Myeloid Heterogeneity Mediates Acute Exacerbations of Pulmonary Fibrosis.

Epidemiological evidence indicates that exposure to particulate matter is linked to the development of idiopathic pulmonary fibrosis (IPF) and increases the incidence of acute exacerbations of IPF. In addition to accelerating the rate of lung function decline, exposure to fine particulate matter (particulate matter smaller than 2.5 µm [PM2.5]) is a risk factor for increased mortality in subjects with IPF. In this article, we show that exposure to PM2.5 mediates monocyte recruitment and fibrotic progression in mice with established fibrosis. In mice with established fibrosis, bronchoalveolar lavage cells showed monocyte/macrophage heterogeneity after exposure to PM2.5. These cells had a significant inflammatory and anti-inflammatory signature. The mixed heterogeneity of cells contributed to the proinflammatory and anti-inflammatory response. Although monocyte-derived macrophages were recruited to the lung in bleomycin-injured mice treated with PM2.5, recruitment of monocytes expressing Ly6Chi to the lung promoted progression of fibrosis, reduced lung aeration on computed tomography, and impacted lung compliance. Ly6Chi monocytes isolated from PM2.5-exposed fibrotic mice showed enhanced expression of proinflammatory markers compared with fibrotic mice exposed to vehicle. Moreover, IPF bronchoalveolar lavage cells treated ex vivo with PM2.5 showed an exaggerated inflammatory response. Targeting Ly6Chi monocyte recruitment inhibited fibrotic progression in mice. Moreover, the adoptive transfer of Ly6Chi monocytes exacerbated established fibrosis. These observations suggest that enhanced recruitment of Ly6Chi monocytes with a proinflammatory phenotype mediates acute exacerbations of pulmonary fibrosis, and targeting these cells may provide a potential novel therapeutic target to protect against acute exacerbations of IPF.

NOX4-TIM23 interaction regulates NOX4 mitochondrial import and metabolic reprogramming.

Pulmonary fibrosis is a progressive lung disease characterized by macrophage activation. Asbestos-induced expression of nicotinamide adenine dinucleotide phosphate hydrogen oxidase 4 (NOX4) in lung macrophages mediates fibrotic progression by the generation of mitochondrial reactive oxygen species (ROS), modulating mitochondrial biogenesis, and promoting apoptosis resistance; however, the mechanism(s) by which NOX4 localizes to mitochondria during fibrosis is not known. Here, we show that NOX4 localized to the mitochondrial matrix following asbestos exposure in lung macrophages via direct interaction with TIM23. TIM23 and NOX4 interaction was found in lung macrophages from human subjects with asbestosis, while it was absent in mice harboring a conditional deletion of NOX4 in lung macrophages. This interaction was localized to the proximal transmembrane region of NOX4. Mechanistically, TIM23 augmented NOX4-induced mitochondrial ROS and metabolic reprogramming to oxidative phosphorylation. Silencing TIM23 decreased mitochondrial ROS and oxidative phosphorylation. These observations highlight the important role of the mitochondrial translocase TIM23 interaction with NOX4. Moreover, this interaction is required for mitochondrial redox signaling and metabolic reprogramming in lung macrophages.

Tinosporaside from Tinospora cordifolia Encourages Skeletal Muscle Glucose Transport through Both PI-3-Kinase- and AMPK-Dependent Mechanisms.

The stem of Tinospora cordifolia has been traditionally used in traditional Indian systems of medicine for blood sugar control, without the knowledge of the underlying mechanism and chemical constitution responsible for the observed anti-diabetic effect. In the present study, Tinosporaside, a diterpenoid isolated from the stem of T. cordifolia, was investigated for its effects on glucose utilization in skeletal muscle cells, which was followed by determining the anti-hyperglycemic efficacy in our diabetic db/db mice model. We found that tinosporaside augmented glucose uptake by increasing the translocation of GLUT4 to the plasma membrane in L6 myotubes, upon prolonged exposure for 16 h. Moreover, tinosporaside treatment significantly increased the phosphorylation of protein kinase B/AKT (Ser-473) and 5' AMP-activated protein kinase (AMPK, Thr-172). These effects were abolished in the presence of the wortmannin and compound C. Administration of tinosporaside to db/db mice improved glucose tolerance and peripheral insulin sensitivity associated with increased gene expression and phosphorylation of the markers of phosphoinositide 3-kinases (PI3Ks) and AMPK signaling in skeletal muscle tissue. The findings revealed that tinosporaside exerted its antidiabetic efficacy by enhancing the rate of glucose utilization in skeletal muscle, mediated by PI3K- and AMPK-dependent signaling mechanisms.

ß-Sitosterol-D-Glucopyranoside Mimics Estrogenic Properties and Stimulates Glucose Utilization in Skeletal Muscle Cells.

Estrogenic molecules have been reported to regulate glucose homeostasis and may be beneficial for diabetes management. Here, we investigated the estrogenic effect of ß-sitosterol-3-O-D-glucopyranoside (BSD), isolated from the fruits of Cupressus sempervirens and monitored its ability to regulate glucose utilization in skeletal muscle cells. BSD stimulated ERE-mediated luciferase activity in both ERα and ERß-ERE luc expression system with greater response through ERß in HEK-293T cells, and induced the expression of estrogen-regulated genes in estrogen responsive MCF-7 cells. In silico docking and molecular interaction studies revealed the affinity and interaction of BSD with ERß through hydrophobic interaction and hydrogen bond pairing. Furthermore, prolonged exposure of L6-GLUT4myc myotubes to BSD raised the glucose uptake under basal conditions without affecting the insulin-stimulated glucose uptake, the effect associated with enhanced translocation of GLUT4 to the cell periphery. The BSD-mediated biological response to increase GLUT4 translocation was obliterated by PI-3-K inhibitor wortmannin, and BSD significantly increased the phosphorylation of AKT (Ser-473). Moreover, BSD-induced GLUT4 translocation was prevented in the presence of fulvestrant. Our findings reveal the estrogenic activity of BSD to stimulate glucose utilization in skeletal muscle cells via PI-3K/AKT-dependent mechanism.

Naturally Occurring Carbazole Alkaloids from Murraya koenigii as Potential Antidiabetic Agents.

This study identified koenidine (4) as a metabolically stable antidiabetic compound, when evaluated in a rodent type 2 model (leptin receptor-deficient db/db mice), and showed a considerable reduction in the postprandial blood glucose profile with an improvement in insulin sensitivity. Biological studies were directed from the preliminary in vitro evaluation of the effects of isolated carbazole alkaloids (1-6) on glucose uptake and GLUT4 translocation in L6-GLUT4myc myotubes, followed by an investigation of their activity (2-5) in streptozotocin-induced diabetic rats. The effect of koenidine (4) on GLUT4 translocation was mediated by the AKT-dependent signaling pathway in L6-GLUT4myc myotubes. Moreover, in vivo pharmacokinetic studies of compounds 2 and 4 clearly showed that compound 4 was 2.7 times more bioavailable than compound 2, resulting in a superior in vivo efficacy. Therefore, these studies suggested that koenidine (4) may serve as a promising lead natural scaffold for managing insulin resistance and diabetes.

Murraya koenigii (L.) Spreng. ameliorates insulin resistance in dexamethasone-treated mice by enhancing peripheral insulin sensitivity.

BACKGROUND: Murraya koenigii (L.) Spreng. is an important medicinal plant used traditionally as an antiemetic, antidiarrhoeal agent and blood purifier and as a medicine for a variety of ailments. This study investigated the effects of ethanolic extract of M. koenigii (MK) on diabetes-associated insulin resistance induced in mice by chronic low-dose injection of dexamethasone. RESULTS: Mice treated with dexamethasone exhibited hyperglycaemia and impaired glucose tolerance. Treatment with MK reduced the extent of dexamethasone-induced hyperglycaemia and decreased insulin resistance as indicated by improved glucose tolerance and increased insulin-stimulated AKT phosphorylation in skeletal muscle tissue. Further evaluation in clonal skeletal muscle cell lines suggested that MK increased glucose uptake in L6 skeletal muscle cells by increasing cell surface GLUT4 density via an AKT-mediated pathway. CONCLUSION: MK can ameliorate dexamethasone-induced hyperglycaemia and insulin resistance in part by increasing glucose disposal into skeletal muscle.

Comparative evaluation of liquid-liquid extraction and nanosorbent extraction for HPLC-PDA analysis of cabazitaxel from rat plasma.

A precise, sensitive, accurate, and validated reverse-phase high-performance liquid chromatography (RP-HPLC) method with a bioanalytical approach was utilized to analyze Cabazitaxel (CBZ) in rat plasma. Comparative research on extraction recoveries was performed between traditional liquid-liquid extraction (LLE) and synthesized graphene oxide (GO) based magnetic solid phase extraction (GO@MSPE). The superparamagnetic hybrid nanosorbent was synthesized using the combination of iron oxide and GO and subsequently applied for extraction and bioanalytical quantification of CBZ from plasma by (HPLC-PDA) analysis. Fourier- transform infrared spectroscopy (FT-IR), particle size, scanning electron microscopy (SEM), and x-ray diffraction (XRD) analysis were employed in the characterization of synthesized GO@MSPE nanosorbent. The investigation was accomplished using a shim pack C18 column (150 mm×4.6 mm, 5 µm) with a binary gradient mobile phase consisting of formic acid: acetonitrile: water (0.1:75:25, v/v/v) at a 0.8 mL/min flow rate, and a λmax of 229 nm. The limits of detection (LOD) and quantitation (LOQ) have been determined to be 50 and 100 ng/mL for both LLE and SPE techniques. The linearity range of the approach encompassed from 100 to 5000 ng/mL and was found to be linear (coefficient of determination > 0.99) for CBZ. The proposed method showed extraction recovery of 76.8-88.4% for the synthesized GO@MSPE and 69.3-77.4% for LLE, suggesting that the proposed bioanalytical approach was robust and qualified for all validation parameters within the acceptable criteria. Furthermore, the developed hybrid GO@MSPE nanosorbent with the help of the proposed RP-HPLC method, showed a significant potential for the extraction of CBZ in bioanalysis.

Cell Adhesion Molecules in Fibrotic Diseases.

Mechanisms underlying the pathogenesis of tissue fibrosis remain incompletely understood. Emerging evidence suggests that cell adhesion molecules (CAMs) are critical in fibrotic progression in many organs, including lung, kidney, skin, and liver. CAMs promote cell-cell and cell-extracellular matrix (ECM) interactions to maintain tissue architecture and normal function in homeostasis. However, dysregulated expression and function of CAMs can lead to chronic inflammation and tissue fibrosis. The major families of CAMs include integrins, cadherins, selectins, and immunoglobulins. Here, we review the role of the CAMs in fibrosis development across various organs with a focus on integrins and cadherins, and discuss their respective roles in the development of pulmonary fibrosis.

Design, synthesis, and evaluation of benzofuran-based chromenochalcones for antihyperglycemic and antidyslipidemic activities.

A series of benzofuran-based chromenochalcones (16-35) were synthesized and evaluated for in vitro and in vivo antidiabetic activities in L-6 skeletal muscle cells and streptozotocin (STZ)-induced diabetic rat models, respectively, and further in vivo dyslipidemia activity of the compounds was evaluated in a Triton-induced hyperlipidemic hamster model. Among them, compounds 16, 18, 21, 22, 24, 31, and 35 showed significant glucose uptake stimulatory effects in skeletal muscle cells and were further evaluated for in vivo efficacy. Compounds 21, 22, and 24 showed a significant reduction in blood glucose levels in STZ-induced diabetic rats. Compounds 16, 20, 21, 24, 28, 29, 34, 35, and 36 were found active in antidyslipidemic studies. Furthermore, compound 24 effectively improved the postprandial and fasting blood glucose levels, oral glucose tolerance, serum lipid profile, serum insulin level, and the HOMA-index of db/db mice, following 15 days of successive treatment.

Assessment of the dual role of Lyonia ovalifolia (Wall.) Drude in inhibiting AGEs and enhancing GLUT4 translocation through LC-ESI-QTOF-MS/MS determination and in silico studies.

Introduction: Diabetes mellitus (DM) is a metabolic disorder that results in glucose accumulation in the blood, accompanied by the production of advanced glycation end products (AGEs) through glycation of cellular proteins. These AGEs interfere with insulin signaling and prevent GLUT4 membrane translocation, thereby promoting the accumulation of more glucose in the blood and causing post-diabetic complications. Methods: In this study, we examine the anti-diabetic potential of Lyonia ovalifolia (Wall.) Drude, a well-known ethnomedicinal plant of the Indian Himalayas. Considering its various medicinal properties, we analyzed its ethanolic extract and various solvent fractions for in vitro antiglycation activity and antidiabetic potential, i.e., stimulation of GLUT4 translocation. Result and Discussions: The results showed that the extract and fractions exhibited increased antiglycation activity and an increased level of GLUT4 translocation. Analysis of a further 12 bioactive compounds of ethanolic extract, identified through LC-ESI-QTOF-MS/MS, revealed the presence of three new compounds: leucothol B, rhodoterpenoids A, and leucothol A. Moreover, we performed molecular docking of identified compounds against key proteins of diabetes mellitus: the sirtuin family of NAD (+)-dependent protein deacetylases 6 (SIRT6), aldose reductase (AR), and tyrosine kinase (TK). The results showed that flavonoid luteolin showed the best binding affinity ((-12.3 kcal/mol), followed by eriodictyol, astilbin, and syringaresinol. An ADMET study showed that luteolin, eriodictyol, astilbin, and syringaresinol may be promising drug candidates belonging to the flavonoid class of compounds, with no harmful effects and complying with all the drug-likeness guidelines. Furthermore, molecular dynamics (MD) simulations on a 50 ns timescale revealed that AR protein was most stable with luteolin throughout the simulation period. Therefore, this study reveals for the first time that L. ovalifolia plays an important role in insulin homeostasis, as shown in in vitro and in silico studies.

Synthesis of novel imbricatolic acid analogues via insertion of N-substituted piperazine at C-15/C-19 positions, displaying glucose uptake stimulation in L6 skeletal muscle cells.

A new class of N-substituted piperazine analogues of imbricatolic acid have been designed and synthesized by using the appropriate synthetic routes in excellent yield. All synthesised compounds were screened for their in vitro glucose uptake stimulatory activity. Among them compounds 4b, 4e, 8b, and 8e triggered L6 skeletal muscle cells for glucose uptake at 54.73%, 40.79%, 40.90%, and 39.55% stimulation, respectively. Compound 4b has emerged as important lead compound showing potential antidiabetic activity. Illustration about their synthesis and in vitro glucose uptake activity is described.

In vitro, In vivo and In silico Antihyperglycemic Activity of Some Semi-Synthetic Phytol Derivatives.

BACKGROUND: Due to the prevalence of type-2 diabetes across the globe, there is an unmet need to explore new molecular targets for the development of cost-effective and safer antihyperglycemic agents. OBJECTIVE: Structural modification of phytol and evaluation of in vitro, in vivo and in silico antihyperglycemic activity of derivatives establishing the preliminary structure activity relationship (SAR). METHODS: The semi-synthetic derivatives of phytol were prepared following previously described methods. The antihyperglycemic potential was measured in vitro in terms of increase in 2- deoxyglucose (2-DG) uptake by L-6 rat skeletal muscle cells as well as in vivo in sucrose-loaded (SLM) and streptozotocin (STZ)-induced diabetic rat models. The blood glucose profile was measured at 30, 60, 90, 120, 180, 240, 300 and 1440 min post administration of sucrose in rats. The in silico docking was performed on peroxisome proliferator-activated receptor gamma (PPARγ) as antidiabetic target along with absorption, distribution, metabolism, excretion and toxicity (ADMET) studies. RESULTS: Nine semi-synthetic ester derivatives: acetyl (1), lauroyl (2), palmitoyl (3), pivaloyl (4), trans-crotonyl (5), benzoyl (6), m-anisoyl (7), 3,4,5-trimethoxy benzoyl (8) cinnamoyl (9) along with bromo derivative (10) of phytol were prepared. The derivatives 9, 8 and 2 caused 4.5, 3.2 and 2.7 times more in vitro uptake of 2-DG respectively than rosiglitazone (ROSI). The derivatives showed significant improvement in oral glucose tolerance both in SLM (29.6-21%) as well as STZ-induced diabetic (30.8-19.0%) rats. The in silico ADMET, docking studies showed non-toxicity and high binding affinity with PPARγ. CONCLUSION: The potent antihyperglycemic activity with favorable pharmacokinetics supports phytol derivatives as a suitable antidiabetic lead.

SGLT2 inhibitors for the treatment of diabetes: a patent review (2013-2018).

INTRODUCTION: The sodium-glucose co-transporter 2 (SGLT2) is ascribed to target renal tubular glucose re-absorption, and its inhibition has been proved to induce glucosuria which improves the glycemic index. Accordingly, SGLT2 inhibitors have found to be the promising class of antidiabetic agents for the management of type 2 diabetes mellitus. A large number of SGLT2 inhibitors have developed through structural modification and investigated for their ability to selectivity inhibit SGLT2 transporters with better bioavailability. AREAS COVERED: This review comprises a summary of patent applications (2013-2018) of SGLT2 inhibitors with focus on chemical structural advancement and therapeutic potentials in the management of diabetes and related disorders. EXPERT OPINION: SGLT2 inhibitors exert multiple metabolic benefits, including reduced glycated hemoglobin (HbA1c), improved glycemic control (fasting and postprandial), reduced body weight, reduced systolic and diastolic blood pressure and improved HDL cholesterol. Due to the virtue of no interference with insulin action and secretion, their efficacy remains the same even in presence of progressive ß cell failure in type 2 diabetes. Additionally, few members of this class have been reported to exhibit cardioprotective, renoprotective, and anticancer activity. However, more study on the long-term outcomes in patients taking SGLT2 inhibitors is warranted.

SVMDLF: A novel R-based Web application for prediction of dipeptidyl peptidase 4 inhibitors.

Dipeptidyl peptidase 4 (DPP4) is a well-known target for the antidiabetic drugs. However, currently available DPP4 inhibitor screening assays are costly and labor-intensive. It is important to create a robust in silico method to predict the activity of DPP4 inhibitor for the new lead finding. Here, we introduce an R-based Web application SVMDLF (SVM-based DPP4 Lead Finder) to predict the inhibitor of DPP4, based on support vector machine (SVM) model, predictions of which are confirmed by in vitro biological evaluation. The best model generated by MACCS structure fingerprint gave the Matthews correlation coefficient of 0.87 for the test set and 0.883 for the external test set. We screened Maybridge database consisting approximately 53,000 compounds. For further bioactivity assay, six compounds were shortlisted, and of six hits, three compounds showed significant DPP4 inhibitory activities with IC50 values ranging from 8.01 to 10.73 µm. This application is an OpenCPU server app which is a novel single-page R-based Web application for the DPP4 inhibitor prediction. The SVMDLF is freely available and open to all users at http://svmdlf.net/ocpu/library/dlfsvm/www/ and http://www.cdri.res.in/svmdlf/.

Multiple machine learning based descriptive and predictive workflow for the identification of potential PTP1B inhibitors.

In insulin and leptin signaling pathway, Protein-Tyrosine Phosphatase 1B (PTP1B) plays a crucial controlling role as a negative regulator, which makes it an attractive therapeutic target for both Type-2 Diabetes (T2D) and obesity. In this work, we have generated classification models by using the inhibition data set of known PTP1B inhibitors to identify new inhibitors of PTP1B utilizing multiple machine learning techniques like naïve Bayesian, random forest, support vector machine and k-nearest neighbors, along with structural fingerprints and selected molecular descriptors. Several models from each algorithm have been constructed and optimized, with the different combination of molecular descriptors and structural fingerprints. For the training and test sets, most of the predictive models showed more than 90% of overall prediction accuracies. The best model was obtained with support vector machine approach and has Matthews Correlation Coefficient of 0.82 for the external test set, which was further employed for the virtual screening of Maybridge small compound database. Five compounds were subsequently selected for experimental assay. Out of these two compounds were found to inhibit PTP1B with significant inhibitory activity in in-vitro inhibition assay. The structural fragments which are important for PTP1B inhibition were identified by naïve Bayesian method and can be further exploited to design new molecules around the identified scaffolds. The descriptive and predictive modeling strategy applied in this study is capable of identifying PTP1B inhibitors from the large compound libraries.

Synthesis of substituted 2H-benzo[e]indazole-9-carboxylate as a potent antihyperglycemic agent that may act through IRS-1, Akt and GSK-3ß pathways.

Based on high throughput screening of our chemical library, we identified two 4,5-dihydro-2H-benzo[e]indazole derivatives (5d and 5g), which displayed a significant effect on glucose uptake in L6 skeletal muscle cells. Based on these lead molecules, a series of benzo[e]indazole derivatives were prepared. Among all the synthesized dihydro-2H-benzo[e]indazoles, 8-(methylthio)-2-phenyl-6-p-tolyl-4,5-dihydro-2H-benzo[e]indazole-9-carboxylate (5e) showed significant glucose uptake stimulation in L6 skeletal muscle cells, even better than lead compounds. Additionally, 5e decreased glucagon-induced glucose release in HepG2 hepatoma cells. The 2H-benzo[e]indazole 5e exerted an antihyperglycemic effect in normal, sucrose challenged streptozotocin-induced diabetic rats and type 2 diabetic db/db mice. Treatment with 5e at a dose of 30 mg kg-1 in db/db mice caused a significant decrease in triglyceride and total cholesterol levels and increased the HDL-C level in a significant manner. The mechanistic studies revealed that the 2H-benzo[e]indazole 5e significantly stimulated insulin-induced signaling at the level of IRS-1, Akt and GSK-3ß in L6 skeletal muscle cells, possibly by inhibiting protein tyrosine phosphatase-1B. This new 2H-benzo[e]indazole derivative has potential for the treatment of diabetes with improved lipid profile.

In vitro anti-hyperglycemic activity of 4-hydroxyisoleucine derivatives.

The nonproteinogenic amino acid, 4-hydroxyisoleucine (1) has been isolated in large quantities from the fenugreek (T. foenum-graecum) seeds. Few novel derivatives (3-11 and 13-18) were prepared from the naturally occurring 4-hydroxyisoleucine (1) and screened for their in vitro glucose uptake stimulatory effect in L-6 skeletal muscle cells. The derivatives 6, 7, 8, 10 and 11 exhibited better glucose uptake stimulatory activity than parent compound, 4-hydroxyisoleucine at 5 and 10µM concentrations and compounds 7 and 11 enhanced translocation of insulin sensitive glucose transporters-4 in skeletal muscle cells.

NOD2 activation induces oxidative stress contributing to mitochondrial dysfunction and insulin resistance in skeletal muscle cells.

Nucleotide-binding oligomerization domain protein-2 (NOD2) activation in skeletal muscle cells has been associated with insulin resistance, but the underlying mechanisms are not yet clear. Here we demonstrate the implication of oxidative stress in the development of mitochondrial dysfunction and insulin resistance in response to NOD2 activation in skeletal muscle cells. Treatment with the selective NOD2 ligand muramyl dipeptide (MDP) increased mitochondrial reactive oxygen species (ROS) generation in L6 myotubes. MDP-induced ROS production was associated with increased levels of protein carbonyls and reduction in citrate synthase activity, cellular ATP level, and mitochondrial membrane potential, as well as altered expression of genes involved in mitochondrial function and metabolism. Antioxidant treatment attenuated MDP-induced ROS production and restored mitochondrial functions. In addition, the presence of antioxidant prevented NOD2-mediated activation of MAPK kinases and the inflammatory response. This was associated with reduced serine phosphorylation of insulin receptor substrate-1 (IRS-1) and improved insulin-stimulated tyrosine phosphorylation of IRS-1 and downstream activation of Akt phosphorylation. These data indicate that oxidative stress plays a role in NOD2 activation-induced inflammatory response and that MDP-induced oxidative stress correlates with impairment of mitochondrial functions and induction of insulin resistance in skeletal muscle cells.

Synthesis of heteroaryl/aryl kojic acid conjugates as stimulators of glucose uptake by GLUT4 translocation.

Insulin exerts its metabolic actions through the insulin receptor (IR) and plays an essential role in treatment of diabetes. The inconvenience of daily injections and the undesirable side-effects associated with insulin injections demand novel drugs for the disease. To search for bioactive insulin mimetic, we synthesized a chemical library of small molecules (2a-3f) based on the indolylkojic acid scaffold (B). An In vitro screening assay was performed to stimulate glucose transport in rat L6 skeletal muscle cells, post treatment of the compounds (2a-3f) for the time period incubation of 16 h. Compounds 2f, 2g, 2l, 3a, 3b, 3c and 3d have shown significant glucose uptake stimulation as compared to the controls at micromolar concentrations. In mechanistic studies, we observed that these compounds exert their biological action by enhancing GLUT4 translocation to cell surface via PI3K-dependent signalling pathway in agreement to the insulin mode of action. Hence, these promising conjugates should be useful for further drug development in diabetes treatment.