Investigation of chemical reactivity of 2-alkoxy-1,4-naphthoquinones and their anticancer activity

To establish the structure-activity relationship of 5-hydroxy-1,4-naphthoquinones toward anticancer activity, a series of its derivatives were prepared and tested for the activity (IC₅₀ in µM) against three cell lines; colo205 (colon adenocarcinoma), T47D (breast ductal carcinoma) and K562 (chronic myelogenous leukemia). Among them 2 (IC₅₀: 2.3; 2.0; 1.4?µM), 6 (IC₅₀: 1.9; 2.2; 1.3?µM), 9 (IC₅₀: 0.7; 1.7; 0.9?µM) and 10 (IC₅₀:1.7; 1.0; 1.2?µM) showed moderate to excellent activity. Our perception toward the DNA substitution of alkoxy groups at the C2 position of these naphthoquinones for the anticancer activity led us to investigate their reactivity of substitution toward dimethylamine as a nucleophile. The ease of the substitution of alkoxy groups at the C2 position with dimethylamine is strongly accelerated by hydroxyl group at C5 position and is well correlated with the found anticancer activity results.     

Structure-activity relationship studies on vitamin D lactam derivatives as vitamin D receptor antagonist

Structure–activity relationship studies on 1α,25-dihydroxyvitamin D₃-26,23-lactams (DLAMs), antagonists of vitamin D, were conducted, focusing on the substituents of the phenyl group. One of the derivatives (23S,25S)-DLAM-1P-3,5(OEt)₂, showed potent antagonistic activity with an IC₅₀ of 90 nM.     

2-(3′-Hydroxypropylidene)-1α-hydroxy-19-norvitamin D compounds with truncated side chains

Our recent studies with 2-(3′-hydroxypropylidene) analogs of 1α,25-dihydroxy-19-norvitamin D₃ showed that this 2-substituent creates compounds with very potent biological activity. In the continuing search for vitamin D compounds with selective activity profiles, we prepared a series of 1α-hydroxy-19-norvitamin D analogs characterized by the presence of a 3′-hydroxypropylidene substituent at C-2 and a truncated side chain. These vitamin D compounds were efficiently prepared using convergent syntheses. The C,D-fragments, namely the Grundmann ketones 19, 20, 27, 36 and 37 were synthesized from the known 8β-benzoyloxy-22-aldehydes 12 and 29. These hydrindanones were subjected to Lythgoe type Wittig–Horner coupling with phosphine oxide 21, prepared by us previously, and after hydroxyl deprotection the set of 19-norvitamins 7–11 was successfully obtained. According to our expectations, all analogs (with an exception of the 20R-compound 7) have pronounced in vitro activity. When compared to the natural hormone 1α,25-(OH)₂D₃ (1), they show the same or only slightly reduced affinity for the vitamin D receptor while being similarly effective as 1 in differentiation of HL-60 cells into monocytes.     

Design,synthesis, and bioactivity of dihydropyrimidine derivatives as kinesin spindle protein inhibitors

A series of twenty-one 3,4-dihydropyrimidine derivatives bearing the heterocyclic 1,3-benzodioxole at position 4 in addition to different substituents at positions 2, 3 and 5 were designed and synthesized as monastrol analogs. The novel synthesized compounds were screened for their cytotoxic activity towards 60 cancer cell lines according to NCI (USA) protocol. Compounds 10b and 15 showed the best antitumor activity against most cell lines. Compound 15 was subsequently tested in 5-doses mode and displayed high selectivity towards CNS, prostate and leukemia subpanel with selectivity ratios of 22.30, 15.38 and 12.56, respectively at GI₅₀ level. The IC₅₀ of compounds 9d, 10b, 12, 15 and 16 against kinesin enzyme were 3.86 ± 0.12, 10.70 ± 0.35, 3.95 ± 0.12, 4.36 ± 0.14, and 14.07 ± 0.45 μM respectively, while the prototype compound, monastrol, reported IC₅₀ value of 20 ± 0.42 μM. The safest compound among test compounds against normal cell line (HEK 293) is 10b with IC₅₀ value of 62.02 ± 2.42 µM/ml in comparison to doxorubicin (IC₅₀ = 11.34 ± 0.44 µM/ml). Cell cycle analysis of SNB-75 cells treated with compound 15 showed cell cycle arrest at G2/M phase. Further, the assay of levels of active caspase-3 and caspase-9 was investigated. Moreover, Molecular docking of compounds, 9d, 10b, 12, 15, 16, monastrol and mon-97 was performed to study the interaction between inhibitors and the kinesin spindle protein allosteric binding site.     

Cytotoxicity of synthesized 1,4-naphthoquinone analogues on selected human cancer cell lines

In an effort to establish new candidates with enhanced anticancer activity of 5-hydroxy-7-methyl-1,4-naphthoquinone scaffold (7-methyljuglone) previously isolated from the root extract of Euclea natalensis, a series of 7-methyljuglone derivatives have been synthesized and assessed for cytotoxicity on selected human cancer lines. These compounds were screened in vitro for anticancer activity on MCF-7, HeLa, SNO and DU145 human cancer cell lines by MTT assay. Most of them exhibited significant toxicity on cancer cell lines with lower IC₅₀ values. The most potent derivative (19) exhibited the toxicity on HeLa and DU145 cell lines with IC₅₀ value of 5.3 and 6.8 μM followed by compound (5) with IC₅₀ value of 10.1 and 9.3 μM, respectively. Structure–activity relationship reveals that the fluoro substituents at position C-8 while hydroxyl substituents at C-2 and C-5 positions played an important role in toxicity.     

Tetrahydroprotoberberine alkaloids with dopamine and σ receptor affinity

Two series of analogues of the tetrahydroprotoberberine (THPB) alkaloid (±)-stepholidine that (a) contain various alkoxy substituents at the C10 position and, (b) were de-rigidified with respect to (±)-stepholidine, were synthesized and evaluated for affinity at dopamine and σ receptors in order to evaluate effects on D3 and σ2 receptor affinity and selectivity. Small n-alkoxy groups are best tolerated by D3 and σ2 receptors. Among all compounds tested, C10 methoxy and ethoxy analogues (10 and 11 respectively) displayed the highest affinity for σ2 receptors as well as σ2 versus σ1 selectivity and also showed the highest D3 receptor affinity. De-rigidification of stepholidine resulted in decreased affinity at all receptors evaluated; thus the tetracyclic THPB framework is advantageous for affinity at dopamine and σ receptors. Docking of the C10 analogues at the D3 receptor, suggest that an ionic interaction between the protonated nitrogen atom and Asp110, a H-bond interaction between the C2 phenol and Ser192, a H-bond interaction between the C10 phenol and Cys181 as well as hydrophobic interactions of the aryl rings to Phe106 and Phe345, are critical for high affinity of the compounds.     

Anti-inflammatory drug approach: Synthesis and biological evaluation of novel pyrazolo[3,4-d]pyrimidine compounds

In this study, the acid chlorides of pyrazolo[3,4-d]pyrimidine compounds were prepared and reacted with a number of nucleophiles. The novel compounds were experimentally tested via enzyme assay and they showed cyclooxygenase-2 inhibition activity in the middle micro molar range (4b had a COX-1 IC₅₀ of 26 µM and a COX-2 IC₅₀ of 34 µM, 3b had a COX-1 IC₅₀ of 19 µM and a COX-2 IC₅₀ of 31 µM, 3a had a COX-2 IC₅₀ of 42 µM). These compounds were analyzed via docking and were predicted to interact with some of the COX-2 key residues. Our best hit, 4d (COX-1 IC₅₀ of 28 µM, COX-2 IC₅₀ of 23 µM), appears to adopt similar binding modes to the standard COX-2 inhibitor, celecoxib, proposing room for possible selectivity. Additionally, the resultant novel compounds were tested in several in vivo assays. Four compounds 3a (COX-2 IC₅₀ of 42 µM), 3d, 4d and 4f were notable for their anti-inflammatory activity that was comparable to that of the clinically available COX-2 inhibitor celecoxib. Interestingly, they showed greater potency than the famous non-steroidal anti-inflammatory drug, Diclofenac sodium. In summary, these novel pyrazolo[3,4-d]pyrimidine analogues showed interesting anti-inflammatory activity and could act as a starting point for future drugs.     

Structural development of p-carborane-based potent non-secosteroidal vitamin D analogs

Non-secosteroidal vitamin D receptor (VDR) ligands are promising candidates for many clinical applications. We recently developed novel non-secosteroidal VDR agonists based on p-carborane (an icosahedral carbon-containing boron cluster) as a hydrophobic core structure. Here, we report the design, synthesis and biological evaluation of carborane-based vitamin D analogs bearing various substituents at the diol moiety. Among the synthesized compounds, methylene derivative 31 exhibited the most potent vitamin D activity, which was comparable to that of the natural hormone, 1α,25(OH)₂D₃. This compound is one of the most potent non-secosteroidal VDR agonists reported to date, and is a promising lead for development of novel drug candidates.     

Melanogenesis inhibitory activity of a 7-O-9′-linked neolignan from Alpinia galanga fruit

An aqueous acetone extract from the fruit of Alpinia galanga (Zingiberaceae) demonstrated inhibitory effects on melanogenesis in theophylline-stimulated murine B16 melanoma 4A5 cells (IC₅₀ = 7.3 μg/mL). Through bioassay-guided separation of the extract, a new 7-O-9′-linked neolignan, named galanganol D diacetate (1), was isolated along with 16 known compounds including 14 phenylpropanoids (2–15). The structure of 1, including its absolute stereochemistry in the C-7 position, was elucidated by means of extensive NMR analysis and total synthesis. Among the isolates, 1 (IC₅₀ = 2.5 μM), 1′S-1′-acetoxychavicol acetate (2, 5.0 μM), and 1′S-1′-acetoxyeugenol acetate (3, 5.6 μM) exhibited a relatively potent inhibitory effect without notable cytotoxicity at effective concentrations. The following structural requirements were suggested to enhance the inhibitory activity of phenylpropanoids on melanogenesis: (i) compounds with 4-acetoxy group exhibit higher activity than those with 4-hydroxy group; (ii) 3-methoxy group dose not affect the activity; (iii) acetylation of the 1′-hydroxy moiety enhances the activity; and (iv) phenylpropanoid dimers with the 7-O-9′-linked neolignan skeleton exhibited higher activity than those with the corresponding monomer. Their respective enantiomers [1′ (IC₅₀ = 1.9 μM) and 2′ (4.5 μM)] and racemic mixtures [(±)-1 (2.2 μM) and (±)-2 (4.4 μM)] were found to exhibit melanogenesis inhibitory activities equivalent to those of the naturally occurring optical active compounds (1 and 2). Furthermore, the active compounds 1–3 inhibited tyrosinase, tyrosine-related protein (TRP)-1, and TRP-2 mRNA expressions, which could be the mechanism of melanogenesis inhibitory activity.     

Synthesis and structure-activity relationship study of multi-target triazine derivatives as innovative candidates for treatment of Alzheimer's disease

The complex pathogenesis of Alzheimer’s disease (AD) requires using multi-target ligands (MTLs) for disease management. We synthesized, characterized and evaluated a series of novel triazine analogues as MTLs for AD. The biological screening results indicated that most of our compounds displayed potent inhibitory activities against β-site APP-cleaving enzyme 1 (BACE1) using a FRET-based assay. Compounds 6c and 6m were found to possess significant BACE1 inhibitory properties with IC₅₀ values of 0.91 (±0.25) µM and 0.69 (±0.20) µM, respectively. DPPH radical scavenging activity evaluation showed that compounds with hydroxyl and pyrrole moieties had antioxidant effects. Docking evaluations provided insight into enzyme inhibitory interactions of novel synthesized compounds with the BACE1 active site involving a critical role for Gln73 and/or Phe108 alongside of Asp32. Metal chelation tests confirmed that compound 6m is a chelator for Fe²⁺, Fe³⁺, Zn²⁺, Cu²⁺. Moreover 6m as the most potent BACE1 inhibitor did not show any toxicity against PC12 neuronal cells. These findings demonstrate the high potential of triazine scaffolds in the design of MTLs for treatment of AD.     

Design and synthesis of tetraol derivatives of 1,12-dicarba-closo-dodecaborane as non-secosteroidal vitamin D analogs

Vitamin D receptor (VDR), a nuclear receptor for 1α,25-dihydroxyvitamin D₃ (1α,25(OH)₂D₃, 1), is a promising target for multiple clinical applications. We recently developed non-secosteroidal VDR ligands based on a carbon-containing boron cluster, 1,12-dicarba-closo-dodecaborane (p-carborane), and examined the binding of one of them to VDR by means of crystallographic analysis. Here, we utilized that X-ray structure to design novel p-carborane-based tetraol-type vitamin D analogs, and we examined the biological activities of the synthesized compounds. Structure–activity relationship study revealed that introduction of an ω-hydroxyalkoxy functionality enhanced the biological activity, and the configuration of the substituent significantly influenced the potency. Among the synthesized compounds, 4-hydroxybutoxy derivative 9a exhibited the most potent activity, which was equal to that of the secosteroidal vitamin D analog, 19-nor-1α,25-dihydroxyvitamin D₃ (2).     

VEGFR-2 inhibiting effect and molecular modeling of newly synthesized coumarin derivatives as anti-breast cancer agents

Twenty five newly synthesized coumarin scaffold based derivatives were assayed for their in vitro anticancer activity against MCF-7 breast and PC-3 prostate cancer cell lines and were further assessed for their in vitro VEGFR-2 kinase inhibitory activity. The in vitro cytotoxic studies revealed that most of the synthesized compounds possessed very promising cytotoxicity against MCF-7, particularly; compounds 4a (IC₅₀ = 1.24 µM) and 3d (IC₅₀ = 1.65 µM) exhibited exceptional activities superior to the positive control staurosporine (IC₅₀ = 8.81 µM). Similarly, the majority of the compounds exhibited higher antiproliferative activities compared to the reference standard with IC₅₀ values ranging from 2.07 to 8.68 µM. The two cytotoxic derivatives 4a and 3d were selected to evaluate their inhibitory potencies against VEGFR-2 kinase. Remarkably, compound 4a, exhibited significant IC₅₀ of 0.36 µM comparable to staurosporine (IC₅₀; 0.33 µM). Moreover, it was capable of inducing preG1 apoptosis, cell growth arrest at G2/M phase and activating caspase-9. On the other hand, insignificant cytotoxic activity was observed for all compounds towards PC-3 cell line. Molecular docking study was carried out for the most active anti-VEGFR-2 derivative 4a, which demonstrated the ability of the tested compound to interact with the key amino acids in the target VEGFR-2 kinase binding site. Additionally, the ADME parameters and physicochemical properties of compound 4a were examined in silico.     

Yohimbine as a Starting Point to Access Diverse Natural Product-Like Agents with Re-programmed Activities against Cancer-Relevant GPCR Targets

G protein-coupled receptors (GPCRs) constitute the largest protein superfamily in the human genome. GPCRs play key roles in mediating a wide variety of physiological events including proliferation and cancer metastasis. Given the major roles that GPCRs play in mediating cancer growth, they present promising targets for small molecule therapeutics. One of the principal goals of our lab is to identify complex natural products (NPs) suitable for ring distortion, or the dramatic altering of the inherently complex architectures of NPs, to rapidly generate an array of compounds with diverse molecular skeletal systems. The overarching goal of our ring distortion approach is to re-program the biological activity of select natural products and identify new compounds of importance to the treatment of disease, such as cancer. Described herein are the results from biological screens of diverse small molecules derived from the indole alkaloid yohimbine against a panel of GPCRs involved in various diseases. Several analogues displayed highly differential antagonistic activities across the GPCRs tested. We highlight the re-programmed profile of one analogue, Y7g, which exhibited selective antagonistic activities against AVPR2 (IC₅₀ = 459 nM) and OXTR (IC₅₀ = 1.16 µM). The activity profile of Y7g could correlate its HIF-dependent anti-cancer activity to its GPCR antagonism since these receptors are known to be upregulated in hypoxic cellular environments. Our findings demonstrate that the ring distortion of yohimbine can lead to the identification of new compounds capable of interacting with distinct cancer-relevant targets.     

Drug-Induced Up-Regulation of Dopamine D2 Receptors on Cultured Cells

Abstract: Ligand-induced up-regulation of recombinant dopamine D2 receptors was assessed using C₆ glioma cells stably expressing the short (415-amino-acid; D2_S) and long (444-amino-acid; D2_L) forms of the receptor. Overnight treatment of C6-D2_L cells with N-propylnorapomorphine (NPA) caused a time- and concentration-dependent increase in the density of receptors, as assessed by the binding of radioligand to membranes prepared from the cells, with no change in the affinity of the receptors for the radioligand. The effect of 10 µM NPA was maximal after 10 h, at which time the density of D2_L receptors was more than doubled. The agonists dopamine and quinpirole also increased the density of D2_L receptors. The receptor up-regulation was not specific for agonists, because the antagonists epidepride, sulpiride, and domperidone caused smaller (30–60%) increases in receptor density. Prolonged treatment with 10 µM NPA desensitized D2_L receptors, as evidenced by a reduced ability of dopamine to inhibit adenylyl cyclase, whereas treatment with sulpiride was associated with an enhanced responsiveness to dopamine. The magnitude of NPA-induced receptor up-regulation in each of four clonal lines of C6-D2_L cells (mean increase, 80%) was greater than in all four lines of C6-D2_S cells (33%). Inactivation of pertussis toxin-sensitive G proteins had no effect on the basal density of D2_L receptors or on the NPA-induced receptor up-regulation. Treatment with 5 µg/ml of cycloheximide, on the other hand, decreased the basal density of receptors and attenuated, but did not prevent, the NPA-induced increase. Chimeric D1/D2 receptors were used to identify structural determinants of dopamine receptor regulation. Treatment with the D1/D2 agonist NPA decreased the density of D1 and chimeric CH4 and CH3 receptors. The latter two receptors have D1 sequence from the amino-terminus to the amino-terminal end of transmembrane region (TM) VII and VI, respectively. CH2, with D1 sequence up to the amino-terminal end of TM V, and thus the third cytoplasmic loop of the D2 receptor, was up-regulated by NPA or the D2-selective agonist quinpirole. Quinpirole treatment decreased the density of CH3 and had no effect on CH4 or D1 receptors. The different responses of CH2 and CH3 to agonist treatment suggest a role for TM V and the third cytoplasmic loop in the direction of receptor regulation.     

New class of alkynyl glycoside analogues as tyrosinase inhibitors

A new series of alkynyl glycoside analogues were designed and synthesized from cheap and a commercially available sugar by introduction of various alkynyl and alkyl groups at C-1 and C-6 positions of the sugar ring. The inhibitory abilities of alkynyl glycosides were investigated in vitro on mushroom tyrosinase for the catalysis of l-Tyrosine and l-DOPA as substrates and comparing with arbutin and kojic acid. Non-terminal alkyne compound 2d showed excellent tyrosinase inhibitory activity (IC₅₀ 54.0 μM) against l-Tyrosine comparable to arbutin (IC₅₀ 1.46 mM) while 2b exhibited potent activities (IC₅₀ 34.3 μM) against L-DOPA higher than kojic acid (IC₅₀ 0.11 mM) and arbutin (IC₅₀ 13.3 mM). Kinetic studies revealed that compound 2d was a non-competitive inhibitor with the best Ki value of 21 μM and formed an irreversible receptor complex with mushroom tyrosinase. The SARs results showed that the type of alkyne and alkyl groups at position C-6 on sugar and the stereoisomer played an important role in determining their inhibitory activities. The potent activity of alkynyl glycosides identified in this study highlight the importance of this scaffold and these compounds are very modestly potent to the development of new class for tyrosinase inhibitor.     

Synthesis and dopaminergic activity of a series of new 1-aryl tetrahydroisoquinolines and 2-substituted 1-aryl-3-tetrahydrobenzazepines

A series of new 1-aryl-6,7-dihydroxy tetrahydroisoquinolines with several substitution patterns in the 1-aryl group at C-1 were prepared in good yields. The influence of each substituent on the affinity and selectivity for D₁ and D₂ dopaminergic receptors was studied. Moreover, N-alkyl salts of these tetrahydroisoquinolines were used as starting material to synthesize a series of new 1-aryl-7,8-dihydroxy 3-tetrahydrobenzazepines derivatives with electron-withdrawing substituents at C-2 position by the diastereoselective Stevens rearrangement. The structure-activity relationship of these compounds was explored to evaluate the effect of the functional group at C-2 in benzazepines and the modification in the aryl group at the isoquinoline C-1 position towards the affinity and selectivity for the mentioned receptors. The 1-aryl-6,7-dihydroxy tetrahydroisoquinoline 4c shows significant affinity towards D₂ receptor, with K_i value of 31 nM. This significant affinity can be attributed to the presence of a thiomethyl group, and it is the most active 1-aryl-6,7-dihydroxy tetrahydroisoquinoline derivative reported to date.     

Two new polyhydroxylated steroidal glycosides from Paris polyphylla var. yunnanensis

Chemical investigation of Paris polyphylla smith var. yunnanensis afforded two new polyhydroxylated steroidal glycosides, named Parisyunnanosides K and L (1–2), together with nine known ones. The chemical structures of the new compounds were elucidated by 1D, 2D NMR and HR-ESI-MS techniques, together with chemical methods. Parisyunnanosides K and L (1–2) are rare C₂₇ steroidal glycosides with two double bonds located at C-5, 6 and C-25, 26 of the aglycone, respectively. The cytotoxic activities of the isolated compounds were evaluated against Caco-2 cells by CCK-8 assay. Among them, compounds 5–11 exhibited potent cytotoxic activity with IC₅₀ values ranging from 1.10 to 14.14 μM compared with the positive control oxaliplatin (1.38 μM).     

Synthesis,biological evaluation,and molecular docking study of novel allyl-retrochalcones as a new class of protein tyrosine phosphatase 1B inhibitors

We describe herein the design, synthesis, and biological evaluation of a series of novel protein tyrosine phosphatase 1B (PTP1B) inhibitor retrochalcones having an allyl chain at the C-5 position of their B ring. Biological screening results showed that the majority of these compounds exhibited an inhibitory activity against PTP1B. Thus, preliminary structure-activity relationship (SAR) and quantitative SAR analyses were conducted. Among the compounds, 23 was the most potent inhibitor, exhibiting the highest in vitro inhibitory activity against PTP1B with an IC₅₀ of 0.57?µM. Moreover, it displayed a significant hepatoprotective property via activation of the IR pathway in type 2 diabetic db/db mice. In addition, the results of our docking study showed that 23, as a specific inhibitor of PTP1B, effectively transformed the WPD loop from “close” to “open” in the active site. These results may reveal suitable compounds for the development of PTP1B inhibitors.     

Six new dihydrobenzofuran lignans from the branches and leaves of Illicium wardii and their cytotoxic activities

Six new dihydrobenzofuran lignans, named illiciumlignans A⿿F (compounds 1⿿6), along with 15 known compounds (7⿿21) were isolated from the branches and leaves of Illicium wardii. The structures of 1⿿6 were determined using a combination of 1D and 2D NMR, HR-ESI⿿-MS, and CD spectroscopic data. Illiciumlignan D (4) is the first reported dihydrobenzofuran lignan arabinofuranoside that is derivatized with the arabinofuranose moiety on C-9⿲. Compounds 1⿿21 were evaluated for cytotoxic activity against four human cancer cell lines. Compounds 8, 12 and 20 exhibited significant activity against human cancer cell lines (A549, SKOV3, HepG2 and HCT116), with IC₅₀ values ranging from 2.7 to 14.9 μM.     

Substituted phenyl[(5-benzyl-1,3,4-oxadiazol-2-yl)sulfanyl]acetates/acetamides as alkaline phosphatase inhibitors: Synthesis,computational studies,enzyme inhibitory kinetics and DNA binding studies

Substituted phenyl[(5-benzyl-1,3,4-oxadiazol-2-yl)sulfanyl]acetates/acetamides 9a-j were synthesized as alkaline phosphatase inhibitors. Phenyl acetic acid 1 through a series of reactions was converted into 5-benzyl-1,3,4-oxadiazole-2-thione 4. The intermediate oxadiazole 4 was then reacted with chloroacetyl derivatives of phenols 6a-f and anilines derivatives 8a-d to afford the title oxadiazole derivatives 9a-j. All of the title compounds 9a-j were evaluated for their inhibitory activity against human alkaline phosphatise (ALP). It was found that compounds 9a-j exhibited good to excellent alkaline phosphatase inhibitory activity especially 9h displayed potent activity with IC₅₀ value 0.420 ± 0.012 µM while IC₅₀ value of standard (KH₂PO₄) was 2.80 µM. The enzyme inhibitory kinetics of most potent inhibitor 9h was determined by Line-weaever Burk plots showing non-competitive mode of binding with enzyme. Molecular docking studies were performed against alkaline phosphatase enzyme (1EW2) to check the binding affinity of the synthesized compounds 9a-j against target protein. The compound 9h exhibited excellent binding affinity having binding energy value (−7.90 kcal/mol) compared to other derivatives. The brine shrimp viability assay results proved that derivative 9h was non-toxic at concentration used for enzyme assay. The lead compound 9h showed LD₅₀ 106.71 µM while the standard potassium dichromate showed LD₅₀ 0.891 µM. The DNA binding interactions of the synthesized compound 9h was also determined experimentally by spectrophotometric and electrochemical methods. The compound 9h was found to bind with grooves of DNA as depicted by both UV–Vis spectroscopy and cyclic voltammetry with binding constant values 7.83 × 10³ and 7.95 × 10³ M⁻¹ respectively revealing significant strength of 9h-DNA complex. As dry lab and wet lab results concise each other it was concluded that synthesized compounds, especially compound 9h may serve as lead compound to design most potent inhibitors of human ALP.