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AMP-Activated Protein Kinase: the Current Landscape for Drug Development

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AMP-Activated Protein Kinase: the Current Landscape for Drug Development REVIEWS AMP-activated protein kinase: the current landscape for drug development Gregory R. Steinberg 1* and David Carling2 Abstract | Since the discovery of AMP-activated protein kinase (AMPK) as a central regulator of energy homeostasis, many exciting insights into its structure, regulation and physiological roles have been revealed. While exercise, caloric restriction, metformin and many natural products increase AMPK activity and exert a multitude of health benefits, developing direct activators of AMPK to elicit beneficial effects has been challenging. However, in recent years, direct AMPK activators have been identified and tested in preclinical models, and a small number have entered clinical trials. Despite these advances, which disease(s) represent the best indications for therapeutic AMPK activation and the long-term safety of such approaches remain to be established. Cardiovascular disease Dramatic improvements in health care coupled with identifying a unifying mechanism that could link these (CVD). A term encompassing an increased standard of living, including better nutri- changes to multiple branches of metabolism followed diseases affecting the heart tion and education, have led to a remarkable increase in the discovery that the AMP-activated protein kinase or circulatory system. human lifespan1. Importantly, the number of years spent (AMPK) provided a common regulatory mechanism in good health is also increasing2. Despite these positive for inhibiting both cholesterol (through phosphoryla- Non-alcoholic fatty liver disease developments, there are substantial risks that challenge tion of HMG-CoA reductase (HMGR)) and fatty acid (NAFLD). A very common continued improvements in human health. Perhaps the (through phosphorylation of acetyl-CoA carboxylase disease in humans in which greatest threat to future health is a chronic energy imbal- (ACC)) synthesis8 (BOx 1). there is an excessive ance in which intake exceeds expenditure3. This energy Over the last 30 years since formally naming AMPK9, accumulation of fat in the liver (steatosis) in individuals who imbalance contributes to a global epidemic of overweight interest in the enzyme as a drug target has continued to 4 are not alcoholic. and obesity that accelerates ageing and also increases the grow. This interest has been stimulated in part by the risk of developing type 2 diabetes, cardiovascular disease discovery that AMPK is activated by physiological reg- (CVD), non-alcoholic fatty liver disease (NAFLD), chronic ulators that are associated with health and longevity such kidney disease and certain cancers5. And although life- as caloric restriction10 and exercise11, hormones such as style interventions that target this energy imbalance leptin12 and adiponectin13 and many natural products through caloric restriction and/or endurance exercise can that have been used as traditional herbal medicines REF.14 1Centre for Metabolism, reduce ageing, obesity and related diseases, implement- (reviewed in ). Many of these herbal medicines such 15 15–17 Obesity and Diabetes ing and maintaining these changes for prolonged periods as resveratrol and berberine are being reformulated Research, Department of is challenging given the pervasiveness of calorically dense to enhance their bioavailability and efficacy. Two of the Medicine and Department foods and sedentary lifestyle-enabling technologies. most widely used medicines in the world, metformin18, of Biochemistry and Therefore, developing new pharmacological strategies the mostly commonly used drug for type 2 diabetes, Biomedical Sciences, 19 McMaster University, that mimic a low energy state, such as that elicited by and salicylate , the active ingredient in aspirin and sal- Hamilton, Ontario, Canada. exercise and caloric restriction, could be potentially salate, also activate AMPK, suggesting that some of their 2Cellular Stress Group, beneficial for treating chronic metabolic diseases. beneficial effects may be mediated in part by AMPK. Medical Research Council At the cellular level, the balance between energy The sodium/glucose cotransporter 2 (SGLT2) inhibi- London Institute of Medical intake and demand can be inferred by the relative levels tor canagliflozin, which has recently been approved for Sciences, Hammersmith of the adenine nucleotides AMP, ADP and ATP that are treating type 2 diabetes, also indirectly activates AMPK Hospital, Imperial College, 20,21 London, UK. continuously produced and consumed through numer- in a manner similar to metformin . More recently, a 6 *e-mail: gsteinberg@ ous metabolic reactions . Given their central role as a number of small molecules have been identified that mcmaster.ca measure of cellular energy balance, a number of enzymes directly activate AMPK. Most of these compounds bind https://doi.org/10.1038/ were known for many years to be regulated by alter- in a specific pocket formed by an interaction between s41573-019-0019-2 ations in adenine nucleotides7. However, a pivotal step in the α and β subunits22, termed the allosteric drug and NATURE REVIEWS | DRUG DISCOVERY VOLUME 18 | JULY 2019 | 527 REVIEWS Box 1 | Discovery of AMPK kinases and plays a pivotal role in their regulation. In AMPK, phosphorylation of Thr172 (pThr172) within Although the first citation of AMP-activated protein kinase (AMPK) was published in the activation loop is required for maximal activity of 9 1988 (REF. ), papers describing the activity of AMPK can be traced back to work AMPK29,30. Two upstream kinases account for the phys- stemming from the early 1970s. Studies investigating the regulation of HMG-CoA iological phosphorylation of Thr172: liver kinase B1 reductase identified a cytosolic factor that inactivated HMGR in the presence of ATP 31–33 and ADP330. Subsequent work identified this factor as a protein kinase and found that (LKB1) and calcium/calmodulin-dependent pro- AMP was a much more potent allosteric activator than ADP331. Other studies reported tein kinase kinase 2 (CAMKK2; sometimes referred to 34–36 an HMGR kinase that was itself regulated by phosphorylation, forming a protein kinase as CAMKKβ) . Numerous studies have shown that cascade72,332,333, although it was not clear whether these activities were catalysed by the Thr172 phosphorylation increases AMPK activity both same enzyme. Around the same time as these studies, acetyl-CoA carboxylase (ACC) in vitro and in vivo, and regulation of Thr172 phos- was shown to be inactivated by a protein kinase that was activated by AMP334. With the phorylation is a central component of the mechanism benefit of hindsight, it seems surprising that there was no suggestion that this enzyme of activation of AMPK by small molecules (see below). might be related to the HMGR kinase activities reported previously. Several years later The kinase domain is followed by a flexible region, in 1987, however, it was reported that the same bicyclic protein kinase cascade termed the α-linker37. Within the α-linker is a region inactivates both ACC and HMGR. The evidence for this claim was based on several of ~60 amino acids that has been named the auto- observations. First, the ACC and HMGR kinase activities co-purified following isolation from rat liver through six purification steps335. Second, the phosphorylation and inhibitory domain (AID), so named because addition inactivation of both ACC and HMGR were stimulated equivalently by AMP, and third, of this sequence to the isolated kinase domain reduces 38–40 protein phosphatase treatment of the partially purified protein kinase preparation AMPK activity by approximately tenfold . Closer to reduced the ACC and HMGR kinase activities by the same extent335. In addition to the carboxy-terminus than the AID is a short sequence clarifying the field and providing the rationale for assigning the name for AMPK, the that has been termed the α-hook37, or α-regulatory sub- finding that the same protein kinase could inactivate key regulatory enzymes in unit interacting motif 2 (α-RIM2)41,42, which plays an the biosynthesis of both fatty acids and cholesterol provided the first clue that AMPK important role in the nucleotide-dependent regulation has an important role in metabolism. of AMPK. Another region within the α subunit that The first study to formally reference AMPK described the identification of a protein may play a regulatory role is the serine/threonine-rich kinase isolated from rat liver that phosphorylated and inactivated ACC9. The kinase was loop43. This region is not conserved in all eukary- named AMPK because it was allosterically activated by AMP8. An important finding in that original study was that AMPK phosphorylated ACC at a different site than did otic species and so might represent a somewhat late 43 cAMP-dependent protein kinase A (PKA)9, which led to two major developments that evolutionary adaptation . provided essential tools to study AMPK. The first, based on the AMPK phosphorylation The β subunit isoforms undergo constitutive amino- site of ACC at Ser79, was the development of a specific peptide substrate (the SAMS terminal myristoylation, and this modification is peptide) that could be used to measure AMPK activity336. The second was the generation required for AMP-stimulated and ADP-stimulated phos- of phospho-specific antibodies to ACC (ACC1 Ser79 and ACC2 Ser221)309, which phorylation of Thr172 (REFS44,45). Within the β subunit appears to be unique for AMPK337, providing a convenient and robust surrogate for is a region termed the carbohydrate binding module monitoring the cellular activity
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