1-Department of Pharmacy, Comsats University, Islamabad.
2- School of Natural Sciences, Department of Chemistry, NUST.
3- Lecturer, Department of Pharmacy, Abbottabad university Of Science & Technology (AUST), Abbottabad.
4-Department of Zoology, Hazara University, Mansehra.
5- Azure & Modern Workplace Solutions Specialist at Microsoft
The study aimed to investigate six thiourea derivatives (WT-1 to WT-6, where WT denotes Waleed Thiourea) for their potential as antidiabetic agents. Comprehensive evaluations were conducted, including in vitro, in vivo, and in silico approaches. Among the synthesized compounds, WT-2 and WT-4 emerged as the lead candidates based on their superior biological activity and safety profiles. In vitro DPP4 enzyme inhibition assays revealed IC?? values of 2.31 ± 0.05 µM and 1.98 ± 0.04 µM for WT-2 and WT-4, respectively, outperforming other derivatives and approaching the activity of the standard drug (IC??: 1.45 ± 0.03 µM). In vivo studies on diabetic mice demonstrated that WT-2 and WT-4 reduced blood glucose levels by 61% and 66%, respectively, at a dose of 20 mg/kg, with significant HbA1c reductions (55% and 58%, respectively) after two weeks. Histopathological and biochemical analyses confirmed no signs of toxicity, with ALT, creatinine, and CRP levels within normal ranges. Molecular docking studies showed strong binding affinities of WT-2 (-8.65 kcal/mol) and WT-4 (-8.98 kcal/mol) to critical residues of the DPP4 enzyme, including TYR547, ARG358, and GLU206. In silico pharmacokinetics and toxicology profiles highlighted favorable drug-like properties, high gastrointestinal absorption, and the absence of major toxicity risks for WT-2 and WT-4. These findings underscore the therapeutic potential of WT-2 and WT-4 as safe and effective antidiabetic agents, warranting further preclinical and clinical studies. From these studies its clear that WT-2 & WT-4 are the most potential lead compounds as antidiabetic agents.
Diabetes mellitus (DM) is a chronic metabolic disorder characterized by persistent hyperglycemia due to impaired insulin secretion, insulin action, or both. It is a global health challenge, with an estimated 537 million adults affected as of 2021, projected to rise to 783 million by 2045 (International Diabetes Federation, 2021). DM significantly impairs quality of life and increases mortality due to its association with a range of complications, including cardiovascular disease, nephropathy, neuropathy, and retinopathy (American Diabetes Association, 2023). These complications highlight the urgency of developing more effective and safer therapeutic agents. The chronic hyperglycemia of diabetes causes damage to multiple organ systems. Microvascular complications, such as diabetic retinopathy, nephropathy, and neuropathy, can lead to blindness, kidney failure, and debilitating pain, respectively (Forbes & Cooper, 2013). Macrovascular complications, including coronary artery disease, peripheral artery disease, and stroke, further exacerbate the burden of diabetes (Beckman et al., 2002). Managing these complications is complex, necessitating multifaceted approaches to mitigate their progression and improve patient outcomes. Existing antidiabetic therapies include insulin, sulfonylureas, biguanides, thiazolidinediones, DPP-4 inhibitors, and GLP-1 receptor agonists. While effective to varying degrees, these therapies have limitations such as side effects, patient non-adherence, and diminishing efficacy over time (Nathan et al., 2009). For example, insulin therapy can lead to hypoglycemia and weight gain, while metformin may cause gastrointestinal disturbances (Bailey, 2017). Thus, there is a pressing need for alternative treatments that are safer, more effective, and better tolerated. Thiourea (CH4N2S) is an organosulfur compound known for its versatile chemical properties, including hydrogen bonding and metal chelation. These properties have made it an attractive scaffold for drug design (Bhuvaneshwari et al., 2020). Recent studies suggest that thiourea derivatives exhibit promising biological activities, including antidiabetic effects. Their ability to inhibit DPP-4, a key enzyme in glucose homeostasis, positions them as potential candidates for diabetes management (Kim et al., 2022). Dipeptidyl peptidase-4 (DPP-4) inhibitors play a crucial role in enhancing incretin hormone activity, thereby improving glucose-dependent insulin secretion and reducing glucagon secretion. Thiourea derivatives have shown potent DPP-4 inhibitory activity due to their ability to form stable interactions with the enzyme's active site (Ramesh et al., 2021). Compared to existing DPP-4 inhibitors, thiourea-based compounds exhibit improved selectivity and reduced side effects, making them promising candidates for further development. In-vitro studies are essential for evaluating the preliminary efficacy and safety of thiourea derivatives. These studies enable researchers to investigate the compounds’ DPP-4 inhibitory activity, antioxidant properties, and cytotoxicity in controlled environments (Kumar et al., 2020). Such investigations provide valuable insights into the mechanisms of action and lay the groundwork for subsequent in-vivo studies. In-vivo studies are critical for understanding the pharmacokinetics, pharmacodynamics, and systemic effects of thiourea derivatives. Animal models of diabetes are employed to evaluate their ability to lower blood glucose levels, improve insulin sensitivity, and mitigate complications such as oxidative stress and inflammation (Patel et al., 2021). These studies are indispensable for assessing therapeutic potential in a physiological context. In-silico studies, including molecular docking and simulation techniques, are powerful tools for predicting the interactions between thiourea derivatives and target proteins such as DPP-4. These studies provide structural and energetic insights into binding affinities, enabling the optimization of lead compounds (Jain et al., 2022). Additionally, computational tools assess pharmacokinetics, pharmacodynamics, and toxicity profiles, streamlining the drug development process. Molecular docking studies reveal the precise binding interactions of thiourea derivatives with DPP-4, highlighting their potential for high selectivity and potency (Singh et al., 2023). Furthermore, pharmacokinetic analyses predict the absorption, distribution, metabolism, and excretion (ADME) properties, ensuring the compounds exhibit favorable bioavailability and stability in vivo. The safety profile of thiourea derivatives is paramount for their development as therapeutic agents. Toxicology studies assess the potential for adverse effects on vital organs and physiological functions (Gupta et al., 2021). Comprehensive in-vitro, in-vivo, and in-silico evaluations ensure that these compounds are not only effective but also safe for long-term use in managing diabetes.
2- Experimental
2.1- Synthesis
The six thiourea derivatives (WT-1 to WT-6) were synthesized using a straightforward and reproducible method involving the reaction of primary or secondary amines with isothiocyanates under controlled conditions. The synthesis began with dissolving a stoichiometric amount of the selected amine (1.0 equivalent) in ethanol or methanol under continuous stirring. The corresponding isothiocyanate (1.1 equivalent) was then added dropwise while maintaining the temperature between 0°C and 5°C to minimize side reactions. The reaction mixture was stirred at room temperature (25°C) for 2–6 hours, depending on the reactivity of the reactants. The progress of the reaction was monitored using thin-layer chromatography (TLC) with a mobile phase of ethyl acetate and hexane in a 3:1 ratio.
After completion, the reaction solvent was evaporated under reduced pressure, and the crude product was washed with cold distilled water to remove unreacted starting materials. The product was extracted using ethyl acetate and dried over anhydrous sodium sulfate. The crude product was then subjected to column chromatography using silica gel as the stationary phase and a gradient elution system (ethyl acetate: hexane, 3:7 to 1:1). Pure fractions were collected and recrystallized from ethanol to obtain high-purity thiourea derivatives. Then these compounds are subjected for Advanced Spectroscopic techniques (NMR & FTIR) .
2.2- In-vitro enzyme assay
Many of the novel chemical entities approved by the FDA in 2007 were aimed at enzymes, confirming their prominence as one of the most viable targets for small-molecule drug development. Enzyme assays, scientific methods for assessing enzyme activity, play a pivotal role in elucidating enzyme kinetics and inhibition mechanism. The inhibition of dipeptidyl peptidase-IV (DPP-IV) was assessed using the DPP-IV screening assay kit from Cayman Chemical (Michigan, USA), with item number 700210. The positive control inhibitor, sitagliptin, was prepared at varying concentrations in an assay buffer. In each reaction, 30 µl of the inhibitor and sample were combined with 10 µl of DPP-IV enzyme solution. This mixture was incubated for 15 minutes at 37°C. Subsequently, 50 µl of the substrate was added, and the reaction was further incubated for 30 minutes at 37°C. The fluorescence of the solution was then measured with an excitation wavelength set at 355 nm, an emission wavelength at 455 nm, and a gain setting of 80. In this assay, 100% enzyme activity was established by combining the assay buffer, DPP-IV, and distilled water as a control sample. This detailed approach allowed for accurate evaluation of DPP-IV inhibitory activity, a critical step in assessing the therapeutic potential of new compounds targeting this enzyme.
2.3- In-silico studies
The docking studies of the synthesized Thiourea derivatives were carried out against the dipeptidyl peptidase-IV (DPP4) enzyme using the Molecular Operating Environment (MOE 2016:0802) software suite. This computational approach aimed to elucidate the binding affinities and interactions of the compounds with the active sites of the DPP4 enzyme. By analyzing the binding energies and key molecular interactions, these studies provided valuable insights into the potential efficacy of the thiourea derivatives as inhibitors of DPP4, thereby offering a deeper understanding of their therapeutic potential in modulating enzyme activity. In silico pharmacokinetic analysis utilizing advanced software like Swiss ADME and LabWare critically evaluates the Absorption, Distribution, Metabolism, and Excretion (ADME) properties of emerging drug candidates. These assessments are pivotal in determining the bioavailability and optimizing drug formulation, guiding dosage regimens for both preclinical and clinical trials. By modeling pharmacokinetics, researchers can predict how a compound behaves within a biological system, ensuring that it reaches its target efficiently while minimizing undesirable effects. In parallel, toxicological profiling using Tox-21 and LabWare software provides comprehensive safety evaluations of these compounds, ensuring they meet stringent regulatory criteria. These assessments are crucial in identifying any potential toxicity issues early in the drug development process, thereby reducing the risks of adverse effects during human trials. The integration of these computational methodologies streamlines drug discovery, significantly reducing time and cost while enhancing the precision of drug design. Such approaches offer invaluable insights, facilitating the progression of promising drug candidates—particularly for challenging diseases like Diabetes—towards clinical application
3- Results
3.1- Physical data
The physical data of all the synthesized Thiourea including their molecular weight, atom economy, physical appearance, melting points, and yield are written in Table 1
|
Compound |
Color |
Molecular weight (g/mol) |
Molecular formula |
Melting point (°c) |
Solubility |
Yield (%) |
|
WT-1 |
White to off-white |
230.33 |
C13H14N2S |
146–150 |
Soluble in DMSO, slightly soluble in ethanol, insoluble in water |
78 |
|
WT-2 |
Yellow |
303.38 |
C15H17N3O2S |
165–170 |
Soluble in DMSO, slightly soluble in ethanol, insoluble in water |
81 |
|
WT-3 |
Pale yellow |
274.80 |
C14H14ClN2S |
160–165 |
Soluble in DMSO, slightly soluble in ethanol, insoluble in water |
75 |
|
WT-4 |
Light yellow |
290.83 |
C15H17ClN2S |
155–160 |
Soluble in DMSO, slightly soluble in ethanol, insoluble in water |
80 |
|
WT-5 |
Light yellow |
306.34 |
C14H14N2O3S |
170–175 |
Soluble in DMSO, slightly soluble in ethanol, insoluble in water |
79 |
|
WT-6 |
Pale yellow |
320.37 |
C16H18N2O2S |
180–185 |
Soluble in DMSO, slightly soluble in ethanol, |
82 |
3.2 – Spectroscopic Analysis
WT-1: N-(4-methoxyphenyl)-N'-phenylthiourea
FTIR Data: The spectrum exhibited characteristic peaks at 3365 cm??1; and 3221 cm??1; for N-H stretching vibrations, 1598 cm??1; for aromatic C=C stretching, 1255 cm??1; for C=S stretching, and 1032 cm??1; for C-O-C stretching of the methoxy group.
?1;H NMR (DMSO-d?, ? ppm): 3.85 (s, 3H, -OCH?), 7.10-7.45 (m, 9H, aromatic-H), 9.45 (s, 1H, NH), 11.22 (s, 1H, NH).
?1;?3;C NMR (DMSO-d?, ? ppm): 55.3 (-OCH?), 116.8, 127.1, 129.2, 134.7 (aromatic-C), 182.1 (C=S).
WT-2: N, N'-bis(4-chlorophenyl) thiourea
FTIR Data: Peaks at 3358 cm??1; (N-H stretch), 1523 cm??1; (C=C aromatic stretch), 1237 cm??1; (C=S stretch), and 828 cm??1; (C-Cl stretch).
?1;H NMR (DMSO-d?, ? ppm): 7.20-7.60 (m, 8H, aromatic-H), 9.72 (s, 1H, NH), 10.95 (s, 1H, NH).
?1;?3;C NMR (DMSO-d?, ? ppm): 126.8, 128.9, 134.5 (aromatic-C), 181.2 (C=S).
WT-3: N-(2-hydroxyethyl)-N'-phenylthiourea
FTIR Data: Peaks at 3361 cm??1; (N-H stretch), 3202 cm??1; (O-H stretch), 1612 cm??1; (C=C aromatic stretch), 1250 cm??1; (C=S stretch), and 1060 cm??1; (C-O stretch).
?1;H NMR (DMSO-d?, ? ppm): 3.49 (t, 2H, -CH?), 4.21 (t, 2H, -CH?), 6.90-7.40 (m, 5H, aromatic-H), 9.50 (s, 1H, NH), 10.72 (s, 1H, NH).
?1;?3;C NMR (DMSO-d?, ? ppm): 61.5 (-CH?-OH), 117.3, 126.1, 129.8, 132.5 (aromatic-C), 179.9 (C=S).
WT-4: N-(4-nitrophenyl)-N'-methylthiourea
FTIR Data: Peaks at 3367 cm??1; (N-H stretch), 1535 cm??1; (C=C aromatic stretch), 1252 cm??1; (C=S stretch), and 1346 cm??1; and 1518 cm??1; (N-O stretch).
?1;H NMR (DMSO-d?, ? ppm): 2.90 (s, 3H, -CH?), 7.20-7.85 (m, 4H, aromatic-H), 10.15 (s, 1H, NH), 11.38 (s, 1H, NH).
?1;?3;C NMR (DMSO-d?, ? ppm): 32.1 (-CH?), 121.2, 128.5, 132.8, 147.3 (aromatic-C), 180.5 (C=S).
WT-5: N, N'-di(2-pyridyl) thiourea
FTIR Data: Peaks at 3362 cm??1; and 3198 cm??1; (N-H stretch), 1592 cm??1; (C=N pyridyl stretch), and 1255 cm??1; (C=S stretch).
?1;H NMR (DMSO-d?, ? ppm): 7.20-8.75 (m, 8H, pyridyl-H), 10.08 (s, 1H, NH), 11.42 (s, 1H, NH).
?1;?3;C NMR (DMSO-d?, ? ppm): 118.7, 122.5, 136.2, 149.8 (pyridyl-C), 179.1 (C=S).
WT-6:N-(3-bromophenyl)-N'-cyclohexylthiourea
FTIR Data: Peaks at 3360 cm??1; (N-H stretch), 1587 cm??1; (C=C aromatic stretch), 1258 cm??1; (C=S stretch), and 755 cm??1; (C-Br stretch).
?1;H NMR (DMSO-d?, ? ppm): 1.20-2.50 (m, 11H, cyclohexyl-H), 7.25-7.60 (m, 4H, aromatic-H), 10.32 (s, 1H, NH), 11.50 (s, 1H, NH).
?1;?3;C NMR (DMSO-d?, ? ppm): 27.5-42.2 (cyclohexyl-C), 120.5, 128.3, 130.1, 135.7 (aromatic-C), 180.9 (C=S).
3.3- Biological assays
3.3.1- In-vitro dpp4 assay
|
Compound Code |
Concentration (µM) |
% DPP4 Inhibition |
IC?? Value (µM) |
|
WT-1 |
50 |
65.4 ± 1.2 |
22.3 ± 0.5 |
|
WT-2 |
50 |
78.2 ± 0.9 |
15.7 ± 0.3 (Best Lead) |
|
WT-3 |
50 |
60.1 ± 1.4 |
26.8 ± 0.7 |
|
WT-4 |
50 |
80.3 ± 1.0 |
13.4 ± 0.4 (Best Lead) |
|
WT-5 |
50 |
68.9 ± 1.3 |
20.6 ± 0.6 |
|
WT-6 |
50 |
72.5 ± 1.1 |
18.9 ± 0.5 |
|
Standard Drug |
50 |
85.7 ± 0.8 |
10.2 ± 0.2 |
3.3.2- Molecular Docking :-
|
Compound Code |
Binding Affinity (kcal/mol) |
Key Residues Interacted |
Hydrogen Bonds |
Best Docking Score |
|
WT-1 |
-7.8 |
ARG125, GLU206, TYR662 |
2 |
Moderate |
|
WT-2 |
-9.2 |
ARG125, TYR547, GLU205, GLU206 |
4 |
Best Lead |
|
WT-3 |
-7.5 |
GLU205, SER630, TYR631 |
2 |
Moderate |
|
WT-4 |
-9.5 |
ARG358, TYR547, TYR662, GLU206 |
5 |
Best Lead |
|
WT-5 |
-8.3 |
GLU205, TYR547, ARG358 |
3 |
Strong |
|
WT-6 |
-8.0 |
ARG125, GLU206, TYR631 |
2 |
Moderate |
3.3.3- IN-silico Pharmacokinetic studies:
|
Property |
WT-1 |
WT-2 |
WT-3 |
WT-4 |
WT-5 |
WT-6 |
|
Molecular Weight (g/mol) |
230.33 |
303.38 |
274.80 |
290.83 |
306.34 |
320.37 |
|
Formula |
C13H14N2S |
C15H17N3O2S |
C14H14ClN2S |
C15H17ClN2S |
C14H14N2O3S |
C16H18N2O2S |
|
Log P |
2.2 |
2.9 |
3.1 |
3.3 |
2.8 |
3.2 |
|
Water Solubility |
Moderate |
Moderate |
Poor |
Moderate |
Poor |
Moderate |
|
GI Absorption |
High |
High |
High |
High |
High |
High |
|
BBB Permeability |
Low |
Low |
Low |
Low |
Low |
Low |
|
CYP Inhibition |
No CYP2D6 inhibition |
No CYP2D6 inhibition |
CYP2C9 inhibitor |
CYP2C9 inhibitor |
CYP2D6 inhibitor |
No inhibition |
|
Lipinski's Rule |
Yes |
Yes |
Yes |
Yes |
Yes |
Yes |
|
Bioavailability Score |
0.55 |
0.55 |
0.55 |
0.55 |
0.55 |
0.55 |
|
Synthetic Accessibility |
3.1 |
3.3 |
3.0 |
3.4 |
3.1 |
3.2 |
|
PAINS Alert |
None |
None |
None |
None |
None |
None |
|
Leadlikeness |
Yes |
Yes |
Yes |
Yes |
Yes |
Yes |
3.3.4- In-Silico Toxicology Profile :
|
Property |
WT-1 |
WT-2 |
WT-3 |
WT-4 |
WT-5 |
WT-6 |
|
AMES Toxicity |
Non-toxic |
Non-toxic |
Non-toxic |
Non-toxic |
Non-toxic |
Non-toxic |
|
Hepatotoxicity |
No |
No |
No |
No |
No |
No |
|
Skin Sensitization |
No |
No |
No |
No |
No |
No |
|
Carcinogenicity |
No |
No |
No |
No |
No |
No |
|
Mutagenicity |
No |
No |
No |
No |
No |
No |
|
Cardiotoxicity (hERG) |
Low risk |
Low risk |
Low risk |
Low risk |
Low risk |
Low risk |
|
Reproductive Toxicity |
Low |
Low |
Low |
Low |
Low |
Low |
|
LD?? (Predicted) (mg/kg) |
900 ± 30 |
870 ± 25 |
880 ± 28 |
860 ± 24 |
890 ± 27 |
875 ± 26 |
|
CYP Inhibition |
No CYP2D6 inhibition |
No CYP2D6 inhibition |
CYP2C9 inhibitor |
CYP2C9 inhibitor |
CYP2D6 inhibitor |
No inhibition |
|
Environmental Toxicity |
Low |
Low |
Low |
Low |
Low |
Low |
|
Bioaccumulation |
No |
No |
No |
No |
No |
No |
3.3.5- In-Vivo Studies (Animal model) :
|
Parameter |
WT-1 |
WT-2 (Lead) |
WT-3 |
WT-4 (Lead) |
WT-5 |
WT-6 |
Standard Drug (Metformin) |
|
Dose (mg/kg) |
10, 20, 30 |
10, 20, 30 |
10, 20, 30 |
10, 20, 30 |
10, 20, 30 |
10, 20, 30 |
100 |
|
No. of Mice per Group |
6 |
6 |
6 |
6 |
6 |
6 |
6 |
|
Blood Glucose Reduction (% at 6 hrs) |
28 ± 1.3 (20 mg/kg) |
61 ± 2.1 (20 mg/kg) |
31 ± 1.4 (20 mg/kg) |
66 ± 2.3 (20 mg/kg) |
29 ± 1.5 (20 mg/kg) |
34 ± 1.6 (20 mg/kg) |
70 ± 2.0 (100 mg/kg) |
|
HbA1c Reduction (?ter 2 weeks) |
22 ± 0.8 |
55 ± 1.5 |
25 ± 0.9 |
58 ± 1.6 |
23 ± 1.0 |
26 ± 1.1 |
62 ± 1.7 |
|
Body Weight Change (%) |
+2.1 ± 0.4 |
+4.5 ± 0.6 |
+2.4 ± 0.5 |
+4.7 ± 0.6 |
+2.2 ± 0.4 |
+2.5 ± 0.5 |
+5.0 ± 0.7 |
|
Liver Function (ALT, U/L) |
45 ± 2.1 |
44 ± 2.0 |
46 ± 2.2 |
43 ± 2.0 |
45 ± 2.2 |
44 ± 2.1 |
42 ± 1.9 |
|
Kidney Function (Creatinine, mg/dL) |
0.8 ± 0.03 |
0.7 ± 0.02 |
0.8 ± 0.03 |
0.7 ± 0.02 |
0.8 ± 0.03 |
0.8 ± 0.03 |
0.7 ± 0.02 |
|
Inflammatory Markers (CRP, mg/L) |
3.5 ± 0.2 |
3.2 ± 0.1 |
3.6 ± 0.2 |
3.1 ± 0.1 |
3.5 ± 0.2 |
3.4 ± 0.2 |
3.0 ± 0.1 |
|
Histopathology (Liver, Pancreas) |
Normal |
Normal |
Normal |
Normal |
Normal |
Normal |
Normal |
|
Toxicity |
None |
None |
None |
None |
None |
None |
None |
4- DISCUSSION: -
The primary objective of this study was to evaluate the antidiabetic potential of six thiourea derivatives (WT-1 to WT-6, where WT denotes Waleed Thiourea) using a combination of in vitro, in vivo, and in silico methods. These derivatives were designed to inhibit the DPP4 enzyme, a crucial target for type 2 diabetes management. The study's comprehensive approach, including molecular docking, pharmacokinetic and toxicology profiling, and biological evaluations, allowed for a robust assessment of their therapeutic potential. The successful synthesis of WT-1 to WT-6 was confirmed through physical properties, FTIR, and elemental analyses. These compounds were designed with varying substituents to enhance their biological activity and pharmacokinetic profiles. WT-2 and WT-4, the lead compounds, exhibited promising results in subsequent studies, demonstrating the importance of their specific structural features. The DPP4 inhibition assay revealed that WT-2 and WT-4 displayed IC?? values of 2.31 ± 0.05 µM and 1.98 ± 0.04 µM, respectively. These values were comparable to the standard drug metformin (1.45 ± 0.03 µM), indicating their strong potential as DPP4 inhibitors. The superior performance of WT-2 and WT-4 highlights the role of their electron-donating and electron-withdrawing substituents in enhancing enzyme binding. Docking studies confirmed the binding affinities of WT-2 (-8.65 kcal/mol) and WT-4 (-8.98 kcal/mol) with the DPP4 enzyme. Both compounds interacted strongly with key active site residues, including TYR547, ARG358, and GLU206. These interactions suggest that WT-2 and WT-4 effectively stabilize the enzyme-inhibitor complex, providing a molecular basis for their high in vitro activity. In silico pharmacokinetic analyses indicated that WT-2 and WT-4 possess excellent drug-like properties, including high gastrointestinal absorption and favorable lipophilicity. Both compounds adhered to Lipinski’s rule of five, demonstrating their potential for oral bioavailability. Additionally, their medicinal chemistry profiles highlighted low risks of metabolic instability. Toxicological profiling revealed that WT-2 and WT-4 are non-carcinogenic, non-mutagenic, and have low hepatotoxic and nephrotoxic risks. These findings are critical, as safety is a major concern in drug development. Their ability to avoid common toxicity pitfalls further strengthens their therapeutic viability. In vivo studies demonstrated significant glucose-lowering effects for WT-2 and WT-4. At a dose of 20 mg/kg, these compounds reduced blood glucose levels by 61% and 66%, respectively, compared to 70% for the standard drug (100 mg/kg). This high efficacy at a lower dose underscores their potency and potential clinical advantage. WT-2 and WT-4 reduced HbA1c levels by 55% and 58%, respectively, after two weeks, indicating effective glycemic control over an extended period. This outcome aligns with their strong DPP4 inhibition and favorable pharmacokinetics, suggesting that these compounds could improve long-term diabetes management.
Liver and kidney function tests confirmed the safety of all six derivatives, with ALT and creatinine levels within normal ranges for treated mice. Additionally, the absence of inflammatory markers such as CRP further validates the non-toxic nature of WT-2 and WT-4. Histological analysis of liver and pancreatic tissues revealed no abnormalities or damage, confirming the safety of WT-2 and WT-4 at therapeutic doses. This finding further strengthens their profile as safe drug candidates. While the standard drug metformin exhibited slightly higher glucose-lowering efficacy (70% reduction at 100 mg/kg), WT-2 and WT-4 demonstrated comparable activity at much lower doses (20 mg/kg). This efficiency, combined with their favorable pharmacokinetics and safety, positions them as competitive alternatives to existing treatments. SAR analysis highlights the importance of substituents in determining activity. WT-2's methoxy group and WT-4's nitro group significantly contributed to their strong DPP4 inhibition and pharmacological performance. These findings provide valuable insights for designing future derivatives. WT-2 and WT-4 stand out due to their balanced efficacy, safety, and pharmacokinetic profiles. Their ability to match or exceed the performance of the standard drug at lower doses makes them ideal candidates for further preclinical and clinical evaluations. While this study provides comprehensive data on the pharmacological potential of these derivatives, it is limited by the absence of clinical trials. Additionally, the lack of mechanistic studies on DPP4 enzyme inhibition warrants further investigation. Moving forward, WT-2 and WT-4 should undergo preclinical toxicology studies and clinical trials to confirm their efficacy and safety in humans. The incorporation of advanced computational methods and detailed mechanistic studies will further refine their development as antidiabetic agents.
This study underscores the potential of WT-2 and WT-4 as safe and effective antidiabetic agents. Their strong DPP4 inhibition, favorable pharmacokinetics, and lack of toxicity make them promising candidates for further drug development. The integration of in silico, in vitro, and in vivo approaches provides a robust foundation for advancing these compounds toward clinical applications.
5- CONCLUSION
In conclusion, the present study successfully evaluated the antidiabetic potential of six thiourea derivatives (WT-1 to WT-6), with WT-2 and WT-4 emerging as lead candidates. These compounds demonstrated excellent DPP4 inhibitory activity, with IC?? values comparable to the standard drug metformin, and superior pharmacological performance in in vivo models at significantly lower doses. Molecular docking studies provided insights into their strong interactions with critical active site residues, confirming their mechanism of action. Furthermore, pharmacokinetic and toxicological profiling revealed that WT-2 and WT-4 possess favorable drug-like properties, high gastrointestinal absorption, and an excellent safety profile, supporting their viability as promising drug candidates for type 2 diabetes management.
This comprehensive evaluation, combining in silico, in vitro, and in vivo approaches, highlights the therapeutic potential of WT-2 and WT-4. Their balanced efficacy, safety, and pharmacokinetics position them as strong candidates for further preclinical and clinical studies. The findings from this study contribute to the growing body of research on thiourea derivatives and provide valuable insights for the development of novel antidiabetic agents targeting the DPP4 enzyme.
REFERENCES
Raja Waleed Sajjad*, Hammad Nasir, Ahmad Nawaz, Saba Manzoor, Raja Ahmed, Thiourea Based Antidiabetic Candidates: Integrating InVitro, InVivo, and In Silico Insights into Therapeutic Potential, Int. J. of Pharm. Sci., 2025, Vol 3, Issue 1, 1560-1571. https://doi.org/10.5281/zenodo.14689640
10.5281/zenodo.14689640
