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Published on: November 17, 2018
Multimodal evaluation of lipoxygenase-targeting NSAIDs using integrated in vitro, SAR, in silico, cytotoxicity
Ambar Iqbal1, Wasim Abbas2, Samina Ejaz3
1Institute of Chemistry, Baghdad-ul-Jadeed Campus, The Islamia University of Bahawalpur, Bahawalpur 63100, Pakistan; Department of Biochemistry, Institute of Biochemistry, Biotechnology & Bioinformatics (IBBB), B.J. Campus, The Islamia University of Bahawalpur, Bahawalpur 63100, Pakistan.
Abstract:
Lipoxygenase (LOX) and cyclooxygenase (COX) pathways generate biologically active mediators implicated in inflammatory disorders and several classes of cancer. Non-steroidal anti-inflammatory drugs (NSAIDs) inhibit the COX pathway by inhibiting the COX-1 and COX-2 enzymes. We reported earlier that several NSAIDs, including naproxen, aspirin and acetaminophen, inhibited lipoxygenase (LOX) enzyme at sub-micromolar concentrations. In continuation, the present work demonstrates the anti-LOX activity of nine more NSAIDs supported by in vitro, in silico, MD simulation and breast cancer cell line studies. All tested drugs displayed potent to excellent inhibitory profiles with IC50 values <24.93 ± 0.64 microM. Aceclofenac (IC50 0.85 ± 0.06 microM) was the most active drug, followed by indomethacin (IC50 1.13 ± 0.07 microM), meloxicam (IC50 1.94 ± 0.07 microM) and ketorolac (IC50 9.26 ± 0.82 microM). Celecoxib (IC50 15.81 ± 0.71 microM), lornoxicam (IC50 16.54 ± 0.28 microM) and nimesulide (IC50 19.87 ± 0.85 microM) showed excellent inhibitory profiles. Flurbiprofen (IC50 21.73 ± 0.93 microM) and etoricoxib (IC50 24.93 ± 0.64 microM) moderately inhibited the target enzyme. SAR studies revealed that active molecules decorated with the carboxylate group afforded strong binding interactions as observed by in vitro assays and structural features. Other drugs, including enol derivatives and celecoxib, also showcased enhanced binding interactions. However, the cytotoxic effects of NSAIDs against the MCF-7 breast cancer cell line did not disclose significant anticancer activity. Molecular docking studies against human 5-LOX offered the best binding affinities for aceclofenac (-13.54 kcal/mol), accompanied by conventional hydrogen bonding and hydrophobic interactions as supported by the in vitro results. Docking studies with DNA dodecamer established minor groove binding with their possible role in DNA replication and gene expression. Density functional theory (DFT) and ESP studies, MD simulations and MMPBSA free energy calculations further reiterated the stability of ligand-receptor complexes. Overall, these findings highlight the potential of targeted NSAIDs as dual COX/LOX inhibitors with broader therapeutic relevance in inflammatory disorders.
Insights
Non-steroidal anti-inflammatory drugs (NSAIDs) show potent inhibition of the lipoxygenase (LOX) enzyme, with aceclofenac being the most active. While effective against LOX, these NSAIDs did not demonstrate significant anticancer activity in breast cancer cell lines.
Area of Science:
- Biochemistry
- Pharmacology
- Medicinal Chemistry
Background:
- Lipoxygenase (LOX) and cyclooxygenase (COX) pathways produce mediators involved in inflammation and cancer.
- Non-steroidal anti-inflammatory drugs (NSAIDs) are known COX inhibitors.
- Previous studies indicated NSAIDs can inhibit LOX.
Purpose of the Study:
- To investigate the anti-LOX activity of nine additional NSAIDs.
- To explore the in vitro, in silico, and molecular dynamics of NSAID-LOX interactions.
- To assess the cytotoxic effects of these NSAIDs on breast cancer cell lines.
Main Methods:
- In vitro enzyme inhibition assays to determine IC50 values.
- In silico molecular docking and molecular dynamics (MD) simulations.
- Density Functional Theory (DFT) and ESP studies.
- Cytotoxicity assays using the MCF-7 breast cancer cell line.
Main Results:
- All nine tested NSAIDs exhibited potent to excellent LOX inhibition (IC50 < 24.93 µM).
- Aceclofenac, indomethacin, meloxicam, and ketorolac were the most potent inhibitors.
- Structural analysis revealed strong binding interactions for NSAIDs with carboxylate groups.
- Molecular docking identified aceclofenac with the highest binding affinity to human 5-LOX.
- No significant anticancer activity was observed against the MCF-7 cell line.
Conclusions:
- NSAIDs can act as dual COX/LOX inhibitors, offering potential for inflammatory disorders.
- The carboxylate group is crucial for strong binding interactions with LOX.
- Further research into NSAIDs as anti-inflammatory agents is warranted, but not for direct anticancer therapy.
- NSAIDs may interact with DNA minor grooves, influencing replication and gene expression.
