Can J Diabetes 37 (2013) 319e326
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Canadian Journal of Diabetes journal homepage: www.canadianjournalofdiabetes.com
Review
Diabetic Dyslipidemia: From Evolving Pathophysiological Insight to Emerging Therapeutic Targets Dominic S. Ng PhD, MD, FRCPC * Li Ka Shing Knowledge Institute, Keenan Research Center, St. Michael’s Hospital, University of Toronto, Toronto, Ontario, Canada
a r t i c l e i n f o
a b s t r a c t
Article history: Received 26 May 2013 Received in revised form 22 July 2013 Accepted 22 July 2013
Diabetic dyslipidemia is characterized by hepatic very low density lipoprotein (VLDL) and intestinal chylomicron overproduction, reduced high density lipoprotein cholesterol (HDL-C) level, increased propensity of small dense LDL (sdLDL) and increased postprandial lipemia. This dyslipidemic profile is also strongly linked to other features of the metabolic syndrome. Diabetic dyslipidemia is a well-recognized risk factor for atherosclerotic cardiovascular diseases. Currently, statins remain the first line therapy primarily through reducing the atherogenic LDL. Clinical trials on other lipid modifying agents were met with variable success in selective patient populations. Emerging new insights into the pathophysiology of lipid metabolism, in general, and diabetic dyslipidemia, in particular, have opened up potentially novel therapeutic strategies to further reduce the risk associated with diabetic dyslipidemia and insulin resistant state. Ó 2013 Canadian Diabetes Association
Keywords: cardiovascular diseases cholesterol efflux diabetes dyslipidemia ER stress HDL insulin resistance triglycerides Mots clés : maladies cardiovasculaires efflux du cholestérol diabète dyslipidémie stress du RE HDL insulinorésistance triglycérides
r é s u m é La dyslipidémie du diabétique est caractérisée par la surproduction hépatique de lipoprotéines de très faible densité (VLDL : very low density lipoprotein) et la surproduction intestinale de chylomicrons, la réduction de la concentration du cholestérol à lipoprotéines de haute densité (cholestérol HDL), l’augmentation de la propension des LDL petites et denses, et l’augmentation de la lipémie postprandiale. Ce profil dyslipidémique est aussi fortement lié à d’autres caractéristiques du syndrome métabolique. La dyslipidémie du diabétique est un facteur de risque bien connu des maladies cardiovasculaires athérosclérotiques. Actuellement, les statines semblent être le traitement de première intention, principalement pour la réduction des LDL athérogènes. Les essais cliniques sur d’autres agents modifiant les lipides ont obtenu un succès inégal chez certaines populations de patients. Les perspectives émergentes en physiopathologie du métabolisme des lipides, en général, et la dyslipidémie du diabétique, en particulier, ont possiblement amené les nouvelles stratégies thérapeutiques à réduire davantage le risque associé à la dyslipidémie du diabétique et à l’état d’insulinorésistance. Ó 2013 Canadian Diabetes Association
Introduction Type II diabetes mellitus is a chronic metabolic disorder that predisposes an individual to be high risk for atherosclerotic cardiovascular diseases (CVD). This is particularly the case if the patient also carries clinical features of the metabolic syndrome, characterized by central obesity, hypertension, impaired fasting glucose, elevated triglyceride (TG) and reduced levels of high density lipoprotein cholesterol (HDL-C), with the latter 2 entities being clinical surrogates for diabetic dyslipidemia (1). Diabetic dyslipidemia in type II diabetes is characterized by overproduction
* Address for correspondence: Dominic S. Ng, PhD, MD, FRCPC, Associate Professor of Medicine, University of Toronto, Keenan Research Center, Li Ka Shing Knowledge Institute, St Michael’s Hospital, Shuter Wing Room 3-041, 30 Bond Street, Toronto, Ontario M5B 1W8, Canada. E-mail address: [email protected]. 1499-2671/$ e see front matter Ó 2013 Canadian Diabetes Association http://dx.doi.org/10.1016/j.jcjd.2013.07.062
of triglyceride-rich lipoproteins (TRL) both by the liver and the intestine, reduced plasma levels HDL-C and increased preponderance of small dense low density lipoproteins (sdLDL) (Figure) (2). This dyslipidemic profile is well recognized to be one of the proatherogenic factors in type II diabetic subjects; but the underlying pathophysiologic mechanisms remain inadequately understood and amelioration of this phenotype remains elusive. In type 1 diabetes subjects, presence of dyslipidemia, which includes elevated TG, LDL-C and low HDL-C, predicts future CVD, but the prevalence of dyslipidemia in type 1 diabetes subjects is not higher than their nondiabetic counterparts (3,4). Overproduction of triglyceride-rich lipoproteins in insulin-resistant state In the fasting state, the liver constitutively synthesizes apolipoprotein (apo) B100, an essential protein component of VLDL.
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Figure. Key pathways that may be altered in diabetic dyslipidemia. (1) Hepatic VLDL overproduction: increased TG availability, increased MTP activity, reduced apoB100 degradation, reduced LPL clearance. (2) Intestinal chylomicron overproduction: increased TG availability, increased MTP activity, reduced GLP-1 action, reduced LPL clearance. (3) Small dense LDL: increased CETP activity, increased TRL. (4) Reduced HDL-C: increased CETP activity, increased TRL, reduced ABCA1 level or activity; reduced apoAI synthesis. ABCA1, ATPase binding cassette A1; apoB100, apolipoprotein B100; CETP, cholesterol ester transfer protein; LPL, lipoprotein lipase; MTP, microsomal transfer protein; TRL, triglyceride rich lipoproteins; VLDL, Very low density lipoproteins.
Newly synthesized apoB100 is co-translationally lipidated by microsomal transfer protein (MTP) with TG and cholesterol ester delivered in the endoplasmic reticulum. Continuing lipidation of these primordial lipoproteins result in maturation of the VLDL particle which will be transported to the Golgi, the site of secretion of the mature VLDL. ApoB100 is constitutively synthesized and the excess, un-lipidated molecules are destined for degradation. Several groups have established that hepatic VLDL over-production is a cardinal feature of diabetic dyslipidemia based on kinetic studies (5,6). In diabetic states, several factors contribute mechanistically to the observed hepatic overproduction of VLDL. First, lipidation of apoB100 is dependent on the availability of triglyceride in the cytosol. Insulin is a strong inducer of sterol response element binding protein 1c (SREBP1c), which is the master transcription factor for promoting de novo lipogenesis, resulting in increased fatty acid as substrate for TG synthesis. In insulin resistant state, the activation of SREBP1c and its downstream lipogenic gene program, through mechanisms that remain incompletely understood (7,8), are accentuated differentially by hyperinsulinemia, resulting in increased fatty acid substrate availability for hepatic TG synthesis. Second, MTP, the central player in mediating the lipidation of apoB100, is commonly activated in a variety of mouse or rat models of diabetes or insulin resistance although
the mechanism remains to be fully elucidated (9). Third, insulin resistant state per se has been shown to attenuate apoB100 degradation. Multiple mechanisms have been identified to mediate apoB100 degradation including authophagy (10). Intestinal lipoprotein overproduction in insulin-resistant state Chylomicrons are lipoproteins synthesized and secreted by intestinal enterocytes, as a response to dietary fat ingestion. In the intestinal cells, free fatty acids of dietary source are resynthesized into TG and are available to lipidate a truncated form of apoB, namely the apoB48, also mediated by MTP. Several lines of experimental evidence support a pro-atherogenic nature of chylomicron remnants (CMR) (11). Botham et al reported that CMR may be taken up directly by macrophages, promoting foam cell formation (12). A recent study reported the detection of apoB48 along with apoB100 proteins in atherosclerotic plaques from autopsy of subjects who suffered sudden cardiac death, implicating a pro-atherogenic role for the intestinally derived, apoB48-containing TRL (13). This finding is consistent with the pro-atherogenic phenotype seen in a mouse model of chylomicronemia (14). Similar to their hepatic counterparts, each chylomicron particle contains 1 molecule of apoB48. In a lipid-rich situation, chylomicrons
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such formed are highly buoyant with high content of lipids for each apoB48. Unlike their VLDL counterpart, degradation of apoB48 is considerably less of a regulatory factor, as the enterocytes can form and secrete more small dense chylomicrons. Thus lipid availability and MTP activity are considered as the major regulatory factors for chylomicron production. In patients with diabetes and subjects with insulin resistance, several lines of evidence strongly suggest that the intestine overproduces chylomicrons when compared to the healthy controls (15). Mechanistically, increased MTP mass and activity increased de novo lipogenesis, as well as increased apoB stability, have all been implicated. In addition to intracellular regulatory mechanisms, it has been shown that circulating free fatty acids also stimulate intestinal chylomicron production, although the underlying mechanism remains uncertain. Increased plasma free fatty acid level is commonly observed in subjects with insulin resistance and obesity, in part due to either impaired uptake by peripheral tissues upon lipolysis of circulating TRL or enhanced lipolysis in the dysfunctional white adipose tissues (16). Thus, a free fatty acid mediated intestinal lipoprotein overproduction is expected to be a common lipid phenotype in subjects with diabetes. Modulation of intestinal lipoprotein production by incretin hormones Glucagon-like peptide (GLP)-1 and GLP-2 are 2 peptides expressed by the proglucagon gene in the intestinal L cells. A major biological action of GLP-1 is its ability to lower blood glucose by stimulating insulin secretion and suppressing glucagon secretion in the islets. The secretion of GLP-1 from the L cells is triggered by the presence of nutrients in the intestine, and, therefore, plays a key role in glucose homeostasis. In light of the very short half-life of endogenously produced GLP-1, GLP-1 analogues, GLP-R agonists and the dipeptidylpeptidase-4 (DPP-IV) inhibitors, the latter through delaying the degradation of endogenous GLP-1 have been formulated and used widely for treatment of hyperglycemia in subjects with diabetes. In humans, short-term treatment with GLP-1 analogues or DPP-IV inhibitors result in not only improvement in glycemia but also improvement in postprandial hypertriglyceridemia. For example, acute treatment with exenatide in patients with either impaired glucose tolerance or new onset type 2 diabetes results in significant reduction in the postprandial rise in lipoproteins (17). In a separate study, exenatide injection acutely suppresses intestinal lipoprotein production in healthy humans (18). In hamster models, infusion of GLP-1 resulted in significant reduction in chylomicrons. On the other hand, infusion of GLP-2, which co-secretes with GLP-1 in a 1:1 molar ratio physiologically, caused significant increases in TRL. The authors suggest that the combined physiological actions of GLP-1 and GLP-2 favours the latter but from the pharmacological therapy perspective, data to date suggest that the currently available GLP1-R agonists and DPPIV inhibitors are sufficient to tip the balance and achieve net lowering of intestinally derived TRL (19). Taken together, emerging data support the notion that intestinal lipoprotein production is accentuated in insulin resistant and diabetic states and is a potential therapeutic target for CVD prevention. Small dense LDL and low HDL-C in diabetic dyslipidemiadphysiologic consequence of hypertriglyceridemia and beyond Increased preponderance of small dense LDL is commonly observed in subjects with type 2 diabetes and insulin resistance. Presence of large VLDL (VLDL1) is the best predictor for the presence of sdLDL. The mechanisms underlying the formation of sdLDL involve actions of both cholesterol ester transfer protein (CETP) and hepatic lipase (HL). CETP mediates the transfer of TG from the large VLDL to LDL. This is followed by increased lipolysis of the LDL-TG by HL, resulting in a net loss of the neutral lipid core and
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smaller sized LDL with lower buoyancy. In vitro studies suggest that these LDL particles are more pro-atherogenic in part due to their increased flux into the subendothelial space, increased susceptibility to oxidative modification and thus increased uptake by lesion macrophages and longer circulatory half life (20). Reduction of HDL-C is commonly observed as a consequence of hypertriglyceridemia (HTG). Mechanistically, in the presence of excess large VLDL, involvement of CETP and HL similar to those described for LDL will generate small dense HDL which is more rapidly cleared from the circulation, resulting in a reduced circulating level of HDL-C and apoAI. In addition to CETP and HL, activities of other HDL modulators are also altered in subjects with either insulin resistance or type 2 diabetes, potentially contributing to the heightening of the CVD risks. In addition to the regulation by HTG, HDL-C level in insulin resistance and diabetic states has also been shown to be regulated by 2 of the major lipid modulators, namely ATP-binding cassette A1 (ABCA1) and lecithin cholesterol acyltransferase (LCAT). ABCA1 is the transporter first cloned by 3 groups independently when studying the genetic basis of Tangier disease (21e23). ABCA1 mediates the transport of unesterified cholesterol and phospholipid from peripheral tissues onto the circulating HDL with lipid free ApoA-I and nascent HDL being the favoured acceptors. ABCA1 is also a major modulator of HDL-C, as homozygotes with loss of function mutations uniformly developed profound HDL deficiency. Amongst those affected kindreds, predisposition to premature CHD is variable, depending on the nature of the mutations. ABCA1 is widely expressed, but it is the hepatic ABCA1 and to a lesser extent the intestinal ABCA1 that play the most important role in sustaining circulating HDL-C level through promoting HDL biogenesis (24). On the other hand, macrophage derived ABCA1 is crucial for mediating cholesterol efflux from the plaque lesion macrophages and protects from progression of atherogenesis even without impacting on the HDL-C level (25,26). A recent study also revealed that hyperinsulinemia, in the context of insulin resistance, lowers HDL-C in part via lowering the ABCA1 level or reducing its specific activity through phosphorylation at the Tyr1206 site (27). These findings provide 1 additional mechanistic link in the lowering of HDL-C in insulin resistant state independent of hypertriglyceridemia. Lecithin cholesterol acyltransferase (LCAT) is a key lipoproteinassociated enzyme in the circulation, and its major function is to mediate the lipolysis of phosphatidylcholine (PC) at the sn-2 position and transfer the fatty acid moiety to the free cholesterol (FC) as acceptor and form cholesterol ester (CE). LCAT is a potent regulator of HDL metabolism, strongly modulating the HDL-C level as well as lipoprotein compositions, namely the FC/CE ratio. Increased LCAT activity results in increased CE content in the HDL particles as well as overall increase in HDL-C level. Likewise, LCAT deficiency directly results in lowering of HDL-C as well as a modest elevation of triglyceride, a relationship best exemplified by individuals with genetic LCAT deficiency (28). In patients with diabetes, LCAT activities were generally increased and the differences had been found to be significant in some studies (29). These observations suggest that LCAT also contributes to the modulation of the HTG/low HDL phenotype seen in subjects with diabetes. However, the impact of altered LCAT activity in this population is not yet established, although a recent study suggest that increased cholesterol esterification per se is linked to increased incidence of coronary event and sudden death (30). Reciprocal relationship between major HDL modifying genes and the development of cardiometabolic phenotypes The impact of altered ABCA1 and LCAT activities in diabetic states on cardiometabolic risks beyond their effects on dyslipidemia, as highlighted above, have begun to emerge from recent studies.
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Under most circumstances, hepatic and intestinal overproduction of VLDL and chylomicrons, respectively, are the dominant proximal lipid phenotypes, resulting in significant but modest changes in sdLDL and lowering of HDL-C. Studying of the relatively rare genetic causes of profound low HDL has unmasked a reciprocal TG/HDL-C relationship. Early preclinical and human studies suggest that alterations of expression of these HDL modifying genes may in turn modulate risks for development of diabetes and obesity. ABCA1 is important for pancreatic beta cell function Hayden et al examined the impact of pancreatic beta cell specific ABCA1 knockout mice and described a novel role of ABCA1 regulating the beta cell function. Absence of ABCA1 in the beta cell led to accumulation of cellular cholesterol which in turn resulted in an impairment of beta cell function based on impaired insulin exocytosis (31). The translatability of this finding was tested in human subjects homozygous with ABCA1 mutations and showed that patients with absence of ABCA1 have impaired beta cell function and are susceptible to the development of diabetes (32). Likewise, studies of genetic variants of ABCA1 in general population is also consistent with a functional role of ABCA1 in the risk of diabetes, as reviewed by Brunham et al (33). LCAT deficiency is associated with protection from obesity and insulin resistance Recently, studies on the metabolic consequences of LCAT deficiency in mouse models led to a number of unexpected phenotypes, suggesting that LCAT deficiency per se may constitute a protective milieu from insulin resistance, diabetes and obesity (34,35). This protective effect was further augmented if the mice were bred into the LDL receptor knockout background. In the liver, LCAT deficiency was found to be associated with improved glucose tolerance attributable to suppression of hepatic ER stress, the latter being strongly correlated specifically with cholesterol content in the ER, but not other cellular compartments (36). In skeletal muscle, the LCAT-deficient mice were found to develop ectopic brown adipose tissues located in the inter-myofiber areas, in association with increased energy expenditure and resistance to diet-induced obesity (35). The mechanisms underlying how absence of LCAT activity promotes such metabolic protective effects are currently under investigation. Taken together, these apparent reciprocal relationships between beta-cell dysfunction and diabetic dyslipidemia (37) may provide novel therapeutic windows of opportunity for the prevention of diabetes and CVD and warranty further investigations.
Treatment strategies for diabetic dyslipidemia (Table 1) Currently approved lipid modifying agents The use of currently available lipid modifying agents in diabetic subjects have been investigated extensively based on clinical trial findings. This topic has been covered thoroughly by a number of excellent reviews (eg, Matikainen et al (38)) and is summarized briefly below. Statins Although elevated LDL-C is not considered as a cardinal feature of diabetic dyslipidemia, lowering of primarily LDL-C using various statins have provided compelling evidence that statins should also be first-line therapy. In patients with diabetes, statins reduce cardiovascular events by approximately 20% for every 1 mmol/L lowering of LDL-C (39). Although the primary lipid modifying effect of statin is through the general mechanims of LDL lowering by upregulating the LDL receptor, studies suggest statins potently reduce apoB100 and sdLDL, 2 of the cardinal risk factors in diabetic dyslipidemia (40).
Fibrates Fibrate drugs act primarily as synthetic ligand of PPARa. In the case of bezafibrate, it functions as a combined PPARa and PPARg, or so-called “pan-PPAR” agonist. Earlier fibrate trials showed promise for this class of drugs for CVD prevention (41). More recently, the Veterans Affairs HDL Intervention Trial (VA-HIT) reported that gemfibrozil significantly improved primary CVD outcomes versus placebo in high-risk patients initially recruited on the basis of low HDL-C (42). Unfortunately, the unique drug interaction between gemfibrozil and statins somewhat dampened its routine use. The Fenofibrate Intervention and Event Lowering in Diabetes (FIELD) study (43) and Action to Control Cardiovascular Risk in Diabetes (ACCORD-LIPID) trial (44) failed to achieve significance reduction in primary CVD endpoints. However, subgroup analyses as well as meta-analyses on fibrate trials suggest that fibrates may be beneficial for individuals with high TG and low HDL-C. Niacin Early clinical trials with niacin showed cardiovascular benefits, even in face of promoting impaired glucose tolerance or worsening of hyperglycemia in patients with preexisting metabolic syndrome (45). However, results from 2 recent randomized control trials testing the benefits of niacin as add on therapy to statins yielded neutral results in spite of favourable lipid changes (46,47). These findings have significantly dampened the enthusiasm for initiating niacin treatment in at-risk patients. Emerging therapeutic strategies Targeting the hepatic overproduction of apoB/VLDL A number of agents targeting the hepatic production of VLDL are currently in development (Table). MTP inhibition. MTP plays a central role in the assembly and secretion of apoB-containing lipoproteins in the liver and intestine. As mutations in the gene that encodes for MTP were the cause of the rare genetic disorder abetalipoproteinemia, inhibition of MTP for treatment of hyperlipidemia has attracted great interest. Despite their favourable effects on lowering of LDL-C in phase 2 studies, and in some preclinical models, evidence of improved insulin sensitivity (48) and rapid regression of atherosclerotic plaques (49) the concurrent development of gastrointestinal side effects including accumulation of liver fat requires careful evaluation of the longterm impact of such potential adverse effects (50). The development of enterocyte-specific MTP inhibitor is an attractive strategy to more specifically target postprandial lipoprotein production, one of the main features of diabetic dyslipidemia, and in the mean time reduced the incidence of hepatic steatosis. Several candidate agents are currently at various stages of clinical development. ApoB100 inhibition with antisense oligonuceotides. Antisense oligonucleotides (ASO) are short synthetic analogues of natural nucleic acids designed to specifically bind to a target messenger RNA, inducing either selective degradation of the mRNA or inhibiting its translation into protein. Therefore, the inhibition of the gene of interest with this technique is highly specific. Mipomersen, the ASO against apoB100, is by far the most studied ASO in drug development. In phase 3 trials, mipomersen has been shown to efficaciously lower atherogenic lipids and lipoproteins, including LDL-C, TG, small dense LDL and Lp(a) both as monotherapy or as adjunct to statins (51). One of the major concerns is accumulation of hepatic fat and to date, development of hepatic steatosis has been infrequent and reversible upon discontinuation of the drug. Long-term monitoring of this and other adverse effects are warranted. The incretins. Beneficial effects of incretin on dyslipidemia in both human studies and rodent models have begun to emerge in recent
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Table Currently approved and emerging lipid modifying agents, their respective targets and effects on lipid profiles Drug class A. Existing lipid modifying agents Statins
Fibrates
Niacin
B. Emerging strategies in clinical development a. Inhibition of lipoprotein productions MTP inhibitor
ApoB100 antisense oligonucleotide b. Incretins GLP1-R agonists DPP-IV inhibitors c. HDL as primary target CETP inhibitor ApoAI “stimulator” Recombinant HDL infusion
Targets and mechanism
Lipid modifying effects
HMGcoA reductase: Up-regulation of LDL receptor
LDL-C, sdLDL, apoB100 Lowers TG Variable lowering in HDL-C Lowers TG, increases HDL-C effect on LDL-C lowering of sdLDL
PPARa and variable PPARg: Activation of LPL, apoAI, apoAII Reduction of apoCIII Activation of PPARg target genes Hepatic chain ATP synthase Hepatic ABCA1 mediated cholesterol efflux: Increases hepatic derived HDL biogenesis and reduced HDL clearance
MTP: Reduced lipidation of ApoB ApoB mRNA: Degradation of apoB mRNA Enhanced activation of GLP1 receptor signalling
Inhibits CETP activity Up-regulates apoAI gene expression Increase exogenous preb-like HDL
HDL-C, lowers TG, LDL-C and Lp(a)
Lowers VLDL-C Lowers LDL-C Lowers postprandial lipoproteins Lowers VLDL Lowers LDL Lowers postprandial intestinal lipoprotein production
Marked increase in HDL-C Lowers LDL-C Increases apoAI and HDL-C Increases HDL-C
apo, apolipoprotein; apoB100, apolipoprotein B100; CETP, cholesterol ester transfer protein; DPP-IV, dipeptidyl peptidase IV; GLP, glucagon like peptide; HDL, high density lipoprotein; HDL-C, high density lipoprotein cholesterol; HMGcoA reductase, 3-hydroxy-3-methyl-glutaryl-CoA reductase; LDL, Low density lipoprotein; LDL-C, low density lipoprotein cholesterol; mRNA, messenger ribonucleic acid; MTP, microsomal transfer protein; PPARa, peroxisome proliferator-activated receptor alpha; PPARg, peroxisome proliferator-activated receptor gamma; sdLDL, small dense low density lipoproteins; TG, triglyceride; VLDL, very low density lipoprotein; VLDL-C, very low density lipoprotein cholesterol.
years (17e19). In the context of diabetic dyslipidemia, the effect of GLP-1 receptor agonism on post-prandial lipoprotein production is of particular relevance (19). In addition, incretins have also been shown to exert a variety of direct cardiovascular effects (52). Meanwhile, several randomized controlled trials are underway examining the effect of various incretins on cardiovascular outcomes (53,54). Thus the outcome of these studies will provide insight into the relative role of lipid modifying effects of the study drugs on cardiovascular events.
Targeting HDL CETP inhibition. The physiological action of CETP of simultaneously raising HDL-C and lowering of LDL-C continues to make CETP inhibtion an attractive target despite the 2 previous failed attempts with torceptrapib and dalceptrapib, the former with serious off target effets and the latter with general lack of efficacy. Currently, 2 phase 3 trials are on-going evaluating CETP inhibitors anaceptrapib and evaceptrapib for their efficacies in reducing cardiovascular events in high-risk patients. Anaceptrapib: The safety and efficacy of anaceptrapib in highrisk patients were evaluated in an 18-month randomized, double blind, placebo controlled trial (55). In addition to achieving a 138% increase in HDL-C and a 39% reduction in LDL-C after treatment for 24 weeks, anaceptrapib treated subjects were free of adverse cardiovascular events. This study set the stage for the larger outcome trial “The Randomized EValuation of the Effects of Anacetrapib Through Lipid-modification (REVEAL) trial” aims to determine whether lipid modification with anacetrapib 100 mg daily reduces the risk of major coronary events in patients with circulatory problems who have their low-density lipoprotein (LDL) cholesterol level treated with a statin. ClinicalTrials.gov Identifier:NCT00685776.
Evaceptrapib: The purpose of the “Assessment of Clinical Effects of Cholesteryl Ester Transfer Protein Inhibition With Evacetrapib in Patients at a High-Risk for Vascular Outcomes” (ACCELERATE) study is to evaluate the efficacy and safety of evacetrapib in participants with high-risk vascular disease. Primary outcomes are the time to first occurrence of the composite endpoint of cardiovascular (CV) death, myocardial infarction (MI), stroke, coronary revascularization or hospitalization for unstable angina (UA). The REVEAL trial is expected to recruit 30 000 subjects and the ACCELERATE trial to recruit 11 000. Diabetes will be well represented in both trials, as it is one of the major recruitment criteria in both cases. Use of either agent will result in not only potent elevation of HDL-C but also significant reduction in LDL-C. The outcome of the 2 studies will address whether CETP is a viable target. ClinicalTrials.gov Identifier NCT01687998. ApoAI stimulator. In light of the selective avidity of nascent HDL in accepting ABCA1-mediated efflux of cellular cholesterol, it is of interest to test the strategy of apoA1 over-expression in the reversal of atherosclerotic heart disease. Homozygotes for apoAI mutations uniformly cause profound HDL deficiency, but their linkage to premature atherosclerotic heart disease is highly variable in the affected kindreds. That being the case, mutations resulting in inability to synthesize apoAI seem more likely to be associated with premature CVD (56). In mice, transgenic over-expression of apoAI has been shown to raise HDL-C and protects the susceptible animals to accelerated atherosclerosis (57). Furthermore, this model revealed that apoAI expression confers generalized antiinflammatory properties, including the vasculature as well as adipose tissue (58). Based on this hypothesis, a small molecule compound RSV-208 has been developed as a stimulator of apoAI gene expression for treatment of cardiovascular disease (59). Presently, 2 clinical studies are underway. The Study of Quantitative Serial Trends in Lipids with Apolipoprotein A-I Stimulation
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(SUSTAIN, NCT01423188) study aims to evaluate the lipid modifying efficacy, safety and tolerability of an apoA-I inducer RVX-208. The ApoA-I Synthesis Stimulation and Intravascular Ultrasound for Coronary Atheroma Regression Evaluation (ASSURE, NCT01067820) study aims to evaluate the effect of RVX-208 on plaque burden. These 2 studies will likely provide important proof of principle data to further explore apoA-I synthesis activation as a novel therapeutic strategy. If proven effective in the general population, this strategy will be attractive to counter the atherogenicity of diabetic dyslipidemia, in part through raising the nascent preb HDL, the most effective cholesterol acceptor in the efflux pathway. HDL infusion. Numerous preclinical and small human studies support the notion that synthetic reconstituted HDL resemble nascent HDL in their avidity in promoting efflux of cholesterol from peripheral tissues, and lesion macrophages in particular. Proof of principle studies had been reported both in animal models and in humans. ApoAI Milano, a recombinant HDL derived from combining the naturally occurring apoAI Milano mutant with phospholipids, was shown to effectively reduce atherosclerotic plaque volume after 5 weekly injections, beginning within 2 weeks of an acute coronary syndrome (ACS) event (60). Drug development program for clinical use is ongoing. Tardif et al reported an rHDL formulation using native apoAI and phosphotidylcholine (CSL111) in a larger trial, the Effect of rHDL on Atherosclerosis-Safety and efficacy (ERASE). One-hundred and eighty-three post-ACS patients were randomized to 4 weekly infusions of placebo or 1 of 2 doses of CSL-111 (61). The greater dose was discontinued because of a high incidence of hepatic enzyme elevation, but among the 136 patients with evaluable endpoint data, percent change in atheroma volume, the primary endpoint, improved significantly in the CSL-111 group but not in the placebo group. Extension of this line of therapeutic strategy includes several ongoing trials: 1) By using a second-generation CSL-based rHDL with apparently more efficacious cholesterol efflux induction, the study “A Single Ascending Dose Study Examining the Safety and Pharmacokinetic Profile of Reconstituted High Density Lipoprotein (CSL112) Administered to Patients,” (ClinicalTrials.gov Identifier: NCT01499420) is currently underway. 2) A related study entitled “Effect of CER-001 on Atherosclerosis in Acute Coronary Syndrome (ACS) Patients - Efficacy and Safety: The CHI SQUARE Trial” (ClinicalTrials.gov Identifier: NCT01201837) will assess the effects of CER-001, an ApoA-Ibased HDL mimetic, on indices of atherosclerotic plaque progression and regression as assessed by intravascular ultrasound (IVUS) measurements in patients with ACS. 3) The study entitled “Safety and Efficacy of APL180 in Healthy Volunteers and Patients With Coronary Heart Disease (CHD)” will assess the safety and tolerability, pharmacokinetics and effects on biomarkers of HDL function of APL180 after a single and 7-daily infusions in healthy volunteers and in patients with coronary heart disease (CHD) (ClinicalTrials.gov Identifier: NCT00568594). Evolving novel strategies Activation of ABCA1da target with an expanding scope of therapeutic benefits Numerous preclinical studies suggest that regulation of macrophage/foam cell specific ABCA1 and ABCG1 is sufficient to modulate atherogenesis, independent of circulating level of HDL (62). On the other hand, the role of hepatic derived ABCA1, though strongly influencing the HDL-C levels, in atherosclerosis seem less relevant (63). Effective pharmacological strategies to modulate ABCA1 and/or ABCG1 in combating atherosclerosis continue to be
elusive. Although activation of the transcription factor liver X receptor (LXR) has shown promise as an upstream target for its role in activating ABCA1 expression, drug development along this pathway was hampered by the concomitant activation of hepatic lipogenesis, resulting in significant hepatic steatosis and hypertriglyceridemia. Very recently, ABCA1 was found to be a specific target of micro RNA 33 (miR33) a/b. Mature miRNAs are w22-nucleotide singlestranded RNAs that exert their function via perfect Watson-Crick base pairing, with sequences most commonly located within the 3’untranslated region (3’-UTR) of target mRNAs. Interaction of a miRNA with its target mRNA results in inhibition of translation and/ or degradation of mRNAs. Preclinical studies revealed that inhibition of miR33 resulted in upregulation of Abca1, increased serum HDL-C (likely as a result of activation of hepatic ABCA1) and reduction in atherosclerotic burden (64). Interestingly, miR33 has also been shown to modulate ABCA1 expression and insulin secretory function of pancreatic beta cells. Taken together, inhibition of miR33 is a potentially promising therapeutic target to prevent atherosclerosis through promoting ABCA1 expression without the issue of HTG on the one hand, and prevent beta cell failure on the other. LCAT-related targetsdnew lessons from an old molecule that does not follow the script Traditionally, LCAT has been regarded as key mediator of the reverse cholesterol transport and modulator of HDL-C levels. The controversial, or sometimes paradoxical relationship between LCAT and atherosclerosis, has led to the increasing recognition of its complex physiological function in vivo and that this enzyme per se being an elusive therapeutic target. However, with the recent findings made in the rodent models of LCAT deficiency, a number of potential targetable metabolic pathways have emerged. A recent report by Hager et al (36) demonstrated that dietary cholesterol being an important promoter of hepatic ER stress, a cellular process known to mediate a network of metabolic-inflammatory responses, including insulin resistance. The deletion of LCAT in this experimental model protects the mice from ER stress by preventing accumulation of cholesterol in the ER, independent of cholesterol accumulation in other cellular compartments. This unexpected observation, therefore, raised the possibility for targeting ER cholesterol for preventing insulin resistance, through modulating the cellular cholesterol sensing machinery, namely the SREBP processing machinery. Additional research is required to further establish this experimental paradigm. Conclusion Diabetic dyslipidemia is a significant proatherogenic factor. It is postulated that the residual dyslipidemia, namely elevated TG and low HDL-C after maximal statin therapy, contribute significantly to the residual risk. However, effective therapeutic tools beyond statin treatment remain elusive. Our understanding of the pathophysiology of diabetic dyslipidemia continues to evolve. Novel regulatory factors of the high TG/low HDL phenotype continues to emerge. With the newer tools either undergoing clinical trials or at various stages of drug discovery, the current emerging development continues to raise hope to fill the therapeutic gap in cardiometabolic complications of diabetes. Acknowledgements This work was supported in part by an operating grant from the Canadian Institutes for Health Research (CIHR; MOP275369), a CIHR ChinaeCanada Joint Health Research Initiative grant and a Grant-in-aid (NA 6331) by the Heart and Stroke Foundation of Ontario.
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Author Disclosures None to disclose.
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Eur J Clin Invest 2000;30:685e94. 6. Kissebah AH, Alfarsi S, Evans DJ, et al. Integrated regulation of very low density lipoprotein triglyceride and apolipoprotein-B kinetics in non-insulindependent diabetes mellitus. Diabetes 1982;31:217e25. 7. Yecies JL, Zhang HH, Menon S, et al. Akt stimulates hepatic SREBP1c and lipogenesis through parallel mTORC1-dependent and independent pathways. Cell Metab 2011;14:21e32. 8. Pajvani UB, Qiang L, Kangsamaksin T, et al. Inhibition of Notch uncouples Akt activation from hepatic lipid accumulation by decreasing mTorc1 stability. Nat Med; 2013 [Epub ahead of print]. 9. Arca M, Pigna G, Favoccia C. Mechanisms of diabetic dyslipidemia: relevance for atherogenesis. Curr Vasc Pharmacol 2012;10:684e6. 10. Fisher EA. The degradation of apolipoprotein B100: multiple opportunities to regulate VLDL triglyceride production by different proteolytic pathways. Biochim Biophys Acta 2012;1821:778e81. 11. Goldberg IJ, Eckel RH, McPherson R. Triglycerides and heart disease: still a hypothesis? Arterioscler Thromb Vasc Biol 2011;31:1716e25. 12. Botham KM, Moore EH, De Pascale C, et al. The induction of macrophage foam cell formation by chylomicron remnants. Biochem Soc Trans 2007;35(Pt 3):454e8. 13. Nakano T, Nakajima K, Niimi M, et al. Detection of apolipoproteins B-48 and B-100 carrying particles in lipoprotein fractions extracted from human aortic atherosclerotic plaques in sudden cardiac death cases. Clin Chim Acta 2008; 390:38e43. 14. Weinstein MM, Yin L, Tu Y, et al. Chylomicronemia elicits atherosclerosis in miceebrief report. Arterioscler Thromb Vasc Biol 2010;30:20e3. 15. Xiao C, Lewis GF. Regulation of chylomicron production in humans. Biochim Biophys Acta 2012;1821:736e46. 16. Pan X, Hussain MM. Gut triglyceride production. Biochim Biophys Acta 2012; 1821:727e35. 17. Schwartz EA, Koska J, Mullin MP, et al. Exenatide suppresses postprandial elevations in lipids and lipoproteins in individuals with impaired glucose tolerance and recent onset type 2 diabetes mellitus. Atherosclerosis 2010;212:217e22. 18. Xiao C, Bandsma RH, Dash S, et al. Exenatide, a glucagon-like peptide-1 receptor agonist, acutely inhibits intestinal lipoprotein production in healthy humans. Arterioscler Thromb Vasc Biol 2012;32:1513e9. 19. Hein GJ, Baker C, Hsieh J, et al. GLP-1 and GLP-2 as yin and yang of intestinal lipoprotein production: evidence for predominance of GLP-2-stimulated postprandial lipemia in normal and insulin-resistant states. Diabetes 2013; 62:373e81. 20. Toth PP. Insulin resistance, small LDL particles, and risk for atherosclerotic disease. Curr Vasc Pharmacol; 2013 [Epub ahead of print]. 21. Brooks-Wilson A, Marcil M, Clee SM, et al. Mutations in ABC1 in Tangier disease and familial high-density lipoprotein deficiency. Nat Genet 1999;22:336e45. 22. Rust S, Rosier M, Funke H, et al. Tangier disease is caused by mutations in the gene encoding ATP-binding cassette transporter 1. Nat Genet 1999;22:352e5. 23. Marcil M, Brooks-Wilson A, Clee SM, et al. Mutations in the ABC1 gene in familial HDL deficiency with defective cholesterol efflux. Lancet 1999;354:1341e6. 24. Singaraja RR, Van Eck M, Bissada N, et al. Both hepatic and extrahepatic ABCA1 have discrete and essential functions in the maintenance of plasma highdensity lipoprotein cholesterol levels in vivo. Circulation 2006;114:1301e9. 25. Van Eck M, Singaraja RR, Ye D, et al. Macrophage ATP-binding cassette transporter A1 overexpression inhibits atherosclerotic lesion progression in low-density lipoprotein receptor knockout mice. Arterioscler Thromb Vasc Biol 2006;26:929e34. 26. Westerterp M, Murphy AJ, Wang M, et al. Deficiency of ABCA1 and ABCG1 in macrophages increases inflammation and accelerates atherosclerosis in mice. Circ Res; 2013 [Epub ahead of print]. 27. Nonomura K, Arai Y, Mitani H, et al. 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29. Kappelle PJ, de Boer JF, Perton FG, et al. Increased LCAT activity and hyperglycaemia decrease the antioxidative functionality of HDL. Eur J Clin Invest 2012;42:487e95. 30. Tanaka S, Yasuda T, Ishida T, et al. Increased serum cholesterol esterification rates predict coronary heart disease and sudden death in a general population. Arterioscler Thromb Vasc Biol 2013;33:1098e104. 31. Kruit JK, Wijesekara N, Westwell-Roper C, et al. Loss of both ABCA1 and ABCG1 results in increased disturbances in islet sterol homeostasis, inflammation, and impaired b-cell function. Diabetes 2012;61:659e64. 32. Koseki M, Matsuyama A, Nakatani K, et al. Impaired insulin secretion in four Tangier disease patients with ABCA1 mutations. J Atheroscler Thromb 2009;16: 292e6. 33. Brunham LR, Kruit JK, Hayden MR, et al. Cholesterol in beta-cell dysfunction: the emerging connection between HDL cholesterol and type 2 diabetes. Curr Diab Rep 2010;10:55e60. 34. Ng DS, Xie C, Maguire GF, et al. Hypertriglyceridemia in lecithin-cholesterol acyltransferase-deficient mice is associated with hepatic overproduction of triglycerides, increased lipogenesis, and improved glucose tolerance. J Biol Chem 2004;279:7636e42. 35. Li L, Hossain MA, Sadat S, et al. Lecithin cholesterol acyltransferase null mice are protected from diet-induced obesity and insulin resistance in a genderspecific manner through multiple pathways. J Biol Chem 2011;286:17809e20. 36. Hager L, Li L, Pun H, et al. Lecithin:cholesterol acyltransferase deficiency protects against cholesterol-induced hepatic endoplasmic reticulum stress in mice. J Biol Chem 2012;287:20755e68. 37. Bardini G, Rotella CM, Giannini S. Dyslipidemia and diabetes: reciprocal impact of impaired lipid metabolism and Beta-cell dysfunction on micro- and macrovascular complications. Rev Diabet Stud 2012;9:82e93. 38. Matikainen N, Taskinen MR. Management of dyslipidemias in the presence of the metabolic syndrome or type 2 diabetes. Curr Cardiol Rep 2012;14:721e31. 39. de Vries FM, Denig P, Pouwels KB, et al. Primary prevention of major cardiovascular and cerebrovascular events with statins in diabetic patients: a meta-analysis. Drugs 2012;72:2365e73. 40. Kei A, Liberopoulos E, Tellis K, Rizzo M, Elisaf M, Tselepis A. Effect of hypolipidemic treatment on emerging risk factors in mixed dyslipidemia: a randomized pilot trial. Eur J Clin Invest 2013;43:698e707. 41. Steiner G. Lipid intervention trials in diabetes. Diabetes Care 2000;23(Suppl 2): B49e53. 42. Rubins HB, Robins SJ, Collins D, et al. Gemfibrozil for the secondary prevention of coronary heart disease in men with low levels of high-density lipoprotein cholesterol. Veterans Affairs High-Density Lipoprotein Cholesterol Intervention Trial Study Group. N Engl J Med 1999;341:410e8. 43. Keech A, Simes RJ, Barter P, et al, FIELD study investigators. Effects of long-term fenofibrate therapy on cardiovascular events in 9795 people with type 2 diabetes mellitus (the FIELD study): randomised controlled trial. Lancet 2005; 366:1849e61. Erratum in: Lancet 2006;368:1415; 1420. 44. ACCORD Study Group, Ginsberg HN, Elam MB, Lovato LC, et al. Effects of combination lipid therapy in type 2 diabetes mellitus. N Engl J Med 2010;362: 1563e74. 45. Phan BA, Muñoz L, Shadzi P, et al. Effects of niacin on glucose levels, coronary stenosis progression, and clinical events in subjects with normal baseline glucose levels (
Contents lists available at ScienceDirect
Canadian Journal of Diabetes journal homepage: www.canadianjournalofdiabetes.com
Review
Diabetic Dyslipidemia: From Evolving Pathophysiological Insight to Emerging Therapeutic Targets Dominic S. Ng PhD, MD, FRCPC * Li Ka Shing Knowledge Institute, Keenan Research Center, St. Michael’s Hospital, University of Toronto, Toronto, Ontario, Canada
a r t i c l e i n f o
a b s t r a c t
Article history: Received 26 May 2013 Received in revised form 22 July 2013 Accepted 22 July 2013
Diabetic dyslipidemia is characterized by hepatic very low density lipoprotein (VLDL) and intestinal chylomicron overproduction, reduced high density lipoprotein cholesterol (HDL-C) level, increased propensity of small dense LDL (sdLDL) and increased postprandial lipemia. This dyslipidemic profile is also strongly linked to other features of the metabolic syndrome. Diabetic dyslipidemia is a well-recognized risk factor for atherosclerotic cardiovascular diseases. Currently, statins remain the first line therapy primarily through reducing the atherogenic LDL. Clinical trials on other lipid modifying agents were met with variable success in selective patient populations. Emerging new insights into the pathophysiology of lipid metabolism, in general, and diabetic dyslipidemia, in particular, have opened up potentially novel therapeutic strategies to further reduce the risk associated with diabetic dyslipidemia and insulin resistant state. Ó 2013 Canadian Diabetes Association
Keywords: cardiovascular diseases cholesterol efflux diabetes dyslipidemia ER stress HDL insulin resistance triglycerides Mots clés : maladies cardiovasculaires efflux du cholestérol diabète dyslipidémie stress du RE HDL insulinorésistance triglycérides
r é s u m é La dyslipidémie du diabétique est caractérisée par la surproduction hépatique de lipoprotéines de très faible densité (VLDL : very low density lipoprotein) et la surproduction intestinale de chylomicrons, la réduction de la concentration du cholestérol à lipoprotéines de haute densité (cholestérol HDL), l’augmentation de la propension des LDL petites et denses, et l’augmentation de la lipémie postprandiale. Ce profil dyslipidémique est aussi fortement lié à d’autres caractéristiques du syndrome métabolique. La dyslipidémie du diabétique est un facteur de risque bien connu des maladies cardiovasculaires athérosclérotiques. Actuellement, les statines semblent être le traitement de première intention, principalement pour la réduction des LDL athérogènes. Les essais cliniques sur d’autres agents modifiant les lipides ont obtenu un succès inégal chez certaines populations de patients. Les perspectives émergentes en physiopathologie du métabolisme des lipides, en général, et la dyslipidémie du diabétique, en particulier, ont possiblement amené les nouvelles stratégies thérapeutiques à réduire davantage le risque associé à la dyslipidémie du diabétique et à l’état d’insulinorésistance. Ó 2013 Canadian Diabetes Association
Introduction Type II diabetes mellitus is a chronic metabolic disorder that predisposes an individual to be high risk for atherosclerotic cardiovascular diseases (CVD). This is particularly the case if the patient also carries clinical features of the metabolic syndrome, characterized by central obesity, hypertension, impaired fasting glucose, elevated triglyceride (TG) and reduced levels of high density lipoprotein cholesterol (HDL-C), with the latter 2 entities being clinical surrogates for diabetic dyslipidemia (1). Diabetic dyslipidemia in type II diabetes is characterized by overproduction
* Address for correspondence: Dominic S. Ng, PhD, MD, FRCPC, Associate Professor of Medicine, University of Toronto, Keenan Research Center, Li Ka Shing Knowledge Institute, St Michael’s Hospital, Shuter Wing Room 3-041, 30 Bond Street, Toronto, Ontario M5B 1W8, Canada. E-mail address: [email protected]. 1499-2671/$ e see front matter Ó 2013 Canadian Diabetes Association http://dx.doi.org/10.1016/j.jcjd.2013.07.062
of triglyceride-rich lipoproteins (TRL) both by the liver and the intestine, reduced plasma levels HDL-C and increased preponderance of small dense low density lipoproteins (sdLDL) (Figure) (2). This dyslipidemic profile is well recognized to be one of the proatherogenic factors in type II diabetic subjects; but the underlying pathophysiologic mechanisms remain inadequately understood and amelioration of this phenotype remains elusive. In type 1 diabetes subjects, presence of dyslipidemia, which includes elevated TG, LDL-C and low HDL-C, predicts future CVD, but the prevalence of dyslipidemia in type 1 diabetes subjects is not higher than their nondiabetic counterparts (3,4). Overproduction of triglyceride-rich lipoproteins in insulin-resistant state In the fasting state, the liver constitutively synthesizes apolipoprotein (apo) B100, an essential protein component of VLDL.
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Figure. Key pathways that may be altered in diabetic dyslipidemia. (1) Hepatic VLDL overproduction: increased TG availability, increased MTP activity, reduced apoB100 degradation, reduced LPL clearance. (2) Intestinal chylomicron overproduction: increased TG availability, increased MTP activity, reduced GLP-1 action, reduced LPL clearance. (3) Small dense LDL: increased CETP activity, increased TRL. (4) Reduced HDL-C: increased CETP activity, increased TRL, reduced ABCA1 level or activity; reduced apoAI synthesis. ABCA1, ATPase binding cassette A1; apoB100, apolipoprotein B100; CETP, cholesterol ester transfer protein; LPL, lipoprotein lipase; MTP, microsomal transfer protein; TRL, triglyceride rich lipoproteins; VLDL, Very low density lipoproteins.
Newly synthesized apoB100 is co-translationally lipidated by microsomal transfer protein (MTP) with TG and cholesterol ester delivered in the endoplasmic reticulum. Continuing lipidation of these primordial lipoproteins result in maturation of the VLDL particle which will be transported to the Golgi, the site of secretion of the mature VLDL. ApoB100 is constitutively synthesized and the excess, un-lipidated molecules are destined for degradation. Several groups have established that hepatic VLDL over-production is a cardinal feature of diabetic dyslipidemia based on kinetic studies (5,6). In diabetic states, several factors contribute mechanistically to the observed hepatic overproduction of VLDL. First, lipidation of apoB100 is dependent on the availability of triglyceride in the cytosol. Insulin is a strong inducer of sterol response element binding protein 1c (SREBP1c), which is the master transcription factor for promoting de novo lipogenesis, resulting in increased fatty acid as substrate for TG synthesis. In insulin resistant state, the activation of SREBP1c and its downstream lipogenic gene program, through mechanisms that remain incompletely understood (7,8), are accentuated differentially by hyperinsulinemia, resulting in increased fatty acid substrate availability for hepatic TG synthesis. Second, MTP, the central player in mediating the lipidation of apoB100, is commonly activated in a variety of mouse or rat models of diabetes or insulin resistance although
the mechanism remains to be fully elucidated (9). Third, insulin resistant state per se has been shown to attenuate apoB100 degradation. Multiple mechanisms have been identified to mediate apoB100 degradation including authophagy (10). Intestinal lipoprotein overproduction in insulin-resistant state Chylomicrons are lipoproteins synthesized and secreted by intestinal enterocytes, as a response to dietary fat ingestion. In the intestinal cells, free fatty acids of dietary source are resynthesized into TG and are available to lipidate a truncated form of apoB, namely the apoB48, also mediated by MTP. Several lines of experimental evidence support a pro-atherogenic nature of chylomicron remnants (CMR) (11). Botham et al reported that CMR may be taken up directly by macrophages, promoting foam cell formation (12). A recent study reported the detection of apoB48 along with apoB100 proteins in atherosclerotic plaques from autopsy of subjects who suffered sudden cardiac death, implicating a pro-atherogenic role for the intestinally derived, apoB48-containing TRL (13). This finding is consistent with the pro-atherogenic phenotype seen in a mouse model of chylomicronemia (14). Similar to their hepatic counterparts, each chylomicron particle contains 1 molecule of apoB48. In a lipid-rich situation, chylomicrons
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such formed are highly buoyant with high content of lipids for each apoB48. Unlike their VLDL counterpart, degradation of apoB48 is considerably less of a regulatory factor, as the enterocytes can form and secrete more small dense chylomicrons. Thus lipid availability and MTP activity are considered as the major regulatory factors for chylomicron production. In patients with diabetes and subjects with insulin resistance, several lines of evidence strongly suggest that the intestine overproduces chylomicrons when compared to the healthy controls (15). Mechanistically, increased MTP mass and activity increased de novo lipogenesis, as well as increased apoB stability, have all been implicated. In addition to intracellular regulatory mechanisms, it has been shown that circulating free fatty acids also stimulate intestinal chylomicron production, although the underlying mechanism remains uncertain. Increased plasma free fatty acid level is commonly observed in subjects with insulin resistance and obesity, in part due to either impaired uptake by peripheral tissues upon lipolysis of circulating TRL or enhanced lipolysis in the dysfunctional white adipose tissues (16). Thus, a free fatty acid mediated intestinal lipoprotein overproduction is expected to be a common lipid phenotype in subjects with diabetes. Modulation of intestinal lipoprotein production by incretin hormones Glucagon-like peptide (GLP)-1 and GLP-2 are 2 peptides expressed by the proglucagon gene in the intestinal L cells. A major biological action of GLP-1 is its ability to lower blood glucose by stimulating insulin secretion and suppressing glucagon secretion in the islets. The secretion of GLP-1 from the L cells is triggered by the presence of nutrients in the intestine, and, therefore, plays a key role in glucose homeostasis. In light of the very short half-life of endogenously produced GLP-1, GLP-1 analogues, GLP-R agonists and the dipeptidylpeptidase-4 (DPP-IV) inhibitors, the latter through delaying the degradation of endogenous GLP-1 have been formulated and used widely for treatment of hyperglycemia in subjects with diabetes. In humans, short-term treatment with GLP-1 analogues or DPP-IV inhibitors result in not only improvement in glycemia but also improvement in postprandial hypertriglyceridemia. For example, acute treatment with exenatide in patients with either impaired glucose tolerance or new onset type 2 diabetes results in significant reduction in the postprandial rise in lipoproteins (17). In a separate study, exenatide injection acutely suppresses intestinal lipoprotein production in healthy humans (18). In hamster models, infusion of GLP-1 resulted in significant reduction in chylomicrons. On the other hand, infusion of GLP-2, which co-secretes with GLP-1 in a 1:1 molar ratio physiologically, caused significant increases in TRL. The authors suggest that the combined physiological actions of GLP-1 and GLP-2 favours the latter but from the pharmacological therapy perspective, data to date suggest that the currently available GLP1-R agonists and DPPIV inhibitors are sufficient to tip the balance and achieve net lowering of intestinally derived TRL (19). Taken together, emerging data support the notion that intestinal lipoprotein production is accentuated in insulin resistant and diabetic states and is a potential therapeutic target for CVD prevention. Small dense LDL and low HDL-C in diabetic dyslipidemiadphysiologic consequence of hypertriglyceridemia and beyond Increased preponderance of small dense LDL is commonly observed in subjects with type 2 diabetes and insulin resistance. Presence of large VLDL (VLDL1) is the best predictor for the presence of sdLDL. The mechanisms underlying the formation of sdLDL involve actions of both cholesterol ester transfer protein (CETP) and hepatic lipase (HL). CETP mediates the transfer of TG from the large VLDL to LDL. This is followed by increased lipolysis of the LDL-TG by HL, resulting in a net loss of the neutral lipid core and
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smaller sized LDL with lower buoyancy. In vitro studies suggest that these LDL particles are more pro-atherogenic in part due to their increased flux into the subendothelial space, increased susceptibility to oxidative modification and thus increased uptake by lesion macrophages and longer circulatory half life (20). Reduction of HDL-C is commonly observed as a consequence of hypertriglyceridemia (HTG). Mechanistically, in the presence of excess large VLDL, involvement of CETP and HL similar to those described for LDL will generate small dense HDL which is more rapidly cleared from the circulation, resulting in a reduced circulating level of HDL-C and apoAI. In addition to CETP and HL, activities of other HDL modulators are also altered in subjects with either insulin resistance or type 2 diabetes, potentially contributing to the heightening of the CVD risks. In addition to the regulation by HTG, HDL-C level in insulin resistance and diabetic states has also been shown to be regulated by 2 of the major lipid modulators, namely ATP-binding cassette A1 (ABCA1) and lecithin cholesterol acyltransferase (LCAT). ABCA1 is the transporter first cloned by 3 groups independently when studying the genetic basis of Tangier disease (21e23). ABCA1 mediates the transport of unesterified cholesterol and phospholipid from peripheral tissues onto the circulating HDL with lipid free ApoA-I and nascent HDL being the favoured acceptors. ABCA1 is also a major modulator of HDL-C, as homozygotes with loss of function mutations uniformly developed profound HDL deficiency. Amongst those affected kindreds, predisposition to premature CHD is variable, depending on the nature of the mutations. ABCA1 is widely expressed, but it is the hepatic ABCA1 and to a lesser extent the intestinal ABCA1 that play the most important role in sustaining circulating HDL-C level through promoting HDL biogenesis (24). On the other hand, macrophage derived ABCA1 is crucial for mediating cholesterol efflux from the plaque lesion macrophages and protects from progression of atherogenesis even without impacting on the HDL-C level (25,26). A recent study also revealed that hyperinsulinemia, in the context of insulin resistance, lowers HDL-C in part via lowering the ABCA1 level or reducing its specific activity through phosphorylation at the Tyr1206 site (27). These findings provide 1 additional mechanistic link in the lowering of HDL-C in insulin resistant state independent of hypertriglyceridemia. Lecithin cholesterol acyltransferase (LCAT) is a key lipoproteinassociated enzyme in the circulation, and its major function is to mediate the lipolysis of phosphatidylcholine (PC) at the sn-2 position and transfer the fatty acid moiety to the free cholesterol (FC) as acceptor and form cholesterol ester (CE). LCAT is a potent regulator of HDL metabolism, strongly modulating the HDL-C level as well as lipoprotein compositions, namely the FC/CE ratio. Increased LCAT activity results in increased CE content in the HDL particles as well as overall increase in HDL-C level. Likewise, LCAT deficiency directly results in lowering of HDL-C as well as a modest elevation of triglyceride, a relationship best exemplified by individuals with genetic LCAT deficiency (28). In patients with diabetes, LCAT activities were generally increased and the differences had been found to be significant in some studies (29). These observations suggest that LCAT also contributes to the modulation of the HTG/low HDL phenotype seen in subjects with diabetes. However, the impact of altered LCAT activity in this population is not yet established, although a recent study suggest that increased cholesterol esterification per se is linked to increased incidence of coronary event and sudden death (30). Reciprocal relationship between major HDL modifying genes and the development of cardiometabolic phenotypes The impact of altered ABCA1 and LCAT activities in diabetic states on cardiometabolic risks beyond their effects on dyslipidemia, as highlighted above, have begun to emerge from recent studies.
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Under most circumstances, hepatic and intestinal overproduction of VLDL and chylomicrons, respectively, are the dominant proximal lipid phenotypes, resulting in significant but modest changes in sdLDL and lowering of HDL-C. Studying of the relatively rare genetic causes of profound low HDL has unmasked a reciprocal TG/HDL-C relationship. Early preclinical and human studies suggest that alterations of expression of these HDL modifying genes may in turn modulate risks for development of diabetes and obesity. ABCA1 is important for pancreatic beta cell function Hayden et al examined the impact of pancreatic beta cell specific ABCA1 knockout mice and described a novel role of ABCA1 regulating the beta cell function. Absence of ABCA1 in the beta cell led to accumulation of cellular cholesterol which in turn resulted in an impairment of beta cell function based on impaired insulin exocytosis (31). The translatability of this finding was tested in human subjects homozygous with ABCA1 mutations and showed that patients with absence of ABCA1 have impaired beta cell function and are susceptible to the development of diabetes (32). Likewise, studies of genetic variants of ABCA1 in general population is also consistent with a functional role of ABCA1 in the risk of diabetes, as reviewed by Brunham et al (33). LCAT deficiency is associated with protection from obesity and insulin resistance Recently, studies on the metabolic consequences of LCAT deficiency in mouse models led to a number of unexpected phenotypes, suggesting that LCAT deficiency per se may constitute a protective milieu from insulin resistance, diabetes and obesity (34,35). This protective effect was further augmented if the mice were bred into the LDL receptor knockout background. In the liver, LCAT deficiency was found to be associated with improved glucose tolerance attributable to suppression of hepatic ER stress, the latter being strongly correlated specifically with cholesterol content in the ER, but not other cellular compartments (36). In skeletal muscle, the LCAT-deficient mice were found to develop ectopic brown adipose tissues located in the inter-myofiber areas, in association with increased energy expenditure and resistance to diet-induced obesity (35). The mechanisms underlying how absence of LCAT activity promotes such metabolic protective effects are currently under investigation. Taken together, these apparent reciprocal relationships between beta-cell dysfunction and diabetic dyslipidemia (37) may provide novel therapeutic windows of opportunity for the prevention of diabetes and CVD and warranty further investigations.
Treatment strategies for diabetic dyslipidemia (Table 1) Currently approved lipid modifying agents The use of currently available lipid modifying agents in diabetic subjects have been investigated extensively based on clinical trial findings. This topic has been covered thoroughly by a number of excellent reviews (eg, Matikainen et al (38)) and is summarized briefly below. Statins Although elevated LDL-C is not considered as a cardinal feature of diabetic dyslipidemia, lowering of primarily LDL-C using various statins have provided compelling evidence that statins should also be first-line therapy. In patients with diabetes, statins reduce cardiovascular events by approximately 20% for every 1 mmol/L lowering of LDL-C (39). Although the primary lipid modifying effect of statin is through the general mechanims of LDL lowering by upregulating the LDL receptor, studies suggest statins potently reduce apoB100 and sdLDL, 2 of the cardinal risk factors in diabetic dyslipidemia (40).
Fibrates Fibrate drugs act primarily as synthetic ligand of PPARa. In the case of bezafibrate, it functions as a combined PPARa and PPARg, or so-called “pan-PPAR” agonist. Earlier fibrate trials showed promise for this class of drugs for CVD prevention (41). More recently, the Veterans Affairs HDL Intervention Trial (VA-HIT) reported that gemfibrozil significantly improved primary CVD outcomes versus placebo in high-risk patients initially recruited on the basis of low HDL-C (42). Unfortunately, the unique drug interaction between gemfibrozil and statins somewhat dampened its routine use. The Fenofibrate Intervention and Event Lowering in Diabetes (FIELD) study (43) and Action to Control Cardiovascular Risk in Diabetes (ACCORD-LIPID) trial (44) failed to achieve significance reduction in primary CVD endpoints. However, subgroup analyses as well as meta-analyses on fibrate trials suggest that fibrates may be beneficial for individuals with high TG and low HDL-C. Niacin Early clinical trials with niacin showed cardiovascular benefits, even in face of promoting impaired glucose tolerance or worsening of hyperglycemia in patients with preexisting metabolic syndrome (45). However, results from 2 recent randomized control trials testing the benefits of niacin as add on therapy to statins yielded neutral results in spite of favourable lipid changes (46,47). These findings have significantly dampened the enthusiasm for initiating niacin treatment in at-risk patients. Emerging therapeutic strategies Targeting the hepatic overproduction of apoB/VLDL A number of agents targeting the hepatic production of VLDL are currently in development (Table). MTP inhibition. MTP plays a central role in the assembly and secretion of apoB-containing lipoproteins in the liver and intestine. As mutations in the gene that encodes for MTP were the cause of the rare genetic disorder abetalipoproteinemia, inhibition of MTP for treatment of hyperlipidemia has attracted great interest. Despite their favourable effects on lowering of LDL-C in phase 2 studies, and in some preclinical models, evidence of improved insulin sensitivity (48) and rapid regression of atherosclerotic plaques (49) the concurrent development of gastrointestinal side effects including accumulation of liver fat requires careful evaluation of the longterm impact of such potential adverse effects (50). The development of enterocyte-specific MTP inhibitor is an attractive strategy to more specifically target postprandial lipoprotein production, one of the main features of diabetic dyslipidemia, and in the mean time reduced the incidence of hepatic steatosis. Several candidate agents are currently at various stages of clinical development. ApoB100 inhibition with antisense oligonuceotides. Antisense oligonucleotides (ASO) are short synthetic analogues of natural nucleic acids designed to specifically bind to a target messenger RNA, inducing either selective degradation of the mRNA or inhibiting its translation into protein. Therefore, the inhibition of the gene of interest with this technique is highly specific. Mipomersen, the ASO against apoB100, is by far the most studied ASO in drug development. In phase 3 trials, mipomersen has been shown to efficaciously lower atherogenic lipids and lipoproteins, including LDL-C, TG, small dense LDL and Lp(a) both as monotherapy or as adjunct to statins (51). One of the major concerns is accumulation of hepatic fat and to date, development of hepatic steatosis has been infrequent and reversible upon discontinuation of the drug. Long-term monitoring of this and other adverse effects are warranted. The incretins. Beneficial effects of incretin on dyslipidemia in both human studies and rodent models have begun to emerge in recent
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Table Currently approved and emerging lipid modifying agents, their respective targets and effects on lipid profiles Drug class A. Existing lipid modifying agents Statins
Fibrates
Niacin
B. Emerging strategies in clinical development a. Inhibition of lipoprotein productions MTP inhibitor
ApoB100 antisense oligonucleotide b. Incretins GLP1-R agonists DPP-IV inhibitors c. HDL as primary target CETP inhibitor ApoAI “stimulator” Recombinant HDL infusion
Targets and mechanism
Lipid modifying effects
HMGcoA reductase: Up-regulation of LDL receptor
LDL-C, sdLDL, apoB100 Lowers TG Variable lowering in HDL-C Lowers TG, increases HDL-C effect on LDL-C lowering of sdLDL
PPARa and variable PPARg: Activation of LPL, apoAI, apoAII Reduction of apoCIII Activation of PPARg target genes Hepatic chain ATP synthase Hepatic ABCA1 mediated cholesterol efflux: Increases hepatic derived HDL biogenesis and reduced HDL clearance
MTP: Reduced lipidation of ApoB ApoB mRNA: Degradation of apoB mRNA Enhanced activation of GLP1 receptor signalling
Inhibits CETP activity Up-regulates apoAI gene expression Increase exogenous preb-like HDL
HDL-C, lowers TG, LDL-C and Lp(a)
Lowers VLDL-C Lowers LDL-C Lowers postprandial lipoproteins Lowers VLDL Lowers LDL Lowers postprandial intestinal lipoprotein production
Marked increase in HDL-C Lowers LDL-C Increases apoAI and HDL-C Increases HDL-C
apo, apolipoprotein; apoB100, apolipoprotein B100; CETP, cholesterol ester transfer protein; DPP-IV, dipeptidyl peptidase IV; GLP, glucagon like peptide; HDL, high density lipoprotein; HDL-C, high density lipoprotein cholesterol; HMGcoA reductase, 3-hydroxy-3-methyl-glutaryl-CoA reductase; LDL, Low density lipoprotein; LDL-C, low density lipoprotein cholesterol; mRNA, messenger ribonucleic acid; MTP, microsomal transfer protein; PPARa, peroxisome proliferator-activated receptor alpha; PPARg, peroxisome proliferator-activated receptor gamma; sdLDL, small dense low density lipoproteins; TG, triglyceride; VLDL, very low density lipoprotein; VLDL-C, very low density lipoprotein cholesterol.
years (17e19). In the context of diabetic dyslipidemia, the effect of GLP-1 receptor agonism on post-prandial lipoprotein production is of particular relevance (19). In addition, incretins have also been shown to exert a variety of direct cardiovascular effects (52). Meanwhile, several randomized controlled trials are underway examining the effect of various incretins on cardiovascular outcomes (53,54). Thus the outcome of these studies will provide insight into the relative role of lipid modifying effects of the study drugs on cardiovascular events.
Targeting HDL CETP inhibition. The physiological action of CETP of simultaneously raising HDL-C and lowering of LDL-C continues to make CETP inhibtion an attractive target despite the 2 previous failed attempts with torceptrapib and dalceptrapib, the former with serious off target effets and the latter with general lack of efficacy. Currently, 2 phase 3 trials are on-going evaluating CETP inhibitors anaceptrapib and evaceptrapib for their efficacies in reducing cardiovascular events in high-risk patients. Anaceptrapib: The safety and efficacy of anaceptrapib in highrisk patients were evaluated in an 18-month randomized, double blind, placebo controlled trial (55). In addition to achieving a 138% increase in HDL-C and a 39% reduction in LDL-C after treatment for 24 weeks, anaceptrapib treated subjects were free of adverse cardiovascular events. This study set the stage for the larger outcome trial “The Randomized EValuation of the Effects of Anacetrapib Through Lipid-modification (REVEAL) trial” aims to determine whether lipid modification with anacetrapib 100 mg daily reduces the risk of major coronary events in patients with circulatory problems who have their low-density lipoprotein (LDL) cholesterol level treated with a statin. ClinicalTrials.gov Identifier:NCT00685776.
Evaceptrapib: The purpose of the “Assessment of Clinical Effects of Cholesteryl Ester Transfer Protein Inhibition With Evacetrapib in Patients at a High-Risk for Vascular Outcomes” (ACCELERATE) study is to evaluate the efficacy and safety of evacetrapib in participants with high-risk vascular disease. Primary outcomes are the time to first occurrence of the composite endpoint of cardiovascular (CV) death, myocardial infarction (MI), stroke, coronary revascularization or hospitalization for unstable angina (UA). The REVEAL trial is expected to recruit 30 000 subjects and the ACCELERATE trial to recruit 11 000. Diabetes will be well represented in both trials, as it is one of the major recruitment criteria in both cases. Use of either agent will result in not only potent elevation of HDL-C but also significant reduction in LDL-C. The outcome of the 2 studies will address whether CETP is a viable target. ClinicalTrials.gov Identifier NCT01687998. ApoAI stimulator. In light of the selective avidity of nascent HDL in accepting ABCA1-mediated efflux of cellular cholesterol, it is of interest to test the strategy of apoA1 over-expression in the reversal of atherosclerotic heart disease. Homozygotes for apoAI mutations uniformly cause profound HDL deficiency, but their linkage to premature atherosclerotic heart disease is highly variable in the affected kindreds. That being the case, mutations resulting in inability to synthesize apoAI seem more likely to be associated with premature CVD (56). In mice, transgenic over-expression of apoAI has been shown to raise HDL-C and protects the susceptible animals to accelerated atherosclerosis (57). Furthermore, this model revealed that apoAI expression confers generalized antiinflammatory properties, including the vasculature as well as adipose tissue (58). Based on this hypothesis, a small molecule compound RSV-208 has been developed as a stimulator of apoAI gene expression for treatment of cardiovascular disease (59). Presently, 2 clinical studies are underway. The Study of Quantitative Serial Trends in Lipids with Apolipoprotein A-I Stimulation
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(SUSTAIN, NCT01423188) study aims to evaluate the lipid modifying efficacy, safety and tolerability of an apoA-I inducer RVX-208. The ApoA-I Synthesis Stimulation and Intravascular Ultrasound for Coronary Atheroma Regression Evaluation (ASSURE, NCT01067820) study aims to evaluate the effect of RVX-208 on plaque burden. These 2 studies will likely provide important proof of principle data to further explore apoA-I synthesis activation as a novel therapeutic strategy. If proven effective in the general population, this strategy will be attractive to counter the atherogenicity of diabetic dyslipidemia, in part through raising the nascent preb HDL, the most effective cholesterol acceptor in the efflux pathway. HDL infusion. Numerous preclinical and small human studies support the notion that synthetic reconstituted HDL resemble nascent HDL in their avidity in promoting efflux of cholesterol from peripheral tissues, and lesion macrophages in particular. Proof of principle studies had been reported both in animal models and in humans. ApoAI Milano, a recombinant HDL derived from combining the naturally occurring apoAI Milano mutant with phospholipids, was shown to effectively reduce atherosclerotic plaque volume after 5 weekly injections, beginning within 2 weeks of an acute coronary syndrome (ACS) event (60). Drug development program for clinical use is ongoing. Tardif et al reported an rHDL formulation using native apoAI and phosphotidylcholine (CSL111) in a larger trial, the Effect of rHDL on Atherosclerosis-Safety and efficacy (ERASE). One-hundred and eighty-three post-ACS patients were randomized to 4 weekly infusions of placebo or 1 of 2 doses of CSL-111 (61). The greater dose was discontinued because of a high incidence of hepatic enzyme elevation, but among the 136 patients with evaluable endpoint data, percent change in atheroma volume, the primary endpoint, improved significantly in the CSL-111 group but not in the placebo group. Extension of this line of therapeutic strategy includes several ongoing trials: 1) By using a second-generation CSL-based rHDL with apparently more efficacious cholesterol efflux induction, the study “A Single Ascending Dose Study Examining the Safety and Pharmacokinetic Profile of Reconstituted High Density Lipoprotein (CSL112) Administered to Patients,” (ClinicalTrials.gov Identifier: NCT01499420) is currently underway. 2) A related study entitled “Effect of CER-001 on Atherosclerosis in Acute Coronary Syndrome (ACS) Patients - Efficacy and Safety: The CHI SQUARE Trial” (ClinicalTrials.gov Identifier: NCT01201837) will assess the effects of CER-001, an ApoA-Ibased HDL mimetic, on indices of atherosclerotic plaque progression and regression as assessed by intravascular ultrasound (IVUS) measurements in patients with ACS. 3) The study entitled “Safety and Efficacy of APL180 in Healthy Volunteers and Patients With Coronary Heart Disease (CHD)” will assess the safety and tolerability, pharmacokinetics and effects on biomarkers of HDL function of APL180 after a single and 7-daily infusions in healthy volunteers and in patients with coronary heart disease (CHD) (ClinicalTrials.gov Identifier: NCT00568594). Evolving novel strategies Activation of ABCA1da target with an expanding scope of therapeutic benefits Numerous preclinical studies suggest that regulation of macrophage/foam cell specific ABCA1 and ABCG1 is sufficient to modulate atherogenesis, independent of circulating level of HDL (62). On the other hand, the role of hepatic derived ABCA1, though strongly influencing the HDL-C levels, in atherosclerosis seem less relevant (63). Effective pharmacological strategies to modulate ABCA1 and/or ABCG1 in combating atherosclerosis continue to be
elusive. Although activation of the transcription factor liver X receptor (LXR) has shown promise as an upstream target for its role in activating ABCA1 expression, drug development along this pathway was hampered by the concomitant activation of hepatic lipogenesis, resulting in significant hepatic steatosis and hypertriglyceridemia. Very recently, ABCA1 was found to be a specific target of micro RNA 33 (miR33) a/b. Mature miRNAs are w22-nucleotide singlestranded RNAs that exert their function via perfect Watson-Crick base pairing, with sequences most commonly located within the 3’untranslated region (3’-UTR) of target mRNAs. Interaction of a miRNA with its target mRNA results in inhibition of translation and/ or degradation of mRNAs. Preclinical studies revealed that inhibition of miR33 resulted in upregulation of Abca1, increased serum HDL-C (likely as a result of activation of hepatic ABCA1) and reduction in atherosclerotic burden (64). Interestingly, miR33 has also been shown to modulate ABCA1 expression and insulin secretory function of pancreatic beta cells. Taken together, inhibition of miR33 is a potentially promising therapeutic target to prevent atherosclerosis through promoting ABCA1 expression without the issue of HTG on the one hand, and prevent beta cell failure on the other. LCAT-related targetsdnew lessons from an old molecule that does not follow the script Traditionally, LCAT has been regarded as key mediator of the reverse cholesterol transport and modulator of HDL-C levels. The controversial, or sometimes paradoxical relationship between LCAT and atherosclerosis, has led to the increasing recognition of its complex physiological function in vivo and that this enzyme per se being an elusive therapeutic target. However, with the recent findings made in the rodent models of LCAT deficiency, a number of potential targetable metabolic pathways have emerged. A recent report by Hager et al (36) demonstrated that dietary cholesterol being an important promoter of hepatic ER stress, a cellular process known to mediate a network of metabolic-inflammatory responses, including insulin resistance. The deletion of LCAT in this experimental model protects the mice from ER stress by preventing accumulation of cholesterol in the ER, independent of cholesterol accumulation in other cellular compartments. This unexpected observation, therefore, raised the possibility for targeting ER cholesterol for preventing insulin resistance, through modulating the cellular cholesterol sensing machinery, namely the SREBP processing machinery. Additional research is required to further establish this experimental paradigm. Conclusion Diabetic dyslipidemia is a significant proatherogenic factor. It is postulated that the residual dyslipidemia, namely elevated TG and low HDL-C after maximal statin therapy, contribute significantly to the residual risk. However, effective therapeutic tools beyond statin treatment remain elusive. Our understanding of the pathophysiology of diabetic dyslipidemia continues to evolve. Novel regulatory factors of the high TG/low HDL phenotype continues to emerge. With the newer tools either undergoing clinical trials or at various stages of drug discovery, the current emerging development continues to raise hope to fill the therapeutic gap in cardiometabolic complications of diabetes. Acknowledgements This work was supported in part by an operating grant from the Canadian Institutes for Health Research (CIHR; MOP275369), a CIHR ChinaeCanada Joint Health Research Initiative grant and a Grant-in-aid (NA 6331) by the Heart and Stroke Foundation of Ontario.
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Author Disclosures None to disclose.
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