**3. New drugs for type 2 diabetes**

### **3.1. Currently available**

The Incretin hormones released by the gut, gastric inhibitory peptide (GIP) and Glucagon like peptide1 (GLP-1) (liraglutide) stimulate insulin secretion upon nutrient entry into the gut, suppression of glucagon release,slow gastric emptying and decrease food intake [128, 129]. Therefore, they have an antidiabetogenic potential. Incretin mimetics e.g. Exenatide LAR from exendin 4 is currently in use and most resistant to DPP4 degredation. GIP has also been shown enhancing β cell proliferation and inhibiting apoptosis in islet cell lines [130, 131]. Additionally functional GIP receptors have been identified on adipocytes and shown to stimulate glucose transport, accelerating fatty acid synthesis and stimulating lipoprotein lipase activity in animal models [131, 133, 134]. Several novel GIP analogues have been developed which act as stronger GIP agonists, showing resistance to degradation by Dipeptidyl Peptidase-4 (DIPP-4) [135] and demonstrating increased insulinotropic and blood glucose lowering activity [135]. Dipeptidyl peptidase Inhibitors (vildagliptin & sitagliptin) suppress breakdown of Glucagon like peptide1 (GLP-1) show great potential and are undergoing clinical testing. Antihyperglycemic synthetic analogs of amylin a hormone which are produced by the pancreas to lower blood sugar levels are available in injectable form and require close monitoring. Dapagliflozin a renal glucose reabsorption inhibitor reduces glycemic reabsorption independent of insulin, promises to be a new drug for type 2 diabetes treatment [136]. Testosterone replacement therapy in diabetic hypogonadal men decreases insulin resistance [137] probably by protective effect on pancreatic beta cells through its action on inflammatory cytokines [138].

#### **3.2. Experimental new drugs**

A vanadium and allixin based drug [139] and macrophage migration inhibitory factor MIF blocking inhibitory synthetic oral drug reducing blood sugar levels was tried in the mouse model and found to be effective in both the type 1&2 diabetc model [140]. Lisofylline, a fat metabolism inhibitor which prevents buildup of ceramide a by product of fat metabolism in mouse skeletal muscle decreased the insulin resistance and thus appears to be a novel new approach for type 2 diabetes [141]. It also has the ability to protect insulin producing cells by inhibiting cytokines produced by immune cells leading to apoptosis and cellular dysfunction and is thus effective in type1 diabetes [142]. LXR agonists have shown potential and require further testing in human and model systems [143]. Growth factors and protein kinase C inhibitors may act as innovative therapies for diabetic retinopathy [144].

#### **3.3. Surgical interventions**

Recently a type of gastric bypass surgery has been successful in normalizing blood sugar in a small number of normal to moderately obese type 2 diabetics [145, 146]. This surgery may possibly reduce death rate by 40% from all causes in morbidly obese people [147].

#### **3.4. Micronutrient approaches to treatment of diabetic complications**

People with diabetes have reduced antioxidant capacity which lays the basis for usage of antioxidant vitamins such as β carotene or vitamin C or E. A reduced level of ascorbic acid (Vitamin C) leaves the body more at mercy of the detrimental effects of aldose reductase, an enzyme responsible for many diabetic complications, such as cataracts and peripheral neuropathy [148]. Quercetin is another powerful aldose reductase inhibitor. It has been shown to inhibit aldose reductase by upto 50% [149]. Vitamin E is a free radical scavenger. It may play a preventive role in diabetic retinopathy by decreasing DAG levels, normalizing protein kinase C activation, normalizing blood flow in retinal and renal microvasculature and restoring NO mediated endothelium dependent relaxation [150, 151]. Renal and retinal vascular flows and responses were normalized in individuals who had diabetes of less than 10 years duration with high dose oral vitamin E therapy given for short periods while unchanged glycaemic control was observed [152].

Magnesium and chromium deficiency have been associated with poor diabetic control, insulin resistance, macro vascular disease and hypertension [153] and decreased glucose tolerance respectively [154]. Reduction of neuronal damage in diabetics by inhibiting glutamate dehydrogenase via vitamin B6 therapy has also been observed [155].N-Reduced glutathione precursor NAcetyl Cysteine is a gene expression and cellular metabolism modulating antiox‐

idant and its role in prevention of β cell oxidatory damage by acting as NFKB (a genetic regulator) inhibitor and subsequent deintensification of inflammatory responses is well documented [156]. Trace element vanadyl sulfate that behaves like insulin normalized hyperglycemic levels in diabetic animals and decreased the insulin need by upto 75% [157]. In human with Type 2 diabetes, low doses of vanadyl sulfate enhanced insulin responsive glucose uptake, glycogen production and decreased endogenous glucose formation. This resulted in reduced lipid oxidation and plasma free fatty acids levels [158]. Alpha lipoic Acid has powerful antioxidant activity, insulinomimetic action and provides protection from insulin resistance linked diabetic stress while improving glucose utilization [159]. Hyperglycemia reduction in diabetic rats was observed along with improvement in GSH levels with selenium therapy [160].Calcium AEP has benefited both type 1 and type 2 diabetics as it is alpha cell membrane integrity factor required for cellular membrane function. The hormone dehydroepiandroster‐ one (DHEA) undergoes a decrease in levels with aging that many researchers have linked to impair glucose metabolism. It was found to be as effective in reducing body fats and main‐ taining insulin responsiveness as exercise [161]. Thiamine is also now showing potential as therapy for type 2 diabetes.

**Figure 7.** Structure of thiamine diphosphate molecule.

are available in injectable form and require close monitoring. Dapagliflozin a renal glucose reabsorption inhibitor reduces glycemic reabsorption independent of insulin, promises to be a new drug for type 2 diabetes treatment [136]. Testosterone replacement therapy in diabetic hypogonadal men decreases insulin resistance [137] probably by protective effect on pancreatic

A vanadium and allixin based drug [139] and macrophage migration inhibitory factor MIF blocking inhibitory synthetic oral drug reducing blood sugar levels was tried in the mouse model and found to be effective in both the type 1&2 diabetc model [140]. Lisofylline, a fat metabolism inhibitor which prevents buildup of ceramide a by product of fat metabolism in mouse skeletal muscle decreased the insulin resistance and thus appears to be a novel new approach for type 2 diabetes [141]. It also has the ability to protect insulin producing cells by inhibiting cytokines produced by immune cells leading to apoptosis and cellular dysfunction and is thus effective in type1 diabetes [142]. LXR agonists have shown potential and require further testing in human and model systems [143]. Growth factors and protein kinase C

Recently a type of gastric bypass surgery has been successful in normalizing blood sugar in a small number of normal to moderately obese type 2 diabetics [145, 146]. This surgery may

People with diabetes have reduced antioxidant capacity which lays the basis for usage of antioxidant vitamins such as β carotene or vitamin C or E. A reduced level of ascorbic acid (Vitamin C) leaves the body more at mercy of the detrimental effects of aldose reductase, an enzyme responsible for many diabetic complications, such as cataracts and peripheral neuropathy [148]. Quercetin is another powerful aldose reductase inhibitor. It has been shown to inhibit aldose reductase by upto 50% [149]. Vitamin E is a free radical scavenger. It may play a preventive role in diabetic retinopathy by decreasing DAG levels, normalizing protein kinase C activation, normalizing blood flow in retinal and renal microvasculature and restoring NO mediated endothelium dependent relaxation [150, 151]. Renal and retinal vascular flows and responses were normalized in individuals who had diabetes of less than 10 years duration with high dose oral vitamin E therapy given for short periods while unchanged glycaemic

Magnesium and chromium deficiency have been associated with poor diabetic control, insulin resistance, macro vascular disease and hypertension [153] and decreased glucose tolerance respectively [154]. Reduction of neuronal damage in diabetics by inhibiting glutamate dehydrogenase via vitamin B6 therapy has also been observed [155].N-Reduced glutathione precursor NAcetyl Cysteine is a gene expression and cellular metabolism modulating antiox‐

possibly reduce death rate by 40% from all causes in morbidly obese people [147].

beta cells through its action on inflammatory cytokines [138].

inhibitors may act as innovative therapies for diabetic retinopathy [144].

**3.4. Micronutrient approaches to treatment of diabetic complications**

**3.2. Experimental new drugs**

36 Treatment of Type 2 Diabetes

**3.3. Surgical interventions**

control was observed [152].

Thiamine (termed aneurin or antineuritic vitamin initially) was the premier discovery of the B vitamins and thus ranked vitamin B1(Fig 7). It has relative temperature, acid stability and water solubility containing a pyrimidine ring and a thiazole nucleus linked with a methylene bridge. Thiamine is an essential micronutrient with a dietary reference intake (DRI) for normal healthy subjects of 1.1 mg/day for females and 1.3 mg/day for males [162]. Found in range of foodstuffs such as cereal grains. Its rich sources are brown rice, bran, oat meal, flax, poultry, egg yolks, beef, pork, liver, nuts, fruits and vegetables such as oranges, asparagus, kale, cauliflower, potatoes [163].UK law demands compulsary fortification of flour with thiamin of not less than 0.24mg/100g flour to replace losses during milling. In Pakistan no compulsory fortification is done and the general public consumes milled white flour which is easily available and probably thiamine deficient. Thiamine is naturally found in 4 forms in varying degrees of phosphorylation in TMP thiamine monophosphate, TPP thiamine pyrophosphate or diphosphate and TTP=thiamine triphosphate. It is commercially available as salt in its mononitrate HCl (also natural byproduct) and relatively inaccessible semi lipid soluble form S-acyl derivative benfotiamine and truly lipid soluble thiamine disulphide derivatives sulbutiamine and fursultiamine. Out of these, Thiamine HCl is the water soluble, easily accessible and commonly used vitamin supplement available with the trade name Benerva.

#### **3.5. Pharmacokinetics**

#### *3.5.1. Thiamine absorption in normal conditions*

Thiamine is released from its administered form by phosphatase and pyrophosphatase in the proximal part of the small intestine, following which absorption occurs mainly from this site with some from the stomach and the colon; thiamine absorbed in the colon may originate from intestinal microflora. Its absorption is hindered by alcohol consumption and folic acid deficiency [164].High affinity organic anion transporters THTR1 [165], THR2 for thiamine and reduced folate transporter RFC-1 transports both folic acid thiamine monophosphate (TMP) intracellularly [166, 168] at normal physiological concentrations. At high expression levels RFC1 also transports TPP out of the cells [167]. At higher concentrations thiamine crosses cell membranes in its open unionized form of the thiazolium ring even by passive diffusion.THTR2 is placed on the luminal surface of the gastrointestinal epithelial cells and THTR1 is on the basolateral surface mainly but not exclusively [169]. THTRI is expressed widely in human tisseues with particular high expression in skeletal muscles, placenta, heart liver and kidney [168, 170].Mutations in the SLC19A2 (D93H, S143F and G172D) cause malfunctioning of the thiamine transporter THTR1, thiamine deficiency and thiamine responsive megaloblastic anaemia (TRMA) [171, 172]. THTR2 is widely expressed most abundantly in placenta, kidney and liver [173]. Also highly expressed RFC-1, is in human tissues including mitochondrial membranes [168, 174]. It has affinities for TMP and TPP of 26µM and 32µM respectively [167, 168]. Cellular efflux is the probable reason for the presence of thiamine in plasma and cere‐ brospinal fluid [175-177].Thiamine in the glomerular filtrate is reabsorbed by the renal brush border membrane high affinity transporters where influx is increased by an outward directed H+gradient [178] RFC-1 is expressed on the apical and basolateral surface of the proximal tubular epithelial cells [179]; it may mediate the reuptake of TMP and provide a solution to the normal absence of TMP in the urine. Proton antiport membrane transport may operate in both intestinal and renal proximal tubular thiamine uptake [180]

#### *3.5.2. Assessment of thiamine status*

Erythrocytes contain approximately 90% of total thiamine in the blood and therefore conven‐ tionally their transketolase levels have generally been considered to be the measure of thiamine status in the body [181].Thiamine deficiency is assessed conventionally by measuring the percentage below complete saturation of the thiamine dependant enzyme transketolase (TK) in RBCs-"thiamine effect". The normal value of the thiamine effect in human subjects is in the range 0-15%, mild deficiency is 15-25% and severe thiamine deficiency >25% [182]. Latest research has however questioned its reliability as thiamine transporters THTR1 and RFC1 in erythrocytes are upregulated in thiamine deficieny and RBC TK levels are not decreased in tandem [183]. Furthermore it doesn't account for changes in TK expression in RBC and other precursor cells. The expression of TK is decreased in thiamine deficiency [184]. Currently assessment of mononuclear TK activity and plasma thiamine concentration determination using HPLC flourimetric determination with respect to normal healthy controls gives greater insight into thiamine status [185-187]. More recently in capillary enzyme reaction and capillary electrophoresis methods are emerging as potential alternative monitoring and determining techniques for thiamine in samples [188].

### *3.5.3. Thiamine metabolism within the cells*

sulbutiamine and fursultiamine. Out of these, Thiamine HCl is the water soluble, easily accessible and commonly used vitamin supplement available with the trade name Benerva.

Thiamine is released from its administered form by phosphatase and pyrophosphatase in the proximal part of the small intestine, following which absorption occurs mainly from this site with some from the stomach and the colon; thiamine absorbed in the colon may originate from intestinal microflora. Its absorption is hindered by alcohol consumption and folic acid deficiency [164].High affinity organic anion transporters THTR1 [165], THR2 for thiamine and reduced folate transporter RFC-1 transports both folic acid thiamine monophosphate (TMP) intracellularly [166, 168] at normal physiological concentrations. At high expression levels RFC1 also transports TPP out of the cells [167]. At higher concentrations thiamine crosses cell membranes in its open unionized form of the thiazolium ring even by passive diffusion.THTR2 is placed on the luminal surface of the gastrointestinal epithelial cells and THTR1 is on the basolateral surface mainly but not exclusively [169]. THTRI is expressed widely in human tisseues with particular high expression in skeletal muscles, placenta, heart liver and kidney [168, 170].Mutations in the SLC19A2 (D93H, S143F and G172D) cause malfunctioning of the thiamine transporter THTR1, thiamine deficiency and thiamine responsive megaloblastic anaemia (TRMA) [171, 172]. THTR2 is widely expressed most abundantly in placenta, kidney and liver [173]. Also highly expressed RFC-1, is in human tissues including mitochondrial membranes [168, 174]. It has affinities for TMP and TPP of 26µM and 32µM respectively [167, 168]. Cellular efflux is the probable reason for the presence of thiamine in plasma and cere‐ brospinal fluid [175-177].Thiamine in the glomerular filtrate is reabsorbed by the renal brush border membrane high affinity transporters where influx is increased by an outward directed H+gradient [178] RFC-1 is expressed on the apical and basolateral surface of the proximal tubular epithelial cells [179]; it may mediate the reuptake of TMP and provide a solution to the normal absence of TMP in the urine. Proton antiport membrane transport may operate in

**3.5. Pharmacokinetics**

38 Treatment of Type 2 Diabetes

*3.5.1. Thiamine absorption in normal conditions*

both intestinal and renal proximal tubular thiamine uptake [180]

Erythrocytes contain approximately 90% of total thiamine in the blood and therefore conven‐ tionally their transketolase levels have generally been considered to be the measure of thiamine status in the body [181].Thiamine deficiency is assessed conventionally by measuring the percentage below complete saturation of the thiamine dependant enzyme transketolase (TK) in RBCs-"thiamine effect". The normal value of the thiamine effect in human subjects is in the range 0-15%, mild deficiency is 15-25% and severe thiamine deficiency >25% [182]. Latest research has however questioned its reliability as thiamine transporters THTR1 and RFC1 in erythrocytes are upregulated in thiamine deficieny and RBC TK levels are not decreased in tandem [183]. Furthermore it doesn't account for changes in TK expression in RBC and other precursor cells. The expression of TK is decreased in thiamine deficiency [184]. Currently

*3.5.2. Assessment of thiamine status*

When TMP enters cells by RFC-1 it is hydrolyzed to thiamine by phosphatases [168]. Thiamine deficiency decreased the activity of TPPK [189] and was implicated in decreased hepatic levels of TPP with normal levels of thiamine in STZ diabetic rats [190].Within mitochondria TPP is slowly hydrolyzed to TMP by phosphatases which may leave the mitochondria via the same transporter. High concentrations of thiamine monophosphate inhibit thiamine pyrophspho‐ kinase activity noncompetitively [191] and inhibit the entry of TPP into the mitochondria competitively [189]. A small amount of TPP is further phosphorylated to thiamine triphosphate (TTP) by thiamine pyrophosphate kinase and hydrolyzed to TPP by TPPphosphatase [192] [168]. Plasma half life is relatively short (2days) [193] but its tissue half life is approximately 9-18 days [194]. Thiamine is stored largely in skeletal muscle and the highly perfused organs such as heart, brain, liver and kidneys [163]. Subcellularly only 10% of total TPP is available for binding to transketolase most of it is associated with the mitochondria [185].Thiamine and its acid metabolites are are excreted primarily in the urine [195].
