PAR4 and dLKB1, respectively, regulate cell polarity. Recently, mammalian LKB1 was found to be active only in a complex with two other proteins--STRAD and MO25--and to induce complete polarization of intestinal epithelial cells in a cell-autonomous fashion. In this article, we summarize the findings regarding LKB1 over the past six years. In addition, we discuss LKB1 in polarity in the context of both the other PAR proteins and its tumor suppressive activities.

Tuberous sclerosis complex (TSC) and Peutz-Jeghers syndrome (PJS) are dominantly inherited benign tumor syndromes that share striking histopathological similarities. Here we show that LKB1, the gene mutated in PJS, acts as a tumor suppressor by activating TSC2, the gene mutated in TSC. Like TSC2, LKB1 inhibits the phosphorylation of the key translational regulators S6K and 4EBP1. Furthermore, we show that LKB1 activates TSC2 through the AMP-dependent protein kinase (AMPK), indicating that LKB1 plays a role in cell growth regulation in response to cellular energy levels. Our results suggest that PJS and other benign tumor syndromes could be caused by dysregulation of the TSC2/mTOR pathway.

The Peutz-Jegher syndrome tumor-suppressor gene encodes a protein-threonine kinase, LKB1, which phosphorylates and activates AMPK [adenosine monophosphate (AMP)-activated protein kinase]. The deletion of LKB1 in the liver of adult mice resulted in a nearly complete loss of AMPK activity. Loss of LKB1 function resulted in hyperglycemia with increased gluconeogenic and lipogenic gene expression. In LKB1-deficient livers, TORC2, a transcriptional coactivator of CREB (cAMP response element-binding protein), was dephosphorylated and entered the nucleus, driving the expression of peroxisome proliferator-activated receptor-gamma coactivator 1alpha (PGC-1alpha), which in turn drives gluconeogenesis. Adenoviral small hairpin RNA (shRNA) for TORC2 reduced PGC-1alpha expression and normalized blood glucose levels in mice with deleted liver LKB1, indicating that TORC2 is a critical target of LKB1/AMPK signals in the regulation of gluconeogenesis. Finally, we show that metformin, one of the most widely prescribed type 2 diabetes therapeutics, requires LKB1 in the liver to lower blood glucose levels.

LKB1, a unique serine/threonine kinase tumor suppressor, modulates anabolic and catabolic homeostasis, cell proliferation, and organ polarity. Chemically reactive lipids, e.g. cyclopentenone prostaglandins, formed a covalent adduct with LKB1 in MCF-7 and RKO cells. Site-directed mutagenesis implicated Cys210 in the LKB1 activation loop as the residue modified. Notably, ERK, JNK, and AKT serine/threonine kinases with leucine or methionine, instead of cysteine, in their activation loop did not form a covalent lipid adduct. 4-Hydroxy-2-nonenal, 4-oxo-2-nonenal, and cyclopentenone prostaglandin A and J, which all contain alpha,beta-unsaturated carbonyls, inhibited the AMP-kinase kinase activity of cellular LKB1. In turn, this attenuated signals throughout the LKB1 --> AMP kinase pathway and disrupted its restraint of ribosomal S6 kinases. The electrophilic beta-carbon in these lipids appears to be critical for inhibition because unreactive lipids, e.g. PGB1, PGE2, PGF2alpha, and TxB2, did not inhibit LKB1 activity (p > 0.05). Ectopic expression of cyclooxygenase-2 and endogenous biosynthesis of eicosanoids also inhibited LKB1 activity in MCF-7 cells. Our results suggested a molecular mechanism whereby chronic inflammation or oxidative stress may confer risk for hypertrophic or neoplastic diseases. Moreover, chemical inactivation of LKB1 may interfere with its physiological antagonism of signals from growth factors, insulin, and oncogenes.

The LKB1/STK11 serine/threonine kinase is mutated in Peutz-Jeghers syndrome and various sporadic cancers such as lung adenocarcinoma. We show here that LKB1 forms a complex with LMO4, GATA-6, and Ldb1, and enhances GATA-mediated transactivation in a kinase-dependent manner. We further demonstrate that LKB1 has the potential to induce p21 expression in collaboration with LMO4, GATA-6, and Ldb1 through the p53-independent mechanism. Our findings suggest that LKB1 regulates GATA-mediated gene expression and that this activity of LKB1 may be important for its tumor suppressor function.

Inactivating germline mutations in the LKB1 gene underlie Peutz-Jeghers syndrome characterized by hamartomatous polyps and an elevated risk for cancer. Recent studies suggest the involvement of LKB1 also in more common human disorders including diabetes and in a significant fraction of lung adenocarcinomas. These observations have increased the interest towards signaling pathways of this tumor suppressor kinase. The recent breakthroughs in understanding the molecular functions of the LKB1 indicate its contribution as a regulator of cell polarity, energy metabolism and cell proliferation. Here we review how the substrates and cellular functions of LKB1 may be linked to Peutz-Jeghers syndrome and other diseases, and discuss how some of the molecular changes associated with altered LKB1 signaling might be used in therapeutic approaches.

Aberrant phosphorylation of tau is associated with a number of neurodegenerative diseases, including Alzheimers disease (AD). The molecular mechanisms by which tau phosphorylation is regulated under normal and disease conditions are not well understood. Microtubule affinity regulating kinase (MARK) and PAR-1 have been identified as physiological tau kinases, and aberrant phosphorylation of MARK/PAR-1 target sites in tau has been observed in AD patients and animal models. Here we show that phosphorylation of PAR-1 by the tumor suppressor protein LKB1 is required for PAR-1 activation, which in turn promotes tau phosphorylation in Drosophila. Diverse stress stimuli, such as high osmolarity and overexpression of the human beta-amyloid precursor protein, can promote PAR-1 activation and tau phosphorylation in an LKB1-dependent manner. These results reveal a new function for the tumor suppressor protein LKB1 in a signaling cascade through which the phosphorylation and function of tau is regulated by diverse signals under physiological and pathological conditions.

The tumor suppressor LKB1 (STK11) is a cytoplasmic/nuclear serine/threonine kinase, defects in which cause Peutz-Jeghers syndrome (PJS) in humans and animals. Recent studies showed that loss of function of LKB1 is associated with sporadic forms of lung, pancreatic, and ovarian cancer. In cancer cells, LKB1 is inactivated by two mechanisms: mutations in its central kinase domain or complete loss of LKB1 expression. Inactivation of LKB1 is associated with progression of PJS and transformation of benign polyps into malignant tumors. This study examines the effect of LKB1 on regulation of STAT3 and expression of transcriptional targets of STAT3. The results show that LKB1 inhibits rearranged in transformation (RET)/papillary thyroid carcinoma (PTC)-dependent activation of signal transducer and activator of transcription 3 (STAT3), which is mediated by phosphorylation of STAT3 tyrosine 705 by RET/PTC. suppression of STAT3 transactivation by LKB1 requires the kinase domain but not the kinase activity of LKB1. The centrally located kinase domain of LKB1 is an approximately 260-amino-acid region that binds to the linker domain of STAT3. Chromatin immunoprecipitation studies indicate that expression of LKB1 reduces the binding of STAT3 to its target promoters and suppresses STAT3-mediated expression of Cyclin D1, VEGF, and Bcl-xL. Knockdown of LKB1 by specific small interfering RNA led to an increase in STAT3 transactivation activity and promoted cell proliferation in the presence of RET/PTC. Thus, this study suggests that LKB1 suppresses tumor growth by inhibiting RET/PTC-dependent activation of oncogenic STAT3.

The human LKB1 tumor suppressor has been implicated as an important regulator of many cellular processes and signaling pathways, indicating that it could be a good candidate for anticancer drugs. The failure of its obtain high-level expression has been a major obstacle to study its protein structure and function in vitro. Here, we describe the high-level expression of human LKB1 in Escherichia coli and show its kinase activity and anticancer effects on a tumor cell line. The gene encoding LKB1 was optimized by replacing rare codons with codons frequently used in E. coli and synthesized with overlapping primers. The recombinant His-LKB1 was expressed in hosts BL21(DE3) (BL) and Rosetta-gami(DE3)pLysS (RG). His-LKB1 from BL was present mainly as inclusion body. The soluble His-LKB1 from RG accounted for 34.1% of total proteins and the yield of purified His-LKB1 was approximately 92 microg/ml. Purified His-LKB1 protein from both hosts was functionally active, as shown by reversible autophosphorylation and kinase activity in the absence of any other associated kinase. The growth inhibitory ratio of the purified BL-derived and RG-derived His-LKB1 on hepatic carcinoma SMMC-7721 cells was 24.97% and 45.68%, respectively, and both could produce significant cell-cycle arrest.

The tumor suppressor LKB1 is mutated in 30% of non-small cell lung cancer (NSCLC) tumors and cell lines and is proposed to be a key regulator of epithelial cell polarity; however, how LKB1 regulates cancer cell polarity is not known. The experiments described herein show for the first time that LKB1 is a dynamic, actin-associated protein that rapidly polarizes to the leading edge of motile cancer cells. LKB1 proves to be essential for NSCLC polarity, because LKB1 depletion results in classic cell polarity defects, such as aberrant Golgi positioning, reduced lamellipodia formation, and aberrant morphology. To probe how LKB1 regulates these events, we show that LKB1 colocalizes at the cellular leading edge with two key components of the polarity pathway - the small rho GTPase cdc42 and its downstream binding partner p21-activated kinase (PAK). Importantly, LKB1 functionality is required for cdc42 polarization to the leading edge, maintaining active cdc42 levels, and downstream PAK phosphorylation. To do this, LKB1 interacts only with active form of cdc42 and PAK, but not with inactive cdc42. Taken together, these results show that LKB1 is a critical mediator of the NSCLC polarity program in lung cancer cells through a novel LKB1-cdc42-PAK pathway.

mutation of LKB1 is the key molecular event underlying Peutz-Jeghers syndrome, a dominantly inherited condition characterized by a predisposition to a range of malignancies, including those of the reproductive system. We report here the use of a Cre-LoxP strategy to directly address the role of Lkb1 in prostate neoplasia. Recombination of a LoxP-flanked Lkb1 allele within all four murine prostate lobes was mediated by spontaneous activation of a p450 CYP1A1-driven Cre recombinase transgene (termed AhCre). Homozygous mutation of Lkb1 in males expressing AhCre reduced longevity, with 100% manifesting atypical hyperplasia and 83% developing prostate intraepithelial neoplasia (PIN) of the anterior prostate within 2 to 4 months. We also observed focal hyperplasia of the dorsolateral and ventral lobes (61% and 56% incidence, respectively), bulbourethral gland cysts associated with atypical hyperplasia (100% incidence), hyperplasia of the urethra (39% incidence), and seminal vesicle squamous metaplasia (11% incidence). PIN foci overexpressed nuclear beta-catenin, p-Gsk3 beta, and downstream Wnt targets. Immunohistochemical analysis of foci also showed a reduction in Pten activation and up-regulation of both p-PDK1 (an AMPK kinase) and phosphorylated Akt. Our data are therefore consistent with deregulation of Wnt and phosphoinositide 3-kinase/Akt signaling cascades after loss of Lkb1 function. For the first time, this model establishes a link between the tumor suppressor Lkb1 and prostate neoplasia, highlighting a tumor suppressive role within the mouse and raising the possibility of a similar association in the human.

Germ line mutations in the LKB1 tumor suppressor gene are associated with the Peutz-Jeghers polyposis and cancer syndrome. Somatic mutations in Lkb1 are observed in sporadic pulmonary, pancreatic and biliary cancers and melanomas. The LKB1 serine-threonine kinase functionally and biochemically links control of cellular structure and energy utilization through activation of the AMPK family of kinases. Lkb1 regulates cell polarity through downstream kinases including AMPKs, MARKs and BRSKs, and nutrient utilization and cellular metabolism through the AMPK-mTOR pathway. LKB1 has been shown to affect normal chromosomal segregation, TGF-beta signaling in the mesenchyme and WNT and p53 activity. Although each of the LKB1-dependent processes and downstream pathways have been individually delineated through work across a range of experimental systems, how they relate to Lkb1s role as a tumor suppressor remains to be fully explored and elucidated. The recent development of mouse cancer models harboring engineered mutations in Lkb1 have offered insights into how LKB1 may be functioning to restrain tumorigenesis and how its role as a master regulator of polarity and metabolism could contribute to its tumor suppressor function.

Germline mutations of the LKB1 tumor suppressor gene result in Peutz-Jeghers syndrome (PJS) characterized by intestinal hamartomas and increased incidence of epithelial cancers. Inactivating mutations in LKB1 have also been found in certain sporadic human cancers and with particularly high frequency in lung cancer. LKB1 has now been demonstrated to play a crucial role in pulmonary tumorigenesis, controlling initiation, differentiation, and metastasis. Recent evidences showed that LKB1 is a multitasking kinase, with great potential in orchestrating cell activity. Thus far, LKB1 has been found to play a role in cell polarity, energy metabolism, apoptosis, cell cycle arrest, and cell proliferation, all of which may require the tumor suppressor function of this kinase and/or its catalytic activity. This review focuses on remarkable recent findings concerning the molecular mechanism by which the LKB1 protein kinase operates as a tumor suppressor and discusses the rational treatment strategies to individuals suffering from PJS and other common disorders related to LKB1 signaling.

Human Papilloma Virus (HPV) is the etiologic agent for cervical cancer. Yet, infection with HPV is not sufficient to cause cervical cancer, because most infected women develop transient epithelial dysplasias that spontaneously regress. Progression to invasive cancer has been attributed to diverse host factors such as immune or hormonal status, as no recurrent genetic alterations have been identified in cervical cancers. Thus, the pressing question as to the biological basis of cervical cancer progression has remained unresolved, hampering the development of novel therapies and prognostic tests. Here we show that at least 20% of cervical cancers harbor somatically-acquired mutations in the LKB1 tumor suppressor. Approximately one-half of tumors with mutations harbored single nucleotide substitutions or microdeletions identifiable by exon sequencing, while the other half harbored larger monoallelic or biallelic deletions detectable by multiplex ligation probe amplification (MLPA). Biallelic mutations were identified in most cervical cancer cell lines; HeLa, the first human cell line, harbors a homozygous 25 kb deletion that occurred in vivo. LKB1 inactivation in primary tumors was associated with accelerated disease progression. Median survival was only 13 months for patients with LKB1-deficient tumors, but >100 months for patients with LKB1-wild type tumors (P = 0.015, log rank test; hazard ratio = 0.25, 95% CI = 0.083 to 0.77). LKB1 is thus a major cervical tumor suppressor, demonstrating that acquired genetic alterations drive progression of HPV-induced dysplasias to invasive, lethal cancers. Furthermore, LKB1 status can be exploited clinically to predict disease recurrence.

The tumor suppressor serine-threonine kinase LKB1 is mutated in Peutz-Jeghers syndrome (PJS) and in epithelial cancers, including hormone-sensitive organs such as breast, ovaries, testes, and prostate. Clinical studies in breast cancer patients show low LKB1 expression is related to poor prognosis, whereas in PJS, the risk of breast cancer is similar to the risk from germline mutations in breast cancer (BRCA) 1/BRCA2. In this study, we investigate the role of LKB1 in estrogen receptor alpha (ERalpha) signaling. We demonstrate for the first time that LKB1 binds to ERalpha in the cell nucleus in which it is recruited to the promoter of ERalpha-responsive genes. Furthermore, LKB1 catalytic activity enhances ERalpha transactivation compared with LKB1 catalytically deficient mutants. The significance of our discovery is that we demonstrate for the first time a novel functional link between LKB1 and ERalpha. Our discovery places LKB1 in a coactivator role for ERalpha signaling, broadening the scientific scope of this tumor suppressor kinase and laying the groundwork for the use of LKB1 as a target for the development of new therapies against breast cancer.

The LKB1 serine-threonine kinase is a tumor suppressor that is inactivated in a large number of sporadic human lung non-small cell carcinomas (NSCLCs) and cervical cancers. genetic deletion of LKB1 in various mouse tissues results in tumorigenesis, and loss of LKB1 increases metastasis in a mouse model of NSCLC. LKB1 directly activates a family of 14 kinases related to AMPK [adenosine monophosphate (AMP)-activated protein kinase] to control cell metabolism, growth, and polarity, though which of these are critical to its tumor suppressor functions remain undefined. The LKB1-dependent kinase SIK1 (salt-inducible kinase 1) has now been identified as a key modulator of anoikis (apoptosis induced by cell detachment) and transformation in culture, and its modulation of the tumor suppressor p53 controls metastasis in transplanted tumor cells. Reduced SIK1 expression is correlated with poor prognosis in two large human breast cancer data sets. These findings suggest that SIK1 is a key upstream regulator of p53-dependent anoikis that may be targeted in tumorigenesis.

Adiponectin is an adipocytokine involved in the pathogenesis of various obesity-related disorders. Also, it has been shown that adiponectin has therapeutic potential for metabolic syndrome, systemic insulin resistance, cardiovascular disease and more recently carcinogenesis. Adiponectin can modulate breast cancer cell growth and proliferation. Anti-metastatic effects of adiponectin have also been elucidated. It has been shown that adiponectin inhibits important metastatic properties such as adhesion, invasion and migration of breast cancer cells. Examination of the underlying molecular mechanisms has shown that adiponectin treatment increases AMP-activated protein kinase (AMPK) phosphorylation and activity. Adiponectin also increases phosphorylation of downstream target of AMPK, Acetyl-CoA Carboxylase (ACC) and decreases phosphorylation of p70S6 kinase (S6K). Importantly, adiponectin treatment increases the expression of tumor suppressor gene, LKB1 in breast cancer cells. LKB1 is required for adiponectin-mediated modulation of AMPK-S6K axis and more importantly, its biological functions including inhibition of adhesion, migration and invasion of breast cancer cells. Although further studies are required to analyze the effect of adiponectin on LKB1-AMPK-S6K axis, these data present a novel mechanism involving specific upregulation of tumor suppressor gene LKB1 by which adiponectin inhibits adhesion, invasion and migration of breast cancer cells. These results highlight a new role for LKB1 in adiponectin action and may have significant implication for development of novel therapeutic options. cancer research has largely focused on the molecular basis of oncogenic transformation and tumorigenesis for many years. Recent progress in cancer research has put the metastatic process at the center stage because higher metastatic potential of tumor cells is the major cause of mortality from solid tumors. Metastasis is a complex process that involves modulation of various molecular signaling networks. Tumor cells alter the microenvironment, attain greater cellular adhesion along with better ability to invade and migrate to gain access to circulation. These wandering tumor cells defy anoikis, survive in the circulation, exit into new permissive organ site and colonize distant organs. The microenvironment in which the tumor originates plays an important role in tumor initiation, progression and metastasis.

Finding an effective treatment for acute myeloid leukemia (AML) remains a challenge, and all cellular processes that are deregulated in AML cells should be considered in the design of targeted therapies. We show in our current study that the LKB1/AMPK/TSC tumor suppressor axis is functional in AML and can be activated by the biguanide molecule metformin, resulting in a specific inhibition of mammalian target of rapamycin (mTOR) catalytic activity. This induces a multisite dephosphorylation of the key translation regulator, 4E-BP1, which markedly inhibits the initiation step of mRNA translation. Consequently, metformin reduces the recruitment of mRNA molecules encoding oncogenic proteins to the polysomes, resulting in a strong antileukemic activity against primary AML cells while sparing normal hematopoiesis ex vivo and significantly reducing the growth of AML cells in nude mice. The induction of the LKB1/AMPK tumor-suppressor pathway thus represents a promising new strategy for AML therapy.

BACKGROUND: Germline mutations in LKB1 result in Peutz-Jeghers Syndrome characterized by intestinal hamartomas and increased incidence of epithelial cancers. LKB1 encodes a serine/threonine kinase that plays an important role in regulating energy metabolism through the AMPK/mTOR signaling pathway. In addition, LKB1 is homologous to PAR-4, a polarity protein first described in C. elegans, while activation of LKB1 in mammalian epithelial cells induces the polarized assembly of actin filaments. RESULTS: To explore the mechanism by which LKB1 interacts with the actin cytoskeleton, we introduced LKB1 into HeLa cells that lack endogenous LKB1. This results in activation of the small GTPase Rho and the assembly of linear actin filaments associated with focal adhesions. These effects on the actin cytoskeleton are attenuated by siRNA-mediated depletion of the guanine nucleotide exchange factor Dbl. Co-expression of the LKB1 with the adaptor protein STRAD induces actin filament puncta associated with phospho-ezrin. CONCLUSIONS: This study reveals that LKB1 regulates the actin cytoskeleton through a Dbl/Rho pathway.

Hypothesis-generating epidemiological research has suggested that cancer burden is reduced in diabetics treated with metformin and experimental work has raised questions regarding the role of direct adenosine monophosphate-activated protein kinase (AMPK)-mediated anti-neoplastic effects of metformin as compared with indirect effects attributable to reductions in circulating insulin levels in the host. We treated both tumor LKB1 expression and host diet as variables, and observed that metformin inhibited tumor growth and reduced insulin receptor activation in tumors of mice with diet-induced hyperinsulinemia, independent of tumor LKB1 expression. In the absence of hyperinsulinemia, metformin inhibited only the growth of tumors transfected with short hairpin RNA against LKB1, a finding attributable neither to an effect on host insulin level nor to activation of AMPK within the tumor. Further investigation in vitro showed that cells with reduced LKB1 expression are more sensitive to metformin-induced adenosine triphosphate depletion owing to impaired ability to activate LKB1-AMPK-dependent energy-conservation mechanisms. Thus, loss of function of LKB1 can accelerate proliferation in contexts where it functions as a tumor suppressor, but can also sensitize cells to metformin. These findings predict that any clinical utility of metformin or similar compounds in oncology will be restricted to subpopulations defined by host insulin levels and/or loss of function of LKB1.

Histone deacetylases 4 (HDAC4), -5, -7, and -9 form class IIa within the HDAC superfamily and regulate diverse physiological and pathological cellular programs. With conserved motifs for phosphorylation-dependent 14-3-3 binding, these deacetylases serve as novel signal transducers that are able to modulate histone acetylation and gene expression in response to extracellular cues. Here, we report that in a PKA-sensitive manner the tumor suppressor kinase LKB1 acts through salt-inducible kinase 2 (SIK2) and SIK3 to promote nucleocytoplasmic trafficking of class IIa HDACs. Both SIK2 and SIK3 phosphorylate the deacetylases at the conserved motifs and stimulate 14-3-3 binding. SIK2 activates MEF2-dependent transcription and relieves repression of myogenesis by the deacetylases. Distinct from SIK2, SIK3 induces nuclear export of the deacetylases independent of kinase activity and 14-3-3 binding. These findings highlight the difference among members of the SIK family and indicate that LKB1-dependent SIK activation constitutes an important signaling module upstream from class IIa deacetylases for regulating cellular programs controlled by MEF2 and other transcription factors.